Bacteriological examination of specified-pathogen-free animals: a procedure for small rodents

Organs of 10 mice or 5 rats were pooled and homogenized. A portion of the homogenate was cultured on several selective and enrichment media. The method was tested by adding various pathogenic organisms to organ homogenates. 10(3)-10(4) microorganisms in homogenates of 10 organs were readily detected. This procedure allows even a small microbiological laboratory to detect a low frequency of infected animals in a colony.     

Reconstitution of the gastrointestinal microflora of lactobacillus-free mice.

A colony of mice that do not harbor lactobacilli in their digestive tracts but whose intestinal microflora is otherwise functionally similar to that of conventional animals was derived. Methods used to reconstitute the intestinal microflora of the mice included inoculation of the animals with cultures of specific microbes, noncultivable microbes attached to epithelial cells, and cecal contents from conventional mice treated with chloramphenicol. Twenty-six microflora-associated characteristics were monitored by using relatively simple tests to determine the microflora status of the mice.     

Development changes to gut microflora metabolism in mice.

Developmental changes in the activities of bacterial nitrate reductase, nitroreductase and beta-glucuronidase and their response to fermentable dietary fibre, were investigated in caecal contents from suckling mice (2-week-old) and in mice aged 4-24 weeks fed either a purified fibre-free diet or that diet supplemented with 5% (w/w) pectin. There was no apparent age-related trend common to the three enzymes studied. Nitrate reductase activity in the mice fed the fibre-free diet did not markedly alter with age. Pectin administration, however, was associated with a significant increase in nitrate reductase activity, particularly in 4-week-old mice. Nitroreductase activity exhibited an overall upward trend in mice from 2 to 12 weeks and thereafter decreased. Caecal beta-glucuronidase activity in mice increased sharply between 2 weeks and 4 weeks of age, thereafter not changing significantly until the 24th week. Pectin feeding had no consistent effect on activities either of nitroreductase or beta-glucuronidase. The changes in enzyme activities with age were not related to the concentration of bacteria in the caecum, which was highest in the 2-week-old mice. We conclude that the weaning is a period in which marked changes in caecal bacterial enzyme activities can occur.     

Intestinal microflora and bile acids. Effect of bile acids on the distribution of microflora and bile acid in the digestive tract of the rat

The population levels of intestinal microflora and bile acid composition in the digestive tract were examined in rats fed bile acids to determine the relationships between gastrointestinal microflora and the host. The population level of Bacteroides was increased in the ceca of rats fed cholic acid or deoxycholic acid. In the ileum, the concentration of conjugated bile acid in rats fed cholesterol, cholic acid, hyodeoxycholic acid or lithocholic acid was higher than that in control rats, and was very low in ceca and feces of all the rats. The concentration of total free bile acid was much higher in the ceca than in the ilea of rats fed hyodeoxycholic acid or lithocholic acid. Cholic acid and deoxycholic acid were found in the ilea, ceca and feces of the cholic acid-fed rats. In the deoxycholic acid-fed rats, cholic acid was localized in the ileum. 7-Ketodeoxycholic acid was also found in the ceca of the cholic acid-fed rats. 12-Ketolithocholic acid was found in the feces of rats fed cholic acid or deoxycholic acid. 3-Ketocholanic acid was found in some samples from the lithocholic acid-fed rats. Therefore, some kinds of bile acids influence the population levels of gastrointestinal microflora and bile acid composition in the intestine.     

Detection of QTL controlling digestive efficiency and anatomy of the digestive tract in chicken fed a wheat-based diet

Background

Improving digestive efficiency is a major goal in poultry production, to reduce production costs, make possible the use of alternative feedstuffs and decrease the volume of manure produced. Since measuring digestive efficiency is difficult, identifying molecular markers associated with genes controlling this trait would be a valuable tool for selection. Detection of QTL (quantitative trait loci) was undertaken on 820 meat-type chickens in a F2 cross between D- and D+ lines divergently selected on low or high AMEn (apparent metabolizable energy value of diet corrected to 0 nitrogen balance) measured at three weeks in animals fed a low-quality diet. Birds were measured for 13 traits characterizing digestive efficiency (AMEn, coefficients of digestive utilization of starch, lipids, proteins and dry matter (CDUS, CDUL, CDUP, CDUDM)), anatomy of the digestive tract (relative weights of the proventriculus, gizzard and intestine and proventriculus plus gizzard (RPW, RGW, RIW, RPGW), relative length and density of the intestine (RIL, ID), ratio of proventriculus and gizzard to intestine weight (PG/I); and body weight at 23 days of age. Animals were genotyped for 6000 SNPs (single nucleotide polymorphisms) distributed on 28 autosomes, the Z chromosome and one unassigned linkage group.

Results

Nine QTL for digestive efficiency traits, 11 QTL for anatomy-related traits and two QTL for body weight at 23 days of age were detected. On chromosome 20, two significant QTL at the genome level co-localized for CDUS and CDUDM, i.e. two traits that are highly correlated genetically. Moreover, on chromosome 16, chromosome-wide QTL for AMEn, CDUS, CDUDM and CDUP, on chromosomes 23 and 26, chromosome-wide QTL for CDUS, on chromosomes 16 and 26, co-localized QTL for digestive efficiency and the ratio of intestine length to body weight and on chromosome 27 a chromosome-wide QTL for CDUDM were identified.

Conclusions

This study identified several regions of the chicken genome involved in the control of digestive efficiency. Further studies are necessary to identify the underlying genes and to validate these in commercial populations and breeding environments.     

Sterilization efficiency of a cascaded dielectric barrier discharge

AIMS: To investigate the microbial inactivation efficiency of a newly developed cascaded dielectric barrier discharge (CDBD) set-up against various micro-organisms on polyethylene terephthalate (PET) foils. METHODS AND RESULTS: Inactivation kinetics in dependency of time were produced with air as process gas and test strains like Salmonella serotype Mons, Staphylococcus aureus and Escherichia coli and spores of Bacillus atrophaeus, Aspergillus niger and Clostridium botulinum, which were homogeneously distributed on the sample surface by a spray method. Highest count reduction was observed for the vegetative cells with at least 6.6 log(10) within 1 s. Aspergillus niger was the most resistant test strain with an inactivation rate of about 5 log(10) in 5 s. CONCLUSIONS: For industrial applications it is necessary to evaluate new sterilization methods against a broad range of different micro-organisms. SIGNIFICANCE AND IMPACT OF THE STUDY: CDBD plasma is a fast and effective technology for decontamination of heat sensitive materials in few seconds.     

Reconstitution of the gastrointestinal microflora of lactobacillus-free mice

A colony of mice that do not harbor lactobacilli in their digestive tracts but whose intestinal microflora is otherwise functionally similar to that of conventional animals was derived. Methods used to reconstitute the intestinal microflora of the mice included inoculation of the animals with cultures of specific microbes, noncultivable microbes attached to epithelial cells, and cecal contents from conventional mice treated with chloramphenicol. Twenty-six microflora-associated characteristics were monitored by using relatively simple tests to determine the microflora status of the mice.     

Radioautographic studies on radiosulfate incorporation in the digestive organs of mice.

The sulfate uptake and accumulation in mouse digestive organs were studied by light microscopic radioautography. Two litters of normal ddY mice 30 days after birth, each consisting of 3 animals, were studied. One litter of animals were sacrificed 30 min after the intraperitoneal injections with phosphate buffered Na2(35)SO4, and the other litter animals were sacrificed 12 hr after the injections. Then several digestive organs, the parotid gland, the submandibular gland, the sublingual gland, antrum and fundus of the stomach, the duodenum, the jejunum, the ileum, the caecum, the ascending colon and the descending colon were taken out. The tissues were fixed, dehydrated, embedded in epoxy resin, sectioned, picked up onto glass slides, coated with radioautographic emulsion by a dipping method. AFter the exposure, they were developed, stained with toluidine blue and analyzed by light microscopy. As the results, many silver grains were observed on serous cells of the salivary glands, mucosa and submucosa of the stomach, villous cells and crypt cells of the small intestines and whole mucosa of the large intestines at 30 min after the injection. Then at 12 hr after the injection silver grains were observed on mucous cells of the salivary glands, some of the stomach glands, and mucigen granules of goblet cells in the small intestines and the large intestines. The numbers of silver grains observed in respective organs at 30 min were less than those at 12 hr. From these results, it is concluded that glycoprotein synthesis was demonstrated in several digestive organs by radiosulfate incorporation. In the salivary glands the silver grains were more observed in serous cells at 30 min, while in mucous cells more at 12 hr than 30 min after the injection. In other organs the silver grains were more at 30 min than at 12 hr. These results show the time difference of glycoprotein synthesis in respective organs.     

Chitinolytic enzymes and the bacterial microflora in the digestive tract of cod, Gadus morhua

The levels of chitinolytic enzymes and chitinolytic bacteria in the digestive tract of feeding and fasting cod were compared. Enzyme activities within a given tissue were of a similar order irrespective of the presence of chitinolytic bacteria and/or chitin. Cod chitinase and chitobiase are therefore endogenous and constitutive enzymes. Fasting cod had similar numbers of bacteria within the gastro-intestinal compartments as feeding fish. Representative bacterial isolates from fasting fish were characterized.     

Absence of transformation of beta-muricholic acid by human microflora implanted in the digestive tracts of germfree male rats.

Germfree rats biosynthetize cholic and beta-muricholic acids. The latter does not exist in humans. Germfree rats were given human fecal suspensions. These rats degraded cholic acid into deoxycholic acid but failed to metabolize beta-muricholic acid.     

Peritrophic membrane role in enhancing digestive efficiency. Theoretical and experimental models

The peritrophic membrane (PM) is an anatomical structure surrounding the food bolus in most insects. Rejecting the idea that PM has evolved from coating mucus to play the same protective role as it, novel functions were proposed and experimentally tested. The theoretical principles underlying the digestive enzyme recycling mechanism were described and used to develop an algorithm to calculate enzyme distributions along the midgut and to infer secretory and absorptive sites. The activity of a Spodoptera frugiperda microvillar aminopeptidase decreases by 50% if placed in the presence of midgut contents. S. frugiperda trypsin preparations placed into dialysis bags in stirred and unstirred media have activities of 210 and 160%, respectively, over the activities of samples in a test tube. The ectoperitrophic fluid (EF) present in the midgut caeca of Rhynchosciara americana may be collected. If the enzymes restricted to this fluid are assayed in the presence of PM contents (PMC) their activities decrease by at least 58%. The lack of PM caused by calcofluor feeding impairs growth due to an increase in the metabolic cost associated with the conversion of food into body mass. This probably results from an increase in digestive enzyme excretion and useless homeostatic attempt to reestablish destroyed midgut gradients. The experimental models support the view that PM enhances digestive efficiency by: (a) prevention of non-specific binding of undigested material onto cell surface; (b) prevention of excretion by allowing enzyme recycling powered by an ectoperitrophic counterflux of fluid; (c) removal from inside PM of the oligomeric molecules that may inhibit the enzymes involved in initial digestion; (d) restriction of oligomer hydrolases to ectoperitrophic space (ECS) to avoid probable partial inhibition by non-dispersed undigested food. Finally, PM functions are discussed regarding insects feeding on any diet.     

A comparison of digestive efficiency in birds of prey

Differences in how raptors hunt and what they catch are correlated with size-independent differences in length of the small intestine, the region of the digestive tract responsible for food absorption. The study examined the functional significance of these differences among ten raptor species. Dry matter apparent digestive efficiency was calculated for each species fed a diet of day-old cockerels. For Falconiformes and Strigiformes, efficiencies varied between 75% and 82%, digestive efficiency being positively correlated with intestine length.
Generalist species, with a wide prey spectrum and feeding on relatively easily caught prey and carrion, had long intestines and high digestive efficiencies. Raptors specializing on fast-moving, avian prey had short intestines and reduced digestive efficiency. The Peregrine Falco peregrinus and the Common Buzzard Buteo buteo were used as examples of specialist and generalist feeders, respectively. Rabbit and pigeon were fed to both. Buzzards digested both diets more efficiently than Peregrines. Body-mass changes were used to examine the nutritional value of the two diets to each species. Buzzards gained body-mass when eating rabbit, Peregrines lost mass. Both species gained mass when feeding on pigeon. It seems that consideration of diet quality, not just quantity, is essential in understanding raptor food requirements. Generalist raptors have high efficiencies on several diets, specialists compensate for their reduced efficiency by eating food of high nutritional quality. Various aspects of prey quality are examined.