In vitro antibiotic susceptibility of field isolates of Mycoplasma hyopneumoniae and Mycoplasma hyorhinis from Korea

Abstract

: The present study was conducted to determine the antibiotic susceptibilities of local Mycoplasma hyopneumoniae (Mhp) and Mycoplasma hyorhinis (Mhr) filed isolates. Minimum inhibitory concentrations (MICs) of Mhp and Mhr field isolates (twelve each) obtained from enzootic pneumonia-like lung lesions during 2009-2011 from Korea were determined using the broth microdilution method. Tylvalosin showed the highest activity against Mhp and Mhr field isolates, with MIC 90 values of 0.06 µg/mL and 0.12 µg/mL, respectively. Therefore, Korean Mhp and Mhr isolates are highly susceptible to tylvalosin.Keywords : Mycoplasma hyopneumoniae, Mycoplasma hyorhinis, antimicrobials, minimum inhibitory concentration, porcine enzootic pneumonia Swine mycoplasma Mycoplasma hyopneumoniae (Mhp) and Mycoplasma hyorhinis (Mhr) are respiratory pathogens in pigs. They are associated with enzootic pneumonia (EP) and porcine respiratory disease complex (PRDC) causing huge loses to the porcine industry. Currently commercial vaccines are available only against Mhp; however, their protection ability is not complete [7]. Therefore, use of antimicrobials often becomes necessary to limit the disease in the event of outbreak or as an additional measure to prevent mycoplasma disease. Although considerable data have been obtained on the in vitro antibiotic susceptibility of Mhp and Mhr around the world [6,[8][9][10][11], there are few reports about their susceptibility, and to our knowledge, there is no recent information on antibiotic susceptibility of these mycoplasmas from Korea. In the present study, minimum inhibitory concentrations (MICs) of Korea Mhp and Mhr isolates to tylvalosin and other commonly used antibiotics (tiamulin, lincomycin, tilmicosin and chlortetracycline) was determined.The reference strains used in this study were Mhp (ATCC 25934) and Mhr (ATCC 27717) (American Type Culture Collection, USA). Field isolates (twelve each) of Mhp and Mhr which were obtained from EP-like lung lesions during 2009-2011 were used in this study [1,2]. Each of these isolates was identified by multiplex polymerase chain reaction (mPCR) [3] and passaged 6 times before using in the antimicrobial assay. Both Mycoplasma species were propagated in Friis broth [4] and used at final concentration of approximately 1 × 10 5 CCU/mL. The antimicrobials chlortetracycline, tiamulin and tilmicosin were obtained from Sigma (USA). The lincomycin and tylvalosin were obtained from Dong Bang (Korea) and ECO Animal Health (UK), respectively. The antimicrobials were prepared by binary micro dilution method, in 96-well micro plate (ranging between 64 and 0.0038 µg/mL; SPL Life Sciences, Korea). MICs-were determined using the broth microdilution method as described by Hannan et al. [5]. Readings were taken after incubation of 7 days at 37 o C, and the lowest concentration of antimicrobial inhibiting color change from red to yellow/orange of the medium was defined as MIC of the drug.The results of the in vitro antimicrobial testing for mycoplasma field isolates and t...

Development of a molecular biological assay for the detection of markers related to decreased susceptibility to macrolides and lincomycin in Mycoplasma hyorhinis

“…However, a targeted antibiotic treatment is usually inaccessible. Elevated MICs of macrolides or lincomycin to M. hyorhinis isolates have been reported from Japan, Korea, Hungary, Spain, Italy and Portugal since 1996 (Kobayashi et al, 1996(Kobayashi et al, , 2005Jang et al, 2016;Bek} o et al, 2019;Rosales et al, 2020), and point mutations associated with elevated MIC values were described by Kobayashi et al (2005). In the present study, among Hungarian M. hyorhinis field isolates a transversion substitution in the previously examined region, at position 2066 (2058 E. coli numbering) of the 23S rRNA was found, an adenine was replaced by a guanine in the isolates with elevated MIC values.…”

“…Resistances to both of these groups are often linked because of the overlap of the binding sites at the 23S rRNA of the 50S subunit (Py€ or€ al€ a et al, 2014). Mycoplasma hyorhinis is intrinsically resistant to 14-membered macrolides (Ter Laak et al, 1991;Wu et al, 2000) and a decreased susceptibility to 15-and 16-membered macrolides has also been reported (Kobayashi et al, 1996(Kobayashi et al, , 2005Jang et al, 2016;Bek} o et al, 2019;Rosales et al, 2020). In vitro selected resistant mutants and Japanese field isolates have been investigated for resistance-related point mutations (SNPs) in the 23S rRNA gene and the L4, L22 protein genes of the 50S ribosome by Kobayashi et al (2005).…”

Evaluation of a Method for Standardized Antimicrobial Susceptibility Testing with Mycoplasma hyorhinis Field Isolates

“…Reduced efficacies toward several antimicrobial agents have been observed during the last decades, involving both human as well as veterinary Mycoplasma spp. [5,10,[14][15][16][17]20,[24][25][26]. In Mycoplasma spp., resistance development is mainly conferred by chromosomal mutations, such as the well-described point mutation within the 23S rRNA, conferring resistance to 14-membered macrolides [15,18,19,23,25,[27][28][29][30][31].…”

“…[5,10,[14][15][16][17]20,[24][25][26]. In Mycoplasma spp., resistance development is mainly conferred by chromosomal mutations, such as the well-described point mutation within the 23S rRNA, conferring resistance to 14-membered macrolides [15,18,19,23,25,[27][28][29][30][31].…”

“…The use of diverging AST methodologies generally leads to inconsistent data, rendering the obtained results incomparable and, when considering monitoring studies, unusable. For M. hyorhinis, the available literature indicates several methodologies, whereas most studies agree on using the broth microdilution method [10,[14][15][16]20,25]. On the other hand, a great variety of broth media is used for AST, wherefore the minimal inhibitory concentrations (MICs) of the tested isolates differ and are consequently incomparable or lead to misinformation, especially concerning the in vivo activity of antimicrobial agents against M. hyorhinis infections [9,[14][15][16]18,19,[32][33][34].…”