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Development of hydrolysis probe-based real-time PCR for identification of virulent gene targets of Burkholderia pseudomallei and B. mallei--a retrospective study on archival cases of service members with melioidosis and glanders.
Mil Med. 2012 Feb; 177(2):216-21.MM

Abstract

Burkholderia pseudomallei and B. mallei are two highly pathogenic bacteria responsible for melioidosis and glanders, respectively. Our laboratory developed hydrolysis probe-based real-time polymerase chain reaction assays targeting type three secretion system (TTS) and transposase family protein (TFP) of B. pseudomallei and B. malli, respectively. The assays were validated for target specificity, amplification sensitivity, and reproducibility. A bacterial DNA panel, composed of B. pseudomallei (13 strains), B. mallei (11 strains), Burkholderia species close neighbors (5 strains), and other bacterial species (17 strains), was prepared for specificity testing. Reference DNAs from B. pseudomallei and B. mallei bacterial cultures were used as controls for amplification, limit of detection, and reproducibility testing. The two TaqMan assays, Bp-TTS 1 and Bm-TFP, were optimized and applied in a retrospective study of archived cases from the Armed Forces Institute of Pathology. We tested 10 formalin-fixed paraffin-embedded blocks originally from autopsy specimens of patients who died of melioidosis or glanders during or after overseas tours in 1960s. Polymerase chain reaction results confirmed that DNA samples from formalin-fixed paraffin-embedded blocks of eight patients with melioidosis were positive for Bp-TTS 1 target and two patients with glanders were positive for Bm-TFP target.

Authors+Show Affiliations

Henry M. Jackson Foundation, 401 Professional Drive, Gaithersburg, MD 20879, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Validation Study

Language

eng

PubMed ID

22360070

Citation

Zhang, Binxue, et al. "Development of Hydrolysis Probe-based Real-time PCR for Identification of Virulent Gene Targets of Burkholderia Pseudomallei and B. Mallei--a Retrospective Study On Archival Cases of Service Members With Melioidosis and Glanders." Military Medicine, vol. 177, no. 2, 2012, pp. 216-21.
Zhang B, Wear DJ, Kim HS, et al. Development of hydrolysis probe-based real-time PCR for identification of virulent gene targets of Burkholderia pseudomallei and B. mallei--a retrospective study on archival cases of service members with melioidosis and glanders. Mil Med. 2012;177(2):216-21.
Zhang, B., Wear, D. J., Kim, H. S., Weina, P., Stojadinovic, A., & Izadjoo, M. (2012). Development of hydrolysis probe-based real-time PCR for identification of virulent gene targets of Burkholderia pseudomallei and B. mallei--a retrospective study on archival cases of service members with melioidosis and glanders. Military Medicine, 177(2), 216-21.
Zhang B, et al. Development of Hydrolysis Probe-based Real-time PCR for Identification of Virulent Gene Targets of Burkholderia Pseudomallei and B. Mallei--a Retrospective Study On Archival Cases of Service Members With Melioidosis and Glanders. Mil Med. 2012;177(2):216-21. PubMed PMID: 22360070.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Development of hydrolysis probe-based real-time PCR for identification of virulent gene targets of Burkholderia pseudomallei and B. mallei--a retrospective study on archival cases of service members with melioidosis and glanders. AU - Zhang,Binxue, AU - Wear,Douglas J, AU - Kim,H S, AU - Weina,Peter, AU - Stojadinovic,Alexander, AU - Izadjoo,Mina, PY - 2012/2/25/entrez PY - 2012/3/1/pubmed PY - 2012/4/6/medline SP - 216 EP - 21 JF - Military medicine JO - Mil Med VL - 177 IS - 2 N2 - Burkholderia pseudomallei and B. mallei are two highly pathogenic bacteria responsible for melioidosis and glanders, respectively. Our laboratory developed hydrolysis probe-based real-time polymerase chain reaction assays targeting type three secretion system (TTS) and transposase family protein (TFP) of B. pseudomallei and B. malli, respectively. The assays were validated for target specificity, amplification sensitivity, and reproducibility. A bacterial DNA panel, composed of B. pseudomallei (13 strains), B. mallei (11 strains), Burkholderia species close neighbors (5 strains), and other bacterial species (17 strains), was prepared for specificity testing. Reference DNAs from B. pseudomallei and B. mallei bacterial cultures were used as controls for amplification, limit of detection, and reproducibility testing. The two TaqMan assays, Bp-TTS 1 and Bm-TFP, were optimized and applied in a retrospective study of archived cases from the Armed Forces Institute of Pathology. We tested 10 formalin-fixed paraffin-embedded blocks originally from autopsy specimens of patients who died of melioidosis or glanders during or after overseas tours in 1960s. Polymerase chain reaction results confirmed that DNA samples from formalin-fixed paraffin-embedded blocks of eight patients with melioidosis were positive for Bp-TTS 1 target and two patients with glanders were positive for Bm-TFP target. SN - 0026-4075 UR - https://www.unboundmedicine.com/medline/citation/22360070/Development_of_hydrolysis_probe_based_real_time_PCR_for_identification_of_virulent_gene_targets_of_Burkholderia_pseudomallei_and_B__mallei__a_retrospective_study_on_archival_cases_of_service_members_with_melioidosis_and_glanders_ L2 - https://academic.oup.com/milmed/article-lookup/doi/10.7205/milmed-d-11-00232 DB - PRIME DP - Unbound Medicine ER -