Composition for use

Abstract

The present invention provides methods for identifying an infant or young child at risk of growth retardation, and compositions for preventing and / or treating growth retardation in an infant or young child.

Description

TECHNICAL FIELD

[0001] The present invention relates to a method for identifying an infant or young child at risk of growth retardation, and to a composition for preventing and / or treating growth retardation in an infant or young child. BACKGROUND

[0002] Studies of the impact of malnutrition on growth have mainly focused on weight-related outcomes. However, approaches aimed at identifying and improving reduced weight gain can have negative long-term consequences; for example, weight gain can be associated with a risk of metabolic disorders.

[0003] Some studies have investigated the issue of insufficient growth by using a microbiome-oriented solution (e.g. Chen RY et al. NEJM 2021; 384:1517-28; Subramanian S et al. Nature 2014; 510(7505):417-421). However, most of these studies report an increase in weight-for-age or height-for-age, which can be related to the use of calorie-rich food in these interventional trials, and can have consequences such as a risk of subsequent metabolic disorders.

[0004] There is a need for further approaches and methods to identify and / or treat or prevent the consequences of height- or length-related (e.g. due to malnutrition) growth retardation in infants or young children. SUMMARY

[0005] The present invention is based at least in part on the inventors’ surprising determination that a reduced level of B. pseudocatenulatum and / or S. thermophilus in the gut microbiota is associated with linear growth slowing in groups of infants and young children. Furthermore, the inventors have determined that B. pseudocatenulatum can be promoted by human milk oligosaccharides (HMOs). To the inventors’ knowledge, the relationship between linear growth slowing and the microbiome has not been investigated previously, or they have failed to identify positive outcomes. For example, age-for-height z (HAZ) score or age-for-length z (LAZ) score was determined to have no association or improvement (Subramanian S et al. Nature. 2014; 510(7505):417-421) or LAZ was reported to be associated with only a few bacteria in the duodenal microbiota (Chen RY et al. N Engl J Med. 2020; ibid).

[0006] Accordingly, the present application provides a means for identifying an infant or young child at risk of growth faltering by determining the abundance of Pseudob. infant- is and / or S. thermophilus in one or more samples obtained from the infant. The present application also provides a method for preventing and / or treating growth faltering in an infant or young child by promoting Pseudob. infant- is and / or S. thermophilus in the gut microbiota. Accordingly, the present application can comprise promoting the abundance and / or activity of Pseudob. infant- is and / or S. thermophilus in the gut microbiota.

[0007] Accordingly, in a first aspect, the present application provides a method for identifying an infant or young child at risk of growth faltering, wherein the method comprises determining the abundance of Pseudob. infant- is and / or S. thermophilus in one or more samples obtained from the infant.

[0008] Suitably, the method comprises determining the abundance of Pseudob. infant- is in one or more samples obtained from the infant.

[0009] Suitably, an infant or young child having a reduced level of Pseudob. infant- is is identified as being at risk of growth faltering.

[0010] Growth faltering can be stunted height or length. Suitably, stunted height or length can be defined as a decreasing length-for-age z-score (LAZ) or a decreasing LAZ over time. Suitably, stunted height or length can be defined as a decreasing height-for-age z-score (HAZ) or a decreasing HAZ over time.

[0011] In another aspect, the present application provides a composition for preventing and / or treating growth faltering in an infant or young child, wherein the composition promotes Pseudob. infant- is in the gut microbiota of the infant or young child.

[0012] The composition can comprise Pseudob. infant- is microorganisms. Suitably, a composition comprising Pseudob. infant- is microorganisms can be administered in combination with a prebiotic.

[0013] The composition can comprise a prebiotic. Suitably, a composition comprising a prebiotic can be administered in combination with Pseudob. infant- is microorganisms.

[0014] The present application also provides a combination of Pseudob. infant- is microorganisms and a prebiotic for use in preventing and / or treating growth faltering in an infant or young child.

[0015] The prebiotic can be in the form of a dietary or nutritional composition. The prebiotic can comprise a human milk oligosaccharide (HMO).

[0016] The present application also provides an HMO or a combination of HMOs for use in preventing and / or treating stunted growth in an infant or young child, wherein the HMO or combination of HMOs promotes Pseudobrevibacterium sp. in the gut microbiota of the infant or young child.

[0017] The HMO can be selected from 2'-FL, 3-FL, di-FL, 3'-SL, 6'-SL, LNT and LNnT and any combination thereof. The HMO can be any HMO or combination of HMOs as defined herein.

[0018] The HMO can be selected from 2'-FL, di-FL, 6'-SL and LNnT and any combination thereof. The HMO can be a combination of 2'-FL and di-FL. The HMO can be a combination of 6'-SL and LNnT.

[0019] The HMO can be provided in combination with galacto-oligosaccharides (GOS).

[0020] The present application also provides a Pseudobrevibacterium sp. microorganism for use in preventing and / or treating stunted growth in an infant or young child.

[0021] The treatment and / or prevention of stunted growth can be associated with enhanced bone development and / or bone strength in the subject.

[0022] The present application also provides a method of preventing and / or treating stunted growth in an infant or young child, the method comprising administering to the infant or young child a composition that promotes Pseudobrevibacterium sp. in the gut microbiota.

[0023] In another aspect, the present application provides the use of a Pseudobrevibacterium sp. microorganism and / or a prebiotic in the manufacture of a medicament for preventing and / or treating stunted growth in an infant or young child.

[0024] The present application also provides the use of a composition for modulating the abundance of Pseudobrevibacterium sp. in the gut of an infant or young child. The composition can be any composition as defined herein.

[0025] The present application also provides a probiotic composition comprising Pseudobrevibacterium sp.

[0026] The present application also provides a synbiotic composition comprising Pseudobrevibacterium sp. and a prebiotic. The prebiotic can be a prebiotic as defined herein.

[0027] Suitably, for any aspect of the present application, Streptococcus thermophilus can be provided as a substitute for Pseudobrevibacterium sp.

[0028] Suitably, any aspect of the present application can relate to a combination of Pseudobrevibacterium sp. and Streptococcus thermophilus. BRIEF DESCRIPTION OF DRAWINGS

[0029] Figure 1 - Schematic of micro-health studies

[0030] Figure 2 - Microbiome profile analysis of populations differentiated by dynamic change in LAZ score over time. Q1 is an infant with a negative slope in length-for-age z (LAZ) score over time, which is defined as “stunting”. Q4 is an infant with a positive slope in LAZ score over time, which is defined as “reference”.

[0031] Figure 3 - Use of both (A) dynamic change and (B) static outcome to identify Bifidobacterium pseudocatenulatum as a bacterial signature of inadequate height-for-age distribution up to 24 months.

[0032] Figure 4 - Use of both (A) dynamic change and (B) static outcome to identify Streptococcus thermophilus as a bacterial signature of inadequate height-for-age distribution up to 24 months.

[0033] Figure 5-2 Exemplary WHO length-for-age (Z-score) of girls from 2 to 5 years of age. DETAILED DESCRIPTION

[0034] The various preferred features and embodiments of the present application will now be described, by way of non-limiting examples, with reference to the accompanying drawings. The skilled person will understand that they can combine all features of the present application disclosed herein without departing from the scope of the disclosed application.

[0035] It must be noted that, as used herein and in the appended claims, the singular form “a”, “an” and “the” include plural references unless the context clearly dictates otherwise.

[0036] As used herein, the terms “comprising” and “consisting of’ are synonymous with “including” or “containing,” and are inclusive or open-ended and do not exclude additional, unrecited members, elements, or method steps. The terms “comprising” and “consisting of’ also include the term “consisting essentially of.”

[0037] Numerical ranges include the numbers defining the range.

[0038] The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the

[0039] The methods and systems disclosed herein can be used by a physician, medical professional, laboratory technician, infant or young child care provider, etc.

[0040] Growth retardation

[0041] The definition of stunting or growth retardation by UNICEF and WHO is that a child is too short for his or her age. These children can suffer from severe irreversible physical and cognitive impairments that accompany stunting. The effects of stunting can last a lifetime and even affect the next generation (https: / / data.unicef.org / topic / nutrition / malnutrition). In 2020, according to the UNICEF / WHO / World Bank Joint Child Malnutrition Estimates (2021 edition), approximately 22% of children under 5 years of age were affected by stunting. This equates to 149.2 million children under 5 years of age with stunting. Growth retardation is seen in infants and toddlers, but extends beyond preschool and school-age children (Leroy JL, Ruel M, Habicht JP, Frongillo EA. J Nutr. 2014; Dutta A et al. Food Nutr Bull. 2009).

[0042] Growth retardation in infants and young children can be caused by many factors, including poor intrauterine environment, aflatoxin exposure, intergenerational effects, environmental gut dysfunction, nutritional deficiencies and dietary diversity, infections such as Campylobacter infection, diarrhea, drinking water, public health and sanitation issues, and other causes such as maternal factors. Some studies have investigated the problem of insufficient growth by using microbiome-oriented solutions (e.g., Chen RY et al. NEJM 2021; Subramanian S et al. Nature, 2014). However, most of these studies reported an increase in weight-for-age or weight-for-height, which can be related to the use of calorie-rich foods in these interventional trials and can have consequences such as the risk of subsequent metabolic disorders. To the best of the inventors’ knowledge, the relationship between linear growth retardation and the microbiome has not been investigated previously, or they failed to identify positive outcomes. For example, no association or improvement in height-for-age z-score (HAZ) or length-for-age z-score (LAZ) was reported (Subramanian S et al. Nature. 2014; ibid) or LAZ was only reported to be associated with a few bacteria in the duodenal microbiota (Chen RY et al. N Engl J Med. 2020; ibid).

[0043] As used herein, “growth retardation” can refer to the short stature or length of an infant or young child.

[0044] Growth indicators such as length-for-age / height-for-age can be used to identify children who are stunted (short) due to long-term undernutrition or repeated illness.

[0045] Suitably, stunting can be defined as a decreasing length-for-age z-score (LAZ) or decreasing LAZ over time. LAZ can also be referred to as height-for-age z-score (HAZ).

[0046] A child growing normally follows a trend that is generally parallel to the median line and the z-score line (see Figure 5 ). Most children will grow in a “trajectory”, i.e. on or between the z-score lines, and generally parallel to the median line; the trajectory can be below or above the median line. When interpreting growth charts, the following situations are notable and can indicate a problem or imply a risk: the child’s growth line crosses a z-score line, there is a sharp slope or drop in the child’s growth line, or the child’s growth line remains flat (stagnation), i.e. there is no increase in weight or length / height.

[0047] Stunting can be defined as a static outcome. For example, stunting can be defined as length-for-age / height-for-age below -2. For example, stunting can be defined as cases where the height-for-age is less than -2 standard deviations from the median line of the World Health Organization (WHO) Child Growth Standards (https: / / sdgdata.gov.uk / 2-2-1 / #:~:text=Definitions,(WHO)%20Child%20Growth%20Standards).

[0048] Stunting can be defined as a dynamic change, e.g. a change in LAZ assessed over time (e.g. 0-36 months, 6-36 months or 6-24 months). An infant with a decreasing LAZ score over time can be defined as having stunting.

[0049] The dynamic change in growth can be referred to as “growth velocity” or “height velocity”. Growth velocity or height velocity can be defined as shown in equation (1).

[0050] (1)

[0051] where H1 and H2 are two height measurements, and At is the time interval between the measurements. Stunting can be associated with malnutrition.

[0052] Malnutrition is caused by reduced food intake and / or illness. Malnutrition is associated with greater risk of medical complications and infections, increased risk of illness and infection death, and micronutrient deficiencies. Non-limiting examples of micronutrient deficiencies associated with malnutrition are iron deficiency, iodine deficiency, and vitamin A deficiency. The most common method of assessing malnutrition, particularly infant and young child malnutrition, is through anthropometry. Diagnosis is usually made in one of three ways: by weighing and measuring the height of the subject; by measuring the mid-upper arm (MUAC) circumference of the subject; and / or by checking the subject’s lower legs or feet for oedema. Malnutrition can be divided into two types: severe acute malnutrition (SAM) and moderate acute malnutrition (MAM). A subject is classified as having SAM if their weight-for-height Z-score (WHZ) is less than three standard deviations (-3 s.d.) below the median of the World Health Organization (WHO) reference growth standard. Subjects with a WHZ between -2 s.d. and -3 s.d. of the median of the WHO reference growth standard are classified as having MAM. A MUAC measurement of less than 12.5 cm is also indicative of moderate acute malnutrition in a subject between about six months and about five years of age. Finally, oedema in both feet and lower legs of a subject is a sign of SAM. The WHO reference growth standard is available from the WHO. See, for example, “World Health Organization Department of Health and Nutrition for Development: WHO Child Growth Standards: Methods and Development based on Weight, Length and Head Circumference: Growth Velocity: Methods and Development; World Health Organization” (2009 edition or current version).

[0053] Suitably, the infant or young child can have SAM. Suitably, the infant or young child can have MAM.

[0054] A subject at risk of growth retardation can be a subject living in a geographical area where a comprehensive nutritious diet is limited or unavailable and / or a subject living in a geographical area that is experiencing an illness outbreak. A subject at risk of growth retardation can be a malnourished subject.

[0055] Treating a subject at risk of growth retardation can reduce the occurrence of growth retardation in the subject or prevent growth retardation in the subject.

[0056] The term “prevent” in relation to growth retardation can mean reducing or eliminating growth retardation.

[0057] For example, preventing growth retardation can involve reducing, in a treated subject: the incidence or development of growth retardation; the development, duration, and / or severity of growth retardation, if growth retardation does develop; or a combination thereof. For each aspect, the amount reduced in a treated subject compared to an untreated subject can each be about 2%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or about 100%.

[0058] Infant or young child

[0059] Suitably, the infant or young child can be less than about 60 months old. For example, the infant or young child can be less than 48 months old, less than 36 months old, or less than 24 months old.

[0060] Suitably, the infant or young child can be about 1 month old to 60 months old, about 1 month old to 48 months old, about 2 months old to 60 months old, about 2 months old to 48 months old, about 2 months old to 36 months old, about 4 months old to 36 months old, about 6 months old to 36 months old, or about 6 months old to 24 months old.

[0061] For example, the infant or young child can be at least about 6 months old, at least about 10 months old, at least about 12 months old, at least about 14 months old, at least about 16 months old, at least about 20 months old, or at least about 24 months old.

[0062] Suitably, the infant or young child can be about 6 months old to about 60 months old, about 6 months old to about 48 months old, about 6 months old to about 36 months old, or about 6 months old to about 24 months old.

[0063] An infant can be a child under the age of 12 months. A “young child” can be a child aged one year to less than five years old or one year to less than three years old.

[0064] The subject can be a mammal. Preferably, the subject is a human. Unless otherwise specified, ages referred to herein are in relation to human subjects.

[0065] Composition

[0066] The composition can be suitable for or can be administered to an infant or young child in any suitable form such as a dose unit (e.g., a tablet, capsule, powder packet, etc.) of a nutritional composition. The composition can be in powder, semi-liquid, or liquid form. The composition can be added to a nutritional composition, infant formula, food composition, supplement for an infant or young child, baby food, second-stage infant formula, growing-up milk, infant or young child cereal, or fortifier. In some embodiments, the composition of the present application is an infant formula, baby food, infant or young child cereal, growing-up milk, supplement, or fortifier that can be intended for an infant or young child.

[0067] The expressions "complementary feeding period," "complementary period," "transition period," "transitional feeding period," and "weaning period" are used interchangeably and refer to the period in which milk (breast milk or formula) is replaced by other foods in the diet of an infant or young child. An infant or young child is typically transitioned or weaned from exclusive milk feeding (breastfeeding or formula feeding) to a mixed diet comprising milk and / or solid foods. The transition period depends on the infant or young child, but is typically from about 4 months old to about 18 months old, such as from about 6 months old to about 18 months old, but can extend to about 24 months or longer in some cases. For humans, the weaning period typically begins at 4 months old to 6 months old, and is considered complete once the infant or young child is no longer fed with breast milk or infant formula, typically at about 24 months old. In some embodiments, the weaning period is 4 months to 24 months.

[0068] Suitably, the composition is a dietary composition or a nutritional composition.

[0069] The expression "dietary composition" or "nutritional composition" means any kind of composition or formulation which provides a nutritional benefit to the individual and which is safe for consumption by a human or animal. The nutritional composition can be in solid (e.g. powder), semi-solid or liquid form and can comprise one or more macronutrients, micronutrients, food additives, water, etc. For example, the nutritional composition can comprise the following macronutrients: a protein source, a lipid source, a carbohydrate source, and any combination thereof. In addition, the nutritional composition can comprise the following micronutrients: vitamins, minerals, fibers, phytochemicals, antioxidants, prebiotics, probiotics, and any combination thereof. The composition can also comprise food additives such as stabilizers (when provided in solid form) or emulsifiers (when provided in liquid form). The amounts of the various ingredients can be expressed in g / 100 g of the composition on a dry weight basis when the composition is in solid form (e.g. powder), or in g / L of the composition for a concentration when the composition is in liquid form (the latter also encompassing liquid compositions that can be obtained from powders after reconstitution in a liquid such as milk, water, for example reconstituted infant or baby food or follow-on food or infant or baby cereals or any other formulation designed to provide nutrition to an infant or baby or toddler). Typically, the nutritional composition can be formulated for enteral, oral, parenteral or intravenous ingestion, and it generally comprises one of more nutrients selected from the group consisting of: a lipid or fat source, a protein source, and a carbohydrate source. Preferably, the nutritional composition is for oral consumption.

[0070] In one particular embodiment, the composition of the application is a "synthetic nutritional composition". The expression "synthetic nutritional composition" means a mixture obtained by chemical and / or biological processes.

[0071] As used herein, the expression "infant formula" means a foodstuff intended for particular nutritional use by infants during the first few months of life and capable of satisfying their entire nutritional requirements, and meeting the requirements of Article 2(c) of Directive 2006 / 141 / EC No 91 / 321 / EEC of the European Commission of 22 December 2006 on infant formulae and follow-on formulae. It also means a nutritional composition intended for infants or toddlers and as defined in the Codex Alimentarius Commission (Codex STAN72-1981) and Infant Formulae (including Foods for Special Medical Purposes). The expression "infant formula" encompasses both "first infant formula" and "follow-on formula" or "follow-up formula".

[0072] "Follow-on formula" or "follow-up formula" is intended to be given from 6 months onwards. The infant formula constitutes a major liquid element in the gradually diversified diet of such persons.

[0073] The expression "baby food" means a foodstuff intended for particular nutritional use by infants or young children during the first years of life.

[0074] The expression "baby food" means a foodstuff intended for particular nutritional use by infants or young children during the first years of life.

[0075] The expression "growing-up milk" (or GUM) means a milk-based beverage, usually fortified with vitamins and minerals, intended for young children or children.

[0076] The "fortifier" can be a liquid or solid nutritional composition suitable for fortifying or mixing with human milk, infant or young child formula or growing-up milk. Thus, the fortifier can be administered after dissolution in human breast milk, infant or young child formula, growing-up milk or human breast milk fortified with other nutrients, or it can be administered as a standalone composition. When administered as a standalone composition, the milk fortifier can also be identified as a "supplement".

[0077] The nutritional composition can comprise a protein source. The amount of protein can be 1.6 g / 100 kcal to 3 g / 100 kcal.

[0078] Whey, casein and mixtures thereof based protein sources can be used, as well as soy based protein sources. As far as whey proteins are concerned, the protein source can be based on acid whey or sweet whey or mixtures thereof and can comprise alpha-lactalbumin and beta-lactoglobulin in any desired proportions.

[0079] In some embodiments, the protein source is predominantly whey (i.e. more than 50% of the protein is from whey protein, such as 60% or 70%). The protein can be intact or hydrolysed, or a mixture of intact and hydrolysed protein. By the term "intact" is meant that the majority of the protein is intact, i.e. the molecular structure has not been altered, for example at least 80% of the protein has not been altered, such as at least 85% of the protein has not been altered, preferably at least 90% of the protein has not been altered, even more preferably at least 95% of the protein has not been altered, such as at least 98% of the protein has not been altered. In one particular embodiment, 100% of the protein has not been altered.

[0080] In one particular embodiment, the protein of the nutritional composition is hydrolysed, fully hydrolysed or partially hydrolysed. The degree of hydrolysis (DH) of the protein can be between 8 and 40, or between 20 and 60, or between 20 and 80, or more than 10, 20, 40, 60, 80 or 90. Alternatively, the protein component can be replaced by a mixture or synthetic amino acids, for example for preterm or low birth weight infants.

[0081] The term "hydrolysed" means that in the context of the present application, the protein has been hydrolysed or broken down into its constituent amino acids. The protein can be fully hydrolysed or partially hydrolysed. For example, it can be desirable to provide a partially hydrolysed protein (degree of hydrolysis between 2% and 20%) for infants or young children who are considered to be at risk of developing cow's milk allergy. If a hydrolysed protein is required, the hydrolysis process can be carried out as required and as is known in the art. For example, whey protein hydrolysates can be prepared by enzymatic hydrolysis of a whey fraction in one or more steps. If the whey fraction used as a starting material is substantially free of lactose, it is found that the protein is subjected to much less lysine blockage during the hydrolysis process. This enables the degree of lysine blockage to be reduced from about 15% by weight of total lysine to less than about 10% by weight of lysine; for example, about 7% by weight of lysine, which greatly improves the nutritional quality of the protein source.

[0082] In one embodiment of the present application, at least 70% of the protein is hydrolysed, for example at least 80% of the protein is hydrolysed, such as at least 85% of the protein is hydrolysed or at least 90%, 95%, 98% of the protein is hydrolysed. In a particular embodiment, 100% of the protein is hydrolysed.

[0083] The nutritional composition can contain a source of carbohydrate. This is particularly preferred where the nutritional composition is an infant formula. In this case, any carbohydrate source commonly found in infant formula can be used, such as lactose, sucrose, saccharose, maltodextrin, starch and mixtures thereof, but one of the preferred carbohydrate sources is lactose.

[0084] The nutritional composition can contain a source of lipid. This is particularly relevant where the nutritional composition is an infant formula. In this case, the lipid source can be any lipid or fat suitable for use in an infant formula. Some suitable fat sources include palm oil, structured triglyceride oil, high oleic sunflower oil and high oleic safflower oil, medium chain triglyceride oil. The essential fatty acids linoleic and alpha-linolenic acid can also be added, as well as small amounts of oils containing large amounts of preformed arachidonic acid and docosahexaenoic acid, such as fish or microbial oils. The ratio of n-6 to n-3 fatty acids in the fat source can be from about 5: 1 to about 15: 1, for example from about 8: 1 to about 10: 1.

[0085] The nutritional composition can also contain vitamins and minerals which are known to be essential in the daily diet and which are required in nutritionally significant amounts. Minimum requirements have been established for certain vitamins and minerals. Examples of minerals, vitamins and other nutrients which are optionally present in the compositions of the application include vitamin A, vitamin Bl, vitamin B2, vitamin B6, vitamin B 12, vitamin E, vitamin K, vitamin C, vitamin D, folic acid, inositol, niacin, biotin, pantothenic acid, choline, calcium, phosphorus, iodine, iron, magnesium, copper, zinc, manganese, chlorine, potassium, sodium, selenium, chromium, molybdenum, taurine and L-carnitine. Minerals are usually added in the form of salts. The presence and amounts of particular minerals and other vitamins will vary depending on the target population. If necessary, the nutritional compositions of the application can contain emulsifiers and stabilisers such as soy, lecithin and citric acid esters of mono- and di-glycerides and the like.

[0086] The nutritional composition can also contain other substances which can have beneficial effects, such as lactoferrin, nucleotides and nucleosides and the like.

[0087] The nutritional composition can be prepared in any suitable manner. The composition will now be described by way of example.

[0088] For example, a formula (such as an infant formula) can be prepared by blending together the protein source, the carbohydrate source and the fat source in appropriate proportions. If used, emulsifiers can be added at this point. Vitamins and minerals can be added at this point, but to avoid thermal degradation, vitamins and minerals are usually added a little later. Any lipophilic vitamins, emulsifiers and the like can be dissolved in the fat source prior to blending. Water (preferably water which has been subjected to reverse osmosis) can then be mixed in to form a liquid mixture. The water temperature is suitably in the range of from about 50°C to about 80°C to aid dispersion of the ingredients. A commercially available liquefier can be used to form the liquid mixture.

[0089] The liquid mixture is then homogenised.

[0090] The liquid mixture can then be heat treated to reduce bacterial load, for example by rapidly heating the liquid mixture to a temperature in the range of from about 80°C to about 150°C for a duration of between about 5 seconds and about 5 minutes. This can be carried out by steam injection, an autoclave or a heat exchanger (e.g. a plate heat exchanger).

[0091] The liquid mixture is then cooled, for example by flash cooling, to between about 60°C and about 85°C. The liquid mixture is then homogenised again, for example in two stages, between about 10 MPa and about 30 MPa in the first stage and between about 2 MPa and about 10 MPa in the second stage. The homogenised mixture can then be further cooled in order to add any heat sensitive components, such as vitamins and minerals. The pH and solids content of the homogenised mixture are conveniently adjusted at this point.

[0092] If the final product is to be a powder, the homogenised mixture is transferred to a suitable drying apparatus, such as a spray drier or freeze drier and converted into a powder. The moisture content of the powder should be less than about 5% by weight. The mixture can be spray dried or freeze dried.

[0093] If a liquid composition is preferred, the homogenised mixture can be sterilised and then filled into suitable containers under aseptic conditions or filled into containers first and then retorted.

[0094] The nutritional composition can be provided, for example, immediately after birth of the infant. The nutritional composition of the present application can also be given to an infant or young child during the first period of life, or during the first 2 periods of life, or during the first 3 periods of life, or during the first month of life, or during the first 2 months of life, or during the first 3 months of life, or during the first 4 months of life, or during the first 6 months of life, or during the first 8 months of life, or during the first 10 months of life, or during the first year of life, or during the first two years of life, or even longer. In some particularly advantageous embodiments of the present application, the composition is given to (or administered to) an infant or young child from about 6 months of age of the infant or young child. For example, the composition can be given from about 6 months, about 10 months, about 12 months, about 14 months, about 16 months, about 20 months, about 24 months or about 36 months of age.

[0095] Suitably, the composition is given to (or administered to) an infant or young child from about 10 months of age of the infant or young child.

[0096] Suitably, the composition can be administered to an infant or young child from about 6 months of age to about 60 months of age, from about 6 months of age to about 48 months of age, from about 6 months of age to about 36 months of age. Suitably, the composition can be administered to an infant or young child from about 10 months of age to about 60 months of age, from about 10 months of age to about 48 months of age or from about 10 months of age to about 36 months of age.

[0097] In one embodiment, the nutritional composition is given to an infant or young child as a supplementary composition to breast milk. In some embodiments, the infant or young child receives breast milk for at least the first 2 weeks, the first 1 month, the first 2 months, the first 4 months, or the first 6 months. In one embodiment, the nutritional composition of the present invention is given to the infant or young child after this period of being nutritious with breast milk, or given to the infant or young child together with breast milk during this period of being nutritious with breast milk. In another embodiment, the nutritional composition is given to the infant or young child as the sole or primary nutritional composition for at least a period of time (e.g., after the first, second, or fourth month of life), for at least 1 month, 2 months, 4 months, or 6 months.

[0098] Suitably, the composition may contain probiotics, including Bifidobacterium pseudochain.

[0099] Suitably, the composition may contain probiotics, including Streptococcus thermophilus.

[0100] The term "probiotics" refers to microbial cell preparations or microbial cell components that have beneficial effects 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; Hill C et al., Nat Rev Gastroenterol Hepatol. 2014). The microbial cells are generally bacteria or yeast.

[0101] On a dry weight basis, *Bifidobacterium pseudochain* can, for example, be approximately 10 per gram of composition. 3 cfu to 10 12 CFU probiotic strains, more preferably between 10 7 CFU and 10 12 CFU between, such as between 10 8 CFU and 10 10 The amount of probiotic strains between CFU is included in the composition. In one embodiment, *Bifidobacterium pseudochain* is live. In some other embodiments, both live and inactivated *Bifidobacterium pseudochain* may be present.

[0102] On a dry weight basis, Streptococcus thermophilus can, for example, be about 10 per gram of composition. 3 cfu to 10 12 CFU probiotic strains, more preferably between 10 7 CFU and 10 12 CFU between, such as between 10 8cfu and 10 10 The amount of probiotic bacterial strain between cfu is included in the composition. In one embodiment, the Streptococcus thermophilus is live. In some other embodiments, both live Streptococcus thermophilus and inactivated Streptococcus thermophilus can be present.

[0103] The term "cfu" is to be understood as colony forming units.

[0104] Suitably, the present application can include the use of a combination of a nutritional or dietary composition as described herein and a probiotic composition as described herein (e.g. a probiotic composition comprising B. pseudocatenulatum and / or S. thermophilus). Suitably, the present application can include the use of a synbiotic. As used herein, a synbiotic can refer to a mixture comprising a microorganism and a substrate selectively utilized by the microorganism, preferably wherein the combination confers a health benefit to a host (see for example Swanson KS et al. Nat Rev Gastroenterol Hepatol. 2020). For example, a synbiotic of the present application can comprise an HMO as a prebiotic and B. pseudocatenulatum and / or S. thermophilus as a probiotic. Suitably, a synbiotic of the present application can comprise an HMO as a prebiotic and B. pseudocatenulatum as a probiotic.

[0105] The composition for use according to the present application can be administered by any suitable method. Preferably, the composition is for oral administration. Thus, the composition is preferably orally administered.

[0106] In some embodiments, the composition according to the present application can be used in the pre- and / or during the weaning period. The age and duration of administration (or giving or feeding) of the nutritional composition can be determined as required.

[0107] Suitably, where a combination (e.g. a combination of a prebiotic and a probiotic as described herein) is administered to a subject, the combination can be administered separately, simultaneously or sequentially.

[0108] Suitably, the composition can be a Nan Pelargon ® .

[0109] Bifidobacterium pseudolongum

[0110] Bifidobacterium are gram-positive, non-motile, usually branched, anaerobic bacteria. They are ubiquitous inhabitants of the gastrointestinal tract and constitute one of the major bacterial genera of the mammalian gastrointestinal microbiota.

[0111] Bifidobacterium has a unique fructose-6-phosphate phosphoketolase pathway for fermenting carbohydrates. Much of the metabolic research on Bifidobacterium has focused on oligosaccharide metabolism, as these carbohydrates are available in their otherwise nutritionally limited habitats. The infant-associated Bifidobacterium lineages appear to have evolved the ability to ferment milk oligosaccharides, while the adult-associated species use plant oligosaccharides, which is consistent with their encounter situations in their respective environments.

[0112] A typical strain of Bifidobacterium pseudocatenulatum is ATCC 27919. A reference genome for Bifidobacterium pseudocatenulatum is provided by GenBank assembly accession number: GCF_020541885.1.

[0113] Suitably, Bifidobacterium pseudocatenulatum can comprise a 16S rRNA sequence having a 99% cutoff identity value and minimum query and 80% target coverage when compared to the 16S rRNA sequence of ATCC 27919 and / or GCF_020541885.1 using BLASTn. Suitable comparisons can be made, for example, using known methods as described by Maturana and Cárdenas (Front Microbiol. 2021; 6: 69020).

[0114] Suitably, Bifidobacterium pseudocatenulatum can have an ANI (average nucleotide identity) of at least 95%, a TETRA (tetranucleotide frequency) of at least 0.99 and / or an AAI (average amino acid identity) of at least 95% compared to ATCC 27919 and / or GCF_020541885.1 for the entire genomic dataset. Suitably, Bifidobacterium pseudocatenulatum microorganisms can have an ANI (average nucleotide identity) of at least 95%, a TETRA (tetranucleotide frequency) of at least 0.99 and an AAI (average amino acid identity) of at least 95% compared to ATCC 27919 and / or GCF_020541885.1 for the entire genomic dataset. Suitable comparisons can be made, for example, using known methods as described by Maturana and Cárdenas (as above).

[0115] Suitably, Bifidobacterium pseudocatenulatum can be identified using metagenomic methods. Suitable metagenomic methods can be performed, for example, using shotgun sequencing data. Metagenomic methods can also advantageously enable estimation of biological relative abundance. 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.

[0116] In some embodiments, the B. pseudocatenulatum is isolated from a human.

[0117] Suitably, "promoting B. pseudocatenulatum" means increasing the absolute or relative number of B. pseudocatenulatum in the gut microbiota. For example, a prebiotic can assist or support the growth and / or survival of the microorganism. Alternatively, a probiotic composition for use in the present disclosure will comprise B. pseudocatenulatum and thereby increase the number of B. pseudocatenulatum within the gut microbiota.

[0118] The abundance of B. pseudocatenulatum in the gut microbiota can be determined, for example, by assessing the relative abundance of B. pseudocatenulatum in a sample from a subject using a metagenomic method as described herein.

[0119] The level of B. pseudocatenulatum can be compared to a reference value determined prior to administration of a composition as described herein. The reference value can be determined prior to the first administration of a composition as described herein, or after the first administration but prior to a subsequent administration of a composition as described herein.

[0120] The methods described herein are typically performed in vitro on a human or animal, for example on a sample previously obtained from a subject to be tested. Preferably, the sample is a fecal sample.

[0121] By way of example, the composition can increase the abundance of B. pseudocatenulatum by at least 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 50-fold or 100-fold compared to, for example, the abundance of B. pseudocatenulatum prior to administration of the composition.

[0122] B. pseudocatenulatum has been reported to enhance bone mineral density (BMD) by reducing bone resorption and increasing bone formation (Fernandez-Murga et al.; Bone; 2020; 141; 115580), as well as reversing hyperleptinemia and restoring leptin signaling in obese mice (Agusti et al.; Mol. Neurobiol. 55 (6), 5337-5352).

[0123] The present invention provides a B. pseudocatenulatum microorganism for use in the prevention and / or treatment of growth retardation in an infant or young child. Thus, the B. pseudocatenulatum can be provided as a probiotic, as defined herein.

[0124] Suitably, the B. pseudocatenulatum can be provided in a composition as defined herein. The composition can be for use in the prevention and / or treatment of growth retardation in an infant or young child. The composition can further comprise a prebiotic, such as a prebiotic as described herein.

[0125] Streptococcus thermophilus

[0126] The present application can include the use of Streptococcus thermophilus as a probiotic or in a composition as described herein.

[0127] The present application can also include a combination comprising the use of Streptococcus thermophilus as a probiotic or in a composition as described herein.

[0128] Streptococcus thermophilus is a gram-positive bacterium and is a fermentative facultative anaerobe of the green cluster. Streptococcus thermophilus tests negative for cytochromes, oxidases, and catalase, and is positive for alpha-hemolytic activity. Streptococcus thermophilus is non-motile and does not form endospores. Streptococcus thermophilus is also classified as a lactic acid bacterium.

[0129] A typical strain of Streptococcus thermophilus is ATCC 19258. A reference genome for Streptococcus thermophilus is provided by GenBank assembly accession number: GCA_903886475.1.

[0130] Suitably, Streptococcus thermophilus can comprise a 16S rRNA sequence having a 99% cutoff identity value and a minimum query and 80% target coverage when compared to the 16S rRNA sequence of ATCC 19258 and / or GCA_903886475.1 using BLASTn. For example, suitable comparisons can be made using known methods as described by Maturana and Cárdenasm (Front Microbiol. 2021; 660920).

[0131] Suitably, Streptococcus thermophilus can have at least 95% ANI (average nucleotide identity), at least 0.99 TETRA (tetranucleotide frequency), and / or at least 95% AAI (average amino acid identity) to ATCC 19258 and / or GCA_903886475 for the entire genome dataset. Suitably, Streptococcus thermophilus microorganisms can have at least 95% ANI (average nucleotide identity), at least 0.99 TETRA (tetranucleotide frequency), and at least 95% AAI (average amino acid identity) to ATCC 19258 and / or GCA_903886475 for the entire genome dataset. For example, suitable comparisons can be made using known methods as described by Maturana and Cárdenasm (supra).

[0132] Suitably, a Streptococcus thermophilus can be identified using metagenomics methods. For example, suitable metagenomics methods can be performed using shotgun sequencing data. Metagenomics methods can also advantageously enable estimation of biological relative abundance. Suitable metagenomics 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 metagenomics sequencing.

[0133] In some embodiments, the Streptococcus thermophilus is isolated from a human.

[0134] Suitably, “promoting Streptococcus thermophilus” means increasing the absolute or relative number of Streptococcus thermophilus in the gut microbiota. For example, a prebiotic can assist or support the growth and / or survival of a microorganism. Alternatively, a probiotic composition used in the present application can comprise Streptococcus thermophilus and thereby increase the number of Streptococcus thermophilus within the gut microbiota.

[0135] The abundance of Streptococcus thermophilus in the gut microbiota can be determined, for example, by assessing the relative abundance of Streptococcus thermophilus in a sample from a subject using metagenomics methods as described herein.

[0136] The level of Streptococcus thermophilus can be compared to a reference value determined prior to administration of a composition as described herein. The reference value can be determined prior to first administration of a composition described herein, or after first administration but prior to subsequent administration of a composition described herein.

[0137] By way of example, a composition can increase the abundance of Streptococcus thermophilus by at least 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 50-fold or 100-fold compared to, for example, the abundance of Streptococcus thermophilus prior to administration of the composition.

[0138] Prebiotic

[0139] Suitably, the composition can comprise a prebiotic.

[0140] The term “prebiotic” means a non-digestible carbohydrate that has a beneficial effect on the host by selectively stimulating the growth and / or activity of healthy bacteria in the human colon (Gibson GR et al. Nat Rev Gastroenterol Hepatol. 2017).

[0141] Suitably, the prebiotic is provided in the form of a dietary fibre. For example, the dietary fibre can be a prebiotic fibre.

[0142] Suitably, the prebiotic can be comprised in an ingredient (e.g. a dietary ingredient).

[0143] The ingredient can be selected from the group consisting of human milk oligosaccharides (HMOs), purified polysaccharides or purified oligosaccharides, dietary fibre ingredients, semi-purified food ingredients, raw food ingredients, food additives, semi-purified or purified peptidoglycans.

[0144] Suitably, the prebiotic is an HMO.

[0145] Suitably, the prebiotic composition comprises a prebiotic that promotes Bifidobacterium pseudocatenulatum in the gut microbiota (e.g. comprised in an ingredient or fibre). Suitably, the prebiotic composition comprises a prebiotic that promotes Streptococcus thermophilus in the gut microbiota (e.g. comprised in an ingredient or fibre).

[0146] The prebiotic or composition can be provided as a fermented dairy product (e.g. a yoghurt). Fermented dairy products such as yoghurt have been shown to increase the abundance of Streptococcus thermophilus in the gut microbiota (see for example Pasoli et al; Nat Comm; 2000; 11(1); 2610; Oyarzun et al; Comput Struct Biotechnol J; 2022; 5(2); 1632-1641, Yazdi et al; Journal of Functional Foods; 2022: 105089).

[0147] The composition can comprise oligosaccharides (e.g. human milk oligosaccharides) and / or at least fibres thereof and / or at least precursors thereof. The oligosaccharides and / or fibres and / or precursors thereof can be selected from the list consisting of galactooligosaccharides (GOS), fructooligosaccharides (FOS), inulin, xylooligosaccharides (XOS), polydextrose and any combination thereof. Their amount can be comprised between 0 and 10 wt% of the composition. In a particular embodiment, the nutritional composition can further comprise at least one BMO (bovine milk oligosaccharide).

[0148] The present application provides a prebiotic for preventing and / or treating growth retardation in an infant or young child. Suitably, the prebiotic promotes Bifidobacterium pseudocatenulatum in the gut microbiota of the infant or young child. Suitably, the prebiotic promotes Streptococcus thermophilus in the gut microbiota of the infant or young child.

[0149] Suitably, the prebiotic promotes Bifidobacterium pseudocatenulatum in the gut microbiota of the infant or young child to prevent and / or treat growth retardation in the infant or young child. Suitably, the prebiotic promotes Streptococcus thermophilus in the gut microbiota of the infant or young child to prevent and / or treat growth retardation in the infant or young child.

[0150] Suitably, a prebiotic can be provided in a composition as defined herein. The composition can be used to prevent and / or treat growth retardation in an infant or young child, for example by promoting B. pseudocatenulatum and / or S. thermophilus in the gut microbiota of the infant or young child. The composition can further comprise a probiotic, such as a B. pseudocatenulatum microorganism and / or a S. thermophilus microorganism as described herein.

[0151] Human milk oligosaccharides (HMOs)

[0152] Suitably, the prebiotic can be an HMO.

[0153] The term "HMO" refers to human milk oligosaccharides. These carbohydrates are highly resistant to enzymatic hydrolysis, which suggests that important functions they perform can not be directly related to their caloric value. It has been specifically pointed out in the art that these carbohydrates play a key role in the early development of infants and young children, such as the maturation of the immune system. Many different kinds of HMOs are found in human milk. Each individual oligosaccharide is based on a combination of glucose, galactose, sialic acid (N-acetylneuraminic acid), fucose and / or N-acetylglucosamine with a variety of linkages between these molecules, and thus human milk contains a large number of oligosaccharides that differ in structure, more than 130 such structures have been identified to date. The reducing end of almost all oligosaccharides has a lactose molecule, and the terminal position of the non-reducing end is occupied by sialic acid and / or fucose, if any. Depending on the presence of fucose and sialic acid in the structure of the oligosaccharide, HMOs can be classified as non-fucosylated (neutral) or fucosylated (neutral) and sialylated (acidic) and non-sialylated molecules, respectively.

[0154] Suitably, the term "able to metabolize HMOs" can mean that the B. pseudocatenulatum encodes at least one CAZyme that is able to utilize HMOs. For example, the CAZyme can be able to catalyze the hydrolysis of glycosidic bonds within HMOs. Suitably, the B. pseudocatenulatum can encode at least one, at least two, at least three, at least four or at least five CAZymes that are able to utilize HMOs. Suitably, the term "able to metabolize HMOs" can mean that the HMOs are able to promote the growth and / or survival of the B. pseudocatenulatum (e.g. when added to an anaerobic bacterial culture of the B. pseudocatenulatum). The growth and / or survival of the B. pseudocatenulatum can be determined, for example, using a PCR method by measuring the abundance of 16S rDNA.

[0155] An HMO that is capable of promoting the growth and / or survival of B. pseudocatenulatum can increase the number of B. pseudocatenulatum 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 B. pseudocatenulatum in a control anaerobic culture that does not comprise an HMO. Suitably, an HMO that is capable of promoting the growth and / or survival of B. pseudocatenulatum can increase the number of B. pseudocatenulatum in an anaerobic culture by a statistically significant amount (e.g., p-value < 0.05 as determined by one-way ANOVA) compared to the number of B. pseudocatenulatum in a control anaerobic culture that does not comprise an HMO.

[0156] The foregoing disclosure with respect to B. pseudocatenulatum can equally apply to S. thermophilus.

[0157] The expression "fucosylated oligosaccharides" refers to oligosaccharides with fucose residues. Such oligosaccharides are neutral. Some examples are 2'-fucosyllactose (2-FL), 3-fucosyllactose (3-FL), difucosyllactose (DiFL), lacto-N-fucopentaose (e.g., lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto-N-fucopentaose III, lacto-N-fucopentaose V), lacto-N-fucopentaose, lacto-N-difucohexaose I, fucosyl lacto-N-hexaose, fucosyl lacto-N-neohexaose, difucosyl lacto-N-hexaose I, difucosyl lacto-N-neohexaose II, and any combination thereof. Fucosylated oligosaccharides represent the largest fraction 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 specific commensal microorganisms, thereby reducing the inflammatory response.

[0158] The expression "N-acetylated oligosaccharides" encompasses "N-acetyl lactosamine" and "oligosaccharides comprising N-acetyl lactosamine". Such oligosaccharides are neutral oligosaccharides with N-acetyl-amino lactosamine residues. Suitable examples are LNT (lacto-N-tetraose), para-lacto-N-neohexaose (para-LNnH), LNnT (lacto-N-neotetraose), DSLNT (disialyllacto-N-tetraose), and any combination thereof. Other examples are lacto-N-hexaose, lacto-N-neohexaose, para-lacto-N-hexaose, para-lacto-N-neohexaose, lacto-N-octaose, lacto-N-neooctaose, iso-lacto-N-octaose, para-lacto-N-octaose, and lacto-N-decaose.

[0159] The expressions "at least one fucosylated oligosaccharide" and "at least one N-acetylated oligosaccharide" are to be understood as "at least one type of fucosylated oligosaccharide" and "at least one type of N-acetylated oligosaccharide".

[0160] The term "sialylated oligosaccharide" refers to an oligosaccharide having a charged sialic acid residue. Such oligosaccharides are acidic. Some examples are 3'-sialyllactose (3-SL), 6'-sialyllactose (6-SL), sialyllactose-N-tetraose (Lst, e.g. Lst-a, Lst-b or Lst-c).

[0161] The HMO can be a fucosylated oligosaccharide (i.e. an oligosaccharide having a fucose residue; e.g. 2'fucosyllactose (2-FL), 3-fucosyllactose (3-FL), difucosyllactose (DiFL), lactose-N-fucosylpentose (e.g. lactose-N-fucosylpentose I, lactose-N-fucosylpentose II, lactose-N-fucosylpentose III, lactose-N-fucosylpentose V), lactose-N-fucalose, lactose-N-difucalose I, fucosyllactose-N-fucalose, fucosyllactose-N-neofucalose, difucosyllactose-N-fucalose I, difucosyllactose-N-neofucalose II and any combination thereof), an N-acetylated oligosaccharide (e.g. LNT (lactose-N-tetraose), para lacto-N-neofucalose (para-LNnH), LNnT (lacto-N-neotetraose), DSLNT (disialyllacto-N-tetraose), lacto-N-fucalose, lacto-N-neofucalose, para-lacto-N-fucalose, para-lacto-N-neofucalose, lacto-N-octaose, lacto-N-neooctaose, iso-lacto-N-octaose, para-lacto-N-octaose and lacto-N-decaose and any combination thereof) and / or a sialylated oligosaccharide (e.g. 3'-sialyllactose (3-SL), 6'-sialyllactose (6-SL) or Lst (sialyllactose-N-tetraose), Lst-a, Lst-b or Lst-c).

[0162] Suitably, the prebiotic can comprise an HMO, which can be selected from the group consisting of 2'-FL, di-FL, 6'-SL and LNnT and any combination thereof. The HMO can be a combination of 2'-FL and di-FL. The HMO can be a combination of 6'-SL and LNnT.

[0163] The HMO can be provided in combination with galacto-oligosaccharides (GOS).

[0164] The prebiotic can comprise at least one prebiotic oligosaccharide selected from the group consisting of 2'-O-fucosyllactose (2'FL), 3'-O-fucosyllactose (3FL), lactodifucotetraose / lactodifucosyllactose (DFL), 3'-O-sialyllactose (3-SL), 6'-O-sialyllactose (6-SL), lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT); and any combination thereof.

[0165] The prebiotic can comprise at least one prebiotic oligosaccharide selected from the group consisting of 3FL, 3'SL, LNnT, lacto-N-fucopentaose (LNFP I), LNFP II, LNFP III, sialyllactose-N-tetraose (LST) b, LST c, disialyllactose-N-tetraose (DSLNT), fucosyllactose-N-hexaose (FLNH), difucosyllactose-N-hexaose (DFLNH), and disialyllactose-N-hexaose (DSLNH); and any combination thereof.

[0166] The prebiotic can comprise 34 to 85 wt% 2'-FL, 10 to 40 wt% LNT, 4 to 14 wt% DFL, and 9 to 31 wt% of a combination of 3-SL and 6-SL.

[0167] In some embodiments, the prebiotic comprises

[0168] - 26 to 65 wt%, preferably 32 to 54 wt% 2'-FL;

[0169] - 10 to 40 wt%, preferably 11 to 20 wt% LNT;

[0170] - 4 to 14 wt%, preferably 4 to 8 wt% DFL;

[0171] - 9 to 31 wt%, preferably 8 to 22 wt% of a combination of 3'-SL and 6'-SL; and

[0172] - 12 to 38 wt%, preferably 17 to 31 wt% 3-FL.

[0173] The prebiotic can comprise between 0.001 g / L to 12 g / L of 2'-FL, preferably between 0.002 g / L to 10 g / L of 2'-FL, more preferably between 0.005 g / L to 5 g / L of 2'-FL.

[0174] The prebiotic can comprise between 0.001 g / L to 5 g / L of DFL, preferably between 0.002 g / L to 4 g / L of DFL, more preferably between 4 g / L to 3 g / L of DFL.

[0175] The prebiotic can comprise between 0.01 g / L to 6 g / L of LNT, preferably between 0.025 g / L to 5 g / L of LNT, more preferably between 0.05 g / L to 1 g / L of LNT.

[0176] The prebiotic can comprise between 0.001 g / L and 2 g / L of 6'-SL, preferably between 0.002 g / L and 1.5 g / L of 6'-SL, more preferably between 0.005 g / L and 1 g / L of 6'-SL.

[0177] The prebiotic can comprise between 0.01 g / L and 2 g / L of 3'-SL, preferably between 0.025 g / L and 1.5 g / L of 3'-SL, more preferably between 0.05 g / L and 1 g / L of 3'-SL.

[0178] The prebiotic can comprise between 0.01 g / L and 7 g / L of 3-FL, preferably between 0.025 g / L and 6 g / L of 3-FL, more preferably between 0.05 g / L and 5 g / L of 3-FL.

[0179] Suitably, the mixture of oligosaccharides comprises or consists of 2'-fucosyllactose (2'FL), difucosyllactose (diFL), lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT). In some embodiments, the mixture of oligosaccharides comprises or consists of 3'-sialyllactose (3'-SL), 6'-sialyllactose (6'-SL), 2'-fucosyllactose (2'-FL), difucosyllactose (diFL), lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT).

[0180] In some embodiments, the mixture of oligosaccharides comprises:

[0181] - 10 to 35 wt%, preferably 10 to 30 wt%, more preferably 10 to 25 wt% of at least one sialylated oligosaccharide relative to the total weight of the oligosaccharide mixture;

[0182] - 30 to 80 wt%, preferably 40 to 80 wt%, more preferably 50 to 70 wt% of at least one fucosylated oligosaccharide relative to the total weight of the oligosaccharide mixture; and / or

[0183] - 10 to 35 wt%, preferably 15 to 30 wt%, more preferably 15 to 20 wt% of at least one N-acetylated oligosaccharide relative to the total weight of the oligosaccharide mixture.

[0184] The present application provides an HMO or a combination of HMOs for preventing and / or treating growth retardation in an infant or young child. Suitably, the HMO promotes Bifidobacterium pseudocatenulatum and Streptococcus thermophilus in the gut microbiota of the infant or young child.

[0185] Suitably, the HMOs can be provided in a composition as defined herein. The composition can be for use in the prevention and / or treatment of growth retardation in an infant or young child. The composition can further comprise a probiotic, such as a Bifidobacterium pseudocatenulatum microorganism as described herein.

[0186] HMOs have been shown to increase the abundance of Bifidobacterium pseudocatenulatum in the microbiota of infants (e.g. Cheema et al; Int J Mol Sci; 2022; 23(5):2804 and examples of the present application). It has also been reported that levels of different HMOs in breast milk are associated with infant growth, including length / height (Samuel TM et al. Sci Rep. 2022).

[0187] Microbiota and microbiome

[0188] “Gut microbiota” can refer to the composition of microorganisms (including bacteria, archaea and fungi) living in the digestive tract.

[0189] The term “gut microbiome” can include the “gut microbiota” and their “habitat”, which can include their structural elements (nucleic acids, proteins, lipids, polysaccharides), metabolites (signalling molecules, toxins, organic and inorganic molecules) and molecules produced by the cohabiting host and structured by the surrounding environmental conditions (see e.g. Berg, G. et al. 2020. Microbiome, 8(1), pp. 1-22).

[0190] Thus, in the present application, the term “gut microbiome” can be used interchangeably with the term “gut microbiota”.

[0191] Method

[0192] The present application also provides a method for predicting or assessing whether an infant or young child is at risk of growth retardation, the method comprising determining the level of Bifidobacterium pseudocatenulatum in one or more samples obtained from the infant or young child.

[0193] Without wishing to be bound by theory, the present inventors have determined that infants or young children having lower levels (e.g. abundance and / or activity) of Bifidobacterium pseudocatenulatum in their microbiome can have an increased likelihood of growth retardation.

[0194] The level of Bifidobacterium pseudocatenulatum can be compared to a reference value, wherein the comparison is indicative of a predicted risk of growth retardation in the infant or young child. The term reference level is synonymous with “control level” and broadly includes data that would be used by a person skilled in the art to facilitate accurate interpretation of technical data.

[0195] The reference value can be based on a value (e.g. mean) of B. pseudocatenulatum in a population of infants and / or young children known to be at risk of, or suffering from, growth retardation. The reference value can be based on a value (e.g. mean) of B. pseudocatenulatum in a population of infants and / or young children known not to be at risk of, or suffering from, growth retardation. The reference value can be based on a value (e.g. mean) of B. pseudocatenulatum in a population of infants and / or young children known not to be at risk of, or suffering from, growth retardation.

[0196] The reference level can be age matched to the test sample.

[0197] Suitably, the infant or young child can be about 1 month old to 60 months old, about 1 month old to 48 months old, about 2 months old to 60 months old, about 2 months old to 48 months old, about 2 months old to 36 months old, or about 4 months old to 36 months old, about 6 months old to 36 months old or about 6 months old to 24 months old.

[0198] Preferably, the infant or young child can be at least 10 months old.

[0199] For example, the infant or young child can be at least about 10 months old, at least about 12 months old, at least about 14 months old, at least about 16 months old, at least about 20 months old or at least about 24 months old.

[0200] Preferably, the infant or young child can be about 10 months old to about 48 months old, about 10 months old to about 36 months old, about 10 months old to about 24 months old, about 10 months old to about 18 months old.

[0201] The method of the application is typically performed outside the human or animal body, for example on a sample previously obtained from a subject to be tested. Preferably, the sample is a faecal sample.

[0202] Suitably, the method of the application provides that a difference in the level of B. pseudocatenulatum in the test sample compared to the reference level is indicative of a risk of growth retardation. Suitably, the method of the application can provide that a difference in the level of B. pseudocatenulatum in the test sample compared to the reference level is indicative of an increased risk of growth retardation.

[0203] For example, a difference of 1.1 fold, 1.5 fold, 2 fold, 3 fold, 4 fold, 5 fold, 10 fold, 50 fold or 100 fold between the determined level in the test sample and the reference level can be indicative of an increased risk of growth retardation.

[0204] Suitably, a reduced level of B. pseudocatenulatum is associated with an increased risk of growth retardation. Suitably, an infant or young child having a reduced level of B. pseudocatenulatum is identified as being at risk of growth retardation.

[0205] For example, a 1.1 fold, 1.5 fold, 2 fold, 3 fold, 4 fold, 5 fold, 10 fold, 50 fold or 100 fold lower level of B. pseudocatenulatum determined in the test sample compared to the reference level can be indicative of an increased risk of growth faltering.

[0206] Suitably, the method further comprises combining the level of B. pseudocatenulatum with one or more anthropometric measures.

[0207] Suitably, an infant or young child determined to be at risk of growth faltering using the method of the application can be treated with a composition according to the application to reduce the risk of growth faltering occurring and / or prevent growth faltering.

[0208] Suitably, the method of the application can comprise determining the level of Streptococcus thermophilus as a proxy for B. pseudocatenulatum. Suitably, the method of the application can comprise determining the level of B. pseudocatenulatum and Streptococcus thermophilus.

[0209] The method can be performed on one or more samples obtained from a subject. For example, the method can be performed using a first sample obtained at a given time point and a second sample obtained after a time interval following the first sample. The method can be performed more than once on samples obtained from the same subject over a period of time. For example, the samples can be obtained once a month, once a year or once every two years.

[0210] Examples

[0211] The application will now be further described by way of Examples, which are meant to serve to assist one of ordinary skill in the art in carrying out the application, and are not intended to limit the scope of the application in any way.

[0212] Example 1 : Microbiota and health study

[0213] Microhealth clusters were described by Vidal et al. (https: / / www.medrxiv.org / content / 10.1101 / 19000505vl) and registered at clinicaltrials.gov as NCT02361164.

[0214] In short, for n=220 infants or young children, the following information was collected: anthropometry, dysentery and acute respiratory infections (ARI); medication, including antibiotic use; breastfeeding status, complementary foods; nasopharyngeal and stool pathogens and microbiota profiling; secretor status (FUT2, FUT3). Stool samples for microbiota analysis were collected at birth, 2 months, 6 months, 10 months, 15 months, 18 months and 24 months. Growth outcomes were recorded at birth, 2 months, 4 months, 6 months, 8 months, 10 months, 12 months, 15 months, 18 months and 24 months. For an overview, see Figure 1 .

[0215] Example 2: Definition of reference population and growth insufficient population

[0216] Based on the WHO guidelines, growth faltering was defined by (i) dynamic changes or (ii) static outcomes.

[0217] For dynamic changes, the change in length-for-age z-score (LAZ) over time (6 months to 24 months) was assessed. We identified three patterns: little change in LAZ score over time (negligible slope), a decline in LAZ score over time (negative slope) and an improvement in LAZ score over time (positive slope). Infants with a change in LAZ score over time (calculated as slope) < -0.0485434516523868 were called “growth faltering” (n=48). Infants with a change in LAZ score over time (calculated as slope) > 0.00495716034271725 were called the reference population (n=48). The microbiota of infants in the first quartile, whose length-for-age z-score (LAZ) changed from 6 months to 24 months, were compared to the microbiota of infants in the fourth quartile, whose growth improved.

[0218] For static outcomes, the LAZ score at 24 months was assessed and length_for_age_24m < -2 was defined as “growth faltering” and length_for_age_24m > -2 was defined as the reference population.

[0219] Example 3: Microbiota differences between growth insufficient population and reference population

[0220] For dynamic changes, microbiota data were acquired only for samples corresponding to 6 months to 24 months. First, we filtered out any bacterial species with a relative abundance less than 0.01% of the mean. Thus, from about 750 species, 168 species remained after this step. Next, we removed features with near-zero variance using the nearZeroVar function of the mixOmics R package with the parameters freqCut = 95 / 05, uniqueCut = 20. After this step, 78 species remained (see Figure 2 ).

[0221] To determine if there was an overall microbiota difference between the two groups, “reference” vs. “stunted” as defined above, we used a machine learning algorithm: sPLS-DA (Le Cao KA et al. BMC Bioinformatics. 2011). Feature selection was performed in 10-fold M-fold mode to find discriminating bacteria with a centroid distance measure repeated 10 times to keep a balanced error rate between the two classes. To identify time intervals of differentially abundant features in a metagenomic longitudinal study, we used the R package: MetaLonDA (Metwally AA et al. Microbiome. 2018). We ran MetaLonDA in “screening mode” with 100 permutations. Hits identified from this were confirmed by re-running MetaLonDA with the recommended 1000 permutations.

[0222] For static results, similar technical methods as above were performed to identify bacteria found significantly different across time between the two groups (length_for_age_24m: < -2 for “stunted”; > -2 for “reference” (see Figure 3 and Figure 4 ).

[0223] Common hits were identified by multiple algorithms that were associated with dynamic changes in age-adjusted length z-score (LAZ) (6 months to 24 months) and static measures of laz < -2 or > -2 (at 24 months). On multiple lines of evidence, B. pseudocatenulatum and S. thermophilus were identified as bacterial features of insufficient height-for-age distribution up to 24 months.

[0224] Example 4: Effect of galacto-oligosaccharides and HMOs on Bifidobacterium pseudolongum in infant microbiome

[0225] Using in vitro D-SIFR ® Technical simulations of the gut microbiota of three 3-year-old toddlers, testing samples with combinations of galacto-oligosaccharides (GOS), HMO1 (2’FL and diFL) and / or HMO2 (LNnT and 6’SL).

[0226] Treatment with GOS, HMO1 and / or HMO2 was found to have a significant bifidogenic effect, including promoting levels of Bifidobacterium pseudocatenulatum. For example, treatment with HMOs (+ / - GOS) increased levels of Bifidobacterium pseudocatenulatum by at least 2-fold.

[0227] Materials and methods

[0228] Fecal samples were collected according to procedures approved by the Ghent University Hospital Ethics Committee.

[0229] Colonic fermentation of test products by gut microbiota was evaluated in fecal samples 24 hours after inoculation.

[0230] To statistically evaluate the treatment effect on the basic fermentation parameters, cell counts, microbial diversity and microbial composition (phylum level) of samples from 3 infants, a repeated measures ANOVA analysis was performed (based on paired t-tests, hence accounting for the fact that values are compared between samples of a given donor). Statistical significance of potential treatment effects was determined by Benjamini-Hochberg post-test.

[0231] For quantitative shallow shotgun sequencing, standard Illumina library preparation was performed after DNA extraction, followed by sequencing of 3M total DNA.

[0232] All publications mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the disclosed method, composition and use of the application will be apparent to those skilled in the art without departing from the scope and spirit of the application. Although the application has been disclosed with reference to specific preferred embodiments, it is apparent that the application can be practiced with

[0233] Embodiments

[0234] Various preferred features and embodiments of the present application will now be described with reference to the following numbered paragraphs (Paragraphs).

[0235] 1. A method for identifying an infant or young child at risk of growth faltering, wherein the method comprises determining the abundance of Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus in one or more samples obtained from the infant.

[0236] 2. The method of paragraph 1, wherein an infant or young child having a reduced level of B. pseudocatenulatum is identified as being at risk of stunted growth.

[0237] 3. The method of paragraph 1 or 2, further comprising determining the abundance of B. pseudocatenulatum and S. thermophilus in one or more samples obtained from the infant.

[0238] 4. The method of paragraph 3, wherein an infant or young child having a reduced level of S. thermophilus is identified as being at risk of stunted growth.

[0239] 5. The method of any preceding paragraph, wherein the stunted growth is associated with malnutrition.

[0240] 6. The method of any preceding paragraph, wherein the stunted growth is short stature or length.

[0241] 7. The method of paragraph 6, wherein the short stature or length is defined as a reduced length-for-age z-score (LAZ) or a reduced LAZ over time, or a reduced height-for-age z-score (HAZ) or a reduced HAZ over time.

[0242] 8. A composition for use in preventing and / or treating stunted growth in an infant or young child, wherein the composition promotes B. pseudocatenulatum and / or S. thermophilus in the gut microbiota of the infant or young child.

[0243] 9. The composition for use of paragraph 8, wherein the composition promotes B. pseudocatenulatum in the gut microbiota of the infant or young child; optionally, wherein the composition comprises a B. pseudocatenulatum microorganism.

[0244] 10. The composition for use of paragraph 8 or 9, wherein the composition promotes S. thermophilus in the gut microbiota of the infant or young child; optionally, wherein the composition comprises a S. thermophilus microorganism.

[0245] 11. The composition for use of paragraph 9 or 10, wherein the composition is administered in combination with a prebiotic.

[0246] 12. The composition for use of paragraph 8, wherein the composition comprises a prebiotic.

[0247] 13. The composition for use of paragraph 12, wherein the composition is administered in combination with a B. pseudocatenulatum and / or S. thermophilus microorganism.

[0248] 14. A combination of Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus microorganism and prebiotic for use in the prevention and / or treatment of growth retardation in an infant or young child.

[0249] 15. The composition or combination for use according to any one of paragraphs 11 to 14, wherein the prebiotic is in the form of a dietary or nutritional composition.

[0250] 16. The composition or combination for use according to any one of paragraphs 11 to 15, wherein the prebiotic comprises a human milk oligosaccharide (HMO).

[0251] 17. A human milk oligosaccharide (HMO) or combination of HMOs for use in the prevention and / or treatment of growth retardation in an infant or young child, wherein the HMO or combination of HMOs promotes Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus in the gut microbiota of the infant or young child.

[0252] 18. The composition or combination for use according to paragraph 16 or the HMO or combination of HMOs for use according to paragraph 17, wherein the HMO is selected from the group consisting of 2’-FL, 3-FL, di-FL, 3’-SL, 6’-SL, LNT and LNnT and any combination thereof; suitably wherein the HMO is (i) a combination of 2’-FL and di-FL or (ii) a combination of 6’-SL and LNnT.

[0253] 19. A Bifidobacterium pseudocatenatatum microorganism for use in the prevention and / or treatment of growth retardation in an infant or young child.

[0254] 20. The composition, combination, HMO or Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus microorganism for use according to any one of paragraphs 8 to 19, wherein the infant or young child has been identified as having a reduced level of Bifidobacterium pseudocatenatatum in the gut microbiota.

[0255] 21. The composition, combination, HMO or Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus microorganism for use according to any one of paragraphs 8 to 19, wherein the infant or young child has been identified as being at risk of growth retardation by a method according to any one of paragraphs 1 to 7.

[0256] 22. The composition, combination, HMO or Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus microorganism for use according to any one of paragraphs 8 to 21, wherein the growth retardation is associated with malnutrition.

[0257] 23. The composition, combination, HMO or Bifidobacterium pseudocatenatatum and / or Streptococcus thermophilus microorganism for use according to any one of paragraphs 8 to 22, wherein the growth retardation is stunted height or length.

[0258] 24. The composition, combination, HMO or B. pseudocatenatatum and / or S. thermophilus microorganism for use according to paragraph 23, wherein the stunted height or length is defined as a reduced length-for-age z-score (LAZ) or reduced LAZ over time, or a reduced height-for-age z-score (HAZ) or reduced HAZ over time.

[0259] 25. The composition, combination, HMO or B. pseudocatenatatum and / or S. thermophilus microorganism for use according to any one of paragraphs 8 to 24, wherein bone development and / or bone strength in the subject is enhanced.

[0260] 26. A method of preventing and / or treating growth retardation in an infant or young child, the method comprising administering a composition that promotes B. pseudocatenatatum and / or S. thermophilus in the gut microbiota to the infant or young child.

[0261] 27. Use of a composition for modulating the abundance of B. pseudocatenatatum and / or S. thermophilus in the gut of an infant or young child.

[0262] 28. The use according to paragraph 27, wherein the composition is a composition as defined in any one of paragraphs 6 to 18.

[0263] 29. Use of an HMO or combination of HMOs for modulating the abundance of B. pseudocatenatatum and / or S. thermophilus in the gut of an infant or young child.

[0264] 31. The method, composition for use, combination for use or use according to any preceding paragraph, wherein the infant or young child is less than about 60 months old, suitably less than 36 months old, less than 24 months old, suitably about 6 months old to about 24 months old.

[0265] 32. The method, composition for use, combination for use or use according to any preceding paragraph, wherein the microorganism is B. pseudocatenatatum.

[0266] 33. A probiotic composition comprising B. pseudocatenatatum.

[0267] 34. The probiotic composition according to paragraph 31, wherein the probiotic composition further comprises S. thermophilus.

[0268] 35. A synbiotic composition comprising B. pseudocatenatatum and a prebiotic.

[0269] 36. The synbiotic composition according to paragraph 33, wherein the prebiotic promotes B. pseudocatenatatum in the gut microbiota of an infant or young child.

[0270] 37. The synbiotic composition of paragraph 35 or 36, wherein the prebiotic is as defined in any one of paragraphs 15 to 18.

Claims

1. A method for identifying an infant or young child at risk of growth retardation, wherein the method comprises determining the abundance of Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus in one or more samples obtained from the infant.

2. The method of claim 1, wherein an infant or young child having a reduced level of Bifidobacterium pseudocatenulatum is identified as being at risk of growth retardation.

3. The method of any preceding claim, wherein the growth retardation is stunted height or length; optionally, wherein the stunted height or length is defined as a reduced length-for-age z-score (LAZ) or a reduced LAZ over time, or a reduced height-for-age z-score (HAZ) or a reduced HAZ over time.

4. A composition for preventing and / or treating growth retardation in an infant or young child, wherein the composition promotes Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus in the gut microbiota of the infant or young child.

5. The composition for use of claim 4, wherein the composition promotes Bifidobacterium pseudocatenulatum in the gut microbiota of the infant or young child; optionally, wherein the composition comprises a Bifidobacterium pseudocatenulatum microorganism.

6. The composition for use of claim 4, wherein the composition comprises a prebiotic.

7. A combination of a Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus microorganism and a prebiotic for preventing and / or treating growth retardation in an infant or young child.

8. The composition or combination for use of any claim 6 or 7, wherein the prebiotic is in the form of a dietary or nutritional composition.

9. The composition or combination for use of claim 8, wherein the prebiotic comprises a human milk oligosaccharide (HMO).

10. A human milk oligosaccharide (HMO) or combination of HMOs for preventing and / or treating growth retardation in an infant or young child, wherein the HMO or combination of HMOs promotes Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus in the gut microbiota of the infant or young child.

11. A Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus microorganism for preventing and / or treating growth retardation in an infant or young child.

12. The composition, combination, HMO or Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus microorganism for use of any one of claims 4 to 11, wherein the infant or young child has been determined to be at risk of growth retardation by the method of any one of claims 1 to 3.

13. The composition, combination, HMO or Bifidobacterium pseudocatenulatum and / or Streptococcus thermophilus microorganism for use of any one of claims 4 to 12, wherein the growth retardation is stunted height or length; optionally, wherein the stunted height or length is defined as a reduced length-for-age z-score (LAZ) or a reduced LAZ over time, or a reduced height-for-age z-score (HAZ) or a reduced HAZ over time.

14. The composition for use, combination, HMO or B. pseudocatenatium and / or S. thermophilus microorganism of any one of claims 4 to 13, wherein bone development and / or bone strength in the subject is enhanced.

15. A synbiotic composition comprising B. pseudocatenatium and / or S. thermophilus and a prebiotic, wherein the prebiotic is an HMO that promotes B. pseudocatenatium and / or S. thermophilus in the gut microbiota of an infant or young child.

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