Antimicrobial Susceptibilities of Four Species of Mycoplasma Isolated in 2008 and 2009 from Cattle in Japan

Uemura, Ryoko; Sueyoshi, Masuo; Nagatomo, Hiroshi

doi:10.1292/jvms.10-0165

Cited by 29 publications

(41 citation statements)

References 11 publications

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“…Three studies reported highly resistant isolates, with MICs of Ͼ8 g/ml, in the United Kingdom, Japan, and Belgium (9-11), while several others indicated a low to no increase in resistance, except for a few isolates, in Japan, the United States, and several European countries (6,(12)(13)(14)(15). Some of these studies involved isolates collected before 2005 (11, 13), but the majority referred to recent ones (6,9,10,12,14,15), suggesting a recent phenomenon. In the United Kingdom, a shift of resistance from an MIC 90 of 1 g/ml to one of 32 g/ml occurred during the last decade (9, 16).…”

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confidence: 99%

Alterations in the Quinolone Resistance-Determining Regions and Fluoroquinolone Resistance in Clinical Isolates and Laboratory-Derived Mutants of Mycoplasma bovis: Not All Genotypes May Be Equal

Khalil

Becker

Tardy

2016

Appl Environ Microbiol

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Mycoplasma bovis is considered a major contributor to respiratory diseases in young cattle. Resistant M. bovis isolates have increasingly been reported worldwide due to extensive use of antimicrobials to treat bovine pneumonia. The frequency of isolates resistant to fluoroquinolones varies considerably from one country to another. The MICs of isolates collected in France have only increased from "very low" to "low." The present study was conducted to investigate whether alterations in the quinolone resistance-determining regions (QRDRs) could account for this slight modification in susceptibility. No correlation between QRDR alterations and increased MICs was evidenced in clinical isolates. In addition, all clinical isolates were subtyped, and the tendencies of the different sequence types to develop resistance through mutations in QRDRs under selective pressure in vitro were examined. In vitro, 3 hot spots for mutations in QRDRs (position 83 in GyrA and positions 80 and 84 in ParC) were associated with a high level of resistance when cumulated. We showed that the point mutations in the QRDRs observed in vitro were different (in location and selection rapidity) between the different subtypes. Our in vitro observations were corroborated by the recent detection of a clinical isolate highly resistant to fluoroquinolones (MIC > 16 g/ml) and belonging to the subtype which easily accumulates QRDR alterations in vitro. The current increased prevalence of this subtype in clinical isolates highlights the urgent need to control fluoroquinolone usage in veterinary medicine. Mycoplasma bovis is a wall-less bacterium responsible for severe infections in cattle, including pneumonia, mastitis, arthritis, and otitis (1). In young cattle, it is now recognized as a major contributor to economic losses associated with bovine respiratory diseases (BRD). The infection pressure of BRD usually peaks 2 to 3 weeks after calves are mingled in fattening units following transportation from their respective birth farms (2). Since no efficient vaccines are available and licensed for use outside the United States (3), efforts to control M. bovis infections often rely on antimicrobial treatments, administered either prophylactically or in the early stages of the disease. Antimicrobials used for the treatment or prevention of BRD usually include broad-spectrum cephalosporins (cefquinome and ceftiofur), extended-spectrum fluoroquinolones (enrofloxacin, danofloxacin, and marbofloxacin), florfenicol, and long-lasting macrolides (tulathromycin, gamithromycin, and tildipirosin) (4). This extensive use of antibiotics has predictably resulted in an increase in resistant isolates over time. In France, it was recently shown that contemporary M. bovis strains had become significantly less susceptible than archival strains to 9 of the 12 antimicrobials tested (5). With regard to fluoroquinolones, the decrease in susceptibility was limited (only 1 dilution of the MIC), and no highly resistant isolates were observed in a set of more than 90 strains. This wa...

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Alterations in the Quinolone Resistance-Determining Regions and Fluoroquinolone Resistance in Clinical Isolates and Laboratory-Derived Mutants of Mycoplasma bovis: Not All Genotypes May Be Equal

Khalil

Becker

Tardy

2016

Appl Environ Microbiol

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“…of veterinary importance (reviewed in reference 8), including M. bovis (9)(10)(11)(12)(13)(14)(15), the genetic background for decreased susceptibility has not been elucidated in either field isolates or mutants selected in vitro. M. bovis contains one or two rRNA operons (rrn).…”

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16S rRNA Gene Mutations Associated with Decreased Susceptibility to Tetracycline in Mycoplasma bovis

Amram

Mikula²,

Schnee

et al. 2015

Antimicrob Agents Chemother

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dMycoplasma bovis isolates with decreased susceptibilities to tetracyclines are increasingly reported worldwide. The acquired molecular mechanisms associated with this phenomenon were investigated in 70 clinical isolates of M. bovis. Sequence analysis of the two 16S rRNA-encoding genes (rrs3 and rrs4 alleles) containing the primary binding pocket for tetracycline (Tet-1 site) was performed on isolates with tetracycline hydrochloride MICs of 0.125 to 16 g/ml. Mutations at positions A965T, A967T/C (Escherichia coli numbering) of helix 31, U1199C of helix 34, and G1058A/C were identified. Decreased susceptibilities to tetracycline (MICs, >2 g/ml) were associated with mutations present at two (A965 and A967) or three positions (A965, A967, and G1058) of the two rrs alleles. No tet(M), tet(O), or tet(L) determinants were found in the genome of any of the 70 M. bovis isolates. The data presented correlate (P < 0.0001) the mutations identified in the Tet-1 site of clinical isolates of M. bovis with decreased susceptibility to tetracycline. The bacterial pathogen Mycoplasma bovis causes a variety of clinical manifestations in cattle, including respiratory disease, mastitis, arthritis, and otitis, which result in substantial economic losses (1). The tetracyclines are among the few important antimicrobial agents that may be used to treat M. bovis infections (2).Tetracyclines are broad-spectrum antimicrobials that have been widely used in human and veterinary medicine (3). They inhibit protein synthesis by binding to the 30S ribosomal subunit and blocking an attachment of aminoacyl-tRNA to the A site (3). Resistance to tetracyclines is common in many bacterial species and may be achieved by (i) an energy-dependent efflux of the drug across the cell membrane mediated by efflux pumps, (ii) the presence of ribosomal protection proteins that confer tetracycline resistance, either by a reduction of the affinity of ribosomes to tetracyclines or by releasing the bound antimicrobial from the ribosome, (iii) the enzymatic inactivation of the drug, or (iv) the mutations in the 16S rRNA genes that affect the binding sites of tetracyclines (4).In Mollicutes, two mechanisms of resistance to tetracyclines have been identified so far, both of which are in Mollicutes species that infect humans. These include ribosomal protection by tet(M) determinants, described in naturally tetracycline-resistant strains of Mycoplasma hominis and Ureaplasma spp. (5, 6), as well as target modification with point mutation(s) in the 16S The aim of this study was to investigate the mechanisms associated with acquired decreased susceptibilities to tetracyclines in M. bovis isolates. ; 1980 [19]) were tested in this study, and their details are given in Table 1. Each isolate originated from different farms, was selected at random, and had no epidemiological link to other isolates unless indicated otherwise. The reference type strain M. bovis PG45 was obtained from the National Collection of Type Cultures, United Kingdom (strain NCTC10131, corresponding to ATC...

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“…Fluoroquinolones are broad-spectrum antimicrobials, and enrofloxacin (ENR), orbifloxacin (ORB) and danofloxacin (DAN) have been approved for use in cattle in Japan. Fluoroquinolone minimum inhibitory concentrations (MICs) against Mb have previously been investigated [6,8]. Mb isolated from cattle in Japan showed various ENR MICs (range≤0.125-4 µg/ml) [8].…”

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“…Fluoroquinolone minimum inhibitory concentrations (MICs) against Mb have previously been investigated [6,8]. Mb isolated from cattle in Japan showed various ENR MICs (range≤0.125-4 µg/ml) [8]. Ten of eleven ENR-resistant (ENR MIC≥2 µg/ml) Mb isolates derived from cattle in Israel carried 2 missense mutations within quinolone resistance-determining regions (QRDRs), causing the amino acid substitutions (AASs) Ser83Phe in GyrA and Asp84Asn in ParC [6].…”

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Amino Acid Substitutions in GyrA and ParC are Associated with Fluoroquinolone Resistance in <i>Mycoplasma bovis</i> Isolates from Japanese Dairy Calves

Sato

Okubo

Usui

et al. 2013

The Journal of Veterinary Medical Science

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We investigated the contribution of quinolone resistance-determining region (QRDR) mutations to fluoroquinolone (enrofloxa-cin, orbifloxacin and danofloxacin) susceptibility in 58 Mycoplasma bovis isolates from dairy cattle in Japan. Fluoroquinolone non-resistant isolates (fluoroquinolone-MICs≤2 µg/ml) possessed no QRDR mutations (19 isolates) or Ser83Leu in GyrA (32 isolates). Fluoroquinolone-resistant isolates (fluoroquinolone-MICs≥4 µg/ml) possessed Ser83Leu in GyrA and Ser81Pro in ParC (3 isolates) or Ser83Phe in GyrA and Ser80Ile in ParC (4 isolates). Laboratory-derived fluoroquinolone-resistant mutants selected from 2 isolates (possessing Ser83Leu in GyrA) had an amino acid substitution in ParC at the same position (Ser80Ile or Ser81Tyr) as fluoroquinolone-resistant isolates, suggesting a concurrent amino acid substitution in ParC (Ser80 or Ser81) is important in fluoroquinolone resistance in M. bovis isolates. Mycoplasma bovis (Mb), a causative agent of bovine respiratory disease, causes high morbidity, and has an economic impact on the cattle industry [3]. Antimicrobials, such as fluoroquinolones, macrolides and tetracyclines, are important for treating Mb infections and preventing its spread in dairy calves. Fluoroquinolones are broad-spectrum anti-microbials, and enrofloxacin (ENR), orbifloxacin (ORB) and danofloxacin (DAN) have been approved for use in cattle in Japan. Fluoroquinolone minimum inhibitory concentrations (MICs) against Mb have previously been investigated [6, 8]. Mb isolated from cattle in Japan showed various ENR MICs (range≤0.125-4 µg/ml) [8]. Ten of eleven ENR-resistant (ENR MIC≥2 µg/ml) Mb isolates derived from cattle in Israel carried 2 missense mutations within quinolone resistance-determining regions (QRDRs), causing the amino acid substitutions (AASs) Ser83Phe in GyrA and Asp84Asn in ParC [6]. Although studies have suggested that QRDR mutations contribute to elevated ENR MICs, the association between QRDR mutations and fluoroquinolone MICs has yet to be confirmed in Mb [6]. We investigated QRDR mutations in Mb isolates and in vitro fluoroquinolone-selected mutants obtained from Mb isolates with various ENR, ORB and DAN MICs to clarify the effect of AAS in GyrA and ParC on fluoroquinolone resistance in Mb isolates from dairy calves in Japan. Fifty-eight Mb isolates were derived from nasal swabs of asymptomatic dairy calves from 51 farms located throughout Hokkaido during 2010-2011. These isolates were provided by 7 branches of the Hokkaido Agricultural Insurance Scheme (Dounan, Iburi-Hidaka, Ishikari-Sorachi, Kitami-Ohotsuku, Kushiro-Nemuro, Souya-Rumoi-Kamikawa and Tokachi) and were cultured in 1 ml NK broth (Kanto Chemical , Tokyo, Japan). After culture on modified Hayflick agar (35.5 g/l of PPLO, 15% equine serum, 2.5% yeast extract and 0.0024% deoxyribonucleic acid; Kanto Chemical) at 37°C in 5% CO 2 for 3 days to 1 week, a single colony was picked, inoculated into 1 ml of NK broth and further cultured at 37°C in 5% CO 2 for 3 days. ENR, ORB and DAN MICs were determined using ...

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Antimicrobial Susceptibilities of Four Species of Mycoplasma Isolated in 2008 and 2009 from Cattle in Japan

Cited by 29 publications

References 11 publications

Alterations in the Quinolone Resistance-Determining Regions and Fluoroquinolone Resistance in Clinical Isolates and Laboratory-Derived Mutants of Mycoplasma bovis: Not All Genotypes May Be Equal

Alterations in the Quinolone Resistance-Determining Regions and Fluoroquinolone Resistance in Clinical Isolates and Laboratory-Derived Mutants of Mycoplasma bovis: Not All Genotypes May Be Equal

16S rRNA Gene Mutations Associated with Decreased Susceptibility to Tetracycline in Mycoplasma bovis

Amino Acid Substitutions in GyrA and ParC are Associated with Fluoroquinolone Resistance in <i>Mycoplasma bovis</i> Isolates from Japanese Dairy Calves

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