Synbiotic combination for brain development

Abstract

The invention relates to nutritional compositions and nutritional supplements comprising probiotics, prebiotics, and glutamine for reducing the risk of occurrence and / or treatment of an impaired motor development and for use in preventing long term motor dysfunction. The nutritional compositions and nutritional supplements are in particular suitable for preterm infants.

Description

[0001] SYMBIOTIC COMBINATION FOR BRAIN DEVELOPMENT

[0002] FIELD OF THE INVENTION

[0003] The present invention relates to nutritional compositions comprising synbiotics and glutamine for reducing the risk of occurrence and / or treatment of an impaired motor development and for use in preventing long term motor dysfunction. The invention is in particular suitable for preterm infants.

[0004] BACKGROUND OF THE INVENTION

[0005] Preterm infants, infants born before 37 weeks of gestation, face significantly higher risks of neurodevelopmental impairments compared to term infants. These risks include cognitive deficits, lower IQ scores, and academic challenges that can persist into adolescence and adulthood. Preterm birth is also associated with delays in motor developmental and increased likelihood of cerebral palsy, developmental coordination disorder and perinatal strokes, all of which may impair motor function. The earlier the gestational age at birth, the greater the risk of these adverse outcomes. Preterm infants may experience persistent motor difficulties that can affect their daily activities and overall quality of life.

[0006] Also term infants can have motor development impairments due to brain injury, for example by trauma or hypoxic ischemic encephalopathy (HIE). HIE is caused by impaired cerebral blood flow and oxygen delivery to the brain and can be caused by various maternal and / or pregnancy adversities and other problems at birth such as umbilical cord prolapse, placental abruption or prolonged labor. A recent study shows prevalence of HIE is about 2 in 1000 live births (Park et al. Sci Rep, 2023, 13:23100).

[0007] Preterm infants have significantly higher nutritional needs compared to term infants due to their rapid growth and development requirements. Breast feeding is preferred, but often the nutritional need of the preterm infant is higher. Similarly, infants with neonatal brain injury have higher nutritional demands as the brain needs to be repaired. Human milk fortifiers are available to supplement the human milk, with additional protein, calories, vitamins, and minerals essential for growth and development. When breastfeeding is not possible, preterm infants require specialized formulas, preterm or post-discharge formulas, that provide higher levels of calories, proteins, vitamins, and minerals, when compared to formulas forterm infants.

[0008] Kieviet et al. in BJN, 2012, 108:2215-2220 describes the outcome of neonatal enteral glutamine supplementation in very preterm and / or very low birth weight infants on cognitive, motor and behavioural skills at school age. With one single exception of visuomotor abilities relating to ball skills as measured using the Movement Assessment Battery for Children (MABC), it was found that short term glutamine supplementation had no detrimental effects on long-term cognitive, motor and behavioural outcomes of very preterm and / or VLBW children at school age.WO 2013 / 105851 describes a nutritional composition enriched in glutamine to improve structural and brain development in preterm and / or low birth weight infant. In particular increases in cortical brain volume and subcortical brain volume was observed in children who were born preterm and were supplemented in the first month after birth with L-glutamine when compared to the control group of preterm infants not having received L-glutamine.

[0009] WO 2018 / 143806 describes the results of a nutritional intervention with prebiotic fiber, a probiotic bacterium and glutamine in piglets. Brain white matter microstructure was assessed and increased fractional anisotropy values in the cortico-cortical tract were found which were specifically correlated to cognitive function.

[0010] Keunen et al. in Pediatric Research, 2014 77:148-155 study the impact of nutrition on brain development and mentions that no correlation between an increase in white matter and an improvement in motor development was observed.

[0011] Hortensius et al. in Nutrients, 2021 , 13, 3409 describes an association of higher protein intake by extremely preterm infants and higher fractional anisotropy in several white matter tracts but lower motor scores were observed in infants at 2 years corrected age. There is no mentioning of the corticospinal tract.

[0012] Preterm infants and HIE infants are at risk of adverse motor outcomes, including cerebral palsy, developmental coordination disorder and perinatal strokes. At present no therapeutic application is known to address long term motor development, thereby providing enhanced efficacy and improved patient outcomes.

[0013] SUMMARY OF THE INVENTION

[0014] In a double-blind, randomized clinical trial, preterm infants (24 to < 30 weeks gestational age) were provided with either a combination of Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine or a placebo from birth until 36 weeks postmenstrual age. At term equivalent age (TEA) infants underwent magnetic resonance imaging (MRI). Surprisingly, the infants that received the product according to the invention demonstrated significantly higher Fractional Anisotropy (FA) values specifically in the right corticospinal tract (rCST) compared to infants that received the placebo.

[0015] The corticospinal tract is one of the major fiber pathways in the central nervous system projecting from the brain to the spinal cord. The corticospinal tract plays a crucial role in motor control. Impaired development of the corticospinal tract in infants is indicative of impaired motor functions later in life. Therefore, it wasconcluded that a combination of Bifidobacterium breve, scGOS, IcFOS and glutamine enhances motor development and reduces the risk of motor dysfunction later in life.

[0016] While in the art upon supplementing with prebiotic fiber such as GOS and FOS, probiotic bacterium and glutamine improvements on cognitive functioning were found, it was expected to observe better Fractional Anisotropy (FA) values in area’s linked to cognition, in particular cerebral white matter. Unexpectedly however, such a general improvement in white matter was not observed while the present study demonstrated very specifically improved FA only in the right corticospinal tract. The corticospinal tract plays a crucial role in motor control and development.

[0017] DETAILED DESCRIPTION

[0018] The present invention thus concerns a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for use in reducing the risk of occurrence and / or treatment of an impaired motor development in an infant.

[0019] In some jurisdictions, the invention can be defined as the use of Bifidobacterium breve, short chain galactooligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for the manufacture of a nutritional composition or nutritional supplement for reducing the risk of occurrence and / or treatment of an impaired motor development in an infant.

[0020] In yet other jurisdictions, the invention may be defined as a method for reducing the risk of occurrence and / or treatment of an impaired motor development in an infant, wherein the method comprises administration of a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine.

[0021] The present invention furthermore concerns a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine, for use in preventing long term motor dysfunction in an infant.

[0022] In some jurisdictions, the invention can be defined as the use of Bifidobacterium breve, short chain galactooligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for the manufacture of a nutritional composition or nutritional supplement for preventing long term motor dysfunction in an infant.

[0023] In yet other jurisdictions, the invention may be defined as a method for preventing long term motor dysfunction in an infant, wherein the method comprises administration of a nutritional composition ornutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine.

[0024] The present invention furthermore concerns a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for use in enhancing motor development and / or long term motoric function in an infant.

[0025] In some jurisdictions, the invention can be defined as the use of Bifidobacterium breve, short chain galactooligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for the manufacture of a nutritional composition or nutritional supplement for enhancing motor development and / or long term motoric function in an infant.

[0026] In yet other jurisdictions, the invention may be defined as a method for enhancing motor development and / or long term motoric function in an infant, wherein the method comprises administration of a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galactooligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine.

[0027] Alternatively this aspect of the invention may also be seen as the use of Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for enhancing motor development and / or long term motoric function in an infant or as a non-therapeutic method for enhancing motor development and / or long term motoric function in an infant, comprising administration of a nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galactooligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine.

[0028] Definitions

[0029] ’Nutritional composition’ and ‘nutritional supplement’ means a substance or formulation or combination of ingredients that can serve as at least a portion of a subject's nutrition. The terms ”nutritional(s)", "nutritional formula(s)", "enteral nutritional(s)", are used as non-limiting examples of nutritional composition(s) or nutritional supplement throughout the present disclosure. Moreover, ‘nutritional composition(s)’ may refer to liquids, powders, concentrates, or ready-to-use forms of enteral formulas, oral formulas, formulas for infants and / or formulas for pediatric subjects.

[0030] Degree of polymerization (DP) refers to the number of monomeric saccharide units in a poly- or oligosaccharide that are linked by glycosidic bonds.For the purpose of this invention, when amounts or ranges are expressed per volume this refers to the nutritional composition in a ready-to-use form, unless indicated otherwise. When amounts are expressed as wt%, this refers to the wt% based on dry weight, unless indicated otherwise.

[0031] In this document and in its claims, the verb "to comprise" and its conjugations is used in its non-limiting sense to mean that items following the word are included, but items not specifically mentioned are not excluded. In addition, reference to an element by the indefinite article "a" or "an" does not exclude the possibility that more than one of the elements is present, unless the context clearly requires that there be one and only one of the elements. The indefinite article "a" or "an" thus usually means "at least one".

[0032] Probiotic

[0033] In the use or method according to the present invention, the nutritional composition or nutritional supplement comprises Bifidobacterium Breve. Bifidobacterium Breve (B. Breve) is a probiotic. The term probiotic is known in the art and refers to micro-organisms, preferably bacteria, which have a beneficial effect on a host when ingested by or administered to that host. In the method or use according to the present invention, the probiotic is preferably a lactic acid producing bacteria.

[0034] The nutritional composition or nutritional supplement according to the present invention preferably comprises Bifidobacterium Breve. Bifidobacterium breve is a Gram-positive, anaerobic, branched rodshaped bacterium. The B. breve preferably has at least 95 % identity of the 16 S rRNA sequence when compared to the type strain of B. breve ATCC 15700, more preferably at least 97% identity (Stackebrandt & Goebel, 1994, Int. J. Syst. Bacteriol. 44:846-849). Preferred B. breve strains are those isolated from the faeces of healthy human milk-fed infants. Typically, these are commercially available from producers of lactic acid bacteria, but they can also be directly isolated from faeces, identified, characterized and produced. According to a preferred embodiment, the present nutritional composition or supplement contains at least one B. breve selected from the group consisting of B. breve Bb-03 (Rhodia / Danisco), B. breve M-16V (Morinaga), B. breve R0070 (Institute Rosell, Lallemand), B. breve BR03 (Probiotical), B. breve BR92 (Cell Biotech), DSM 20091, LMG 11613, YIT4065, FERM BP-6223 and CNCM 1-2219. Most preferably, the B. breve is selected from the group consisting of B. breve M-16V and B. breve CNCM 1-2219, most preferably M-16V. B. breve 1-2219 was published in WO 2004 / 093899 and was deposited at the Collection Nationale de Cultures de Microorganisms, Institute Pasteur, Paris, France on 31 May 1999 by Compagnie Gervais Danone. B. breve M-16V was deposited as BCCM / LMG23729 and is commercially available from Morinaga Milk Industry Co., Ltd.

[0035] The nutritional composition or nutritional supplement that is administered according to the present use or according to the present method preferably comprises 103to 1013colony forming units (cfu) of B. breve,per gram dry weight of the nutritional composition or supplement, preferably 104to 1012, more preferably 105to 1010, most preferably from 105to 108cfu. The amount of B. breve, according to the present use or method is preferably administered at a daily dose of 102to 1013, more preferably from 105to 1012, most preferably from 107to 5x109colony forming units (cfu). Preferably the nutritional composition or supplement comprises 103to 1013cfu of B. breve, per 100 ml, more preferably 106to 1011cfu per 100 ml, most preferably 107to 109cfu per 100 ml.

[0036] The nutritional composition or nutritional supplement according to the present use or method preferably comprises viable B. breve. Alternatively, the nutritional composition or supplement according to the present use or method preferably comprises non-viable B. breve equivalent to the amounts of cfu as described above. The equivalent of cfu can be determined by performing the 5’nuclease assay with appropriate probes and primers, preferably the B. breve probes and primers as disclosed in WO 2005 / 039319 in the nutritional composition or supplement comprising non-viable B. breve and compare this with a calibration curve obtained from a comparable nutritional composition to which known amounts in cfu of viable B. breve have been added. Viable bifidobacteria can be commercially obtained as described above. B. breve cells can be made non-viable by methods known in the art, including heat treatment steps (including sterilization, pasteurization, UHT treatment), radiation (UV), treatment with oxygen, treatment with bactericidals such as ethanol, sonication, ultra-high pressure application, high pressure homogenization and use of a cell disruptor. Preferably B. breve is heat-killed. The presence of non-viable B. breve, advantageously provides many product technological benefits, including increased shelf-life, a reduced incidence of bacterial contamination, decreased post-acidification of the product, improved dosage control and improved convenience of reconstitution.

[0037] Non-digestible oligosaccharides

[0038] In the use or method according to the present invention, the nutritional composition or nutritional supplement comprises short chain galacto-oligosaccharides (scGOS) and long chain fructooligosaccharides (IcFOS). scGOS and IcFOS are prebiotic fibers also referred to as dietary fibers or non-digestible oligosaccharides. Non-digestible saccharides are nutritional components that are typically resistant to digestion and absorption in the small intestine, hence which are not or only partially digested by the action of acids or digestive enzymes such as present in for instance small intestine and stomach, with preferably a complete or partial fermentation in the large intestine, in particular by intestinal flora. For example galactose (as monomer), sucrose, lactose, maltose and maltodextrins are considered digestible.

[0039] Short chain galacto-oligosaccharides are non-digestible oligosaccharides preferably having the formula ([galactose]n-glucose; wherein n is an integer ranging from 2 to 10, i.e. 2, 3, 4, 5, 6, ....,10;), wherein the galacto-oligosaccharides are transgalacto-oligosaccharides. Transgalacto-oligosaccharides are for example available under the trade name VivinalOGOS (FrieslandCampina Ingredients), Bimuno®(Clasado), Cup-oligo® (Nissin Sugar) and Oligomate55® (Yakult). Preferably the nutritional composition or nutritional supplement comprises scGOS with an average degree of polymerization (DP) ranging from 3 to 7, more preferably ranging from 3 to 5.

[0040] Long chain fructo-oligosaccharides are a non-digestible oligosaccharides comprising a chain of beta-linked fructose units with an average degree of polymerization (DP) ranging from 6 to 1000, more preferably 10 to 100, even more preferably 20 to 40. IcFOS include inulin, levan and / or a mixed type of polyfructan. An especially preferred IcFOS is inulin. Inulin has a structure of chain-terminating glucosyl moieties and a repetitive fructosyl moiety, which are linked by beta-2,1 linkages. IcFOS suitable for use in the present nutritional composition or nutritional supplement is also commercially e.g OraftiOHP (Beneo GmbH). Preferably the IcFOS has an average DP above 20.

[0041] Preferably the combination of scGOS and IcFOS is present in a weight ratio of from 1 :99 to 99:1 , more preferably from 1:19 to 19:1 , more preferably from 1:1 to 19:1, more preferably from 2:1 to 15:1 , more preferably from 5:1 to 12:1 , even more preferably from 8:1 to 10:1 , even more preferably in a ratio of about 9:1.

[0042] In one embodiment, the nutritional composition preferably comprises 0.5 to 20 % of the sum of scGOS and IcFOS based on dry weight of the nutritional composition, more preferably 0.5 to 10 %, even more preferably 1.5 to 7.5 % based on dry weight. In a preferred embodiment, the nutritional composition comprises 80 mg to 2 g of the sum of scGOS and IcFOS per 100 ml, more preferably 150 mg to 1.50 g, even more preferably 300 mg to 1 g per 100 ml.

[0043] In case of a nutritional supplement this comprises at least 10 % of the combination scGOS and IcFOS based on dry weight of the nutritional supplement, preferably at least 15 %, more preferably at least 20 %, more preferably at least 25 %, or at least 30 % or at least 40 % ofthe combination scGOS and IcFOS based on dry weight ofthe nutritional supplement.

[0044] Glutamine

[0045] In the use or method according to the present invention, the nutritional composition or nutritional supplement comprises glutamine. Glutamine in the present invention refers to L-glutamine. Glutamine is one ofthe most abundant amino acids in plasma and human milk and is considered conditionally essential in preterm infants. Glutamine is utilized as a source of energy and for nucleotide synthesis in all rapidly dividing cells, such as the intestinal lining and certain immune cells. In the brain, glutamine is a substrate for neurotransmitters and an important source of energy for the nervous system.Glutamine is preferably present in an easily absorbable form. In particular in infants with an immature intestinal tract, glutamine present in intact protein is less easily absorbed. Therefore the nutritional composition or nutritional supplement preferably comprises glutamine in the form of free amino acid, glutamine containing dipeptide and / or glutamine containing tripeptide, most preferably glutamine containing dipeptide and / or free glutamine. Free glutamine and glutamine containing dipeptide and glutamine containing tripeptide are commercially available, for example at Ajinomoto, USA.

[0046] The nutritional composition in the method or use of the present invention preferably comprises glutamine levels higher then normally present in human milk protein or standard infant formula based on cow’s milk derived protein. Preferably the nutritional composition comprises at least 12 wt.%, more preferably at least 15 wt.%, even more preferably at least 30 wt.% glutamine based on total protein. Preferably the nutritional composition comprises less than 80 wt.%, more preferably less than 50 wt.% glutamine based on total protein.

[0047] Preferably the nutritional composition in the method or use of the present invention comprises at least 1.5 %, more preferably at least 2 %, even more preferably at least 4 % glutamine based on dry weight of the nutritional composition. Preferably the nutritional composition comprises less than 20 %, more preferably less than 10 % glutamine based on dry weight of the nutritional composition.

[0048] In an embodiment, preferably the nutritional composition in the method or use of the present invention comprises at least 0.3 g, more preferably at least 0.5 g, even more preferably at least 1 g glutamine based on 100 kcal of the nutritional composition. Preferably the nutritional composition comprises less than 5 g, even more preferably less than 2 g glutamine based on 100 kcal of the nutritional composition. In a preferred embodiment, the nutritional composition preferably comprises at least 0.4 g, more preferably at least 0.6 g, even more preferably at least 1.25 g glutamine per 100 ml. Preferably the nutritional composition comprises less than 6 g, even more preferably less than 2.5 g glutamine per 100 ml.

[0049] In a preferred embodiment, the weight ratio of glutamine to the sum of scGOS and IcFOS in the nutritional composition is between 2:1 and 1:4, more preferably between 1:1 and 1 :3, most preferably about 1:2.

[0050] In case of a nutritional supplement this preferably comprises glutamine in the form of free amino acid, glutamine containing dipeptide and / or glutamine containing tripeptide, most preferably glutamine containing dipeptide and / or free glutamine.

[0051] Preferably the nutritional supplement is in the form of a powder, preferably in a unit dose.In a preferred embodiment the nutritional supplement comprises at least 5 %, or at least 8 % glutamine in the form of free glutamine, glutamine dipeptide and / or glutamine tripeptide based on dry weight of the nutritional supplement. Preferably the nutritional supplement comprises at least 30 %, more preferably at least 50 %, most preferably more than 75 % glutamine based on weight of total protein.

[0052] In a preferred embodiment, the nutritional supplement is added to human milk, standard preterm formula, or human milk fortified with standard human milk fortifier in such an amount that the final nutritional composition comprises at least 12 wt.%, more preferably at least 15 wt.%, more preferably at least 20 wt.%, even more preferably at least 30 wt.% glutamine based on total protein.

[0053] Preferably the nutritional supplement is added to human milk, standard preterm formula, or human milk fortified with standard human milk fortifier in such an amount that the final glutamine enriched nutritional composition comprises at least 1.5 wt.%, more preferably at least 2 wt.%, even more preferably at least 4 wt.% glutamine based on dry weight of the nutritional composition.

[0054] In a preferred embodiment, the weight ratio of glutamine to the sum of scGOS and IcFOS in the nutritional supplement is between 1 :1 and 1 :8, more preferably between 1 :2 and 1 :6, most preferably about 1 :5.

[0055] Nutritional composition and nutritional supplement

[0056] The terms “nutritional composition” and “composition” are used interchangeably. The nutritional composition according to the invention preferably is an infant formula, a follow-on formula, a preterm formula, a post discharge formula or a pediatric formula. A pediatric formula can be a formula for faltering growth or a formula for tube feeding. Preferably, the nutritional composition is a preterm formula.

[0057] The terms as used herein, “infant formula”, “follow-on formula”, “a preterm formula”, “a post discharge formula” or “a pediatric formula” refer to compositions that are artificially made or that are synthetic This means that the composition that is administered is not human milk. It also means that the administered composition is not native cow’s milk or native milk from another mammal.

[0058] In the present context, infant formula refers to nutritional compositions, artificially made, intended for infants of 0 to about 4 to 6 months of age and are intended as a substitute for human milk. Typically, infant formulas are suitable to be used as sole source of nutrition. Such formulas are also known as starter formula. Formula for infants starting for 4 to 6 months of life to 12 months of life are intended to be supplementary feedings to infants that start weaning on other foods. Such formulas are also known as follow-on formulas. Infant formulas and follow-on formulas are subject to strict regulations, for example the EU regulations no.

[0059] 609 / 2013 and no. 2016 / 127.The nutritional composition in the method or use according to the present invention advantageously concerns a composition comprising lipid, protein and carbohydrates, wherein the lipid provides 5 to 50% of the total calories, the protein provides 5 to 50% of the total calories, and the carbohydrate provides 15 to 90% of the total calories. Preferably, in the nutritional composition the lipid provides 35 to 50% of the total calories, the protein provides 7.5 to 12.5% of the total calories, and the carbohydrate provides 40 to 55% of the total calories. For calculation of the % of total calories for the protein component, the total of energy provided by the proteins, peptides and amino acids needs to be taken into account.

[0060] The nutritional composition preferably comprises at least one lipid selected from the group consisting of animal lipid (excluding human lipids) and vegetable lipids. Preferably the present composition comprises a combination of vegetable lipids and at least one oil selected from the group consisting offish oil, animal oil, algae oil, fungal oil, and bacterial oil.

[0061] The nutritional composition preferably comprises protein. The protein in the nutritional composition is preferably selected from the group consisting of non-human animal proteins (preferably milk proteins, preferably proteins from cow’s milk), vegetable proteins (preferably soy protein and / or rice protein), free amino acids and mixtures thereof. The nutritional composition preferably contains casein, whey, hydrolyzed casein and / or hydrolyzed whey protein. Preferably the protein comprises intact proteins, more preferably intact bovine whey proteins and / or intact bovine casein proteins. As the present invention is preferably for use by preterm or HIE infants, the protein is preferably selected from the group consisting of hydrolyzed milk protein, more preferably selected from the group consisting of hydrolyzed whey protein and hydrolyzed casein.

[0062] The nutritional composition preferably comprises digestible carbohydrates. The nutritional composition preferably comprises a digestible carbohydrate, wherein at least 35 wt.%, more preferably at least 50 wt.%, more preferably at least 75 wt.%, even more preferably at least 90 wt.%, most preferably at least 95 wt.% is lactose. The nutritional composition preferably comprises at least 25 grams lactose per 100 gram dry weight of the nutritional composition, preferably at least 40 grams lactose per 100 gram.

[0063] When in liquid from, the nutritional composition preferably has a caloric density between 0.1 and 2.5 kcal / ml, even more preferably a caloric density of between 0.5 and 1.5 kcal / ml, most preferably between 0.6 and 0.8 kcal / ml. The amount of nutritional composition administered per day is preferably between 50 and 2000 ml, more preferably between 200 and 1500, most preferably between 400 and 1000 ml.

[0064] In one embodiment the nutritional composition in the method or use according to the present invention is a preterm infant formula, or briefly preterm formula. The preterm formula comprises all macro- andmicronutrients needed for preterm infants so as to achieve a growth similar to fetal growth coupled with satisfactory functional development.

[0065] In a preferred embodiment the preterm formula comprises from 5 to 25 wt.% protein, preferably 9 to 20 wt.%, more preferably 13 to 18 wt.% protein based on the dry weight of the preterm formula. In a preferred embodiment the preterm formula comprises from 1.8 to 3.0 g protein, preferably 2.0 to 3.0 g, more preferably 2.5 g to 2.6 g protein per 100 ml.

[0066] The preterm formula in ready to drink form has in a preferred embodiment about 70 to 90 kcal, preferably 75 to 85 kcal per 100 ml. The preterm formula preferably has an osmolarity below 450 mOsmol / l, more preferably below 400, even more preferably below 350. Particularly in preterm infants, a too high osmolarity is a disadvantage.

[0067] In one embodiment the present invention concerns a nutritional supplement, suitable to fortify human milk, to fortify human milk fortified with a standard human milk fortifier or to fortify a standard preterm formula. In the context of this invention, a nutritional supplement does not comprise all macro- and micronutrients needed for preterm infants so as to achieve a growth similar to fetal growth coupled with satisfactory functional development.

[0068] The nutritional supplement in the method or use according to the present invention is intended to supplement the diet of an infant and in particular is suitable to supplement human milk or to supplement an infant formula including to supplement a standard preterm formula. Such supplementation may also be referred to as fortifying, hence in the context of the present invention the nutritional supplement is intended and suitable to fortify human milk, or fortify an infant formula, for example to fortify a standard preterm formula.

[0069] Preferably the nutritional supplement in the method or use according to the present invention additionally comprises an additional component other than B. breve, scGOS, IcFOS and glutamine. Preferably such an additional component is a carrier. In a preferred embodiment less than 75 %, preferably less than 60 % based on dry weight of the nutritional supplement is comprised of additional components other than B. breve, scGOS, IcFOS and glutamine. Preferably less than 75 %, preferably less than 60 % based on dry weight of the nutritional supplement is a carrier. Suitable carrier components are for example maltodextrin, lactose, maltose, sucrose, glucose, glucose syrup, starch, in particular precooked or gelatinized starch, calcium phosphate. Preferably the nutritional supplement comprises maltodextrin as a carrier component. Preferably less than 75 %, more preferably less than 60 % based on dry weight of the nutritional supplement is comprised of maltodextrin.In one embodiment, the nutritional composition or nutritional supplement is in dry form, preferably in the form of a powder. This powder is suitable for reconstitution with water or another aqueous phase. When the nutritional composition or nutritional supplement is in powder form it advantageously has a better shelf life.

[0070] In an embodiment, the nutritional supplement is in the form of a kit of parts wherein one part comprises B. breve and another part comprises scGOS, IcFOS and glutamine. Preferably both elements in the kit of parts are in powder form. In an embodiment the kit of parts comprises a third part which is an infant formula in liquid form that is to be fortified with the ingredients provided by the other parts of the kit of parts comprising B. breve and scGOS, IcFOS, glutamine. The latter two parts of the kit of parts are to be added to the third part of the kit of parts which is the liquid infant formula, just before use.

[0071] LC-PUFA

[0072] Preferably the nutritional composition in the method or use according to the present invention comprises long chain poly unsaturated fatty acids (LC-PUFA), more preferably n-3 and n-6 LC-PUFA, even more preferable arachidonic acid (ARA) and docosahexaenoic acid (DHA). LC-PUFA is an important part of the fatty acyl chain composition of the brain membranes and therefore advantageously enhances brain microstructure maturation. The presence of LC-PUFA, in particular ARA and DHA, will have a further improved, or even synergistic, beneficial effect together with the prebiotic fiber, the probiotic and the glutamine. This is especially important for preterm infants as the maternal-fetal transfer of LC-PUFAs such as DHA and ARA in the third trimester is interrupted.

[0073] n-3 LC-PUFA, in particular docosahexaenoic acid (DHA), is an important part of the fatty acyl chain composition of the brain membranes and advantageously enhances brain microstructure maturation. More preferably, the nutritional composition comprises n-3 LC-PUFA, even more preferably DHA. Since a low concentration of DHA is already effective, the content of n-3 LC-PUFA in the nutritional composition, preferably does not exceed 15 wt.% of the total fatty acid content, preferably does not exceed 10 wt.%, even more preferably does not exceed 5 wt.%. Preferably the nutritional composition comprises at least 0.2 wt.%, preferably at least 0.5 wt.%, more preferably at least 0.75 wt.% n-3 LC-PUFA of the total fatty acid content. Preferably the nutritional composition comprises at least 0.2 wt.%, preferably at least 0.5 wt.%, more preferably at least 0.75 wt.% DHA of the total fatty acid content. The DHA content preferably does not exceed 5 wt.%, more preferably does not exceed 1 wt.%, but is preferably at least 0.1 wt.% of the total fatty acid content. Preferably as a source of n-3 LC-PUFA single cell oil, preferably algal oil, fungal oil and / or microbial oil is used, since these oil sources have a low EPA / DHA ratio, which results in a beneficial effect on the brain. More preferably the nutritional composition comprises fish oil, more preferably tuna oil.n-6 LC-PUFA, in particular arachidonic acid (ARA) is an important part of the fatty acyl chain composition of the brain membranes and advantageously enhances brain microstructure maturation. The nutritional composition preferably comprises relatively low amounts of ARA. The n-6 LC-PUFA content preferably does not exceed 5 wt.%, more preferably does not exceed 0.8 wt.%, more preferably does not exceed 0.75 wt.%, even more preferably does not exceed 0.5 wt.% based on total fatty acids. Since ARA is important in infants for optimal functional membranes, especially membranes of neurological tissues, the amount of n-6 LC-PUFA is preferably at least 0.02 wt.%, more preferably at least 0.05 wt.%, even more preferably at least 0.1 wt.% based on total fatty acid content, more preferably at least 0.25 wt.%. Preferably the nutritional composition comprises at least 0.02 wt.%, more preferably at least 0.05 wt.%, even more preferably at least 0.1 wt.% ARA based on total fatty acid content, more preferably at least 0.25 wt.% ARA based on total fatty acid content. The ARA content preferably does not exceed 5 wt.%, more preferably does not exceed 0.8 wt.%, more preferably does not exceed 0.75 wt.%, even more preferably does not exceed 0.5 wt.% based on total fatty acid content.

[0074] The weight ratio n-6 LC-PUFA I n-3 LC-PUFA, in particular the weight ratio of ARA / DHA in the nutritional composition is preferably from 3 to 0.5, more preferably from 2 to 0.8. Preferably the weight ratio is 1. These ratio’s ensure an optimal brain functioning.

[0075] LC-PUFA are preferably provided as free fatty acids, in triglyceride form, in diglyceride form, in monoglyceride form, in phospholipid form, or as a mixture of one of more of the above. Preferably the nutritional composition contains LC-PUFA in triglyceride and / or phospholipid form, even more preferably phospholipid form since LC-PUFA in phospholipid form are better incorporated into membranes. Therefore a dietary source of LC-PUFA in the form of phospholipids will have a further improved effect on the brain than when administered in form of triglycerides. A preferred source of LC-PUFA therefore is egg phospholipid. Commercial sources of egg oil, rich in phospholipids, and having arachidonic and docosahexaenoic fatty acyl chains in the phospholipid molecules are known.

[0076] Application

[0077] Motor development and motor function in infants involves the development of both gross motor skills, such as crawling and walking and fine motor skills such as grasping and manipulating objects. Preterm birth is associated with motor developmental delays and increased likelihood of cerebral palsy, developmental coordination disorder and perinatal strokes. All of which may impair motor function.

[0078] The inventors have found that early nutritional supplementation with B. breve, scGOS, IcFOS and glutamine supports microstructural development of the right corticospinal tract (rCST) at term equivalent age in preterm infants. The corticospinal tract is a major fiber pathway in the central nervous system projectingfrom the brain and terminates in the spinal court. The corticospinal tract is the principal motor system for controlling movements that require the greatest skill and flexibility and is important for motor development.

[0079] Development of the corticospinal tract occurs during a protracted postnatal period, increasing the period of vulnerability for damage. This damage can have a long-term consequence for motor development and / or impaired motor functions later in life.

[0080] The nutritional composition or nutritional supplement according to the invention comprising Bifidobacterium breve, short chain galacto-oligosaccharides, long chain fructo-oligosaccharides and glutamine for use in reducing the risk of occurrence and / or treatment of an impaired motor development in an infant.

[0081] In another preferred embodiment, the nutritional composition or nutritional supplement according to the invention comprising Bifidobacterium breve, short chain galacto-oligosaccharides, long chain fructooligosaccharides and glutamine for use in preventing long term motor dysfunction in an infant.

[0082] Impaired development of the corticospinal tract in infants is indicative of impaired motor development and / or of impaired motor functions later in life. In a preferred embodiment, motor development and / or motor dysfunction is selected from neuronal motor disabilities, motor disability, cerebral palsy, tight or resistant muscle control, weak muscle control, spasms, coordination disorder, tremors, dystonia, spasticity, hyperreflexia and / or ataxia.

[0083] In another preferred embodiment, the nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for use in enhancing motor development and / or long term motoric function in an infant.

[0084] The effects of preventing long term motor dysfunction as described above are preferably established later in life. Preferably, when the infant is older than 1 year of age, more preferably when the infant is older than 2 years of age, most preferably when the infant is older than 3 years of age.

[0085] With reducing the risk of occurrence and / or treatment of an impaired motor development and / or preventing long term motor dysfunction is meant to obtain motor development and / or motor function more similar to a term infant control group. Alternatively, with reducing the risk of occurrence and / or treatment of an impaired motor development and / or preventing long term motor dysfunction is meant an improvement in motor development or motor function compared to the motor development or motor function of a preterm control group not having received the nutritional composition or nutritional supplement according to the invention.Motor development can be measured by the Motor Scales in the Bayley Scales of Infant and Toddler Development. The Bayley Scales of Infant and Toddler Development is a well-known and standard series of measurements used to assess the development of infants and toddlers between the age of 1 -42 months. In the Bayley Scales gross and fine motor skills such as grasping, sitting, stacking blocks and climbing stairs are assessed with the Motor Scale subtest. The most recent version of the Bayley scales is the Bayley IV, however also previous version III is suitable that also included the Motor Scale subtest. In particular motor development and / or motor function is measured by the Motor Scales in the Bayley Scales of Infant and Toddler Development. The test is preferably assessed by an experienced paediatric psychologist or educational therapist. Preferably toddlers are tested around 24 months corrected age.

[0086] Damage or risk for damage of motor function and development is higher in preterm infants. In a preferred embodiment, the infant is a preterm infant, a small for gestational age (SGA) infant, a low birth weight infant, an infant with hypoxic-ischemic encephalopathy (HIE) and / or an infant with brain damage caused by trauma. Preferably, the infant is a preterm infant.

[0087] A preterm infant can also be named a premature infant. A subgroup of premature infants are very preterm infants. A preterm infants is born before the end of the 37th week of pregnancy. A very preterm infant is born before the end of the 32th week of pregnancy. In a preferred embodiment, the method or use according to the present invention is in a very preterm infant.

[0088] A small for gestational age (SGA) infant is an infant whose birth weight lies below the 10th percentile for that gestational age. A SGA infant has usually been the subject of intrauterine growth restriction (IUGR). In one embodiment, the nutritional composition in the present method or use is preferably intended for an infant that is born small for gestational age (SGA).

[0089] Premature and / or SGA infants include low birth weight infants (LBW infants), very low birth weight infants (VLBW infants), and extremely low birth weight infants (ELBW infants). LBW infants are defined as infants with a weight less than 2500 g. VLBW infants as infants with a weight which is less than 1500 g, and ELBW infants as infants with a weight less than 1000 g. In a preferred embodiment, the method or use according to the present invention is in a VLBW. Also in a preferred embodiment, the method or use according to the present invention is in an ELBW.

[0090] Hypoxic-ischemic encephalopathy is a type of brain injury that occurs when the brain does not receive enough oxygen and blood flow. HIE can occur before birth, during labour and delivery or after birth. Brain injury from HIE can cause developmental delays including impaired motor development and / or motor function.Administration of the present nutritional composition to the infant is preferably enterally or parenterally, more preferably enterally. Preferably the nutritional composition is fed orally via a bottle, even more preferably enterally by naso-gastric tube feeding. Enteral feeding has the advantage that the B. breve, scGOS, IcFOS and glutamine will provide a beneficial effect in the intestinal tract as well regarding intestinal infections and improving barrier function.

[0091] Preterm infants have an immature intestinal tract, hence it is preferred that the nutritional composition or supplement according to the invention is administered to the preterm infant starting at least in the first two weeks after birth, preferably within the first week after birth, more preferably at least within 5 days after birth, even more preferably at least within 3 days after birth, most preferably at least within 2 days after birth.

[0092] In a preferred embodiment, the nutritional composition or the nutritional supplement is administered to the infant for a duration of at least 4 weeks, preferably at least 6 weeks, in the period of 0 to 4 months after birth, more preferably in the period of 0 to 3 months after birth.

[0093] Daily dose

[0094] The nutritional composition or nutritional supplement is preferably administered to the infant so that the infant receives an effective daily dose of B. Breve, scGOS, IcFOS and glutamine.

[0095] In a preferred embodiment according to the use or method of the present invention the daily dose of B. breve is 102to 1013cfu / kg, more preferably from 105to 1012cfu / kg, most preferably from 107to 1010cfu / kg.

[0096] In a preferred embodiment according to the use or method of the present invention the daily dose of scGOS and IcFOS are administered to the infant in a daily dose of 0.02 to 1.0 g / kg body weight, preferably of 0.1 to 0.8 g / kg body weight, wherein the weight ratio of scGOS to Ic FOS ranges from 2:1 to 15:1, preferably from 5:1 to 12:1, even more preferably from 8:1 to 10:1, even more preferably in a ratio of about 9:1.

[0097] In a preferred embodiment according to the use or method of the present invention the daily dose of glutamine provided to the infant is 0.01 to 0.5 g / kg body weight, preferably 0.05 to 0.4 g / kg body weight.

[0098] In a preferred embodiment the nutritional composition or nutritional supplement for use according to the invention is administered to the infant in a daily dose providing

[0099] a) B. breve in a dose of 105to 1012cfu / kg body weight / day, preferably 107to 101° / kg body weight / day, b) scGOS and IcFOS in a dose of 0.02 to 1.0 g / kg body weight / day, preferably of 0.1 and 0.8 g / kg body weight / day, wherein the weight ratio of scGOS to Ic FOS ranges from 5:1 to 12:1, preferably from 8:1 to 10:1 , even more preferably in a ratio of about 9:1, andc) glutamine in a dose of 0.01 to 0.5 g / kg body weight / day, preferably of 0.05 to 0.4 g / kg body weight / day.

[0100] Examples

[0101] Example 1: Effect of B. breve, glutamine, GOS and IcFOS on brain development

[0102] A double-blind, randomized, controlled, parallel group, single centre clinical trial was performed. The study design and setting has been described in Hortensius et al., 2021, Nutribrain: protocol for a randomized, double-blind, controlled trial to evaluate the effects of a nutritional product on brain integrity in preterm infants, BMC Pediatrics 21 :132.

[0103] Preterm infants born at 24 to 30 weeks gestational age (GA) less than 72 hours old were enrolled. In total 99 infants born before 30 weeks GA were enrolled in the study. 50 infants were randomized in the test group and 49 infants in the control group. The modified intention to treat (mITT) population (intervention started and MRI scan present) consisted of 81 infants, of which 40 infants in the test group and 41 infants in the control group. Of this group 74 had a MRI scan of sufficient quality to determine FA values.78 infants completed the study protocol without major deviations leading to a Per Protocol (PP) group of 78, wherein 39 infants in the test group and 39 infants in the control group. The Per Protocol group (PP) group with a high quality MRI scan was 71 : 36 in the test group and 35 in the control group. A high quality MRI scan was needed to determine the FA values.

[0104] The infants received either the nutritional intervention or a placebo. The test group received the nutritional composition in two sachets, Part A containing B. breve M-16V and Part B containing scGOS (VivinalOGOS, FrieslandCampina Ingredients) and IcFOS (OraftiOHP) in a wt / wt ratio of 9:1 and L-glutamine. The control group received two separate sachets, Part A containing maltodextrin and Part B containing maltodextrin, casein and whey hydrolysates. Enteral nutritional supplementation started within 72 hours after birth. The supplement was complementary to regular hospital feeding regimes of preterm infants. Supplementation continued until the MRI was taken.

[0105] B. breve was administered once a day as single daily dose. The dosage depended on birth weight. Infants with a birth weight > 1000 g received 3 x 109colony-forming units (cfu) per day. Infants with a birth weight < 1000 g received 1.5 x 1 o9cfu per day, which was increased to 3 x 1 o9cfu per day when they had reached enteral feeding intakes of 50 ml / kg / day. scGOS, IcFOS and glutamine were administered enterally supplemented to the regular milk feed (breast milk or preterm formula, depending on parents’ choice). Dosage was increased in a stepwise fashion. Based on the infant’s weight, daily intake of enteral feeding and the desired (maximum) osmolarity of the feeding, a supplementation schema was created, starting with a minimum dosage of 0.05 g / kg / day L-glutamine and 0.1 g / kg / day scGOS / lcFOS and aiming for a final doseof 0.3 g / kg / day L-glutamine and 0.6 g / kg / day scGOS / lcFOS when the infant receives full enteral feeding (120 ml / kg / day for 1 day).

[0106] Infants underwent magnetic resonance imaging (MRI) at TEA. Segmented fractography analysis of Fractional Anisotropy (FA) in the corticospinal tract (CST) from Posterior Limb of the Internal Capsule (PLIC) to the spine was conducted. Sensitivity analyses on percentage product intake were performed. Tract-Based Spatial Statistics (TBBS) was applied and FA values were compared between infants in the intervention group and infants in the control group, controlling for sex, GA at birth, age at time of scan, and birth weight Z-score. Higher FA indicates increased white matter integrity.

[0107] Results

[0108] There were no differences between groups in clinical characteristics and the occurrence of neonatal infections. Diffusion Tensor Imaging (DTI; an advanced MRI technique) was used to map white matter fractography in the infant brains. Tract-Based Spatial Statistics (TBSS) on fractional anisotropy (FA) maps throughout the centers of the white matter suggested that FA values were higher in the right corticospinal tract (CST) in the intervention group compared to the control group. Segmented fractography analysis on this tract revealed significantly higher FA values in the test compared to the control group in the mITT and PP populations (with and without controlling for the covariates: sex of child, gestational age at birth, weight-for-age z-score at birth and postmenstrual age at scan), see table 1.

[0109] Table 1 : FA values in right corticospinal tract (rCST)

[0110] &

[0111]

[0112] * p value <0.05 is significant.

[0113] #not controlled for covariates;&controlled for covariates: sex of child, gestational age at birth, weight-forage z-score at birth and postmenstrual age at scan

[0114] Bayley Scales of Infant and Toddler Development - Motor Scale

[0115] Motor function was assessed by the Bayley’s Motor Scale tests to confirm effects on motor development. Motor development was tested at 24 months corrected age.At 24 months corrected age, gross motor function was assessed using the Bayley Scales III (Dutch version). Motor impairment was defined as a motor composite score below 85 (below 1 SD). The incidence of motor impairment (n (%)) was calculated for each group

[0116] After controlling for sex, gestational age, birthweight z-score, maternal education level and severe illness the incidence of motor impairment was significantly lower in the intervention group compared to the placebo in both the ITT and PP population, see table 2.

[0117] Table 2: Incidence of motor impairment at 24 months corrected age

[0118] &

[0119]

[0120] *p value < 0.05 is significant

[0121] &Controlled for covariates: sex of child, gestational age at birth, weight-for-age z-score at birth, maternal education level and severe illness (Severe illness was defined as two or more unplanned hospital admissions due to illness between term equivalent age and follow-up at 24 months corrected age).

[0122] Conclusion

[0123] Early nutritional supplementation with B. breve, scGOS, IcFOS and L-glutamine supports microstructural development of the rCST at TEA in preterm infants.

[0124] The corticospinal tract plays a crucial role in motor control and development. Therefore, an improved microstructural development of the corticospinal tract leads to an improvement in motor development and motor function.

[0125] Example 2: Preterm formula with B. breve, scGOS, IcFOS and glutamine

[0126] Preterm formula in powder form comprising per 100 g about 475 kcal, 15.8 g protein including L-glutamine in a total amount of about 2.4 g both in free form and incorporated in proteins, 49.8 g digestible carbohydrates (mainly lactose), 23 g lipid, 3.4. g non-digestible oligosaccharides and 2.9 109cfu Bifidobacterium breve M-16V (Morinaga). The non-digestible oligosaccharides are short chain galactooligosaccharides (VivinalOGOS, FrieslandCampina Ingredients) and long chain fructo-oligosaccharides(OraftiOHP) in a weight ratio of 9:1. For a ready to drink formula 16.9 g powder is to be reconstituted with water until a final volume of 100 ml.

[0127] Example 3: Faltering growth composition

[0128] Faltering growth composition in powder form. For a ready to drink form about 19.1 g powder is reconstituted with water until a final volume of 100 ml comprising 101 kcal, 2.6 g protein including 0.4 g L-glutamine in the form of L-alanyl-L-glutamine, 10.3 g digestible carbohydrates, 5.4 g lipid, 0.8 g scGOS / lcFOS in a 9:1 ratio, 6.107cfu B. breve R-70 (Lallemand).

[0129] Example 4: Nutritional supplement

[0130] Human milk fortifier in powder form packed in sachets comprising about 1 gram. The sachet is to be reconstituted per 25 ml of expressed human milk. The supplement comprises 109cfu B. breve M16-V (Lallemand), 0.5 g non-digestible oligosaccharides, 0.1 g L-glutamine and about 0.4 g maltodextrin as carrier material per gram / sachet. The non-digestible oligosaccharides are galacto-oligosaccharides (VivinalOGOS) and fructo-oligosaccharides (OraftiOHP) in a weight ratio of 9:1.

Claims

CLAIMS1. A nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine, for use in reducing the risk of occurrence and / or treatment of an impaired motor development in an infant.

2. A nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine, for use in preventing long term motor dysfunction in an infant.

3. The nutritional composition or supplement according to claim 1 or 2, wherein motor development and / or motor dysfunction is selected from the group consisting of neuronal motor disabilities, motor disability, cerebral palsy, tight or resistant muscle control, weak muscle control, spasms, coordination disorder, tremors, dystonia, spasticity, hyperreflexia and / or ataxia.

4. A nutritional composition or nutritional supplement comprising Bifidobacterium breve, short chain galacto-oligosaccharides (scGOS), long chain fructo-oligosaccharides (IcFOS) and glutamine for use in enhancing motor development and / or long term motoric function in an infant.

5. The nutritional composition or nutritional supplement for use according to any one of the preceding claims, wherein the infant is selected from the group consisting of a preterm infant, a small for gestational age infant, a low birth weight infant, an infant with hypoxic-ischemic encephalopathy (HIE) and / or an infant with brain damage caused by trauma.

6. The nutritional composition or nutritional supplement for use according to any one of the preceding claims, wherein the infant is a very preterm infant or a very low birth weight infant.

7. The nutritional composition or nutritional supplement for use according to any one of the preceding claims wherein the motor development and / or motor function is measured by the Motor Scales in the Bayley Scales of Infant and Toddler Development.

8. The nutritional composition or the nutritional supplement for use according to any of the preceding claims wherein the composition or supplement is administered to the infant for a duration of at least 4 weeks, preferably at least 6 weeks, in the period of 0 to 4 months after birth, more preferably in the period of 0 to 3 months after birth.

9. The nutritional composition for use according to any one of the preceding claims wherein- B. breve is present in an amount of 103to 1013cfu per g dry weight of the composition- scGOS and IcFOS are present in a total amount of 0.5 to 20 wt% in a weight ratio scGOSJcFOS of 1 :19 to 19:1 based on dry weight of the composition, and- glutamine is present in an amount of at least 1.5 % based on dry weight of the nutritional composition.

10. The nutritional composition for use according to any one of the preceding claims wherein the weight ratio of glutamine to the sum of scGOS and IcFOS is between 2:1 and 1 :4, more preferably between 1 :1 and 1:3.

11. The nutritional composition for use according to any of the preceding claims wherein the composition is an infant formula, a follow-on formula, a preterm formula, a post discharge formula, or a pediatric formula.

12. The nutritional supplement for use according to any one of claims 1-8, wherein- B. breve is present in an amount of 103to 1013cfu per g dry weight of the supplement,- scGOS and IcFOS are present in a total amount of at least 10 % in a weight ratio scGOSJcFOS of 1 :19 to 19:1 based on dry weight of the supplement, and- glutamine is in the form of free amino acid, glutamine containing dipeptide and / or glutamine containing tripeptide and is present in an amount of at least 5 % based on dry weight of the nutritional supplement.13 The nutritional supplement for use according to any claim 12, wherein the weight ratio of glutamine to the sum of scGOS and IcFOS is between 1:1 and 1 :8, more preferably between 1 :2 and 1 :6.14 The nutritional supplement for use according to any one of claims 12-13, wherein the supplement is intended to fortify human milk or to fortify an infant formula.15 The nutritional composition or nutritional supplement according to anyone of the preceding claims which is a powder suitable for reconstitution with water to a ready to drink form.

Images