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In JoVE (1)
Other Publications (7)
- Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America
- The Medical Journal of Australia
- Journal of Clinical Microbiology
- Journal of Clinical Microbiology
- Medical Mycology : Official Publication of the International Society for Human and Animal Mycology
- Journal of Clinical Microbiology
- The Journal of Infectious Diseases
Articles by Matthew V. N. O'Sullivan in JoVE
Multiplex PCR and Reverse Line Blot Hybridization Assay (mPCR/RLB)
Matthew V. N. O'Sullivan, Fei Zhou, Vitali Sintchenko, Fanrong Kong, Gwendolyn L. Gilbert
Centre for Infectious Diseases and Microbiology, University of Sydney
An inexpensive, high throughput method for simultaneous detection of up to 43 molecular targets is described. Applications of mPCR/RLB include microbial typing and detection of multiple pathogens from clinical samples.
Other articles by Matthew V. N. O'Sullivan on PubMed
Disseminated Pyogenic Mycoplasma Pneumoniae Infection in a Renal Transplant Recipient, Detected by Broad-range Polymerase Chain Reaction
Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America. Nov, 2004 | Pubmed ID: 15494902
Although extrapulmonary manifestations of Mycoplasma pneumoniae infection are generally immune-mediated, disseminated infections occasionally occur. We describe a renal transplant recipient who developed disseminated pyogenic M. pneumoniae infection that was detected by broad-range polymerase chain reaction and that manifested by prosthetic arterial graft infection, psoas abscess, and septic arthritis.
The Medical Journal of Australia. May, 2006 | Pubmed ID: 16646749
Influence of Disk Separation Distance on Accuracy of the Disk Approximation Test for Detection of Inducible Clindamycin Resistance in Staphylococcus Spp
Journal of Clinical Microbiology. Nov, 2006 | Pubmed ID: 17005747
We undertook this study to assess the accuracy of the clindamycin-erythromycin disk approximation test (D-test) for detection of inducible clindamycin resistance in Staphylococcus spp. One hundred sixty-three Staphylococcus aureus and 68 coagulase-negative Staphylococcus (CoNS) spp. which were erythromycin nonsusceptible but clindamycin susceptible were tested using the D-test performed at both 15-mm and 22-mm disk separations and compared with genotyping as the "gold standard." The rate of inducible clindamycin resistance was 96.3% for S. aureus and 33.8% for CoNS spp. The sensitivities of the D-tests performed at 15 mm and 22 mm were 100% and 87.7%, respectively, and specificities were 100% for both. The use of 22-mm disk separation for the D-test to detect inducible clindamycin resistance results in an unacceptably high very major error rate (12.3%). All isolates with false-negative results harbored the ermA gene, and the majority were methicillin-resistant Staphylococcus aureus. False-negative results were associated with smaller clindamycin zone sizes and double-edged zones. We recommend using a disk separation distance of =15 mm. There is wide geographic variation in the rates of inducible clindamycin resistance, and each laboratory should determine the local rate before deciding whether to either perform the D-test routinely or else report that all erythromycin-resistant S. aureus isolates are also clindamycin resistant.
Journal of Clinical Microbiology. Dec, 2007 | Pubmed ID: 17942656
We describe the development and validation of an agar dilution method for the detection of inducible clindamycin resistance by using 227 previously characterized erythromycin-resistant, clindamycin-susceptible Staphylococcus sp. isolates. Mueller-Hinton agar with defibrinated horse blood containing a range of erythromycin concentrations (1 to 8 mg/liter) combined with clindamycin at 0.5 mg/liter was used to determine the optimal concentration that produced growth of inducible isolates while inhibiting that of isolates without the inducible phenotype. A concentration of clindamycin of 0.5 mg/liter with erythromycin at 1 mg/liter was the optimal combination for detection of inducible resistance and resulted in a sensitivity of 100% (95% confidence interval [CI], 97.9 to 100) and a specificity of 100% (95% CI, 93.0 to 100). Attention must be paid to ensuring that a sufficient inoculum has been used, since an inoculum below the standard 10(7) bacteria/ml may result in false-negative results. This method has been incorporated into routine use in our laboratory.
Mannose-binding Lectin Deficiency Does Not Appear to Predispose to Cryptococcosis in Non-immunocompromised Patients
Medical Mycology : Official Publication of the International Society for Human and Animal Mycology. Jun, 2008 | Pubmed ID: 18415846
While most patients with cryptococcosis have obvious cellular immune deficiency, a minority have no apparent predisposing factors. However, in the latter there may be subtle innate immune system deficiencies which go unrecognized. Mannose-binding lectin (MBL) deficiency is associated with increased susceptibility to infectious diseases and may predispose to cryptococcosis, particularly when it disseminates to the central nervous system (CNS) in apparently immunocompetent patients. MBL function and levels, as well as MBL2 genotype were determined in 36 HIV-negative cryptococcosis patients (25 with CNS involvement) using C4 deposition and mannan-binding ELISA. MBL deficiency was defined using C4 deposition level < 0.2 U/microl or mannan-binding level < 0.5 microg/ml. MBL results were compared between patients with cryptococcosis and healthy controls and among the cryptococcosis patients according to the site of their disease. There was no difference in MBL function, mannan-binding level or increase in the frequency of MBL deficiency or low producing MBL2 genotypes in any of these comparisons. Patients with CNS cryptococcosis were no more likely to be MBL deficient than those with non-CNS disease. It appears that MBL deficiency is not associated with cryptococcosis in non-immunocompromised hosts. Beta errors consequent on the small number of patients studied may account for the lack of association.
Rapid Identification of Methicillin-resistant Staphylococcus Aureus Transmission in Hospitals by Use of Phage-derived Open Reading Frame Typing Enhanced by Multiplex PCR and Reverse Line Blot Assay
Journal of Clinical Microbiology. Aug, 2010 | Pubmed ID: 20519463
The relatively high-level clonality of methicillin-resistant Staphylococcus aureus (MRSA) and its frequent high-level endemicity in nosocomial settings complicate the development of methods for rapid subtyping of MRSA strains that are capable of identifying person-to-person transmission in hospitals. Phage-derived open reading frame (PDORF) typing is an MRSA typing method targeting mobile genetic elements that was recently described and evaluated using a geographically restricted set of isolates. The objective of this study was to develop a multiplex PCR-reverse line blot (mPCR/RLB) assay for PDORF typing and to test its applicability on a broad range of isolates and in an environment where MRSA is highly endemic. The 16 targets were identified using a 23-primer-pair mPCR/RLB assay with two probes for each target. The method was evaluated using 42 MRSA reference strains, including those representing major international clones, and 35 isolates from episodes of suspected nosocomial transmission. In vivo stability was explored using 81 isolate pairs. Pulsed-field gel electrophoresis (PFGE) and spa typing were performed for comparison. Among the 42 reference strains, there were 33 PFGE subtypes, 30 PDORF types, and 22 spa types. Simpson's index of diversity was 0.987, 0.971, and 0.926 for PFGE subtyping, PDORF typing, and spa typing, respectively. Typing of clinical isolates by PDORF typing and PFGE demonstrated concordant results. mPCR/RLB-based PDORF typing has similar discriminatory power to that of PFGE, can assist in tracking MRSA transmission events in a setting of high-level endemicity, and has the advantage of being a high-throughput technique.
Antibiotic Choice May Not Explain Poorer Outcomes in Patients with Staphylococcus Aureus Bacteremia and High Vancomycin Minimum Inhibitory Concentrations
The Journal of Infectious Diseases. Aug, 2011 | Pubmed ID: 21742831
There are concerns about reduced efficacy of vancomycin in patients with Staphylococcus aureus bacteremia (SAB), especially when the minimum inhibitory concentration (MIC) nears the upper limit of the susceptible range.