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Methicillin-resistant Staphylococcus aureus (MRSA), in which acquisition of mecA [which encodes the cell wall peptidoglycan biosynthesis component penicillin-binding protein 2a (PBP2a)] confers resistance to β-lactam antibiotics, is of major clinical concern. We show that, in the presence of antibiotics, MRSA adopts an alternative mode of cell division and shows an altered peptidoglycan architecture at the division septum. PBP2a can replace the transpeptidase activity of the endogenous and essential PBP2 but not that of PBP1, which is responsible for the distinctive native septal peptidoglycan architecture. Successful division without PBP1 activity requires the alternative division mode and is enabled by several possible chromosomal potentiator ( pot ) mutations. MRSA resensitizing agents differentially interfere with the two codependent mechanisms required for high-level antibiotic resistance, which provides opportunities for new interventions.

The aim of this study was to estimate the prevalence of antimicrobial resistance (AMR) in Escherichia coli from a dog population in Spain and assess specific virulence factors. Susceptibility to 22 antimicrobials was tested along with the production of extended-spectrum β-lactamases (ESBLs) and AmpC in faecal isolates from 100 dogs. Virulence-related genes associated with attaching and effacing E. coli (eae, Stx1, Stx2) and extraintestinal pathogenic E. coli - ExPEC - (papC, hlyA and cnf1) were detected by PCR. At least one kind of AMR was observed in 73% of the isolates. The highest prevalences corresponded to penicillin (45%), aminoglycoside (40%) and non-extended spectrum cephalosporin (39%) classes. Multidrug resistance (MDR) was observed in 53.4% of the resistant isolates. No resistance to colistin was found. Production of ESBL/AmpC enzymes was detected in 5% of E. coli. Shiga toxin-producing E. coli were not observed, enteropathogenic E. coli were identified in only 12% of them, and ExPEC were found in 25%. Dog faeces can be a source of E. coli strains potentially presenting a threat to humans through their virulence factors or AMR. The non-hygienic keeping of animals may increase the risk of colonisation of such pathogens in humans.

We assessed the ceftazidime-avibactam disk diffusion breakpoints that provide the lowest discrepancy error rates by testing an Enterobacterales isolate collection with ceftazidime-avibactam MIC values near the breakpoints. Isolates ( n  = 112) were susceptibility tested by broth microdilution and disk diffusion methods in 3 laboratories. Current disk diffusion breakpoints (≥21/≤20 mm for susceptible/resistant) provided the lowest error rates, but confirmatory MIC testing is indicated for isolates with inhibition zones of 20 to 22 mm., (Copyright © 2020 American Society for Microbiology.)

Limited methods for colistin MIC determination are available to clinical microbiology laboratories. The purpose of this study was to evaluate the accuracy of the colistin broth disk elution (CBDE) test compared to that of broth microdilution (BMD) for identifying colistin MICs. CBDE was compared to colistin BMD using a collection of Gram-negative bacilli tested at two U.S. microbiology laboratories. The isolates tested included 121 retrospective clinical isolates, 45 prospective clinical isolates, and 6 mcr-1 -positive Escherichia coli isolates. CBDE was performed with four 10-ml cation-adjusted Mueller-Hinton broth tubes per isolate, to which 0, 1, 2, and 4 colistin 10-µg disks were added, generating final concentrations in the tubes of 0 (growth control), 1, 2, and 4 µg/ml, respectively. MICs were evaluated visually and interpreted using Clinical and Laboratory Standards Institute breakpoints. Site 2 also compared CBDE to the reference broth macrodilution (BMAD) method ( n = 110 isolates). Overall, CBDE yielded a categorical agreement (CA) and essential agreement (EA) of 98% and 99%, respectively, compared to the results of colistin BMD. Very major errors occurred for mcr-1 -producing strains, with MICs fluctuating from 2 to 4 µg/ml on repeat testing. The results for all other isolates were in CA with those of BMD. CBDE versus BMAD had an EA of 100% and a CA of 100%. Compared to currently used techniques, CBDE is an easy and practical method to perform colistin MIC testing. Some mcr-1 -producing isolates yielded MICs of 2 µg/ml by CBDE and 4 µg/ml by BMD. As such, the results for isolates with colistin MICs of 2 µg/ml by CBDE should be confirmed by the reference BMD method, and isolates with MICs of ≥2 µg/ml should be evaluated for the presence of mcr genes., (Copyright © 2019 American Society for Microbiology.)

Carbon monoxide-releasing molecules (CORMs) are a promising class of new antimicrobials, with multiple modes of action that are distinct from those of standard antibiotics. The relentless increase in antimicrobial resistance, exacerbated by a lack of new antibiotics, necessitates a better understanding of how such novel agents act and might be used synergistically with established antibiotics. This work aimed to understand the mechanism(s) underlying synergy between a manganese-based photoactivated carbon monoxide-releasing molecule (PhotoCORM), [Mn(CO)3(tpa-κ 3 N)]Br [tpa=tris(2-pyridylmethyl)amine], and various classes of antibiotics in their activities towards Escherichia coli EC958, a multi-drug-resistant uropathogen. The title compound acts synergistically with polymyxins [polymyxin B and colistin (polymyxin E)] by damaging the bacterial cytoplasmic membrane. [Mn(CO)3(tpa-κ 3 N)]Br also potentiates the action of doxycycline, resulting in reduced expression of tetA, which encodes a tetracycline efflux pump. We show that, like tetracyclines, the breakdown products of [Mn(CO)3(tpa-κ 3 N)]Br activation chelate iron and trigger an iron starvation response, which we propose to be a further basis for the synergies observed. Conversely, media supplemented with excess iron abrogated the inhibition of growth by doxycycline and the title compound. In conclusion, multiple factors contribute to the ability of this PhotoCORM to increase the efficacy of antibiotics in the polymyxin and tetracycline families. We propose that light-activated carbon monoxide release is not the sole basis of the antimicrobial activities of [Mn(CO)3(tpa-κ 3 N)]Br.

Aims: To determine the presence of Clostridium difficile on fattening pig farms in north-eastern Spain., Methods and Results: Twenty-seven farms were sampled. Pools of pig faecal samples (n = 210), samples of intestinal content from common farm pest species (n = 95) and environment-related samples (n = 93) were collected. Isolates were tested for toxin genes of C. difficile, and typed by PCR-ribotyping and toxinotyping. The minimal inhibitory concentrations of six antimicrobial agents were determined using Etest. Thirty-four isolates were obtained from 12 farms, and 30 (88·2%) had toxin genes. Seven ribotypes were identified. Ribotype 078 and its variant 126 were predominant (52·9%). The same ribotypes were isolated from different animal species on the same farm. None of the isolates were resistant to metronidazole or vancomycin., Conclusions: Clostridium difficile was common within the pig farm environment. Most of the positive samples came from pest species or were pest-related environmental samples., Significance and Impact of the Study: Pest species were colonized with toxigenic and antimicrobial-resistant C. difficile strains of the same ribotypes that are found in humans and pigs. Rodents and pigeons may transmit toxigenic and antimicrobial-resistant C. difficile strains that are of the same ribotypes as those occuring in humans., (© 2016 The Society for Applied Microbiology.)

Aims: We set out to investigate the antibacterial activity of a new Mn-based photoactivated carbon monoxide-releasing molecule (PhotoCORM, [Mn(CO)3(tpa-κ(3)N)](+)) against an antibiotic-resistant uropathogenic strain (EC958) of Escherichia coli., Results: Activated PhotoCORM inhibits growth and decreases viability of E. coli EC958, but non-illuminated carbon monoxide-releasing molecule (CORM) is without effect. NADH-supported respiration rates are significantly decreased by activated PhotoCORM, mimicking the effect of dissolved CO gas. CO from the PhotoCORM binds to intracellular targets, namely respiratory oxidases in strain EC958 and a bacterial globin heterologously expressed in strain K-12. However, unlike previously characterized CORMs, the PhotoCORM is not significantly accumulated in cells, as deduced from the cellular manganese content. Activated PhotoCORM reacts avidly with hydrogen peroxide producing hydroxyl radicals; the observed peroxide-enhanced toxicity of the PhotoCORM is ameliorated by thiourea. The PhotoCORM also potentiates the effect of the antibiotic, doxycycline., Innovation: The present work investigates for the first time the antimicrobial activity of a light-activated PhotoCORM against an antibiotic-resistant pathogen. A comprehensive study of the effects of the PhotoCORM and its derivative molecules upon illumination is performed and mechanisms of toxicity of the activated PhotoCORM are investigated., Conclusion: The PhotoCORM allows a site-specific and time-controlled release of CO in bacterial cultures and has the potential to provide much needed information on the generality of CORM activities in biology. Understanding the mechanism(s) of activated PhotoCORM toxicity will be key in exploring the potential of this and similar compounds as antimicrobial agents, perhaps in combinatorial therapies with other agents. Antioxid. Redox Signal. 24, 765-780.

The possibility of a "post-antibiotic era" in the 21st century, in which common infections may kill, has prompted research into radically new antimicrobials. CO-releasing molecules (CORMs), mostly metal carbonyl compounds, originally developed for therapeutic CO delivery in animals, are potent antimicrobial agents. Certain CORMs inhibit growth and respiration, reduce viability, and release CO to intracellular hemes, as predicted, but their actions are more complex, as revealed by transcriptomic datasets and modeling. Progress is hindered by difficulties in detecting CO release intracellularly, limited understanding of the biological chemistry of CO reactions with non-heme targets, and the cytotoxicity of some CORMs to mammalian cells., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Staphylococcus aureus is one of the most common causes of infection in people and is increasingly recognized in dogs. The increasing prevalence of methicillin resistant S. aureus (MRSA) is complicating the treatment of these infections. Panton Valentine leukocidin (PVL), a toxin involved in the pathogenesis of necrotic syndromes in people may be partially responsible for the rise of MRSA. Canine and human S. aureus from the same geographic area are genetically similar, indicating a common population and likely transmission. The implications of increasing antimicrobial resistance complicated by interspecies transmission, necessitates including both dogs and humans in S. aureus resistance surveillance studies. A collection of 126 S. aureus isolates from people (n = 99) and dogs (n = 27) were included, minimum inhibitor concentrations to a panel of 33 antimicrobials used in human and veterinary medicine were determined. No resistance to vancomycin, linezolid, daptomycin, quinupristin/dalfopristin or nitrofurantoin was found. A wide range of antibiograms were found; including resistance to 0-12 drugs (0-6 drug classes). Outstanding antibiograms included a canine MRSA resistant to rifampin and a human MRSA resistant to chloramphenicol. Inducible clindamycin resistance was found among 78% and 4% of canine and human MRSA and 17% and 25% of canine colonizing and human methicillin susceptible S. aureus (MSSA), respectively. Resistance to mupirocin was only found among human isolates including 20% of MRSA and 4% of MSSA. While no canine isolates were PVL positive, 39% of human MRSA and 2% of MSSA carried the gene. The bidirectional transmission of S. aureus between people and dogs necessitates the inclusion of isolates from both species in future studies., (© 2011 Blackwell Verlag GmbH.)

An observational study determined the normal fecal bacterial flora of clinically healthy bison, detected the presence of common potential zoonotic pathogens, and determined the antimicrobial susceptibility of isolated E. coli strains. Ninety-six fecal samples from 10 captive herds were cultured for aerobic, anaerobic, facultative, and microaerophillic bacteria. Nineteen major genera of gram-positive and 8 genera of gram-negative bacteria were identified. Salmonella spp. were not detected but some of the isolated bacteria are potential gastrointestinal pathogens. Minimum inhibitory concentrations (MIC) of 24 antimicrobials were determined for the E. coli isolated. Nearly all were susceptible to 23 of the 24 antimicrobials but there was a reduced susceptibility to sulphonamide. There were fewer resistant strains than were reported in recent studies of generic E. coli from cattle living in the same area.

This study characterized the antimicrobial susceptibility of 221 Staphylococcus aureus isolated from various species, and 60 canine Staphylococcus pseudintermedius isolated from 1986 through 2000 at the Western College of Veterinary Medicine (WCVM). Resistance of S. aureus was most common to penicillin (31%) and tetracycline (14%); resistance of S. pseudintermedius to penicillin was present in 8% and to tetracycline in 34% of isolates. Resistance to trimethoprim/sulfamethoxazole was only seen among S. pseudintermedius, and there was no resistance to amoxicillin/clavulanate, ampicillin/sulbactam, cephalothin, amikacin, gentamicin, enrofloxacin, chloramphenicol, or rifampin among any isolate. Inducible clindamycin resistance was found in both S. aureus and S. pseudintermedius, highlighting the need for careful interpretation of culture and susceptibility test results. There were significant differences in the minimum inhibitory concentrations of penicillin, ciprofloxacin, enrofloxacin, clindamycin, erythromycin, chloramphenicol, and tetracycline between avian, bovine, equine, and porcine isolates.

A randomized, controlled, blinded clinical trial was performed at a research feedlot in western Canada. Auction-market-derived steers (n = 288) were randomly assigned to 1 of 3 treatments: 1) no antimicrobials on arrival; 2) oxytetracycline in the starter ration for 14 d; and 3) long-acting oxytetracycline subcutaneously on day 0. Minimal inhibitory concentrations of 7 antimicrobials were determined for 3 generic fecal E. coli isolates per animal on arrival and throughout the feeding period. There was a low prevalence of antimicrobial resistance in generic E. coli isolates from calves on arrival. There were increased proportions of cattle with resistant E. coli isolates early in the feeding period among calves in groups 2 and 3. Individual animal treatments were not associated with increased proportions of cattle with resistant E. coli isolates preslaughter. There was no difference in the proportion of animals with E. coli isolates resistant to tetracycline between the treatment groups preslaughter. However, there were significantly more animals with tetracycline resistant isolates of E. coli preslaughter than at arrival.

Between January 2002 and June 2007, uropathogens were isolated from 473 of 1557 canine urine samples submitted to Prairie Diagnostic Services from the Western College of Veterinary Medicine Veterinary Teaching Hospital. Culture and susceptibility results were analyzed, retrospectively, to estimate the prevalence of common bacterial uropathogens in dogs with urinary tract infections and to identify changes in antimicrobial resistance. The most common pathogens identified were Escherichia coli, Staphylococcus intermedius, Enterococcus spp., and Proteus spp. Antimicrobial resistance increased during the study period, particularly among recurrent E. coli isolates. Using the formula to help select rational antimicrobial therapy (FRAT), bacterial isolates were most likely to be susceptible to gentamicin, fluoroquinolones, amoxicillin-clavulanic acid, and groups 4 and 5 (third generation) cephalosporins.

All bacterial samples of equine origin submitted to the diagnostic laboratory at the Western College of Veterinary Medicine from January 1998 to December 2003 from either "in-clinic" or Field Service cases were accessed (1323 submissions). The most common bacterial isolates from specific presenting signs were identified, along with their in vitro antimicrobial susceptibility patterns. The most common site from which significant bacterial isolates were recovered was the respiratory tract, followed by wounds. Streptococcus zooepidemicus was the most common isolate from most infections, followed by Escherichia coli. Antimicrobial resistance was not common in the isolates and acquired antimicrobial resistance to multiple drugs was rare. The results are compared with previous published studies from other institutions and used to suggest appropriate antimicrobial treatments for equine infections in western Canada.

A prospective observational study was carried out to examine antimicrobial resistance patterns of fecal Escherichia coli isolates of calves on arrival at the feedlot, and then evaluate the associations between the total volume of antimicrobial used for disease treatment and changes in antimicrobial resistance, during the feeding period. No macrolides or tetracyclines were administered in the feed during this study. On arrival, at the animal level, all 3 isolates obtained from 36.6% [95% confidence interval (CI): 29.0 to 44.8] of all cattle sampled (n = 153), were susceptible to all antimicrobials, while 5.9% (95% CI: 2.7 to 10.9) of cattle had at least 1 isolate that was resistant to--3 antimicrobials out of the 7 antimicrobials tested. The most frequent antimicrobials for which resistance was observed were sulphamethoxazole, ampicillin, and tetracycline where, of all cattle, 44.4% (95% CI: 36.4 to 52.7), 20.3% (95% CI: 14.2 to 27.5), and 17.7% (95% CI: 12.0 to 24.6), respectively had at least 1 resistant isolate. All cattle received antimicrobial metaphylaxis on arrival at the feedlot. Antimicrobial use was described for a cohort of 95 cattle. Antimicrobials were given to 42 of the 95 cattle during the feeding period, to treat disease. Amongst the 42 treated cattle, there were a total of 133 animal daily doses (ADD(Feedlot)), where 1 ADD(Feedlot) represented 1 day of antimicrobial treatment received by a feedlot animal at the approved dose. Only 1 ADD(Feedlot) was given in the 100 days immediately prior to slaughter. There were no associations found between antimicrobial use and antimicrobial resistance in this study.

Bacterial cultures from 32 living and dead farmed white-tailed deer (Odocoileus virginianus) with necrobacillosis yielded Fusobacterium necrophorum from nine individuals, F. varium from six individuals, and Arcanobacterium pyogenes from 16 individuals. The isolates were characterized biochemically using automated identification systems. Gram-stained smears suggested the presence of Fusobacterium spp. in eight cases from which organisms were not cultured. Minimum inhibitory concentration determinations in 23 strains of gram-negative anaerobic bacteria detected resistance to enrofloxacin and clindamycin. Enrofloxacin resistance was detected in A. pyogenes isolates, and although biochemical profiling indicated that the deer strains of A. pyogenes could be grouped, it is uncertain whether these biochemical characteristics correlate with antigenic or virulence factors. Deer-specific or autogenous vaccines may provide a useful alternative to generic vaccines.