Distribution of extended-spectrum cephalosporin resistance determinants in Salmonella enterica and Escherichia coli isolated from broilers in southern Japan.

This study was conducted to investigate the distribution and diversity of extended-spectrum cephalosporin (ESC) resistance determinants in Salmonella enterica and Escherichia coli obtained from the same cecal samples and to provide evidence of transmission of the resistance determinants among these bacteria in broiler farms in southern Japan. Salmonella enterica and E. coli were characterized by serotyping and multilocus sequence typing, respectively. An antimicrobial susceptibility test, plasmid analysis, and identification and localization of resistance genes were performed to determine the relatedness of ESC resistance determinants among the isolates. Of 48 flocks examined, 14 had S. enterica. In total, 57 S. enterica isolates were obtained, 45 of which showed ESC resistance. Extended-spectrum cephalosporin-resistant E. coli were also obtained from all of these ESC-resistant Salmonella-positive samples. ß-Lactamase genes, blaTEM-52 (38 isolates), blaCTX-M-14 (1 isolate), and blaCMY-2 (6 isolates), were carried by conjugative untypable or IncP plasmids detected in the S. enterica serovars Infantis and Manhattan. The ß-lactamase genes blaCTX-M-14 (3 isolates), blaCTX-M-15 (3 isolates), blaSHV-2 (1 isolate), blaSHV-12 (2 isolates), and blaCMY-2 (32 isolates) associated with IncI1-Iγ, IncFIB, IncFIC, IncK, IncB/O, and IncY plasmids were detected in E. coli co-isolates. Restriction mapping revealed similar plasmids in Salmonella Infantis and Salmonella Manhattan and in different sequence types of E. coli. Intraspecies transmission of plasmids was suggested within S. enterica and E. coli populations, whereas interspecies transmission was not observed. This study highlights the importance of plasmids as carriers of ESC resistance determinants.

Measurement of urinary zearalenone concentrations for monitoring natural feed contamination in cattle herds: On-farm trials.

The aims of the present study were to investigate the efficacy of measuring bovine urinary zearalenone (ZEN) concentrations by using a commercially available ELISA method in cattle kept under different feeding conditions to monitor the natural contamination of feeds at the farm level, and to investigate the effects of supplementation of a mycotoxin adsorbent (MA) product in the feed based on urinary ZEN concentration. First, Japanese Black cattle herds kept for breeding (4 herds) and fattening (4 herds) purposes were provided with similar feeding conditions. Then, urinary samples from 5 cows in each herd were collected and analyzed. Second, dairy cows from 1 herd fed with total mixed rations (TMR) were selected. After thorough mixing of the MA (40 g/d) with TMR, the supplemented TMR was fed according to the following schedule: with MA for 2 wk, without MA for 3 wk; then with MA for 2 wk and without MA for 6 wk. Urine samples were collected from cows (n = 6 to 7) and examined before and after each interval. Zearalenone concentrations were measured by the ELISA and liquid chromatography-tandem mass spectrometry methods. The concentration of ZEN and its metabolites was expressed after creatinine (Crea) correction [ZEN or metabolites (pg/mL)/Crea (mg/dL); pg/mg of Crea]. In the first experiment, the urinary concentrations of ZEN and its metabolites were variable in all herds, and significant differences were observed between herds. In 1 fattening herd, in particular, urinary ZEN concentrations were greater (P < 0.001) than in the other 3 herds. This might reflect significant natural ZEN contamination of the feed at the farm level. In Exp. 2, urinary ZEN concentrations displayed peculiar trends after supplementation with MA. After 2 wk of supplementation, a significant decrease of ZEN (P < 0.05) was observed. Zearalenone concentrations remained at a reduced amount during 3 wk without MA supplementation and 2 wk with MA supplementation. When MA was not added to the feed for the next 6 wk, the concentrations increased to the original quantity. These findings indicate the usefulness of measuring concentrations of urinary ZEN and its metabolites not only for monitoring the natural ZEN contamination of cattle feed at the farm level but also for in vivo evaluation of MA function after supplementing feeds with MA.

Temporal distribution and genetic fingerprinting of Salmonella in broiler flocks from southern Japan.

During the 1998 to 2003 period, cecal contents of 4,024 broiler chickens from 252 flocks raised in 63 holdings were examined for Salmonella. The aims were to establish the actual status of the infection, its temporal distribution, prevalent serotype, and common genotype among broiler flocks brought at the slaughterhouse. Collected samples were preenriched in Hajna tetrathionate broth, and after 24 h of incubation, 10 microL of the broth was streaked on selective Rambach agar plate. Suspected scarlet color colonies of Salmonella were cloned on nutrient agar, confirmed through biochemical tests and sero-typed using O and H antigens. Pulsed field gel electrophoresis technique generated DNA fragments banding patterns and established their clonal relatedness. Salmonella was isolated from 563 (14%) samples in 179 (71%) flocks. The flock situation varied from Salmonella-negative holdings (n = 9), positive-flocks from persistently infected holdings (n = 21), and holdings (n = 19) that showed fluctuations with alternating negative and positive flocks for variable time periods. Fourteen holdings (negative, n = 5 and positive, n = 9) were sampled once throughout the study period. Seasonality component was not observed, and salmonellae were found colonizing broiler ceca in warm and cold months. Predominant serovar was Salmonella Infantis (93.3%; n = 525). Macrorestriction fingerprints of Salmonella Infantis using XbaI presumed the isolates to be derived from a common parent. Enhanced discrimination by BlnI digestion produced 3 banding patterns that were closely related genetically and hence epidemiologically related. Such epidemiological information may enable producers to formulate effective control action plan tailored for individual holdings with special emphasis on biosecurity, hygiene, and pest control.