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• W2549555959 abstract "The present doctoral thesis aimed at investigating a multi-resistant Pasteurella multocida strain obtained from a case of bovine respiratory disease (BRD) in the U.S.A. for its genomic structure and the genetic basis of multi-resistance. Particular emphasis was put on the identification of novel genes that confer resistance to macrolides and triamilides as members from these classes are frequently used to combat BRD and the genetics of resistance to macrolides in BRD pathogens, including P. multocida, were largely unknown. For this doctoral thesis, the representative P. multocida strain 36950 was chosen. Since PCR-directed searches for known erm genes as well as repeated transformation attempts were unsuccessful, P. multocida 36950 was subjected to whole genome sequencing. Contigs obtained from the draft genome led to the identification of a novel rRNA methylase gene erm(42), the macrolide transporter gene msr(E), and the macrolide phosphotransferase gene mph(E). Functional cloning and expression of these genes in a macrolide susceptible P. multocida recipient strain confirmed that erm(42) was mainly responsible for resistance to tilmicosin and lincosamides such as clindamycin and only slightly increased the minimal inhibitory concentrations (MICs) of tulathormycin. In contrast, msr(E)-mph(E) were responsible for resistance to tilmicosin and tulathromycin, but had no effect on the MICs of lincosamides. The results of this study described for the first time the molecular basis of macrolide, triamilide, and lincosamide resistance in P. multocida [Chapter 2]. Further analysis of P. multocida 36950 genome and genomic comparisons revealed that these three genes were located on a mobile genetic element, an integrative and conjugative element (ICE), designated ICEPmu1. It was also the first report of an ICE in P. multocida [Chapter 3]. In addition to the three macrolide/triami-lide resistance genes, another nine antimicrobial resistance genes were found to be part of ICEPmu1. Eleven of the 12 resistance genes conferred resistance to streptomycin (strA and strB), streptomycin/spectinomycin (aadA25), gentamicin (aadB), kanamycin/neomycin (aphA1), tetracycline [tetR-tet(H)], chloramphenicol/ florfenicol (floR), sulphonamides (sul2), tilmicosin/clindamycin [erm(42)] or tilmicosin/tulathromycin [msr(E)-mph(E)]. In addition, a complete β-lactamase gene blaOXA-2 was detected, which, however, appeared to be functionally inactive in P. multocida. These resistance genes were organized in two regions of approximately 15.7 and 9.8 kb. Furthermore, resistance to nalidixic acid and enrofloxacin was due to point mutations within the quinolone-resistance determining region (QRDR) of the genes gyrA and parC [Chapter 3]. Such an expanded multi-resistance phenotype has very rarely been observed in P. multocida and other bovine respiratory tract pathogens. And the resistance genes and resistance-mediating mutations detected could fully explain this multi-resistance phenotype [Chapter 3]. The 82,214 bp ICEPmu1 harbours 88 genes. The core genes of ICEPmu1, which are involved in excision/integration and conjugative transfer, resemble those found in a 66,641 bp ICE from Histophilus somni. ICEPmu1 integrates into a tRNALeu and is flanked by 13 bp direct repeats. It is able to transfer by conjugation to P. multocida, M. haemolytica and E. coli, where it also uses a tRNALeu for integration and produces closely related 13 bp direct repeats at the integration site. The presence of ICEPmu1 and its circular intermediate in the transconjugands was confirmed by PCR and sequence analysis. PCR assays and susceptibility testing confirmed the presence and the functional activity of the ICEPmu1-associated resistance genes in the transconjugands. The gene blaOXA-2 proved to be inactive in P. multocida and M. haemolytica recipients, but was functionally active in the E. coli recipient strain [Chapter 4]. The novel macrolide and triamilide resistance genes were tested for their ability to confer resistance to gamithromycin and tildipirosin, two novel macrolides approved during the course of this doctoral thesis. Based on the observed increases in the MIC values in P. multocida B130 carrying the cloned erm(42) or msr(E)-mph(E), it appears as if erm(42) has mainly an effect on the tildipirosin MIC (128-fold increase) whereas msr(E)-mph(E) increases the gamithromycin MIC 256-fold in P. multocida B130. These observations were confirmed with P. multocida and M. haemolytica field isolates that carried the three genes in different combinations [CHAPTER 5]. ICEPmu1 proved to move across species and genus boundaries and since it carries 12 resistance genes, some of which confer resistance to the most recently approved antimicrobial agents for treatment of BRD, its dissemination drastically limits the treatment options. As such, the spread of ICEPmu1 is considered an emerging issue in antimicrobial resistance of food-producing animals [Chapter 6]." @default.
• W2549555959 created "2016-11-30" @default.
• W2549555959 creator A5050581449 @default.
• W2549555959 date "2015-01-01" @default.
• W2549555959 modified "2023-09-27" @default.
• W2549555959 title "Molecular analysis of a multi-resistant bovine Pasteurella multocida strain from the U.S.A." @default.
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