Yi-Wei Tang

Publications of Yi-Wei Tang

• Comparison of the Luminex xTAG RVP FAST and the Idaho Technology FilmArray RP Assays for the Detection of Respiratory viruses in Pediatric Patients at a Cancer Hospital.

  Authors: N Esther Babady, Peter Mead, Jeffrey Stiles, Carrie Brennan, Haijing Li, Susan Shuptar, Charles W Stratton, Yi-Wei Tang, Mini Kamboj

  Journal of clinical microbiology. 04/2012;

  Background: Respiratory viruses have become increasingly recognized as serious causes of morbidity and mortality in immunocompromised patients. Rapid and sensitive detection of respiratory viruses isBackground: Respiratory viruses have become increasingly recognized as serious causes of morbidity and mortality in immunocompromised patients. Rapid and sensitive detection of respiratory viruses is essential for early diagnosis and administration of appropriate antiviral therapy as well as for effective implementation of infection control measures.Methods: We compared the performance of two commercial assays, the xTAG RVP FAST (Luminex Diagnostics, Toronto, Canada) and the FilmArray RVP (FA RVP, Idaho Technology, Salt Lake City, Utah) in pediatric patients at Memorial Sloan-Kettering Cancer Center. These assays detect the following viruses: respiratory syncytial virus, influenza A and B, parainfluenza 1, 2, 3, 4, human metapneumovirus, adenovirus, enterovirus-rhinovirus, coronaviruses NL63, HKU1, 229E and OC43 and bocavirus. We tested a total of 358 respiratory specimens from 173 pediatric patients previously tested by direct fluorescence assay (DFA) and viral culture.Results: The overall detection rate (positive specimen/total specimen) for viruses tested by all methods was 24% for DFA/culture, 45% for xTAG RVP FAST and 51% for FA RVP. The agreement between the two multiplex assays was 84.5%, and the difference in detection rate was statistically significant (p<0.0001).Conclusion: Overall, the FA RVP was more sensitive than the xTAG RVP FAST and had a turn-around time of approximately 1 hour. The sensitivity, simplicity, and the random access platform make the FA RVP an excellent choice for the laboratory on-demand service with low to medium volume.

• High Proportion of Fluoroquinolone-Resistant M. tuberculosis Isolates with Novel Gyrase Polymorphisms and a gyrA Region Associated with Fluoroquinolone Susceptibility.

  Authors: Rose Devasia, Amondrea Blackman, Svetlana Eden, Haijing Li, Fernanda Maruri, Ayumi Shintani, Charles Alexander, Anne Kaiga, Charles W Stratton, Jon Warkentin, Yi-Wei Tang, Timothy R Sterling

  Journal of clinical microbiology. 12/2011;

  Fluoroquinolone resistance in M. tuberculosis can be conferred by mutations in gyrA or gyrB. The prevalence of resistance mutations outside the quinolone-resistance determining region (QRDR) of gyrAFluoroquinolone resistance in M. tuberculosis can be conferred by mutations in gyrA or gyrB. The prevalence of resistance mutations outside the quinolone-resistance determining region (QRDR) of gyrA or gyrB is unclear, since such regions are rarely sequenced. M. tuberculosis isolates from 1,111 patients with newly-diagnosed culture-confirmed tuberculosis diagnosed in Tennessee 2002-2009 were screened for phenotypic ofloxacin resistance (>2 μg/ml). For each resistant isolate, two ofloxacin-susceptible isolates were selected: one with antecedent fluoroquinolone exposure and one without. The complete gyrA and gyrB genes were sequenced and compared with M. tuberculosis H37Rv. Of 25 ofloxacin-resistant isolates, 11 (44%) did not have previously-reported resistance mutations, of which 10 had novel polymorphisms: 3 in the QRDR of gyrA, 1 in the QRDR of gyrB, and 6 outside the QRDR of gyrA or gyrB; 1 did not have any gyrase polymorphisms. Polymorphisms in gyrA codons 1-73 were more common in fluoroquinolone-susceptible than -resistant strains (20% vs. 0%; P = 0.016). In summary, almost half of fluoroquinolone-resistant M. tuberculosis isolates did not have previously described resistance mutations, which has implications for genotypic diagnostic tests.

• Pearls and Pitfalls of HIV-1 Serologic Laboratory Testing

  Authors: Jiasheng Shao, Yunzhi Zhang, Yi-Wei Tang, Hongzhou Lu

  10/2011;

  ISBN: 978-953-307-601-0

• Methicillin-resistant Staphylococcus aureus bloodstream infections in a Japanese University Hospital between 1987 and 2004.

  Authors: Tohru Takata, Motoyasu Miyazaki, Haijing Li, Mimi Healy, Stacie Frye, Koko Tanaka, Shuji Hara, Hidetoshi Kamimura, Hisae Yoshimura, Akira Matsunaga, Daiki Ohta, Hiroyasu Ishikura, Kazuo Tamura, Yi-Wei Tang

  Journal of infection and chemotherapy : official journal of the Japan Society of Chemotherapy. 10/2011;

  Methicillin-resistant Staphylococcus aureus (MRSA) infections have been the most common cause of nosocomial infections in Japan, but their genetic characteristics related to bloodstream infectionsMethicillin-resistant Staphylococcus aureus (MRSA) infections have been the most common cause of nosocomial infections in Japan, but their genetic characteristics related to bloodstream infections have not been well studied. The aim of this study was to investigate a comprehensive molecular characterization of MRSA blood isolates during the historical 18-year study period between 1987 and 2004 in a tertiary care university hospital. A total of 137 MRSA isolates recovered from the blood of inpatients at Fukuoka University Hospital were analyzed. Clinical information and antimicrobial susceptibility profiles were reviewed, and staphylococcal chromosomal cassette mec (SCCmec), accessory gene regulator (agr), and a battery of bacterial genes were tested by PCR-based assays. The relatedness of these isolates was determined by the repetitive sequence-based PCR (rep-PCR) and pulsed-field gel electrophoresis (PFGE). Although low numbers of agr type III/SCCmec type IV isolates circulated between 1987 and 1992, agr type II/SCCmec type II isolates started circulating in 1993 and were responsible for the increased MRSA isolates until 2004. The rep-PCR and PFGE identified 104 epidemic and 33 sporadic isolates. Among the 104 epidemic isolates, six major rep-PCR/PFGE types were identified, which occupied 67.3% of epidemic isolates. The SCCmec type II and agr type II isolates were observed in significantly higher proportion in epidemic isolates than in sporadic isolates (P = 0.0318, P = 0.0123, respectively). In contrast, SCCmec type IV strains were observed in significantly higher proportion in sporadic isolates than in epidemic isolates (P = 0.0494). Although isolates with sec were detected in higher rates in epidemic isolates (P = 0.0397), seh was detected in higher rates in sporadic isolates (P = 0.0350). Multivariate logistic regression analysis with forward stepping revealed that SCCmec type II was independently associated with epidemic isolates (P = 0.0067; odds ratio, 1.75; 95% confidence interval, 1.17-2.64). These data indicated that SCCmec type II MRSA isolates were responsible for the increased MRSA bloodstream infections for inpatients during the 18-year study period in the hospital.

• Comparative evaluation of Bruker Biotyper and BD Phoenix systems for identification of bacterial pathogens associated with urinary tract infections.

  Authors: Yingjun Yan, Shufang Meng, Dongmo Bian, Criziel Quinn, Haijing Li, Charles W Stratton, Yi-Wei Tang

  Journal of clinical microbiology. 09/2011; 49(11):3936-9.

  The Bruker Biotyper and BD Phoenix systems were evaluated for identification of 1,024 bacterial urinary tract isolates. The Biotyper and Phoenix systems correctly identified 99.9% and 99.5% to theThe Bruker Biotyper and BD Phoenix systems were evaluated for identification of 1,024 bacterial urinary tract isolates. The Biotyper and Phoenix systems correctly identified 99.9% and 99.5% to the genus level and 99.1% and 98.5% to the species level, respectively. Both systems provide reliable results, and the Biotyper system offers a rapid tool for urine bacterial isolate identification.

• Molecular assays for the detection and characterization of respiratory viruses.

  Authors: Yingjun Yan, Shu Zhang, Yi-Wei Tang

  Seminars in respiratory and critical care medicine. 08/2011; 32(4):512-26.

  Patient care providers face an enormous challenge in diagnosing vial respiratory diseases because of similar clinical manifestations, as well as insensitivity and/or slow conventional laboratoryPatient care providers face an enormous challenge in diagnosing vial respiratory diseases because of similar clinical manifestations, as well as insensitivity and/or slow conventional laboratory detection methods. Nucleic acid-targeted molecular assays are playing critical roles in rapid detection, screening, and identification of respiratory viral pathogens due to their high sensitivity and specificity, short test turnaround time, as well as automatic and high-throughput processing. User-developed and commercial molecular methods have gradually been developed and become available for detection and identification of common viral pathogens causing respiratory tract infections. Incorporated with cutting edge techniques, these methods can be used to detect one or more pathogens in a single reaction tube qualitatively and quantitatively. Although the molecular assays have been increasingly used in the clinical setting, laboratorians and clinicians should well know the limitations of these molecular assays to wisely choose the right tests and correctly interpret test results.

• Matrix-assisted laser desorption ionization time-of-flight mass spectrometry and database for identification of Legionella species.

  Authors: Ying He, Tsung C Chang, Haijing Li, Gongyi Shi, Yi-Wei Tang

  Canadian journal of microbiology. 07/2011; 57(7):533-8.

  More than 20 species of Legionella have been identified in relation to human infections. Rapid detection and identification of Legionella isolates is clinically useful to differentiate betweenMore than 20 species of Legionella have been identified in relation to human infections. Rapid detection and identification of Legionella isolates is clinically useful to differentiate between infection and contamination and to determine treatment regimens. We explored the use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) Biotyper system (Bruker Daltonik GmbH, Bremen, Germany) for the identification of Legionella species. The MALDI MS spectra were generated and compared with the Biotyper database, which includes 25 Legionella strains covering 22 species and four Legionella pneumophila serogroups. A total of 83 blind-coded Legionella strains, consisting of 54 reference and 29 clinical strains, were analyzed in the study. Overall, the Biotyper system correctly identified 51 (61.4%) of all strains and isolates to the species level. For species included in the Biotyper database, the method identified 51 (86.4%) strains out of 59 Legionella strains to the correct species level, including 24 (100%) L. pneumophila and 27 (77.1%) non-L. pneumophila strains. The remaining 24 Legionella strains, belonging to species not covered by the Biotyper database, were either identified to the Legionella genus level or had no reliable identification. The Biotyper system produces constant and reproducible MALDI MS spectra for Legionella strains and can be used for rapid and accurate Legionella identification. More Legionella strains, especially the non-L. pneumophila strains, need to be included in the current Biotyper database to cover varieties of Legionella species and to increase identification accuracy.

• Improved identification of yeast species directly from positive blood culture media by combining Sepsityper specimen processing and Microflex analysis with the matrix-assisted laser desorption ionization Biotyper system.

  Authors: Yingjun Yan, Ying He, Thomas Maier, Criziel Quinn, Gongyi Shi, Haijing Li, Charles W Stratton, Markus Kostrzewa, Yi-Wei Tang

  Journal of clinical microbiology. 05/2011; 49(7):2528-32.

  Current methods for identification of yeast from blood cultures may take several days after these microorganisms have been observed by Gram stain smears from positive blood cultures. We explored theCurrent methods for identification of yeast from blood cultures may take several days after these microorganisms have been observed by Gram stain smears from positive blood cultures. We explored the use of a matrix-assisted laser desorption ionization (MALDI) Biotyper system in combination with Sepsityper specimen processing and Microflex analysis for improved detection and identification of yeast species directly from positive blood culture specimens demonstrating yeast-like organisms by Gram stain. The limit of detection of yeast species in blood culture medium was determined to be 5.9 × 10(5) CFU, with intra- and interstrain coefficients of variation of 1.8 to 3.6% and 2.9%, respectively. A total of 42 yeast-containing positive blood culture specimens were processed, and the identification results were compared to those obtained by routinely used phenotypic methods. Specimens with discrepant results between the Biotyper and phenotypic methods were identified on the basis of internal transcribed spacer region sequencing. The MALDI Biotyper system correctly identified the 42 specimens to species level, including 28 (66.7%) Candida albicans, 8 (19.0%) Candida parapsilosis, and 5 (11.9%) Candida tropicalis isolates and 1 (2.4%) Cryptococcus neoformans isolate. The entire procedure, from specimen extraction to final result reporting, can be completed within 1 h. Our data indicated that the Sepsityper specimen processing and Microflex analysis by the MALDI Biotyper system provide a rapid and reliable tool for yeast species identification directly from positive blood culture media.

• Molecular characterization of drug-resistant Mycobacterium tuberculosis isolates circulating in China by multilocus PCR and electrospray ionization mass spectrometry.

  Authors: Feifei Wang, Christian Massire, Haijing Li, Lendell L Cummins, Feng Li, Jialin Jin, Xiaoping Fan, Sen Wang, Lingyun Shao, Shu Zhang, Shufang Meng, Jing Wu, Chanyi Lu, Lawrence B Blyn, Rangarajan Sampath, David J Ecker, Wenhong Zhang, Yi-Wei Tang

  Journal of clinical microbiology. 04/2011; 49(7):2719-21.

  We used multilocus PCR and electrospray ionization mass spectrometry (PCR/ESI-MS) to determine the genotype and drug resistance profiles for 96 Mycobacterium tuberculosis isolates circulating inWe used multilocus PCR and electrospray ionization mass spectrometry (PCR/ESI-MS) to determine the genotype and drug resistance profiles for 96 Mycobacterium tuberculosis isolates circulating in regions of high and low tuberculosis (TB) endemicity in China. The dominant principal genetic group (PGG) circulating in China was PGG1, and drug-resistant gene mutations were more diversified in the region of low rather than high TB endemicity.

• Molecular diagnosis of viral respiratory infections.

  Authors: Shu Zhang, Wenhong Zhang, Yi-Wei Tang

  Current infectious disease reports. 04/2011; 13(2):149-58.

  In clinical practice, a rapid and accurate identification of pathogens causing viral respiratory tract infections can be problematic because of nonspecific clinical presentations, lack of rapid andIn clinical practice, a rapid and accurate identification of pathogens causing viral respiratory tract infections can be problematic because of nonspecific clinical presentations, lack of rapid and sensitive tests, and the emergence of new and mutating viral pathogens. Nucleic acid-targeted molecular techniques are increasingly being used to provide high sensitivity and specificity, short test turnaround time, and automatic and high-throughput processing. In-house and commercially available molecular methods have been developed to qualitatively and quantitatively detect and identify a single or a panel of clinically encountered respiratory tract viruses in a single reaction. Molecular techniques are being gradually introduced in routine laboratory diagnosis of viral respiratory tract infections. However, their performance characteristics and limitations must be clearly understood by both laboratory personnel and clinicians to ensure proper utilization and interpretation.

• Evaluation of a rapid and completely automated real-time reverse transcriptase PCR assay for diagnosis of enteroviral meningitis.

  Authors: Frederick S Nolte, Beverly B Rogers, Yi-Wei Tang, M Steven Oberste, Christine C Robinson, K Sue Kehl, Kenneth A Rand, Harley A Rotbart, Jose R Romero, Ann-Christine Nyquist, David H Persing

  Journal of clinical microbiology. 02/2011; 49(2):528-33.

  Nucleic acid amplification tests (NAATs) for enterovirus RNA in cerebrospinal fluid (CSF) have emerged as the new gold standard for diagnosis of enteroviral meningitis, and their use can improve theNucleic acid amplification tests (NAATs) for enterovirus RNA in cerebrospinal fluid (CSF) have emerged as the new gold standard for diagnosis of enteroviral meningitis, and their use can improve the management and decrease the costs for caring for children with enteroviral meningitis. The Xpert EV assay (Cepheid, Sunnyvale, CA) is a rapid, fully automated real-time PCR test for the detection of enterovirus RNA that was approved by the U.S. Food and Drug Administration for in vitro diagnostic use in March 2007. In this multicenter trial we established the clinical performance characteristics of the Xpert EV assay in patients presenting with meningitis symptoms relative to clinical truth. Clinical truth for enteroviral meningitis was defined as clinical evidence of meningitis, the absence of another detectable pathogen in CSF, and detection of enterovirus in CSF either by two reference NAATs or by viral culture. A total of 199 prospectively and 235 retrospectively collected specimens were eligible for inclusion in this study. The overall prevalence of enteroviral meningitis was 26.04%. The Xpert EV assay had a sensitivity of 94.69% (90% confidence interval [CI] = 89.79 to 97.66%), specificity of 100% (90% CI = 99.07 to 100%), positive predictive value of 100%, negative predictive value of 98.17, and an accuracy of 98.62% relative to clinical truth. The Xpert EV assay demonstrated a high degree of accuracy for diagnosis of enteroviral meningitis. The simplicity and on-demand capability of the Xpert EV assay should prove to be a valuable adjunct to the evaluation of suspected meningitis cases.

• Simultaneous detection and differentiation of respiratory syncytial virus and other respiratory viral pathogens.

  Authors: Susan E Sefers, Haijing Li, Yi-Wei Tang

  Methods in molecular biology (Clifton, N.J.). 01/2011; 665:309-23.

  Rapid and accurate detection of respiratory syncytial virus (RSV) provides pathogen-specific diagnosis, allows implementation of appropriate infection control measures, and improves patientRapid and accurate detection of respiratory syncytial virus (RSV) provides pathogen-specific diagnosis, allows implementation of appropriate infection control measures, and improves patient management. One diagnostic challenge is that respiratory infections, which can be caused by several viral pathogens including RSV, usually present with similar signs and symptoms that are nearly indistinguishable by clinical diagnosis. We have described in the chapter a rapid, high-throughput laboratory technique that can detect a panel of common viral pathogens in one single reaction. With the combination of target-enriched multiplexing PCR amplification and Luminex suspension array identification, 12 common respiratory viruses, including RSV A and B, influenza virus A and B, parainfluenza virus 1, 2, 3, and 4, human metapneumovirus, rhinoviruses, enteroviruses, and SARS coronavirus, are detected and differentiated simultaneously within five hours.

• Basaloid squamous cell carcinoma of the skin.

  Authors: Alan S Boyd, Thomas S Stasko, Yi-Wei Tang

  Journal of the American Academy of Dermatology. 01/2011; 64(1):144-51.

  Basaloid squamous cell carcinoma (BSCC), an aggressive tumor of the aerodigestive tract, was described over 20 years ago, and its defining histologic parameters remain largely unchanged. While rareBasaloid squamous cell carcinoma (BSCC), an aggressive tumor of the aerodigestive tract, was described over 20 years ago, and its defining histologic parameters remain largely unchanged. While rare reports have noted cutaneous metastatic deposition, primary tumors have not been previously described. Although most cutaneous malignancies with basaloid features comprise variants of basal cell carcinomas, a subset exhibit histologic attributes suggestive of more aggressive tumors. We evaluated 3 such tumors submitted to our dermatopathology service over a 6-month period. Immunohistochemical stains useful in differentiating the lineage of cutaneous malignancies with basaloid-appearing tumor cells were employed. Human papillomavirus (HPV) detection and typing were performed by using polymerase chain reaction and sequencing. The tumor cells expressed high molecular weight cytokeratin (34βE12) and cytokeratin 5/6 but not Ber-EP4 or bcl-2. This pattern of immunohistochemical staining and the histologic attributes of the neoplasms are inconsistent with those expected in better defined cutaneous basaloid malignancies but are characteristic of BSCC. Two of the tumors arose in the inguinal crease of middle-aged men, and two patients were known to be immunosuppressed. HPV genotype 33 was detected in the tumor tissue from both inguinal lesions. The number of cases available for evaluation is small and any prognostic implications therefore tenuous. The differential diagnosis of cutaneous malignancies exhibiting basaloid cells should include BSCC, a tumor with an unusual pattern of immunohistochemical staining and a potentially poor prognosis.

• Serum microRNA expression profile distinguishes enterovirus 71 and coxsackievirus 16 infections in patients with hand-foot-and-mouth disease.

  Authors: Lunbiao Cui, Yuhua Qi, Haijing Li, Yiyue Ge, Kangchen Zhao, Xian Qi, Xiling Guo, Zhiyang Shi, Minghao Zhou, Baoli Zhu, Yan Guo, Jun Li, Charles W Stratton, Yi-Wei Tang, Hua Wang

  PloS one. 01/2011; 6(11):e27071.

  Altered circulating microRNA (miRNA) profiles have been noted in patients with microbial infections. We compared host serum miRNA levels in patients with hand-foot-and-mouth disease (HFMD) caused byAltered circulating microRNA (miRNA) profiles have been noted in patients with microbial infections. We compared host serum miRNA levels in patients with hand-foot-and-mouth disease (HFMD) caused by enterovirus 71 (EV71) and coxsackievirus 16 (CVA16) as well as in other microbial infections and in healthy individuals. Among 664 different miRNAs analyzed using a miRNA array, 102 were up-regulated and 26 were down-regulated in sera of patients with enteroviral infections. Expression levels of ten candidate miRNAs were further evaluated by quantitative real-time PCR assays. A receiver operating characteristic (ROC) curve analysis revealed that six miRNAs (miR-148a, miR-143, miR-324-3p, miR-628-3p, miR-140-5p, and miR-362-3p) were able to discriminate patients with enterovirus infections from healthy controls with area under curve (AUC) values ranged from 0.828 to 0.934. The combined six miRNA using multiple logistic regression analysis provided not only a sensitivity of 97.1% and a specificity of 92.7% but also a unique profile that differentiated enterovirial infections from other microbial infections. Expression levels of five miRNAs (miR-148a, miR-143, miR-324-3p, miR-545, and miR-140-5p) were significantly increased in patients with CVA16 versus those with EV71 (p<0.05). Combination of miR-545, miR-324-3p, and miR-143 possessed a moderate ability to discrimination between CVA16 and EV71 with an AUC value of 0.761. These data indicate that sera from patients with different subtypes of enteroviral infection express unique miRNA profiles. Serum miRNA expression profiles may provide supplemental biomarkers for diagnosing and subtyping enteroviral HFMD infections.

• Identification and characterization of OXA-48 producing, carbapenem-resistant Enterobacteriaceae isolates in Turkey.

  Authors: Abdullah Kilic, Zerin Aktas, Orhan Bedir, Ramazan Gumral, Yasemin Bulut, Charles Stratton, Yi-Wei Tang, A Celal Basustaoglu

  Annals of clinical and laboratory science. 01/2011; 41(2):161-6.

  Carbapenem resistance in Enterobacteriaceae isolates has been reported from Turkey and is most often mediated by OXA-48 type carbapenemases. We report the identification and characterization of fourCarbapenem resistance in Enterobacteriaceae isolates has been reported from Turkey and is most often mediated by OXA-48 type carbapenemases. We report the identification and characterization of four carbapenem-resistant isolates (three Klebsiella pneumoniae and one Escherichia coli) among 515 clinical Enterobacteriaceae isolates collected during a 7-month study period in Ankara, Turkey. The four isolates were recovered from blood and urine specimens in patients with varied clinical manifestations. They had distinct pulsed-field gel electrophoresis patterns and harbored a variety of β-lactamases including bla(TEM-1), bla(SHV-12) genes, bla(SHV-11), and/or bla(CTX-M-15). PCR and sequencing analysis revealed that the bla(OXA-48) gene was present in all four isolates. Our data indicated that the OXA-48-type carbapenemase was the only mechanism for carbapenem resistance in our hospital.

• Simultaneous identification of mycobacterial isolates to the species level and determination of tuberculosis drug resistance by PCR followed by electrospray ionization mass spectrometry.

  Authors: Christian Massire, Cristina Agasino Ivy, Robert Lovari, Natalia Kurepina, Haijing Li, Lawrence B Blyn, Steven A Hofstadler, George Khechinashvili, Charles W Stratton, Rangarajan Sampath, Yi-Wei Tang, David J Ecker, Barry N Kreiswirth

  Journal of clinical microbiology. 12/2010; 49(3):908-17.

  Mycobacterium tuberculosis that is resistant to both isoniazid (INH) and rifampin (RIF) is spreading. It has become a public health problem in part because the standard culture methods used toMycobacterium tuberculosis that is resistant to both isoniazid (INH) and rifampin (RIF) is spreading. It has become a public health problem in part because the standard culture methods used to determine the appropriate treatment regimen for patients often take months following the presumptive diagnosis of tuberculosis. Furthermore, the misidentification of nontuberculosis mycobacteria (NTM) in patients presumably suffering from tuberculosis results in additional human and health care costs. The mechanisms of resistance for several drugs used to treat Mycobacterium tuberculosis are well understood and therefore should be amenable to determination by rapid molecular methods. We describe here the use of PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) in an assay that simultaneously determines INH and RIF resistance in Mycobacterium tuberculosis and identifies and determines the species of NTMs. The assay panel included 16 primer pairs in eight multiplexed reactions and was validated using a collection of 1,340 DNA samples from cultured specimens collected in the New York City area, the Republic of Georgia, and South Africa. Compared with phenotypic data, the PCR/ESI-MS assay had 89.3% sensitivity and 95.8% specificity in the determination of INH resistance and 96.3% sensitivity and 98.6% specificity in the determination of RIF resistance. Based on a set of 264 previously characterized liquid culture specimens, the PCR/ESI-MS method had 97.0% sensitivity and 99.9% specificity for determination of NTM identity. The assay also provides information on ethambutol, fluoroquinolone, and diarylquinoline resistance and lineage-specific polymorphisms, to yield highly discriminative digital signatures potentially suitable for epidemiology tracking.

• Assessment of Clostridium difficile infections by quantitative detection of tcdB toxin by use of a real-time cell analysis system.

  Authors: Alex B Ryder, Ying Huang, Haijing Li, Min Zheng, Xiaobo Wang, Charles W Stratton, Xiao Xu, Yi-Wei Tang

  Journal of clinical microbiology. 11/2010; 48(11):4129-34.

  We explored the use of a real-time cell analysis (RTCA) system for the assessment of Clostridium difficile toxins in human stool specimens by monitoring the dynamic responses of the HS27 cells toWe explored the use of a real-time cell analysis (RTCA) system for the assessment of Clostridium difficile toxins in human stool specimens by monitoring the dynamic responses of the HS27 cells to tcdB toxins. The C. difficile toxin caused cytotoxic effects on the cells, which resulted in a dose-dependent and time-dependent decrease in cell impedance. The RTCA assay possessed an analytical sensitivity of 0.2 ng/ml for C. difficile toxin B with no cross-reactions with other enterotoxins, nontoxigenic C. difficile, or other Clostridum species. Clinical validation was performed on 300 consecutively collected stool specimens from patients with suspected C. difficile infection (CDI). Each stool specimen was tested in parallel by a real-time PCR assay (PCR), a dual glutamate dehydrogenase and toxin A/B enzyme immunoassay (EIA), and the RTCA assay. In comparison to a reference standard in a combination of the three assays, the RTCA had a specificity of 99.6% and a sensitivity of 87.5% (28 of 32), which was higher than the EIA result (P = 0.005) but lower than the PCR result (P = 0.057). In addition, the RTCA assay allowed for quantification of toxin protein concentration in a given specimen. Among RTCA-positive specimens collected prior to treatment with metronidazole and/or vancomycin, a significant correlation between toxin protein concentrations and clinical CDI severities was observed (R(2) = 0.732, P = 0.0004). Toxin concentrations after treatment (0.89 ng/ml) were significantly lower than those prior to the treatment (15.68 ng/ml, Wilcoxon P = 0.01). The study demonstrates that the RTCA assay provides a functional tool for the potential assessment of C. difficile infections.

• Mass spectrometry biotyper system identifies enteric bacterial pathogens directly from colonies grown on selective stool culture media.

  Authors: Ying He, Haijing Li, Xuedong Lu, Charles W Stratton, Yi-Wei Tang

  Journal of clinical microbiology. 11/2010; 48(11):3888-92.

  We evaluated the performance and cost-effectiveness of a matrix-assisted laser desorption ionization time-of-flight mass spectrometry-based Biotyper system for the routine identification of commonWe evaluated the performance and cost-effectiveness of a matrix-assisted laser desorption ionization time-of-flight mass spectrometry-based Biotyper system for the routine identification of common enteric bacterial pathogens seen in middle Tennessee from suspicious colonies grown on selective stool culture media. A total of 304 suspicious colonies were selected and further identified from 605 stool specimens. The suspicious colonies were analyzed by the Biotyper system, and the results were compared to those from routine phenotypic methods, which identified 22 Salmonella species, 39 Shigella species, 3 enterohemorrhagic Escherichia coli (EHEC) isolates, 2 Yersinia enterocolitica isolates, 2 Campylobacter species, and 236 gastrointestinal normal flora isolates. The Biotyper system correctly identified the Salmonella species, Yersinia enterocolitica, and Campylobacter species but failed to distinguish the Shigella species and EHEC isolates from E. coli. Among the 236 normal flora isolates, 233 (98.7%) and 228 (96.6%) agreed at the genus and species levels, respectively, between the phenotypic and Biotyper methods. Organism identification scores were insignificantly different between colonies directly from selective media and subsequently from pure subculture. The entire Biotyper identification procedure, from smear preparation to final result reporting, can be completed within 30 min. The Biotyper system provides a rapid and simple screening tool for identifying many, but not all, suspicious colonies grown on selective media within 24 h after inoculation, which shortens test turnaround time by 2 to 3 days.

• Complex febrile seizures followed by complete recovery in an infant with high-titer 2009 pandemic influenza A (H1N1) virus infection.

  Authors: Mandy F O'Leary, James D Chappell, Charles W Stratton, Robert M Cronin, Mary B Taylor, Yi-Wei Tang

  Journal of clinical microbiology. 10/2010; 48(10):3803-5.

  We describe a 2009 H1N1 virus infection with a high viral load in a previously healthy infant who presented with complex febrile seizures and improved on oseltamivir without neurologic sequelae.We describe a 2009 H1N1 virus infection with a high viral load in a previously healthy infant who presented with complex febrile seizures and improved on oseltamivir without neurologic sequelae. Febrile seizures may be a complication in young children experiencing infection with high viral loads of 2009 H1N1 influenza virus.

• Mycobacterium tuberculosis Beijing lineage favors the spread of multidrug-resistant tuberculosis in the Republic of Georgia.

  Authors: Stefan Niemann, Roland Diel, George Khechinashvili, Medea Gegia, Nino Mdivani, Yi-Wei Tang

  Journal of clinical microbiology. 10/2010; 48(10):3544-50.

  High rates and transmission of multidrug-resistant (MDR) tuberculosis (TB) have been associated with the Mycobacterium tuberculosis complex (MTBC) Beijing lineage, pointing to the importance ofHigh rates and transmission of multidrug-resistant (MDR) tuberculosis (TB) have been associated with the Mycobacterium tuberculosis complex (MTBC) Beijing lineage, pointing to the importance of pathogen genetic factors for the modulation of infection outcome and epidemiology. We present here an in-depth analysis of the population structure of MTBC strains from the Republic of Georgia, a high-incidence setting at the Black Sea Coast. Phylogenetic lineages were identified based on 24-locus MIRU-VNTR (for mycobacterial interspersed repetitive unit-variable number tandem repeat) and spoligotyping analysis. Clusters of strains with identical genotyping profiles were determined as an indicator for the rate of recent transmission. Among the 183 M. tuberculosis isolates investigated, the most prominent lineage found was Beijing (26%), followed by the LAM (18%), Ural (12%), and Haarlem (5%) strains. A closely related previously undefined phylogenetic group (62 strains) showed a genotyping pattern similar to laboratory strain H37RV and was denominated as "Georgia-H37RV-like." Although isoniazid resistance was found among strains of different lineages, MDR TB was nearly completely restricted to Beijing strains (P < 0.0001). Approximately 50% of the isolates were grouped in clusters, indicating a high rate of recent transmission. Our data indicate that, in addition to the confirmation of the importance of Beijing genotype strains for the TB epidemiology in former Soviet Union countries, a high-population diversity with strains of the LAM, Ural, Haarlem, and a previously undefined lineage represents nearly two-thirds of the strains found in Georgia. Higher rates among previously treated and MDR TB patients point to a higher potential of lineage Beijing to escape therapy and develop MDR TB.