Performance Analysis of Novel Nucleic Acid Detection Kit for Mycoplasma pneumoniae

  • Respiratory tract infections caused by Mycoplasma pneumoniae is a serious risk for child health.
  • It has been difficult to prevent and control for a variety of reasons; therefore, timely diagnosis is particularly important for treatment of patients.
  • At present, the rapid M pneumoniae test kits based on nucleic acid amplification have been commercialized and used as primary diagnostic tools for M pneumoniae infection, but current kits are time-consuming, which is difficult to meet the requirement for accurate and rapid diagnosis of M pneumoniae during epidemics.
  • Rapid and accurate test kits are urgently required to diagnose M pneumoniae infection.
  • In this article, we evaluated the performance of a novel nucleic acid detection kit (A) for M pneumonia from feasibility and sensitivity, and compared it with kit B.
  • Results showed this kit has the advantage of being rapid, sensitive, and specific, which meets the demands for the diagnosis of M pneumoniae infection in clinical settings.

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Mycoplasma detection in a historical arbovirus repository: Commercial kit comparison and implications for improved repository management.

The Centers for Disease Control and Prevention, Arbovirus Reference Collection (ARC) contains viral isolates from both environmental and human sources that are maintained in the laboratory through passage in suckling mouse brain and/or vertebrate and invertebrate cell culture. There has been increased concern regarding the effect of mycoplasma contamination on virus growth and its impact on research and phenotypic analysis.
Therefore, quality control testing of virus preparations has become a routine part of the ARC quality assurance program.
We compared the performance of three kits – the PCR Mycoplasma Detection Kit (ABM), the VenorGem Mycoplasma Detection Kit (Sigma), and the MycoAlert Mycoplasma Detection Kit (Lonza) – against a reference mycoplasma detection assay from the American Tissue Culture Collection (ATCC) using 744 virus preparations in the ARC, representing 721 unique viruses comprising twelve families and unclassified viruses.
We found the ABM kit had the highest sensitivity and specificity, followed by the Sigma kit and Lonza kit, when compared to the ATCC kit.
An increase in false positives was observed for the Lonza kit for preparations recently passaged in suckling mouse. Our data supports previously reported observations; that once introduced a specific species of mycoplasma is maintained within a lab.

Evaluation of novel nucleic acid detection kit for Mycoplasma pneumoniae

  • For diagnosis of Mycoplasma pneumoniae infection, highly sensitive and rapid diagnosis is important. Because antibiotics are limited for the treatment of M. pneumoniae infection.
  • In this study, we evaluated new rapid nucleic acid detection kit for M. pneumoniae.
  • This kit does not require excessive pretreatment of specimens and molecular diagnosis of M. pneumoniae is possible within 40 min. Using 120 nasopharyngeal specimens, we compared this kit with a commercially available molecular diagnostic reagent (LAMP). 51 of 120 cases were M. pneumoniae positive, and the results of both assays were all consistent.
  • In addition, sequencing of 23S rRNA gene was performed on 51 cases positive for M. pneumoniae. As a result, macrolide resistance mutation (2063A>G) was observed in 19 cases (37.3%).
  • The gene mutations estimated by this kit coincided completely with the sequencing. In conclusion, new rapid nucleic acid detection kit could detect M. pneumoniae with the same sensitivity as other molecular diagnostics, in a simple process.

Validation of the Kit for Detecting Mycoplasma Genitalium from the Male Urethritis.

Mycoplasma genitalium is one of the pathogenic microorganisms in male urethritis as a sexually transmitted infection (STI). M.genitalium is detected in the urine specimens of 15-25% male patients with urethritis.
The emergence of macrolide- or fluoroquinolone-resistant M.genitalium has become a serious problem in the treatment of male urethritis worldwide, but there is no commercial-based detecting kits accepted by the national insurance in Japan. In this study, we tested the validity of a molecular kit for detecting seven microorganisms related to STI (Anyplex™ II STI-7 Detection which detects Neisseria gonorrhoeae, Chlamydia trachomatis, M.genitalium, Mycoplasma hominis, Ureaplasma urealyticum, Ureaplasma parvum, Trichomonas vaginalis) produced by Seegene company in Korea.
Seventeen M.genitalium strains were used to determine the detection limit of M.genitalium. M.genitalium DNA samples were extracted from M.genitalium strains and the diluted DNA samples were reacted to detect M.genitalium by the Anyplex™ II STI-7 Detection. The detection limit was determined as the maximum dilution of DNA samples and the number of M.genitalium DNA copies calculated.
In this study, the minimum DNA copies to detect M.genitalium by the Anyplex™ II STI-7 Detection was determined to be around 50 per reaction.
The detection rates of M.genitalium in urine specimens were compared between MgPa gene PCR and the Anyplex™ II STI-7 Detection.
The positive and negative concordant rates were high as 96.4% (27/28) and 98.6% (71/72), respectively. The validity of the kit for detecting seven microorganisms related to STI (Anyplex™ II STI-7 Detection) was high and thought to be useful for clinical uses.

Evaluation of the Mycoplasma genitalium Resistance Plus kit for the detection of M. genitalium and mutations associated with macrolide resistance.

To compare performance of the ResistancePlus kit (SpeeDx, Australia) with in-house methods for the detection of Mycoplasma genitalium-specific DNA and mutations associated with resistance to macrolide antimicrobials, directly from clinical specimens.
Assay specificity and sensitivity was analysed using DNA from 46 non-M. genitalium organisms and standard curve analysis, respectively.
A panel of archived DNA extracted from 97 M. genitalium-positive clinical specimens, for which the macrolide susceptibility genotype had been previously determined, were tested on the assay and results were compared.
Final analytical specificity was 100%. Sensitivity was detected to at least 140 genome copies/µL. The assay detected M. genitalium in 92/97 (94.9%, 95% CI 88.4% to 98.3%) previously positive specimens.
The genetic macrolide susceptibility assigned was concordant with previous results in 85/92 (92.4%, 95% CI 85.0% to 96.9%) specimens or 85/97 (87.6%, 95% CI: 79.4% to 93.4%) when the false-negative specimens were included. On seven (7/92, 7.6%) occasions, resistant specimens were called susceptible.
Further testing resolved discrepancies for all but five (5.2%) specimens.
The ResistancePlus assay generally performed well in comparison to methods currently employed at the reference laboratory.
It detected a range of different mutations; however, a small number of specimens that were genotyped as macrolide resistant by Sanger sequencing were either not detected by the assay or were genotyped as susceptible. This could impact on treatment outcomes if assay results were used for patient management.

Evaluation of a Rapid Antigen Detection Kit Targeting L7/L12 Ribosomal Protein for Mycoplasma pneumoniae

We evaluated the usefulness of a rapid antigen detection assay for L7/L12 ribosomal protein (Ribotest Mycoplasma; Asahi Kasei Pharma) for diagnosis of Mycoplasma pneumoniae (M. pneumoniae) infection.
Nasopharyngeal swabs were obtained from patients with pneumonia and/or bronchitis; real-time PCR and the L 7/L12 antigen assays were performed with each sample.
Serum was also taken from each patient, and the particle agglutination (PA) method was used to detect anti-M. pneumoniae antibody in these samples.
Macrolide-resistance genes were detected and M. pneumoniae P1 protein subtyping was performed on PCR-positive samples.
PCR assays were positive for 85 of 212 specimens (40.1%).
Sensitivity and specificity of the L7/L12 antigen assays relative to the PCR standard were 74.1% (63/85) and 81.1% (103/127), respectively. For PCR-positive specimens with a large quantity of M. pneumoniae nucleic acid, sensitivity of the L7/L12 antigen assays seemed to be high. In PCR-positive specimens with fewer than 1.0 x 10(6) copies/mL of M. pneumoniae nucleic acid, sensitivity of the L7/L12 antigen assays seemed to be low.

PCR Mycoplasma Detection Kit

M034-Kit TOKU-E Kit 266 EUR

Luciferase Mycoplasma Detection Kit

20-abx098882 Abbexa
  • 1233.00 EUR
  • 523.00 EUR
  • 648.00 EUR
  • 200rxns
  • 25 rxns
  • 50 rxns

PCR Mycoplasma Detection Kit

abx098883-100rxns Abbexa 100 rxns 537 EUR

Luciferase Mycoplasma Detection Kit

20-abx298016 Abbexa
  • 1010.00 EUR
  • 300.00 EUR
  • 425.00 EUR
  • 200 rxns
  • 25 rxns
  • 50 rxns

PCR Mycoplasma Detection Kit

abx298017-100rxns Abbexa 100 rxns 314 EUR

Mycoplasma Detection Kit (OKSB00010)

OKSB00010 Aviva Systems Biology 50 Tests 382 EUR

Mycoplasma PCR Detection Kit

K1476-100 Biovision 100 Rxns 298 EUR

Formaldehyde Detection Kit

K001-F1 Arbor Assays 2x96 well plate 425 EUR

Ubiquitin Detection Kit

SKT-131-20 Stressmarq 20 assays 495 EUR
When the PA method was used as the standard, the relative sensitivity and specificity of the L7/L12 antigen assays were 41.7% (5/12) and 75.3% (58/77), respectively, for single serum and 60.9% (14/23) and 85.7% (18/21), respectively, for paired sera.
The macrolide-resistance gene A2063G was detected in 20 of the 30 tested PCR-positive specimens (66.7%). Of these 20 A2063G-positive specimens, 13 (65.0%) were positive for the L7/L12 antigen assays.
The numbers of M. pneumoniae P1 subtypes were as follows: types I (22), IIa(2), IIc(1), and untypable (5). The L7/L12 antigen assays gave positive results for 17 of 21 (81.0%) subtype I, 1 of 2 (50.0%) IIa, and 1 of 1(100%) IIc specimens.