Macrolide-resistant Mycoplasma Research Articles

Comparison of three real-time PCR methods for detection of macrolide-resistant Mycoplasma genitalium in Sweden

There is a worldwide increase in macrolide-resistant Mycoplasma genitalium strains, with severe impacts on treatment. The aim of this study was to compare three real-time PCR methods for the detection of macrolide resistance: an in-house PCR described by Touati et al., ResistancePlus® MG (SpeeDx), and S-DiaMGRes™ (Diagenode Diagnostics). One hundred M. genitalium-positive patient samples collected in Sweden and a quantitated M. genitalium DNA control were analyzed. Macrolide resistance was detected in 18, 15, and 16 of the samples with the respective methods. Sequencing of the 23S rRNA gene confirmed resistance in 16 (16%) of 100 samples in which it was detected with any of the three methods. ResistancePlus® MG and S-DiaMGRes™ falsely determined one sample as macrolide-sensitive, but this sample was determined as resistant when retested. The sensitivity of the methods was comparable, although there should be awareness of possible incorrect determination of macrolide resistance, especially of low-positive samples.

Clinical Efficacy of Doxycycline for Treatment of Macrolide-Resistant Mycoplasma pneumoniae Pneumonia in Children.

In areas with high prevalence of macrolide-resistant Mycoplasma pneumoniae (MRMP) pneumonia, treatment in children has become challenging. This study aimed to analyze the efficacy of macrolides and doxycycline with regard to the presence of macrolide resistance. We analyzed children with MP pneumonia during the two recent epidemics of 2014–2015 and 2019–2020 from four hospitals in Korea. Nasopharyngeal samples were obtained from children with pneumonia for MP cultures and polymerase chain reaction (PCR). Macrolide resistance was determined by the analysis of 23S rRNA gene transition. Time to defervescence and to chest X-ray improvement were analyzed. Of 145 cases, the median age was 5.0 years and MRMP accounted for 59 (40.7%). Among macrolide-susceptible MP (MSMP), 78 (90.7%) were treated with macrolides and 21 (35.6%) in the MRMP group with doxycycline. In MRMP pneumonia, shorter days to defervescence (2 vs. 5 days, p < 0.001) and to chest X-ray improvement (3 vs. 6 days, p < 0.001) in the doxycycline group than in the macrolide group was observed, whereas no differences were observed among children with MSMP pneumonia. Compared to macrolides, treatment with doxycycline resulted in better outcomes with a shorter time to defervescence and to chest X-ray improvement among children with MRMP pneumonia.

Usefulness of Q-Probe PCR in Detecting Macrolide-Resistant Mycoplasma pneumoniae Infection in Children.

To investigate the usefulness of quenching probe polymerase chain reaction (Q-probe PCR) for the detection of macrolide-resistant Mycoplasma pneumoniae (MP), we retrospectively analyzed the clinical course of 21 children with MP infection. The rate of macrolide-resistant MP was 66.7%. The duration of pyrexia after the initial antibiotic treatment was longer in patients with macrolide-resistant MP infection than in those with macrolide-sensitive MP infection. The duration of pyrexia after Q-probe PCR was not significantly different between patients with macrolide-resistant and -sensitive MP infection. Antibiotic use based on qPCR may reduce the duration of pyrexia. Q-probe PCR is useful in determining the appropriate antibiotics and improves the clinical course of MP infections.

The treatment of macrolide-resistant Mycoplasma pneumoniae pneumonia in children.

In recent years, the resistance of Mycoplasma pneumoniae to macrolide antibiotics has increased significantly. The health systems are facing significant challenges in carrying out the diagnosis and treatment of refractory Mycoplasma pneumoniae pneumonia in children. Levofloxacin is suitable for treating infectious diseases in various systems but limited in children due to arthropathy issues in weight-bearing joints. This study aimed to evaluate the efficacy and safety of levofloxacin in children with macrolide-resistant Mycoplasma pneumoniae pneumonia. We retrospectively enrolled six confirmed cases of refractory Mycoplasma pneumoniae pneumonia who were admitted in the paediatric respiratory ward of Shandong provincial hospital Affiliated to Shandong first Medical University between 1st January 2020 and 29th February 2020. Levofloxacin was given to the patients through the intravenous or oral route as per the following dosages :<5years, 8-10mg/kg q12h; >5years, 8-10mg/kg, qd for 10days. The clinical data were collected and analysed. The average age of the enrolled cases was six years and nine months (range, four years, and seven months to eleven years and seven months). All cases were found to be drug-resistant and were treated with azithromycin combined with antibacterial drugs. Levofloxacin was used in the patient's refractory to macrolide antibiotics. The temperature of five cases returned to normal 1-2days after treatment with levofloxacin, and the imaging of the four cases showed expected improvements. The gastrointestinal symptoms, neurological manifestations, joint symptoms, blood parameters, liver and kidney functions, and exercise conditions of the children were closely monitored. The follow-up time of the patients ranged from one week to five months. No drug-related adverse reactions were observed in patients during treatment or during follow-up. The clinical symptoms and imaging significantly improved after treatment with levofloxacin, and no drug-related adverse reactions were observed. Levofloxacin proved to be an effective and safe drug in the treatment of children with macrolide-resistant mycoplasma pneumonia. This study will provide a reference for evaluating the efficacy and safety of levofloxacin in the paediatric population.

Risk factors for the development of post-infectious bronchiolitis obliterans after Mycoplasma pneumoniae pneumonia in the era of increasing macrolide resistance

BackgroundThe prevalence of macrolide-resistant Mycoplasma pneumoniae (MP) pneumonia has been rapidly increased. MP pneumonia is a risk factor for the development of post-infectious bronchiolitis obliterans (PIBO). The aim of the present study was to identify the risk factors for the development of PIBO after MP pneumonia in the era of increasing macrolide resistance of MP. Materials and methodsThis retrospective study enrolled 150 children with a mean age of 6.0 years admitted to the hospital due to MP pneumonia between May 2019 and February 2020 at a tertiary hospital. The clinical, radiologic, and laboratory data were obtained using retrospective chart review. ResultsEighteen children (12%) were diagnosed with PIBO after MP pneumonia. PIBO was diagnosed after a mean duration of 100.0 days (range, 6–268 days) from symptom onset. The respiratory virus co-infection (adjusted odds ratio [aOR], 4.069; 95% confidence interval [95% CI], 1.224–13.523), adenovirus co-infection (aOR, 5.607; 95% CI, 1.801–17.454), longer duration between symptom onset and admission (aOR, 1.150; 95% CI, 1.020–1.298), higher levels of serum lactate dehydrogenase (LDH) at the time of admission (aOR, 1.001; 95% CI, 1.000–1.003), and poor response to stepwise treatment increased the risk for development of PIBO after MP pneumonia. However, macrolide resistance of MP was not associated with development of PIBO after MP pneumonia. ConclusionThe present study suggests that respiratory virus co-infection, including adenovirus, poor response to the treatment of MP pneumonia, and higher levels of serum LDH, but not macrolide resistance of MP, are risk factors of PIBO after MP pneumonia.

Recent acute reduction in macrolide-resistant Mycoplasma pneumoniae infections among Japanese children

IntroductionMycoplasma pneumoniae contributes to numerous pneumonia cases among children and young adults. Therefore, this study aimed to investigate the prevalence of M. pneumoniae infections among Japanese children, occurring since 2008. MethodsNasopharyngeal swab specimens were obtained from all cases, following which real-time PCR was performed to identify M. pneumoniae. Further, the p1 genotypes of isolates were determined using the PCR restriction fragment length polymorphism typing method. ResultsThe annual rate of macrolide-resistant M. pneumoniae (MRMP) infections peaked at 81.8% in 2012 and decreased annually until 2015. Although the infection rate increased to 65.3% in 2016, it decreased again to 14.3% in 2018. Although >90% of isolates harbored the type 1 genotype until 2012, this rate decreased, and approximately 80% harbored p1 genotypes other than type 1 in 2018. Furthermore, the occurrence rate of MRMP among the type 1 isolates was very high (82.4%), whereas that among p1 genotypes other than type 1 was very low (6.5%). ConclusionsMRMP occurrence potentially decreased owing to changes in not only antibiotic usage but also in the distribution of p1 genotype among isolates.

Clonal spread of macrolide-resistant Mycoplasma pneumoniae sequence type-3 and type-17 with recombination on non-P1 adhesin among children in Taiwan

ObjectivesMycoplasma pneumoniae is currently the most commonly detected bacterial cause of childhood community-acquired pneumonia in several countries. Of note, clonal expansion of macrolide-resistant ST3 occurred in Japan and South Korea. An alarming surge in macrolide resistance complicates the treatment of pneumonia. We aimed to evaluate the clinical manifestation and clonal relatedness of M. pneumoniae circulating among children in Taiwan. MethodsWe prospectively enrolled 626 children with radiologically confirmed pneumonia between 2017 and 2019. An M. pneumoniae infection was suspected on clinical grounds, and tested by real-time PCR and oropharyngeal swab cultures. We used multilocus sequence typing and whole-genome sequencing to characterize the genetic features of M. pneumoniae. ResultsA total of 226 children with M. pneumoniae pneumonia were enrolled. Macrolide resistance was found in 77% (174/226) of patients. Multi-locus sequence typing revealed that ST3 (n = 93) and its single-locus variant ST17 (n = 84) were the predominant clones among macrolide-resistant strains. ST17 presented clinical characteristics comparable to its ancestor ST3. On multivariate analysis, macrolide resistance (OR 3.5; 95% CI 1.4–8.5; p 0.007) was independently associated with fever >72 hours after macrolide treatment. By whole-genome sequencing, prediction analysis of recombination sites revealed one recombination site in ST3 and ST17 compared with M29 (a macrolide-sensitive ST3 strain isolated from China in 2005) containing cytadhesin MgpC-like protein, RepMP4 and RepMP5. ST17 had another recombination site containing an adhesin and RepMP2/3. ConclusionsIn addition to macrolide resistance, ST3 and its ST17 variant might evolve through recombination between repetitive sequences and non-P1 cytadhesins for persistent circulation in Taiwan.

Sequence Type Changes Associated with Decreasing Macrolide-Resistant Mycoplasma pneumoniae, Japan.

We compared sequence types (STs) of Mycoplasma pneumoniae isolates from Japan during 2002–2019. ST3 and ST14 dominated during 2002–2016, and ST7 and ST33 dominated during 2018–2019. These STs were associated with a decrease in macrolide-resistant strains after an epidemic of infection with M. pneumoniae during 2011–2012.

Epidemiology of macrolide-resistant Mycoplasma pneumoniae by age distribution in Japan

IntroductionMycoplasma pneumoniae (M. pneumoniae) is the major pathogen involved in community-acquired pneumonia in all age groups. Resistance to macrolides, the first-line treatment for M. pneumoniae infection, is a major global public health concern. However, studies evaluating macrolide-resistant M. pneumoniae infection simultaneously in all ages are limited. This study aimed to determine the prevalence and clinical characteristics of macrolide-resistant M. pneumoniae infection in terms of age distribution. MethodsWe enrolled 292 patients in Tokyo, Japan, who visited Eiju General Hospital or Zama Children's Clinic in 2015–2016. Patients were tested using real-time PCR for M. pneumoniae DNA. PCR-positive patients (n = 151) were further selected and sequentially divided into preschool-aged children (≤5 years, n = 31), school-aged children (6–15 years, n = 101), adolescents (16–19 years, n = 5), and adults (≥20 years, n = 14). We then analyzed the M. pneumoniae infection clinical characteristics, prevalence of macrolide-resistant infection, and 23S rRNA domain V resistance-associated mutation status. ResultsWe found insignificant differences in the prevalence of macrolide-resistant M. pneumoniae infection among all groups, clinical characteristics, and resistance-associated mutation status in patients with macrolide-resistant M. pneumoniae infection. We also found statistically higher prevalence of mutation-positive (n = 85) M. pneumoniae in patients previously treated with macrolide compared to the mutation-negative group (n = 66); 63.8% vs 11.1% (p < 0.001), respectively. ConclusionsWe found no significant differences in both clinical characteristics and prevalence of macrolide-resistant M. pneumoniae infection among all ages. Also, previous macrolide treatment contributes to drug-resistance.

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

BackgroundCases of refractory Mycoplasma pneumoniae pneumonia have been increasing recently; however, whether viral coinfection or macrolide-resistant M. infection contribute to the development of refractory M. pneumoniae pneumonia remains unclear. This study aimed to investigate the impacts of viral coinfection and macrolide-resistant M. pneumoniae infection on M. pneumoniae pneumonia in hospitalized children and build a model to predict a severe disease course.MethodsNasopharyngeal swabs or sputum specimens were collected from patients with community-acquired pneumonia meeting our protocol who were admitted to Shanghai Children’s Medical Center from December 1, 2016, to May 31, 2019. The specimens were tested with the FilmArray Respiratory Panel, a multiplex polymerase chain reaction assay that detects 16 viruses, Bordetella pertussis, M. pneumoniae, and Chlamydophila pneumoniae. Univariate and multivariate logistic regression models were used to identify the risk factors for adenovirus coinfection and macrolide-resistant mycoplasma infection.ResultsAmong the 107 M. pneumoniae pneumonia patients, the coinfection rate was 56.07%, and 60 (60/107, 56.07%) patients were infected by drug-resistant M. pneumoniae. Adenovirus was the most prevalent coinfecting organism, accounting for 22.43% (24/107). The classification tree confirmed that viral coinfection was more common in patients younger than 3 years old. Adenovirus coinfection and drug-resistant M. pneumoniae infection occurred more commonly in patients with refractory M. pneumoniae pneumonia (P = 0.019; P = 0.001). A prediction model including wheezing, lung consolidation and extrapulmonary complications was used to predict adenovirus coinfection. The area under the receiver operating characteristic curve of the prediction model was 0.795 (95% CI 0.679–0.893, P < 0.001). A prolonged fever duration after the application of macrolides for 48 h was found more commonly in patients infected by drug-resistant M. pneumoniae (P = 0.002). A fever duration longer than 7 days was an independent risk factor for drug-resistant Mycoplasma infection (OR = 3.500, 95% CI = 1.310–9.353, P = 0.012).ConclusionsThe occurrence of refractory M. pneumoniae pneumonia is associated with adenovirus coinfection and infection by drug-resistant M. pneumoniae. A prediction model combining wheezing, extrapulmonary complications and lung consolidation can be used to predict adenovirus coinfection in children with M. pneumoniae pneumonia. A prolonged fever duration indicates drug-resistant M. pneumoniae infection, and a reasonable change in antibiotics is necessary.

Macrolide-Resistant Mycoplasma pneumoniae Infections in Pediatric Community-Acquired Pneumonia.

A high prevalence rate of macrolide-resistant Mycoplasma pneumoniae (MRMP) has been reported in Asia. We performed a systematic review and meta-analysis to investigate the effect of macrolide resistance on the manifestations and clinical judgment during M. pneumoniae infections. We found no difference in clinical severity between MRMP and macrolide-sensitive Mycoplasma pneumoniae (MSMP) infections. However, in the pooled data, patients infected with MRMP had a longer febrile period (1.71 days), length of hospital stay (1.61 day), antibiotic drug courses (2.93 days), and defervescence time after macrolide treatment (2.04 days) compared with patients infected with MSMP. The risk of fever lasting for >48 hours after macrolide treatment was also significantly increased (OR 21.24), and an increased proportion of patients was changed to second-line treatment (OR 4.42). Our findings indicate diagnostic and therapeutic challenges after the emergence of MRMP. More precise diagnostic tools and clearly defined treatment should be appraised in the future.

Evaluation of Commercial Molecular Diagnostic Methods for Detection and Determination of Macrolide Resistance in Mycoplasma pneumoniae.

We evaluated six commercial molecular tests targeting Mycoplasma pneumoniae, namely, the BioFire FilmArray respiratory panel (RP), the Meridian Alethia Mycoplasma Direct, the GenMark ePlex respiratory pathogen panel (RPP), the Luminex NxTAG RPP, the ELITech ELITe InGenius Mycoplasma MGB research use only (RUO) PCR, and the SpeeDx Resistance Plus MP assays. Laboratory-developed PCR assays at the University of Alabama at Birmingham and the Centers for Disease Control and Prevention were used as reference standards. Among 428 specimens, 212 were designated confirmed positives for M. pneumoniae The highest clinical sensitivities were found with the InGenius PCR (99.5%) and the FilmArray RP (98.1%). The Resistance Plus MP identified 93.3% of the confirmed-positive specimens, whereas 83.6, 64.6, and 55.7% were identified by the ePlex RPP, NxTAG RPP, and Mycoplasma Direct assays, respectively. There was no significant difference between the sensitivity of the reference methods and that of the FilmArray RP and InGenius assays, but the remaining four assays detected significantly fewer positive specimens (P < 0.05). Specificities of all assays were 99.5 to 100%. The Resistance Plus MP assay detected macrolide resistance in 27/33 specimens, resulting in a sensitivity of 81.8%. This study provides the first large-scale comparison of commercial molecular assays for detection of M. pneumoniae in the United States and identified clear differences among their performance. Additional studies are necessary to explore the impact of various test performances on patient outcome.

Macrolide-resistant Mycoplasma pneumoniae pneumonia in transplantation: Increasingly typical?

Mycoplasma pneumoniae is one of the most common bacterial causes of pneumonia. Macrolide-resistant Mpneumoniae (MRMP) was documented in 7.5% of isolates in the United States. Resistance portends poor outcomes to macrolide therapy, yet patients respond well to fluoroquinolones or tetracyclines such as minocycline. However, MRMP may be under-appreciated because Mpneumoniae generally causes relatively mild infections in non-immunosuppressed adults that may resolve without effective therapy and because microbiological confirmation and susceptibility are not routinely performed. We report two cases of pneumonia due to MRMP in kidney transplant recipients. Both patients required hospital admission, worsened on macrolide therapy, and rapidly defervesced on doxycycline or levofloxacin. In one case, Mpneumoniae was only identified by multiplex respiratory pathogen panel analysis of BAL fluid. Macrolide resistance was confirmed in both cases by real-time PCR and point mutations associated with macrolide resistance were identified. Mpneumoniae was isolated from both cases, and molecular genotyping revealed the same genotype. In conclusion, clinicians should be aware of the potential for macrolide resistance in Mpneumoniae, and may consider non-macrolide-based therapy for confirmed or non-responding infections in patients who are immunocompromised or hospitalized.

Systematic research to overcome newly emerged multidrug-resistant bacteria.

In the 1980s, I found that the chromosomal β-lactamase of Klebsiella pneumoniae LEN-1 showed a very high similarity to the R-plasmid-mediated penicillinase TEM-1 on the amino acid sequence level, and this strongly suggested the origination of TEM-1 from the chromosomal penicillinases of K. pneumoniae or related bacteria. Moreover, the chromosomal K1 β-lactamase (KOXY) of Klebsiella oxytoca was found to belong to the class A β-lactamases that include LEN-1 and TEM-1, although KOXY can hydrolyze cefoperazone (CPZ) like the chromosomal AmpC-type cephalosporinases of various Enterobacteriaceae that can hydrolyze several cephalosporins including CPZ. Furthermore, my collaborators and I found plural novel serine-type β-lactamases, such as MOX-1, SHV-24, TEM-91, CTX-M-64, CMY-9, CMY-19, GES-3, GES-4, and TLA-3, mediated by plasmids. Besides these serine-type β-lactamases, we also first identified exogenously acquired metallo-β-lactamases (MBLs), IMP-1 and SMB-1, in imipenem-resistant Serratia marcescens, and the IMP-1-producing S. marcescens TN9106 became the index case for carbapenemase-producing Enterobacteriaceae. I developed the sodium mercaptoacetic acid (SMA)-disk test for the simple identification of MBL-producing bacteria. We were also the first to identify a variety of plasmid-mediated 16S ribosomal RNA methyltransferases, RmtA, RmtB, RmtC, and NpmA, from various Gram-negative bacteria that showed very high levels of resistance to a wide range of aminoglycosides. Furthermore, we first found plasmid-mediated quinolone efflux pump (QepA) and fosfomycin-inactivating enzymes (FosA3 and FosK). We also first characterized penicillin reduced susceptible Streptococcus agalactiae, macrolide-resistant Mycoplasma pneumoniae, as well as Campylobacter jejuni, and Helicobacter pylori, together with carbapenem-resistant Haemophilus influenzae. We constructed a PCR-based open reading frame typing method for rapid identification of Acinetobacter baumannii international clones.

In vitro activities of acetylmidecamycin and other antimicrobials against human macrolide-resistant Mycoplasma pneumoniae isolates.

To assess the in vitro activities of acetylmidecamycin, a 16-membered macrolide, and 11 other antimicrobial agents against human mycoplasmas. A total of 187 clinical isolates, Mycoplasma pneumoniae (n = 110), Mycoplasma hominis (n = 26) and Ureaplasma species (n = 51), were included in this study. The MICs of 12 antimicrobial agents, including acetylmidecamycin, thiamphenicol, chloramphenicol and some other macrolides, fluoroquinolones and tetracyclines, for these clinical isolates were determined by the broth microdilution method. For M. pneumoniae, the MIC90 values of the tested macrolides were: acetylmidecamycin (1 mg/L)<josamycin (4 mg/L)<midecamycin (8 mg/L)<azithromycin (16 mg/L)<erythromycin (>128 mg/L). Thiamphenicol and chloramphenicol had the same MIC90 (2 mg/L). For Ureaplasma species, the MIC90 values were: acetylmidecamycin (0.25 mg/L)<josamycin (0.5 mg/L)=midecamycin<azithromycin (1 mg/L)=erythromycin. Chloramphenicol had a lower MIC90 (2 mg/L) than that of thiamphenicol (4 mg/L). For M. hominis, the MIC90 values were: acetylmidecamycin (0.25 mg/L)<josamycin (0.5 mg/L)<midecamycin (2 mg/L)<azithromycin (>128 mg/L)=erythromycin. The MIC90 values of chloramphenicol and thiamphenicol were 2 and 4 mg/L, respectively. The results indicated that acetylmidecamycin and thiamphenicol are active in vitro against the most common mycoplasma species infecting humans, including those resistant to macrolides and fluoroquinolones. Acetylmidecamycin and thiamphenicol might be a promising option for clinicians to treat infections caused by Mycoplasma and Ureaplasma spp., particularly macrolide-resistant M. pneumoniae in paediatrics and fluoroquinolone-resistant M. hominis in adults. Further investigation of their clinical roles in treating infections caused by these organisms is warranted.

Macrolide-Resistant Mycoplasma pneumoniae in the United States as Determined from a National Surveillance Program.

There are sparse data to indicate the extent that macrolide-resistant Mycoplasma pneumoniae (MRMp) occurs in the United States or its clinical significance. Between 2015 and 2018, hospitals in 8 states collected and stored respiratory specimens that tested positive for M. pneumoniae and sent them to the University of Alabama at Birmingham, where real-time PCR was performed for detection of 23S rRNA mutations known to confer macrolide resistance. MRMp was detected in 27 of 360 specimens (7.5%). MRMp prevalence was significantly higher in the South and East (18.3%) than in the West (2.1%). A2063G was the predominant 23S rRNA mutation detected. MICs for macrolide-susceptible M. pneumoniae (MSMp) were ≤0.008 μg/ml, whereas MICs for MRMp were 16 to 32 μg/ml. Patients with MRMp infection were more likely to have a history of immunodeficiency or malignancy. Otherwise, there were no other significant differences in the clinical features between patients infected with MRMp and those infected with MSMp, nor were there any differences in radiographic findings, hospitalization rates, viral coinfections, the mean duration of antimicrobial treatment, or clinical outcomes. There was no significant change in MRMp incidence over time or according to age, sex, race/ethnicity, or status as an inpatient or an outpatient. Patients with MRMp were more likely to have received a macrolide prior to presentation, and their treatment was more likely to have been changed to a fluoroquinolone after presentation. This is the first national surveillance program for M. pneumoniae in the United States. Additional surveillance is needed to assess the clinical significance of MRMp and to monitor changes in MRMp prevalence.

Outbreak of macrolide-resistant mycoplasma pneumoniae in a primary school in Beijing, China in 2018

BackgroundOn 7th June, 2018, a primary school in Beijing, China notified Shunyi CDC of an outbreak of acute respiratory disease characterized by fever and cough among students and resulting in nine hospitalization cases during the preceding 2 weeks. We started an investigation to identify the etiologic agent, find additional cases, develop and implement control measures.MethodsWe defined probable cases as students, teachers and other staffs in the school developed fever (T ≥ 37.5 °C) with cough or sore throat; or a diagnosis of pneumonia during May 1–June 31, 2018. Confirmed cases were probable cases with Mycoplasma pneumoniae detected in oropharyngeal (OP) swabs by quantitative real-time polymerase chain reaction (qPCR). We searched case by reviewing school absenteeism records and interviewing students, teachers and staff in this school. Oropharyngeal swabs were collected from symptomatic students. Two qPCR) assay, a duplex qPCR assay, and sequencing were performed to determine the pathogen, genotype and macrolide resistance at the gene level, respectively.ResultsFrom May 1st to June 31st, 2018, we identified 55 cases (36 probable and 19 confirmed), of whom 25 (45%) were hospitalized for complications. All cases were students, none of the teachers and other staffs in the school were with similar symptoms. The attack rate (AR) was 3.9% (55/1398) for all students. The cases were mainly male (58%), with an age range of 7–8 years (median: 7 years). 72% (18/25) of inpatients had radiograph findings consistent with pneumonia, and some cases were hospitalized for up to 4 weeks. Pathogen detection results indicated that Mycoplasma pneumonia (M. pneumoniae) P1 type 1 was the causative agent in this outbreak, and the strain harbored one point mutation of A to G at position 2063.ConclusionsThe infections by macrolide-resistant M. pneumoniae are not always mild and pneumonia was common and M. pneumoniae could causes serious complications which require long-term hospitalization. In the future infectious disease prevention and control practice, M. pneumoniae should be paid more attention. It is necessary to establish and improve the pathogen and drug resistance surveillance system in order to prevent and control such mutated strains of M. pneumoniae from causing future outbreaks or epidemics in China.

ATYPICAL ATYPICALS: TWO CASES OF MACROLIDE-RESISTANT MYCOPLASMA PNEUMONIAE INFECTIONS

SESSION TITLE: Monday Fellow Case Report Posters SESSION TYPE: Fellow Case Report Posters PRESENTED ON: 10/21/2019 02:30 PM - 03:15 PM INTRODUCTION: Mycoplasma pneumoniae pneumonia is self-limiting in most cases, and is usually treated with macrolide antibiotics. However, macrolide resistance is increasingly recorded especially in South East Asia, where prevalence has been reported to be as high as 90%. The following cases describe two patients at a U.S. hospital who had severe community-acquired M. pneumoniae pneumonia (CAP) that did not respond to macrolide therapy. CASE PRESENTATION: Patient 1: A 45-year-old female was admitted to the hospital with acute hypoxemic respiratory failure and a left lower lobe pneumonia on imaging. She reported a week of respiratory symptoms, which had been treated with amoxicillin-clavulanate without improvement. The serum Mycoplasma IgM antibody titer was elevated (4.89, threshold 0.91) on admission. Patient 2: A 74-year-old female was re-admitted to the hospital with worsening hypoxemic respiratory failure. She had been admitted a week prior with similar symptoms. During that hospitalization her Mycoplasma IgM antibody titer was below diagnostic threshold (0.09). On re-admission, the Mycoplasma IgM titer had significantly increased (3.59), and she had a right middle lobe predominant pneumonia.. In each case, Mycoplasma PCR was positive, and azithromycin was started. However the patients remained profoundly hypoxemic requiring oxygen via high flow nasal cannula and the antibiotic regimen was expanded to include vancomycin and piperacllin-tazobactam without much clinical change. Azithromycin was switched to levofloxacin around hospital day 3 in each case, considering the possibility of macrolide resistance. The patients' hypoxemia and radiographic abnormalities began to improve significantly with fluoroquinolone therapy. DISCUSSION: M. pneumoniae lacks a cell wall and is thus intrinsically resistant to antibiotics that target the cell wall. The mainstays of therapy include macrolides, ketolides, tetracyclines and fluroquinolones. Resistance to antibiotic therapy occurs through acquired mutations affecting the ribosomal targets of macrolide antibiotics, most commonly the A2058G mutation in the 23S rRNA. The prevalence of macrolide resistance is extremely high in Asia, with rates up to 100% being reported in countries like China. In North America, macrolide resistance rates are 3-13% across various studies. Europe, in contrast, has lower rates of resistance (0-10%) with the exception of Italy (26%). While most studies have reported similar clinical severity amongst M. pneumoniae infections regardless of susceptibility, the patients in our series had significant hypoxemia and were at risk of poor outcomes without the change in antibiotic regimen based on recognition of potential macrolide resistance. CONCLUSIONS: Macrolide-resistant M. pneumoniae infections represent a clinical entity that should be recognized early to initiate prompt antibiotic changes and initiate effective treatment. Reference #1: Pereyre S, Goret J, Bébéar C. Mycoplasma pneumoniae: Current Knowledge on Macrolide Resistance and Treatment. Front Microbiol. 2016;7:974. Reference #2: To KKW, Chan KH, Fung YF, Yuen KF, Ho PL. Azithromycin treatment failure in macrolide-resistant Mycoplasma pneumoniae pneumonia. Eur Resp Journal 2010 36: 969-971. Reference #3: Cao B, Zhao CJ, Yin YD, et. al. High prevalence of macrolide resistance in Mycoplasma pneumoniae isolates from adult and adolescent patients with respiratory tract infection in China. Clin Infect Dis. 2010 Jul 15;51(2):189-94. DISCLOSURES: No relevant relationships by Alok Bhatt, source=Web Response No relevant relationships by John Munger, source=Web Response

Four-color multiplex real-time PCR assay prototype targeting azithromycin resistance mutations in Mycoplasma genitalium

BackgroundThe worldwide expansion of macrolide-resistant Mycoplasma genitalium (MG) in cases of genital infections has led to an increased recurrence rate of these infections after first-line azithromycin treatment. By detecting the presence of azithromycin-resistant MG, the patient’s antibiotic treatment can be targeted and the spread of resistance prevented. With this aim in mind, macrolide-resistance detection kits are helpful tools for the physician.MethodsAzithromycin resistance mutations in MG are targeted using a four-color multiplex real-time RT-PCR assay. Tested targets include plasmid DNA (as positive controls) as well as macrolide-sensitive and macrolide-resistant genomic DNA from characterized cell lines and clinical samples.ResultsThe analytical data presented here were generated from plasmid DNA and genomic RNA/DNA and include adaptation to an internal control, specificity between targets, specificity vs non-MG species, limit of detection (LoD) and interference studies (co-infection and endogenous substances). The clinical data were based on the application of the assay to clinical samples characterized by sequencing.ConclusionsA new NAAT (nucleic acid amplification test) prototype has been developed in collaboration with the Diagenode s.a. company, this prototype targets MG and azithromycin-resistance mutations in that pathogen.

First Detection and Characterization of Macrolide-Resistant Mycoplasma pneumoniae from People with Community-Acquired Pneumonia in Iran.

Objectives: Increasing macrolide resistance of Mycoplasma pneumoniae strains is becoming a public health concern worldwide. Nevertheless, no comprehensive genomic background of circulating isolates is available in our region. We aimed to study the genetic diversity of this microorganism using the multiple-locus variable-number tandem-repeat analysis method and to investigate the relationships between MLVA types and macrolide susceptibility profiles of the isolates. Materials and Methods: A total of 270 patients attending Tehran general university hospitals were included in this study. One throat swab was taken from each patient. M. pneumoniae was identified using culture and PCR assay. Macrolide resistance was determined using the broth microdilution method. The MLVA was performed by amplification of four variable-number tandem-repeat loci. Results: Of 270 specimens, M. pneumoniae was detected in 25.2% (n = 68) and 21.8% (n = 59) samples using PCR and culture, respectively. Approximately 56.9% of isolates were resistant to macrolides. Fifty-one of 59 M. pneumoniae isolates were divided into 6 distinct MLVA types. Conclusion: The macrolide-resistant M. pneumoniae (MRMP) rate in this study was relatively high and most of the MRMP isolates were assigned into the type 4/5/7/2. Since a significant association between MLVA type 4/5/7/2 and macrolide resistance of M. pneumoniae isolates was observed, further monitoring of genetic diversity of MRMP isolates might facilitate better understanding of epidemiology of this microorganism. Besides surveillance of the antibiotic susceptibility might be helpful to make necessary reconsiderations on guidelines for treatment of M. pneumoniae infection.