A human flora-associated rat model of the breast-fed infant gut.

OBJECTIVES: Bacterial colonization of the infant gut may have important influences on the development of gastrointestinal, respiratory, and allergic disease. Early diet is a major determinant of the gut microflora. It is very difficult to carry out studies in human infants that can investigate the interaction of diet, flora, and mucosa. In this study we have developed an infant human flora-associated (IHFA) rat model to allow such investigation. METHODS: Germ-free infant rats were infected with fecal bacteria from exclusively breast-fed infants and were maintained on a modified infant formula for 8 weeks. The fecal and cecal contents were collected and compared with feces of breast-fed infants for bacterial populations, bacterial metabolites, and enzymes and for the ability to inhibit adhesion of pathogenic bacteria to human mucosal cells. RESULTS: The IHFA cecum and feces were dominated by lactic acid bacteria, Bifidobacterium, and lactobacilli, which were representative of the infant feces. The fecal short-chain fatty acid profile was dominated by acetic and lactic acid in a similar manner to human infant feces. Other bacterial metabolites were similar to those of the human infant. Rat intestinal samples were able to inhibit the adhesion of pathogens to mucosal cells, but to a lesser extent than the human samples. CONCLUSIONS: This IHFA infant model of the intestinal flora of the breast-fed infant is considered valid for studying the effect of diet on bacterial colonization and metabolism.

Dietary fibre in infancy and childhood.

There is very little evidence for the effects of dietary fibre in young children and current dietary guidelines are based on assumptions and data extrapolated from studies in adults. The first years of life may be critical for the establishment of a healthy colonic microflora, as well as good eating habits. The lack of clear and well-founded guidelines for the intake of dietary fibre in childhood may hinder both factors. The fears that a high-fibre diet in children < 5 years of age will lead to growth faltering and mineral imbalance are not well supported in the literature, especially for children in the developed world. Indeed, with the rising levels of obesity, fibre intake may be of benefit in reducing energy intake. A low-fibre diet may also be implicated in the aetiology of childhood constipation and appendicitis. The latest proposals for the definition of dietary fibre include oligosaccharides, which may act as prebiotics. There are potential health benefits of including oligosaccharides in the diets of infants and children, but more research is needed to consider the long-term effects. The immature intestine of the infant may also result in a greater amount of starch entering the colon during weaning, and this starch would now be considered dietary fibre under the new definitions. Much new research is needed to allow adequate recommendations for the intake of dietary fibre in childhood based on data collected in appropriate age-groups.

Intestinal flora during the first months of life: new perspectives.

Increasing awareness that the human intestinal flora is a major factor in health and disease has led to different strategies to manipulate the flora to promote health. The complex microflora of the adult is difficult to change in the long term. There is greater impact of diet on the infant microflora. Manipulation of the flora particularly with probiotics has shown promising results in the prevention and treatment of diarrhoea and allergy. Before attempting to change the flora of the infant population in general, a greater understanding of the gut bacterial colonisation process is required. The critical stages of gut colonisation are after birth and during weaning. Lactic acid bacteria dominate the flora of the breast-fed infant. The formula-fed infant has a more diverse flora. The faeces of the breast-fed infant contain mainly acetic and lactic acid whereas the formula fed-infant has mainly acetic and propionic acid. Butyric acid is not a significant component in either group. The formula-fed infant also has higher faecal ammonia and other potentially harmful bacterial products. The composition of the microflora diversifies shortly before and particularly after weaning. The flora of the formula-fed infant develops more quickly than that of the breast-fed infant. Before embarking on any strategy to change the flora, the following questions should be considered: Should we retain a breast-fed style flora with limited ability to ferment complex carbohydrates? Can pro- and prebiotics achieve a flora with adult characteristics but with more lactic acid bacteria in weaned infants? Are there any health risks associated with such manipulations of the flora?

Colonic fermentation capacity in vitro: development during weaning in breast-fed infants is slower for complex carbohydrates than for sugars.

Fresh feces from 27 healthy infants-12 breastfed (complete, exclusive breast-feeding), 7 in early weaning (partial, high breast-feeding), and 8 in late weaning (partial, low breast-feeding)-were cultured with simple and complex carbohydrates in vitro to test the hypothesis that colonic fermentation capacity for carbohydrates increases during weaning. Infants in all three groups were able to ferment sugars, with no significant differences in median total short-chain fatty acid (SCFA) concentrations (mmol/L): preweaning, 56.4(range: 0-77.6); early weaning 68.5(range: 57.9-98.8); late weaning, 61.3(range: 28.6-120.4) for glucose. Preweaned infants were less able to ferment oligosaccharides and complex carbohydrates than were weaned infants (P < 0.05). Ability to ferment raftilose was higher in early weaning; median total SCFA concentrations (mmol/L) were as follows: preweaning 31.0 (range: 3.6-48.9), early weaning 57.1 (range: 2.5-70.6), late weaning 68.6 (range: 22.0-113.4) (P < 0.05). Ability to ferment complex carbohydrates did not develop until late weaning; median total SCFA concentrations for guar gum (mmol/L) were as follows: preweaning 6.4 (range: 0.1-57.3), early weaning 18.4 (range: 0.0-40.5), late weaning 45.4 (range: 15.6-62.1) (P < 0.05, preweaning and early weaning compared with late weaning). Development of the ability to ferment complex carbohydrate was slow. Cultures of feces from preweaned infants produced eight times more SCFAs with glucose than with complex carbohydrates, at early weaning there was a threefold difference and by late weaning the difference was only 25%, but this was still only 42% of the SCFAs produced by cultures of adult feces. These data suggest that for the complex carbohydrates tested, colonic fermentation is likely to contribute only a small proportion of daily energy needs of weaning infants.

In vitro fermentation of carbohydrate by breast fed and formula fed infants.

Unabsorbed carbohydrates are fermented by colonic bacteria to short chain fatty acids (SCFA) which are rapidly absorbed, salvaging energy and reducing stool output. There are marked differences between the faecal flora and SCFA of breast fed (BF) and formula fed (FF) infants which may be related to the higher incidence of diarrhoea in FF infants. Part of this effect may be caused by a difference in the ability of the microflora to ferment carbohydrate. To test the hypothesis that BF and FF have different fermentation capacities for simple and complex carbohydrates, in vitro cultures of faeces from healthy infants (2-10 weeks; 11 BF, 11 FF) containing glucose, lactose, raftilose (a fructo-oligosaccharide), or soybean polysaccharide were incubated anaerobically. Results were compared with those of adult faecal cultures using the same carbohydrates. Cultures of faeces from BF and FF infants produced comparable amounts of total SCFA in all cultures. These cultures produced less SCFA than those from adult faeces and produced very little SCFA from complex carbohydrate. BF cultures produced more acetic acid than FF in all cultures, whereas FF cultures produced more propionate with sugars and more butyrate with raftilose. Both groups of infants produced less butyrate than adults in all cultures. Thus it is unlikely that a lower ability to ferment carbohydrate is a major cause of increased risk of diarrhoea in FF fed infants but individual SCFA production may be important.

Faecal short chain fatty acids in breast-fed and formula-fed babies.

The intestinal flora of breast-fed infants differs from that of formula-fed infants. It is thought that this difference in flora may be one important reason why breast-fed babies suffer less from gastrointestinal disease. Differences in intestinal flora are reflected in the profile of faecal short chain fatty acids (SCFA). Very little is known about faecal concentrations of SCFA in babies fed breast milk or infant formula. In this study, faecal SCFA were measured in babies at two and four weeks of age who had been either exclusively breast fed or bottle fed from birth. There was no significant difference in total faecal SCFA concentrations between breast-fed and formula-fed babies when lactate was included. The formula-fed group, however, had less lactic acid and higher concentrations of propionic and n-butyric acids than breast-fed babies. Very few babies had significant levels of n-butyric acid, although this SCFA is believed to be important for the health of the colonic mucosa of adults.

Characterisation of a series of human immunodeficiency virus isolates derived sequentially from a single patient.

Five HIV-1 isolates were obtained sequentially from a single seropositive individual during the later stages of AIDS. Four of these isolates were adapted to grow in a continuous human T-lymphocytic cell line. Comparative biological and biochemical studies of the virus isolates were made using persistently infected cultures or virus derived from these systems respectively. The data obtained clearly shows that viruses with different biological properties can be isolated from the same individual at different times during the course of clinical AIDS.

Establishment and characterization of a human CD4 positive cell bank for HIV related studies.

The human CD4 positive cell lines JM, CCRF, CEM, U937, HL60 and THP-1 have been cleared of mycoplasma contamination and defined by DNA fingerprinting and cell surface phenotype marker analysis. These cells have been banked and are now available as a source of standardized cell lines for HIV related research.