Influence of diets high and low in animal fat on bowel habit, gastrointestinal transit time, fecal microflora, bile acid, and fat excretion.

Epidemiological observations and animal experiments suggest that large bowel cancer is related to serveral factors. Among them, high dietary intakes of animal fat, the presence in the colon of relatively high levels of bile acids, specific patterns of intestinal microflora, slow transit through the gut, and low stool weights. Under metabolic conditions we have observed the effect on these variables of dietes containing 62 or 152 g/day of fat mainly of animal origin in six healthy young men over 4-wk periods. No change attributable to the diet was observed in the subjects' bowel habit, fecal weight, mean transit time through the gut, or in the excretion of dry matter. Total fecal bile acid excretion was significantly higher on the high fat diet (320 +/- 120 mg/day) than on the low fat diet (139.7) +/- 63 mg/day) t test = 7.78 P less than 0.001 as also was the total fecal fatty acid excretion, 3.1+/-0.71 and 1.14+/-0.35 g/day, respectively t test = 11.4 P less than 0.001). The fecal microflora including the nuclear dehydrogenating clostridia were unaltered by the dietary changes as was fecal beta-glucuronidase activity. Dietary changes which increase animal fat intake clearly influence fecal bile acid excretion in a way that would favor the development of large bowel cancer if current theories prove to be true. Dietary fat however has no effect on overall colonic function so other components of the diet must be responsible for the observed associations of bowel cancer with slow transit and reduced fecal bulk.

Bacteria, diet, and large bowel cancer.

The influence of various dietary supplements on the fecal bacteria was studied. Substances were chosen for study because they were known to influence body metabolism or had been implicated in the various hypotheses as the cause of large bowel cancer. No alteration in the fecal flora was demonstrated. The implications of these findings are discussed in the light of other investigations with special reference to our theory concerning the cause of large bowel cancer.

Relative carriage rates of nuclear dehydrogenating clostridia in two populations of different colorectal cancer risk.

Carriage of nuclear dehydrogenating clostridia has been associated with colon cancer and implicated in its aetiology. This study has compared the carriage of these organisms in a British population at high risk for the development of colon cancer with a low risk Nigerian population. Clostridia were found in all of the stools from both populations. Nuclear dehydrogenating clostridia were only found in the stools of the British subjects (32%). These results support the suggestion that the carriage rate of nuclear dehydrogenating clostridia in a population is related to the risk of colon cancer.

Metabolic responses to dietary supplements of bran.

During a metabolic ward study, the addition of dietary fiber in the form of wheat bran biscuits to the diet of five volunteer subjects resulted in an increase in the stool wet weight and fecal solids. The excretion of fecal solids was highly correlated with the intake of unavailable carbohydrates, and fecal losses of water were similarly correlated with fecal excretion of these constituents. The major component of the increase in fecal solids was due to the noncellulosic polysaccharide fraction of dietary fiber. There was an increased fecal excretion of nitrogen fat and energy by most subjects when the supplement was eaten. However, the increased loss of energy in the feces was only 40-80 kcal/day, and therefore a large supplemental intake of dietary fiber had only minor effects on energy metabolism. Supplemental fiber is thus unlikely to induce a useful loss of calories in the management of obesity. The addition of dietary fiber caused an increased excretion of most inorganic constituents, particularly sodium and phosphorus; increased excretion of iron and magnesium was also found in two subjects.

The effects of two dietary fiber supplements on gastrointestinal transit, stool weight and frequency, and bacterial flora, and fecal bile acids in normal subjects.

The effect of dietary supplements of sugar-cane fiber (bagasse), on stool weight, solids, and water content were studied in normal ambulant volunteers over a 9-mo period; a second inpatient study was done with bran supplements. The addition on 10.5 g of bagasse containing 5.1 g of crude fiber to a normal diet containing 3.7 g of crude dietary fiber daily raised the mean fecal weight from 88.3 +/- 6.4 g to 139.7 +/- 10.2 g/day (p less than 0.005). There was also a significant rise in fecal solids and fecal water, although the percentage of water in the stools remained unchanged. Bagasse supplements accelerated gastrointestinal transit when measured by the carmine marker technique. Radiopaque "shapes" showed a trend toward more rapid transit with bagasse supplements. Daily supplements of 39 g of wheat bran or 10.5 g of bagasse increased the total daily excretion of fecal bacteria, but there were no changes in bacteria excreted per gram of feces. The composition of the bacterial flora showed no change. There was increased excretion of fecal acid sterols on the bagasse supplement, but this failed to occur with bran. No changes attributable to fiber supplements occurred in the plasma triglycerides or cholesterol. Future work may define specific dietary fiber supplements for different therapeutic purposes. One fiber may be used as a bulk expander in diverticular disease and another as a hypocholesteremic fiber.

Evidence for a DNA inversion system in Bordetella pertussis.

The expression of virulence-associated genes in Bordetella pertussis can be lost in three ways: phase variation, antigenic modulation, or serotype conversion. The mechanism(s) of these alterations in gene expression is unclear. B. pertussis chromosomal DNA was probed with cloned pin genes from Escherichia coli and cloned hin genes from Salmonella typhimurium. DNA duplex melting temperature experiments indicated significant homology between B. Pertussis chromosomal DNA and both DNA inversion genes. Southern blots using the hin gene probe showed homology with a 15 kb EcoRI fragment of B. pertussis chromosomal DNA. We postulate here that B. pertussis contains a DNA inversion system which may be responsible for serotype conversion or virulence phase change in this organism.

Transferable or mobilisable antibiotic resistance in Shigella dysenteriae types 1, 2, 3, 4, 6 and 7 isolated in Ethiopia during 1974-85.

A total of 199 Shigella dysenteriae isolates resistant to one or more antibiotics and belonging to types 1, 2, 3, 4, 6 and 7 was examined by one-step broth mating with Escherichia coli K12 and, if non-conjugative, additionally by triparental crosses with the conjugative plasmids X and delta. Of the S. dysenteriae type 1 (Shiga's bacillus) strains, 96% harboured conjugative plasmids. During 1974-79, isolates of Shiga's bacillus carried conjugative plasmids coding for ACSSuT (ampicillin, chloramphenicol, streptomycin, sulphonamide, tetracycline) resistance that transferred at low frequencies (less than 10(-4). After 1980, however, about 50% of isolates of Shiga's bacillus with this resistance (R)-type carried conjugative plasmids that transferred at high frequencies (10 degrees-10(-2)) and that expressed the ACT determinant only. The introduction of a new clone of Shiga's bacillus into Ethiopia in 1980 is suspected. Conjugative plasmids coding for SSuT resistance were detected in S. dysenteriae types 2, 3, and 4. Non-conjugative SSu determinants in S. dysenteriae type 3 were mobilised by conjugative plasmids X and delta. R-type CSSuT in strains of types 2 and 7, and R-type ACST in type-3 strains were neither transferable nor mobilisable and are probably determined chromosomally.

The influence of drugs on the response of a cell culture preparation to bacterial toxins.

The influence was studied of lanthanum chloride, chlorpromazine hydrochloride, indomethacin and sodium cromoglycate on the morphological changes induced in Vero cells by the action of the cholera toxin, the thermolabile enterotoxin (LT) and the Vero cell cytotoxin (VT) of Escherichia coli, the enterotoxin of Clostridium perfringens, and the cytotoxin of Clostridium difficile. These drugs were able to inhibit the effects produced by C. difficile cytotoxin but not by the other toxins examined.

Plasmid profiles of Shigella dysenteriae type 1 isolates from Ethiopia with special reference to R-plasmids.

Plasmid profiles of 80 Shigella dysenteriae type 1 (Shiga's bacillus) strains, collected between 1974 and 1985 in Ethiopia, were examined. Strains with the dominant antibiotic-resistance (R-) type--resistance to ampicillin (A), chloramphenicol (C), streptomycin (S), sulphadiazine (Su) and tetracycline (T)--showed two distinct plasmid profiles (PP). Six plasmid bands were demonstrated in "Ethiopian strains" with PP-1A isolated between 1974 and 1982. In mating experiments with these strains, Escherichia coli K12 recipients showed plasmids pYH10a (72 Mda, atypical Inc FIme, coding for ACSSuT resistance). Eight plasmid bands were demonstrated in strains with PP-2A. These strains were first isolated in 1980 and carried plasmid pYH11a (40 Mda, Inc X, coding for ACT resistance). Strains with PP-2A were identical with a "Zairian strain" described elsewhere. Strains with R-type ACT were "Zairian strains" lacking the 5.1- and 4.2-Mda plasmids. Those with R-type CSSuT were temporally clustered in 1978-1980 and carried plasmid pYH12 (58 Mda, Inc B, coding for the same R-type). A trimethoprim-resistant strain (Gimira strain) had a pattern of small plasmids similar to those of the "Zairian strain" and is probably a subclone of the latter. The fact that a limited number of plasmid profiles have remained constant over many years shows the limitation of plasmid profile analysis as an epidemiological tool. However, when the usual profile is known for a given area, identification of a distinctly different pattern becomes easy and epidemiologically useful.

Association of Vibrio cholerae with fresh water amoebae.

An investigation was undertaken to determine whether Acanthamoeba polyphaga SHI and Naegleria gruberi 1518/1e could affect the survival of various strains of Vibrio cholerae in laboratory microcosms. In microcosms pre-inoculated with trophozoites of amoebae, all six strains of V. cholerae tested survived and multiplied during 24 h. In control microcosms without trophozoites of amoebae, survival of the V. cholerae strains was much decreased. Two strains of V. cholerae were used to determine whether V. cholerae might survive ingestion within amoebae and subsequent encystment. Strain 152 was re-isolated from encysting N. gruberi 1518/1e but not from A. polyphaga SHI. Strain 9112 could not be isolated from cysts of either species of amoebae.

The influence of a diet rich in wheat fibre on the human faecal flora.

The effect on the faecal flora of adding wheat fibre to a controlled diet in four healthy volunteers for a 3-week period has been observed. No change in the concentration of the bacteria in the bacterial groups counted was found, although there was a slight increase in total output associated with increased faecal weight. The predominant organisms in all subjects were non-sporing anaerobes, but the dominant species in each subject was different and was unaffected by changing the diet. Similarly, the concentration of faecal beta-glucuronidase detected in two subjects was unaltered and the concentration of clostridia able to dehydrogenate the steroid nucleus found in one subject was unaltered. It is suggested that the faecal microflora is not primarily controlled by the presence of undigested food residues in the large bowel.

Detection of Clostridium perfringens enterotoxin by tissue culture and double-gel diffusion methods.

Culture filtrates of four enterotoxin-producing strains of Clostridium perfringens type A produced morphological changes in cells of 10 of 12 established mammalian cell lines tested, and particularly Vero (African green-monkey kidney) and MDCK (dog kidney) cells. Double-gel diffusion and Vero-cell tests detected enterotoxin in culture filtrates of, respectively, 29 (28%) and 56 (54%) of 103 strains of C. perfringens tested. The feasibility of cell-culture assays for the routine detection of C. perfringens enterotoxin is indicated.

Clostridia isolated from faeces.

Clostridia were isolated from human faecal samples from England, Scotland, Wales, USA, Hong Kong, Uganda and Japan. Strains were identified on the basis of various biochemical reactions and on the type of fatty acid produced from glucose. Analysis of these acids was by gas-liquid chromatography. Clostridium paraputrificum was common in stool samples from England, Scotland, Wales, and USA, but rare in samples from Hong Kong, Uganda and Japan.

Comparison of five assays for the heat-labile enterotoxin of Escherichia coli.

Enzyme-linked immunosorbent assay (ELISA), DNA-DNA hybridisation, Vero cell assay, the Biken test and a new membrane-filter method were compared in the detection of heat-labile enterotoxin (LT) of Escherichia coli. Six subcultures of each of 50 strains of E. coli from the Biken collection were evaluated "blind" in the laboratory. The combined results of the most reproducible tests (ELISA and DNA-DNA hybridisation) were used to calculate the sensitivity and specificity of the other assays. The Vero-cell assay had a high sensitivity (98%) but a lower specificity (91%). The Biken and membrane-filter assays had sensitivities of 58-71% and 77-84% respectively, depending on the type of antiserum used. Only one false positive result was obtained with the Biken test; specificity of the membrane-filter assay was 94-95%. The membrane-filter assay, with anti-cholera toxin, is specific and reasonably sensitive. It has particular advantages over DNA-DNA hybridisation and the Biken test, and it may prove suitable for screening large numbers of E. coli isolates in epidemiological studies in developing countries.

Diploma in Hospital Infection Control (Dip HIC)

The London School of Hygiene and Tropical Medicine (LSHTM) has established a Diploma in Hospital Infection Control (Dip-HIC). The course for this new Diploma is run under the auspices of the Hospital Infection Society (HIS) and the Public Health Laboratory Service (PHLS) and will commence in October 1997. The aim of this course is to provide infection control staff with systematic training in the sciences relevant to hospital infection control which will allow them to provide, and to take responsibility for, a broad-based infection control service. Topics will include the epidemiology of infectious diseases, clinical microbiology, health care economics, statistics, surveillance methods and patient management. The course will be multi-disciplinary and open to UK and overseas students, both medical and non-medical.

Anaerobic bacteria in tropical ulcer--the application of a new transport system for their isolation.

The successful isolation of anaerobes largely depends on adequate transportation of the specimens after collection. We are investigating the role of anaerobes in the pathogenesis of tropical ulcer and have used Hungate tubes with pre-reduced peptone yeast broth and agar as a transportation system. The medium included 0.05% cysteine and 0.03% sodium formaldehyde sulphoxylate as an Eh buffer. Resazurin 1 microgram/ml seved as an Eh indicator. We found the tubes convenient to use and very reliable at maintaining anaerobiasis.

A comparison of the prevalence of campylobacter, Shigellae and Salmonellae in faeces of malnourished and well nourished children in The Gambia and Northern Nigeria.

The proportion of children in whom bacterial pathogens were isolated in faeces was compared in 59 malnourished and 59 age-matched well nourished children in Fajara, The Gambia, with 38 malnourished and 38 age-matched well nourished children in Malumfashi, Northern Nigeria. Campylobacters were identified more frequently in malnourished (10.2%) and well nourished (6.8%) children in The Gambia than in malnourished (2.6%) and well nourished (2.6%) children in Nigeria. Salmonellae were more frequently isolated in the malnourished Nigerian children (28.9%) than in the malnourished Gambian children (3.4%) and were present in similar proportions (5.1% and 5.3%) in well nourished children in both communities. Shigellae were isolated in a few individuals in each group. The differences in the epidemiology of these enteropathogens are discussed with respect to the differences in ecology between these two sub-Saharan, savanna communities.

Bacterial colonization of the upper intestine in mild tropical malabsorption.

The bacterial flora of the upper intestine has been examined in symptomatic expatriate adults with mild tropical malabsorption, without steatorrhoea, persisting for many months after return to a western environment. Seven of the 11 patients had enterobacteria in luminal fluid or mucosal samples in numbers ranging from 10(3) to 10(8) per ml or per g. The most common isolate was Klebsiella pneumoniae (in four cases); Citrobacter freundii, Serratia marcescens and Pseudomonas spp. were also detected. The signficance of bacterial colonization in the pathogenesis of mild tropical malabsorption is discussed.

Survival of toxigenic Vibrio cholerae O1 with a common duckweed, Lemna minor, in artificial aquatic ecosystems.

Cholera epidemics occur twice a year in Bangladesh. During epidemics, Vibrio cholerae O1 are isolated from patients, as well as from the surface water, but the bacteria disappear during inter-epidemic periods. Their reservoirs or sites of survival and multiplication during inter-epidemic period are still unknown. The present survival study in the laboratory explored the role of an aquatic plant, Lemna minor (duckweed), as a possible reservoir. L. minor was added to sea-salt solution at pH 8.5, containing V. cholerae. Survival of V. cholerae on L. minor, in water on which L. minor was floating, and in control water (without L. minor) was monitored at regular intervals. Survival of both environmental and clinical strains of V. cholerae was assessed by viable counts on thiosulphate-citrate-bile salt-sucrose agar. It was observed that both strains survived better on L. minor than in water on which L. minor was floating or in control water. It is suggested that plants may serve as an effective environmental reservoir for V. cholerae either through a non-specific association or by interaction with V. cholerae in commensal relationship.

Effect of fermentation of Ghanaian maize dough on the survival and proliferation of 4 strains of Shigella flexneri.

Fermented and non-fermented Ghanaian maize dough was seeded with approximately 10(7) colony forming units of 4 strains of Shigella flexneri which had been isolated from patients with dysentery. In the non-fermented maize dough (pH 6.2) the shigellae were detectable in large numbers for up to 24 h after exposure. In the maize dough that had been fermented for 3 d (pH 3.2) 3 strains were detectable in small numbers for up to 6 h after inoculation. Thereafter none was isolated. The fourth strain, though detectable for up to 24 h after inoculation, had its numbers reduced considerably. This suggests that traditional methods of food preparation using fermentation have important anti-diarrhoeal functions and the current decline in popularity of such food technologies in certain developing countries may increase the risk of childhood diarrhoea.