Drug susceptibility patterns of Mycobacterium abscessus and Mycobacterium massiliense isolated from respiratory specimens.

In this study, we aimed to retrospectively investigate and compare the drug susceptibility patterns of two major Mycobacterium abscessus complex (MABC) species; M. abscessus and M. massiliense. A total of 546 MABC respiratory isolates (277 M. abscessus and 269 M. massiliense) from 2011 to 2016 were analyzed in this study. We estimated minimum inhibitory concentrations (MICs) using the broth microdilution method, and we calculated MIC50 and MIC90 values from the MIC distribution. Both M. abscessus and M. massiliense were highly susceptible to amikacin and linezolid. For M. abscessus, the proportions of inducible and acquired resistance to clarithromycin were 68.6% and 12.3%, respectively. Only 15.2% of M. abscessus remained susceptible at day 14. On the other hand, none of the M. massiliense showed inducible resistance and 6.3% showed acquired resistance to clarithromycin. A total of 92.6% of the M. massiliense remained susceptible at day 14. The resistance rate of M. abscessus to moxifloxacin (90.3%) was significantly higher than that of M. massiliense (83.3%; p = 0.016). These susceptibility differences may explain the divergent treatment outcomes between patients with pulmonary disease caused by these two species.

Comparative Evaluation Between the RealStar Pneumocystis jirovecii PCR Kit and the AmpliSens Pneumocystis jirovecii (carinii)-FRT PCR Kit for Detecting P. jirovecii in Non-HIV Immunocompromised Patients.

BACKGROUND: Real-time PCR is more sensitive than microscopic examination for detecting Pneumocystis jirovecii. We compared the performance of two assays for detecting P. jirovecii DNA: the RealStar Pneumocystis jirovecii PCR Kit 1.0 CE (Altona Diagnostics, Hamburg, Germany) and the AmpliSens Pneumocystis jirovecii (carinii)-FRT PCR kit (InterLabService Ltd., Moscow, Russia). METHODS: We used 159 samples from the lower respiratory tract (112 bronchoalveolar lavage [BAL] fluid, 37 sputum, and 10 endotracheal aspirate [ETA] samples) of non-HIV immunocompromised patients. Nested PCR and sequencing were used to resolve discordant results. The performance of the two assays was evaluated according to clinical categories (clinical Pneumocystis pneumonia [PCP], possible PCP, or unlikely PCP) based on clinical and radiological observations. RESULTS: The positive and negative percent agreement values were 100% (95% confidence interval [CI], 85.4-100%) and 96.6% (95% CI, 90.9-98.9%), respectively, and kappa was 0.92 (95% CI, 0.84-0.99). P. jirovecii DNA load was significantly higher in the clinical PCP group than in the other groups (P<0.05). When stratified by sample type, the positive rate for BAL fluids from the clinical PCP group was 100% using either assay, whereas the positive rate for sputum/ETA samples was only 20%. CONCLUSIONS: The two assays showed similar diagnostic performance and detected low P. jirovecii burden in BAL fluids. Both assays may be useful as routine methods for detecting P. jirovecii DNA in a clinical laboratory setting, though their results should be interpreted considering sample type.

Simultaneous Detection of Clostridioides difficile Glutamate Dehydrogenase and Toxin A/B: Comparison of the C. DIFF QUIK CHEK COMPLETE and RIDASCREEN Assays.

Various commercial assays have recently been developed for detecting glutamate dehydrogenase (GDH) and/or toxin A/B to diagnose Clostridioides difficile infection (CDI). We compared the performance of two assays for the simultaneous detection of C. difficile GDH and toxin A/B, using 150 stool samples: C. DIFF QUIK CHEK COMPLETE (QCC; TechLab, Blacksburg, VA, USA) and RIDASCREEN Clostridium difficile GDH (RC-GDH) and Toxin A/B (RC-Toxin A/B; R-Biopharm, Darmstadt, Germany). For GDH detection, QCC and RC-GDH showed satisfactory sensitivity (95.7% and 94.3%, respectively) and specificity (92.5% and 93.8%, respectively) compared with C. difficile culture. For toxin A/B detection, QCC showed higher sensitivity than RC-Toxin A/B (60.0% vs 33.3%, P<0.001) compared with toxigenic C. difficile culture. When the results of QCC or RC-GDH+RC-Toxin A/B were used as the first step of a two-step algorithm for diagnosing CDI, QCC permitted more accurate discrimination than RC of positive or negative results for CDI (77.3% and 65.3%, respectively). QCC is useful for the simultaneous detection of C. difficile GDH and toxin A/B as a part of the two-step algorithm for diagnosing CDI.

Performance evaluation of the QMAC-dRAST for staphylococci and enterococci isolated from blood culture: a comparative study of performance with the VITEK-2 system.

Objectives: To evaluate the performance of a rapid antimicrobial susceptibility testing (AST) platform based on microfluidic chip technology, the QMAC-dRAST, which enables AST from colony isolates or positive blood culture broth (PBCB), and to compare the performance of the QMAC-dRAST for staphylococci and enterococci with that of the VITEK-2 system based on reference broth microdilution (BMD). Methods: A total of 110 staphylococcal and enterococcal isolates from blood cultures were included. AST was performed directly using the QMAC-dRAST with PBCB. Thereafter, colony isolates derived from subculture of PBCB were used for the QMAC-dRAST, the VITEK-2 system and BMD. Results: The overall agreement between the QMAC-dRAST with PBCB and BMD was 91.5%. There were 1.2% very major errors (VMEs), 4.3% major errors (MEs) and 5.4% minor errors (mEs). The QMAC-dRAST with colony isolates yielded 94.6% agreement and error rates of 1.0% VMEs, 1.8% MEs and 4.0% mEs. The VITEK-2 system showed 96.2% agreement and error rates of 2.3% VMEs, 0.5% MEs and 2.6% mEs. The incubation time in the QMAC-dRAST was significantly shorter than in the VITEK-2 system (median of 6 versus 10 h; P < 0.0001). Conclusions: The QMAC-dRAST system provided rapid results and represents an alternative to conventional AST methods. The QMAC-dRAST with colony isolates produced more reliable results for staphylococci and enterococci than the QMAC-dRAST with PBCB. The QMAC-dRAST system also performed comparably to BMD and the VITEK-2 system.

Differences in drug susceptibility pattern between Mycobacterium avium and Mycobacterium intracellulare isolated in respiratory specimens.

Mycobacterium avium complex (MAC) is the most common etiologic organisms of nontuberculous mycobacteria (NTM) lung disease. In this study, we aimed to retrospectively investigate the differences in drug susceptibility patterns of two major MAC species; Mycobacterium avium and Mycobacterium intracellulare. A total of 1883 major two MAC isolates (1060 M. avium and 823 M. intracellulare) from respiratory specimens were included in this study during the period 2011─2016. The minimum inhibitory concentrations (MICs) were determined by broth microdilution method and MIC50/MIC90 values were derived from MIC distribution. M. intracellulare had generally low susceptible rates than M. avium for almost all tested antimicrobials except ethambutol and amikacin. The susceptible rate to clarithromycin was >94% of the MAC without significant differences between the two species. The MIC50 values of ciprofloxacin, clarithromycin, linezolid, moxifloxacin, and rifampicin were higher in M. intracellulare than in M. avium, contrary to the results of ethambutol with a higher MIC50 in M. avium. In general, M. intracellulare showed a higher resistance rate and higher MIC50 values than M. avium. Differences between this study and previous reports suggest regional differences in drug susceptibility profile of MAC species.

Resistance mechanisms of linezolid-nonsusceptible enterococci in Korea: low rate of 23S rRNA mutations in Enterococcus faecium.

PURPOSE: To investigate linezolid-resistance mechanisms in linezolid-nonsusceptible enterococci (LNSE) isolated from a tertiary hospital in Korea. METHODOLOGY: Enterococcal isolates exhibiting linezolid MICs ≥4 mg l-1 that were isolated between December 2011 and May 2016 were investigated by PCR and sequencing for mutations in 23S rRNA or ribosomal proteins (L3, L4 and L22) and for the presence of cfr, cfr(B) and optrA genes.Results/Key findings. Among 135 LNSE (87 Enterococcus faecium and 48 Enterococcus faecalis isolates), 39.1 % (34/87) of E. faecium and 18.8 % (9/48) of E. faecalis isolates were linezolid-resistant. The optrA carriage was the dominant mechanism in E. faecalis: 13 isolates, including 10 E. faecalis [70 % (7/10) linezolid-resistant and 30 % (3/10) linezolid-intermediate] and three E. faecium [33.3 % (1/3) linezolid-resistant and 66.7 % (2/3) linezolid-intermediate], contained the optrA gene. G2576T mutations in the 23S rRNA gene were detected only in E. faecium [14 isolates; 71.4 % (10/14) linezolid-resistant and 28.6 % (4/14) linezolid-intermediate]. One linezolid-intermediate E. faecium harboured a L22 protein alteration (Ser77Thr). No isolates contained cfr or cfr(B) genes and any L3 or L4 protein alterations. No genetic mechanism of resistance was identified for 67.6 % (23/34) of linezolid-resistant E. faecium. CONCLUSION: A low rate of 23S rRNA mutations and the absence of known linezolid-resistance mechanisms in the majority of E. faecium isolates suggest regional differences in the mechanisms of linezolid resistance and the possibility of additional mechanisms.

Performance evaluation of the Cobas TaqMan MTB assay on respiratory specimens according to clinical application.

OBJECTIVE: To evaluate the performance of the Cobas TaqMan MTB assay (Cobas assay) with respect to its clinical application. METHODS: This was a retrospective analysis of 1154 results from 1034 patients for whom mycobacterial cultures and the Cobas assay were performed simultaneously. Based on the patient medical records, two categories of clinical application were defined: (1) the diagnosis of patients with a high probability of pulmonary tuberculosis according to clinical and radiological features (n=128), and (2) the exclusion of tuberculosis in clinically indeterminate patients (n=1026). Standard culture was used as the reference method. RESULTS: The sensitivity of the Cobas assay for the detection of Mycobacterium tuberculosis was 70.4% (95% confidence interval (CI) 49.7-85.5%) for category 1, but only 25.0% (95% CI 4.5-64.4%) for category 2. The specificity was ≥95.0% for both categories. The positive predictive value was 79.2% (95% CI 57.3-92.1%) for category 1 and 33.3% (95% CI 6.0-75.9%) for category 2, while the negative predictive value was 92.3% (95% CI 85.0-96.4%) for category 1 and 99.4% (95% CI 98.7-99.8%) for category 2. CONCLUSIONS: The results of this study indicate that Cobas assay results must be interpreted carefully according to the clinical purpose of the assay.

Laboratory Identification of Leptotrichia Species Isolated From Bacteremia Patients at a Single Institution.

We describe the laboratory identification of Leptotrichia species from clinical isolates collected over a six-year period. Five isolates from blood cultures were identified as Leptotrichia species. Gram stain showed large, fusiform, gram-negative or -variable bacilli. Identification based on biochemical testing was unsuccessful; however, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry proved to be a useful tool for identifying Leptotrichia species to the genus level. Species level identification was successfully achieved by using 16S ribosomal RNA gene sequencing.

Comparative evaluation of the AdvanSure Mycobacteria GenoBlot assay and the GenoType Mycobacterium CM/AS assay for the identification of non-tuberculous mycobacteria.

In this study, to assess the performance of the AdvanSure Mycobacteria GenoBlot assay (AdvanSure assay), we compared its performance with that of the GenoType Mycobacterium CM/AS assay (GenoType assay) for the identification of non-tuberculous mycobacteria (NTM). Twenty-four reference strains and 103 consecutive clinical NTM isolates were analysed. The accuracy rates for the 24 reference strains were 87.5 and 95.8 % for the AdvanSure and GenoType assays, respectively. For the 103 clinical isolates, a 91.3 % (94/103) concordance rate was observed between the two assays. The majority (7/9) of discrepancies were isolates identified as Mycobacterium avium complex (MAC) by only the AdvanSure assay. All of these isolates except one were confirmed as MAC by sequence-based typing. The AdvanSure assay showed comparable performance to the GenoType assay and can be useful as a routine method for NTM identification in the clinical setting, especially where MAC is the main cause of NTM infection.

A case of Bacteroides pyogenes bacteremia secondary to liver abscess.

Bacteroides pyogenes, a non-spore-forming, anaerobic, gram-negative rod, is a component of the oral flora of animals and has, on occasion, been reported to cause human infection through dog or cat bites. We report the first case of B. pyogenes bacteremia secondary to liver abscess with no history of an animal bite. The microorganism was identified by 16S rRNA sequencing.

Comparison of BacT/Alert FAN and FAN Plus Bottles with Conventional Medium for Culturing Cerebrospinal Fluid.

We compared the BacT/Alert system FAN and FAN Plus media to conventional media for culturing cerebrospinal fluid (CSF) with 2,545 samples. FAN/FAN Plus bottles showed better performance for isolating microorganisms in CSF than conventional media (positive rate, 7.2% [182/2,545] versus 3.1% [80/2,545]). The incremental recovery rate of Cryptococcus neoformans from FAN Plus bottles was higher than that from FAN bottles.

Evaluation of three real-time PCR assays for differential identification of Mycobacterium tuberculosis complex and nontuberculous mycobacteria species in liquid culture media.

We evaluated the analytical performance of M. tuberculosis complex (MTBC)/nontuberculous mycobacteria (NTM) PCR assays for differential identification of MTBC and NTM using culture-positive liquid media. Eighty-five type strains and 100 consecutive mycobacterial liquid media cultures (MGIT 960 system) were analyzed by a conventional PCR assay (MTB-ID(®) V3) and three real-time PCR assays (AdvanSure™ TB/NTM real-time PCR, AdvanSure; GENEDIA(®) MTB/NTM Detection Kit, Genedia; Real-Q MTB & NTM kit, Real-Q). The accuracy rates for reference strains were 89.4%, 100%, 98.8%, and 98.8% for the MTB-ID V3, AdvanSure, Genedia, and Real-Q assays, respectively. Cross-reactivity in the MTB-ID V3 assay was mainly attributable to non-mycobacterium Corynebacterineae species. The diagnostic performance was determined using clinical isolates grown in liquid media, and the overall sensitivities for all PCR assays were higher than 95%. In conclusion, the three real-time PCR assays showed better performance in discriminating mycobacterium species and non-mycobacterium Corynebacterineae species than the conventional PCR assay.

Evaluation of the Cobas TaqMan MTB test for the detection of Mycobacterium tuberculosis complex according to acid-fast-bacillus smear grades in respiratory specimens.

We evaluated the performance of the Cobas TaqMan MTB test (Roche Diagnostics, Basel, Switzerland), stratified by acid-fast bacilli (AFB) smear grades. The sensitivity of this test in smear-positive specimens was >95% in all grades, while that in trace and negative specimens was 85.3% and 34.4%, respectively.