Effect of growth promotant usage on enterococci species on a poultry farm

The species population of enterococci isolated from four poultry houses for six grow-outs on one farm was determined. Two houses on the farm were control houses and did not use any antimicrobials, while two other houses on the farm used flavomycin, virginiamycin, and bacitracin during different poultry grow-outs. Litter, chick boxliners, feed, and poultry carcass rinses were obtained from each house and cultured for the presence of enterococci. Nine species of enterococci (Enterococcusfaecalis, E. faecium, E. avium, E. casselifiavus, E. cecorum, E. durans, E. gallinarum, E. hirae, and E. malodoratus) were identified from the study. Enterococcus faecalis was isolated more frequently from chick boxliners (n=176; 92%) and carcass rinses (n=491; 69%), whereas E. faecium was found more frequently in litter (n=361; 77%) and feed (n=67; 64%). Enterococcus faecalis (n=763; 52%) and E. faecium (n=578; 39%) were isolated most often from the farm and houses, regardless of antimicrobial treatment. Fifty-two percent of E. faecalis and 39% of E. faecium were isolated from both control (n=389 and 295, respectively) and treatment (n=374 and 283, respectively) samples. This study indicates that antimicrobial usage on this farm did not alter the resident population of enterococci.     

Antimicrobial susceptibility of Clostridium perfringens isolates of bovine, chicken, porcine, and turkey origin from Ontario

Antimicrobial susceptibilities and toxin types were determined for 275 Clostridium perfringens isolates collected in Ontario in the spring of 2005. Minimal inhibitory concentrations (MICs) of C. perfringens isolates for 12 antimicrobials used in therapy, prophylaxis, and/or growth promotion of cattle (n = 40), swine (n = 75), turkeys (n = 50), and chickens (n = 100) were determined using the microbroth dilution method. Statistical analyses and MIC distributions showed reduced susceptibility to bacitracin, clindamycin, erythromycin, florfenicol, and tetracycline for some isolates. Reduced susceptibility to bacitracin was identified in chicken (64%) and turkey (60%) isolates. Swine isolates had predominantly reduced susceptibility to clindamycin (28%) and erythromycin (31%), whereas bovine isolates had reduced susceptibility to clindamycin (10%) and florfenicol (10%). Reduced susceptibility to tetracycline was spread across all species. No clear reduced susceptibility, but elevated MIC(50) for virginiamycin was found in chicken isolates in comparison with isolates from other species. Toxin typing revealed that C. perfringens type A is the dominant toxin type isolated in this study across all 4 host species.     

Characterization of Salmonella spp. isolated from an integrated broiler chicken operation in Korea

The purpose of this study was to investigate the biological and genetic characterization of persistent Salmonella isolates in an integrated broiler chicken operation, in an attempt to elucidate the source of contamination. From the breeder farm, the hatchery, the broiler farm and the chicken slaughter house of an integrated broiler chicken operation, a total of 6 serotypes were observed. Although S. Heidelberg was not detected in the broiler farm, it was consistently found in the breeder farm, the hatchery and the chicken slaughter house. Also, S. Enteritidis and S. Senftenberg were found in the hatchery and the chicken slaughter house, and the hatchery and the broiler farm, respectively. S. Gallinarum and S. Blockley were found only in the broiler farm, and S. Virchow was only recovered in the chicken slaughter house. Isolated S. Heidelberg, S. Enteritidis and S. Senftenberg strains were divided into 3, 5 and 7 types, respectively, on the basis of all properties. Especially, S. Senftenberg isolates, divided into four types by their antimicrobial resistance patterns, were all obviously the XbaI PFGE pattern. Also, four S. Enteritidis isolates resistant to nalidixic acid showed a difference in phage type and PFGE pattern. Such a different pattern was shown despite Salmonella isolates originating from an integrated broiler operation, suggesting that further epidemiological studies on many integrated chicken companies in Korea are needed.     

Antimicrobial and heavy metal resistance in commensal enterococci isolated from pigs

Antibiotic resistance in animal isolates of enterococci is of public health concern because of the risk of transfer of antibiotic resistance isolates or resistance determinants to consumers via the food chain. In this study, phenotypic and genotypic resistance in 192 pig isolates of enterococci to ampicillin, avilamycin, avoparcin, bacitracin, flavophospholipol, gentamicin, narasin, tetracycline, tiamulin, tylosin, vancomycin, virginiamycin, copper and zinc were investigated by susceptibility test and molecular methods. Resistance rates varied between the species but all isolates were susceptible to ampicillin, avilamycin, avoparcin, gentamicin and narasin but resistant to tetracycline and tylosin and intermediately resistant to copper. Only Enterococcus gallinarum and Enterococcus casseliflavus were resistant to vancomycin and virginiamycin resistance was present in less than half the Enterococcus faecium isolates. Zinc resistance was largely confined to Enterococcus faecalis but bacitracin resistance was uncommon in E. faecalis in comparison with the other species. Tiamulin resistance was common in all species except E. casseliflavus. Resistance to flavophospholipol was detected in most E. faecium isolates and in a high proportion of E. gallinarum, E. casseliflavus and E. hirae/durans but was only found in one isolate of E. faecalis. No tetO, rplC, rplD, vanA, vanB, vatA and vatD genes were found. The presence of ermB, tetL, tetM, tcrB, aac6-aph2, tetK, tetS, vanC1, vanC2, lsaA, lsaB and vatE varied between the species and largely corresponded to the susceptibility phenotype. The findings show that resistance to antibiotics of high clinical significance for nosocomial Enterococcus infections is absent, whereas antimicrobial resistance was detected for some other antibiotics including bacitracin, flavophospholipol, tetracycline, tiamulin, tylosin and virginiamycin.     

Dynamics of Salmonella contamination in a commercial quail operation.

Control of carcass contamination requires knowledge of the source and dynamics of spread of Salmonella in commercial poultry production. We examined Salmonella contamination at a U.S. commercial quail operation. Pulsed-field gel electrophoresis (PFGE) was used to type isolates in order to trace Salmonella throughout this production environment. During a 6-mo survey, Salmonella serotypes hadar, typhimurium, typhimurium variant Copenhagen, and paratyphi were encountered within this poultry operation. Ninety-four percent of the Salmonella isolated from breeder and production houses and from carcass rinses belonged to Salmonella serotypes typhimurium variant Copenhagen and hadar. There were six distinct S. typhimurium variant Copenhagen genetic types, as identified by PFGE, present within this particular poultry operation. Seventy-nine percent of S. typhimurium variant Copenhagen identified from the environment of the breeder and production houses produced the same PFGE pattern. Thirty-eight percent of S. typhimurium Copenhagen isolated from carcass rinses and the breeder house shared the same PFGE DNA pattern. This study demonstrates the transmission of salmonellae throughout this production environment, from the breeders to their progeny and to the birds ultimately processed for human consumption.     

Pulsed-field gel electrophoresis-based subtyping of DNA degradation-sensitive Salmonella enterica subsp. enterica serovar Livingstone and serovar Cerro isolates obtained from a chicken layer farm

Salmonella enterica serovar subsp. enterica Livingstone and serovar Cerro isolates from a commercial egg-producing farm, which had previously been untypeable by pulsed-field gel electrophoresis (PFGE) because of DNA degradation during the PFGE process, successfully gave banding patterns using electrophoresis buffer supplemented with 50 microM thiourea. By PFGE in the presence of thiourea, DNA degradation-sensitive S. enterica serovar Cerro isolates from the commercial egg-producing farm were found to be genetically unrelated to S. enterica serovar Cerro isolates that gave the patterns in the absence of thiourea. Forty-five of 50 (90%) S. enterica serovar Livingstone isolates from the farm showed arbitrarily designated XbaI-digested patterns X1 and X2 that were distinguished by one-band difference and had an identical BlnI-digested pattern. In one of the two layer houses in the farm, the numbers of isolates having the pattern X2 increased from 57% in 1997 to 89% in 1998, whereas virtually all the isolates obtained from the other house in the same period showed the profile X1. This suggests that strains having the pattern X2 might have an advantage to preferentially colonize in the former house.     

Antimicrobial resistant Escherichia coli isolates in cattle and house sparrows on two Czech dairy farms

Rectal smears of calves, cows and young bulls, as well as cloacal smears of house sparrows (Passer domesticus), from farms at the villages of Sumice and Troskotovice, Czech Republic, were examined for E. coli resistant to 12 antimicrobials. The resistant isolates were tested for antimicrobial-resistance genes and integrons. Totals of 40% (n=183), 3% (n=95), 0% (n=33), and 9% (n=54) of Escherichia coli isolates from calves, cows, young bulls and house sparrows, respectively, were antimicrobial resistant. The following genes were identified in cattle E. coli isolates: tetA, tetB (isolates resistant to tetracycline), bla(TEM) (beta-lactams), strA, aadA (streptomycin), sul1, sul2 (sulphonamides), and cat, floR (chloramphenicol). Seven of 16 antimicrobial-resistant calf isolates from the Sumice farm possessed class 1 integrons with the aadA1 gene cassette integrated, 1 kb in size. On the Troskotovice farm, eight of 57 antimicrobial-resistant calf isolates possessed class 1 integrons. Integrons of 1.5kb with the dhfr1- aadA1 gene cassette were found in four isolates, followed by a 1kb integron with the aadA1 gene found in three isolates, and a 1.7kb integron with the dhfr17-aadA5 gene cassette and the phenotype ASSuTSxtNaCipCCfG. The prevalence of resistant E. coli in calves compared to adult cattle was much higher and probably was influenced by oral antimicrobial usage in calves, feeding with milk and colostrum from treated cows, as well as mechanisms unrelated to antimicrobial drug selection. Although house sparrows lived together with the cattle and came into contact with cattle waste on the farm, they were not infected by resistant E. coli isolates with the same characteristics as those found in cattle.     

Antibiotic resistance in faecal bacteria isolated from horses receiving virginiamycin for the prevention of pasture-associated laminitis

Enterococcus faecium, a major cause of potentially life-threatening hospital-acquired human infections, can be resistant to several antimicrobials, such that streptogramin quinupristin-dalfopristin (Q/D) is one of the few antibiotics still effective. Consequently use of the streptogramin virginiamycin as an animal growth promoter was banned in the EU in 1999 as some believed this contributed to the emergence of Q/D resistant E. faecium. Virginiamycin is advocated for preventing equine pasture-associated laminitis, but its effect on equine faecal bacterial Q/D resistance has not been determined. Faecal samples were obtained from horses receiving virginiamycin, horses co-grazing and horses not exposed to virginiamycin. Streptogramin resistant E. faecium were cultured from 70% (21/30) of animals treated with virginiamycin, 75% (18/24) of co-grazing animals and 69% (11/16) of animals not exposed. ermB and vatD genes were detected using real time PCR in 63% and 66% of animals treated with virginiamycin, 75% and 71% of co-grazing animals and 63% and 69% of animals not exposed. Antimicrobial resistance genes were present only in samples which had cultured Q/D resistant E. faecium. There was no significant difference between groups with respect to antimicrobial resistance. The gene load of vatD was significantly (p=0.04) greater in unexposed animals compared to those treated with virginiamycin. The use of virginiamycin to prevent pasture-associated laminitis does not appear to be related to an increased Q/D resistance frequency. However, in view of the high frequency of resistance within all groups, the horse is a reservoir of Q/D resistant genes and clones that potentially could be transferred transiently to humans.     

Genetic diversity of Clostridium perfringens isolated from healthy broiler chickens at a commercial farm

Clostridium perfringens is an important commensal and bacterial pathogen of many animal species. It has particular significance in poultry, where it may cause necrotic enteritis. Our objective was to characterize the population diversity of C. perfringens colonizing healthy birds, and to observe how diversity changed over time. Isolates were obtained from broiler chicken cecal samples in two barns on a single farm, on days 7, 14, 22, 27, 30 and 34 of a single 42-day rearing cycle. Bacitracin was used as a feed additive in one of the barns and withdrawn from the second barn for the duration of the experiment. Each isolate was typed using pulsed-field gel electrophoresis (PFGE) using SmaI restriction endonuclease. A total of 205 cecal isolates from 49 birds were typed, as well as 93 isolates from the barn environment (bedding, drinking water and feces). Eight major PFGE types and 17 subtypes were found in the 298 total isolates. The results show that an optimal sampling strategy would involve a large number of birds, with only a few isolates sampled per bird. The diversity of C. perfringens in this study appears to be low within a single bird, and increases as the bird matures. There was no significant difference in genetic diversity between the two barns. In addition, isolates from fresh fecal samples appear to represent the cecal C. perfringens population accurately, although this was not proven statistically. Antimicrobial susceptibility testing was performed on selected isolates (n=41) representing a cross-section of PFGE types. Based on minimum inhibitory concentration distributions, 95% of the isolates tested were deemed resistant to bacitracin, with a 16 microg/mL breakpoint. Three new cpb2 (beta2 toxin gene) variants were found in the study.     

Phenotypic and genotypic antimicrobial resistance patterns of Escherichia coli isolated from dairy cows with mastitis

Pulsed field gel electrophoresis (PFGE) patterns, susceptibility to 26 antimicrobial agents used in veterinary and human medicine, and prevalence of antimicrobial resistance genes of Escherichia coli isolated from cows with mastitis were evaluated. Among 135 E. coli isolates, PFGE analysis revealed 85 different genetic patterns. All E. coli were resistant to two or more antimicrobials in different combinations. Most E. coli were resistant to antimicrobials used in veterinary medicine including ampicillin (98.4%, >or=32 microg/ml) and many E. coli were resistant to streptomycin (40.3%, >or=64 microg/ml), sulfisoxazole (34.1%, >or=512 microg/ml), and tetracycline (24.8%, >or=16 microg/ml). Most E. coli were resistant to antimicrobials used in human medicine including aztreonam (97.7%, >or=32 microg/ml) and cefaclor (89.9%, >or=32 microg/ml). Some E. coli were resistant to nitrofurantoin (38%, >or=128 microg/ml), cefuroxime (22.5%, >or=32 microg/ml), fosfomycin (17.8%, >or=256 microg/ml). All E. coli were susceptible to ciprofloxacin and cinoxacin. Almost 97% (123 of 127) of ampicillin-resistant isolates carried ampC. Eleven of 52 (21.2%) streptomycin-resistant isolates carried strA, strB and aadA together and 29 streptomycin-resistant isolates (55.8%) carried aadA alone. Among 44 sulfisoxazole-resistant E. coli, 1 isolate (2.3%) carried both sulI and sulII, 12 (27.3%) carried sulI and 10 (22.7%) isolates carried sulII. Among 32 tetracycline-resistant isolates, 14 (43.8%) carried both tetA and tetC and 14 (43.8%) carried tetC. Results of this study demonstrated that E. coli from cows with mastitis were genotypically different, multidrug resistant and carried multiple resistance genes. These bacteria can be a reservoir for antimicrobial resistance genes and can play a role in the dissemination of antimicrobial resistance genes to other pathogenic and commensal bacteria in the dairy farm environment.     

Sequence analysis of E2 glycoprotein genes of classical swine fever viruses: identification of a novel genogroup in Thailand

Thirty classical swine fever viruses (CSFV) isolated in Thailand between 1988 and 1996 were characterised by genetic sequence analysis of a part of their E2 coding regions, comparing the new data with that for representative reference viruses from other countries and continents. Thai isolates were divided into three distinct genogroups, indicating multiple origins for the outbreaks. Eighteen isolates from 1988-1995 form a new genogroup not previously described from any other geographical region. Eleven isolates from 1988-1995 are in the same genogroup as old US and European strains represented by reference strains Alfort 187 and Brescia. The viruses of this group seem to have died out in Europe but still persist in Thailand. One recent isolate from 1996 represents another previously described genogroup being closely related to Italian viruses isolated in the same year.     

Antimicrobial resistance, virulence profiles, and phylogenetic groups of fecal Escherichia coli isolates: a comparative analysis between dogs and their owners in Japan

In this study, fecal Escherichia coli isolates (n=188) from 34 dog-owner pairs and 26 healthy control humans (2 isolates per individual) were tested for susceptibility to 6 antimicrobials and screened for virulence genes. Genetic diversity between canine and owner isolates was evaluated by pulsed-field gel electrophoresis (PFGE). Canine isolates exhibited significantly different rates of resistance to four and two antimicrobials, compared to control and owner isolates, respectively. Of the genes examined, the prevalence of sfa, hly, and cnf genes in canine isolates were higher than in control isolates, but not than in owner isolates. These results suggest that characteristics of owner isolates are somewhat similar to canine isolates, compared to isolates from non-dog owners. In addition, PFGE analysis revealed that transfer of E. coli between owners and their dogs had occurred within 3/34 (8.8%) households. Considering the effects of dog ownership on the population of E. coli isolates from owners, further epidemiological studies are required.     

In-vitro antimicrobial susceptibility of Clostridium perfringens from commercial turkey and broiler chicken origin

The minimum inhibitory concentrations (MIC) of eight antibiotics and two anticoccidial agents were determined for Clostridium perfringens strains isolated from 26 commercial broiler farms and 22 commercial turkey farms. Isolates were obtained from the intestines of birds on the farm or at the processing plant using standard culture and identification techniques. The microbroth dilution test was used to determine the MIC for each compound. Most isolates from chickens had MICs in the range of 2–16 mg/L for tilmicosin, tylosin and virginiamycin, whereas the MICs for avilamycin, avoparcin, monensin, narasin and penicillin were ≤1 mg/L. Most strains from chickens had high MICs (≥64 mg/L) and appeared to be resistant to bacitracin and lincomycin. Most turkey isolates had MICs in the range of 2–16 mg/L for bacitracin, tilmicosin, tylosin and virginiamycin, with strains exhibiting MICs ≤ 1 mg/L for avilamycin, avoparcin, monensin, narasin and penicillin. Several turkey isolates had MICs ≥ 64 mg/L to lincomycin. No attempt was made to associate farm usage of a particular antibiotic to the antibiograms.     

IncN plasmids carrying bla CTX-M-1 in Escherichia coli isolates on a dairy farm

The aim of the study was to compare the prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli bovine isolates on a conventional dairy cattle farm with high consumption of parenteral and intramammary cephalosporins (farm A) and on an organic dairy farm with no cephalosporin use (farm B). ESBL-producing E. coli were isolated from rectal swabs and milk filters by selective cultivation on MacConkey agar with cefotaxime (2mg/l). ESBL genes were identified by polymerase chain reaction (PCR) and sequencing, and the genetic diversity of the isolates was determined by XbaI pulsed field gel electrophoresis (PFGE). Conjugative transfer, incompatibility group, and restriction fragment length polymorphism (RFLP) profiles of the ESBL-carrying plasmids were studied. Higher prevalence (39%, n(rectal samples in cows)=309) of CTX-M-1-producing E. coli isolates was found on farm A compared to farm B (<1%, n(rectal samples in cows)=154; 0%, n(rectal samples in calves)=46). Using PFGE, the isolates from farm A were divided into nine pulsotypes. In all ESBL-positive isolates, the bla(CTX-M-1) gene was carried on 40 kb IncN conjugative plasmids of three related HincII restriction profiles. Horizontal gene transfer through transmission of IncN plasmids harboring bla(CTX-M-1) as well as clonal dissemination of a particular clone seems to be involved in dissemination of CTX-M-1-producing E. coli isolates in cows on the farm using cephalosporins in treating bacterial infections. The study demonstrates a possible role of cephalosporin use in the widespread occurrence of CTX-M-1-producing E. coli on the conventional dairy cattle farm compared to the organic farm.     

Detection of Feline calicivirus (FCV) from Vaccinated Cats and Phylogenetic Analysis of its Capsid Genes

We analysed genogroups of four feline calcivirus (FCV) isolates (FCV-S, H10, Ao198-1 and ML89) obtained from cats that experienced FCV infection after having been vaccinated against FCV. New PCR primer sets (8F/8R, Ao-S/Ao-A, cp-S/cp-A) were also designed, since the conventional Seal primer failed to amplify the target sequences in two samples. The genogroups of the four isolates as well as eight global and 17 domestic strains were determined by phylogenetic analysis of their amino acid sequences. One out of the four strains (25%) isolated in this study, H10, was grouped into genogroup I, along with the vaccine strains F9 and FCV-255. The other three isolates (75%) belonged to genogroup II. Thus, there were more isolates in genogroup II than in genogroup I. However, the antibody values of the four isolates against cat anti-F9 antisera were significantly decreased. There may be no relationship between the neutralizing antibody titre and genogroup. Amino acid sequence alignment of the four isolates showed that only a single amino acid in region C, which is involved in neutralization epitopes, was different in ML89 strain from that of F9. The other three strains, H10, Ao198-1 and FCV-B, shared the same amino acid sequence with F9. Alignment of amino acids for linear epitopes in the F9 strain, which are located at regions D and E, showed variations in 5' hypervariable region (HVR) of E, whereas D and conE had only synonymous substitutions i.e. no change in the amino acid sequence. This mutation in 5' HVR of region E suggested a vaccine breakdown, as the region is known to be essential for antigenicity. The genogroup II FCV is likely to be the cause of the FCV infection in this study, while the vaccine strains belong to genogroup I. Thus, the existing vaccine may need reevaluation for its effectiveness.     

Antimicrobial resistance in Swiss laying hens, prevalence and risk factors

Antimicrobial resistance is an emerging concern to public health, and food-producing animals are known to be a potential source for transmission of resistant bacteria to humans. As legislation of the European Union requires to ban conventional cages for the housing of laying hens on the one hand, and a high food safety standard for eggs on the other hand, further investigations about the occurrence of antimicrobial resistance in alternative housing types are required. In this study, we determined antimicrobial resistance in indicator bacteria from 396 cloacal swabs from 99 Swiss laying hen farms among four alternative housing types during a cross-sectional study. On each farm, four hens were sampled and exposure to potential risk factors was identified with a questionnaire. The minimal inhibitory concentration was determined using broth microdilution in Escherichia coli (n=371) for 18 antimicrobials and in Enterococcus faecalis (n=138) and Enterococcus faecium (n=153) for 16 antimicrobials. All antimicrobial classes recommended by the European Food Safety Authority for E. coli and enterococci were included in the resistance profile. Sixty per cent of the E. coli isolates were susceptible to all of the considered antimicrobials and 30% were resistant to at least two antimicrobials. In E. faecalis, 33% of the strains were susceptible to all tested antimicrobials and 40% were resistant to two or more antimicrobials, whereas in E. faecium these figures were 14% and 39% respectively. Risk factor analyses were carried out for bacteria species and antimicrobials with a prevalence of resistance between 15% and 85%. In these analyses, none of the considered housing and management factors showed a consistent association with the prevalence of resistance for more than two combinations of bacteria and antimicrobial. Therefore we conclude that the impact of the considered housing and management practices on the egg producing farms on resistance in laying hens is low.     

Drug use and antimicrobial resistance among Escherichia coli and Enterococcus spp. isolates from chicken and turkey flocks slaughtered in Quebec,Canada

An observational study was conducted of chicken and turkey flocks slaughtered at federal processing plants in the province of Quebec, Canada. The objectives were to estimate prevalence of drug use at hatchery and on farm and to identify antimicrobial resistance (AMR) in cecal Escherichia coli and Enterococcus spp. isolates and factors associated with AMR. Eighty-two chicken flocks and 59 turkey flocks were sampled. At the hatchery, the most used antimicrobial was ceftiofur in chickens (76% of flocks) and spectinomycin in turkeys (42% of flocks). Virginiamycin was the antimicrobial most frequently added to the feed in both chicken and turkey flocks. At least 1 E. coli isolate resistant to third-generation cephalosporins was present in all chicken flocks and in a third of turkey flocks. Resistance to tetracycline, streptomycin, and sulfisoxazole was detected in > 90% of flocks for E. coli isolates. Antimicrobial resistance (AMR) was observed to bacitracin, erythromycin, lincomycin, quinupristin-dalfopristin, and tetracycline in both chicken and turkey flocks for Enterococcus spp. isolates. No resistance to vancomycin was observed. The use of ceftiofur at hatchery was significantly associated with the proportion of ceftiofur-resistant E. coli isolates in chicken flocks. In turkey flocks, ceftiofur resistance was more frequent when turkeys were placed on litter previously used by chickens. Associations between drug use and resistance were observed with tetracycline (turkey) in E. coli isolates and with bacitracin (chicken and turkey), gentamicin (turkey), and tylosin (chicken) in Enterococcus spp. isolates. Further studies are needed to provide producers and veterinarians with alternative management practices and tools in order to reduce the use of antimicrobial feed additives in poultry.     

The carry-over of Campylobacter isolates between sequential poultry flocks

The carry-over of Campylobacter strains from one flock to a subsequent flock in the same broiler house has been studied using molecular epidemiological techniques. In all, 524 Campylobacter strains, isolated from two sequential broiler flocks from 60 broiler houses, were typed by restriction fragment polymorphism of the polymerase chain reaction (PCR) product of the flaA and flaB genes (fla typing). Selected strains were also typed using pulsed field gel electrophoresis (PFGE). By fla typing, 15 (21%) of the 60 houses with Campylobacter-positive sequential flocks had identical genotypes. In 10 (16% overall) of these houses the strains were also identical by PFGE. The difference in PFGE patterns in the strains from the three remaining houses may be indicative of genetic instability. Overall, these results suggest that carry-over from one flock to a subsequent flock in the same house is a relatively infrequent event and, therefore, that routine broiler house cleansing and/or disinfection is largely adequate to eliminate Campylobacter contamination. An alternative explanation of the low level carry-over is a persistent source or reservoir, external to the environment of the broiler houses.     

Susceptibility of Clostridium perfringens of animal origin to fifteen antimicrobial agents

The minimal inhibitory concentrations of fifteen antibacterial agents were determined by agar-dilution method against 121 strains of Clostridium perfringens isolated from pigs, cattle and poultry. All strains, regardless of host of origin, were susceptible to avoparcin, furazolidone, monensin, nitrovin, penicillin G, ronidazole and tiamulin and resistant to flavomycin. Poultry strains were also susceptible to carbadox, chloramphenicol, erythromycin and virginiamycin. Bacitracin-resistant poultry strains were susceptible to all tested antibacterial agents except tetracycline, but the bacitracin-resistant cattle strains were polyresistant. Porcine strains were susceptible to bacitracin and bovine strains to carbadox. Carbadox-resistant porcine strains were resistant to erythromycin, lincomycin and tetracycline and susceptible to chloramphenicol. Resistance to erythromycin was associated with resistance to lincomycin. High level erythromycin-lincomycin-resistant strains were susceptible to virginiamycin, but the intermediate level erythromycin-lincomycin-resistant cattle strains were resistant to virginiamycin. Resistance to chloramphenicol or erythromycin-lincomycin was always associated with resistance to tetracycline but the reverse was not always true.     

Phenotypic and genotypic heterogeneity of Campylobacter coli within individual pigs at farm and slaughter in the US

The aim of this study was to determine the phenotypic and genotypic diversity of multiple Campylobacter isolates (n = 3 per sample) present within individual (heterogeneity) pig faecal and carcass samples at farm and slaughter, respectively. We isolated 1459 Campylobacter coli (1110 on farm and 349 from slaughter) from 908 pigs and 757 carcasses and characterized them for their antimicrobial susceptibility profile to a panel of six antimicrobials using the agar dilution method. Overall, we detected a significantly higher Campylobacter prevalence at the farm (54.7%) than at slaughter (19%) level (P < 0.05). C. coli isolates were resistant most commonly to tetracycline (66.2%) and erythromycin (53.6%) while fluoroquinolone resistance was detected in isolates (n = 17) only from the farm level. Phenotypic diversity of C. coli isolates at the 4-fold minimum inhibitory concentration levels within the same sample was detected in 38.6% (n = 192) pigs and 40.2% (n = 58) carcass swabs with no significant difference between the two sources (P = 0.72). Phenotypic heterogeneity based on the antimicrobial resistance patterns was observed in 32.5% (n = 162) of the farm samples and in 30.5% (n = 44) carcass swabs at slaughter (P = 0.64). A subset of 40 isolates representing ten pigs and eight carcass samples (originating from separate pigs) were further genotyped by multi locus sequence typing. The observation of phenotypic diversity was replicated at the genotypic level, as it was highlighted by the 22 sequence types which represented the 40 isolates. In conclusion, we detected multiple C. coli subtypes from individual pig or carcass samples indicating unprecedented level of heterogeneity. Our study clearly signifies the importance of testing multiple colonies to make appropriate and valid conclusions in epidemiological-based studies.