Use of bifidobacterium transitional microorganisms

Abstract

Treatment and / or prevention of allergy and / or allergic sensitization. The present invention relates to Bifidobacterium longum transitional microorganisms for use in the treatment and / or prevention of allergy and / or allergic sensitization in an infant or young child.

Description

Technical Field

[0001] This invention relates to probiotics and prebiotics, particularly Bifidobacterium longum transitional microorganisms or prebiotics that promote the growth and / or survival of Bifidobacterium longum transitional microorganisms for use in the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children. Background Technology

[0002] Allergies are among the most common health problems, affecting patients of all ages. The prevalence of allergies has increased rapidly over the past few decades. It is estimated that more than one-third of the world's population is affected, leading to allergies being considered a novel epidemic in industrialized countries. The reasons for the steady increase in allergic diseases are not yet fully understood. The host's genetic background is a prominent factor, and recently discovered genes have been shown to be associated with respiratory allergies / asthma and skin symptoms. Environmental factors appear to play a significant role in the high prevalence and severity of allergic manifestations; these include lifestyle, pollution, smaller family sizes, and reduced stimulation of the immune system by microbes in early life stages due to improved sanitation.

[0003] Allergic sensitization during childhood, especially early childhood (particularly food allergen sensitization), is dangerous because the development of an "allergic phenotype" or "atopic" has been shown to promote subsequent sensitization to other allergens. Therefore, childhood allergic reactions may be the first step in an allergic cascade that can trigger multiple allergic reactions later in life (a process often referred to as the "atopic process"). For example, children with persistent food hypersensitivity in early childhood have a significantly increased risk of developing allergic rhinitis (hay fever) or asthma later in childhood. Children with mild food hypersensitivity also have an increased risk of respiratory allergies, but to a lesser extent than children with persistent food hypersensitivity. Therefore, alleviating the severity of food hypersensitivity may be crucial to slowing the "atopic process." In this context, managing the onset of allergic reactions and preventing allergic reactions during childhood and infancy are extremely important.

[0004] In the first few years after birth, an infant's immune system is actively developing. Intervening in, preventing, avoiding, managing, mitigating, or regulating allergic reactions in early childhood can affect not only short-term allergic conditions but also long-term allergic conditions later in life.

[0005] Food allergens are among the first allergens infants encounter early in life: milk proteins are commonly encountered by infants who are not exclusively breastfed. Milk proteins are indeed one of the most frequently observed causes of food allergies in infants, followed by egg and wheat proteins. Generally, food allergies in infants and young children can manifest as skin symptoms (rashes, eczema, and others) and gastrointestinal symptoms (abdominal cramps; pain, especially in the abdomen; vomiting). Food allergies are the most common trigger for severe allergic reactions, which can lead to life-threatening anaphylaxis. Other sensitization and allergic reactions can also occur when infants / young children are exposed to new foods (such as grains, vegetables, fruits, nuts, or fish) and airborne allergens (such as pollen, house dust mites, and pet dander). Adults are significantly affected by contact and respiratory allergies. Data from the WHO [Clark, MJ and Million, R. P (2009) Allergic rhinitis: market evolution, Nature Reviews, Drug Discovery, 8, pp. 271-272] indicates that as many as 30% to 40% of the world’s population suffers from some form of respiratory allergy.

[0006] Animals, especially small animals such as pets (and particularly companion animals such as dogs and cats), can also suffer from food allergies and food intolerances, as well as environmental allergens. These often manifest as symptoms similar to those in humans, such as gastrointestinal disturbances like diarrhea, vomiting, and abdominal discomfort, as well as dermatitis or itching. In small animals (especially dogs), the most common cause of chronic diarrhea is food-reactive enteropathy (dietary-reactive enteropathy or food-reactive diarrhea).

[0007] New strategies for treating and / or preventing allergic reactions and / or allergic sensitization still need to be developed. Summary of the Invention

[0008] The inventors have determined that *Bifidobacterium longum* subspecies (*Bifidobacterium longum* transitional microorganisms), which exist in the gut microbiome of mammals, particularly humans, during the transitional feeding period, can have beneficial effects on reducing the risk of allergic reactions and / or allergic sensitization. For example, the inventors have shown that *Bifidobacterium longum* transitional microorganisms may be able to modulate intestinal barrier permeability and / or promote an anti-inflammatory and / or tolerance-inducing environment in the gut microbiome during weaning.

[0009] Therefore, in a first aspect, the present invention provides a Bifidobacterium longum transitional microorganism for use in the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children, wherein the Bifidobacterium longum transitional microorganism comprises a strain deposited at the French National Center for Microbial Collection (CNCM) with accession number CNCM I-5942 or a Bifidobacterium longum transitional strain having the identifying characteristics of the Bifidobacterium longum transitional strain deposited with accession number CNCM I-5942.

[0010] The present invention also provides a prebiotic for use in treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children by promoting the growth and / or survival of the Bifidobacterium longum transitional microorganism according to the invention in the intestine of an infant or young child, wherein the prebiotic is: i. a polysaccharide substrate suitably selected from any of the groups listed in Tables 1 to 3; and / or ii. human milk oligosaccharides (HMOs).

[0011] The present invention also provides a combination of a prebiotic and a Bifidobacterium longum transitional microorganism according to the invention, the combination being used for the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children; wherein the prebiotic is: i. a polysaccharide substrate suitably selected from any of the groups listed in Tables 1 to 3; and / or ii. human milk oligosaccharides (HMOs).

[0012] The present invention also provides a prebiotic for use in treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children by promoting the growth of the Bifidobacterium longum transitional microorganism according to the invention in the intestine of an infant or young child.

[0013] The present invention also provides a combination of a prebiotic and a transitional Bifidobacterium longum microorganism according to the invention, the combination being used for the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children.

[0014] In another aspect, the present invention relates to the use of Bifidobacterium longum transitional microorganisms, prebiotics, or combinations thereof as defined herein for promoting immune tolerance in infants or young children, preferably by promoting the growth and / or survival of Bifidobacterium longum transitional microorganisms in the intestine of infants or young children.

[0015] On the other hand, the present invention relates to the use of Bifidobacterium longum transitional microorganisms, prebiotics or combinations thereof as defined herein for regulating intestinal barrier permeability in infants or young children, preferably by promoting the growth of Bifidobacterium longum transitional microorganisms in the intestine of infants or young children. Attached Figure Description

[0016] Figure 1- UPGMA phylogenetic tree of Bifidobacterium longum genome.

[0017] Figure 2 - Using 2×10 6 Transepithelial electrical resistance (TEER) of Caco-2 monolayers after apical treatment with CFU probiotic strains. TEER was measured at 2, 4, 6, and 24 hours post-treatment, and each value was normalized to its corresponding 0-hour value and displayed as a percentage of the initial value. Data were plotted as mean ± SEM. For each concentration, two-way ANOVA with Dunnett's multiple comparison test was used to assess the difference between the complete medium (CM) control and treatment at each time point, and statistical differences are indicated by (*). (*) = p < 0.05; (**) = p < 0.01; (***) = p < 0.001 and (****) = p < 0.0001. NCC5000-5004: transitional strain of Bifidobacterium longum, NCC2818: Bifidobacterium animalis subsp. lactis, and NCC3001: Bifidobacterium longum subsp. longum.

[0018] Figure 3 -Stimulation of peripheral blood mononuclear cells (PBMCs) for 36 hours in the presence of different bacterial strains, including 10 7 All transitional Bifidobacterium longum isolates and probiotic strains with CFU / ml. Cell culture supernatant was collected to assess cytokine expression of IL-10 and IL-12p40 by ELISA. Standard curves for each cytokine were used to calculate the absolute amount (picograms / ml) from the optical density readings. NCC5000-5004: transitional Bifidobacterium longum strains, NCC2818: Bifidobacterium animalis subsp. lactis, NCC3089: Bifidobacterium longum subsp. infantis, NCC4007: Lactobacillus rhamnosus, and NCC2705: Bifidobacterium longum subsp. longum.

[0019] Figure 4 - Carbohydrate-active enzymes (CAZyme) carried by transitional strains of Bifidobacterium longum (including NCC 5025).

[0020] Figure 5 - An additional illustration of glycoside hydrolases (GH) and polysaccharide lyases (PL) in the genome of the Bifidobacterium longum clade. The heatmap shows the presence (light color) and absence (dark color) of GH and PL genes, and the size of the circles represents the number of these genes in each genome of a particular strain.

[0021] Figure 6 - Growth curves of Bifidobacterium longum transitional strains, including NCC 5025 on 3-FL as the sole carbon source. The last figure shows the growth rate k obtained for each tested strain.

[0022] Figure 7 - Pectin (beetroot) and arabinogalactan (larch wood) promote the growth of Bifidobacterium longum transitional strain NCC 5001 in a complex gut microbiota community. P **** < 0.0001, *** < 0.001, ** < 0.01, * < 0.05, one-way ANOVA with uncorrected Fisher LSD.

[0023] Figure 8 -Arabinogalactan (larch wood) and starch (potato) promote the growth of Bifidobacterium longum transitional strain NCC 5002 in a complex gut microbiota community. P **** < 0.0001, *** <0.001, ** <0.01, * <0.05, one-way ANOVA with uncorrected Fisher LSD.

[0024] Figure 9 - A representative CAZyme sequence.

[0025] Figure 10 - A schematic diagram of the gene organization involved in the degradation and metabolism of fucosylated human milk oligosaccharides in transitional strains of Bifidobacterium longum compared to Bifidobacterium longum subsp. infantum ATCC 15697 and Bifidobacterium karyotype DSM 21854. Values ​​represent the percentage (%) of identity between different genes.

[0026] Figure 11 - Riboflavin biosynthesis genes present in transitional strains of *Bifidobacterium longum*. Subplot A shows the organization of the riboflavin biosynthesis operon as found in *Bifidobacterium longum* NCC 5000. Subplot B depicts a heatmap of these genes in closely related strains of *Bifidobacterium longum*. Genes were stained based on their % identity with the *NCC 5000* genes as identified by Blast.

[0027] Figure 12 - In vitro, transitional strains of *Bifidobacterium longum* reduced IL-5 expression by type 2 skewed helper T cells after 48 hours of stimulation to a degree similar to or even greater than that of the probiotic *Bifidobacterium lactis*. NCC5000-5004: *Bifidobacterium longum* transitional strain; NCC2818: *Bifidobacterium animalis* subsp. *lactobacterium*; NCC3089: *Bifidobacterium longum* subsp. *infant*.

[0028] Figure 13 -Stimulation of peripheral blood mononuclear cells (PBMCs) for 36 hours in the presence of different bacterial strains, including 10 7All transitional Bifidobacterium longum isolates and probiotic strains with CFU / ml. Cell culture supernatant was collected to assess IL-10 cytokine expression by ELISA. Standard curves for each cytokine were used to calculate the absolute amount (picograms / ml) from the optical density readings. NCC5000-5004: transitional Bifidobacterium longum strains, NCC2818: Bifidobacterium animalis subsp. lactis, NCC3089: Bifidobacterium longum subsp. infantis, NCC4007: Lactobacillus rhamnosus, and NCC2705: Bifidobacterium longum subsp. longum.

[0029] Figure 14 - In vitro batch fermentation of infant microbiota containing 3-fucosylated lactose (3FL) with or without supplementation with *Bifidobacterium longum* species or transitional *Bifidobacterium longum*. Total SCFA corresponds to the sum of peak integrals of acetate, butyrate, and propionate. Bar graphs indicate the dynamics of total SCFA consumption and production between 0 and 24 hours (blue bars) and between 24 and 48 hours (red bars). Detailed Implementation

[0030] Unless otherwise specified, all percentages are by weight.

[0031] As used herein, the terms “about” or “approximately” when referring to measurable values ​​such as parameters, quantities, durations, etc., are intended to cover variations in a particular value and variations arising from that particular value, such as variations in a particular value and variations arising from that particular value being 1 / -10% or less, 1 / -5% or less, 1 / -1% or less, and + / 0.1% or less, provided that such variations are suitable for making in the disclosed invention. It should be understood that the values ​​referred to by the modifiers “about” or “approximately” are themselves specifically and preferably disclosed.

[0032] The terms “subject,” “individual,” and “patient” are used interchangeably to refer to vertebrates, preferably mammals, and more preferably humans. Mammals include, but are not limited to, rodents, apes, humans, farm animals, sporting animals, and pets.

[0033] The term "infant" refers to a human individual under 12 months of age or a non-human animal of equivalent age.

[0034] As used herein, the term “toddler” or “walking infant” may refer to human subjects aged between 12 months and 5 years. Appropriately, “toddler” may refer to non-human animals of equivalent age.

[0035] The terms "supplementary feeding period," "supplementary period," "transition period," "transitional feeding period," and "weaning period" are used interchangeably and refer to the period during which milk (breast milk or formula) is replaced by other foods in an infant's or toddler's diet. This typically involves a gradual transition from exclusive breastfeeding (breast milk or formula) to a mixed diet that includes milk and / or solid foods. The transition period varies depending on the infant or toddler, but is generally from about 4 months to about 18 months of age, such as from about 6 months to about 18 months, but in some cases it can extend to about 24 months or longer. For humans, weaning typically begins between 4 and 6 months of age and is considered complete once the infant and / or toddler is no longer breastfed or formula-fed, usually around 24 months of age. In some implementations, the weaning period is from 4 to 24 months.

[0036] The term "composition" or "nutritional composition" refers to any kind of composition or formulation that provides nutritional benefits to an individual and is safe for consumption by humans or animals. A nutritional composition may be in solid (e.g., powder), semi-solid, or liquid form and may contain one or more macronutrients, micronutrients, food additives, water, etc. For example, a nutritional composition may contain the following macronutrients: protein sources, lipid sources, carbohydrate sources, and any combination thereof. Furthermore, a nutritional composition may contain the following micronutrients: vitamins, minerals, fiber, phytochemicals, antioxidants, prebiotics, probiotics, bioactive agents, metabolites (e.g., butyrate, docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), gamma-linolenic acid (GLA)), and any combination thereof. The composition may also contain food additives, such as stabilizers (when provided in liquid or solid form) or emulsifiers (when provided in liquid form). The amounts of various components (e.g., oligosaccharides) may be expressed in g / 100g of composition on a dry weight basis when the composition is in solid form (e.g., powder), or in g / L of composition concentration when the composition is in liquid form (the latter also covers liquid compositions that can be obtained by reconstituted powder in a liquid (such as milk, water), such as reconstituted infant formula or stage 2 infant formula or infant cereal products or any other formulation designed to provide nutrition for infants or young children). Typically, nutritional compositions are formulated for oral, enteral, parenteral, or intravenous administration, and generally include one or more nutrients selected from a variety of nutrients: lipid or fat sources, protein sources, and carbohydrate sources. Preferably, the nutritional composition is intended for oral administration.

[0037] In this specific implementation, the nutrient composition is a "synthetic nutrient composition." The expression "synthetic nutrient composition" means a mixture obtained by chemical and / or biological methods.

[0038] As used herein, the term "infant formula" refers to a food intended for specific nutritional purposes in the first few months after birth and which meets the nutritional needs of such infants (in accordance with Article 2(c) of European Commission Directive 91 / 321 / EEC 2006 / 141 / EC of 22 December 2006 concerning infant formula and follow-up formula). It also refers to nutritional compositions intended for infants, as defined in the Codex Alimentarius Commission (STAN 72-1981) and in provisions concerning infant specialties (including foods for specific medical purposes). The term "infant formula" encompasses both "Stage 1 infant formula" and "Stage 2 infant formula" or "follow-up formula."

[0039] Stage 2 infant formula or follow-up formula is introduced starting from the 6th month. Infant formula constitutes the main liquid component of this group's gradually diversifying diet.

[0040] The term "infant food" refers to food designed for specific nutritional purposes for infants or young children during the first few years of their lives.

[0041] The term "infant cereal composition" refers to a food intended for specific nutritional purposes for infants or young children during the first few years of their lives.

[0042] The term "GUM" refers to a milk-based beverage that is typically fortified with vitamins and minerals and is intended for use by toddlers or children.

[0043] The term "fortifier" refers to a liquid or solid nutritional composition suitable for fortifying or mixing with human milk, infant formula, growing milk, or human breast milk fortified with other nutrients. Therefore, a fortifier may be applied after being dissolved in human breast milk, infant formula, growing milk, or human breast milk fortified with other nutrients, or it may be applied as a standalone composition. When applied as a standalone composition, milk fortifiers may also be identified as "supplements".

[0044] The term "metabolism" is used here to refer to the ability of a substrate to be broken down, adsorbed, and / or utilized by microorganisms. For example, a substrate may promote and / or contribute to the growth and / or survival of microorganisms.

[0045] Suitablely, the term "capable of metabolizing glycan substrates" may mean that a Bifidobacterium longum transitional strain encodes at least one CAZyme capable of utilizing glycan substrates. For example, the CAZyme may be capable of catalyzing the hydrolysis of glycosidic bonds within the glycan substrate. Suitablely, the Bifidobacterium longum transitional strain may encode at least one, at least two, at least three, at least four, or at least five CAZymes capable of utilizing glycan substrates. Suitablely, the term "capable of metabolizing glycan substrates" may mean that the glycan substrate (or fibers or components containing the glycan substrate) is capable of promoting the growth and / or survival of the Bifidobacterium longum transitional strain (e.g., when added to an anaerobic culture of the Bifidobacterium longum transitional strain). The growth and / or survival of the Bifidobacterium longum transitional strain can be determined, for example, by measuring the abundance of 16S rDNA using PCR methods. An exemplary assay for measuring the growth of the Bifidobacterium longum transitional strain in the presence of a glycan substrate (e.g., in fiber form) is provided in this example.

[0046] Suitablely, the glycan substrate can be metabolized by *Bifidobacterium longum* transitional microorganisms. Suitablely, the glycan substrate may be able to promote the growth and / or survival of *Bifidobacterium longum* transitional strains. Glycan substrates that can promote the growth and / or survival of *Bifidobacterium longum* transitional strains can be determined, for example, by an anaerobic culture of *Bifidobacterium longum* transitional strains with the glycan substrate to be tested. The growth and / or survival of *Bifidobacterium longum* transitional strains can be determined by measuring bacterial cell number, cell density (e.g., by optical density measurement), and / or 16S rDNA abundance, for example using PCR methods. Exemplary assays for measuring the growth of *Bifidobacterium longum* transitional strains in the presence of glycan substrates are provided in the examples. Compared to the number of *Bifidobacterium longum* transitional strains in control anaerobic cultures without HMOs, a polysaccharide substrate that promotes the growth and / or survival of *Bifidobacterium longum* transitional strains can increase the number of *Bifidobacterium longum* transitional bacteria in anaerobic cultures by at least 20%, at least 30%, at least 40%, at least 50%, at least 75%, or at least 100%. Suitablely, the polysaccharide substrate that promotes the growth and / or survival of *Bifidobacterium longum* transitional strains can increase the number of *Bifidobacterium longum* transitional bacteria in anaerobic cultures by a statistically significant amount (e.g., p-value < 0.05 determined by one-way ANOVA) compared to the number of *Bifidobacterium longum* transitional bacteria in control anaerobic cultures without a polysaccharide substrate.

[0047] "Glycan substrate" refers to a glycan that can be metabolized by microorganisms. A glycan substrate can be, for example, a glycoconjugate, oligosaccharide, or polysaccharide. Glycan conjugates can include N-linked or O-linked glycans within glycoproteins and proteoglycans or glycolipids. For example, O-linked glycans can include proteins or peptides in which the oxygen atom of a serine or threonine residue is linked to a monosaccharide, oligosaccharide, or polysaccharide, as in the case of glycosaminoglycans (GAGs). Other examples of "glycan substrates" are cellulose, a glycan composed of β-1,4-linked D-glucose, and chitin, a glycan composed of β-1,4-linked N-acetyl-D-glucosamine. Glycans can be homopolymers or heteropolymers of monosaccharide residues and can be linear or branched. As used herein, "glycan substrate" includes, for example, oligosaccharides and polysaccharides.

[0048] "Oligosaccharide" can refer to a carbohydrate having more than two but relatively few monosaccharide units (typically three, four, five, six, and up to ten). Exemplary oligosaccharides include, but are not limited to, fructooligosaccharides, galactooligosaccharides (raffinose, stachyose, verbascose), maltodextrin, gentiosaccharides, cellulose oligosaccharides, lactoosaccharides (e.g., those found in mammary gland secretions), isomaltooligosaccharides, lactulose oligosaccharides, mannan oligosaccharides, melibiose-derived oligosaccharides, pectin oligosaccharides, and xylooligosaccharides.

[0049] The term "polysaccharide" can refer to a carbohydrate having more than ten monosaccharide units. Exemplary polysaccharides include, but are not limited to, starch, arabinogalactan, laminarin, golden kelp laminarin, xylan, arabinoxylan, mannan, fucoidan, and galactomannan. It should be understood that there is no precise boundary or distinction between the terms oligosaccharide and polysaccharide, and such distinction is not necessary for carrying out this invention.

[0050] The term "glycosaminoglycan" (GAG), or mucopolysaccharide, refers to a long, linear polysaccharide composed of repeating disaccharide units (i.e., two sugar units). The repeating disaccharide units consist of aldoses and amino sugars, with galactose present at the aldose position, except for keratin. GAGs are classified into four groups based on their core disaccharide structure.

[0051] As used in this article, "mucins" can refer to a family of high-molecular-weight, highly glycosylated proteins (glycoconjugates). A key characteristic of mucins is their ability to form gels; therefore, they are key components in most gel-like secretions, playing roles ranging from lubrication to cell signaling to the formation of mechanical or chemical barriers.

[0052] The term "HMO" refers to human milk oligosaccharides. These carbohydrates are highly resistant to enzymatic hydrolysis, suggesting that their important functions may not be directly related to their calorific value. It has been specifically noted in the art that these carbohydrates play a crucial role in early infant and toddler development, such as the maturation of the immune system. Many different types of HMOs have been found in human milk. Each individual oligosaccharide is based on a variety of combinations of glucose, galactose, sialic acid (N-acetylneuraminic acid), fucose, and / or N-acetylglucosamine with these molecules, resulting in a large number of diverse oligosaccharides in human milk; over 130 such structures have been identified to date. Almost all oligosaccharides have a lactose molecule at the reducing end, and the non-reducing end is occupied by sialic acid and / or fucose (if present). Based on the presence of fucose and sialic acid in the oligosaccharide structure, HMOs can be classified as unfucosylated (neutral) or fucosylated (neutral) and sialylated (acidic) and non-sialylated molecules, respectively.

[0053] The term "fucosylated oligosaccharide" refers to oligosaccharides containing fucose residues. These oligosaccharides are neutral. Some examples are 2'-fucosylvose (2'-FL), 3-fucosylvose (3-FL), difucosylvose (DiFL), lactose-N-fucopentose (e.g., lactose-N-fucopentose I, lactose-N-fucopentose II, lactose-N-fucopentose III, lactose-N-fucopentose V), lactose-N-fucohexasaccharide, lactose-N-difucohexasaccharide I, fucosylvose-N-hexasaccharide, fucosylvose-N-neohexose, difucosylvose-N-hexasaccharide I, difucosylvose-N-neohexose II, and any combination thereof. Fucosylated oligosaccharides represent the largest portion of human milk, with 2'-FL accounting for up to 30% of total HMOs. Fucosylated oligosaccharides are believed to reduce the risk of infection and inflammation and promote the growth and metabolic activity of certain symbiotic microorganisms, thereby reducing inflammatory responses.

[0054] The term "N-acetylated oligosaccharide" encompasses both "N-acetyl-lactoside" and "oligosaccharides containing N-acetyl-lactoside." Such oligosaccharides are neutral oligosaccharides having N-acetyl-lactoside residues. Suitable examples are LNT (lactose-N-tetrasaccharide), para-lactose-N-neohexose (para-LNnH), LNnT (lactose-N-neohexose), DSLNT (disialyllactose-N-tetrasaccharide), and any combination thereof. Other examples are lactose-N-hexasaccharide, lactose-N-neohexose, para-lactose-N-hexasaccharide, para-lactose-N-neohexose, lactose-N-octasaccharide, lactose-N-neohexose, isol-lactose-N-octasaccharide, para-lactose-N-octasaccharide, and lactose-N-decansaccharide.

[0055] The expressions “at least one fucoidylated oligosaccharide” and “at least one N-acetylated oligosaccharide” should be understood as “at least one type of fucoidylated oligosaccharide” and “at least one type of N-acetylated oligosaccharide”.

[0056] The term "sialylated oligosaccharide" refers to an oligosaccharide containing charged sialic acid residues. This oligosaccharide is acidic. Some examples are 3'-sialyllactose (3-SL), 6'-sialyllactose (6-SL), and sialyllactose-N-tetrasaccharide (Lst, such as Lst-a, Lst-b, or Lst-c).

[0057] Appropriately, the term "capable of metabolizing HMOs" may mean that a Bifidobacterium longum transitional strain encodes at least one CAZyme capable of utilizing HMOs. For example, this CAZyme may be capable of catalyzing the hydrolysis of glycosidic bonds within the HMO. Appropriately, the Bifidobacterium longum transitional strain may encode at least one, at least two, at least three, at least four, or at least five CAZymes capable of utilizing HMOs. Appropriately, the term "capable of metabolizing HMOs" may mean that the HMO promotes the growth and / or survival of the Bifidobacterium longum transitional strain (e.g., when added to an anaerobic culture of the Bifidobacterium longum transitional strain). The growth and / or survival of the Bifidobacterium longum transitional strain can be determined, for example, by measuring the abundance of 16S rDNA using PCR methods.

[0058] The term fiber is used herein to refer to carbohydrates that are indigestible in humans or animals. Such fibers in relation to carbohydrates are also discussed herein. Suitablely, fiber may be fermented by one or more Bifidobacterium longum transitional microorganisms provided in the uses or compositions of this invention and / or fermented in one or more regions of the gastrointestinal tract of an organism such as a human or non-human animal. In the context of this invention, as used herein, the expressions “fiber” or “multiple fibers” or “dietary fiber” or “various dietary fibers” refer to the portion of a plant-derived food that is indigestible in the small intestine, comprising two main components: soluble fiber, which dissolves in water; and insoluble fiber. Mixtures of fibers are included within the scope of the terms mentioned above. Soluble fiber readily ferments in the colon into gases and physiologically active byproducts and may be prebiotic and viscous. Insoluble fiber is insoluble in water, metabolically inert and bloated, or it may be prebiotic and fermented in the large intestine. Chemically, dietary fiber consists of carbohydrate polymers having three or more monomeric units that are not hydrolyzed by endogenous enzymes in the small intestine, such as arabinoxylan, cellulose, and many other plant components such as resistant starch, resistant dextrin, inulin, lignin, chitin, pectin, arabinogalactan, arabinogalactan, galactan, xylan, β-glucan, and oligosaccharides. Non-limiting examples of dietary fiber include: prebiotic fibers such as fructooligosaccharides (FOS), inulin, galactooligosaccharides (GOS), fruit fiber, plant fiber, cereal fiber, and resistant starches such as high-amylose corn starch.

[0059] As used herein, “added fiber” or “added dietary fiber” indicates an ingredient that consists primarily or entirely of fiber added to a nutritional supplement composition, and the amount of fiber in which it constitutes the total fiber content of the composition. The total fiber content of a nutritional supplement composition is provided by the sum of the amount of fiber naturally present in the ingredients used in the formulation (e.g., from whole grain flour) and the amount of added fiber.

[0060] The term "prebiotic" refers to non-digestible carbohydrates that exert a beneficial effect on the host by selectively stimulating the growth and / or activity of beneficial healthy bacteria (such as Bifidobacteria in the human colon) (Gibson GR, Roberfroid MB. Dietary modulation of the human colonic microbiota: introducing the concept of prebiotics. J Nutr. 1995; 125:1401-12).

[0061] The term "probiotics" refers to a microbial cell preparation or microbial cell component that has a beneficial effect on the health or well-being of the host (Salminen S, Ouwehand A. Benno Y. et al., "Probiotics: how should they be defined" Trends Food Sci. Technol. 1999: 10 107-10). The microbial cells according to the present invention are typically bacteria.

[0062] The term "cfu" should be understood as colony-forming unit.

[0063] The gut microbiota is the composition of microorganisms (including bacteria, archaea, and fungi) that live in the digestive tract.

[0064] The term “gut microbiome” can include the “gut microbiota” and their “site of activity”, which can include their structural elements (nucleic acids, proteins, lipids, polysaccharides), metabolites (signaling molecules, toxins, organic and inorganic molecules), and molecules produced by the coexisting host and structured by the surrounding environmental conditions (Berg, G. et al., 2020. Microbiome, 8(1), pp. 1-22).

[0065] Bifidobacterium longum transitional microorganism

[0066] Previously, subspecies of *Bifidobacterium longum* belonging to an evolutionary clade in the gut microbiome of mammals, particularly humans, during the transitional feeding period, have been identified. *Bifidobacterium longum* microorganisms belonging to this clade are referred to herein as *Bifidobacterium longum* transitional. *Bifidobacterium longum* transitional strains NCC 5000, NCC 5001, NCC 5002, NCC 5003, and NCC 5004 were deposited by SOCIÉTÉ DES PRODUITS NESTLÉ SA on May 11, 2021, at the French National Center for Microbial Collections (CNCM), Pasteur Institute, under the Budapest Treaty, with accession numbers CNCM I-5683, CNCM I-5684, CNCM I-5685, CNCM I-5686, and CNCM I-5687, respectively. The transitional strain of *Bifidobacterium longum* referred to herein as NCC 5025 was deposited by Nestlé at the CNCM, Pasteur Institute, on March 29, 2023, under the Budapest Treaty, with accession number CNCM I-5942. Patent application WO2023 / 278441 indicates that the relative abundance of the transitional *Bifidobacterium longum* microorganism during the transitional feeding period (e.g., weaning) is greater than that of *Bifidobacterium longum* subsp. infantis (*Bifidobacterium infantis*) or *Bifidobacterium longum* subsp. longum. In fact, the relative abundance of *Bifidobacterium longum* subsp. infantis decreases at the beginning of the transitional feeding period and continues until the end of the transitional feeding period, while the abundance of *Bifidobacterium longum* subsp. longum begins to increase. Vatanen et al. demonstrated that this unique Bifidobacterium longum clade expanded with the introduction of solid foods and possesses enzymes for utilizing breast milk and solid food substrates (Vatanen et al.; 2022, Cell 185, 1-18; published online November 1, 2022; https: / / doi.org / 10.1016 / j.cell.2022.10.011).

[0067] Suitable, the Bifidobacterium longum transitional microorganism may encode one or more CAZymes selected from the groups listed in Table 1. Suitable, the Bifidobacterium longum transitional microorganism may encode one or two CAZymes selected from the groups listed in Table 1.

[0068] Appropriately, the Bifidobacterium longum transitional microorganism encodes at least one CAZyme selected from the groups listed in Table 1 and one or more CAZymes selected from the groups listed in Tables 2 and 3. For example, the Bifidobacterium longum transitional microorganism may encode at least 2, at least 5, at least 10, at least 20, or at least 30 CAZymes selected from the groups listed in Tables 2 and 3.

[0069] Appropriately, the transitional microbial code for Bifidobacterium longum is (i) at least one CAZyme selected from the groups listed in Table 1 and (ii) each of the CAZymes listed in Table 3 or except GH5_44 of the CAZymes listed in Table 3.

[0070] Appropriately, the transitional microbial code for Bifidobacterium longum is (i) at least one CAZyme selected from the groups listed in Table 1 and (ii) each of the CAZymes listed in Table 3 or except GH25 of the CAZymes listed in Table 3.

[0071] Suitablely, the Bifidobacterium longum transitional microorganism does not encode one or more of the CAZymes listed in Table 4. Suitablely, the Bifidobacterium longum transitional microorganism does not encode any of the CAZymes listed in Table 4.

[0072] The *Bifidobacterium longum* transitional microorganism of the present invention advantageously carries a gene encoding CAZyme, thereby allowing the cleavage of sialic acid residues from glycans such as sialylated oligosaccharides, glycoproteins, and glycolipids. This allows for the efficient utilization of sialylated oligosaccharides present in breast milk at weaning and can therefore participate in the proper development of the gut microbiome in infants and / or young children. It may also help prevent the presence of intestinal pathogens.

[0073] In some implementations, the Bifidobacterium longum transitional microorganism contains a sialidase or neuraminidase family 33 (GH33, sialidase or neuraminidase) gene that has at least 60% identity with the BLON_2348 gene present in Bifidobacterium longum infant subspecies ATCC15697.

[0074] In some embodiments, the Bifidobacterium longum transitional microorganism contains a BLON_2348 gene that has approximately 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity with the sialidase or neuraminidase family 33 (GH33, sialidase or neuraminidase) gene present in Bifidobacterium longum infantis subsp. ATCC15697.

[0075] In some embodiments, the Bifidobacterium longum transitional microorganism contains a BLON_2348 gene with at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least 72%, at least 74%, at least 73%, at least 75%, at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sialidase or neuraminidase family 33 (GH33, sialidase or neuraminidase) gene present in Bifidobacterium longum infantis subsp. ATCC15697.

[0076] In some embodiments, the transitional Bifidobacterium longum microorganism used according to the present invention comprises: a glycosyl hydrolase family 95 (GH95, α-L-galactosidase; α-L-fucosidase; α-1,2-L-fucosidase) gene, which has at least 60% identity with the BLON_2335 gene present in Bifidobacterium longum subsp. infantis ATCC 15697; and / or a glycosyl hydrolase family 29 (GH29, α-L-fucosidase; α-1,3 / 1,4-L-fucosidase; α-1,2-L-fucosidase), which has at least 60% identity with the BLON_2336 gene present in Bifidobacterium longum subsp. infantis ATCC 15697.

[0077] In some embodiments, the Bifidobacterium longum transitional microorganism used according to the present invention comprises a sialidase or neuraminidase family 33 (GH33, sialidase or neuraminidase) gene that has at least 60% identity with the BLON_2348 gene present in Bifidobacterium longum infant subspecies ATCC 15697.

[0078] In some implementations, the Bifidobacterium longum transitional microorganism preferentially utilizes 3-fucosyllactose (3-FL).

[0079] In some implementations, compared with the Bifidobacterium longum strain deposited in CNCM with accession number CNCM I-5942, the transitional strain of Bifidobacterium longum has an ANI of at least 99.0%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9%.

[0080] Preferably, compared with the Bifidobacterium longum strain deposited in CNCM with accession number CNCM I-5942, the transitional strain of Bifidobacterium longum has at least 99.9% ANI.

[0081] Appropriately, as described herein, compared with the Bifidobacterium longum strain deposited in CNCM with accession number CNCM I-5942, the transitional strain of Bifidobacterium longum has an ANI of at least 98.1%, at least 98.2%, at least 98.3%, at least 98.4%, at least 98.5%, at least 98.6%, at least 98.6%, at least 98.7%, at least 98.8%, at least 98.9%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9%, and has at least one identifying characteristic of the transitional strain of Bifidobacterium longum deposited with accession number CNCM I-5942.

[0082] Appropriately, as described herein, compared with the Bifidobacterium longum strain deposited in CNCM with accession number CNCM I-5942, the transitional strain of Bifidobacterium longum has an ANI of at least 98.4%, at least 98.5%, at least 98.6%, at least 98.6%, at least 98.7%, at least 98.8%, at least 98.9%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9%, and has at least one identifying characteristic of the transitional strain of Bifidobacterium longum deposited with accession number CNCM I-5942.

[0083] Methods for microbial genome sequencing are known in the art (see, for example, Segerman; Front. Cell. Infect. Microbiol.; 2020; 10; Article 527102 & Donkor; Genes; 2013;4(4); 556-572). Metagenomic methods can be used by way of example. For example, shotgun sequencing data can be used for suitable metagenomic methods. Suitable metagenomic methods are known in the art and include, for example, MetaPhlAn 3.0 (see Beghini et al.; eLife 2021; 10: e65088; https: / / huttenhower.sph.harvard.edu / metaphlan) for metagenomic sequencing.

[0084] "Average Nucleotide Identity (ANI)" is a term in this field referring to a distance-based method of characterizing species based on pairwise comparisons of genome sequences, and is a computerized alternative to the traditional DNA-DNA hybridization (DDH) technique already used for the phylogenetic definition of species (Goris et al., 2007, "DNA-DNA hybridization values ​​and their relationship to whole-genome sequence similarities", Int. J. Syst. Evol. Microbiol. 57: 81-91). Based on DDH, strains with a correlation greater than 70% are considered to belong to the same species (see, for example, Wayne et al., 1987, Report of the Ad-Hoc-Committee on Reconciliation of Approaches to Bacterial Systematics. Int J Syst Bacteriol 37: 463-464). ANI is similar to the aforementioned 70% DDH cutoff value and can be used for species characterization. ANI has been evaluated in multiple laboratories and has become the gold standard for species demarcation (see, for example, Kim et al., 2014, “Towards ataxonomic coherence between average nucleotide identity and 16S rRNA genesequence similarity for species demarcation of prokaryotes”, Int. J.Syst.Evol.Micr.64: 346-351; Richter et al., 2009, “Shifting the genomic goldstandard for the prokaryotic species definition”, P Natl Acad Sci USA 106:19126-19131; and Chan et al., 2012, “Defining bacterial species in the genomic era:insights from the genus Acinetobacter”, Bmc.Microbiol.12)).

[0085] The ANI (Average Nucleus Indices) of shared genes between two strains is known to be a powerful tool for comparing genetic relatedness between strains, and approximately 95% of ANI values ​​correspond to the 70% DNA-DNA hybridization criterion used to define species. See, for example, Konstantinidis and Tiedje, Proc Natl Acad Sci USA, 102(7):2567-72 (2005); and Goris et al., Int Syst Evol Microbiol.57(Pt 1):81-91 (2007). The ANI between two bacterial genomes is calculated by pairwise comparisons of all sequences common to both strains and can be determined, for example, using any of a number of publicly available ANI tools, including but not limited to OrthoANI with usearch (Yoon et al., Antonie van Leeuwenhoek 110:1281-1286 (2017)); ANI calculator, JSpecies (Richter and Rossello-Mora, Proc Natl Acad Sci USA 106:19126-19131 (2009)); and JSpeciesWS (Richter et al., Bioinformatics 32:929-931 (2016)). Other methods for determining the ANI between two genomes are known in the art. See, for example, Konstantinidis, KT and Tiedje, JM, Proc. Natl. Acad. Sci. USA, 102: 2567-2572 (2005); and Varghese et al., Nucleic Acids Research, 43(14):6761-6771 (2015). In one specific implementation, the ANI between two bacterial genomes can be determined, for example, by averaging the nucleotide identity of homologous genes identified as bidirectional best hits (BBHs). Protein-coding genes in the first genome (genome A) and the second genome (genome B) are compared at the nucleotide level using a similar search tool, such as NSimScan (Novichkov et al., Bioinformatics 32(15): 2380-23811 (2016)). The results are then filtered to show at least 70% sequence identity for BBHs that retain only those shorter than 70% of the shorter sequence in each BBH pair. The ANI between genome A and genome B is defined as the percentage of identity multiplied by the sum of the alignment lengths of all BBH genes, then divided by the sum of the lengths of the BBH genes. These and ANI determination techniques are known in the art.

[0086] In some embodiments, the Bifidobacterium longum transitional microorganism used in this invention is isolated from humans.

[0087] In some other implementations, the transitional microorganism of Bifidobacterium longum is not a subspecies of Bifidobacterium longum or Bifidobacterium longum infantis.

[0088] Suitable, the Bifidobacterium longum transitional microorganism is provided in the form of a probiotic. Suitable, the Bifidobacterium longum transitional microorganism is provided in the composition.

[0089] Treatment and / or prevention of allergic reactions and / or allergic sensitization

[0090] As used in this article, "allergy" can refer to allergic diseases or allergic reactions (including their symptoms).

[0091] Bifidobacterium longum transitional microorganisms and / or prebiotics can be used to treat and / or prevent allergic reactions and / or allergic sensitization in infants or young children.

[0092] As used herein, “treatment” can refer to the administration of Bifidobacterium longum transitional microorganisms and / or prebiotics to a subject with an existing disease or condition in order to reduce, alleviate or improve at least one symptom associated with the disease and / or slow, mitigate or halt the progression of the disease.

[0093] As used herein, “prevention” can refer to the administration of Bifidobacterium longum transitional microorganisms and / or prebiotics to subjects who have not yet contracted the disease and / or do not exhibit any symptoms of the disease, in order to prevent or attenuate the cause of the disease or to reduce or prevent the development of at least one symptom associated with the disease. Individuals may be predisposed to developing the disease, or are considered to be at risk of developing the disease. Appropriately, Bifidobacterium longum transitional microorganisms and / or prebiotics may be administered to subjects to reduce the likelihood of allergic reactions and / or allergic sensitization in infants or young children.

[0094] Allergic sensitization during childhood, especially early childhood (particularly food allergen sensitization), is dangerous because the development of an "allergic phenotype" or "atopic" has been shown to lead to subsequent sensitization to other allergens. Therefore, childhood allergic reactions may be the first step in an allergic cascade that can trigger multiple allergic reactions later in life (a process often referred to as the "atopic process"). For example, children with persistent food hypersensitivity in early childhood have a significantly increased risk of developing allergic rhinitis (hay fever) or asthma later in childhood (Ostblom, E. et al., 2008, Phenotypes of food hypersensitivity and development of allergic diseases during the first 8 years of life, Clinical and Experimental Allergy, 38(8), pp. 1325-1332). Children with mild food hypersensitivity also have an increased risk of respiratory allergies, but to a lesser extent than children with persistent food hypersensitivity. Therefore, alleviating the severity of food hypersensitivity may be crucial to slowing the "atopic reaction process." In this context, managing allergic episodes and preventing allergic reactions during childhood and infancy are extremely important.

[0095] In one implementation, the prevention and / or reduction of the risk of allergic reactions and / or allergic sensitization is achieved through primary prevention. "Primary prevention" refers to the effect of preventing or reducing the risk of a patient being sensitized by an allergen, characterized by the absence of allergen-specific IgE antibodies or a reduction in the level of allergen-specific IgE antibodies. The result of preventing or reducing sensitization can be the absence of allergic symptoms or a reduction in allergic symptoms upon exposure to the same allergen. Subsequent allergic responses can also be modulated by modulating the manner in which a patient is sensitized by one or a group of allergens (primary prevention).

[0096] Food allergens are among the first allergens infants encounter in early life, and milk proteins are a common source, not just for breastfed babies. Milk proteins are indeed one of the most frequently observed causes of food allergies in infants, followed by egg and wheat proteins. Generally, food allergies in infants and young children can manifest as skin symptoms (rashes, eczema, and others) and gastrointestinal symptoms (abdominal colic; pain, especially in the abdomen; vomiting). Food allergies are the most common trigger for severe allergic reactions, which can lead to life-threatening anaphylaxis.

[0097] Other sensitization and allergic reactions may also occur when infants / young children are exposed to new foods (such as grains, vegetables, fruits, nuts, or fish) and airborne allergens (such as pollen, house dust mites, and pet dander). Adults are significantly affected by contact allergies and respiratory allergies. Recent data from the WHO (Clark, MJ, and Million, R. P., 2009, Allergic rhinitis: market evolution, Nature Reviews, DrugDiscovery, 8, pp. 271-272) indicate that as many as 30% to 40% of the world's population is affected by some form of respiratory allergy.

[0098] Animals, especially small animals such as pets (and particularly companion animals such as dogs and cats), can also suffer from food allergies and food intolerances, as well as environmental allergens. These often manifest as symptoms similar to those in humans, such as gastrointestinal disturbances like diarrhea, vomiting, and abdominal discomfort, as well as dermatitis or itching. In small animals (especially dogs), the most common cause of chronic diarrhea is food-reactive enteropathy (dietary-reactive enteropathy or food-reactive diarrhea).

[0099] By treating and / or preventing or reducing the risk of allergen sensitization in subjects, the compounds and compositions of the present invention can be used to prevent or treat food allergies, respiratory allergies and skin allergies.

[0100] In some embodiments, the allergic response is a specific IgE-related immune response and / or a T-cell-dependent hypersensitivity reaction. Therefore, in some embodiments, treating and / or preventing and / or reducing the risk of allergic reactions and / or allergic sensitization includes reducing or preventing specific IgE-related immune responses and / or T-cell-dependent hypersensitivity reactions. In some embodiments, allergic inflammation is reduced and / or tolerability is enhanced (e.g., oral tolerability).

[0101] Appropriately, allergic diseases are selected from one or more of the following groups: food allergies, respiratory allergies, and skin allergies.

[0102] In one implementation, the allergic disease is selected from one or more of the following groups: rhinitis, asthma, dermatitis, atopic dermatitis, contact dermatitis, eczema, atopic eczema, urticaria, psoriasis, eosinophilic esophagitis, and eosinophilic-related gastrointestinal diseases.

[0103] In one implementation, the allergen in an allergic disease is selected from one or more of the following: food allergens, dust mites, pollen, mold or mold spores, weed pollen, tree pollen, grass pollen, fleas, pet hair, feathers, pollution, or pet dander.

[0104] In one implementation, the allergen in allergic diseases is a food allergen. Suitablely, the food allergen is selected from: nuts, tree nuts, peanuts, fish, shellfish, mollusks, crustaceans, milk, eggs, soybeans, gluten, cereals, wheat, oats, barley, rye, celery, corn, lupins, sulfites, sesame, mustard, rice, poultry, and meat.

[0105] In one implementation, the allergen is an airborne sensitizer. Suitablely, the airborne sensitizer is selected from dust mites, pollen, mold or mold spores, weed pollen, tree pollen, grass pollen, fleas, pet hair, feathers, pollution, or pet dander.

[0106] Appropriately, treating, preventing, or reducing the risk of allergic reactions and / or allergic sensitization may refer to reducing or improving one or more symptoms as described herein.

[0107] As used in this article, "food allergy" refers to an abnormal immune response to one or more food allergens, typically an IgE response triggered by histamine release, but also encompassing non-IgE immune responses. Symptoms of food allergies may include itching, swollen tongue, vomiting, diarrhea, hives, difficulty breathing, or low blood pressure. When symptoms are severe, it is referred to as an allergic reaction.

[0108] As used herein, the term "food allergen" refers to a protein or its derivative that elicits an abnormal immune response. Purified food allergens may be named using the systematic nomenclature of the Allergen Nomenclature Sub-Committee of the World Health Organization and the International Union of Immunology. Allergen names consist of an abbreviation of the scientific name of its source (genus: 3 to 4 letters; species: 1 to 2 letters) and Arabic numerals; for example, Der p 1 refers to the first allergen to be described from the house dust mite *Dermatophagoides pteronyssinus*. Food allergens originate from proteins with diverse biological functions, including proteases, ligand-binding proteins, structural proteins, pathogen-associated proteins, lipid transfer proteins, exoproteins, and calcium-binding proteins. The list of food allergens is available on the official website of the WHO / IUIS Allergen Nomenclature Database: http: / / www.allergen.org / index.php. (Radauer, C. et al., 2014, Allergy, 69(4), pp. 413-419; and Pomés, A. et al., 2018. Molecular immunology).

[0109] As used herein, “respiratory allergy” or “airborne sensitizer” refers to an abnormal immune response to one or more airborne allergens. Airborne allergens may include pollen, mold or mold spores, weed pollen, tree pollen, grass pollen, and pet dander. Respiratory allergies may include, for example, allergic rhinitis and allergic asthma. Symptoms of allergic rhinitis (hay fever) include runny nose or nasal congestion, sneezing, red, itchy, and watery eyes, and swelling around the eyes. Symptoms of allergic asthma include wheezing attacks, coughing, chest tightness, and shortness of breath.

[0110] As used herein, “cutaneous allergy” refers to an abnormal immune response caused by exposure to one or more environmental allergens. Environmental allergens may include food allergens, dust mites, pollen, mold or mold spores, weed pollen, tree pollen, grass pollen, fleas, pet dander, feathers, or pet dander. Cutaneous allergies may include, for example, dermatitis, atopic dermatitis, contact dermatitis, eczema, atopic eczema, urticaria, and psoriasis. These are generally a group of conditions that cause inflammation of the skin, and symptoms include itching, redness of the skin, and rash.

[0111] Allergic diseases can also include other allergic inflammatory conditions, such as eosinophilic esophagitis and eosinophil-related gastrointestinal disorders. Eosinophilic esophagitis is an allergic inflammatory condition of the esophagus involving eosinophils (a type of white blood cell). Symptoms include difficulty swallowing, food impaction, vomiting, and heartburn.

[0112] Suitablely, the *Bifidobacterium longum* transitional microorganism and / or prebiotic of the present invention can increase the levels of anti-inflammatory cytokines in infants or young children. Suitablely, the *Bifidobacterium longum* transitional microorganism and / or prebiotic can increase the levels of IL-10 in infants or young children. IL-10 is a key immunomodulatory cytokine secreted by several immune cells, including regulatory T cells, and has been shown to attenuate the pro-inflammatory IL-12p40 / helper T cell type 1 mediated response. This cytokine is crucial in the establishment and maintenance of gut homeostasis between host immune cells and the gut microbiota. Therefore, aberrant IL-10 signaling has been reported in chronic inflammatory bowel diseases such as inflammatory bowel disease (IBD). Changes in the gut microbiota at weaning induce a strong immune response, as well as a regulatory immune response (weaning response). Interference with this weaning response may increase susceptibility to later immunopathologies such as allergic diseases.

[0113] Appropriately, Bifidobacterium longum transitional microorganisms and / or prebiotics can increase the IL-10 / IL-12 ratio in infants or young children.

[0114] Appropriately, Bifidobacterium longum transitional microbes and / or prebiotics can reduce the levels of pro-inflammatory cytokines, particularly those associated with allergic and / or atopic responses. For example, Bifidobacterium longum transitional microbes and / or prebiotics can reduce IL-5 levels in infants or young children. IL-5 is an interleukin produced by type 2 helper T cells and mast cells, which stimulates B cell growth and increases immunoglobulin secretion (primarily IgA). It is also a key regulator in eosinophil activation. IL-5 is associated with the etiology of several allergic diseases, including allergic rhinitis and asthma. Lowering IL-5 levels has been shown to reduce allergic symptoms in diseases such as severe asthma, for example (Casale et al.; 2021; Annals of Allergy, Asthma & Immunology; 127(3); 354-362 and Sposato et al.; 2021; Int Arch Allergy Immunol; 182(4):311-318).

[0115] The cytokine effects mediated by the probiotics and / or prebiotics of this invention can be systemic. Therefore, cytokine effects (e.g., increased IL-10 levels, increased IL-10 / IL-12 ratio, and / or decreased IL-5) can systemically reduce the risk of allergic reactions and / or allergic sensitization and / or promote immune tolerance, as described herein. Cytokine effects can occur locally in the intestines, lungs, and / or skin of infants or young children. Suitablely, cytokine effects can occur in the intestines of infants or young children. Therefore, cytokine effects can reduce the risk of allergic reactions and / or allergic sensitization and / or promote immune tolerance in specific organs or systems.

[0116] Appropriately, Bifidobacterium longum transitional microorganisms and / or prebiotics can modulate the permeability of the intestinal epithelial barrier in infants or young children. Appropriately, Bifidobacterium longum transitional microorganisms and / or prebiotics can reduce the permeability of the intestinal epithelial barrier. Increased permeability of the intestinal epithelial barrier may be associated with, for example, increased transepithelial hapten and antigen crossing of the intestinal epithelium.

[0117] The gut microbiota constitutes the earliest and largest stimuli for the development of gut-associated lymphoid tissue and the related immune system.

[0118] Epidemiological studies have demonstrated a link between Westernized lifestyles (e.g., lower consumption of fermented foods, increased use of antibiotics and other medications, and improved sanitation) and an increase in allergic diseases. This relates to the so-called "hygiene hypothesis," which posits that limited exposure to microbial stimuli in early childhood is a major factor influencing the prevalence of allergic diseases. Epidemiological studies have indeed demonstrated a correlation between the occurrence of allergic diseases and gut microbiota dysbiosis. Epidemiological data show that children with specific allergies have different gut microbiota compared to those with non-specific allergies.

[0119] Such changes in the gut microbiota can also negatively impact intestinal barrier integrity. Impaired barrier function, termed "leaky gut," has long been considered a susceptibility factor for gastrointestinal diseases (Heyman, M. Eur.J. Gastroenterol.Hepatol.17:1279-1285; Odenwald M., Nature Reviews Gastroenterology & Hepatology, (2017), (14), 9 21). Therefore, alterations in intestinal barrier integrity / function have multiple consequences, depending on other incidental findings and genetic and epigenetic conformations that contribute to a variety of diseases. Patients with food allergies often exhibit increased intestinal permeability, which correlates with the severity of their clinical symptoms (Ventura, M. T et al., 2006.Dig.Dis.Sci.38:732-736). Preclinical animal models also provide corroborating evidence supporting the role of intestinal barrier dysfunction and leaky gut in susceptibility to oral sensitization and subsequent development of food allergies. In addition, Western diet-induced changes in intestinal permeability promote food allergen sensitization and clinical allergic symptoms in mice in response to dietary antigens (Hussain M. et al., J. Allergy Clin. Immunol. (2019). Probiotics represent a nutritional approach for improving / enhancing intestinal barrier integrity and / or function (Ewaschuk JB et al., Am JPhysiol Gastrointest Liver Physiol. Nov 2008; 295(5):G1025-34). Therefore, enhancing intestinal barrier integrity through probiotic supplementation could prevent sensitization to orally administered allergens in at-risk individuals. (Tulyeu J, Microorganisms. 2019, Oct 16; 7(10)). In addition to the established role of intestinal barrier function in allergen sensitization, uncontrolled immune responses to dietary or environmental antigens promote the development of type 2 immune-mediated allergic diseases. Probiotic cultures or mixtures have well-known immunomodulatory properties that can prevent or mitigate allergic responses.

[0120] The epithelial barrier of human newborns is not fully mature at birth. Transfer of macromolecules or antigens across the intestinal epithelium of infants or young children induces the differentiation of regulatory T cells (Tregs) and is essential for inducing tolerance and preventing allergic diseases. However, the transepithelial passage of antigens is important in determining the immune response / outcome. Transcellular passage with intestinal epithelial cell processing may be preferred over uncontrolled paracellular passage (leakage), which may instead lead to inflammation and / or sensitization.

[0121] As demonstrated in this example, the transitional Bifidobacterium longum microbe encodes genes associated with the production of riboflavin and folic acid.

[0122] Riboflavin (vitamin B2) is an essential metabolite for host physiological processes. Mammalian hosts cannot produce riboflavin and are therefore strictly dependent on external supplies from diet and the gut microbiome. Once converted into its active forms (FAD and FMN), riboflavin participates in a variety of metabolic pathways, including energy metabolism, fatty acid oxidation, and purine catabolism.

[0123] Preclinical and clinical studies of acute and chronic riboflavin deficiency have shown mucosal damage (EFSA Journal. 2010; 8(10):1814.). The gastrointestinal tract contains one of the largest mucosal epithelial cells, which forms a direct barrier that allows for nutrient absorption and immune sensing while limiting the transport of potentially harmful antigens and microorganisms. Riboflavin has been found to maintain the structure and function of the small intestine, including the size and cellular structure of the duodenal crypts in rat models (Yates et al., Br J Nutr. 2001; 86(5):593-9) and the length and number of villi (Williams et al., Gut. 1996;39:220-225). Yates et al. also demonstrated the importance of luminal riboflavin in early postnatal gastrointestinal development in rats, as luminal riboflavin deficiency led to crypt hypertrophy and reduced crypt bifurcation even when systemic riboflavin status was maintained via injection (Yates et al., Dig Dis Sci 2003; 48(6):1159-64.). Riboflavin depletion affects the proliferation and proliferative potential of adult intestinal cells, which may impair gastrointestinal function (Nakano et al., Dig Dis Sci 2011; 56(4):1007-19.). These results highlight the role of riboflavin in supporting or maintaining the normal development of the mucosal barrier, including the gastrointestinal mucosa. Therefore, during the transition from breast milk to solid foods, Bifidobacterium longum transitional microbes can maintain intestinal epithelial integrity through riboflavin production, thereby promoting a healthy imprint of the immune system.

[0124] Folic acid (vitamin B9) is another essential vitamin for nucleic acid and protein synthesis, and folic acid deficiency has been associated with altered immune responses. Specifically, folic acid receptor 4 is highly expressed in regulatory T (Treg) cells, and folic acid is involved in maintaining Treg cell survival (Kunisawa et al., 2013, Front Immunol 4:189). Mice fed a folic acid-deficient diet had reduced numbers of intestinal Treg cells, leading to increased susceptibility to inflammation (Kunisawa et al., 2012; PLoS One 7(2): e32094). Tregs are essential for controlling pro-allergic TH2 cell responses and for producing anti-inflammatory IL-10. Transitional Bifidobacterium longum can influence tolerant immune responses via folic acid production.

[0125] The present invention also provides the use of prebiotics to increase riboflavin and / or folic acid levels in the gut of infants or children as described herein; wherein riboflavin and / or folic acid are increased due to the prebiotics promoting the growth and / or survival of Bifidobacterium longum transitional microorganisms in the gut of infants or children.

[0126] The present invention also provides a method for increasing riboflavin and / or folic acid levels in the gut of an infant or child; wherein the method comprises administering a prebiotic as described herein to the infant or child in order to promote the growth and / or survival of Bifidobacterium longum transitional microorganisms in the gut of the infant or child.

[0127] Bifidobacterium longum transitional microbes and / or prebiotics can promote immune tolerance in infants and / or young children. For example, Bifidobacterium longum transitional microbes and / or prebiotics can promote immune tolerance in the gut of infants and / or young children. Immune tolerance can be promoted through mechanisms as described herein, such as increased IL-10 production, decreased IL-5 production, and / or reduced permeability of the intestinal epithelial barrier.

[0128] prebiotics

[0129] The present invention also provides a prebiotic for use in treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children by promoting the growth of Bifidobacterium longum transitional microorganisms in the intestine of infants or young children, wherein the prebiotic is:

[0130] i. a polysaccharide substrate, wherein the polysaccharide substrate is suitably selected from any group listed in Tables 1 to 3; and / or

[0131] ii. Human milk oligosaccharides (HMOs).

[0132] Preferably, the HMO is 3'-O-fucosylated lactose (3'-FL).

[0133] On the other hand, the present invention provides a prebiotic for use in treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children by promoting the growth of Bifidobacterium longum transitional microorganisms in the intestine of infants or young children.

[0134] Polysaccharide substrate / carbohydrate-active enzyme (CAZyme)

[0135] The present invention provides a transitional strain of Bifidobacterium longum that encodes a specific spectrum of carbohydrate-active enzymes (CAZyme).

[0136] Carbohydrate active enzymes (CAZymes) are responsible for the synthesis and breakdown of glycoconjugates, oligosaccharides, and polysaccharides. They typically correspond to 1%–5% of the genes in living organisms. Glycoconjugates, oligosaccharides, and polysaccharides play important roles in many biological functions, such as serving as structural and energy storage components, and in many intracellular and intercellular events. The CAZyme classification is a sequence-based family classification system associated with the structure and molecular mechanisms of CAZymes (www.cazy.org).

[0137] CAZyme includes glycoside hydrolases (GH), glycosyltransferases (GT), polysaccharide lyases (PL), carbohydrate esterases (CE), and the carbohydrate binding module family (CBM).

[0138] GH catalyzes the hydrolysis of glycosidic bonds between two or more carbohydrates or between a carbohydrate and a non-carbohydrate moiety. In most cases, the hydrolysis of glycosidic bonds is catalyzed by two amino acid residues of the enzyme: the common acid (proton donor) and the nucleophile / base. Depending on the spatial position of these catalytic residues, the hydrolysis occurs via overall retention or overall inversion of the terminal isomer configuration.

[0139] The GH classification system is provided by the CAZy classification. In this paper, GH are divided into families (e.g., GH1, GH2, GH3, GH4, etc.) based on molecular function. These families are then further divided into subfamilies based on subgroups found within each family, which share a more recent ancestor and are generally more consistent in molecular function (e.g., GH13_1, GH13_2, GH13_3, GH13_4, etc.).

[0140] Suitablely, the present invention provides a transitional strain of *Bifidobacterium longum* encoding a glycosyl hydrolase family 43_17 (GH43_17) enzyme. GH43_17 contains the activities of both α-L-arabinofuranase (EC 3.2.1.55) and endonucleo-β-1,4-xylanase (EC 3.2.1.8), and has the ability to break down complexed carbohydrates such as arabinogalactan, arabinogalactan, and arabinoxylan. Suitablely, the GH43_17 gene contains the sequence of SEQ ID NO: 7 or has at least 60% sequence identity with SEQ ID NO: 7. Suitablely, the GH43_17 gene contains a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 7.

[0141] SEQ ID NO: 7

[0142] ATGAAACGAACTGACATCCACCTGCGCGATCCGTTCGTCCTGCCTCACGACGGTGTCTATTACCTGTATGGCACCCGCGCTGATAACGTGTGGGGCGCGATGGATGGTTTTGATTGCTACACCAGCCGCGACCTTGACAATTGGGAGGGTCCGTTCGAGGTGTTCCACAAGCCGGATGAATTCACGGCCGACCGTGCTTACTGGGCGCCCGAATGCTACGAGCGAGACGGTGTATTCCACCTGATTGCCACGCTCGGCGAGCCGGACGGGCGCAAAAGCGTGCACATGCTACGCGCTGATAGTCCGCTTGATCCGTTCGAATATGTCTGCCGGCTGACCGATCCGAATCAGTCCTGCATTGACGGAACTCTGCATGGTGAAGGTACCGATATGTGGCTTGTCTACTCGCATTCCTTGGAGGATGTGCCCGCCGGAGACATGGATGCCGTACGTCTGTCCTCCGACCTGACTCGGACGGTGGGGGAGAGCATGACATTGTTCCAGGCCTCGGATGCGCCGTGGGCGGTGCCGGTGCCGTTCGCGAAAGCGGAATTCGGCATCGACGAGGACGCCTACTTCTCCGATGGTCCCTGCCTGTGCAGGCTTTCCAACGGACGGCTGGCGATGCTGTGGTCGAGCTGGTCGACGGAAGGCGGATATGCAGTCGGCCAGGCCATCAGCGAATCAGGGTCGATTGCTGGGCCTTGGACGCAATGCCCCGAGCCTCTGCTTAGCCACGGCGGCCACGGCATGCTGTTCAACGGTCTCGATGGCGTGCTGCGTTACGCGGTCCACTCGCCCAACGACCCCGGCCAGGAACGGCCTACGTTTTTGTGCGTCGAAGAACAAGACGGGCTGCTGACGATTACGGAATAG

[0143] Suitablely, the GH43_17 gene may encode a protein as shown in SEQ ID NO: 8 or a sequence having at least 80% sequence identity with SEQ ID NO: 8. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 8.

[0144] SEQ ID NO: 8

[0145] MKRTDIHLRDPFVLPHDGVYYLYGTRADNVWGAMDGFDCYTSRDLDNWEGPFEVFHKPDEFTADRAYWAPECYERDGVFHLIATLGEPDGRKSVHMLRADSPLDPFEYVCRLTDPNQSCIDGTLHGEGTDMWLVYSHSLEDVPAG DMDAVRLSSDLTRTVGESMTLFQASDAPWAVPVPFAKAEFGIDEDAYFSDGPCLCRLSNGRLAMLWSSWSTEGGYAVGQAISESGSIAGPWTQCPEPPLLSHGGHGMLFNGLDGVLRYAVHSPNDPGQERPTFLCVEEQDGLLTITE

[0146] Suitably, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 43_22 (GH43_22) gene. Suitably, the GH43_22 gene comprises SEQ ID NO: 9 and / or SEQ ID NO: 10, or a sequence having at least 60% sequence identity with SEQ ID NO: 9 or SEQ ID NO: 10. Preferably, the *Bifidobacterium longum* transitional strain of the present invention comprises the GH43_22 gene having at least 60% sequence identity with SEQ ID NO: 9 and the GH43_22 gene having at least 60% sequence identity with SEQ ID NO: 10. Suitably, the GH43_22 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 9 or SEQ ID NO: 10.

[0147] SEQ ID NO: 9

[0148]

[0149] SEQ ID NO: 10

[0150]

[0151] Suitablely, the GH43_22 gene may encode the protein shown in SEQ ID NO: 11 or SEQ ID NO: 12, or a sequence having at least 80% sequence identity with SEQ ID NO: 11 or SEQ ID NO: 12. Preferably, the *Bifidobacterium longum* transitional strain comprises the GH43_22 gene encoding the protein shown in SEQ ID NO: 11 or a sequence having at least 80% sequence identity with SEQ ID NO: 11, and the GH43_22 gene encoding the protein shown in SEQ ID NO: 12 or a sequence having at least 80% sequence identity with SEQ ID NO: 12. Suitablely, the protein may comprise a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 11 or SEQ ID NO: 12.

[0152] SEQ ID NO: 11

[0153]

[0154] SEQ ID NO: 12

[0155]

[0156] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 43_27 (GH43_27) gene. Suitablely, the GH43_27 gene comprises the sequence of SEQ ID NO: 13 or has at least 60% sequence identity with SEQ ID NO: 13. Suitablely, the GH43_27 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 13.

[0157] SEQ ID NO: 13

[0158]

[0159] Suitablely, the GH43_27 gene may encode a protein as shown in SEQ ID NO: 14 or a sequence having at least 80% sequence identity with SEQ ID NO: 14. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 14.

[0160] SEQ ID NO: 14

[0161]

[0162] Suitably, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 43_29 (GH43_29) gene. Suitably, the GH43_29 gene comprises the sequence of SEQ ID NO: 15 or has at least 60% sequence identity with SEQ ID NO: 15. Suitably, the GH43_29 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 15.

[0163] SEQ ID NO: 15

[0164]

[0165] Suitablely, the GH43_29 gene may encode a protein as shown in SEQ ID NO: 16 or a sequence having at least 80% sequence identity with SEQ ID NO: 16. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 16.

[0166] SEQ ID NO: 16

[0167]

[0168] Suitably, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 121 (GH121) gene. Suitably, the GH121 gene comprises the sequence of SEQ ID NO: 17 or has at least 60% sequence identity with SEQ ID NO: 17. Suitably, the GH121 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 17.

[0169] SEQ ID NO: 17

[0170]

[0171] Suitablely, the GH121 gene may encode a protein as shown in SEQ ID NO: 18 or a sequence having at least 80% sequence identity with SEQ ID NO: 18. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 18.

[0172] SEQ ID NO: 18

[0173]

[0174] Appropriately, the transitional strain of Bifidobacterium longum contains the GH43_17 gene and one or more genes selected from the GH43_22, GH43_27, GH43_29 and GH121 genes as defined herein.

[0175] Appropriately, the transitional strain of Bifidobacterium longum includes the GH43_17, GH43_22, GH43_27, GH43_29 and GH121 genes as defined herein.

[0176] Suitablely, one or more of the arabinogalactan-degrading GH methods described herein include a signal peptide. A “signal peptide” can refer to a short amino acid sequence typically present at the N-terminus of a polypeptide that allows the polypeptide to be secreted extracellularly into sterile cells. Without being bound by theory, this can advantageously allow the present invention’s *Bifidobacterium longum* transitional strains (typically found in the diet when present at high molecular weight) to act as the primary degrader of the complex structure of arabinogalactan. Suitablely, a “primary degrader” can refer to bacteria capable of depolymerizing a specific polysaccharide into monosaccharides, disaccharides, and oligosaccharides that can absorb themselves and ferment themselves into acidic end products such as acetate or lactate. Suitablely, GH43_22 enzymes, GH43_27 enzymes, GH43_29 enzymes, GH_121 enzymes, GH43_24 enzymes, and / or GH30_5 enzymes can include a signal peptide. Appropriately, each of the enzymes GH43_22, GH43_27, GH43_29, GH_121, GH43_24, and GH30_5 may contain a signal peptide.

[0177] Suitable, the Bifidobacterium longum transitional strain of the present invention contains a gene encoding a glycosyl hydrolase family that targets arabinogalactan.

[0178] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 43_24 (GH43_24) gene. Suitablely, the GH43_24 gene comprises the sequence of SEQ ID NO: 19 or has at least 60% sequence identity with SEQ ID NO: 19. Suitablely, the GH43_24 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 19.

[0179] SEQ ID NO: 19

[0180]

[0181] Suitablely, the GH43_24 gene may encode a protein as shown in SEQ ID NO: 20 or a sequence having at least 80% sequence identity with SEQ ID NO: 20. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 20.

[0182] SEQ ID NO: 20

[0183]

[0184] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 127 (GH127) gene. Suitablely, the GH127 gene comprises the sequence of SEQ ID NO: 21 or a sequence having at least 60% sequence identity with SEQ ID NO: 21. Suitablely, the GH127 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 21.

[0185] SEQ ID NO: 21

[0186]

[0187] Suitablely, the GH127 gene may encode a protein as shown in SEQ ID NO: 22 or a sequence having at least 80% sequence identity with SEQ ID NO: 22. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 22.

[0188] SEQ ID NO: 22

[0189] MNVTITSPFWKRRRDQIVESVIPYQWGVMNDEIDTTVPDDPAGNQLADSKSHAVANLKVAAGELDDEFHGMVFQDSDVYKWLEEAAYALAYHPDPELKALCDRTVDLIARAQQPDGYLDTPYQIKSGVWADRPRFSLIQQSHEMYVMGHYIEAAVAYHQVTGNE QALEVAKKMADCLDANFGPEEGKIHGADGHPEIELALAKLYEETGEKRYLTLSQYLIDVRGQDPQFYTKQLKALNGDNIFPDLGFYKPTYFQAAEPVRDQQTADGHAVRVGYLCTGVAHVGRLLGDRGLIDTAKRFWTNIVARRMYVTGAIGSTHVGESFTYDYD LPNDTMYGETCASVAMSMFAQQMLDLEPKGEYADVLEKELFNGSIAGISLDGKQYYYVNALETTPDGLDNPDRHHVLSHRVDWFGCACCPANIARLIASVDRYIYTERDGGKTVLSHQFIANTAEFASGLTVEQRSNFPWDGHVEYTVSLPASATDSSVRFGLR IPGWSRGSYTLTVNGKPAVGSLEDGFVYLVVNAGDTLEIALELDMSVKFVRANSRVRSDAGQVAVMRGPLVYCAEQVDNPGDLWNYRLADGVTGADAAVAFQADLLGGVDTVDLPAVREHADEDDAPLYVDADEPRAGEPATLRLVPYYSWANREIGEMRVFQRR

[0190] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 30_5 (GH30_5) gene. Suitablely, the GH30_5 gene comprises the sequence of SEQ ID NO: 23 or has at least 60% sequence identity with SEQ ID NO: 23. Suitablely, the GH30_5 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 23.

[0191] SEQ ID NO: 23

[0192]

[0193] Suitablely, the GH30_5 gene may encode a protein as shown in SEQ ID NO: 24 or a sequence having at least 80% sequence identity with SEQ ID NO: 24. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 24.

[0194] SEQ ID NO: 24

[0195]

[0196] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 43_32 (GH42_32) gene. Suitablely, the GH42_32 gene comprises the sequence of SEQ ID NO: 25 or has at least 60% sequence identity with SEQ ID NO: 25. Suitablely, the GH42_32 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 25.

[0197] SEQ ID NO: 25

[0198]

[0199] Suitablely, the GH42_32 gene may encode a protein as shown in SEQ ID NO: 26 or a sequence having at least 80% sequence identity with SEQ ID NO: 26. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 26.

[0200] SEQ ID NO: 26

[0201] MTATISNGVSASYSPAEDELGAADPTALLAESGDLKPLAERTYTNPVPYADGKSHTAPDPFVLKYRDLYYCYATDEHGILVSTSPDMVHWTSHGFCYTEAGRRNFWAPSVILINGVFHMYFSNMPAEETDTHTEIMRVAVSEDPLGPFEKKAELFNTFAIDSQVVYGDDGQLYLLYAD NQVTGLSDDRRPGTSVMIDRLVTPYSRENKPRPLIVPTMDEEIFARNRFGDGRDWHTVEGATYFAYRDRAFITYSANAYEHEDYFVGYSYAQLPNKQADAHIDQLDWTKQLNENRFDPLLIRSPKVEGTGHNSIVKAPNAVDDWIVYHGRNADDELYVGTEQRVMRIDPLYYAEGGLDT PGPTAAAQSAPLYGTVHDDFADGLNAGWSVISGAAHTESDVDGHALVADESSVFIAVSGKSSATQVIDVWAKAPVTPLGARFGIVVRYQDANNLTKLEVDAGRQVISVVDVIGGVASERVTNADLHDFDSHAWHEYRLERRYCRLEIRIDGRFAASCTISDKPGRAGLFSLRTGAAFS AYAATEHVNLWGAGLRDLGRELHADRRLVIDGGVRSSGVCPVTLELAYPLVSNRFVLDFAGQTSRGQALLSLGEYRLSGTASSVEFMRNGKSLPSTPEPARLRVFEDNVRRDRSGRAVLTIRIEALNGTMRLHLRGKTWQVPFADNAARARITLDRASLTGYERTSLESSIEERSASGN

[0202] Appropriately, the transitional strain of Bifidobacterium longum contains one or more genes selected from the GH43_24 gene, GH127 gene, GH30_5 gene and GH43_32 gene as defined herein.

[0203] Appropriately, the transitional strain of Bifidobacterium longum contains the GH43_17 gene and one or more genes selected from the GH43_24, GH127, GH30_5 and GH43_32 genes as defined herein.

[0204] Appropriately, transitional strains of Bifidobacterium longum contain the GH43_17, GH43_24, GH127, GH30_5 and GH43_32 genes as defined herein.

[0205] Appropriately, the transitional strain of Bifidobacterium longum includes the GH43_17, GH43_22, GH43_27, GH43_29, GH121, GH43_24, GH127, GH30_5 and GH43_32 genes as defined herein.

[0206] Appropriately, transitional strains of Bifidobacterium longum include GH43_17, GH43_22, GH43_27, GH43_29, GH121, GH43_24, GH127, GH30_5, and GH43_32 as defined herein.

[0207] GH43_17 gene cluster

[0208] Appropriately, transitional strains of Bifidobacterium longum may contain one or more genes encoding family 31 glucosidase (GH31), ABC transporter, Lac-I type regulator, MFS transporter and / or AraC family transcription regulator.

[0209] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises the glycosyl hydrolase family 31 (GH31) gene. Suitablely, the GH31 gene comprises the sequence of SEQ ID NO: 27 or has at least 60% sequence identity with SEQ ID NO: 27. Suitablely, the GH31 gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 27.

[0210] SEQ ID NO: 27

[0211]

[0212] Suitablely, the GH31 gene may encode a protein as shown in SEQ ID NO: 28 or a sequence having at least 80% sequence identity with SEQ ID NO: 28. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 28.

[0213] SEQ ID NO: 28

[0214] MTTSFTIDGNALIWTGDGETLRIEPWEENSVRVRATRNRGFGPVDWALLEPKNESGRVADIAVGEDGEHASLTNGSITVKADSNHAPLLSAGYETFRCDLSFWNAEGELLFREYPQGGSLLLKARDYTPVSGESFAVTTSFSADPKERLYGMGEYQQDVLDLKGSTFELAH RNSQASVPFVVSSKGYGFLWHNPAIGRATFGRNRTEWAAQSTDQIDYWVTAGDSYAQIESQYADATGHAPVMPEWGMGFWQCKLRYWNQEQLLDVARGFKSRNIPLDLIVIDFFHWPHLGDYKFEDEFWPDPEAMVAELNSMGVKLMVSVWPQVSVSSENFVEMKRNNYLVS AEAGLNLDMMFEEPCVNYDPTNPGARKFVWDKCKANYWDKGVRAFWLDEAEPEYGVYDFRNYRYHMGSDLNVGNVYPQAYNRGFYEGQIEAGMEGEIVNLTRCAWAGSQRYGSLVWSGDVGSTFADLKSQITCAIHMGMAGIPWFTTDMGGFHDGVIDSDSFKELLARWCAF SCFLPVMRNHGDRSLGESTGKQTITKATGEHRSPSGADNEPWSYGPEMESIFRKYIAVREVMRPYTRELFQSAHEQGQPLVRGLFYEFPTDEHVADIADEYLYGPDILVAPVVEAGAASRSVYLPGDETTTWTDLRDGAVYAGGQSIESSAAIDTVPAFARDGRDHGLIGLL

[0215] Suitably, the *Bifidobacterium longum* transitional strain of the present invention comprises one or more ABC transporter genes. Suitably, the ABC transporter gene comprises SEQ ID NO: 29 to SEQ ID NO: 31, or a sequence having at least 60% sequence identity with SEQ ID NO: 29 to SEQ ID NO: 31. Suitably, the ABC transporter gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 29 to SEQ ID NO: 31. Suitably, the *Bifidobacterium longum* transitional strain of the present invention comprises a gene having at least 60% sequence identity with SEQ ID NO: 29, a gene having at least 60% sequence identity with SEQ ID NO: 30, and a gene having at least 60% sequence identity with SEQ ID NO: 31.

[0216] SEQ ID NO: 29

[0217] ATGACGCATCGTAGCACCTGGTGGAAAACCGCTCTCGGCATCATATTGACGCTCATCATGATGTTTCCTGTCTACTGGATGATCAACATCTCGTTCACTGGTAAGGCATCCATTCGTTCCGGCGACCTGTGGCCCAAGGATTTCACCTTTGACAACTACGCCCGCGTAATCGCCGACCAAATGCCCTATCTGGGCACTTCCATCCTCGTAGCGGTATGCTGCGTGATTCTAACGCTGGTCATCGCACTGCCTGCCGCCTACGCACTGGCTTTGCTGCGCTGTCCAGGCAGCGGCGCGCTCAGCTTCCTGCTCATCGTGGCTCAGATGATTCCCGCCGTCGTGATGTCGCTCGGCTTCTACGAGATTTATAACAACATTGGTCTGCTCGATACGTTGCCCGGCCTGATCCTCGCCGACTCGACCATTGCGGTGCCGTTCGCGGTCATGCTCCTGACTTCTTTCATGGCCGGCATCCCGCGGTCCCTGCTTGAGGCCGCCGAAGTGGATGGAGCCTCACGTACCCGTCGCTTCTTTTCCATTGTCATCCCGTTATCGCGCAATTCGATCGTGACCGTCTCCCTGTTCGCTTTCCTATGGTCTTGGAGCGACTTCCTGTTCGCTTCCACCCTTGACTCCGGCGGCGGCAAGATGCGCCCGATCACTATGGGTCTGTACAACTATATCGGTGCGCAGACCCAGGAATGGGGGCCGATGATGGCCACCGCAGTGCTTGCATCCATTCCCGCGACCATCCTGCTTGTCTTCGCCCAGAAGTACGTCGCCGCAGGCGTGACCGCCGGTGCTGTTAAGGACTAA

[0218] SEQ ID NO: 30

[0219] ATGACAGCCTCAACAACAAGCCCCGTTCGCCGGGCAAAGTCCGGCACTCCGGTCCGGGCCAAACTGGCCATCGCCGGATTCATTGCCCCACTGATTATCTACTTGGTAATCTTTTACGCGTTCCCGCTCATCCAGAACGTGTCAATGAGCCTGCACCGATACACGCGACGAACCTTCGTTACCGGAGATGCGCTGTTCGTGGGTCTCGACATCTACAAGGAAGTCATTTCCTCCGTGGAGTTCTGGCCGGTTGTGGGGCAGACCTTCGTGTTCGTGGTCGTCTCGCTGATATTCCAATATGTAATCGGCTTGGCCCTGGCGGTGTTCTTCAACGATAACTTCAAGCTCTCTGGTGTGCTGCGCGGCATCATGCTGGTTCCGTGGCTGTTGCCGCTGATTGTTTCTGGAACCGTCTGGCAGTGGATGATGGACCCTGACTCCGGCATCCTCAACATGTTCCTCGGTCTGTTTGACATCGAACCCATCTGGTGGCTCCAGGCGGATAACTCGCTGTGGGCCGTCATCATCGCCAACATCTGGCTGGGAATCCCCTTCAACCTCGTGATCCTGTATTCCGGCCTACAGAACATCAGCGGCGACCTGTATGAAGCCGCCTCCCTCGATGGCTGCAACGCCTGGCAGCGCTTCTGGAAGATCACCTTCCCTCTCCTGAAGCCCGTCACTTCGATCACCCTGTTGCTCGGCTTCGTCTATACATTGAAGGTCGTTGACGTGATCTGGATGATGTCCCAGGGAACCGGCACCTCGCGTACCCTCGCCACCTGGGCCTATTCGATGGCATTTGGCAAGGGAACTTCAATGACTATCAAATACTCGGAGGCTTCGGTGCTCGGCACGATTCTCATCATCGTGGCGTTGATTTTCGGACTGATTTACCTGCGGGTCCAGAAGACCCAGGAAACCTGCTAA

[0220] SEQ ID NO: 31

[0221]

[0222] Suitablely, the ABC transporter gene may encode a protein as shown in SEQ ID NO: 32 to SEQ ID NO: 34, or a polypeptide having at least 80% sequence identity with SEQ ID NO: 32 to SEQ ID NO: 34. Suitablely, the gene may encode a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 32. Suitablely, the gene may encode a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 33. Suitablely, the gene may encode a polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 34.

[0223] SEQ ID NO: 32

[0224] MTHRSTWWKTALGIILTLIMMFPVYWMINISFTGKASIRSGDLWPKDFFTFDNYARVIADQMPYLGTSILVACCVILTLVIALPAAYALALLRCPGSGALSFLLIVAQMIPAVVMSLGFYEIYNNIGLLDTLPGL ILADSTIAVPFAVMLLTSFMAGIPRSLLEAAEVDGASRTRRFFSIVIPLSRNSIVTVSLFAFLWSWSDFLFASTLDSGGGKMRPITMGLYNYIGAQTQEWGPMMATAVLASIPATILLVFAQKYVAAGVTAGAVKD

[0225] SEQ ID NO: 33

[0226] MTASTTSPVRRAKSGTPVRAKLAIAGFIAPLIIYLVIFYAFPLIQNVSMSLHRYTRRTFVTGDALFVGLDIYKEVISSVEFWPVVGQTFVFVVVSLIFQYVIGLALAVFFNDNFKLSGVLRGIMLVPWLLPLIVSGTVWQWMMDPDSGILNMFL GLFDIEPIWWLQADNSLWAVIIANIWLGIPFNLVILYSGLQNISGDLYEAASLDGCNAWQRFWKITFPLLKPVTSITLLLGFVYTLKVVDVIWMMSQGTGTSRTLATWAYSMAFGKGTSMTIKYSEASVLGTILIIVALIFGLIYLRVQKTQETC

[0227] SEQ ID NO: 34

[0228] MKSNTALKITAALCSCAMLVGVSACGSNSTTDDKVIEWWDDWTRHEDGSEFDKLVKACAPEGYTIERQAIATSDLLNNLTTAIKEDNGPDVAVIDNPMIPSAVDAGLVAGSDETGLDVSAWDENLEAPGVVDGQAYGVPLGGSNTLGLMYNPTIIEAAGVDVSTITDWDSLNAAIKKVVDAGYKGITFSGISGEE GVFQFLPWFWGAGGDLSKLDSQAQKDAEDLLSGWISKGWAPKSATTNTQSASWDLFLAGDYGFAEIGTWMQSEADEAGAKLIPIPAKDGGVATVPTGGEFAMVAYHKKDAESHYKLANQVIECLSEDETLLKVSNALSNLAAKKAVRAEQLAASDGLAQWKESIENAAGRTSDLGLKYEEASASISESLLAALNAA

[0229] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises a Lac-I type regulatory gene. Suitablely, the Lac-I type regulatory gene comprises a sequence of SEQ ID NO: 35 or having at least 60% sequence identity with SEQ ID NO: 35. Suitablely, the Lac-I type regulatory gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 35.

[0230] SEQ ID NO: 35

[0231]

[0232] Suitable, the Lac-I type regulatory gene may encode a protein as shown in SEQ ID NO: 36 or a sequence having at least 80% sequence identity with SEQ ID NO: 36. Suitable, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 36.

[0233] SEQ ID NO: 36

[0234] MVTINDVAREAGVSKTTVSFVLSGSRPVAAATEQRIREAMDRLGYTVNHAARSLSTSKTMTIAVVTSNRQDAYFDIARGTYINGLSRAAAETGYDMLITNDPDGSATENACQSHKADGLVFLDVRQNDPRVPIAAESGIPTVSLGVPVNPMNLDVVDTDFTDMAAST MRTLHDAGHRRVSVITLSSRVIAEQLNDTARFLREIERSGERLGMHATIRHCSTRPGIIDTDIARILDGRGEDTAFVIHNESAVLVFRRAVEHRGLRIPEDISVIAINEKQMSDALYLPYSAYENDVELVTQSAVNTLVDRIEHPELTPRTLIKASYIDRDSVANI

[0235] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention comprises an facilitater superfamily (MFS) gene. Suitablely, the MFS gene comprises a sequence of SEQ ID NO: 37 or having at least 60% sequence identity with SEQ ID NO: 37. Suitablely, the MFS gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 37.

[0236] SEQ ID NO: 37

[0237]

[0238] Suitablely, the MFS gene may encode a protein as shown in SEQ ID NO: 38 or a sequence having at least 80% sequence identity with SEQ ID NO: 38. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 38.

[0239] SEQ ID NO: 38

[0240] MAEFHYAIGHFHCAGHRIGCSGIPGQLALQRADDCFGITLTALVGAWLTGKLASILSRKTVALIGAGGMLLFGLLPYFVHSSLAAVIAFSALMGVCLGFINNVLPTLISVHYEGDERQSIMGQQVAVASIGAMVFMTVAGKLATAQWYHAYLIYLFAAVVLVVCAFTLPTKNGETDEAGRIQGTGPSASIR EVMTGKLWFLVVAGFFFLLANNAYSNNLSLLVEQRGLGDAGTAGLISTIGQFGGLLAGLCVGLMVRFVKNHLLMVGFIVEGLSLLLLGCSASLPLLIIGSFFAGAGLSIYYAQAPFLVTVIEKPYLIPLGIAAMTTANALGGFASPVLVNAINGLFGSHAAGAMFIGAAIALAGAVALGVSGFQKKCLESAK

[0241] Suitablely, the *Bifidobacterium longum* transitional strain of the present invention contains an AraC family transcriptional regulatory gene. Suitablely, the AraC gene contains a sequence of SEQ ID NO: 39 or a sequence having at least 60% sequence identity with SEQ ID NO: 39. Suitablely, the AraC gene contains a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 39.

[0242] SEQ ID NO: 39

[0243] ATGGAGCGCGATGCTTTCCGGCTGCCGGGCCTCACCGCCGGCGATGACAACCAGTATGCCGATCACACGCTCACCGGCATGGCAGCCGATGCGGCGAACGTCATAGCCGCAGGCGGTCCCGCCCCGCTGACTAGCTTCGGCACTGTCGCTCAAGCCGCCCATCTCAATCCAGATGACGGCTTCGGCATCATTGGCCATGATCTTGCACACCCATCGCACCTACACCGGCATGACTATATGGAAATCACGCACGCCATCGCCGGTACGGTACTGGTCTGGGTCGAAGGAGAGACCAACGTGCTGACACAGGGCGGCACCATACTCATCAAGCCTGGAGCCCGTCATCTCATCTCCCCCATCATCGAATACGGGCAAACACCACACGAGGCGGACATCCTGATTAAACCCGAGCTCATCAGGCAATGCCGCATTCCGATTCTGGAAGCAGCCGGCGCCGACCGGATGTTCATTAGCTGGCTTGACGATGACCGGCAGACCCACTGCCTGCTGGCAGCCGGCAAGCACCACGCCGGCGAGGCCGCTATCAGCCGCATGTTCATCGCCTACTGCATCAACGCAACCTACAGGCCAGACTTCACCGTCATCGGCAACCTGCTCGAGCTGTTCCACGAAACGTCCCGAGTCTTGGAACACCAGCCACGTACCGATCCGCTGATCGCCGCCATCATCGAAACCATCACGGCAGATCCCGCCACGGCCCACAACCAGGCCATAGCGGACACACTTGGATACAGCGTGGGATATCTGTCCCGGTACGCGCGCAAGCACAGCGGGCACACACTCGGCCAACTCATCAACGAGGAAAGGCTCCGACTCGGCGCCGAACTGCTCGTCACCACCGACGACACCATTGCCGAAATCACCCGAACCATTGGCTACGAAAGTCCAGCCTATTTCCATAAACTCTTCCGCAGCCGCTACCTCATTACCCCCGACCGCTACCGCAACGACTTCCGTATCGCATTACGTTGCGGATGA

[0244] Suitablely, the AraC gene may encode a protein as shown in SEQ ID NO: 40 or a sequence having at least 80% sequence identity with SEQ ID NO: 40. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 40.

[0245] SEQ ID NO: 40

[0246] MERDAFRLPLGLTAGDDNQYADHTLTGMAADAANVIAAGGPAPLTSFGTVAQAAHLNPDDGFGIIGHDLAHPSHLHRHDYMEITHAIAGTVLVWVEGETNVLTQGGTILIKPGARHLISPIIEYGQTPHEADILIKPELIRQCRIPILEAAGADRMFISWLDDDRQT HCLLAAGKHHAGEAAISRMFIAYCINATYRPDFTVIGNLLELFHETSRVLEHQPRTDPLIAAIIETITADPATAHNQAIADTLGYSVGYLSRYARKHSGHTLGQLINEERLRLGAELLVTDDTIAEITRTIGYESPAYFHKLFRSRYLITPDRYRNDFRIALRCG

[0247] Appropriately, the transitional strain of Bifidobacterium longum contains the MFS transporter and AraC family transcriptional regulatory genes.

[0248] Appropriately, the transitional strain of *Bifidobacterium longum* contains GH43_17, the MFS transporter, and AraC family transcriptional regulatory genes. Appropriately, GH43_17, the MFS transporter, and AraC family transcriptional regulatory genes are contained within a gene cluster.

[0249] As used in this article, a "gene cluster" can refer to a group of genes located adjacent to each other on a chromosome.

[0250] Appropriately, the transitional strain of Bifidobacterium longum contains each of the genes for GH31, ABC transporter, Lac-I type regulator, MFS transporter, and / or AraC family transcription regulator.

[0251] Appropriately, the transitional strain of *Bifidobacterium longum* contains GH43_17, MFS transporter, AraC, GH31, ABC transporter, and Lac-I type regulatory gene. Appropriately, GH43_17, MFS transporter, AraC family transcription regulators, GH31, ABC transporter, and Lac-I type regulatory gene are included in the aa gene cluster as described above.

[0252] Appropriately, the Bifidobacterium longum transitional strain also contains xylulokinase genes and / or xylose isomerase genes. Appropriately, xylulokinase genes and / or xylose isomerase genes are included in the gene clusters defined above.

[0253] Suitablely, the xylulokinase gene comprises SEQ ID NO: 41 or a sequence having at least 60% sequence identity with SEQ ID NO: 41. Suitablely, the xylulokinase gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 41.

[0254] SEQ ID NO: 41

[0255]

[0256] Suitablely, the xylulokine gene may encode a protein as shown in SEQ ID NO: 42 or a sequence having at least 80% sequence identity with SEQ ID NO: 42. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 42.

[0257] SEQ ID NO: 42

[0258] MTRVLVAGVDTSTQSTKVRITDAATGEQVRFGQAKHPDGTSVNPEFWWEAFTKAAEQAGGLDDVAALAVGGQQHGMVILDKQGNVIRDAMLWNDTSSAPQAAALIDKLGATPAEGDEPDDVTARGK QRWVKAVGSSPVASYTLTKVAWVAENEPENAKKIAAVCLPHDWLSWRIAGYGPVAEGEDAHLEALFTDRSDASGTIYYDAAHDEYRRDLIAMVLTPAEGEEAAKAHADAIVLPTVLGPHEAAAVKAD PAIAGKDVEGGCIIGPGGGDNAMASLGLGMAVGDVSVSLGTSGVAAAIAENPVYDLTGAISGFADCTGHYLPLACTINGSRILDAGRAALGVDYDELAELAFKAEPGAGGITLVPYFDGERTPNRP DATASLTGLTLHNTTKENLARAFVEGLLCSQRDCLELIRSLGAEINRILLIGGGAKSVAIRTLAPSILGMDVTRPATDEYVAIGAARQAAWVLSGEAEPLTWQLTIEGVETGEPTEAVYEAYAKARG

[0259] Suitablely, the xylose isomerase gene comprises SEQ ID NO: 43 or a sequence having at least 60% sequence identity with SEQ ID NO: 43. Suitablely, the xylose isomerase gene comprises a sequence having at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 43.

[0260] SEQ ID NO: 43

[0261]

[0262] Suitablely, the xylulose isomerase gene may encode a protein as shown in SEQ ID NO: 44 or a sequence having at least 80% sequence identity with SEQ ID NO: 44. Suitablely, the protein may contain a sequence having at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% sequence identity with SEQ ID NO: 44.

[0263] SEQ ID NO: 44

[0264] MGLWDVDKIEYVGRAKGPKEDFAFHYYDADKVVAGKKMKDWLRFGVAWWHTFNQELVDPFGTGTAHRPYYKYTDPMDQALAKVDYAFELFQKLGVEYFCFHDRDIAPEGDTL RETNANLDKVVDKIDENMKSTGVKLLWNTSSLFTNPRFVSGAATSPFADIYAYAGGQLKKSLEIGKRLGAENYVFWGGREGYENLWNTEMKRETDHIAKFFHMCADYAKEIG FEAQFLIEPKPKEPTLHQYDFDAATAIEEFLRNHDLTDVFKLNLEGNHANLAGHTYQHEIRVARESGFLGSLDANQGDKLIGWDMDEFPTDLYETVAVMWEVLQAGSIGPHGG LNFDAKPRRTSFYEEDLFRSHIAGMDAYAAGLLVADKMNQDGFIQNLQAERYSSYDSGIGKDIDEGNVTLADLEAYSLDKPQSELIAATKSDHLESVKATINNYIIDALAEVE

[0265] Table 1 provides detailed information on CAZymes specific to transitional strains of *Bifidobacterium longum* (i.e., those not coded by *Bifidobacterium longum suis / suillum*, *Bifidobacterium longum longum*, or *Bifidobacterium longum infantis* strains). Table 1 also provides a summary of the glycan substrates metabolized by each CAZyme and exemplary dietary fiber sources / components.

[0266] Table 1

[0267]

[0268] Table 2 lists strains of *Bifidobacterium longum* that are present in at least one transitional strain of *Bifidobacterium longum* but not in strains selected from... Figure 4 and Figure 5 Detailed information on CAZyme in at least one of the groups of *Bifidobacterium longum* subsp. suis, *Bifidobacterium longum* subsp. spp., or *Bifidobacterium longum* subsp. spp. infantis strains is shown. Table 2 also provides a summary of the polysaccharide substrates metabolized by each CAZyme and exemplary dietary fiber sources / components.

[0269] Table 2

[0270]

[0271]

[0272]

[0273]

[0274] Table 3 provides detailed information on CAZymes present in all analyzed Bifidobacterium longum strains (i.e., transitional Bifidobacterium longum, Bifidobacterium longum suis subsp., Bifidobacterium longum subsp. longum, and Bifidobacterium longum infantis subsp.). Table 3 also provides a summary of the glycan substrates metabolized by each CAZyme and exemplary dietary fiber sources / components.

[0275] Table 3

[0276]

[0277]

[0278]

[0279] Table 4 provides detailed information on CAZymes that are not encoded by transitional strains of Bifidobacterium longum but are encoded by one or more of the following: Bifidobacterium longum suis subsp., Bifidobacterium longum subsp. longum, and Bifidobacterium longum infantis subsp.

[0280] Table 4

[0281]

[0282] The representative sequences of CAZyme listed in Tables 1 to 4 are shown in... Figure 9 In appropriate context, the CAZyme referred to in any of Tables 1 through 4 may include... Figure 9 The corresponding sequence shown in the diagram or composed of it. Appropriately, CAZyme may include... Figure 9The variants of the corresponding sequences shown, or those constituting them, retain at least one function of the corresponding CAZyme listed in Tables 1 to 4. Suitablely, the variants may provide each functional activity of the corresponding CAZyme listed in Tables 1 to 4. Suitablely, the variants may include... Figure 9 The sequences listed herein have at least 70% sequence identity with amino acid sequences or are composed of such sequences, and retain at least one functional activity of the corresponding CAZyme listed in Tables 1 to 4, preferably each functional activity. Suitable variants may include amino acid sequences with... Figure 9 The corresponding sequences listed herein have or consist of amino acid sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity. Variants retain at least one functional activity of the corresponding CAZyme listed in Tables 1 to 4, preferably each functional activity.

[0283] Suitable prebiotics used in this invention may include polysaccharide substrates selected from any of the groups listed in Tables 1 to 3.

[0284] Suitable prebiotics used in this invention may include combinations of glycan substrates selected from any of the groups listed in Tables 1 to 3.

[0285] The combination of glycan substrates may include at least 2, at least 4, at least 10, at least 20, at least 30, at least 40, or at least 50 glycan substrates selected from the groups listed in Tables 1 to 3. The combination may include each of the glycan substrates listed in Tables 1 to 3.

[0286] Appropriately, prebiotics may include one or more polysaccharide substrates selected from the groups listed in Table 1 or Table 2.

[0287] Prebiotics may include at least 2, at least 4, at least 10, at least 20, or at least 30 glycan substrates listed in Tables 1 and 2. Prebiotics may include each of the glycan substrates listed in Tables 1 and 2.

[0288] Appropriately, the polysaccharide substrate may include or consist of pectin, arabinogalactan, and / or starch.

[0289] Appropriately, the polysaccharide substrate may include or consist of pectin.

[0290] Appropriately, the polysaccharide substrate may include or consist of arabinogalactan.

[0291] Appropriately, the polysaccharide substrate may include or consist of starch.

[0292] Appropriately, the polysaccharide substrate is provided in the form of dietary fiber. For example, dietary fiber can be prebiotic fiber.

[0293] Where appropriate, the polysaccharide substrate may be included in the ingredients, such as dietary ingredients.

[0294] This ingredient contains one or more polysaccharide substrates, optionally from the group consisting of: purified polysaccharides or purified oligosaccharides, dietary fiber components, semi-purified food components, raw food components, food additives, HMOs, and semi-purified or purified peptidoglycans.

[0295] Semi-purified food ingredients can be fruit, vegetable, or grain extracts.

[0296] Raw food ingredients can be fruits, vegetables, grains, seaweed, or microalgae.

[0297] Food additives can be guar gum or gum arabic.

[0298] Appropriately, peptidoglycan can be GAG.

[0299] Appropriately, the polysaccharide substrate may be included in the purified cellulose.

[0300] Tables 1 through 3 provide exemplary components and / or purified fibers that include suitable glycan substrates. In particular, dietary fibers and / or components that include a given glycan substrate are identified in the same row as the glycan substrate.

[0301] Pectin can be included in fruit or vegetable pectin. Therefore, suitable ingredients that include pectin include, but are not limited to: fruits (e.g., apples, pears), vegetables, legumes (peas), and roots (e.g., sugar beets). Suitable purified fibers that include arabinogalactan include peach pectin. Pectin extracted from sugar beets suitably contains arabinogalactan, galactan, and arabinogalactan, and can be provided as an ingredient.

[0302] Arabinogalactan can be included in fruit or vegetable pectin. Typical suitable ingredients that include arabinogalactan include, but are not limited to, dietary fiber from fruits, vegetables, whole grains, and seaweed. Suitable purified fibers that include arabinogalactan include peach pectin, larch wood arabinogalactan, and gum arabic. Suitablely, arabinogalactan can be provided in larch wood arabinogalactan.

[0303] Starch can be included in resistant starch from grains (whole grains), legumes, vegetables (e.g., corn), and roots (e.g., potatoes). Suitable, illustrative, starch-containing ingredients include, but are not limited to, corn. Suitable purified fibers including starch include high amylose and resistant dextrin. Starch may suitably be provided in potatoes, corn, or other ingredients. Starch may suitably be included in potato ingredients.

[0304] Human milk oligosaccharides (HMOs)

[0305] Appropriately, prebiotics include HMOs.

[0306] Suitablely, HMOs can be metabolized by *Bifidobacterium longum* transitional microorganisms. Suitablely, HMOs may be able to promote the growth and / or survival of *Bifidobacterium longum* transitional strains. HMOs that can promote the growth and / or survival of *Bifidobacterium longum* transitional strains can be determined, for example, by an anaerobic culture of the *Bifidobacterium longum* transitional strain and the HMO to be tested. The growth and / or survival of the *Bifidobacterium longum* transitional strain can be determined by measuring the number of bacterial cells, cell density (e.g., by optical density measurement), and / or the abundance of 16S rDNA, for example using PCR methods. An exemplary assay for measuring the growth of *Bifidobacterium longum* transitional strains in the presence of HMOs is provided in Example 6. HMOs that can promote the growth and / or survival of *Bifidobacterium longum* transitional strains can increase the number of *Bifidobacterium longum* transitional bacteria in an anaerobic culture by at least 20%, at least 30%, at least 40%, at least 50%, at least 75%, or at least 100% compared to the number of *Bifidobacterium longum* transitional bacteria in a control anaerobic culture that does not contain HMOs. Appropriately, HMOs that promote the growth and / or survival of Bifidobacterium longum transitional bacteria can increase the number of Bifidobacterium longum transitional bacteria in anaerobic cultures by a statistically significant amount (e.g., p-value < 0.05 as determined by one-way ANOVA) compared to the number of Bifidobacterium longum transitional bacteria in control anaerobic cultures that do not contain HMOs.

[0307] HMOs can be fucosylated oligosaccharides (i.e., oligosaccharides with fucose residues; for example, 2'-fucosylvose (2'-FL), 3-fucosylvose (3-FL), difucosylvose (DiFL), lactose-N-fucopentose (e.g., lactose-N-fucopentose I, lactose-N-fucopentose II, lactose-N-fucopentose III, lactose-N-fucopentose V), lactose-N-fucohexasaccharide, lactose-N-difucohexasaccharide I, fucosylvose-N-hexasaccharide, fucosylvose-N-neohexose, difucosylvose-N-hexasaccharide I, difucosylvose-N-neohexose II and any combination thereof), N-acetylated oligosaccharides (e.g.) Such as LNT (lactose-N-tetrasaccharide), para-lactose-N-neohexose (para-LNnH), LNnT (lactose-N-neohexose), DSLNT (disialyllactose-N-tetrasaccharide), lactose-N-hexasaccharide, lactose-N-neohexose, para-lactose-N-hexasaccharide, para-lactose-N-neohexose, lactose-N-octasaccharide, lactose-N-neohexose, isolaxose-N-octasaccharide, para-lactose-N-octasaccharide and lactose-N-decasaccharide and any combination thereof) and / or sialylated oligosaccharides (e.g. 3'-sialyllactose (3-SL), 6'-sialyllactose (6-SL) or Lst (sialyllactose-N-tetrasaccharide), Lst-a, Lst-b or Lst-c)).

[0308] Prebiotics may contain 3'-O-fucosylated lactose (3FL).

[0309] In some implementation schemes, prebiotics include

[0310] -12% to 38% by weight, preferably 17% to 31% by weight of 3-FL.

[0311] The prebiotic may contain 3-FL between 0.01 g / L and 7 g / L, preferably between 0.025 g / L and 6 g / L, and more preferably between 0.05 g / L and 5 g / L.

[0312] Appropriately, the 3'-O-fucosylated lactose (3'FL) contained in prebiotics promotes the growth of Bifidobacterium longum transitional microorganisms that preferentially utilize 3-fucosylated lactose (3-FL).

[0313] Combination of Bifidobacterium longum transitional microorganisms and prebiotics

[0314] The present invention also provides a combination of a prebiotic and a Bifidobacterium longum transitional microorganism for the stated purpose according to the present invention.

[0315] Bifidobacterium longum transitional microorganisms and prebiotics can be applied alone, simultaneously, or sequentially.

[0316] Appropriately, Bifidobacterium longum transitional microorganisms and prebiotics can be applied in combination.

[0317] Appropriately, the combination of Bifidobacterium longum transitional microorganisms and prebiotics can be referred to as a "symbiotic preparation".

[0318] In aspects of the invention that use a combination of Bifidobacterium longum transitional microorganisms and prebiotics (e.g., glycan substrates), each aspect may be selected such that the Bifidobacterium longum transitional microorganism is capable of metabolizing the glycan substrate provided in the combination. Such selection may be made, for example, by selecting a Bifidobacterium longum transitional microorganism encoding CAZyme as a glycan substrate (or selecting an ingredient that includes said glycan substrate) from the same row of Tables 1 through 3.

[0319] The combinations of the present invention are not limited to requiring the Bifidobacterium longum transitional microorganism to be able to metabolize the glycan substrate provided in the combination. Therefore, the present invention includes any combination of the Bifidobacterium longum transitional microorganism and glycan substrate disclosed herein.

[0320] Suitable, the composition comprises one or more polysaccharide substrates as described herein.

[0321] Suitable, the composition comprises a Bifidobacterium longum transitional microorganism that preferentially utilizes 3-fucosyllactose (3-FL). On a dry weight basis, the composition may contain between 10 [units of something] per g of composition. 3 cfu to 10 12 Probiotic strains between CFU, more preferably between 10 7 CFU and 10 12 CFU, such as between 10 8 CFU and 10 10 The probiotic strain is mixed with 3-FL in an amount between 0.01 g / L and 7 g / L, preferably between 0.025 g / L and 6 g / L, and more preferably between 0.05 g / L and 5 g / L.

[0322] Composition

[0323] The Bifidobacterium longum transitional microorganism, prebiotic, or symbiotic preparation used in this invention can be provided in the form of a composition.

[0324] The composition can be suitably administered to an individual, such as an infant or toddler, in any suitable form, such as a dosage unit (e.g., tablets, capsules, powder sachets, etc.). The composition can be in powder, semi-liquid, or liquid form. The composition can be added to nutritional compositions, infant formula, food compositions, supplements for infants or toddlers, baby foods, stage 2 infant formula, growing-up milk, infant cereals, or fortifiers. In some embodiments, the composition of the present invention is an infant formula, baby food, infant cereal, growing-up milk, supplement, or fortifier suitable for use in infants or toddlers.

[0325] By way of example, the composition may contain additional components that may help reduce the risk of allergic reactions and / or allergic reactions. Additionally or alternatively, the composition may contain other components that may be beneficial during the weaning period.

[0326] For example, the composition may contain additional probiotics, such as probiotics known to reduce the risk of allergic reactions and / or allergic reactions (e.g., Bifidobacterium lactis, Lactobacillus rhamnosus, Bifidobacterium longum subsp. infantis), formula foods (e.g., partially hydrolyzed formula foods, extensively hydrolyzed formula foods, amino acid-based formula foods, or complete formula foods), infant foods (with or without milk fat), milk fat, cereal foods, docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), butyrate, and / or gamma-linolenic acid (GLA).

[0327] On a dry weight basis, the transitional form of Bifidobacterium longum can be approximately 10 per gram of composition. 3 cfu to 10 12 CFU probiotic strains, preferably between 10 7 cfu with 10 12 CFU, such as between 10 8 cfu with 10 10 The amount of probiotic strains between CFU is included in the composition. In one embodiment, the Bifidobacterium longum transitional microorganism is live. In another embodiment, the Bifidobacterium longum transitional microorganism is non-replicating or inactivated. In some other embodiments, both live and inactivated Bifidobacterium longum transitional microorganisms may be present.

[0328] Suitable, the composition comprises one or more polysaccharide substrates as described herein.

[0329] In some embodiments, the composition comprises at least one prebiotic oligosaccharide, preferably 3'-O-fucosylated lactose (3FL).

[0330] In some embodiments, the composition comprises

[0331] -12% to 38% by weight, preferably 17% to 31% by weight of 3-FL.

[0332] The composition may contain 3-FL in the range of 0.01 g / L to 7 g / L, preferably in the range of 0.025 g / L to 6 g / L, and more preferably in the range of 0.05 g / L to 5 g / L.

[0333] In some embodiments, the composition comprises a Bifidobacterium longum transitional microorganism that preferentially utilizes 3-fucosyllactose (3-FL). On a dry weight basis, the composition may contain between 10 [units of something] per g of composition. 3 cfu to 10 12 Probiotic strains between CFU, more preferably between 10 7 CFU and 10 12 CFU, such as between 10 8 CFU and 10 10 The probiotic strain is mixed with 3-FL in an amount between 0.01 g / L and 7 g / L, preferably between 0.025 g / L and 6 g / L, and more preferably between 0.05 g / L and 5 g / L.

[0334] Suitablely, the 3'-O-fucosylated lactose (3'FL) contained in the composition promotes the growth of Bifidobacterium longum transitional microorganisms that preferentially utilize 3-fucosylated lactose (3-FL).

[0335] method

[0336] On the other hand, the present invention provides a method for treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children; wherein the method comprises administering an effective amount of a prebiotic and a combination of a Bifidobacterium longum transitional microorganism, a prebiotic, or a Bifidobacterium longum transitional microorganism to a subject in need.

[0337] In another aspect, the present invention relates to the use of a combination of a prebiotic and a Bifidobacterium longum transitional microorganism according to the present invention, a prebiotic or a Bifidobacterium longum transitional microorganism, in the preparation of a medicament for treating and / or preventing allergic reactions and / or allergic sensitization in infants or young children.

[0338] The Bifidobacterium longum transitional microorganism according to the present invention can be the Bifidobacterium longum transitional microorganism as described herein.

[0339] Prebiotics can be as described in this article.

[0340] The combination of prebiotics and the Bifidobacterium longum transitional microorganism according to the invention can be provided in any form as described herein. For example, the combination can be provided in a composition as described herein.

[0341] Those skilled in the art will understand that they are free to combine all the features of the invention disclosed herein. In particular, features described for the products of the invention can be combined with the methods of the invention, and vice versa. Furthermore, features described for different embodiments of the invention can be combined. Where known equivalents exist for a particular feature, such equivalents are incorporated as expressly mentioned in this specification.

[0342] Further advantages and features of the invention will become apparent upon reference to the accompanying drawings and non-limiting embodiments.

[0343] Example

[0344] Example 1: Transitional Bifidobacterium longum increases intestinal epithelial barrier resistance

[0345] In vitro experiments using the human colorectal adenocarcinoma cell line (Caco-2) have shown that transitional Bifidobacterium longum strains, when incubated with epithelial cells for 24 hours, can increase transmembrane resistance (TEER) (see...). Figure 2 ).

[0346] These results suggest that transitional Bifidobacterium longum plays a role in reducing paracellular permeability during the transition from breast milk to solid foods, which is associated with increased dietary antigen uptake and thus contributes to the barrier balance required for optimal maturation of the immune system.

[0347] Example 2: Transitional Bifidobacterium longum increases the anti-inflammatory cytokine IL-10, improves the IL-10 / IL-12 ratio, and reduces... Low IL-5 expression in type 2 skewed helper T cells

[0348] In vitro, a transitional strain of *Bifidobacterium longum* increases the immunomodulatory response in human peripheral blood mononuclear cells, characterized by interleukin-10 (IL-10) after 36 hours of stimulation. Figure 13 The IL-10 / IL-12p40 ratio increased to that of the probiotic Lactobacillus rhamnosus (NCC4007) Figure 3 (Similar to or even greater in degree.)

[0349] Furthermore, in vitro, the transitional strain of *Bifidobacterium longum* reduced IL-5 expression on type 2 oblique helper T cells after 48 hours of stimulation to a degree similar to or even greater than that of the probiotic *Bifidobacterium lactis* (which has known anti-inflammatory properties). Figure 12 IL-5 was determined using the following method: peripheral blood mononuclear cells (PBMCs) were isolated from the yellow layer of blood clots obtained from healthy adults using a density gradient. The PBMCs were then... 6Cells / ml were seeded in 48-well plates in complete Isocove modified Dulbecco medium (cIMDM) containing 10% fetal bovine serum, 1% glutamine, 1% penicillin / streptomycin, and 0.1% gentamicin. Cells were cultured in the presence of 50 ng / ml IL-4 and 1 μg / ml anti-CD40. After 3 days of culture, different bacterial strains, including all transitional Bifidobacterium longum isolates, were added at specified concentrations. The cell culture supernatant was collected to assess IL-5 cytokine expression by ELISA. Standard curves for each cytokine were used to calculate the absolute amount (picograms / ml) from the optical density readings.

[0350] Example 3: Analysis of carbohydrate-active enzyme (CAZyme) genes in a transitional microorganism of Bifidobacterium longum

[0351] Combined with the dbCAN2 tool (Zhang et al., Nucleic Acids Res.46(W1):W95-W101 (2018)) and the databases HMMdb (v9) and Diamond (v2.0.8). Figure 4 and Figure 5 The genomes of the subspecies of *Bifidobacterium longum* listed in the documentation were annotated as CAZyme. Query sequences with a coverage >0.50 and an e-value <1e-15 were annotated using HMMER according to the dbCAN CAZyme domain HMM database. Diamond was also used to annotate query sequences with hits in the CAZy database (Drula et al., NucleicAcids Res.50(D1):D571-D577 (2022)) (http: / / www.cazy.org / ), with an identity >0.90 and an e-value <1e-102. In cases where CAZyme annotations for query sequences did not match between the HMMER and DIAMOND tools, HMMER annotation was preferred. Only the CAZyme family and subfamilies encoding glycoside hydrolases (GH) and polysaccharide lyases (PL) were used for comparative analysis of *Bifidobacterium longum* subspecies (see [link to documentation]). Figure 4 and Figure 5 ).

[0352] Example 4: Utilization of polysaccharide substrates

[0353] The pulverized or homogenized fecal sample was mixed 10-fold with PBS / glycerol (1 / 10) (w / v) and then centrifuged at 2000g for 2 minutes. The slurry and pellets were then stored at -80°C. Frozen fecal samples were thawed from -80°C storage before centrifugation at 2000g for 2 minutes. The resulting supernatant was inoculated using a medium based on the medium disclosed in the following literature: Daguet et al. (Journal of Functional Foods; 2016; 20; 369-379). This medium was supplemented with 5 g / L of the specific fiber to be tested and 5 E07 CFU / ml of Bifidobacterium supplement.

[0354] Cultures were established at 37°C under a nitrogen gas flow to ensure anaerobic conditions and gentle stirring. Aliquots were taken and analyzed at specified time points.

[0355] The growth of the Bifidobacterium longum transitional strain NCC 5001 was promoted by pectin (beet) and arabinogalactan (larch wood). Figure 7 ).

[0356] The growth of the Bifidobacterium longum transitional strain NCC 5002 was promoted by arabinogalactan (larch wood) and starch (potato). Figure 8 ).

[0357] Example 5: Characterization of the transitional microorganism Bifidobacterium longum

[0358] Transitional strains of *Bifidobacterium longum* were isolated from the feces of breastfed infants using Eugon tomato agar (ETA). The obtained isolates were sequenced using PacBio to obtain the fully closed-assembled genome of each strain. Each strain, along with its genome sequence data, is deposited at the Nestlé In-House Culture Collection (NCC, Lausanne, Switzerland) and the National Collection of Microorganisms (CNCM) at the Pasteur Institute (Paris, France). OrthoAni (… https: / / www.ezbiocloud.net / tools / orthoani The genomes of the strains were compared with other publicly available genomes representing the overall diversity of Bifidobacterium longum species (Table 5) by average nucleotide identity (ANI) and with metagenomic assembled genomes (MAGs) obtained from metagenomic sequences published from infant feces of the same cohort.

[0359] Table 5 - List of genomes used for ANI analysis and their publicly available references. (T) represents the type strain. .

[0360]

[0361] Analysis showed that the newly described strains, grouped together with MAG from the same cohort, defined a well-defined clade belonging to the *Bifidobacterium longum* species. Two previously isolated strains, BSM11-5 and 3_mod, were found to be grouped within this newly described clade. This clade is genetically distinct from the *Bifidobacterium longum* subspecies (96.40% ANI). This clade is associated with *Bifidobacterium longum* subspecies (98.207%) and with the previously identified group of strains (JDM301, CMCC_P0001, and BXY01) (O'Callaghan et al., 2015), but still exhibits significant differences, sharing 98.260% identity with that group. Figure 1 A phylogenetic tree based on ANI UPGMA is shown. The scale indicates the percentage (%) of identity at each branch point.

[0362] The selection of the above-mentioned genomes, representing the diversity of *Bifidobacterium longum* subspecies, was annotated with carbohydrate-active enzymes (CAZYs) using the dbCAN annotation pipeline (http: / / bcb.unl.edu / dbCAN / ). The results showed that *Bifidobacterium longum* subsp. *suis*, *Bifidobacterium longum* subsp. *suillum*, and *Bifidobacterium longum* subsp. *suillum* strains contain GH20 (lactose-N-glucosidase), which is involved in the degradation and metabolism of lactose-N-tetrasaccharides (LNTs). Similar to *Bifidobacterium longum* subsp. *infant* strains, *Bifidobacterium longum* transitional strains also possess similar enzymes and additionally carry the GH29 (fucosidase) gene, which is involved in the degradation and metabolism of fucosylated human milk oligosaccharides (such as 2'FL, 3'FL, or diFL). In addition, three strains (CNCM I-5684, BSM1-15 and 3_mod) also carry the GH 33 (sialidase) encoding gene involved in the degradation and metabolism of sialylated HMOs (such as 3'SL or 6'SL) (Table 6).

[0363] Table 6 - GH20 (lacto-N-glucosidase) and GH29 (α-fucosidase) are encoded in the genomes of each representative. The genes for GH95 (α-fucosidase / α-galactosidase) and GH33 (sialidase) glucosyl hydroxylase family enzymes. quantity .

[0364]

[0365] All newly obtained genomes were compared and aligned with the genomes of two strains (Bifidobacterium longum subsp. infant ATCC15697 and Bifidobacterium kanamycin DSM 21854), which belong to species in which genes responsible for the utilization of fucosylated HMOs have been elucidated (James et al., 2019).

[0366] result

[0367] like Figure 10 As shown, all newly described strains contain genes responsible for utilizing fucosylated HMOs. The NCC5001 tissue reflects one of the *Bifidobacterium longum* subsp. *infant* ATCC 15697, while all other strains (NCC 5000, NCC5002, NCC 5003, NCC 5004, NCC 5025) carry gene tissues more closely related to *Bifidobacterium kanamycin* DSM 21854. Overall, the similarity of fucosidases to the fully described *Bifidobacterium longum* subsp. *infant* ATCC 15697 was higher than 77% (for BLON_2334) and 88% (for BLON_2335) among all newly described strains.

[0368] Example 6 - NCC 5025 exhibits a high growth rate on 3-FL

[0369] Transitional strains of *Bifidobacterium longum* were obtained from the Nestlé Culture Collection and reactivated from freeze-dried stock in MRS supplemented with 0.05% cysteine ​​(MRSc) using two consecutive culture steps (16 h, 37 °C, anaerobic). The reactivated cultures were then centrifuged, washed, and resuspended in 1 volume of PBS. The washed cells were then inoculated into carbon-free MRS-based medium (MRSc-C) (10 g L⁻¹ bactoproteose peptone n°3, 5 g L⁻¹ bacterial yeast extract, 1 g L⁻¹ Tween 80, 2 g L⁻¹ diammonium citrate, 5 g L⁻¹ sodium acetate, 0.1 g L⁻¹ magnesium sulfate, 0.05 g L⁻¹ manganese sulfate, 2 g L⁻¹ disodium phosphate, 0.5 g L⁻¹ cysteine), with 3-FL added as the sole carbon source at a final concentration of 0.5%. Growth was then carried out in 96-well microplates with a volume of 200 µl per well. Incubation was performed anaerobically for 46 h, during which the optical density was measured at 580 nm using a spectrophotometer. Growth curves were then modeled using a logistic growth model to obtain the relative growth rate k for each variant.

[0370] Of all the Bifidobacterium longum transitional strains tested, NCC 5025 exhibited the highest growth rate on 3FL, indicating that this strain is best adapted to this substrate (see [link]). Figure 6It has been shown that 3-FL is a human milk oligosaccharide, which shows the greatest increase in human breast milk during the transition period between a milk-based diet and a solid diet (Plows, JF, et al., Longitudinal Changes in Human Milk Oligosaccharides (HMOs) Over the Course of 24 Months of Lactation. J Nutr, 2021. 151(4): 876-882), and therefore these results demonstrate the advantage of applying NCC 5025 during this period.

[0371] method

[0372] CACO-2 cell culture and transmembrane resistance measurement

[0373] Caco-2 cells (HTB-37; American Type Culture Collection) were seeded into 24-well semi-permeable inserts. Caco-2 monolayers were cultured for 14 days, with the medium changed three times weekly, until a functional cell monolayer with transmembrane resistance (TEER) was obtained. Cells were maintained in Dulbecco modified Eagle medium (DMEM) containing glucose and glutamine, supplemented with HEPES and 20% (v / v) heat-inactivated fetal bovine serum. The TEER of the Caco-2 monolayer (=0-hour time point) was measured before adding bacteria to the top compartment. The TEER of the empty insert was subtracted from all readings to calculate the residual resistance of the insert. A probiotic strain (directly derived from the glycerol stock) was then diluted in Caco-2 complete medium and added to the top of the Caco-2-containing insert at a colony-forming unit (CFU) of 2 × 10⁶. Cells were also exposed to Caco-2 complete medium (CM) in two control compartments and 0.75% glycerol in the top compartment as a vector control. Cells were treated for 24 hours, and TEER was measured at several time points (2 hours, 4 hours, 6 hours, and 24 hours). After subtracting the TEER of the empty insert, all time point values ​​were normalized to their own 0-hour values ​​(to account for differences in initial TEER between different inserts) and expressed as a percentage of the initial values. Figure 2 ).

[0374] Immunospectral analysis using PBMC cells

[0375] Peripheral blood mononuclear cells (PBMCs) were isolated from the yellow layer of blood clots obtained from healthy adults using a density gradient. The PBMCs were then separated at a density of 1.5 × 10⁻⁶. 6PBMCs were seeded at 100 cells / ml in 48-well Isocove modified Dulbecco medium (cIMDM) containing 10% fetal bovine serum, 1% glutamine, 1% penicillin / streptomycin, and 0.1% gentamicin. PBMCs were stimulated for 36 hours in the presence of different bacterial strains, including 10... 7 Transitional Bifidobacterium longum isolates and probiotic strains were collected at CFU / ml. Cell culture supernatant was collected to assess cytokine expression of IL-10 and IL-12p40 by ELISA. Standard curves for each cytokine were used to calculate the absolute amount (picograms / ml) from optical density readings.

[0376] Example 7: Bifidobacterium longum transitional microorganisms produce precursors of folic acid and riboflavin, thus maintaining the integrity of the intestinal barrier. Two whole metabolites

[0377] Genetic characterization of transitional isolates of Bifidobacterium longum revealed that they possess a specific genetic region containing a set of 6 genes ( Figure 11 As shown in the figure, the transitional strain of *Bifidobacterium longum* and the *Bifidobacterium longum* subsp. *infant* are the only strains carrying this set of genes. Strains belonging to *Bifidobacterium longum* subsp. *Suis* and *Bifidobacterium longum* subsp. *suillum* do not contain the entire operon. This region encompasses two genes (PabA and PabB) involved in the conversion of cladonic acid to 4-amino-4-deoxycladonic acid (a precursor to para-aminobenzoic acid and folic acid) in different microorganisms. Furthermore, this region also contains another set of four genes (RibD, RibE, RibAB, RibH) related to riboflavin biosynthesis, as recently demonstrated in *Bifidobacterium longum* subsp. *infant* (Solopova et al., 2020, Front Microbiol, 2020.11: 573335).

[0378] The presence of these genes suggests that the transitional microorganism Bifidobacterium longum may be able to produce riboflavin and folic acid.

[0379] Example 8: Transitional Bifidobacterium longum produces SCFAs beneficial for the prevention and control of allergic reactions. .

[0380] methodInfant microbiota containing 3-fucosylated lactose (3-FL), supplemented or unsupplemented with *Bifidobacterium longum* species or transitional *Bifidobacterium longum*, were subjected to in vitro batch fermentation for 48 hours. Acetate, butyrate, and propionate levels in the supernatants collected at T0, T24, and T48 hours were measured by 1H-NMR. Total SCFA was correlated with the sum of peak integrals for acetate, butyrate, and propionate. Bar graphs indicate the dynamics of total SCFA consumption and production between 0 and 24 hours (blue bars) and between 24 and 48 hours (red bars).

[0381] result :

[0382] Figure 14 Transitional Bifidobacterium longum showed greater metabolic activity against weaning-associated HMOs (i.e., 3-FL) and produced more total SCFAs than Bifidobacterium infantis, which is beneficial for allergy prevention and management, as demonstrated in numerous studies (Trompette et al., Mucosal Immunology 15, 908-926 (2022); Roduit et al., Allergy, Apr;74(4):799-809 (2019); Canani et al., Sci Rep Aug 21;8(1):12500 (2018); Gio-Batta et al., Sci Rep10, 22449 (2020); Cait et al., 2019 J Allergy Clin Immunol. Dec;144(6):1638-1647).

[0383] All publications mentioned in the foregoing specification are incorporated herein by reference. Various modifications and variations of the methods and systems described herein will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in conjunction with specific preferred embodiments, it should be understood that the invention protected by the claims should not be unduly limited to such specific embodiments. Indeed, various modifications to the described modes of carrying out the invention that will be apparent to those skilled in molecular biology or related fields are intended to fall within the scope of the following claims.

[0384]

[0385]

[0386]

[0387]

Claims

1. A transitional strain of *B. longum* for use in the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children, wherein the transitional strain of *B. longum* comprises a strain deposited at the French National Center for Microbial Collections (CNCM) with accession number CNCMI-5942 or a transitional strain of *B. longum* having the identifying characteristics of a transitional strain of *B. longum* deposited with accession number CNCM I-5942.

2. A transitional strain of Bifidobacterium longum for use in the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children, wherein the transitional strain of Bifidobacterium longum has at least 98.1% average nucleotide identity (ANI) with a Bifidobacterium longum strain deposited in the CNCM with accession number CNCM I-5942.

3. A transitional strain of *Bifidobacterium longum* for the purpose according to any of the preceding claims, wherein the transitional strain of *Bifidobacterium longum* is not resistant to any of tetracycline, erythromycin, clindamycin, and ampicillin; preferably wherein the transitional strain of *Bifidobacterium longum* is not resistant to any of tetracycline, erythromycin, clindamycin, ampicillin, gentamicin, streptomycin, chloramphenicol, and vancomycin.

4. A transitional strain of *Bifidobacterium longum* for the purpose according to any of the preceding claims, wherein the transitional strain of *Bifidobacterium longum* comprises the glycosyl hydrolase family 43_17 (GH43_17) gene; suitably wherein the GH43_17 gene comprises SEQ ID NO: 7 or a sequence having at least 60% sequence identity with SEQ ID NO:

7.

5. The *Bifidobacterium longum* transitional microorganism for the stated purpose according to claim 1, wherein the *Bifidobacterium longum* transitional microorganism is used in combination with a prebiotic, and wherein the prebiotic is: i. a polysaccharide substrate, wherein the polysaccharide substrate is suitably selected from any group listed in Tables 1 to 3; and / or ii. Human milk oligosaccharides (HMOs).

6. A combination of a prebiotic and a Bifidobacterium longum transitional microorganism according to claim 1, said combination being used for the treatment and / or prevention of allergic reactions and / or allergic sensitization in infants or young children; wherein said prebiotic is: i. a polysaccharide substrate, wherein the polysaccharide substrate is suitably selected from any group listed in Tables 1 to 3; and / or ii. Human milk oligosaccharides (HMOs).

7. The Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the said use according to any one of claims 2 to 6, wherein the prebiotic is a polysaccharide substrate selected from any one of the groups listed in Tables 1 to 3.

8. The Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the said use according to any one of claims 2 to 6, wherein the prebiotic is 3'-fucosylated lactose (3'-FL).

9. A Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the purpose according to any one of the preceding claims, wherein the Bifidobacterium longum transitional microorganism is capable of metabolizing the HMO and / or the glycan substrate.

10. A Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the stated purpose according to any one of the preceding claims, wherein the Bifidobacterium longum transitional microorganism preferentially utilizes 3'-fucosylated lactose (3'-FL).

11. A Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the stated purpose according to any one of the preceding claims, wherein the Bifidobacterium longum transitional microorganism: (a) is capable of metabolizing a polysaccharide substrate selected from any of the groups listed in Tables 1 to 3; and / or (b) encodes one or more CAZymes selected from the groups listed in Table 1, preferably wherein the Bifidobacterium longum transitional microorganism further encodes one or more CAZymes selected from Tables 2 and 3.

12. A Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof for the said use according to any one of the preceding claims, wherein the Bifidobacterium longum transitional microorganism and / or prebiotic: (a) Increase IL-10 levels in the infants or young children; (b) Increase the IL-10 / IL-12 ratio in the infants or young children; (c) Reduce IL-5 levels in the infants or young children; (d) Modulating the permeability of the intestinal epithelial barrier; preferably, wherein the Bifidobacterium longum transitional microorganism and / or prebiotic reduces the permeability of the intestinal epithelial barrier; and / or (e) Increase the levels of riboflavin and / or folic acid in the infants or young children.

13. Use of Bifidobacterium longum transitional microorganism, prebiotic or combination thereof as defined in any of the preceding claims for promoting immune tolerance in an infant or young child, preferably by promoting the growth of Bifidobacterium longum transitional microorganism in the intestine of the infant or young child.

14. The use of Bifidobacterium longum transitional microorganism, prebiotic, or combination thereof as defined in any of the preceding claims for regulating the intestinal barrier permeability of an infant or young child, preferably by promoting the growth of Bifidobacterium longum transitional microorganism in the intestine of the infant or young child.

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