Allele-specific real-time PCR testing for minor macrolide-resistant Mycoplasma Pneumoniae

Abstract

Background The point mutations in 23S rRNA gene of Mycoplasma pneumoniae ( M. pneumoniae ) can lead to high-level resistance to macrolides. This study aimed to evaluate allele-specific real-time PCR (ASPCR) to detect the resistance-related mutations located at positions A2063G and A2064G of 23S rRNA gene. Methods We detected 178 pharyngeal swab specimens and calculated the proportions of resistant and sensitive quasispecies using ASPCR assays. ASPCR assays can detect down to 10 copies of 23S rRNA gene and achieved sensitivities of < 0.1% for A2063G and A2064G. We also compared the findings of ASPCR with the results of nested PCR with sequencing. Results Of 178 samples, 164 were found to have M. pneumoniae including 90.85% (149/164) samples with macrolide-resistant M. pneumoniae (MRMP) quasispecies by ASPCR, while 153 were found to be M. pneumoniae -positive including 71.90% (110/153) samples with MRMP quasispecies by nested PCR with sequencing. Of the 164 M. pneumoniae- positive samples, 61.59% (101/164) had the mixed population of wild-type and mutant M. pneumoniae , and 56.44% (57/101) of the latter contained the mutations at low frequency (≤50%). Conclusion ASPCR indicated that sensitive and resistant quasispecies coexisted in most of the M. pneumoniae positive samples. The ASPCR was a highly sensitive, accurate and rapid method for detecting the macrolide resistance-associated mutations and it could provide earlier and more drug-resistant information for M. pneumoniae research and the clinical therapy.

Impact of viral coinfection and macrolide-resistant mycoplasma infection in children with refractory Mycoplasma pneumoniae pneumonia

“…The mutation rate in this study was 56.07%, of which were all A2063G mutations. The rate of drug-resistant MP was lower than those reported in other studies in China, where the rates range from almost 70-90% [20][21][22] . This is the because point mutations in drug-resistance genes were detected by PCR in this study, while drug sensitivity tests were chosen by other researchers.…”

“…1). ADV infection may cause severe disease necessitating ICU admission and mechanical ventilation [20] .…”

Impact of viral coinfection and macrolide-resistant mycoplasma infection in children with refractory Mycoplasma pneumoniae pneumonia

“…The mutation rate in this study was 56.07%, of which were all A2063G mutations. The rate of drug-resistant MP was lower than those reported in other studies in China, where the rates range from almost 70 to 90% [21,22,24]. This is the because point mutations in drug-resistance genes were detected by PCR in this study, while drug sensitivity tests were chosen by other researchers.…”

“…However, in this study, no remarkable differences in clinical characteristics were observed between patients who were and were not coinfected, except for in those with ADV coinfections. ADV infection may cause severe disease necessitating ICU admission and mechanical ventilation [21]. The severity of pneumonia with ADV coinfection is significantly related to viral load and serotypes, and children with ADV genotype 7 develop severe pneumonia more frequently than those with other genotypes [22].…”

Impact of viral coinfection and macrolide-resistant mycoplasma infection in children with refractory Mycoplasma pneumoniae pneumonia

“…ADV infection may cause severe disease necessitating ICU admission and mechanical ventilation [21] .…”