A. H. Linton

The biotyping of Escherichia coli isolated from healthy farm animals

A total of 2973 Escherichia coli, isolated from six different groups of animals, were examined for their ability to ferment adonitol, dulcitol, raffinose, rhamnose and sorbose in solid media. Twenty-nine fermentation patterns were recorded although 2443 (82%) of the E. coli belonged to seven of the 32 possible biotypes. Ninety-six O-serotypes were identified within the 2973 E. coli. The number of O-serotypes represented in the 15 most common biotypes ranged from three to 15. Serotypes O8 and O9 were found most commonly in the different groups of animals and several biotypes amongst these two O-serotypes were identified in two or more groups of the animals. The ability of the E. coli to metabolize aesculin, ornithine, salicin and sucrose was also assessed. These test proved less reproducible and were not included in the primary biotyping scheme although their use allowed the enumeration of additional biotypes. The application of biotyping to the study of the ecology of drug-resistant strains of E. coli in five situations is briefly presented.

The persistence of drug resistant Escherichia coli strains in the majority faecal flora of calves.

Two groups of calves, one of three and the other of two animals, were purchased in markets and reared initially on a commercial veal unit for 1 month and 4 months respectively. They were then moved to the Veterinary School, Langford, and kept for a further 6 and 4 months respectively. The animals were sampled weekly and a continual turnover in the strains forming the majority Escherichia coli faecal flora was demonstrated for all calves. Antibacterial-drug resistance, as measured by an Antibiotic Resistance Index (ARI), increased after arrival on the veal unit and persisted at high levels during the whole of their stay. After moving to Langford the ARI fell. Initially there was a reduction in the average number of resistance determinants per resistant strain and then, after a delay of up to 8 weeks, by an increase in the proportion of isolates that were fully sensitive. The source of the sensitive strains was not ascertained, although their appearance was not associated specifically with either weaning or turning out to pasture.

The effects of oxytetracycline on the intestinal Escherichia coli flora of newly weaned pigs

Four recently weaned pigs were dosed orally with oxytetracycline. This caused a rapid increase in the incidence of tetracycline resistance (TcR) among Escherichia coli isolates from the faecal flora. The isolates were differentiated further on the basis of O-serogroup, biotype and resistance pattern. There was no evidence that the administration of the antibiotic selected for a few TcR clones, but rather a relatively large number of TcR strains were identified during the dosing period. Using selective isolation media a proportion of these strains were demonstrated in the minority faecal Esch. coli flora before dosing, while the remainder were recognized for the first time after dosing commenced. The incidence of TcR among Esch. coli isolates also increased after weaning in other pigs which were not dosed with oxytetracycline or any other antibacterial agent. In a proportion of these animals this increase was associated with the dominance of a TcR enteropathogenic serotype (0149:K 91, K 88a, c) in the faecal Esch. coli flora which was probably ingested in small numbers before weaning. The source of other TcR strains was probably the environment in which each pig was placed after weaning.

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.

An epidemiological study of Salmonella in a closed pig herd.

An epidemiological survey of salmonella infection in a closed herd of pigs is reported. The main cause of cross-infection resulted from the movement of animals, with subsequent mixing of infected and clean stock and exposure of non-infected animals to infected faeces. Pen-to-pen infection carried by farm attendants also proved important. An extended survey is at present being conducted to gain further information on the processes of cross-infection.

The persistence of drug resistant Escherichia coli in the intestinal flora of healthy broiler chicks

Antibacterial drugs (oxytetracycline, streptomycin and sulphonamides) were included in the drinking water of healthy broiler chicks from the sixth to the twentieth day of life to select a resistant gut flora. On the twenty-first day the birds were divided into three groups and reared in separate rooms until 100 days of age. One group was housed in cages with wire floors while the others were reared on litter. Faeces from adult hens were added regularly to the litter of one of these groups to determine its effect on the gut flora of the chicks. The ecology of Escherichia coli was studied using O-serotyping, biotyping and antibacterial drug resistogram typing. The proportion of E. coli in the dominant faecal flora resistant to two to four antibacterial drugs increased with time to reach a peak several days after the drugs were withdrawn. Thereafter, the level of drug resistance in the E. coli declined equally in all three groups. The majority of organisms with multiple resistance were derived from biotypes of O-serotypes initially resistant to only one drug and were identified before the drugs were administered. The decline in the level of resistance in the dominant faecal flora after the fourth week was due to the appearance of either new O-serotypes or new biotypes of O-serotypes previously shown to be multiply resistant, and which were either sensitive or resistant to only one drug. It is probably that these new strains were derived from the food since several O-serotypes appeared simultaneously in all three groups of birds.

The survival of multi-antibacterial drug-resistant Escherichia coli and Salmonella typhimurium in stored static slurry from a veal calf unit

Salmonella typhimurium phage type DT 193 survived in small numbers, in stored static slurry derived from veal calves, for the 7-week period of observation. The viable coliform count fell by 1 1/2 logs during the first 2 weeks of storage, thereafter there were only relatively small fluctuations in the coliform population. In all 735 of 752 Escherichia coli isolates examined from eight samples of slurry were resistant to 3-6 antibacterial drugs. There was no dramatic change in the overall level of drug resistance amongst the E. coli with time. Chloramphenicol resistance was recorded in 400 (55%) of the E. coli. It was always associated with multiple resistance, with 96% of the strains being resistant to 5 or 6 drugs, although the proportion of isolates of each of the ten most prevalent O-serotypes resistant to chloramphenicol was variable and ranged between none and 97.5%. The use of biotyping together with O-serotyping indicated that the E. coli population was extremely complex, although certain components of the population remained relatively stable within the dominant flora with time since several of the more common O-serotype/biotype combinations were isolated from more than half of the eight slurry samples examined.

The effect of the management of calves on the prevalence of antibiotic‐resistant strains of Escherichia coli in their faeces

Escherichia coli isolated from six calves, in two groups of three which were managed differently and which received no antibiotics, were differentiated into strains on the basis of O serotype, biotype and resistance pattern. Keeping the calves in the maternal environment for 10, rather than 4 d, was associated with a delay in the time taken for strains of E. coli resistant to antibacterial agents to outnumber, in the faeces, those that were sensitive. These findings indicate that the prevalence of resistant E. coli strains in the intestinal microbiota reflect, in part, their prevalence in the environment and that their level may be influenced by the management of the animal.

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.