Increased MICs of gamithromycin and tildipirosin in the presence of the genes erm(42) and msr(E)-mph(E) for bovine Pasteurella multocida and Mannheimia haemolytica.

Sir, Bovine respiratory disease (BRD) is one of the economically most important diseases in animal production, with global losses of the feedlot industry due to BRD being estimated to be over US$ 3 billion per year. Antimicrobial agents, particularly macrolides, are commonly used to combat bacteria involved in BRD, such as Mannheimia haemolytica, Pasteurella multocida and Histophilus somni. After the approval of the 16-membered macrolide tilmicosin (Micotil) in 1992 and the 15-membered triamilide tulathromycin (Draxxin) in 2005 for use in BRD, two new macrolides were approved during 2011 for the treatment of BRD. These are the 15-membered macrolide gamithromycin (Zactran) and the 16-membered macrolide tildipirosin (Zuprevo). Little is known about the mechanisms of resistance to these two new veterinary antimicrobial agents. Recently, three genes involved in macrolide resistance of P. multocida and M. haemolytica were identified: erm(42), coding for a novel rRNA methylase; msr(E), coding for an ABC transporter; and mph(E), coding for a macrolide phosphotransferase. The last two genes were present in an operon structure. To determine the role of erm(42) and msr(E)-mph(E) in macrolide and lincosamide resistance, these genes were cloned and expressed in P. multocida B130. A closer analysis of these clones for their MICs of selected macrolides and lincosamides revealed that erm(42) conferred resistance to erythromycin, tilmicosin and clindamycin, but not to the triamilide tulathromycin. Although erm(42)-carrying P. multocida B130 showed an 8-fold increase in the tulathromycin MIC to 16 mg/L (Table S1, available as Supplementary data at JAC Online), this MIC classified the P. multocida isolate as susceptible. In contrast, msr(E)-mph(E) conferred resistance to erythromycin, tilmicosin and tulathromycin, but not to the lincosamide clindamycin (Table S1). When the new macrolides became commercially available, we tested the aforementioned P. multocida B130 clones for their MICs of gamithromycin and tildipirosin by broth macrodilution according to CLSI recommendations. The recipient strain P. multocida B130 showed 8-fold lower MICs of 0.25 mg/L for both gamithromycin and tildipirosin, compared with tulathromycin (2 mg/L) (Table S1). In the presence of erm(42), the MIC of tildipirosin increased 128-fold to 32 mg/L, while that of gamithromycin increased only 16-fold to 4 mg/L. In the presence of msr(E)-mph(E), an opposite observation was made: the MIC of tildipirosin increased only 8-fold to 2 mg/L, while that of gamithromycin increased 256-fold to 64 mg/L. Based on these increases in the MICs, it appears that erm(42) has mainly an effect on the tildipirosin MIC, whereas msr(E)-mph(E) increases the gamithromycin MIC for P. multocida B130. To see whether naturally occurring P. multocida and M. haemolytica isolates from BRD cases that carry the genes erm(42) and/or msr(E)-mph(E) show similar MICs of gamithromycin and tildipirosin, a total of 40 P. multocida and 29 M. haemolytica isolates (Table 1), whose macrolide resistance gene status was determined by previously described PCR assays, were tested. These isolates were collected in the Pfizer Animal Health Susceptibility Surveillance Program for BRD between 1999 and 2007 from various states in the USA. The macrolide-susceptible P. multocida isolates (n1⁄48) showed low MICs of 0.25–0.5 mg/L for both gamithromycin and tildipirosin. Slightly higher MICs of 0.5–1 and 0.5–2 mg/L for gamithromycin and tildipirosin, respectively, were detected for the macrolidesusceptible M. haemolytica isolates (n1⁄47). These MICs are in agreement with the gamithromycin MIC50 and MIC90 values for 144 P. multocida and 142 M. haemolytica isolates obtained from animals enrolled in field studies in the USA. Moreover, the observed tildipirosin MICs are in the same range as determined for 105 P. multocida and 88 M. haemolytica isolates. Research letters