Abstract
CTX-M-2-producing Klebsiella oxytoca (K. oxytoca) has not received much attention in animal husbandry compared with Klebsiella pneumoniae (K. pneumoniae), a major reservoir of extended-spectrum β-lactamase (ESBL) genes. Bacteriological examinations of 1,466 mastitic milk samples between October 2012 and December 2014 were conducted. Ninety-five K. pneumoniae isolates (total prevalence: 6.5%) and 81 K. oxytoca isolates (total prevalence: 5.5%) were obtained. Seventeen K. pneumoniae isolates obtained from 15 animals reared on 11 farms and 9 K. oxytoca isolates obtained from 9 animals reared on the same farm were phenotypically confirmed to be ESBL producers. All nine ESBL-producing K. oxytoca isolates were obtained from one farm between June and November 2013 and related to a significantly (p < 0.05) higher monthly prevalence of mild mastitis (in June, August, September, October, and November 2013). Pulsed-field gel electrophoresis (PFGE) patterns of ESBL-producing K. pneumoniae isolates were distinguished from each other by more than 6-band differences except for two isolates from two animals, whereas all nine K. oxytoca isolates showed an identical PFGE pattern. Transferability of the blaCTX−M−2 gene was found in 14 K. pneumoniae and 9 K. oxytoca isolates by conjugation analysis. Of these isolates, the blaCTX−M−2 gene was detected on plasmids belonging to the incompatibility (Inc) groups P and N derived from five K. pneumoniae and nine K. oxytoca isolates, respectively, although the plasmids from the remaining nine K. pneumoniae were untypeable. All the transconjugants exhibited elevated minimum inhibitory concentrations of ampicillin, cefotaxime, and ceftiofur compared with those in the wild-type, recipient strain. Restriction fragment length polymorphism analysis demonstrated that the IncN plasmids extracted from eight of nine transconjugants, which received resistance against β-lactams from K. oxytoca, showed an identical DraI digestion pattern. These results suggest that the CTX-M-2-producing K. oxytoca strain with the above-mentioned characteristics may have clonally spread within a farm, whereas the blaCTX−M−2 gene in K. pneumoniae possibly disseminated among the farms through different plasmids. Thus, monitoring of ESBL genes, including the blaCTX−M−2 gene, among causative agents of bacterial mastitis in cows can help to develop relevant treatments and control practices.
Highlights
Coliform bacteria are one of the major groups of pathogens associated with mastitis, with an estimated prevalence of 24.0–47.9%, 30.6–47.0%, and 15.0–43.0% in mild, moderate, and severe clinical mastitis, respectively [1,2,3]
K. oxytoca and K. pneumoniae were isolated in all seasons, with a higher prevalence recorded in the summer
Higher monthly incidences of mild mastitis in farm W were strongly correlated with the isolation of cephalosporin-resistant K. oxytoca from April to November 2013, despite there being no prevalence in May 2013
Summary
Coliform bacteria are one of the major groups of pathogens associated with mastitis, with an estimated prevalence of 24.0–47.9%, 30.6–47.0%, and 15.0–43.0% in mild, moderate, and severe clinical mastitis, respectively [1,2,3]. Intramammary infections of coliform bacteria are related to subclinical mastitis in approximately 60% of affected animals [4]. Of Klebsiella species, K. pneumoniae is frequently identified as a causative microbe with significant clinical impact because intramammary infections have a strong association with the occurrence of acute or peracute mastitis [5]. K. oxytoca is recognized as a minor coliform bacterium that is related to subclinical or occasionally mild mastitis, which is defined as the secretion of various forms of denatured milk but without any systemic signs [7]. K. oxytoca comprises 10.7% of gram-negative bacteria isolated from subclinical or clinical mastitis compared with 17.0% for K. pneumoniae [7]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
