Biomedical subjects

David W Warnock

Publications and source records attributed to David W Warnock.

At least 19 recordsLinked to original sources

PubMed2006

Comparison of multilocus sequence typing and Ca3 fingerprinting for molecular subtyping epidemiologically-related clinical isolates of Candida albicans.

Southern hybridization with the complex probe Ca3 is a well established tool for molecular subtyping of Candida albicans. Multilocus sequence typing (MLST) is a DNA sequence-based subtyping method recently applied to C. albicans and shown to have a high degree of intraspecies discriminatory power. However, its utility for studying the molecular epidemiology of sequential isolates from recurrent disease has not been established. We compared Ca3 Southern hybridization and MLST using seven housekeeping genes (CaAAT1a, CaACC1, CaADP1, CaPMI, CaSYA1, CaVPS13, CaZWF1b) for their ability to discriminate among 37 C. albicans isolates from recurrent cases of oropharyngeal candidiasis (OPC) in ten HIV-positive patients from India and the US. Among the 37 isolates, MLST identified 23 distinct genotypes (index of diversity = 97%); Ca3 Southern hybridization identified 21 distinct genotypes (index of diversity = 95%). Both methods clustered isolates into seven genetically-related groups and, with one exception, isolates that were indistinguishable by MLST were indistinguishable or highly related by Ca3 Southern hybridization. These results demonstrate that MLST performs equally well or better compared to Ca3 Southern hybridization for defining genetic-relatedness of sequential C. albicans isolates from recurrent cases of OPC in HIV-positive patients.

Blotting, Southern↗

PubMed2006

Fungal diseases: an evolving public health challenge.

The emergence of new fungal pathogens and the resurgence of mycotic diseases that had previously been uncommon is a serious and growing public health problem. This review examines the factors involved in the emergence or re-emergence of several mycotic diseases, including coccidioidomycosis and cryptococcosis, over the past two decades. New approaches to prevention and control are also discussed.

Communicable Disease Control↗

PubMed2006

Candidemia in neonatal intensive care units: Barcelona, Spain.

BACKGROUND: Candida spp. are increasingly important hospital-acquired pathogens in neonatal intensive care units (NICU) and cause considerable mortality in preterm infants. Most studies have been limited to a single institution. The aim of this study was to determine the epidemiology of candidemia in all Barcelona NICUs. METHODS: We conducted prospective population-based surveillance for candidemia in Barcelona, Spain, during 2002-2003. This report focuses on the results from 5 participating hospitals with NICUs. RESULTS: We detected 24 cases, resulting in an annual incidence of 32.6 cases per 100,000 live births and 1.1 cases per 100 NICU discharges. Median gestational age was 27.5 weeks (range, 24-40.5), and there were 21 cases among very low birth weight infants. Among the 20 (83%) cases evaluated for the presence of end organ infection, endophthalmitis occurred in 2 cases, and endocarditis, meningitis and peritonitis occurred in 1 case each. Candida parapsilosis was the most frequent species isolated (67%). All isolates were fluconazole-susceptible. Crude mortality was 21%. CONCLUSIONS: The preponderance of C. parapsilosis candidemias observed in Barcelona NICUs is similar to reports from the literature. Morbidity and mortality associated with neonatal candidemia remain high.

Candida↗

PubMed2006

Evaluation of amphotericin B interpretive breakpoints for Candida bloodstream isolates by correlation with therapeutic outcome.

One hundred seven Candida bloodstream isolates (51 C. albicans, 24 C. glabrata, 13 C. parapsilosis, 13 C. tropicalis, 2 C. dubliniensis, 2 C. krusei, and 2 C. lusitaniae strains) from patients treated with amphotericin B alone underwent in vitro susceptibility testing against amphotericin B using five different methods. Fifty-four isolates were from patients who failed treatment, defined as death 7 to 14 days after the incident candidemia episode, having persistent fever of >or=5 days' duration after the date of the incident candidemia, or the recurrence of fever after two consecutive afebrile days while on antifungal treatment. MICs were determined by using the Clinical Laboratory Standards Institute (formally National Committee for Clinical Laboratory Standards) broth microdilution procedure with two media and by using Etest. Minimum fungicidal concentrations (MFCs) were also measured in two media. Broth microdilution tests with RPMI 1640 medium generated a restricted range of MICs (0.125 to 1 microg/ml); the corresponding MFC values ranged from 0.5 to 4 microg/ml. Broth microdilution tests with antibiotic medium 3 produced a broader distribution of MIC and MFC results (0.015 to 0.25 microg/ml and 0.06 to 2 microg/ml, respectively). Etest produced the widest distribution of MICs (0.094 to 2 microg/ml). However, none of the test formats studied generated results that significantly correlated with therapeutic success or failure.

Adolescent↗

PubMed2005

An epidemic of coccidioidomycosis in Arizona associated with climatic changes, 1998-2001.

BACKGROUND: Reports of coccidioidomycosis cases in Arizona have increased substantially. We investigated factors associated with the increase. METHODS: We analyzed the National Electronic Telecommunications System for Surveillance (NETSS) data from 1998 to 2001 and used Geographic Information Systems (GIS) to map high-incidence areas in Maricopa County. Poisson regression analysis was performed to assess the effect of climatic and environmental factors on the number of monthly cases; a model was developed and tested to predict outbreaks. RESULTS: The overall incidence in 2001 was 43 cases/100,000 population, a significant (P<.01, test for trend) increase from 1998 (33 cases/100,000 population); the highest age-specific rate was in persons > or =65 years old (79 cases/100,000 population in 2001). Analysis of NETSS data by season indicated high-incidence periods during the winter (November-February). GIS analysis showed that the highest-incidence areas were in the periphery of Phoenix. Multivariable Poisson regression modeling revealed that a combination of certain climatic and environmental factors were highly correlated with seasonal outbreaks (R2=0.75). CONCLUSIONS: Coccidioidomycosis in Arizona has increased. Its incidence is driven by seasonal outbreaks associated with environmental and climatic changes. Our study may allow public-health officials to predict seasonal outbreaks in Arizona and to alert the public and physicians early, so that appropriate preventive measures can be implemented.

Adult↗

PubMed2005

Rapid acquisition of stable azole resistance by Candida glabrata isolates obtained before the clinical introduction of fluconazole.

Five azole-susceptible Candida glabrata isolates obtained before 1975 became resistant to fluconazole, itraconazole, and voriconazole within 4 days of in vitro fluconazole exposure. This cross-resistance was stable for at least 4 months after removal of fluconazole and was associated with increased CgCDR1 and CgCDR2 expression.

ATP-Binding Cassette Transporters↗

PubMed2005

Epidemiology and predictors of mortality in cases of Candida bloodstream infection: results from population-based surveillance, barcelona, Spain, from 2002 to 2003.

We conducted population-based surveillance for Candida bloodstream infections in Spain to determine its incidence, the extent of antifungal resistance, and risk factors for mortality. A case was defined as the first positive blood culture for any Candida spp. in a resident of Barcelona, from 1 January 2002 to 31 December 2003. We defined early mortality as occurring between days 3 to 7 after candidemia and late mortality as occurring between days 8 to 30. We detected 345 cases of candidemia, for an average annual incidence of 4.3 cases/100,000 population, 0.53 cases/1,000 hospital discharges, and 0.73 cases/10,000 patient-days. Outpatients comprised 11% of the cases, and 89% had a central venous catheter (CVC) at diagnosis. Overall mortality was 44%. Candida albicans was the most frequent species (51% of cases), followed by Candida parapsilosis (23%), Candida tropicalis (10%), Candida glabrata (8%), Candida krusei (4%), and other species (3%). Twenty-four isolates (7%) had decreased susceptibility to fluconazole (MIC > or = 16 microg/ml). On multivariable analysis, early death was independently associated with hematological malignancy (odds ratio [OR], 3.5; 95% confidence interval [CI], 1.1 to 10.4). Treatment with antifungals (OR, 0.05; 95% CI, 0.01 to 0.2) and removal of CVCs (OR, 0.3; 95% CI, 0.1 to 0.9) were protective factors for early death. Receiving adequate treatment, defined as having CVCs removed and administration of an antifungal medication (OR, 0.2; 95% CI, 0.08 to 0.8), was associated with lower odds of late mortality; intubation (OR, 7.5; 95% CI, 2.6 to 21.1) was associated with higher odds. The incidence of candidemia and prevalence of fluconazole resistance are similar to other European countries, indicating that routine antifungal susceptibility testing is not warranted. Antifungal medication and catheter removal are critical in preventing mortality.

Adolescent↗

PubMed2005

Assessment of ribosomal large-subunit D1-D2, internal transcribed spacer 1, and internal transcribed spacer 2 regions as targets for molecular identification of medically important Aspergillus species.

Molecular approaches are now being developed to provide a more rapid and objective identification of fungi compared to traditional phenotypic methods. Ribosomal targets, especially the large-subunit RNA gene (D1-D2 region) and internal transcribed spacers 1 and 2 (ITS1 and ITS2 regions), have shown particular promise for the molecular identification of some fungi. We therefore conducted an assessment of these regions for the identification of 13 medically important Aspergillus species: Aspergillus candidus, Aspergillus (Eurotium) chevalieri, Aspergillus (Fennellia) flavipes, Aspergillus flavus, Aspergillus fumigatus, Aspergillus granulosus, Aspergillus (Emericella) nidulans, Aspergillus niger, Aspergillus restrictus, Aspergillus sydowii, Aspergillus terreus, Aspergillus ustus, and Aspergillus versicolor. The length of ribosomal regions could not be reliably used to differentiate among all Aspergillus species examined. DNA alignment and pairwise nucleotide comparisons demonstrated 91.9 to 99.6% interspecies sequence identities in the D1-D2 region, 57.4 to 98.1% in the ITS1 region, and 75.6 to 98.3% in the ITS2 region. Comparative analysis using GenBank reference data showed that 10 of the 13 species examined exhibited a < or = 1-nucleotide divergence in the D1-D2 region from closely related but different species. In contrast, only 5 of the species examined exhibited a < or = 1-nucleotide divergence from sibling species in their ITS1 or ITS2 sequences. Although the GenBank database currently lacks ITS sequence entries for some species, and major improvement in the quality and accuracy of GenBank entries is needed, current identification of medically important Aspergillus species using GenBank reference data seems more reliable using ITS query sequences than D1-D2 sequences, especially for the identification of closely related species.

Aspergillus↗

PubMed2004

In vitro susceptibilities of bloodstream isolates of Candida species to six antifungal agents: results from a population-based active surveillance programme, Barcelona, Spain, 2002-2003.

OBJECTIVES: The antifungal drug susceptibilities of 351 isolates of Candida species, obtained through active laboratory-based surveillance in the period January 2002-December 2003, were determined (Candida albicans 51%, Candida parapsilosis 23%, Candida tropicalis 10%, Candida glabrata 9%, Candida krusei 4%). METHODS: The MICs of amphotericin B, flucytosine, fluconazole, itraconazole, voriconazole and caspofungin were established by means of the broth microdilution reference procedure of the European Committee on Antibiotic Susceptibility Testing. RESULTS AND CONCLUSIONS: Amphotericin B and flucytosine were active in vitro against all strains. A total of 24 isolates (6.8%) showed decreased susceptibility to fluconazole (MIC > or = 16 mg/L) and 43 (12.3%) showed decreased susceptibility to itraconazole (MIC > or = 0.25 mg/L). Voriconazole and caspofungin were active in vitro against the majority of isolates, even those that were resistant to fluconazole.

Antifungal Agents↗

PubMed2004

A large histoplasmosis outbreak among high school students in Indiana, 2001.

BACKGROUND: A histoplasmosis outbreak occurred in an Indiana high school in November-December 2001. METHODS: To describe the risk factors for this outbreak, we conducted a cohort study of all available students and staff (N = 682) and an environmental investigation. RESULTS: Of the 523 (77%) persons who displayed serologic evidence of recent Histoplasma capsulatum infection, 355 (68%) developed symptoms consistent with acute pulmonary histoplasmosis. Rototilling of soil in a school courtyard known to be a bird roosting site had been performed during school hours on November 12, 2001, 14 days before both the peak of the onset of illness and a rise in student absenteeism. Being a student (odds ratio, 3.3; 95% confidence interval, 2.2-5.0) and being a student in a classroom near the courtyard during the rototilling (odds ratio, 3.1; 95% confidence interval, 1.8-5.2) were independently associated with infection and symptomatic illness. H. capsulatum was isolated from environmental samples, including soil from the courtyard and dust collected from a filter of a heating, ventilating and air-conditioning system. CONCLUSIONS: Soil-disrupting activities within a school courtyard caused the largest outbreak to date of histoplasmosis among adolescents. Improved efforts are needed to educate the community in endemic areas about histoplasmosis to prevent the occurrence of such outbreaks in the future. In addition, increased awareness among health care providers of this disease would facilitate appropriate diagnosis and treatment.

Adolescent↗

PubMed2004

Resolution of discrepant results for Candida species identification by using DNA probes.

Candida species bloodstream isolates were collected from institutions participating in an active, population-based surveillance for candidemia. Species identifications were performed locally and then confirmed at the Centers for Disease Control and Prevention (CDC) by phenotype-based methods. Discrepancies in species identification between the referring institution and the CDC were noted for 43 of 935 isolates (4.6%). A DNA probe-based species identification system (PCR-enzyme immunoassay [EIA]) was then used to resolve these discrepancies. The PCR-EIA result was identical to the CDC phenotypic identification method for 98% of the isolates tested. The most frequently misidentified species was Candida glabrata (37% of all discrepant identifications). Such misidentifications could lead to the administration of inappropriate therapy given the propensity of C. glabrata to develop resistance to azole antifungal drugs.

Base Sequence↗

PubMed2004

Incidence of bloodstream infections due to Candida species and in vitro susceptibilities of isolates collected from 1998 to 2000 in a population-based active surveillance program.

To determine the incidence of Candida bloodstream infections (BSI) and antifungal drug resistance, population-based active laboratory surveillance was conducted from October 1998 through September 2000 in two areas of the United States (Baltimore, Md., and the state of Connecticut; combined population, 4.7 million). A total of 1,143 cases were detected, for an average adjusted annual incidence of 10 per 100,000 population or 1.5 per 10,000 hospital days. In 28% of patients, Candida BSI developed prior to or on the day of admission; only 36% of patients were in an intensive care unit at the time of diagnosis. No fewer than 78% of patients had a central catheter in place at the time of diagnosis, and 50% had undergone surgery within the previous 3 months. Candida albicans comprised 45% of the isolates, followed by C. glabrata (24%), C. parapsilosis (13%), and C. tropicalis (12%). Only 1.2% of C. albicans isolates were resistant to fluconazole (MIC, > or = 64 microg/ml), compared to 7% of C. glabrata isolates and 6% of C. tropicalis isolates. Only 0.9% of C. albicans isolates were resistant to itraconazole (MIC, > or = 1 micro g/ml), compared to 19.5% of C. glabrata isolates and 6% of C. tropicalis isolates. Only 4.3% of C. albicans isolates were resistant to flucytosine (MIC, > or = 32 microg/ml), compared to < 1% of C. parapsilosis and C. tropicalis isolates and no C. glabrata isolates. As determined by E-test, the MICs of amphotericin B were > or = 0.38 microg/ml for 10% of Candida isolates, > or =1 microg/ml for 1.7% of isolates, and > or = 2 microg/ml for 0.4% of isolates. Our findings highlight changes in the epidemiology of Candida BSI in the 1990s and provide a basis upon which to conduct further studies of selected high-risk subpopulations.

Adolescent↗

PubMed2004

Rapid quantification of drug resistance gene expression in Candida albicans by reverse transcriptase LightCycler PCR and fluorescent probe hybridization.

We developed a rapid, sensitive, and reproducible assay to quantify Candida albicans ACT1, CDR1, CDR2, ERG11, and MDR1 mRNA using a two-step reverse transcription and LightCycler real-time PCR (RT-LightCycler PCR) method with sequence-specific hybridization probes. We compared RT-LightCycler PCR with Northern hybridization for quantitative analysis of gene expression in isolates with various fluconazole susceptibilities. Specificity of each LightCycler PCR was verified by LightCycler melting curve analysis and agarose gel electrophoresis of amplified products. Correlation of quantification results between RT-LightCycler PCR and Northern hybridization yielded correlation coefficients of > or = 0.91 for all genes except MDR1 (0.74). In this case, reduced correlation was due to the inability of Northern hybridization to accurately quantify the high MDR1 expression in a susceptible dose-dependent isolate which was shown by RT-LightCycler PCR to overexpress MDR1 >200-fold relative to the other isolates tested. In four isolates, low levels of CDR2 mRNA were detected by RT-LightCycler PCR but were undetectable by Northern hybridization. mRNA quantification by RT-LightCycler PCR correlates with Northern hybridization and offers additional advantages, including increased sensitivity and speed of analysis, along with lower RNA concentration requirements and an increased dynamic range of signal detection.

Base Sequence↗

PubMed2004

Interlaboratory comparison of results of susceptibility testing with caspofungin against Candida and Aspergillus species.

Seventeen laboratories participated in a study of interlaboratory reproducibility with caspofungin microdilution susceptibility testing against panels comprising 30 isolates of Candida spp. and 20 isolates of Aspergillus spp. The laboratories used materials supplied from a single source to determine the influence of growth medium (RPMI 1640 with or without glucose additions and antibiotic medium 3 [AM3]), the same incubation times (24 h and 48 h), and the same end point definition (partial or complete inhibition of growth) for the MIC of caspofungin. All tests were run in duplicate, and end points were determined both spectrophotometrically and visually. The results from almost all of the laboratories for quality control and reference Candida and Aspergillus isolates tested with fluconazole and itraconazole matched the NCCLS published values. However, considerable interlaboratory variability was seen in the results of the caspofungin tests. For Candida spp. the most consistent MIC data were generated with visual "prominent growth reduction" (MIC(2)) end points measured at 24 h in RPMI 1640, where 73.3% of results for the 30 isolates tested fell within a mode +/- one dilution range across all 17 laboratories. MIC(2) at 24 h in RPMI 1640 or AM3 also gave the best interlaboratory separation of Candida isolates of known high and low susceptibility to caspofungin. Reproducibility of MIC data was problematic for caspofungin tests with Aspergillus spp. under all conditions, but the minimal effective concentration end point, defined as the lowest caspofungin concentration yielding conspicuously aberrant hyphal growth, gave excellent reproducibility for data from 14 of the 17 participating laboratories.

Antifungal Agents↗

PubMed2004

Coccidioidomycosis among workers at an archeological site, northeastern Utah.

In 2001, an outbreak of acute respiratory disease occurred among persons working at a Native American archeological site at Dinosaur National Monument in northeastern Utah. Epidemiologic and environmental investigations were undertaken to determine the cause of the outbreak. A clinical case was defined by the presence of at least two of the following symptoms: self-reported fever, shortness of breath, or cough. Ten workers met the clinical case definition; 9 had serologic confirmation of coccidioidomycosis, and 8 were hospitalized. All 10 were present during sifting of dirt through screens on June 19; symptoms began 9-12 days later (median 10). Coccidioidomycosis also developed in a worker at the site in September 2001. A serosurvey among 40 other Dinosaur National Monument workers did not find serologic evidence of recent infection. This outbreak documents a new endemic focus of coccidioidomycosis, which extends northward its known geographic distribution in Utah by approximately 200 miles.

Adolescent↗

PubMed2004

Candida parapsilosis characterization in an outbreak setting.

Candida parapsilosis is an important non-albicans species which infects hospitalized patients. No studies have correlated outbreak infections of C. parapsilosis with multiple virulence factors. We used DNA fingerprinting to determine genetic variability among isolates from a C. parapsilosis outbreak and from our clinical database. We compared phenotypic markers of pathogenesis, including adherence, biofilm formation, and protein secretion (secretory aspartic protease [SAP] and phospholipase). Adherence was measured as colony counts on silicone elastomer disks immersed in agar. Biofilms formed on disks were quantified by dry weight. SAP expression was measured by hydrolysis of bovine albumin; a colorimetric assay was used to quantitate phospholipase. DNA fingerprinting indicated that the outbreak isolates were clonal and genetically distinct from our database. Biofilm expression by the outbreak clone was greater than that of sporadic isolates (p < or =0.0005). Adherence and protein secretion did not correlate with strain pathogenicity. These results suggest that biofilm production plays a role in C. parapsilosis outbreaks.

Aspartic Acid Endopeptidases↗

PubMed2004

Histoplasmosis associated with exploring a bat-inhabited cave in Costa Rica, 1998-1999.

Between October 1998 and April 1999, 51 persons belonging to two separate groups developed acute pulmonary histoplasmosis after visiting a cave in Costa Rica. The first group consisted of 61 children and 14 adults from San Jose, Costa Rica; 44 (72%) were diagnosed with acute histoplasmosis. The second group comprised 14 tourists from the United States and Canada; 9 (64%) were diagnosed with histoplasmosis. After a median incubation time of 14 days, the most common symptoms were headache, fever, cough, and myalgias. Risk factors for developing histoplasmosis included crawling (odds ratio [OR] = 17.5, 95% confidence interval [CI] = 2.3-802) and visiting one specific room (OR = 3.4, 95% CI = 1.0-12.3) in the cave. Washing hands (OR = 0.1, 95% CI = 0.01-0.6) after exiting the cave was associated with a decreased risk of developing histoplasmosis. Histoplasma capsulatum was isolated from bat guano collected from inside the cave. Persons who explore caves, whether for recreation or science, should be aware of the risk bat-inhabited caves pose for developing histoplasmosis, especially if they are immunocompromised in any way.

Adolescent↗

PubMed2003

Drug resistance genes and trailing growth in Candida albicans isolates.

OBJECTIVES: To investigate possible molecular mechanisms of azole resistance among fluconazole-susceptible bloodstream isolates of Candida albicans that displayed the trailing growth phenomenon, and to compare these isolates with bloodstream and mucosal isolates that showed reduced susceptibilities to fluconazole. METHODS: Twelve C. albicans isolates-seven trailing and five susceptible dose dependent (SDD) or resistant (R)-were screened for ERG11 mutations by DNA sequencing and quantification of ERG11, CDR1 and MDR1 expression by RT-PCR using the LightCycler high-speed PCR system. RESULTS: SDD and R isolates possessed more homozygous ERG11 mutations than did the trailing isolates. Two of these, V404I and V509M, have not been described previously and were found exclusively in fluconazole SDD and R isolates. Quantification of ERG11 expression revealed that both trailing and SDD and R isolates were capable of ERG11 up-regulation in response to fluconazole, although the SDD and R isolates showed maximal up-regulation at higher fluconazole concentrations. Quantification of CDR1 and MDR1 revealed that all isolates, regardless of in vitro fluconazole response, were capable of CDR1 and MDR1 up-regulation following fluconazole exposure. Furthermore, the SDD and R isolates expressed higher constitutive levels of CDR1 and MDR1 or CDR1, respectively, in the absence of drug compared with trailing isolates. CONCLUSIONS: Trailing isolates, although susceptible to fluconazole, express the same molecular mechanisms as SDD and R isolates following fluconazole exposure but regulate them differently.

Antifungal Agents↗