Gail L Woods

Publications of Gail L Woods

• Multisite Reproducibility of the Broth Microdilution Method for Susceptibility Testing of Nocardia species.

  Authors: Patricia S Conville, Barbara A Brown-Elliott, Richard J Wallace, Frank G Witebsky, Deloris Koziol, Geraldine S Hall, Scott B Killian, Cindy C Knapp, David Warshauer, Tam Van, Nancy L Wengenack, Sharon Deml, Gail L Woods

  Journal of clinical microbiology. 01/2012;

  Antimicrobial susceptibility testing (AST) of clinical isolates of Nocardia is recommended to detect resistance to commonly used antimicrobial agents; such testing is complicated by difficulties inAntimicrobial susceptibility testing (AST) of clinical isolates of Nocardia is recommended to detect resistance to commonly used antimicrobial agents; such testing is complicated by difficulties in inoculum preparation and test interpretation. In this study, six laboratories performed repetitive broth microdilution testing on single strains of Nocardia brasiliensis, Nocardia cyriacigeorgica, Nocardia farcinica, Nocardia nova and Nocardia wallacei. For each isolate, a total of 30 microdilution panels from three different lots were tested at most sites. The goal of the study was to determine the inter- and intra-laboratory reproducibility of susceptibility testing of this group of isolates. Acceptable agreement [> 90% agreement at (±) 1 dilution of the minimum inhibitory concentration (MIC) mode] was found for amikacin, ciprofloxacin, clarithromycin and moxifloxacin. After eliminating MIC values from single laboratories whose results showed the greatest deviation from those of the remaining laboratories, acceptable agreement was also found for amoxicillin/clavulanic acid, linezolid, minocycline and tobramycin. Results showed unsatisfactory reproducibility of broth microdilution testing of ceftriaxone with N. cyriacigeorgica and N. wallacei, tigecycline with N. brasiliensis and N. cyriacigeorgica and sulfonamides with N. farcinica and N. wallacei. N. nova ATCC BAA-2227 is proposed as a quality control organism for AST of Nocardia sp., and the use of a disk diffusion test for sulfisoxazole is proposed as a check of the adequacy of the inoculum and to confirm sulfonamide MIC results.

• Comparison of traditional phenotypic identification methods with partial 5' 16S rRNA gene sequencing for species-level identification of nonfermenting Gram-negative bacilli.

  Authors: Joann L Cloud, Dag Harmsen, Peter C Iwen, James J Dunn, Gerri Hall, Paul Rocco Lasala, Karen Hoggan, Deborah Wilson, Gail L Woods, Alexander Mellmann

  Journal of clinical microbiology. 02/2010; 48(4):1442-4.

  Correct identification of nonfermenting Gram-negative bacilli (NFB) is crucial for patient management. We compared phenotypic identifications of 96 clinical NFB isolates with identifications obtainedCorrect identification of nonfermenting Gram-negative bacilli (NFB) is crucial for patient management. We compared phenotypic identifications of 96 clinical NFB isolates with identifications obtained by 5' 16S rRNA gene sequencing. Sequencing identified 88 isolates (91.7%) with >99% similarity to a sequence from the assigned species; 61.5% of sequencing results were concordant with phenotypic results, indicating the usability of sequencing to identify NFB.

• Repetitive-sequence-based PCR using the DiversiLab system for identification of Aspergillus species.

  Authors: Dewey Hansen, Mimi Healy, Kristy Reece, Cheryl Smith, Gail L Woods

  Journal of clinical microbiology. 05/2008; 46(5):1835-9.

  Repetitive-sequence-based PCR (rep-PCR) using the DiversiLab system was investigated for identification of Aspergillus. Ninety-five clinical isolates, identified by conventional methods, and fiveRepetitive-sequence-based PCR (rep-PCR) using the DiversiLab system was investigated for identification of Aspergillus. Ninety-five clinical isolates, identified by conventional methods, and five ATCC strains were tested. Sequencing of the internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) was performed on 2 isolates with discrepant rep-PCR and conventional results and 15 randomly selected outlier isolates. One isolate not identified by sequencing was excluded from analysis. After resolving discrepancies, all but one A. glaucus strain had >or=85% similarity to one or more strains of the same Aspergillus species in the mold database, thereby providing accurate identification. No isolate was misidentified.

• Identification of Histoplasma capsulatum, Blastomyces dermatitidis, and Coccidioides species by repetitive-sequence-based PCR.

  Authors: June I Pounder, Dewey Hansen, Gail L Woods

  Journal of clinical microbiology. 09/2006; 44(8):2977-82.

  The performance of repetitive-sequence-based PCR (rep-PCR) using the DiversiLab system for identification of Coccidioides species, Blastomyces dermatitidis, and Histoplasma capsulatum was assessed byThe performance of repetitive-sequence-based PCR (rep-PCR) using the DiversiLab system for identification of Coccidioides species, Blastomyces dermatitidis, and Histoplasma capsulatum was assessed by comparing data obtained to colony morphology and microscopic characteristics and to nucleic acid probe results. DNA from cultures of 23 Coccidioides, 24 B. dermatitidis, 24 H. capsulatum, 3 Arthrographis, and 2 Malbranchea isolates was extracted using a microbial DNA isolation kit as recommended by Bacterial Barcodes, Inc. Rep-PCR and probe results agreed for 97.2% of the dimorphic fungi when > or =85% similarity was used as the criterion for identification. Two H. capsulatum isolates were not identified, but no isolates were misidentified. From 43 of those cultures (15 Coccidioides, 14 B. dermatitidis, 14 H. capsulatum, 3 Arthrographis, and 2 Malbranchea), DNA also was extracted using an IDI lysis kit, a simpler method. Rep-PCR and probe results agreed for 97.7% of the dimorphic fungi when a criterion of > or =90% similarity was used for identification. One H. capsulatum isolate could not be identified; no isolates were misidentified. Using > or =85% similarity for identification resulted in one misidentification. These data suggest that the DiversiLab system can be used to identify Coccidioides and B. dermatitidis and, possibly, H. capsulatum isolates.

• Multiplex single-color PCR with amplicon melting analysis for identification of Aspergillus species.

  Authors: Maria Erali, June I Pounder, Gail L Woods, Cathy A Petti, Carl T Wittwer

  Clinical chemistry. 08/2006; 52(7):1443-5.

• Mycobacterium arupense sp. nov., a non-chromogenic bacterium isolated from clinical specimens.

  Authors: Joann L Cloud, Jay J Meyer, June I Pounder, Kenneth C Jost, Amy Sweeney, Karen C Carroll, Gail L Woods

  International journal of systematic and evolutionary microbiology. 07/2006; 56(Pt 6):1413-8.

  Several Mycobacterium-like organisms related to the Mycobacterium terrae complex have been isolated from clinical samples. In the clinical microbiology laboratory, partial 16S rRNA gene sequencingSeveral Mycobacterium-like organisms related to the Mycobacterium terrae complex have been isolated from clinical samples. In the clinical microbiology laboratory, partial 16S rRNA gene sequencing (approximately the first 500 bp) rather than full 16S rRNA gene sequencing is often used to identify Mycobacterium species. Partial 16S rRNA gene sequence analysis revealed 100 % similarity between 65 clinical isolates and Mycobacterium sp. MCRO 6 (GenBank accession no. X93032). Even after sequencing the nearly full-length 16S rRNA gene, closest similarity was only 99.6 % to Mycobacterium nonchromogenicum ATCC 19530(T). Sequencing of the nearly full-length 16S rRNA gene, the 16S-23S internal transcribed spacer region and the hsp65 gene did not reveal genotypic identity with the type strains of M. nonchromogenicum, M. terrae or Mycobacterium triviale. Although sequence analysis suggested that these clinical isolates represented a novel species, mycolic acid analysis by HPLC failed to distinguish them from M. nonchromogenicum. Therefore, phenotypic analysis including growth characterization, antibiotic susceptibility testing and biochemical testing was performed. These strains from clinical samples should be recognized as representing a novel species of the genus Mycobacterium, for which the name Mycobacterium arupense sp. nov. is proposed. The type strain is AR30097(T) (=ATCC BAA-1242(T) = DSM 44942(T)).

• Clinical evaluation of repetitive sequence-based polymerase chain reaction using the Diversi-Lab System for strain typing of vancomycin-resistant enterococci.

  Authors: June I Pounder, Cheryl K Shutt, Barbara J Schaecher, Gail L Woods

  Diagnostic microbiology and infectious disease. 04/2006; 54(3):183-7.

  The reliability of the Diversi-Lab System, an automated method of microbial strain typing using repetitive sequence-based polymerase chain reaction (rep-PCR), was evaluated by comparing results withThe reliability of the Diversi-Lab System, an automated method of microbial strain typing using repetitive sequence-based polymerase chain reaction (rep-PCR), was evaluated by comparing results with those obtained by pulsed-field gel electrophoresis (PFGE). Ninety-five clinical isolates of vancomycin-resistant enterococci (VRE; 13 groups, 2-17 isolates per group) sent to Associated Regional and University Pathologists (ARUP) Laboratories for typing were tested by both methods. Rep-PCR and PFGE results were concordant for 83 isolates: all 32 isolates in 6 of the groups and 51 of the 63 isolates in the other 7 groups. Clustering of the remaining 12 isolates differed. With the Diversi-Lab System, analysis is objective, and results are available in 4 h, compared with a more subjective analysis and a 2- to 3-day turnaround time for PFGE. The Diversi-Lab System may be a viable alternative to PFGE for typing VRE in clinical reference laboratories.

• Comparison of real-time polymerase chain reaction using the Smart Cycler and the Gen-Probe amplified Mycobacterium tuberculosis direct test for detection of M. tuberculosis complex in clinical specimens.

  Authors: June I Pounder, Wade K Aldous, Gail L Woods

  Diagnostic microbiology and infectious disease. 04/2006; 54(3):217-22.

  The performance of a real-time polymerase chain reaction (PCR) assay using the Smart Cycler instrument and a minor groove binding MGB Eclipse probe (Epoch Biosciences, Bothell, WA) for identificationThe performance of a real-time polymerase chain reaction (PCR) assay using the Smart Cycler instrument and a minor groove binding MGB Eclipse probe (Epoch Biosciences, Bothell, WA) for identification of Mycobacterium tuberculosis complex in acid-fast bacillus smear-positive and smear-negative clinical specimens was assessed by comparing results to the Amplified M. tuberculosis Direct Test (MTD) and mycobacterial culture plus clinical diagnosis. After initial testing, the overall sensitivity, specificity, and positive and negative predictive values of PCR for the 172 specimens submitted for mycobacterial culture were 86.3%, 100%, 100%, and 94.5%, respectively. These same values for MTD were 98.0%, 99.2%, 98.0%, and 99.2%. For 83 additional specimens, only MTD and PCR were performed; 5 specimens were positive and 78 were negative by both tests. The sensitivity of the PCR assay was improved by using different primers and probes. The time to a result for real-time PCR, starting with a decontaminated sample, was less than 3 h compared with 5-6 h for the MTD.

• Sequence variant for internal transcribed spacer region of Mycobacterium abscessus.

  Authors: Haleina Neal, Joann L Cloud, June I Pounder, Sam R Page, Gail L Woods

  Journal of clinical microbiology. 01/2006; 43(12):6214.

• Use of the MGB Eclipse system and SmartCycler PCR for differentiation of Mycobacterium chelonae and M. abscessus.

  Authors: Joann L Cloud, Karen Hoggan, Evgeniy Belousov, Samuel Cohen, Barbara A Brown-Elliott, Linda Mann, Rebecca Wilson, Wade Aldous, Richard J Wallace, Gail L Woods

  Journal of clinical microbiology. 09/2005; 43(8):4205-7.

  Although accurate in the identification of Mycobacterium species, partial 16S rRNA gene sequencing does not distinguish Mycobacterium chelonae from M. abscessus. Thus, we designed a SmartCycler PCRAlthough accurate in the identification of Mycobacterium species, partial 16S rRNA gene sequencing does not distinguish Mycobacterium chelonae from M. abscessus. Thus, we designed a SmartCycler PCR assay targeting the 16S-to-23S internal transcribed spacer (ITS) region with use of MGB Eclipse probes to distinguish each species. Comparison with PCR-restriction enzyme analysis of a 441-bp fragment of the hsp65 gene resulted in 100% correlation with 25 isolates of M. chelonae and 25 isolates of M. abscessus. ITS PCR performed on 90 consecutive isolates identified by partial 16S rRNA gene sequencing (26 isolates of the M. chelonae-M. abscessus complex and 64 remaining isolates, including Mycobacterium species, Nocardia species, and other aerobic actinomycetes) showed 100% specificity and sensitivity. The ITS PCR assay is accurate and specific, easy to perform, and a good supplemental test when using partial 16S rRNA gene sequencing to identify M. chelonae and M. abscessus.

• Interpretive criteria for use of AccuProbe for identification of Mycobacterium avium complex directly from 7H9 broth cultures.

  Authors: Joann L Cloud, Karen C Carroll, Samuel Cohen, Clint M Anderson, Gail L Woods

  Journal of clinical microbiology. 07/2005; 43(7):3474-8.

  Rapid identification of Mycobacterium avium complex (MAC) is possible by use of AccuProbe (Gen-Probe, San Diego, Calif.). To evaluate the reliability of the MAC AccuProbe for testing 7H9 culturesRapid identification of Mycobacterium avium complex (MAC) is possible by use of AccuProbe (Gen-Probe, San Diego, Calif.). To evaluate the reliability of the MAC AccuProbe for testing 7H9 cultures inoculated with broth from MGIT cultures positive for acid-fast bacilli or growth on a solid medium, we compared probe results to results obtained by sequencing a portion of the 16S rRNA gene. Isolates were sequenced if the MAC probe result was <100,000 relative light units (RLU) or if it was > or =100,000 RLU and the colony morphology was not classic or there were two colony types. For the 1,389 cultures tested in phase 1, conducted to evaluate cutoff values for the MAC probe in testing of 7H9 cultures inoculated with broth from MGIT cultures, the sensitivity and specificity of the MAC AccuProbe were 97.7% and 88.8%, respectively, according to the manufacturer's interpretive criteria (> or =30,000 RLU is positive). If the cutoff for a positive result were 80,000 RLU, the specificity would be 100% and the sensitivity 92.3%. Of the 344 isolates in phase 2, which was conducted to confirm the 80,000-RLU cutoff for a positive result and therefore included only isolates with a MAC probe result of < or =100,000 RLU, 13 of 16 with results of > or =30,000 but <80,000 RLU were identified as mycobacteria other than MAC, including five Mycobacterium tuberculosis complex isolates. These data support the use of 80,000 RLU as the cutoff for a positive result in testing of 7H9 broth cultures with the MAC AccuProbe.

• Comparison of six methods of extracting Mycobacterium tuberculosis DNA from processed sputum for testing by quantitative real-time PCR.

  Authors: Wade K Aldous, June I Pounder, Joann L Cloud, Gail L Woods

  Journal of clinical microbiology. 06/2005; 43(5):2471-3.

  Six methods of extracting Mycobacterium tuberculosis DNA from sputum for testing by quantitative PCR were compared: Tris-EDTA (TE) buffer, PrepMan Ultra, 2% sodium dodecyl sulfate (SDS)-10% Triton XSix methods of extracting Mycobacterium tuberculosis DNA from sputum for testing by quantitative PCR were compared: Tris-EDTA (TE) buffer, PrepMan Ultra, 2% sodium dodecyl sulfate (SDS)-10% Triton X with and without sonication, Infectio Diagnostics, Inc. (IDI) lysing tubes, and QIAGEN QIAamp DNA mini kit; all included a 15-min boiling step. Pooled digested and decontaminated sputum was spiked with M. tuberculosis ATCC 27294. Each extraction method was repeated eight times. Quantitative PCR was performed on the Smart Cycler and Rotor-Gene 3000 using primers targeting an 83-bp fragment of IS6110. An minor grove binding Eclipse probe with a fluorescent label was used for detection. An internal control was included to detect amplification inhibition. The limit of detection of M. tuberculosis DNA was 0.5 fg with both instruments. Calculated DNA concentrations (picograms) extracted using IDI, PrepMan, QIAGEN, and TE were 42.8, 30.4, 28.2, and 7.4, respectively, when run on the Smart Cycler, and 51.7, 20.1, 14.9, and 8.6, respectively, when run on Rotor-Gene. All extractions using SDS/Triton X with or without sonication were inhibited. Of the extraction methods evaluated, IDI lysis tubes provided the greatest yield of mycobacterial DNA, and the procedure can be completed in less than 1 h versus 2.5-3 h for the QIAGEN extraction.

• Repetitive-sequence-PCR-based DNA fingerprinting using the Diversilab system for identification of commonly encountered dermatophytes.

  Authors: June I Pounder, Sheri Williams, Dewey Hansen, Mimi Healy, Kristy Reece, Gail L Woods

  Journal of clinical microbiology. 05/2005; 43(5):2141-7.

  The performance of repetitive-sequence-based PCR (rep-PCR) using the DiversiLab system for identification of dermatophytes commonly isolated in a clinical laboratory was assessed by comparing resultsThe performance of repetitive-sequence-based PCR (rep-PCR) using the DiversiLab system for identification of dermatophytes commonly isolated in a clinical laboratory was assessed by comparing results to those of conventional tests (colony morphology, microscopic examination of slide cultures, and, for suspected Trichophyton species, use of additional media). Sixty-one cultures were tested in phase 1, the feasibility portion of the study; 64 additional cultures were tested in phase 2, the validation portion conducted to assess reproducibility and confirm accuracy. Discrepancies were resolved by repeating rep-PCR and conventional tests and, in phase 2, sequencing the internal transcribed spacers. After initial testing of the cultures in phase 1 (excluding one contaminated culture), agreement between conventional tests and rep-PCR was 90% (54 of 60). Agreement was 98.3% after resolution of discrepancies, and in all but one case the initial rep-PCR result was correct. After initial testing of cultures in phase 2 (excluding one discarded and one contaminated culture), agreement between rep-PCR and conventional testing was 88.7% (55 of 62). After discrepancies were resolved, agreement was 100%. Initial rep-PCR results were correct, except for one Microsporum canis culture containing two colony variants, which could not be initially identified by rep-PCR. The performance of the DiversiLab system for identification of the dermatophytes commonly encountered in a clinical mycology laboratory-Trichophyton mentagrophytes, Trichophyton rubrum, Trichophyton tonsurans, and M. canis-was excellent. Moreover, the DiversiLab system is technically simple and provides results in < 24 h once a pure culture is available for testing, which is considerably more rapid than conventional identification tests.

• Clinical evaluation of the DiversiLab microbial typing system using repetitive-sequence-based PCR for characterization of Staphylococcus aureus strains.

  Authors: Cheryl K Shutt, June I Pounder, Sam R Page, Barbara J Schaecher, Gail L Woods

  Journal of clinical microbiology. 04/2005; 43(3):1187-92.

  The DiversiLab System, which includes microfluidics-based detection, reagent kits, and software for data processing and analysis, is an automated method using repetitive sequence-based PCR (rep-PCR)The DiversiLab System, which includes microfluidics-based detection, reagent kits, and software for data processing and analysis, is an automated method using repetitive sequence-based PCR (rep-PCR) for microbial strain typing. To assess the reliability of the DiversiLab System for strain characterization of Staphylococcus aureus, we tested clinical isolates sent to ARUP Laboratories for typing and compared results to those of pulsed field electrophoresis (PFGE) aided by the cluster analysis provided by BioNumerics software. spa typing was performed when the results of these two methods for an outbreak were not concordant. The study included 89 S. aureus isolates (65 mecA positive, 24 mecA negative) from 19 outbreaks (2 to 11 isolates/outbreak). The DiversiLab and PFGE-BioNumerics results were concordant for 15 of the 19 outbreaks. For the remaining four outbreaks, there was partial concordance between the two methods. spa typing results were the same as or more similar to rep-PCR results for three of those outbreaks and were more similar to PFGE results for one. With regard to performance, the DiversiLab system was considerably less labor intensive than PFGE and provided results in less than 24 h, compared with 2 to 3 days for PFGE. Additionally, the Web-based DiversiLab software provides standardized comparisons among isolates almost instantaneously and generates user-friendly, customized reports.

• Multisite reproducibility of results obtained by two broth dilution methods for susceptibility testing of Mycobacterium avium complex.

  Authors: Gail L Woods, Natalie Williams-Bouyer, Richard J Wallace, Barbara A Brown-Elliott, Frank G Witebsky, Patricia S Conville, Marianne Plaunt, Geraldine Hall, Priscilla Aralar, Clark Inderlied

  Journal of clinical microbiology. 02/2003; 41(2):627-31.

  A multicenter study was conducted to assess the interlaboratory reproducibility of susceptibility testing of Mycobacterium avium complex (MAC) by broth microdilution using two different mediaA multicenter study was conducted to assess the interlaboratory reproducibility of susceptibility testing of Mycobacterium avium complex (MAC) by broth microdilution using two different media (cation-adjusted Mueller-Hinton broth with 5% oleic acid-albumin-dextrose-catalase and 7H9 broth with casein) and by macrodilution using the BACTEC 460TB and 12B media at pH 6.8 and 7.3 to 7.4. Ten well-characterized strains of MAC (four macrolide susceptible, six macrolide resistant) were tested against clarithromycin and azithromycin (the latter only by BACTEC 460TB, pH 6.8). At each site, strains were tested against clarithromycin three times on each of three separate days (nine testing events per isolate) by using a common lot of microdilution trays and BACTEC 12B medium, pH 6.8; strains were tested once on three separate days against clarithromycin in 12B medium at pH 7.3 to 7.4 and against azithromycin. Agreement among MICs (i.e., mode +/- 1 twofold dilution) was 100% for all strains and both drugs when BACTEC 460TB was used, regardless of the pH of the medium, but varied when microdilution with either medium was used, particularly with susceptible strains. Agreement based on interpretive category, with NCCLS-recommended breakpoints, was 100% for all strains with the BACTEC 460TB method (both drugs and both pH values) and with microdilution using 7H9 broth. With microdilution and Mueller-Hinton broth, agreement by interpretive category was 100% for eight isolates and >90% for two; errors occurred only in laboratories where personnel had minimal experience with this technique. MAC susceptibility testing may be performed by broth macrodilution or microdilution at either pH, with NCCLS-recommended interpretive breakpoints. However, because visual interpretation of broth microdilution end points is subjective, it is more prone to reader error; therefore, this method requires greater expertise than the BACTEC 460TB. Both techniques require appropriate validation and continued documentation of proficiency.

• Differentiation of mycobacterium chelonae and M. abscessus using SmartCycler PCR and MGB eclipse probes

  Authors: Gail L Woods, Evgeniy Belousov

  This poster discusses how the use of partial 16S rDNA sequencing (first one-third of the gene) for the identification of Mycobacterium species from cultured isolates has become recognized as a veryThis poster discusses how the use of partial 16S rDNA sequencing (first one-third of the gene) for the identification of Mycobacterium species from cultured isolates has become recognized as a very accurate method for identification. The distinction between M. chelonae and M. abscessus, however, is not possible with partial 16S rDNA sequencing and therefore requires additional testing. Conventional tests that have been used for the distinction include growth on 5% sodium chloride and growth on citrate medium, both of which have been problematic with test inaccuracies and prolonged turnaround time. Restriction analysis of an amplified fragment of the hsp65 gene can distinguish M. chelonae from M. abscessus, but this requires gel analysis and interpretation of fragment sizes and is sometimes difficult because of the similarity of restriction patterns. In this study we developed a simple and rapid method using real-time PCR with the SmartCycler TM (Cepheid, Sunnyvale, CA) to distinguish M.

• Mycobacterium arupense sp. nov., a novel moderately growing nonchromogenic bacterium isolated from clinical specimens

  Authors: Gail L Woods, Karen C Carroll

  This author manuscript discusses how several isolates of Mycobacterium species related to the M. terrae complex have been isolated from clinical samples. In the clinical microbiology laboratory,This author manuscript discusses how several isolates of Mycobacterium species related to the M. terrae complex have been isolated from clinical samples. In the clinical microbiology laboratory, partial 16S rRNA gene sequencing (approximate first 500 base pairs) is often used to identify Mycobacterium species rather than full 16S rRNA gene sequencing. Partial 16S rRNA gene sequence analysis revealed 100% identity between 65 clinical isolates and Mycobacterium species MCRO 6 (GenBank accession no. X93032). Even after sequencing the nearly full 16S rRNA gene, the closest match to an existing type strain is only 99.6% similar to M. nonchromogenicum (ATCC 19530T). Sequencing of the nearly full 16S rRNA gene, the 16S-23S internal transcribed spacer region, and the hsp65 gene did not reveal genotypic identity with the type strains of M. nonchromogenicum, M. terrae, or M. triviale. manuscript