| The effect of administering feedlot cattle subtherapeutic levels of chlortetracycline (CT) or CT and therapeutic levels of oxytetracycline (CT-OX) on resistance genotype in Escherichia coli was investigated. Fecal samples were collected from cattle at commercial feedlots and had a documented history of antimicrobial use. Isolates were tested for susceptibility to tetracycline, oxytetracycline, and chlortetracycline using disk diffusion or broth microdilution. Detection of tet(A), tet(B), and tet(C) genes encoded by tetracycline-resistant isolates (N=176) was performed by multiplex PCR. All isolates encoded one or a combination of two resistance genes. Prevalence of tet(A) was similar between groups of E. coli, however prevalence of tet(B) was lower(P<0.5) and tet(C) was greater(P<0.5) in CT isolates. The nature of the tet determinants was further assessed in a group of intermediately tetracycline-resistant isolates (N=52). Minimum inhibitory concentrations showed that susceptibility was dependent on tetracycline analogue and the type of resistance determinant. The tet(C) gene was present in92%of these isolates. Copies of tet(C) transcripts, analyzed by real-time PCR, indicated that up-regulation did not occur in tetracycline-resistant isolates when compared to intermediately-resistant isolates. However, sequence analysis of the tet(C) gene revealed a Tâ†'G substitution at position1063in resistant isolates that may have affected phenotype. These data provide insight into the relationship between the type of tetracycline analogue administered to cattle and the prevalence of resistance genes in E. coli.The beef cattle were separated into subtherapeutic tetracycline treated group and non-treated group, and the fecal samples and pharyngeal secretion samples were collected at≥60DOF, and another animals were collected before and after tetracycline treat. The metagenomic DNA was extracted to quantify the tetracycline resistant gene, sulfanilamide resistant gene, and erythrocin resistant gene. The results showed that, the concerntration of tet(B) was affected by subtherapeutic tetracycline administering, and the administration put a selective pression on it. The concerntration of tet(C) gene is lower than others in fecal samples,because of the susceptibility in E.coli. The selective pression on tet(M), tet(O) and tet(W) is present, so that their concerntration in fecal samples showed a higher level. sul2gene show a higher concerntration in fecal samples. And for erythrocin resistant gene, the erm(A) an erm(X) gene were affected by the treat. This study investigated the effects of administering beef cattle antimicrobials at subtherapeutic concentrations on the abundance and persistence of tetracycline resistance genes within the whole microbial community of fecal waste. Cattle were administered chlortetracycline, chlortetracycline plus sulfamethazine and no antimicrobials. Model fecal deposits (N=3) were prepared by mixing fresh feces from pens into a single composite sample. Real-time PCR was used to measure concentrations of tetracycline resistance genes tet(B), tet(C), tet(L), tef(M), tet(W) and16S-rRNA in DNA extracted from composite feces after7,14,28,42,56,70,84,98,112,126, and175days in the field. The concentrations of16S-rRNA in feces were similar across treatments and increased by day56(P<0.05). Generally, initial concentrations of tetracycline resistance genes were greater (P<0.05) in fecal pats from animals fed chlortetracycline. For all fecal treatments, tet(B) and tet(C) increased1～2log units by day56, and then decreased to the initial day7levels by day175(P>0.05). The concentrations of tet(M) and tet(W) were greater than other tetracycline resistance determinants. Tetracycline resistance genes can persist in fecal waste from cattle beyond175days and the initial load of some genes may underestimate concentrations at later time points. Temporal changes in the concentrations of resistance genes were likely due to shifts in microbial populations. |
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