Characterization of Mutations in DNA Gyrase and Topoisomerase IV in Field Strains and In Vitro Selected Quinolone-Resistant Mycoplasma hyorhinis Mutants

Abstract

Mycoplasma hyorhinis is ubiquitous in swine, and it is a common pathogen of swine that causes polyserositis, arthritis, and maybe pneumonia. Fluoroquinolones are effective antimicrobials used for the treatment of mycoplasmal infection. However, a decrease in fluoroquinolones susceptibility in mycoplasma was observed. The molecular mechanisms have been studied in many mycoplasma species, while the mechanism in M. hyorhinis is still unknown. This study aimed to illustrate the in vitro development of fluoroquinolone resistance in M. hyorhinis and unveil the resistance mechanisms in both in vitro selected mutants and field strains. Seven ciprofloxacin-sensitive M. hyorhinis isolates were chosen to induce the fluoroquinolone resistance in vitro, and the point mutations in the quinolone resistance-determining regions (QRDRs) were characterized. The substitutions first occurred in ParC, resulting in a 2- to 8-fold increase in resistance, followed by additional mutations in GyrA and/or ParE to achieve a 32-fold increase. The mutations occurred in hot spots of QRDRs, and they were diverse and variable, including five in ParC (Ser80Phe, Ser80Tyr, Phe80Tyr, Glu84Gly, and Glu84Lys), four in GyrA (Ala83Val, Ser84Pro, Asp87Tyr, and Asp87Asn) and one in ParE (Glu470Lys). Target mutations in field strains were observed in the ParC (Ser80Phe, Ser81Pro, and Glu84Gln) of isolates with MICCIP = 2 μg/mL. This study characterized the point mutations in the QRDRs of M. hyorhinis and could be useful for the rapid detection of fluoroquinolone resistance in M. hyorhinis field isolates.

Aeromonas hydrophilaCobQ is a new type of NAD+- and Zn2+- independent protein lysine deacetylase

“…For example, the bacterial topoisomerases DNA gyrase GyrB is an essential enzyme that controls the topological state of DNA during replication (Tari et al, 2013). Previous studies reported the important role of GyrB in bacterial fluoroquinolone resistance (Chauffour et al, 2021; Li et al, 2022). The 3D structure prediction showed that GyrB K331 is located in the ATPase domain that is responsible for supercoiling DNA, whereas K449 is located in the Toprim domain that is required for the interaction with GyrA (Brino et al, 2000).…”

Whole-genome sequencing of Histophilus somni strains isolated in Russia

“…According to the literature, substitutions at positions 80 and 84 in the topoisomerase IV ( parC ) gene were found to be significant for resistance to fluoroquinolones [ 25 ]. These codon substitutions were not detected in all our sequences that have been analyzed.…”

Towards a Standardized Antimicrobial Susceptibility Testing Method for Mycoplasma hyorhinis

“…Since mycoplasmas are generally very demanding concerning their culture conditions, it is necessary to accept certain difficulties, such as expensive or very complex broth media as well as the extended growth periods, also for AST methods. Several very different agar and broth media have been used over time for AST of porcine mycoplasmas: Friis media [6,8,9,25], modified Friis media [15,25,37], the commercial Liquid Mycoplasma Medium (Mycoplasma Experience Ltd., UK) [19,20,38], variations of YUS and CMRL media [30], M media [14,36,39] and KM2 media [40]. According to the published literature, for M. hyorhinis and M. hyopneumoniae, the (modified) Friis broth appears to be the most reliable culture medium so far.…”

“…Although the AST methodology previously used was different between working groups, most still agree on using the type strain of M. hyorhinis as a reference and for quality control purposes without generally valid quality control ranges [8,9,[14][15][16]19,25,36,37,40]. Due to the variety of broth media used in broth microdilution methods, the resulting MIC values might differ as well.…”

“…Furthermore, the incubation times differ between studies. Most agree on evaluating the microtiter plates on a daily basis until a color change occurs, whereas others read the plates at more or less fixed time points from up to two to seven days of incubation [8,9,15,25,36,40].…”