Chloromed

The article “Effects of in-feed chlortetracyclineprophylaxis in beef cattle on animal health and antimicrobial resistant Escherichia coli ” was written by Agga et al. in 2016 and was published in the journal “Applied and Environmental Microbiology”.

The study was conducted to provide further understanding on the theory that exposing healthy animals to in-feed chlortetracycline (CTC) may increase the spread of antimicrobial resistance (AMR). This study evaluated the effect of a one-time 5-day in-feed CTC treatment to USA feedlot calves(morbidity and body weight gain), occurrence of tetracycline-resistant (TET E. coli , and occurrence of 3 generation cephalosporin-resistant (3GC E. coli over a 4-month follow-up period.

Three hundred weaned calves less than six months old of both sexes were randomly allocated to a CTC or control group. The calves in the CTCgroup were medicated with 10 mg chlortetracycline per lb (equivalent to 22.05mg/kg) for 5 days, starting on day 5 post arrival. The animals were housed in pens of 30 animals per pen. The groups were separated by empty pens, and three empty pens were also included in the study. The detailed study was performed by the United States Department of Agriculture (USDA) with a declaration of no competing interests.

Faecal swabs, pen surface material, feed and water samples were collected on five occasions: at arrival, and on days 5, 27, 75 and 117post treatment. On each sampling occasion, faecal swabs were collected from the rectum, four samples per pen were collected from all pens including the three empty pens, one water trough sample from each trough and one fresh feed sample were collected.

Generic E. coli were isolated from CHROMagar E. coli (CEC) media that was supplemented with 2 mg/l cefotaxime to detect 3GC E. coli and with 32 mg/l tetracycline to detect TET E. coli . Colonies were enumerated with an automated colony counter. After secondary enrichment, cultures were plated in media as described above to detect generic, 3GC and TET E. coli and presumptive colonies were subsequently used to confirm presumptive E.coli by PCR.

The publication used a clinical breakpoint of 32 mg/l to detect resistant strains, and the authors suggest that a lower value may provide false positives, but it is possible that a 32 mg/l breakpoint may have underestimated the number of bacteria that might be considered to be resistant.

Bodyweight measures were assessed using a multilevel mixed effects linear regression. Morbidity was assessed using survival analysis with cox proportional hazards regression model. Pen-level generic, TET and 3GC E. coli mean and95% confidence intervals were determined using a multilevel mixed-effects linear model. Mean prevalence (%) was obtained using a multilevel mixed effects logistic regression model. Multiple comparisons were adjusted by Bonferroni. The experimental unit was the pen. P values less than 5% or 95% confidence intervals were used to make inferences.

Zero values for colony enumeration were replaced by the midpoint between the limit of detection and the limit of enumeration. For samples below the limit of detection the zero values were assigned a -1 log less than the limit of detection.

Groups were balanced at the start of the study for age, weight and prior antimicrobial treatments although there were more females in the CTC group (p

Faecal swabs: Only on day 5 post treatment were generic and TET E. coli significantly higher (P E.coli was higher in the control group(p=0.01). In the CTC group, on day 5 post treatment generic E. coli concentration did not differ from the arrival concentration but TET E. coli concentration was higher than at arrival (P E. coli because only 0.1% of the samples presented values above the limit of enumeration.Prevalence of 3GC E. coli did not differ between the CTC and control group throughout the study and increased from less than 10% to more than 70% in both groups on the last two sampling occasions (P

Pen surface samples: On day 5 post treatment, TET E. coli was significantly higher(P E. coli were higher than their arrival concentration in both groups (p E. coli concentrations were higher than the arrival concentration only in the CTC group (P E.coli concentrations were not significantly different, but were higher than at earlier time points (p=0.01).As for faecal swabs, only prevalence (%) was analysed for 3GC E. coli and prevalence did not differ significantly between the CTC and control group throughout the study.

At arrival, generic and TET E. coli concentrations in the empty pens did not differ from control or CTC pens, and in the subsequent sampling time points empty pens did not differ from their arrival concentration. From day 27 post treatment onwards empty pen concentrations were lower (P E. coli was significantly lower in the empty pens compared to both other groups.

Feed and water samples: Prevalence in feed of generic and TET E.coli concentrations was 100% at all time points, concentrations were higher on days 75 and 117 post treatment in comparison to earlier time points. 3GC E. coli was not detected in feed samples until the last sampling time point when it was found in 100% of the feed samples below the limit of enumeration. In water, 3GC E.coli was detected in 50% of the samples in the CTC group on days 75 and 117post treatment and in 100% of the samples in the control group on day 117 posttreatment.

The choice of sampling time points was not explained in the publication. Based on the peak observed 5 days post treatment an additional sampling time-point before 27 days post treatment could have provided some additional information regarding treatment related effects, on TET E. coli isolates. Similarly, an additional sampling time point between day 27 and 75 post treatment may have provided further information on the development of 3GC E. coli prevalence. The pre-study selection of sampling times is often difficult, and further research would be needed to investigate these issues.

Pen samples wereassessed in this study to address the concerns about the potential environmental impact of in-feed treatment with CTC. Pen samples showed as for faecal samples,that on day 5 post treatment TET E. coli was higher in the CTC group but thereafter differences were not significant between groups, but concentrations were significantly higher than at arrival. All three pens showed a proportional concentration between TET and generic E. coli

It is also noted that the faecal sample concentrations (generic and TET E. coli ) and prevalence (3GC E. coli ) were higher on days 75 and117 post treatment than at earlier time points, but this increase was not discussed in detail and the investigators suggest that it could be attributed to other factors, including ambient temperature and season.

The lack of significant differences between groups was discussed and it was suggested that other factors had greater impact than in-feed CTC for faecal TET and 3GC E. coli prevalence The investigators suggest that deposited E.coli populations or increased nutrients from deposited manure or a combination of these were the principal factors contributing to the increase in resistant E. coli prevalence The investigators suggest this is supported by the published literature stating that antimicrobial resistant bacteria and antimicrobial resistant genes increased in soils from pens holding untreated cattle or pens that were fertilised with manure from cattle that had not received antibiotics.

The reason why resistant E. coli increased as the study progressed in both groups was attributed to the known presence of E.coli in feed which also increased as the study progressed, and presence in water was attributed to contamination by E.coli from the calves’ heads and oral cavities.

The publication noted that the ration fed contained antibiotics tylosin and monensin on occasions, but it was stated that these antimicrobials had no impact on the results due to the intrinsic resistance of E. coli to them.

The results showed that CTC did not influence body weight but significantly reduced morbidity and therapeutic antimicrobial use in the treated group. Morbidities were more likely in the control group and animals had to be treated with antimicrobials that are considered highest priority critical antimicrobials by the World Health Organisation (WHO).

The investigators concluded that in-feed treatment with CTC temporarily increased TET E. coli in faecal samples and pen samples but did not impact on faecal or pen prevalence of 3GC E. coli . Feed and water samples showed increases in generic, TET and 3GC resistant E. coli regardless of CTC treatment in the second half of the study. The investigators concluded that other undefined factors contributed more significantly than CTC to the prevalence of TET and 3GC E. coli , and that confident attribution of antimicrobial-resistant E. coli occurrences to specific factors requires additional study.

These conclusions are also supported by Alexander et al (2008)who found that the proportion of feedlot steers- harbouring tetracycline resistant E. coli strains exceeded 40% despite their having had no previous exposure to tetracycline, even before antimicrobial agents were included in the diet. Alexander etal demonstrated that tetracycline (- and ampicillin) -resistant E. coli were harboured by a large number of cattle shortly after arrival at the feedlot, independent of any direct exposure to antibiotics and that presumably, the majority of these resistant E. coli would have been acquired from the environment in which the calves were raised, either from their dams or other environmental sources. Survival of E. coli has been shown to occur up to 150 days in faecal pats on range land, and stored feed has also been implicated as a source of antimicrobial-resistant E. coli at feedlots.

The authors summarise their findings that CTC administered for 5 days in-feed to USAfeedlot cattle less than 6 months old: reduced morbidity; reduced the therapeutic use of highest priority critically important antimicrobials; temporarily increased faecal TET E. coli concentration; temporarily increased pen surface TET E. coli concentration; did not impact long-term faecal TET E. coli concentration; did not impact faecal 3GC E. coli prevalence; and did not impact pen surface 3GC E. coli prevalence.

Regardless of in-feed CTC exposure, generic, TET and 3GC E. coli occurrences were highest during the last two sampling occasions, 75 and 117days after in-feed CTC administration ceased. Consequently, the paper concludes that other undefined factors contributed more significantly to TET and 3GC E. coli occurrence than the use of a 5-day course of in-feed CTC.

Alexander, T W et al . “Effect of subtherapeutic administration of antibiotics on the prevalence of antibiotic-resistant Escherichiacoli bacteria in feedlot cattle.” Applied and environmental microbiology vol. 74,14 (2008): 4405-16. doi:10.1128/AEM.00489-08

Important Review Article on the Treatment of Respiratory Diseases in Calves Without the use of Critically Important Antibiotics.