Competitive exclusion of salmonellas from the chick caecum using a defined mixture of bacterial isolates from the caecal microflora of an adult bird.

Colonization of the caeca of newly hatched chicks by Salmonella typhimurium was prevented by oral administration of a mixture of cultures comprising 48 different bacterial strains originating from an adult bird known to be free from salmonellas. The treatment conferred protection to the same degree as that obtained previously with a suspension of adult caecal contents or an undefined anaerobic culture from the same source and was demonstrated in four separate laboratory trials. Examination of the caecal microflora of chicks one day after being given the protective treatment showed that the presence of high levels of lactobacilli and Bacteroides spp. which are not found usually at two days of age in chicks produced under commercial conditions was indicative of the successful establishment of an adult-type microflora. Although the usual method of administering the protective organisms was to dose the chicks directly into the crop, it was also found possible to incorporate the organisms in the drinking water given to the birds at dilutions up to one in five, the maximum tested. When chicks were given the bacterial mixture via the crop and fed on a diet containing 10 mg kg-1 nitrovin and 100 mg kg-1 monensin, the bacteroides failed to establish in the caeca and the birds were not protected against salmonella colonization. However, when the bacterial cultures were incorporated in the drinking water and the chicks given the same feed, normal protection was obtained; possible reasons for these observations are discussed.     

The effect of feeding diets containing permitted antibiotics on the faecal excretion of Salmonella typhimurium by experimentally infected chickens.

Groups of 45 chickens were fed continuously on diets containing 10 or 100 mg./kg. of different ''growth-promoting'' antibiotics and infected orally with a nalidixic acid-resistant mutant of Salmonella typhimurium. The amount of S. typhimurium organisms excreted in their faeces was estimated by culturing them at weekly intervals and in a standard manner on plates of brilliant green agar containing sodium nalidixate, both direct and after enrichment in selenite broth. All of four groups fed diets containing 100 mg./kg. of nitrovin in three different experiments excreted much larger amounts of S. typhimurium than did groups fed antibiotic-free diets. In some, but not all, experiments, larger amounts were also excreted by groups fed diets containing 10 mg./kg. of nitrovin or 10 or 100 mg./kg. of flavomycin or tylosin. Feeding diets containing 10 or 100 mg./kg. of virginiamycin or bacitracin either did not influence or slightly increased the amount of S. typhimiurium excreted. Two groups fed continuously on diets containing 100 or 500 mg./kg. of sulphaquinoxaline for 44 days excreted smaller amounts of the S. typhimurium organisms that did groups fed antibiotic-free diets; no sulphonamide-resistant organisms of the S. typhimurium strain were isolated from the faeces.     

Furazolidone resistance in Salmonella gallinarum: the relationship between in vitro and in vivo determinations of resistance.

Of 22 strains of Salmonella gallinarum isolated from recent outbreaks of infection in poultry in Greece (15). Amman (3), Kenya (2), Lebanon (1) and Yemen (1), 20 were more resistant to furazolidone in vitro than 6 strains that had been isolated in the U.K. in the 1950s; the minimum inhibitory concentration of furazolidone was approximately 0.3 microgram/ml for the sensitive strains and 1.3 or 2.5 microgram/ml for the more resistant strains. Furazolidone given continuously in the food did not control experimental infections in chickens caused by most of the strains that had been classed as more resistant by the in vitro tests. Chloramphenicol, trimethoprim and sulphadiazine or mixtures of the latter two were the best antibiotics for treating these infections, but they were less satisfactory than furazolidone for treating infections caused by the furazolidone-sensitive strains. As a group, the furazolidone-resistant strains and furazolidone-resistant mutants of one of the sensitive strains were less virulent for chickens than the sensitive strains.     

The effect of feeding diets containing avoparcin on the excretion of salmonellas by chickens experimentally infected with natural sources of salmonella organisms

Chickens were readily infected with salmonella organisms when fed diets containing unsterilized bone-meal or provided with drinking water containing a suspension of natural salmonella infected chicken faeces. When fed diets containing avoparcin at concentrations of 10 or 100 mg/kg chickens infected in these ways excreted larger numbers of salmonellas for longer periods than did chickens fed a nonmedicated diet.     

The effect of feeding diets containing avoparcin on the excretion of salmonellas by chickens experimentally infected with natural sources of salmonella organisms.

Chickens were readily infected with salmonella organisms when fed diets containing unsterilized bone-meal or provided with drinking water containing a suspension of natural salmonella infected chicken faeces. When fed diets containing avoparcin at concentrations of 10 or 100 mg/kg chickens infected in these ways excreted larger numbers of salmonellas for longer periods than did chickens fed a nonmedicated diet.     

Further observations on the effect of feeding diets containing avoparcin on the excretion of salmonellas by experimentally infected chickens

Chickens which had been inoculated orally with a nalidixic acid-resistant strain of Salmonella typhimurium were reared on a diet containing different concentrations of avoparcin in an attempt to explain the variation in response to commercial levels of this antibiotic observed by different workers. In one experiment small increases in faecal excretion of the inoculated salmonella occurred at 2.5 and 5.0 mg kg-1, with greater increases between 7.5 and 20 mg kg-1. In a second experiment there was a significant increase in excretion at 7.5 mg kg-1 and in a third experiment in which generally higher excretion rates were detected in all groups, significant increases were observed at 10 and 12.5 mg kg-1 only. In addition avoparcin significantly increased the faecal excretion of S. cholerae-suis, S. dublin and S. arizonae, serotypes not usually associated with poultry-derived food-poisoning in the United Kingdom. It did not increase faecal excretion of S. pullorum. Avoparcin at 10 mg kg-1 appeared to have little effect on the normal intestinal flora of S. typhimurium infected chickens despite the fact that when tested in vitro individual organisms were susceptible to this drug concentration. At 100 mg kg-1 viable counts of intestinal enterococci and Gram-negative anaerobic bacteria were considerably reduced while those of S. typhimurium and Escherichia coli increased. Antibiotic activity due to avoparcin was detectable in the alimentary tract and there was some increase in concentration of the antibiotic in the more distal regions.     

Factors influencing salmonella shedding in broiler chickens

Three variables were included in a study to determine their effect on the incidence of Salmonella typhimurium in broilers challenged at four days of age. Variables included the presence or absence of a feed additive, avoparcin; the use of new or used litter and the initiating dose of salmonella. Cloacal swabs were taken from approximately 600 chicks at weekly intervals for 45 days. At 10(4), 10(6) and 10(8) c.f.u./chick there was a direct association of challenge dose and the incidence of positive chicks for the first several weeks. Chicks raised on used litter showed an appreciable reduction in susceptibility to salmonella when compared to control animals on fresh litter. As the birds approached slaughter age, the influence of litter hygiene and challenge dose diminished under the conditions of this study. Avoparcin in the diet at 10 p.p.m. had no enhancing effect on salmonella shedding at any time during the 45-day sampling period. The implications of competitive exclusion are discussed.     

Factors influencing salmonella shedding in broiler chickens.

Three variables were included in a study to determine their effect on the incidence of Salmonella typhimurium in broilers challenged at four days of age. Variables included the presence or absence of a feed additive, avoparcin; the use of new or used litter and the initiating dose of salmonella. Cloacal swabs were taken from approximately 600 chicks at weekly intervals for 45 days. At 10(4), 10(6) and 10(8) c.f.u./chick there was a direct association of challenge dose and the incidence of positive chicks for the first several weeks. Chicks raised on used litter showed an appreciable reduction in susceptibility to salmonella when compared to control animals on fresh litter. As the birds approached slaughter age, the influence of litter hygiene and challenge dose diminished under the conditions of this study. Avoparcin in the diet at 10 p.p.m. had no enhancing effect on salmonella shedding at any time during the 45-day sampling period. The implications of competitive exclusion are discussed.     

Salmonella infection in chicks following the consumption of artificially contaminated feed

Poultry feed was contaminated artificially with either Salmonella kedougou or S. livingstone using a two-stage mixing process. Intestinal infection became established in a small proportion of birds when feed containing between 0.1 and 0.3 salmonellas/g was given continuously for 2 or 3 weeks from the day of their purchase as 'day-olds' while nearly all birds became infected when the feed contained 100-300 salmonellas/g. Between these limits a dose response was demonstrated in that the proportion of birds becoming colonized with salmonellas increased as the numbers of salmonellas added to feed was increased. Nalidixic acid-resistant strains of both serotypes were used to facilitate the recovery of organisms. The isolation rate was higher from dilutions of caecal contents inoculated directly onto brilliant green agar supplemented with nalidixic acid than it was from swabs of cloacal faeces, even when an enrichment technique was used. This observation confirms that the incidence of salmonella carriage in flocks will be under-estimated if only cloacal faeces are cultured. Of the two serotypes used S. kedougou proved the more efficient colonizer although for both serotypes variation in infection rates was demonstrated in different groups of birds given feed containing comparable numbers of salmonellas.     

The epidemiology of Salmonella infection of calves: the role of dealers

Salmonellas were detected in the environment of 10 of the 12 calf dealers' premises studied. The cleaning and disinfection routines were often ineffective and salmonellas were isolated from 7.6% and 5.3% of the wall and floor samples before disinfection and 6.8% and 7.6% afterwards. Eight different salmonella serotypes were detected, of which the commonest were Salmonella typhimurium, predominantly phage type DT204C, and S. dublin. Plasmid profiles were used to fingerprint S. typhimurium DT204C and the results indicated that with the exception of one of the premises, prolonged salmonella-persistence in the environment was not occurring. Three separate epidemics of salmonellosis in calves were studied by use of plasmid profile analysis. The results illustrated the role of delers, and their subcontractors, in the dissemination of salmonellas. The study concludes with suggestions for methods to reduce the spread of salmonellas in the calf marketing chain.     

Effects of varying the content and proportions of gum arabic and cellulose on caecal volatile fatty acid concentrations in the rat

Adult male rats were fed a synthetic diet containing 5, 10 or 15% by weight of gum arabic (GA) or cellulose (C). Food consumption over the 10 d experimental period was similar in rats fed the 5C and 15C and all the GA diets but was raised in the 10C group. Final body weights were similar in the 5C, 10C and 15C groups and were equally lower in rats fed the GA diets. The wet weight of caecal digesta was the same with all three C diets while digesta wet weight rose with increasing dietary GA content and diarrhoea was observed with the 15GA diet. Concentrations of total caecal VFA were similar in the 5C and 5GA groups and were depressed equally by the 10C and 15C diets. Total VFA did not change in the 10GA rats but were depressed in the 15GA group. Calculation of the mass of caecal VFA showed that this was increased with the 10GA diet but substantially reduced with that containing 15GA, suggesting that VFA were not causative of diarrhoea. In animals fed diets containing mixtures of both fibres to a level of 15% by weight, caecal digesta was the same with 10C+5GA and 7.5C+7.5 GA diets but increased with 5C+10GA. Total VFA concentrations and pool size were unaffected by changing from 10C+5GA to 7.5C+7.5GA but the concentration and mass of propionate was increased. In rats fed 5C+10GA VFA concentration was greatly depressed although their mass was unchanged from the 10C+5GA group. We conclude that VFA concentrations are influenced by the type and concentration of dietary fibre and that fibre mixtures favour production of propionate. Below a certain level of fermentable fibre digesta mass seems to reflect enhanced fermentation but above that level, the presence of unfermented fibre. Differences between the present experiments and previous ones may be due to changes in the large bowel microflora.     

Effect of time in lairage on caecal and carcass salmonella contamination of slaughter pigs

A longitudinal study of the effect of time spent in lairage on salmonellas in the caecum and on the skin surface of 450 slaughter pigs from a single producer was conducted. Pigs were tested in 6 groups at 2 abattoirs, with one-third of a group being slaughtered after 18 h, one-third after 42 h and one-third after 66 h spent in lairage. The salmonella isolation rate from caeca and carcass surfaces increased significantly with increasing time spent in lairage. Salmonellas were isolated from the caeca of 18.5% of pigs held less than 24 h in lairage, 24.1% of pigs held a further 24 h and 47.7% of pigs held for 66 h in lairage before slaughter. The salmonella isolation rates from carcasses were 9.3%, 12.8% and 27.3% for the same groups. Thirteen salmonella serotypes were isolated from the caecal contents and carcasses over the 6 weeks of the trial. One abattoir had a higher salmonella isolation rate from pigs than the other and this was probably related to lairage management. It appeared that lairage is an important factor in the manipulation of the salmonella contamination of pig carcasses.     

The effect of antibiotic therapy on the faecal excretion of Salmonella typhimurium by experimentally infected chickens.

Chickens in groups of 40 were infected orally with a nalidixic acid-resistant mutant of Salmonella typhimurium and then fed continuously on diets containing ampicillin, chloramphenicol, furazolidone, neomycin, oxytetracycline, polymixin, spectinomycin, streptomycin or a mixture of trimethoprim and sulphadiazine. The amount of S. typhimurium excreted in their faeces was estimated at intervals by culture on brilliant green agar containing sodium nalidixate, both direct and after enrichment in selenite broth; the amount of Escherichia coli excreted was estimated by culture on MacConkey agar. The feeding of diets containing 500 mg./kg. of ampicillin, furazolidone, neomycin, polymixin, spectinomycin or streptomycin or 100 mg./kg. of trimethoprim and 500 mg./kg. of sulphadiazine for 46 days reduced to a varying degree the amount of S. typhimurium and E. coli excreted, the greatest reduction in S. typhimurium being brought about by the last treatment. The effect was less obvious when the concentration of the antibiotics in the food was decreased fivefold. An important reason for the very limited effect of some of the antibiotics was the emergence of antibiotic-resistant populations of S. typhimurium and E. coli. High concentrations of antibiotic-resistant organisms also arose in the faeces of the chickens fed diets containing tetracyclines and chloramphenicol, treatments which had no apparent effect on the amount of S. typhimurium and E. coli excreted. Much of the antibiotic resistance encountered was determined by R factors, a particular R factor usually being found in the E. coli populations of individual chickens before it was found in their S. typhimurium populations. No S. typhimurium or E. coli were isolated that possessed R factors determining resistance to polymixin, furazolidone or trimethoprim. No S. typhimurium or E. coli were isolated that were polymixin-resistant and no S. typhimurium that were furazolidone-resistant. The few trimethoprim-resistant S. typhimurium isolated were thymine-dependent. The feeding of diets containing the higher concentrations of trimethoprim/sulphadiazine, neomycin, furazolidone or ampicillin for 9 days reduced the amount of S. typhimurium excreted. After the withdrawal of these diets, the amount of S. typhimurium excreted increased to the numbers found in chickens given ordinary diets throughout; the chickens that had been given trimethoprim/sulphadiazine or furazolidone did not remain faecal excreters of S. typhimurium longer than the chickens that had been given ordinary diets. Similar results were obtained with trimethoprim/sulphadiazine when the start of the 9-day treatment period was delayed for an extra 9 days or when it was extended to 18 days.     

Observations on environmental contamination in a microbiological laboratory.

Contamination of a laboratory environment with pathogenic or non-pathogenic micro-organisms may be relevant to safety of technicians and quality of technical performance. Two widely separated incidents in 1968 and 1974 initiated a study of aspects of the laboratory environment. Water-baths, water of syneresis and portions of salmonella cultures spurting out of the sterilizing flame were examined. The water of water-baths was shown to be contaminated from the fluid cultures incubated in them. This raised questions of potential cross-contamination and reporting of false positives. Water of syneresis was sometimes contaminated with salmonellas. A few quantitative counts were made. The range of counts varied between 16 salmonellas per ml. and 13,000,000 salmonellas per ml. Five hundred portions of salmonella cultures and 571 portions of Shigella sonnei cultures which had spurted from the sterilizing flame were examined. All these samples failed to grow salmonellas or shigellas. Precautions necessary to avoid environmental contamination are briefly discussed.     

The excretion of Salmonella typhimurium in the faeces of calves fed milk substitute

A total of 495 calves in 16 batches were examined (117 calves in 4 batches in 1979 and 378 in 12 batches in 1982). They were purchased in markets, transported by road to a farm in Somerset and reared on a milk substitute diet for a period of up to five weeks. Salmonella typhimurium phage type DT 193 was endemic in 1979 and phage type DT 204c in 1982. The mortality rates in the two years were 9.4% and 1.9% respectively. The causes of death were not investigated although the majority were probably due to salmonellosis. The rate of isolation of S. typhimurium from the rectal faeces of calves in all groups was either zero or relatively low on arrival. It rose to a peak (which was higher in 1979) in the second or third weeks before declining to low levels by the end of the fourth week of residence on the farm. Data from 162 calves, examined twice weekly for four weeks in 1982, indicated that the distribution of infected calves, based on the number of times that S. typhimurium was isolated from each, was not random. The calves could be assigned to two main categories; those from which the organism was never isolated and those from which it was isolated at least twice. This suggested that salmonella infected calves actively excreted the organism. The association between salmonella excretion and medication of sick animals with antibacterial drugs was strongest during the second week. Over the four-week period nearly 40% of the calves found to be excreting S. typhimurium were not treated, indicating a high incidence of subclinical infection. Salmonella excretion by the calves followed a regular pattern and infection was self-limiting within five weeks. The peak in the salmonella excretion rate and the mortality rate were higher in 1979 when phage type DT 193 was the endemic strain. However, in 1982 the calves received 100 p.p.m. furazolidone in their milk ration during the first week of their stay on the farm, and this may have contributed to the differences noted between the two years.     

The excretion of Salmonella typhimurium in the faeces of calves fed milk substitute.

A total of 495 calves in 16 batches were examined (117 calves in 4 batches in 1979 and 378 in 12 batches in 1982). They were purchased in markets, transported by road to a farm in Somerset and reared on a milk substitute diet for a period of up to five weeks. Salmonella typhimurium phage type DT 193 was endemic in 1979 and phage type DT 204c in 1982. The mortality rates in the two years were 9.4% and 1.9% respectively. The causes of death were not investigated although the majority were probably due to salmonellosis. The rate of isolation of S. typhimurium from the rectal faeces of calves in all groups was either zero or relatively low on arrival. It rose to a peak (which was higher in 1979) in the second or third weeks before declining to low levels by the end of the fourth week of residence on the farm. Data from 162 calves, examined twice weekly for four weeks in 1982, indicated that the distribution of infected calves, based on the number of times that S. typhimurium was isolated from each, was not random. The calves could be assigned to two main categories; those from which the organism was never isolated and those from which it was isolated at least twice. This suggested that salmonella infected calves actively excreted the organism. The association between salmonella excretion and medication of sick animals with antibacterial drugs was strongest during the second week. Over the four-week period nearly 40% of the calves found to be excreting S. typhimurium were not treated, indicating a high incidence of subclinical infection. Salmonella excretion by the calves followed a regular pattern and infection was self-limiting within five weeks. The peak in the salmonella excretion rate and the mortality rate were higher in 1979 when phage type DT 193 was the endemic strain. However, in 1982 the calves received 100 p.p.m. furazolidone in their milk ration during the first week of their stay on the farm, and this may have contributed to the differences noted between the two years.     

Cockroaches (Blatta and Periplaneta species) as reservoirs of drug-resistant salmonellas

A total of 221 cockroaches (Blatta and Periplaneta spp.), collected in hospitals, houses, animal sheds, grocery stores and restaurants, in various parts of South Kanara District, a south-west coastal region of India, were studied bacteriologically for the presence of various salmonellas. Salmonellas were isolated from 4.1% of these cockroaches. Nine strains of salmonellas were recovered, belonging to five serotypes--Salmonella bovismorbificans, S. oslo, S. typhimurium, S. mbandaka and S. braenderup, the former two being the commonest serotypes. All salmonellas were resistant to one or other of 11 antibacterial drugs used in the susceptibility test. Isolation of salmonellas from cockroaches collected from the livestock premises and human dwellings suggested that they may act as significant reservoirs of salmonella in nature. Recovery of serotypes, phage types and R-types that were commonly isolated from humans and animals of this locality, suggested a transmission role for cockroaches. By harbouring potentially pathogenic, drug-resistant salmonellas, these wandering arthropods may pose dangerous infective hazards to humans and animals.     

Comparative effects of dietary wheat bran and its morphological components (aleurone and pericarp-seed coat) on volatile fatty acid concentrations in the rat

Adult male rats were fed on diets containing 100 g dietary fibre/kg either as alpha-cellulose or wheat bran or the pericarp-seed coat or aleurone layers prepared from that bran by sequential milling and air elutriation and electrostatic separation. After 10 d, concentrations of total volatile fatty acids (VFA) in caecal fluid were significantly different between groups and fell in the order: aleurone greater than wheat bran greater than pericarp-seed coat greater than cellulose. This ranking probably reflected the ease of fermentation of fibre polysaccharides by colonic bacteria which also resulted in a considerably higher faecal bacterial mass in the aleurone group. Because of the differences in the volume of caecal digesta, the mass of caecal VFA was considerably the highest in the aleurone group, intermediate with wheat bran and equally low in the pericarp-seed coat and cellulose groups. The diet based on aleurone gave a relatively higher proportion of propionate but with both pericarp-seed coat and wheat bran the contribution of butyrate was raised. VFA concentrations in hepatic portal venous plasma were proportional to caecal concentrations with very high (greater than 3 mM) values being recorded in the aleurone group. The findings are discussed in relation to the apparent susceptibility of the morphological components of wheat bran to fermentation by large bowel bacteria.     

Further observations on the excretion of salmonella in the faeces of calves fed milk substitute.

Swabs of rectal faeces were obtained daily for 28 days from 90 calves reared in five batches during the summer of 1983. The calves were purchased unweaned in markets and fed a milk-substitute diet. Salmonella typhimurium phage type DT204c was isolated from calves in four batches and Salmonella newport from one. When the data from the 90 calves were considered together the incidence of salmonellas, excretion rose to peak between 5 and 7 days after purchase before declining to low levels during the fourth week. Salmonella was isolated from 55 (61%) calves; 30 were positive on up to four occasions while 21 and 4 animals respectively were positive between 5 and 11 and 15 and 20 times. In the majority of animals infection was probably subclinical since treatment with antibacterial drugs and excretion of S. typhimurium coincided in four calves only.     

Further observations on the excretion of salmonella in the faeces of calves fed milk substitute

Swabs of rectal faeces were obtained daily for 28 days from 90 calves reared in five batches during the summer of 1983. The calves were purchased unweaned in markets and fed a milk-substitute diet. Salmonella typhimurium phage type DT204c was isolated from calves in four batches and Salmonella newport from one. When the data from the 90 calves were considered together the incidence of salmonellas, excretion rose to peak between 5 and 7 days after purchase before declining to low levels during the fourth week. Salmonella was isolated from 55 (61%) calves; 30 were positive on up to four occasions while 21 and 4 animals respectively were positive between 5 and 11 and 15 and 20 times. In the majority of animals infection was probably subclinical since treatment with antibacterial drugs and excretion of S. typhimurium coincided in four calves only.