Longitudinal characterization of antimicrobial resistance genes in feces shed from cattle fed different subtherapeutic antibiotics.

BACKGROUND: Environmental transmission of antimicrobial-resistant bacteria and resistance gene determinants originating from livestock is affected by their persistence in agricultural-related matrices. This study investigated the effects of administering subtherapeutic concentrations of antimicrobials to beef cattle on the abundance and persistence of resistance genes within the microbial community of fecal deposits. Cattle (three pens per treatment, 10 steers per pen) were administered chlortetracycline, chlortetracycline plus sulfamethazine, tylosin, or no antimicrobials (control). Model fecal deposits (n = 3) were prepared by mixing fresh feces from each pen into a single composite sample. Real-time PCR was used to measure concentrations of tet, sul and erm resistance genes in DNA extracted from composites over 175 days of environmental exposure in the field. The microbial communities were analyzed by quantification and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S-rRNA. RESULTS: The concentrations of 16S-rRNA in feces were similar across treatments and increased by day 56, declining thereafter. DGGE profiles of 16S-rRNA differed amongst treatments and with time, illustrating temporal shifts in microbial communities. All measured resistance gene determinants were quantifiable in feces after 175 days. Antimicrobial treatment differentially affected the abundance of certain resistance genes but generally not their persistence. In the first 56 days, concentrations of tet(B), tet(C), sul1, sul2, erm(A) tended to increase, and decline thereafter, whereas tet(M) and tet(W) gradually declined over 175 days. At day 7, the concentration of erm(X) was greatest in feces from cattle fed tylosin, compared to all other treatments. CONCLUSION: The abundance of genes coding for antimicrobial resistance in bovine feces can be affected by inclusion of antibiotics in the feed. Resistance genes can persist in feces from cattle beyond 175 days with concentrations of some genes increasing with time. Management practices that accelerate DNA degradation such as frequent land application or composting of manure may reduce the extent to which bovine feces serves as a reservoir of antimicrobial resistance.

Development of a Health-Related Quality of Life Questionnaire (HRQL) for patients with Extremity Soft Tissue Infections (ESTI).

BACKGROUND: Past clinical trials of antimicrobial treatment in soft tissue infections have focused on non-standardized clinical and physiological outcome variables, and have not considered the subjective experience of patients. The objective of this study was to develop a health-related quality of life questionnaire (HRQL) for patients with extremity soft tissue infections (ESTI) for future use in clinical trials. METHODS: The design of this study followed published guidelines and included item generation, item reduction, and questionnaire preparation. Study subjects were consenting English-speaking adults with acute ESTI requiring prescription of at least two days of outpatient intravenous antibiotic therapy. RESULTS: A list of 49 items that adversely impact the quality of life of patients with ESTI was generated by literature review, informal health professional feedback, and semi-structured interviews with twenty patients. A listing of these items was then administered to 95 patients to determine their relative importance on quality of life. A questionnaire was prepared that included the twenty most important items with a 5-point Likert scale response. Questionnaire domains included physical symptoms, problems performing their activities of daily living, impairment of their emotional functioning, and difficulties in their social interactions as related to their ESTI. The final questionnaire was pre-tested on a further ten patients and was named the ESTI-Score. CONCLUSION: The ESTI-Score is a novel instrument designed to quantify the impact of ESTI on quality of life. Future study is required to determine its validity and responsiveness before use as an outcome measure in clinical trials.

A rare case of pleuropulmonary infection and septic shock associated with Enterococcus faecium endocarditis.

Enterococci rarely cause pleuropulmonary infections. A novel case of definitively diagnosed nosocomial Enterococcus faecium pneumonia, empyema, and endocarditis associated with septic shock is reported.

Biofilm bacteria: formation and comparative susceptibility to antibiotics.

The Calgary Biofilm Device (CBD) was used to form bacterial biofilms of selected veterinary gram-negative and gram-positive pathogenic bacteria from cattle, sheep, pigs, chicken, and turkeys. The minimum inhibitory concentration (MIC) and minimum biofilm eradication concentration (MBEC) of ampicillin, ceftiofur, cloxacillin, oxytetracycline, penicillin G, streptomycin, tetracycline, enrofloxacin, erythromycin, gentamicin, tilmicosin, and trimethoprim-sulfadoxine for gram-positive and -negative bacteria were determined. Bacterial biofilms were readily formed on the CBD under selected conditions. The biofilms consisted of micro-colonies encased in extracellular polysaccharide material. Biofilms composed of Arcanobacterium (Actinomyces) pyogenes, Staphylococcus aureus, Staphylococcus hyicus, Streptococcus agalactiae, Corynebacterium renale, or Corynebacterium pseudotuberculosis were not killed by the antibiotics tested but as planktonic bacteria they were sensitive at low concentrations. Biofilm and planktonic Streptococcus dysgalactiae and Streptococcus suis were sensitive to penicillin, ceftiofur, cloxacillin, ampicillin, and oxytetracycline. Planktonic Escherichia coli were sensitive to enrofloxacin, gentamicin, oxytetracycline and trimethoprim/ sulfadoxine. Enrofloxacin and gentamicin were the most effective antibiotics against E. coli growing as a biofilm. Salmonella spp. and Pseudomonas aeruginosa isolates growing as planktonic populations were sensitive to enrofloxacin, gentamicin, ampicillin, oxytetracycline, and trimethoprim/sulfadoxine, but as a biofilm, these bacteria were only sensitive to enrofloxacin. Planktonic and biofilm Pasteurella multocida and Mannheimia haemolytica had similar antibiotic sensitivity profiles and were sensitive to most of the antibiotics tested. The CBD provides a valuable new technology that can be used to select antibiotics that are able to kill bacteria growing as biofilms.

High-throughput species identification of enterococci using pyrosequencing.

Here we report the development and validation of an automated high-throughput pyrosequencing-based method for the reliable identification of isolated Enterococcus species. This method exploits the discrete species-specificity of hypervariable groES-EL spacer region and utilizes a universal dispensation order optimized for a wide range of Enterococcus species.

Genetic diversity and tetracycline resistance genes of Histophilus somni.

Nasal swabs were collected at three time points from 2378 calves in four feedlots and cultured for Histophilus somni to assess genetic relatedness and tetracycline resistance. The proportions of animals carrying tetracycline resistant isolates were 0.32% at arrival, 14.82% at interim, and 0.80% at exit. The 606 H. somni isolates recovered were compared by pulsed-field gel electrophoresis (PFGE), screened for the presence of plasmids, and assessed for the tetracycline resistance genes tet(A), tet(B), tet(C), tet(E), tet(G), tet(H), tet(K), tet(L), tet(M) and tet(O) using multiplex polymerase chain reaction. Most of the isolates (98.6%) belonged to one of seven PFGE clusters (A-G) of closely related profiles with 77.7% of the isolates belonging to clusters C and D. Clusters A, B and E were associated with a higher proportion of tetracycline susceptible isolates. Genetic diversity of the isolates was highest at entry in the feedlot and lowest after the period when the animals received in-feed chlortetracycline (interim samples). Clusters A and E were more prominently represented at exit from the feedlot than other clusters. All resistant strains harboured the gene tet(H) while no other tetracycline resistance genes and no plasmids were detected with the methodology employed. It appears that genetic variability in H. somni in Alberta feedlots is low, dissemination likely occurs by clonal expansion, and resistance to tetracyclines is mediated by the tet(H) encoded efflux pump. Pulsotypes associated with tetracycline susceptible strains appear more common at exit suggesting that the in-feed oxytetracycline included throughout the feeding period is not sufficient to exert selective pressure for resistant strains.

Characteristics of individuals with male-to-male and heterosexually acquired infectious syphilis during an outbreak in Calgary, Alberta, Canada.

BACKGROUND: Eliminating syphilis is important not only to prevent the sequelae of infection but also to control the spread of HIV. Current prevention and control efforts in Canada have been ineffective in eliminating this disease. GOAL: The goal of the study was to determine the characteristics of individuals with infectious syphilis due to male-to-male and heterosexual contact, diagnosed during an outbreak in Calgary, Alberta, Canada. STUDY DESIGN: This was a prospective study of individuals with infectious syphilis diagnosed at the STD clinic in Calgary between January 2000 and April 2002. RESULTS: The outbreak reported here (September 2000 to April 2002) involves 32 cases of infectious syphilis, corresponding to rates of 0.9/100,000 population during 2000 and 1.8/100,000 population during 2001. Between September 2000 and June 2001, the cases diagnosed were among men who have sex with men (MSM); between May 2001 and April 2002, they were due to locally acquired infections among heterosexuals, including one case of congenital syphilis. Compared to the heterosexuals, MSM tended to be older, be coinfected with HIV, and report excessive alcohol use (versus injection drug use) and had infectious syphilis diagnosed earlier. MSM used the Internet and bars or bathhouses to initiate sexual contact, whereas heterosexually acquired infections were largely among sex workers and their clients. Contact tracing was more successful among the heterosexuals than among MSM. The public health staff at the STD clinic initiated a series of multifaceted interventions in response to the outbreak. These interventions were moderately successful, as measured by the increased numbers of individuals seeking counseling and testing services at the clinic. CONCLUSION: The results highlight key differences in the risk factor-specific characteristics of the outbreak that should be taken into account when designing prevention and control strategies.