WO2019068924A1

WO2019068924A1 - Promoting cognitive function after birth by caesarean section

Info

Publication number: WO2019068924A1
Application number: PCT/EP2018/077275
Authority: WO
Inventors: Ingrid Brunhilde RENES; Jan Knol; Shugui WANG; Kees VAN LIEMPT
Original assignee: N.V. Nutricia
Priority date: 2017-10-06
Filing date: 2018-10-08
Publication date: 2019-04-11
Also published as: WO2019068356A1

Abstract

The present invention concerns a method for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, comprising administering to said infant delivered via Caesarean section a composition comprising a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of agalacto-oligosaccharide and a fructan.

Description

Promoting cognitive function after birth by Caesarean section Field of the invention

[0001] The present invention relates to a composition for promoting cognitive function after birth by Caesarean section.

Background art

[0002] The worldwide rate of infant deliveries via Caesarean section has increased over the last decade, making it the most common surgical procedure performed in women of childbearing age today. While the WHO recommends that Caesarean section deliveries to be below 15 % of all deliveries, in many countries the rate of Caesarean section deliveries significantly exceeds this recommendation. Over the past years, the medical field has started to realise that a Caesarean section delivery introduces health risks, and the obstetrician is thus advised to assess these long- and short-term health consequences for mother and infant, as well as weigh the risks associated with the procedure itself against not performing the procedure. Nowadays, Caesarean section delivery has been associated with delayed intestinal colonization by beneficial bacteria, impaired immune system, and increased risk of asthma and obesity in the infant.

Summary of the invention

[0003] The present invention pertains to a method for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, comprising administering to said infant delivered via Caesarean section a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. While some improvement on cognition in infants may be expected, the inventors surprisingly found that the improvements in terms of cognitive functioning in infants delivered via Caesarean section go beyond reasonable expectations based on observations in infants in general. It is unknown in the art that B. breve and/or the mixture of a galacto-oligosaccharide and a fructan would be capable of normalizing the cognitive functioning of infants delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants.

[0004] The present invention can also be worded as the use of a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan in the manufacture of a composition for promoting cognitive functioning in an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants.

[0005] The present invention can also be worded as a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto- oligosaccharide and a fructan for use in promoting cognitive functioning in an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants. [0006] In one embodiment, the promoting cognitive functioning involves normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery. In one embodiment, the promoting cognitive functioning or normalizing the negative effects thereon involve at least one of: (a) improving social cognition; (b) improving behaviour; (c) reducing anxiety; (d) improving memory; (e) improving learning skills; and (f) reducing susceptibility to stress. In one embodiment, the promoting cognitive functioning or normalizing the negative effects thereon involve at least one of: (g) reducing stress levels and/or stress response; (h) increasing attachment to the mother; (i) improving social interaction; (j) improving novel object recognition; (k) reducing depressive-like symptoms; (I) reducing autism-like repetitive behaviour; and (m) reducing ADHD- like behaviour. In a particularly preferred embodiment, the promoting cognitive functioning or normalizing the negative effects thereon involves reducing depressive-like symptoms.

List of preferred embodiments

1. A method for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, comprising administering to said infant delivered via Caesarean section a composition comprising a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan.

2. The method according to embodiment 1 , wherein promoting cognitive functioning involves normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery.

3. The method according to embodiment 1 or 2, wherein cognitive functioning includes at least one of behaviour, memory, learning skills and sociability.

4. The method according to any one of the preceding embodiments, wherein promoting cognitive functioning or normalizing the negative effects thereon involves at least one of:

(a) improving social cognition;

(b) improving behaviour;

(c) reducing anxiety;

(d) improving memory;

(e) improving learning skills; and

(f) reducing susceptibility to stress.

5. The method according to any one of the preceding embodiments, wherein promoting cognitive functioning or normalizing the negative effects thereon involves at least one of:

(g) reducing stress levels and/or stress response;

increasing attachment to the mother;

improving social interaction;

(j) improving novel object recognition;

(k) reducing depressive-like symptoms;

reducing autism-like repetitive behaviour; and

reducing ADHD-like behaviour. 6. The method according to any one of the preceding embodiments, wherein promoting cognitive functioning or normalizing the negative effects thereon involves reducing depressive-like symptoms.

7. The method according to any one of the preceding embodiments, wherein the composition comprises a therapeutically effective amount of Bifidobacterium breve and a therapeutically effective amount of the mixture of galacto-oligosaccharide and fructan.

8. The method according to any one of the preceding embodiments, wherein the Bifidobacterium breve is Bifidobacterium breve M-16V.

9. The method according to any one of the preceding embodiments, wherein the fructan is selected from fructo-oligosaccharides, fructo-polysaccharides, inulin and mixtures thereof.

10. The method according to embodiment 9, wherein the fructan comprises fructo- oligosaccharides and/or fructo-polysaccharides, preferably fructopolysaccharides.

1 1. The method according to embodiment 9, wherein galacto-oligosaccharide and fructan are present in a weight ratio in the range of 5:1 - 20: 1 , preferably 7: 1 - 15: 1.

12. The method according to any one of the preceding embodiments, wherein administration occurs through human milk or by administering to the infant directly.

13. The method according to embodiment 12, wherein administration occurs directly to the infant and the composition is provided in the form of:

(i) a liquid having a volume between 0.5 to 5 ml for oral administration, wherein said composition is preferably administered to said infant with a syringe, pipette or tube;

(ii) a (reconstituted) infant formula; or

(iii) a suppository, pill or tablet, and if the composition is provided in the form of a suppository, said composition is rectally administered to said infant.

14. The method according to embodiment 12, wherein administration occurs through human milk and the composition is administered to the lactating mother in the form of a nutritional supplement.

15. The method according to any one of the preceding embodiments, wherein the composition is administered to the infant starting at least in the first sixteen weeks after birth, preferably at least within twelve weeks after birth, even more preferably at least within eight weeks after birth, most preferably at least within four weeks after birth.

16. Use of a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan in the manufacture of a composition for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants.

17. A composition comprising a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan for use in promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants. Detailed description

[0007] The inventors have surprisingly found that the administration of a composition comprising β. breve and/or a mixture of a galacto-oligosaccharide and a fructan has a beneficial effect on cognitive function in infants delivered via Caesarean section. The composition according to the invention was found capable of improving or promoting cognitive function of infants delivered via Caesarean section even towards the level found in vaginally-born infants, appreciated as the 'healthy standard'.

[0008] The invention may be worded as a method, a use or a composition for use, wherein the cognitive functioning of infants delivered via Caesarean section is promoted. In the context of the invention, throughout the specification whenever reference is made to the method of promoting cognitive functioning according to the invention, this equally reads on to the use for promoting cognitive functioning and the composition for use in promoting cognitive functioning according to the invention.

Composition

[0009] The method according to the invention involves the administration of a composition comprising B. breve and/or a mixture of a galacto-oligosaccharide and a fructan. This composition is interchangeably referred to as the composition according to the invention or the present composition and is further defined here below. The mixture of a galacto-oligosaccharide and a fructan is also referred to as the non-digestible oligosaccharides.

[0010] The composition according to the invention is typically suitable for enteral administration to the infant. The composition may be in any form known in the art to be suitable for such administration, such as in solid form, in semi-solid form or in liquid form. Preferably, the composition is a nutritional composition or a nutritional supplement. The composition may be referred to as a nutritional composition, preferably a nutritional composition for providing nutrition to infants, in particular infants delivered via Caesarean section. Preferably, the composition is in the form of a liquid or a powder which can be reconstituted with a liquid (typically water) to obtain a liquid composition. Alternatively, the composition is in the form of a capsule or tablet. In one embodiment, the composition is a complete nutrition. In a particularly preferred embodiment, the composition is an infant formula, preferably in powder form suited to be reconstituted with water.

[0011] The present composition comprises B. breve and/or the non-digestible oligosaccharides. In one embodiment, the present composition comprises at least B. breve. In one embodiment, the present composition comprises the non-digestible oligosaccharides. In one embodiment, the present composition comprises B. breve and the non-digestible oligosaccharides. As evidenced in the examples, cognition benefits are obtained with B. breve in the absence of non-digestible oligosaccharides, and with the non-digestible oligosaccharides in the absence of B. breve, but optimal results on promoting cognition are obtained using a combination of B. breve and the non- digestible oligosaccharides. According to a most preferred embodiment, both B. breve and the non- digestible oligosaccharides are comprised in the composition according to the invention. [0012] The inventors have found that B. breve and the non-digestible oligosaccharides both independently and in combination have a beneficial effect on (a) improving social cognition; (b) improving behaviour; (c) reducing anxiety; (d) improving memory; (e) improving learning skills; and (f) reducing susceptibility to stress. More in particular, beneficial effects have been observed in terms of (g) reducing stress levels and/or stress response; (h) increasing attachment to the mother; (i) improving social interaction; (j) improving novel object recognition; (k) reducing depressive-like symptoms; (I) reducing autism-like repetitive behaviour; and (m) reducing ADHD-like behaviour.

Bifidobacterium breve

[0013] In one embodiment, the composition according to the invention comprises Bifidobacterium breve. B. breve is a probiotic, i.e. a bacterial strain bringing a health benefit to the targeted infants. An especially suitably strain of B. breve to be used in the present invention is B. breve M-16V. The present composition may comprise more than one, such as at least two, preferably at least three and more preferably at least four distinct Bifidobacterium species. In one embodiment, the present composition comprises one distinct Bifidobacterium species.

[0014] The composition may comprise one or more further probiotic strains, preferably a Bifidobacterium or a lactic acid bacteria selected from the group consisting of Carnobacterium, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Oenococcus, Pediococcus, Streptococcus, Tetragenococcus, Vagococcus and Weissella. Preferred further probiotics are selected from the genera Lactobacillus, Streptococcus and Bifidobacterium. In case a further Bifidobacterium species is present, it is preferably selected from the group consisting of B. infants, B. bifidum, B. catenulatum, B. adolescentis, B. thermophilum, B. gallicum, B. animalis, B. angulatum, B. pseudocatenulatum, B. thermacidophilum and B. longum, more preferably from the group consisting of B. infantis, B. bifidum, B. catenulatum, B. adolescentis and B. longum, most preferably from the group consisting of B. longum subsp. longum, B. longum subsp. infantis and B. bifidum. In one embodiment, the further probiotic strain is selected from the group consisting of Bifidobacterium longum, Bifidobacterium infantis, Bifidobacterium bifidum, Lactobacillus acidophilus, Lactobacillus paracasei, Lactobacillus johnsonii, Lactobacillus plantarum, Lactobacillus reuteri, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus lactis and Streptococcus thermophiles. In one embodiment, the present composition comprises L. lactis and β. breve.

[0015] If present, B. breve is present in a therapeutically effective amount. The present composition preferably contains between 10³ and 10¹³ colony forming units (cfu) B. breve per gram dry weight of the present composition, preferably between 10⁴ and 10¹², more preferably between 10⁵ and 10¹⁰. Preferably, the present composition contains between 10³ and 10¹³ colony forming units (cfu) Bifidobacteria per g dry weight of the present composition, more preferably between 10⁴ and 10¹², most preferably between 10⁵ and 10¹². In terms of doses, the present composition preferably provides between 10³ and 10¹⁶ cfu, more preferably between 10⁴ and 10¹⁵ cfu, most preferably between 10⁵ and 10¹² cfu B. breve per serving. Preferably, the present composition provides between 10³ and 10¹⁶ cfu, more preferably between 10⁴ and 10¹⁵ cfu, most preferably between 10⁵ and 10¹² cfu Bifidobacteria per serving.

[0016] Preferably, the composition comprises B. breve and the optional further probiotics in freeze- dried form, which is especially suitable when the composition is in powder, capsule or tablet form.

Non-digestible oligosaccharides

[0017] In one embodiment, the composition according to the invention comprises a mixture of a galacto-oligosaccharide and a fructan. Galacto-oligosaccharides and fructans are non-digestible oligosaccharides, also referred to as a "prebiotic" or "prebiotic fibre". In the context of the present invention, the term "non-digestible oligosaccharide" refers to oligosaccharides which are not digested in the intestine by the action of digestive enzymes present in the upper digestive tract (small intestine and stomach) of the C-section infant but instead are fermented by the intestinal microbiota of said infant, thus conferring benefits upon the host wellbeing and health. Preferably the present non-digestible oligosaccharides has a degree of polymerisation (DP) of 2 to 250, preferably an average DP 2 to 100, more preferably 2 to 60. Preferably, at least 50 wt.% of the present non-digestible oligosaccharides have an average degree of polymerisation in the range of 2 to 60. The terms "prebiotic" and "prebiotic fibre" refer to non-digestible fibres that beneficially affects the host by selectively stimulating the growth and/or activity of one or a limited number of bacterial species in the colon. Health effects of prebiotics, also in combination with probiotics, are described in Collins, Am. J. Clin. Nutr. 1999, 69(suppl.), 10525-10575. Preferably, the present non- digestible oligosaccharide is soluble. The term "soluble" as used herein, when having reference to a fibre or oligosaccharide, means that the substance is at least 50% soluble according to the method described by Prosky et al. in J. Assoc. Off. Anal. Chem. 1988, 71 , 1017-1023.

[0018] First of all, the mixture comprises galacto-oligosaccharides, in particular β-galacto- oligosaccharides. Herein, galactose units make up for at least 50% of the monosaccharide units. The galacto-oligosaccharides are preferably [galactose]_n-glucose; wherein n is an integer between 1 and 60, i.e. 2, 3, 4, 5, 6,...., 59, 60; preferably n is 2, 3, 4, 5, 6, 7, 8, 9 and/or 10, a good example being irans-galacto-oligosaccharides. The galactose units are preferably beta linked. The galacto- oligosaccharides preferably comprise saccharides with an average degree of polymerisation (DP) of 2 to 10 (scGOS). (Trans)galactooligosaccharide is for example available under the trade name Vivinal®GOS (Borculo Domo Ingredients, Zwolle, Netherlands), Bimuno (Clasado), Cup-oligo (Nissin Sugar) and Oligomate55 (Yakult).

[0019] Secondly, the mixture comprises fructans. Preferred fructans include fructo- oligosaccharides, fructo-polysaccharides, inulin and mixtures thereof, most preferably fructo- oligosaccharides are comprised in the mixture. Preferred fructo-oligosaccharides are short-chain fructo-oligosaccharides (scFOS), having an average DP in the range of 2 to 10, and long-chain fructo-oligosaccharide (IcFOS), having an average DP in the range of 10 to 60, preferably in the range of 15 - 40. scFOS is commercially available as Beneo® P95 or Raftilose P95 (Orafti). A particular type of IcFOS is inulin, such as Raftilin HP. In an especially preferred embodiment, the present composition comprises galacto-oligosaccharides and fructo-oligosaccharides and/or fructo- polysaccharides, most preferably scGOS and IcFOS (preferably having an average DP > 15).

[0020] The composition according to the invention may comprise further non-digestible oligosaccharide, which are preferably selected from the group consisting of non-digestible dextrin, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto-oligosaccharides, gluco- oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan- oligosaccharides, chito-oligosaccharides, uronic acid oligosaccharides, sialyl-oligosaccharides and fuco-oligosaccharides. In one embodiment, the composition comprises galacto-oligosaccharides, fructans and fucosyllactose. The present composition comprises at least two, more preferably at least three and most preferably at least four distinct non-digestible oligosaccharides. In one embodiment, the present composition comprises two distinct non-digestible oligosaccharides.

[0021] In the present embodiment, galacto-oligosaccharides and fructans are preferably present in a weight ratio 5:1 - 20:1 , even more preferably 7:1 - 15:1 , even more preferably 8:1 - 10:1 , most preferably about 9: 1.

[0022] If present, the non-digestible oligosaccharides are present in therapeutically effective amounts. The present composition preferably comprises 0.05 to 20 wt% of said non-digestible oligosaccharides, more preferably 0.5 to 15 wt%, even more preferably 1 to 10 wt%, most preferably 2 to 10 wt%, based on dry weight of the composition. In terms of doses, the present composition preferably provides between 0.05 and 25 grams non-digestible oligosaccharides, preferably between 0.1 and 5 gram non-digestible oligosaccharides per serving.

Further components

[0023] The composition may contain further components that are typically incorporated in nutritional compositions, such as lipid, protein and digestible carbohydrates. Preferably, the present composition is an infant formula, comprising any further component suitable in infant formulae.

[0024] The composition according to the present invention may contain a protein source, preferably in an amount of not more than 2.5 g/100 kcal, preferably 1.6 to 2.2 g/100 kcal. The protein preferably provides 5 to 15 % of the total calories of the composition (en%). Caloric contents can be calculated based on Atwater constants, using the factors 4 kcal/g for protein and carbohydrates, 9 kcal/g for lipids, and 2 kcal/g for fibres. Preferably the composition comprises protein that provides 6 to 12 en%. More preferably, protein is present in an amount below 9 en%, such as 6 to 9 en%. The source of the protein is typically selected in such a way that the minimum requirements for essential amino acid content are met and satisfactory growth is ensured. Hence, protein sources based on cow's milk proteins such as whey, casein and mixtures thereof and proteins based on soy, potato or pea are preferred. Preferably, the composition comprises casein and whey protein. In case whey proteins are used, the protein source is preferably based on acid whey or sweet whey, whey protein isolate or mixtures thereof and may include alpha-lactalbumin and alpha-lactoglobulin. The protein source may be based on acid whey or sweet whey or mixtures thereof and may include alpha-lactalbumin and beta-lactoglobulin in whatever proportions are desired. More preferably, the protein source is based on acid whey or sweet whey from which caseino-glycomacropeptide (CGMP) has been removed. Preferably the composition comprises casein, preferably it comprises at least 3 wt.% casein based on dry weight. Preferably the casein is intact and/or non-hydrolysed. In the context of the present invention, protein includes peptides and free amino acids. The proteins may be intact or hydrolysed or a mixture of intact and hydrolysed proteins.

[0025] The composition may contain a digestible carbohydrate source. Any digestible carbohydrate source conventionally found in infant formulae such as lactose, saccharose, maltodextrin, starch and mixtures thereof may be used although the preferred source of carbohydrates is lactose. Preferably the carbohydrate sources contribute between 35 and 65 en%.

[0026] The composition may also contain a source of lipids. The lipid source may be any lipid or fat which is suitable for use in infant formulas. Suitably the composition contains at least one, preferably at least two lipid sources selected from the group consisting of rape seed oil (such as colza oil, low erucic acid rape seed oil and canola oil), high oleic sunflower oil, high oleic safflower oil, olive oil, marine oils, microbial oils, coconut oil, palm kernel oil and milk fat. The lipid component of the composition suitably provides 2.9 to 6.0 g, more suitably 4 to 6 g per 100 kcal of the composition. When in liquid form, the composition preferably comprises 2.1 to 6.5 g lipid per 100 ml, more suitably 3.0 to 4.0 g per 100 ml. Based on dry weight the infant formula preferably comprises 12.5 to 40 wt% lipid, more preferably 19 to 30 wt%.

[0027] The fat fraction preferably comprises long chain polyunsaturated fatty acids (LC-PUFA). LC-PUFAs are fatty acids wherein the acyl chain has a length of 20 to 24 carbon atoms, typically 20 or 22 carbon atoms, and wherein the acyl chain comprises at least two unsaturated bonds between said carbon atoms in the acyl chain. More preferably, the present composition comprises at least one LC-PUFA selected from the group consisting of eicosapentaenoic acid (EPA, 20:5 n3), docosahexaenoic acid (DHA, 22:6 n3), arachidonic acid (ARA, 20:4 n6) and docosapentaenoic acid (DPA, 22:5 n3), most preferably the present composition comprises at least DHA. LC-PUFA further have anti-inflammatory effects and promote the adhesion of lactic acid producing bacteria to mucosal surfaces, thereby stimulating the development of a healthy microbiota, which are further advantages for use in caesarean section delivered infants. The LC-PUFA may be 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. The present composition preferably comprises at least one of ARA and DHA in phospholipid form. The weight ratio of n-6 to n-3 polyunsaturated fatty acids is preferably in the range of 5: 1 to 15:1 ; more preferably 8: 1 to 10: 1.

[0028] The composition may also contain all vitamins and minerals understood to be essential in the daily diet and in nutritionally significant amounts. Minimum requirements have been established for certain vitamins and minerals to be present in infant formulae. Examples of minerals, vitamins and other nutrients optionally present in the composition include vitamin A, vitamin B1 , vitamin B2, vitamin B6, vitamin B12, vitamin E, vitamin K, vitamin C, vitamin D, folic acid, inositol, niacin, biotin, pantothenic acid, choline, calcium, phosphorous, iodine, iron, magnesium, copper, zinc, manganese, chloride, potassium, sodium, selenium, chromium, molybdenum, taurine, and L- carnitine. Minerals are usually added in salt form. The composition may optionally contain other substances which may have a beneficial effect for the infant, such as lactoferrin, nucleotides, nucleosides, and the like.

Application

[0029] The composition according to the invention is for promoting cognitive functioning of an infant delivered via Caesarean section (C-Section). In the method according to the invention, the composition according to the invention is administered to an infant delivered via Caesarean section. Caesarean section is a surgical procedure where an infant is delivered through an incision made in the mother's abdominal wall, and then through the wall of the uterus. A Caesarean section is typically performed when it is safer for the mother or the infant than a vaginal delivery, or in case the mother prefers to have a caesarean section rather than deliver her infant vaginally.

[0030] Infants born via Caesarean section are known to have an impaired or delayed cognitive functioning, which is attributed to the mode of delivery. Such impaired cognitive function found in infants born via Caesarean section is distinct from impaired cognitive function caused by a disorder such as (neuro)inflammation, as the underlying cause is completely different. In one embodiment the infant does not suffer from (neuro)inflammation. In one embodiment, the target group is infants delivered via Caesarean section which are in need of promoting cognitive functioning.

[0031] Administration of the present composition to the infant may occur through human milk, or by administering to the infant directly. Administration through human milk involves administering the composition according to the invention to the lactating mother, which transfers the active components through the milk to the infant. In both scenarios, administration typically occurs enterally, i.e. directly into the gastrointestinal tract of the infant. Enteral administration includes oral administration, tube feeding, stoma feeding and rectal administration. Oral administration is preferred. In a preferred embodiment, administration occurs directly to the infant. The composition may be in any form known in the art to be suitable for enteral administration, such as in solid form, in semi-solid form or in liquid form. The composition may be an infant formula, preferably in powder form suited to be reconstituted with water, a nutritional supplement, a suppository, pill or tablet. Wherever doses per serving are defined above for the composition according to the invention, the method according to the inventions preferably involves administration of a serving containing said dose.

[0032] In one embodiment, the composition is administered to the infant, provided in the form of (i) a liquid having a volume between 0.5 to 5 ml for oral administration, wherein said composition is preferably administered to said infant with a syringe, pipette or tube; (ii) a (reconstituted) infant formula; or (iii) a suppository, pill or tablet, and wherein said composition in the form of a suppository is administered to the infant rectally. In one embodiment, administration occurs through the mother and the composition is provided in the form of a nutritional supplement.

[0033] In the context of the present invention, promoting cognitive function in infants delivered via Caesarean section may involve normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery. The negative effects on cognition associated with delivery by Caesarean section may be referred to as impaired cognition. The present invention provides a solution to these negative effects on cognition by improving said impaired cognition towards the healthy level found in vaginally-born infants. In one embodiment, the method according to the invention is for correcting the cognitive alterations (or negative effects on cognition) caused by Caesarean section delivery. In the context of the present invention, "cognitive functioning" or "cognition" includes behaviour, memory, learning skills and sociability. In a preferred embodiment, "cognitive functioning" includes at least behaviour.

[0034] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involve at least one, preferably at least two, more preferably at least three, more preferably at least four, more preferably at least five, most preferably all six of: (a) improving social cognition; (b) improving behaviour; (c) reducing anxiety; (d) improving memory; (e) improving learning skills; and (f) reducing susceptibility to stress. Each of these symptoms or conditions may be referred to as being associated with delivery by Caesarean section. Social cognition may also be referred to as social skills, social interaction or social memory. "Anxiety" in the context of (c) particularly refers to "early-life anxiety". In the context of the present invention, "Improving behaviour" particularly refers to "improving responses to external visual, auditory, environmental and/or physical stimuli". "Stress" in the context of (f) particularly refers to "early-life stress". In one embodiment, the method according to the invention is for improving social cognition. In one embodiment, the method according to the invention is for improving behaviour. In one embodiment, the method according to the invention is for reducing anxiety, in particular early-life anxiety. In one embodiment, the method according to the invention is for improving memory. In one embodiment, the method according to the invention is for improving learning skills. In one embodiment, the method according to the invention is for reducing susceptibility to stress, in particular early-life stress. In one embodiment, the method according to the invention is for (e) improving learning skills; and/or (f) reducing susceptibility to stress.

[0035] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involve at least one, preferably at least two, more preferably at least three, even more preferably at least four, even more preferably at least five, even more preferably at least six, most preferably all seven of (g) reducing stress levels and/or stress response; (h) increasing attachment to the mother; (i) improving social interaction; (j) improving novel object recognition; (k) reducing depressive-like symptoms; (I) reducing autism-like repetitive behaviour; and (m) reducing ADHD-like behaviour. Each of these symptoms or conditions may be referred to as being associated with delivery by Caesarean section. "Stress" in the context of (g) particularly refers to "early-life stress". An improved attachment to the mother may be revealed as an improved recognition of the mother, including recognition of mother's smell and voice, in particular "improved attachment to the mother" refers to "improved recognition of the mother's smell". In one embodiment, the method according to the invention is for reducing stress levels and/or stress response, in particular early-life stress levels and/or response. In one embodiment, the method according to the invention is for increasing attachment to the mother. In one embodiment, the method according to the invention is for improving social interaction, or alternatively worded for improving social memory. In one embodiment, the method according to the invention is for improving novel object recognition. In one embodiment, the method according to the invention is for reducing depressive-like symptoms. In one embodiment, the method according to the invention is for reducing autism-like repetitive behaviour. In one embodiment, the method according to the invention is for reducing ADHD-like behaviour. Of the uses (g) - (m) defined herein, use (k) for reducing depressive-like symptoms is particularly preferred. In an especially preferred embodiment, the method according to the invention is for all of (g) - (m).

[0036] In a preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves one or more of the uses (g) - (m) as defined above. In an especially preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves (g) reducing stress levels and/or stress response; and/or (h) increasing attachment to the mother. The method according to the invention may thus also be worded as a method for reducing stress levels and/or stress response; and/or increasing attachment to the mother, of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. In one embodiment, the method is for reducing stress levels and/or stress response. In one embodiment, the method is for increasing attachment to the mother.

[0037] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving social cognition. The method according to the invention may thus also be worded as a method for improving social cognition of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto- oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0038] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving behaviour. The method according to the invention may thus also be worded as a method for improving behaviour of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto- oligosaccharide and a fructan.

[0039] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing anxiety. The method according to the invention may thus also be worded as a method for reducing anxiety of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. "Anxiety" particularly refers to "early-life anxiety". The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan. [0040] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving memory. The method according to the invention may thus also be worded as a method for improving memory of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto- oligosaccharide and a fructan.

[0041] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving learning skills. The method according to the invention may thus also be worded as a method for improving learning skills of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto- oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructans.

[0042] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing susceptibility to stress. The method according to the invention may thus also be worded as a method for reducing susceptibility to stress of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. "Stress" particularly refers to "early-life stress". The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0043] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing stress levels and/or stress response. The method according to the invention may thus also be worded as a method for reducing stress levels and/or stress response of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0044] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves increasing attachment to the mother. The method according to the invention may thus also be worded as a method for increasing attachment to the mother of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0045] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving social interaction. The method according to the invention may thus also be worded as a method for improving social interaction of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0046] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves improving novel object recognition. The method according to the invention may thus also be worded as a method for improving novel object recognition of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve and a mixture of a galacto-oligosaccharide and a fructan.

[0047] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing depressive-like symptoms. In the context of the present invention, "depressive-like symptoms" may also be referred to as "depression". The method according to the invention may thus also be worded as a method for reducing depressive-like symptoms of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. Alternatively, the method may also be worded as a method for treating or preventing depression, preferably preventing depression, in an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve, more preferably without non-digestible oligosaccharides being present, or a mixture of a galacto- oligosaccharide and a fructan, more preferably without B. breve being present. Most preferably, the composition comprises a mixture of a galacto-oligosaccharide and a fructan and no B. breve.

[0048] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing autism-like repetitive behaviour. The method according to the invention may thus also be worded as a method for reducing autism-like repetitive behaviour of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. The composition preferably comprises B. breve, more preferably without non-digestible oligosaccharides being present, or a mixture of a galacto-oligosaccharide and a fructan, more preferably without B. breve being present. Most preferably, the composition comprises B. breve and no non-digestible oligosaccharides.

[0049] In one embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves reducing ADHD-like behaviour. The method according to the invention may thus also be worded as a method for reducing ADHD-like behaviour of an infant delivered via Caesarean section, comprising administering to said infant a composition comprising a therapeutically effective amount of B. breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan. "ADHD-like behaviour" particularly refers to hyperactivity. The composition preferably comprises B. breve and a mixture of a galacto- oligosaccharide and a fructan.

[0050] Whenever the present use is for "promoting", "improving", "increasing" or "decreasing", this typically refers to normalizing the level for infants delivered via Caesarean section towards the level found in vaginally born infants. "Normalizing" refers to bringing the level for infants delivered via Caesarean section closer to the level found in vaginally born infants, most preferably at the level found in vaginally born infants. "Normalizing" may thus also be referred to as "restoring" or "correcting", i.e. promoting from an impaired status and bringing cognition to a level considered the healthy standard.

[0051] To limit the negative impact associated with delivery by Caesarean section on cognition, it is preferred that administration is started as soon as possible after birth. Preferably, the composition is administered to the infant starting at least within the first sixteen weeks after birth, preferably at least within twelve weeks after birth, even more preferably at least within eight weeks after birth, most preferably at least within four weeks after birth. In one embodiment, the composition is administered to the infant delivered via Caesarean section in the first week of life, preferably at least within 5 days after birth, more preferably at least within 3 days after birth.

[0052] Administration may prolong as long as the negative effects on cognitive functioning associated with delivery via Caesarean section prolong, and preferably lasts for as long as at least one week, more preferably 2 weeks to 6 months, most preferably 4 - 8 weeks. In one embodiment, administration is continued up to the age of 1 year, preferably up to 3, 5 or 10 years of age, or even until the infant is 15 or 18 years old. Administration preferably starts as soon as possible after delivery via Caesarean section, such as in the first month after delivery, preferably in the first week after delivery, most preferably within 48 hours after delivery.

[0053] The beneficial effects of the administration of the composition according to the invention typically occur shortly after administration begins, in that the impaired levels start to increase towards the levels found in vaginally born infants, and these effects may prolong over extended time periods, even after administration of the composition according to the invention has halted. As such, the effects may occur in infanthood, childhood, adolescence and/or adulthood. Although most preferable the effects occur throughout infanthood, childhood, adolescence and adulthood, the timing of the effects may depend on the specific effect strived for. In one embodiment, the effects occur in infanthood, typically when the subject has an age of 0 - 24 months, preferably 5 - 18 months. In one embodiment, the effects occur in childhood, typically when the subject has an age of 1 - 14 years, preferably 2 - 8 years. In one embodiment, the effects occur in adolescence, typically when the subject has an age of 12 - 19 years, preferably 14 - 18 years. In one embodiment, the effects occur in adulthood, typically when the subject has an age above 18 years, preferably 18 - 25 years. In one embodiment, the effects occur in childhood, adolescence and/or adulthood. In one embodiment, the effects occur in infanthood and/or childhood. [0054] In a preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves (c) reducing anxiety, (d) improving memory and/or (f) reducing susceptibility to stress in infanthood and/or childhood, preferably when the subject has an age in the range of 0 - 24 months, most preferably 4 - 18 months. In an alternative preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves (a) improving social cognition, (b) improving behaviour and/or (e) improving learning skills in childhood, adolescence and/or adulthood, preferably when the subject has an age in the range of at least 2 years, most preferably 4 - 25 years.

[0055] In a preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves (g) reducing stress levels and/or stress response and/or (h) increasing attachment to the mother in infanthood and/or childhood, preferably when the subject has an age in the range of 0 - 10 years, most preferably 4 - 24 months. In an alternative preferred embodiment, the promoting cognitive functioning and/or normalizing the negative effects thereon involves (i) improving social interaction; (j) improving novel object recognition; (k) reducing depressive-like symptoms; (I) reducing autism-like repetitive behaviour; and/or (m) reducing ADHD- like behaviour in childhood, adolescence and/or adulthood, preferably when the subject has an age in the range of at least 2 years, most preferably 4 - 25 years.

Figures

[0056] Figure 1 depicts the results of the Isolation-induced Ultrasound Vocalization (USV) test (see Example), in number of vocalisations (n) per 3 minutes. NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto-oligosaccharides/fructans.

[0057] Figure 2 depicts the results of the homing test (see Example), in duration on mother's bedding (t) in seconds. NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto- oligosaccharides/fructans.

[0058] Figure 3 depicts the results of the elevated plus maze (see Example), in number of marbles buried (n). NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto- oligosaccharides/fructans.

[0059] Figure 4 depicts the results of the homing test (see Example), in time spent in the open arms of the maze (in % of total time). NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto-oligosaccharides/fructans.

[0060] Figure 5 depicts the results of the forced swim test (see Example), in duration of immobility (t) in seconds. NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto- oligosaccharides/fructans. [0061] Figure 6 depicts the results of the novel object recognition test (see Example), in discrimination index (Dl). NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto- oligosaccharides/fructans.

[0062] Figure 7 depicts the results of the 3-chamber test (see Example), in time spent interacting with the novel animal (in % of total time). NB = natural born; CS = Caesarean section delivered. Treatment groups are: (1 ) Control; (2) B. breve; (3) galacto-oligosaccharides/fructans; (4) B. breve + galacto-oligosaccharides/fructans. Example

Methodology

[0063] Male and female Swiss mice (Harlan EEUU) were mated in the Preclinical Research Facility, Biosciences Institute, University College Cork. The resulting dams and litters were housed in breeding cages in a temperature and humidity controlled room on a 12 h light, 12 h dark cycle (lights on from 07:00-19:00). All experiments were conducted in accordance with the European Directive 2010/63/EEC, the requirements of the S.I No 543 of 2012, and approved by the Animal Experimentation Ethics Committee of University College Cork. The pups were divided in five experimental groups, a natural born (NB) group and four groups of Caesarean section delivered animals (CS). For CS, the four treatment group were as follows: (1 ) Control; (2) B. breve; (3) GOS/FOS; (4) B. breve + GOS/FOS. Dietary intervention was given to mothers as of birth during lactation period and during weaning, when the pup were switched to their mother's diet. Treatments continued throughout all behavioural testing. The probiotic treatment was administered in the drinking water and prebiotics were included in the diet. Probiotics (B. breve M-16V) were freshly incorporated in the drinking water (Clear H2O, MediDrop 75-02-1001 ) every 24 h at a dose of 10⁹ cfu/ml. Prebiotics (galacto-oligosaccharides (scGOS, Vivinal) / fructo-oligosaccharides (IcFOS, Inuline HP) 9/1 (w/w)) were mixed with standard rodent diet AIN93G (Sniff, Germany) up to a final concentration of 1 wt%. The non-prebiotic groups received the same diet without added prebiotics.

[0064] Mother care behaviour test: On postnatal day 7, the mothers with her respectively pups were recorded during a 30 minute period to evaluate typical maternal behaviour of female mice toward the pups. Recorded behaviours includes pup retrieval, licking of pups, nest building, and crouching over the gathered pups in the nest in a lactation position (Noirot, Anim. Behav. 1969b;17:547-550).

[0065] Isolation-induced Ultrasound Vocalization (USV): Testing for isolation-induced USV was performed on postnatal day 9. Pups were isolated one by one from their mother and littermates and placed in a clean plastic container into a sound attenuating chamber. To record vocalizations, an ultrasound sensitive microphone - a bat detector (US Mini-2 bat detector, Summit, Birmingham, USA) tuned in the range of 60-80 kHz - was suspended above the isolated pup and USVs were recorded for 3 minutes. The number of ultrasonic calls was calculated. [0066] Homing test: In the morning of postnatal day 10, the homing test was performed. This test exploits the tendency of immature pups to maintain contact with their mother and siblings. The floor of a clean mouse cage was subdivided into three areas by wire-mesh dividers, one of which was uniformly covered with wood shavings from the home cage, thus containing familiar odour stimuli ("mother's bedding"). The opposite space was covered with wood shavings from the cage of another litter (born at approximately the same time). The central area was covered with clean bedding material. Individual pups were placed in the central area for 1 minute, after which the dividers were removed and the pups were allowed to freely move around for 2 minutes. Total time spent in each area and numbers of crossings were noted.

[0067] Social interaction test: A 3-chamber social test was performed in a three-chamber apparatus (Desbonnet et al., Mol. Psychiatry. 2014; 19(2), 146-8). Animals were placed in a rectangular apparatus divided into three chambers (left and right and a smaller centre chamber) by transparent partitions with small circular openings allowing easy access to all compartments. The test was composed of three sequential 10 min trials: (1 ) habituation (the test animal is allowed to explore the three empty chambers); (2) sociability (an unfamiliar animal is placed in an inner mesh wire cage in either the left or right chambers); (3) social novelty preference (a novel animal is placed into the previously empty inner cage in the chamber, opposite the now familiar animal). All animals were age- and sex-matched, with each chamber cleaned and lined with fresh bedding between trials. For each of the three stages, the behaviour of the test animal was recorded by a video camera mounted above the apparatus.

[0068] Cognition test: Novel object recognition (NOR) is a highly validated test for recognition memory. On the first day, animals were habituated to a square open field box (Perspex sides and base: 30 * 30 * 20 cm) in a dimly lit room, by individually placing animals into the apparatus for three 10 min exploration periods. On the second day, two identical objects were positioned on adjacent corners approximately 5 cm from each wall of the open field and each animal was introduced for a 5 minute exploration period (test session 1 ). Animals were then placed directly back into their home cages. After a 4 h inter-trial interval, one familiar object is replaced with a novel object and each animal is introduced for a further 5 minute exploration period (test session 2). On each day, animals were acclimatized to the testing room for approximately 1 h prior to being introduced in the open field box. Object exploration is defined as when the animal's nose comes within a 2 cm radius of the object. In between trials, objects and testing arenas are cleaned with alcohol wipes. Results are given as discrimination index (Dl), which quantifies the preference of the mouse for a particular object. If Dl = 0, the mouse has no preference between novel and familiar objects. If Dl > 0, the mouse prefers the novel object and if Dl < 0, the mouse prefers the familiar object.

[0069] Marble burying test: Marble burying test reflects the anxiety, repetitive behaviour and autism-like behaviour. Animals who are more anxious engage in active behaviour (defensive marble burying), while animals prone to autism exhibit repetitive behaviour (repetitive marble burying). Animals were placed individually in small cages, in which marbles had been equally distributed on top of a 5 cm thick bed of sawdust and a wire lid placed on top of the cage. Animals are left undisturbed for up to 30 min, after which the number of buried marble (i.e. those more than three- quarters covered by sawdust) are counted.

[0070] Open field test: Each animal was placed in a grey plastic box (32 cm ^χ 40 cm) and tracked and monitored for 10 minutes using recording software. After 10 minutes the animal was returned to its home cage. The distance moved and velocity of movement in the open field are recorded using Ethovision videotracking system (Noldus Information Technology).

[0071] Elevated plus maze (EPM): The EPM is a rodent model of anxiety that is used as a screening test for putative anxiolytic or anxiogenic compounds and as a general research tool in neurobiological anxiety research. Each animal was placed in the centre of an Elevated Plus Maze (a cross shaped maze with 2 open arms and 2 closed arms) and their behaviour is monitored and tracked for 5 minutes, after which the animal is returned to its home cage.

[0072] Forced swim test (FST): The FST is a standard test for assessing depression-like symptoms and for screening antidepressants. The animal is placed in a water filled cylinder (21 cm in diameter; water depth 15 cm) at 23-25 °C for 6 minutes. The behaviour of the animal, in particular the duration of immobility, was scored by a trained observer for the last 4 minutes. The researcher is in the room with the animal for the whole duration of the test.

[0073] Statistical analysis: The results were analysed using 2-way ANOVA. Differences were considered statistically significant at p<0.05. Data are expressed as mean ± standard error of the mean (sem).

Results

[0074] No significant differences between the five treatment groups were observed in the maternal care test (data not shown). Also the dietary intervention did not show significant differences between all treatment groups on weight gain during the entire duration of the experiment (data not shown).

[0075] The results of the USV test, in number of vocalisations n per 3 minutes, are given in Figure 1. An increased number or frequency of vocalisations indicates a higher level of stress or anxiety. The number of vocalisations was significantly higher in the CS-control group (1 ) over all other treatment groups, including CS treatment groups (2), (3) and (4), indicating that administration of β. breve and/or galacto-oligosaccharides and fructans decreases the amount of ultrasonic vocalisation and the stress/anxiety levels in the subject. The level is normalized towards the level found in natural born subjects.

[0076] The results of the homing test are provided in Figure 2, as duration on mother's bedding t in seconds. The CS-control group (1 ) spent significantly less time in the mother's bedding, whereas all other treatment groups spent most of the 120 seconds of the experiment in the mother's bedding. The recognition of the own mother's stimuli was significantly increased and brought to the level found in natural born animals, when the diet contained B. breve and/or galacto-oligosaccharides and fructans. These results indicate that the maternal attachment of subjects delivered via Caesarean section was significantly increased and promoted towards the level found in natural born subjects by administration of B. breve and/or galacto-oligosaccharides and fructans. [0077] The results of the marble burying test are depicted in Figure 3. The number of marbles buried was significantly increased in the CS-control group vs. the NB-control group, indicating that CS delivered animals exhibited increased anxiety-like behaviour. This increase observed for CS subjects was reduced towards the level observed for NB subjects by administration of B. breve and/or galacto-oligosaccharides and fructans. For the individual components, i.e. the CS-B. breve (2) and the CS-galacto-oligosaccharide/fructan group (3), a significant decrease in number of marbles buried was observed compared to the CS control group (1 ), indicating that these dietary interventions are capable of reversing CS-induced anxiety-like behaviour.

[0078] The results obtained in the elevated plus maze (EPM) are depicted in Figure 4. An increased amount of time spent in the open arms is indicative of decreased anxiety. Dietary intervention with B. breve and/or galacto-oligosaccharides and fructans reduced anxiety-like behaviour in the CS groups. Where animals of the CS control group (1 ) spent a slightly decreased amount of time in the open arms of the maze compared to the NB control group, dietary intervention with either B. breve alone or galacto-oligosaccharides and fructans alone significantly increased the time spent in the open arms beyond the level found in the NB control group. In order to confirm that the increased open arm exploration in the CS treatment groups (2) and (3) was not caused by a general increase in locomotor activity, an open field test was also performed, wherein no differences between all groups in the total distance and the speed of movement were seen (data not shown). The results thus confirmed that animals fed with B. breve and/or galacto- oligosaccharides and fructans were not altering horizontal exploratory behaviour and it did not produce hyperactivity.

[0079] The results of the forced swim test (Figure 5) demonstrated that CS delivered animals exhibit an increased depressive-like symptoms, as the CS control group (1 ) remained immobile for a longer duration, compared to the NB control group. Animals with a depressive-like phenotype stop struggling more quickly and spend a longer time in an immobile posture. The CS delivered animals from the dietary intervention groups (2), (3) and (4) showed decreased immobility compared to the CS-control group, which reached significance for the CS-B. breve (2) and the CS- galacto-oligosaccharide/fructan groups (3). These results corroborate that the CS-induced anxiety is normalized towards the levels found in natural born subject by administration of B. breve and/or galacto-oligosaccharides and fructans.

[0080] The novel object recognition test, the results of which are depicted in Figure 6, demonstrated that CS born animals were significantly less able of discriminating a novel object from a familiar one, while for dietary intervention with B. breve and/or galacto-oligosaccharides and fructans, discrimination indices (Dl) were significantly improved in CS groups (2), (3) and (4) (compared to group (1 )). These results demonstrate that the CS-induced impaired cognition is promoted towards the levels found in natural born subject by administration of B. breve and/or galacto-oligosaccharides and fructans.

[0081] The 3-chamber social test demonstrated that CS delivered animals spent significantly less time interacting with the novel animal in the third stage of the experiment, compared to NB animals (Figure 7). A deficit in social interaction for the CS-control mice was thus observed. This CS-induced impaired sociability was countered by dietary intervention with B. breve and/or galacto- oligosaccharides and fructans, promoting the level of sociability towards the level found in NB subjects.

[0082] An extensive battery of behavioural tests has been performed which show that the composition according to the invention, comprising B. breve and/or galacto-oligosaccharides and fructans, is capable of promoting cognitive functioning of subjects delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born subjects. The composition according to the invention is capable of not only ameliorating the negative behavioural and cognitive effects that are associated with Caesarean delivery, but bringing the respective levels towards those found in vaginally delivered infants.

Claims

A method for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, wherein promoting cognitive functioning involves normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery, the method comprising administering to said infant delivered via Caesarean section a composition comprising a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan.

The method according to claim 1 , wherein cognitive functioning includes at least one of behaviour, memory, learning skills and sociability.

The method according to any one of the preceding claims, wherein promoting cognitive functioning or normalizing the negative effects thereon involves at least one of:

(a) improving social cognition;

(b) improving behaviour;

(c) reducing anxiety;

(d) improving memory;

(e) improving learning skills; and

(f) reducing susceptibility to stress.

(g) reducing stress levels and/or stress response;

(h) increasing attachment to the mother;

(i) improving social interaction;

(j) improving novel object recognition;

(k) reducing depressive-like symptoms;

(I) reducing autism-like repetitive behaviour; and

(m) reducing ADHD-like behaviour.

5. The method according to any one of the preceding claims, wherein promoting cognitive functioning or normalizing the negative effects thereon involves reducing depressive-like symptoms.

6. The method according to any one of the preceding claims, wherein the composition comprises a therapeutically effective amount of Bifidobacterium breve and a therapeutically effective amount of the mixture of galacto-oligosaccharide and fructan.

7. The method according to any one of the preceding claims, wherein the Bifidobacterium breve is Bifidobacterium breve M-16V.

8. The method according to any one of the preceding claims, wherein the fructan is selected from fructo-oligosaccharides, fructo-polysaccharides, inulin and mixtures thereof.

9. The method according to claim 8, wherein the fructan comprises fructo-oligosaccharides and/or fructo-polysaccharides, preferably fructopolysaccharides.

10. The method according to claim 8, wherein galacto-oligosaccharide and fructan are present in a weight ratio in the range of 5: 1 - 20: 1 , preferably 7: 1 - 15: 1.

1 1. The method according to any one of the preceding claims, wherein administration occurs through human milk or by administering to the infant directly.

12. The method according to claim 1 1 , wherein administration occurs directly to the infant and the composition is provided in the form of:

(ii) a (reconstituted) infant formula; or

(iii) a suppository, pill or tablet, and if the composition is provided in the form of a suppository, said composition is rectally administered to said infant. 13. The method according to claim 1 1 , wherein administration occurs through human milk and the composition is administered to the lactating mother in the form of a nutritional supplement.

14. The method according to any one of the preceding claims, wherein the composition is administered to the infant starting at least in the first sixteen weeks after birth, preferably at least within twelve weeks after birth, even more preferably at least within eight weeks after birth, most preferably at least within four weeks after birth.

15. Use of a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan in the manufacture of a composition for promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, wherein promoting cognitive functioning involves normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery. A composition comprising a therapeutically effective amount of Bifidobacterium breve and/or a therapeutically effective amount of a mixture of a galacto-oligosaccharide and a fructan for use in promoting cognitive functioning of an infant delivered via Caesarean section towards a cognitive functioning which is similar to the cognitive functioning observed for vaginally born infants, wherein promoting cognitive functioning involves normalizing the negative effects on cognition associated with delivery by Caesarean section towards those of vaginal delivery.