[Show abstract][Hide abstract] ABSTRACT: Accurate diagnosis is critical to providing appropriate care in infectious diseases (ID). New technologies for infectious disease diagnostics are emerging, but gaps remain in test development and availability. The Emerging Infections Network surveyed ID physicians to assess unmet diagnostic needs. Responses reflected the urgent need to identify drug-resistant infections and highlighted the potential for early diagnosis to improve antibiotic stewardship. Information gained from this survey can help inform recommendations for new diagnostic test development in the future.
[Show abstract][Hide abstract] ABSTRACT: Background: Meningitis is a severe infection that causes significant morbidity and mortality. Rapid etiologic diagnosis is critical to initiation of appropriate therapy. A molecular system to rapidly identify bacteria, viruses, and fungi causing meningitis could improve the medical management of this infection in children.
Methods: The FilmArray® Meningitis / Encephalitis Panel (FA ME; BioFire Diagnostics, Inc., Salt Lake City, UT) performs automated nucleic acid purification and multiplex PCR to identify 17 bacterial, viral and fungal pathogens directly from CSF. We performed a retrospective study of 120 children < = 18 presenting to our children’s hospital with concern for meningitis. Conventional CSF testing (culture and/or viral testing) was ordered at the discretion of the treating physician. FA studies on archived CSF, using a research use only version of the panel, were performed at the University of Utah. FA ME results were compared to conventional testing.
Results: The median age of children in the study was 6 months (range 0-237 months). 18 children had positive CSF cultures, of which 11 grew true pathogens. Nine were included on the FA ME panel. FA detected 8/9 cultured panel bacteria, and also detected bacteria in 6 additional specimens (see Table). The median CSF WBC count in children with positive cultures was 391; median WBC for children with positive FA ME testing was 1749. This difference is related to low CSF WBC in samples culture-positive for presumed contaminants. Only 4 children had viruses detected by conventional methods, including PCR of blood or CSF. FA ME detected viruses in 16 samples, including 4 also positive for bacteria.
Conclusion: The FilmArray ME panel is a sensitive tool for the rapid identification of pathogens from CSF and may identify causative pathogens not detected by conventional testing. Improved detection of pathogens from CSF using FilmArray has the potential to improve treatment and outcomes for children with meningitis.
Identified by Conventional Testing (n)
Identified by FilmArray (n)
K. pneumoniae/C. koseri
Presumed contaminants (skin flora)
IDWeek 2014 Meeting of the Infectious Diseases Society of America; 10/2014
[Show abstract][Hide abstract] ABSTRACT: Background
Levofloxacin is routinely used for the prevention of invasive bacterial infections during autologous peripheral blood stem cell transplantation (APBSCT). However, increasing rates of bacterial sepsis were noted at our institution among multiple myeloma (MM) patients undergoing outpatient APBSCT with melphalan-based chemotherapy and levofloxacin prophylaxis. We assessed the impact of a change in antibacterial prophylaxis from oral levofloxacin (Period 1) to sequential oral levofloxacin followed by ertapenem (Period 2).Methods
Electronic medical records were reviewed to identify MM patients who underwent APBSCT in the outpatient clinic between October 2007 and April 2012.ResultsOver a 4.5-year period, 165 outpatient APBSCTs were eligible for the analysis. Fewer overall bacteremias occurred during Period 2 as compared with Period 1 (0.5 cases per 100 person-days vs. 2.4 cases per 100 person-days, P < 0.001). In addition, fewer patients were hospitalized for neutropenic fever while receiving sequential prophylaxis (45.7% vs. 75.7% of outpatient APBSCT recipients during Periods 2 and 1, respectively; P < 0.001). In Kaplan–Meier analysis, receipt of sequential prophylaxis (Period 2) was significantly associated with overall bacteremia-free survival within 30 days after the APBSCT (P < 0.001). No significant differences were seen in the number of patients developing Clostridium difficile infection or ertapenem-resistant gram-negative bacteremia between study periods.Conclusion
In conclusion, sequential prophylaxis may effectively prevent episodes of bacteremia and hospitalizations in neutropenic MM outpatient APBSCT recipients. Prospective studies that involve larger numbers of MM patients with extended periods of follow-up are ultimately required to define the safety and efficacy of sequential antibacterial prophylaxis.
[Show abstract][Hide abstract] ABSTRACT: Paecilomyces species are emerging fungal pathogens. Morphological identifications are complicated by similarities among the members of
the P. variotii complex as well as to some Rasamsonia and Hamigera species. The purpose of this study was to compare matrix-assisted laser desorption ionization–time-of-flight mass spectrometry
(MALDI–TOF MS) with molecular diagnostic standards (i.e., multilocus DNA sequencing of the internal transcribed spacer regions
1 and 2, D1/D2 regions, and part of the β-tubulin gene) for the identification of Paecilomyces spp. encountered in two clinical mycology laboratories. A total of 77 clinical isolates identified morphologically as P. variotii (n = 21), P. lilacinus (n = 52), and Paecilomyces spp. not otherwise specified (n = 4) were included. In accord with the most recent taxonomy, all P. lilacinus isolates were confirmed as Purpureocillium lilacinum by both sequencing and MALDI–TOF MS. Fungi phenotypically resembling P. variotii or Paecilomyces spp. were identified by molecular techniques as P. variotii sensu stricto (n = 12), P. formosus (n = 3), P. dactylethromorphus (n = 3), Rasamsonia argillacea (n = 4), or R. piperina (n = 1) and at the genus level as an isolate of a Hamigera sp. and a Paecilomyces sp. There was 92.2% (71/77) agreement between the molecular and proteomic methods only after supplementation of the MALDI–TOF
MS database with type strains. Paecilomyces variotii–like organisms required multilocus DNA interrogations for differentiation and account for all of the fungi whose identification
was missed by MALDI–TOF MS. Overall, MALDI–TOF MS was a rapid and reliable alternative to multilocus sequencing. However,
significant augmentation of the commercially available database was required to reproducibly identify this group of important
Medical mycology: official publication of the International Society for Human and Animal Mycology 03/2014; 52(5). DOI:10.1093/mmy/myu001 · 2.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The erm (41) gene causes inducible macrolide resistance in Mycobacterium abscessus but not M. chelonae. Erm (41) sequencing of 285 M. abscessus and 45 M. chelonae isolates was compared to 14 day susceptibility; agreement was 98.9% and 100%, respectively. Extended incubation may not be necessary for M. chelonae and erm (41) genotype is a useful adjunct for M. abscessus.
[Show abstract][Hide abstract] ABSTRACT: Reference broth microdilution methods of Candida echinocandin susceptibility testing are limited by interlaboratory variability in caspofungin MICs. Recently revised Clinical
and Laboratory Standards Institute (CLSI) breakpoint MICs for echinocandin nonsusceptibility may not be valid for commercial
tests employed in hospital laboratories. Indeed, there are limited echinocandin susceptibility testing data from hospital
laboratories. We conducted a multicenter retrospective study of 9 U.S., Australian, and New Zealand hospitals that routinely
tested Candida bloodstream isolates for echinocandin susceptibility from 2005 to 2013. Eight hospitals used Sensititre YeastOne assays.
The Candida spp. were C. albicans (n = 1,067), C. glabrata (n = 911), C. parapsilosis (n = 476), C. tropicalis (n = 185), C. krusei (n = 104), and others (n = 154). Resistance and intermediate rates were ≤1.4% and ≤3%, respectively, for each echinocandin against C. albicans, C. parapsilosis, and C. tropicalis. Resistance rates among C. glabrata and C. krusei isolates were ≤7.5% and ≤5.6%, respectively. Caspofungin intermediate rates among C. glabrata and C. krusei isolates were 17.8% and 46.5%, respectively, compared to ≤4.3% and ≤4.4% for other echinocandins. Using CLSI breakpoints,
18% and 19% of C. glabrata isolates were anidulafungin susceptible/caspofungin nonsusceptible and micafungin susceptible/caspofungin nonsusceptible,
respectively; similar discrepancies were observed for 38% and 39% of C. krusei isolates. If only YeastOne data were considered, interhospital modal MIC variability was low (within 2 doubling dilutions
for each agent). In conclusion, YeastOne assays employed in hospitals may reduce the interlaboratory variability in caspofungin
MICs against Candida species that are observed between reference laboratories using CLSI broth microdilution methods. The significance of classifying
isolates as caspofungin intermediate and anidulafungin/micafungin susceptible will require clarification in future studies.
[Show abstract][Hide abstract] ABSTRACT: In this IDSA policy paper, we review the current diagnostic landscape, including unmet needs and emerging technologies, and assess the challenges to the development and clinical integration of improved tests. To fulfill the promise of emerging diagnostics, IDSA presents recommendations that address a host of identified barriers. Achieving these goals will require the engagement and coordination of a number of stakeholders, including Congress, funding and regulatory bodies, public health agencies, the diagnostics industry, healthcare systems, professional societies, and individual clinicians.
[Show abstract][Hide abstract] ABSTRACT: Background: Broad-range amplification and sequencing of conserved housekeeping genes provides a culture-independent method to detect infectious pathogens in clinical specimens. The Emerging Infections Network (EIN) surveyed ID physicians to assess use of this novel technology.
Methods: 1572 EIN members were surveyed in 03/13. Respondents who reported having performed broad-range PCR were asked about frequencies of submitted specimen types, positive results and their clinical usefulness.
Results: Of the 700 (44.5%) respondents to the survey, 297 (42%) had used broad-range PCR. The most common reason for not using these tests was lack of availability (76%), followed by a lack of knowledge about the test (28%). 201 respondents answered questions about their use of broad-range PCR. 60/201 (30%) had used it more than 10 times; the majority (50%) had used it 1-5 times. The most commonly submitted specimens were osteoarticular, CSF, and endovascular samples, including blood, each submitted by more than 50% of respondents. Most specimens were submitted in the setting of inflammation on histopathology with negative pathogen stains and culture. A majority of respondents (65%) could submit specimens with no laboratory utilization review. Most respondents reported only rare (36%) to occasional (38%) positive results. 89% of respondents who had used broad-range PCR more than 10 times and 80% of respondents who used it less than 10 times reported test results to be helpful (not significant). Contaminant results were reported by similar proportions of respondents regardless of how frequently the test was ordered.
Conclusion: Increasing the use of broad-range PCR for diagnosis of suspected infections will depend on increased availability and awareness of the test as well as increased specificity and decreased frequency of contamination. Positive results need to be interpreted with caution due to risk of contamination. Studies that help physicians correlate test results with clinical decision-making and treatment strategies can help develop guidelines for use of this test.
IDWeek 2013 Meeting of the Infectious Diseases Society of America; 10/2013
[Show abstract][Hide abstract] ABSTRACT: Cytomegalovirus (CMV) infection is one of the most important infectious complications of transplantation. Monitoring CMV-specific CD8 T cell immunity is useful for predicting active CMV infection and for directing targeted antiviral therapy. In this study, we examined four basic parameters for validation of CMV-specific tetramer staining and peptide stimulation assays that cover five most frequent HLA class I alleles. We also examined the potential use of CMV-specific CD8(+) T cell numbers and functional and cytolytic responses in two autologous HSCT recipients treated for multiple myeloma. The coefficient of variation (CV %) of the precision within assays was 3.1-24% for HLA-tetramer staining, 2.5-47% for IFN-γ, and 3.4-59.7% for CD107a/b production upon peptide stimulation. The precision between assays was 5-26% for tetramer staining, 4-24% for IFN-γ, and 5-48% for CD107a/b. The limit of detection was 0.1-0.23 cells/μL of blood for tetramer staining, 0-0.23 cell/μL for IFN-γ, and 0.11-0.98 cells/μL for CD107a/b. The assays were linear and specific. The reference interval with 95% confidence level was 0-18 cells/μL for tetramer staining, 0-2 cells/μL for IFN-γ, and 0-3 cells/μL for CD107a/b. Our results provide acceptable measures of test performance for CMV immune competence assays for the characterization of CD8(+) T cell responses posttransplant measured in the absolute cell count per μL of blood.
[Show abstract][Hide abstract] ABSTRACT: Cryptococcosis is a systemic infection caused by the pathogenic yeasts Cryptococcus neoformans and C. gattii. Detection of cryptococcal capsular antigen (CrAg) in serum and cerebrospinal fluid (CSF) plays an important diagnostic role. We prospectively compared the new IMMY lateral flow assay (LFA) and enzyme immunoassay (EIA) to our current CrAg test (Premier EIA, Meridian Bioscience Inc.). Discordant samples were retested with the Latex-Cryptococcus antigen test (IMMY) and using serotype-specific monoclonal antibodies (mAbs). A total of 589 serum and 411 CSF specimens were tested in parallel. Qualitative agreement across assays was 97.7%. In all, 56 (41 serum and 15 CSF) were positive and 921 (527 serum and 394 CSF) were negative by all three assays. The 23 discrepant specimens were all Meridian EIA negative. Of 23 discordant specimens, 20 (87.0%) were positive by both the IMMY LFA and EIA, 2 were LFA positive only and 1 EIA positive only. Eleven discrepant specimens had adequate volume for latex agglutination (LA) testing; 8 were LA positive and 3 LA negative. LA negative samples (2 CSF and 1 serum) had low IMMY LFA/EIA titers (≤ 1:10). Serotype-specific mAb analysis of the LA positive samples suggested that these specimens contain CrAg epitopes similar to those of serotype C strains. In conclusion, the IMMY assays showed excellent overall concordance with the Meridian EIA. Assay performance differences were related to issues of analytic sensitivity and possible serotype bias. Incomplete access to patient-level data combined with low specimen volumes limited our ability to fully resolve discrepant results.
[Show abstract][Hide abstract] ABSTRACT: Invasive candidiasis (IC) is a devastating disease. While prompt antifungal therapy improves outcomes, empiric treatment based on the presence of fever has little clinical impact. Β-D-Glucan (BDG) is a fungal cell wall component detectable in the serum of patients with early invasive fungal infection (IFI). We evaluated the utility of BDG surveillance as a guide for preemptive antifungal therapy in at-risk intensive care unit (ICU) patients.
Patients admitted to the ICU for ≥ 3 days and expected to require at least 2 additional days of intensive care were enrolled. Subjects were randomized in 3:1 fashion to receive twice weekly BDG surveillance with preemptive anidulafungin in response to a positive test or empiric antifungal treatment based on physician preference.
Sixty-four subjects were enrolled, with 1 proven and 5 probable cases of IC identified over a 2.5 year period. BDG levels were higher in subjects with proven/probable IC as compared to those without an IFI (117 pg/ml vs. 28 pg/ml; p<0.001). Optimal assay performance required 2 sequential BDG determinations of ≥ 80 pg/ml to define a positive test (sensitivity 100%, specificity 75%, positive predictive value 30%, negative predictive value 100%). In all, 21 preemptive and 5 empiric subjects received systemic antifungal therapy. Receipt of preemptive antifungal treatment had a significant effect on BDG concentrations (p< 0.001). Preemptive anidulafungin was safe and generally well tolerated with excellent outcome.
BDG monitoring may be useful for identifying ICU patients at highest risk to develop an IFI as well as for monitoring treatment response. Preemptive strategies based on fungal biomarkers warrant further study.
Clinical Trials.gov NCT00672841.
PLoS ONE 08/2012; 7(8):e42282. DOI:10.1371/journal.pone.0042282 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cytomegalovirus (CMV) is an important pathogen after allogeneic transplantation. However, few studies have examined CMV reactivation after autologous peripheral blood stem cell transplantation (APBSCT) to treat multiple myeloma (MM), especially in the setting of the newer chemotherapeutic agents and/or 2 sequential APBSCTs (ie, tandem transplantation). A retrospective chart review of patients with MM who underwent either single APBSCT or tandem transplantation was conducted to evaluate the incidence, risk factors, and outcomes of CMV infection at a single institution. A total of 104 patients with MM underwent transplantation during the study period, including 66 patients who received tandem transplantation. The majority of patients (66 of 104; 63.5%) were CMV-seropositive, and CMV viremia was frequently detected in this subgroup (32 of 66; 48.5%). No primary CMV infections were identified. CMV reactivation was more common in recipients of tandem transplantation than in recipients of single APBSCT (P < .001). In addition, patients who developed CMV viremia were more likely to have received conditioning therapy with melphalan, bortezomib, dexamethasone, and thalidomide compared with those without CMV reactivation (P = .015). However, on multiple logistic regression analysis, only receipt of tandem transplantation was significantly associated with CMV reactivation (odds ratio, 5.112; 95% confidence interval, 1.27-20.60; P = .022). Febrile episodes of CMV viremia were observed in 17 patients (17 of 32; 53.1%), and invasive CMV disease was diagnosed in 1 patient. Our data suggest that CMV reactivation after APBSCT for MM is relatively common, and that viremia is often associated with fever. CMV surveillance should be considered, especially when tandem transplantation is performed using combination chemotherapy with high-dose melphalan.
Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 06/2012; 18(11):1753-8. DOI:10.1016/j.bbmt.2012.06.008 · 3.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Leptotrichia spp. are anaerobic, pencil-shaped, Gram-negative rods that are part of the normal oral and intestinal human flora. Although
not typically considered pathogenic, invasive Leptotrichia infections have been reported in immunosuppressed patients. A perceived rise in the identification of Leptotrichia spp. at our institution prompted a retrospective evaluation of these infections. Laboratory and clinical records were reviewed
to identify Leptotrichia culture-positive patients. Over a 5-year period, 68 Leptotrichia-positive specimens were identified. Of these, 21% (14/68) were identified in original samples submitted from 13 different
patients at our institution, and the remainder (79% [54/68]) were unknown isolates referred from outside hospitals for molecular
identification. All in-house Leptotrichia were identified from blood cultures. Only 64% (9/14) of these grew on solid media, and 5 were a part of polymicrobial bacteremias
containing other enteric pathogens. All local patients were receiving chemotherapy and a majority received hematopoietic stem
cell transplant (HSCT) (11/13). All had neutropenic fever with symptoms of mucositis and/or enteritis. Most of the HSCT patients
(73% [8/11]) were autologous recipients hospitalized after recent high-dose chemotherapy for multiple myeloma. L. hongkongensis, a novel species, was found in the majority of myeloma cases (63% [5/8]). In conclusion, we suggest that Leptotrichia spp. may be an underappreciated cause of bacteremia, particularly in multiple myeloma patients receiving cytotoxic chemotherapy
for autologous HSCT. In our cohort, these infections were associated with neutropenic fever from an enteric source, and most
isolates remained sensitive to standard antibiotics.
[Show abstract][Hide abstract] ABSTRACT: Rapid and accurate identification of clinically important yeasts is essential given their inherent differences in antifungal susceptibility. We implemented nucleic acid sequencing for those species that could not be identified by phenotypic methods. Internal Transcribed Spacer region 1 and 2 (ITS1 and ITS2) sequences were investigated using SmartGene IDNS software, an rDNA sequence database and analysis program for microbial identification (ID). Over a 2.5-year period, 2,938 specimens were evaluated. Most (94%) isolates were fully identified by conventional methods, with Candida species accounting for the majority of them. Of the 169 organisms that required molecular analysis, 79% were identified to species level, 19% to genus and 2% remained unresolved. Sequenced isolates encompassed 33 unique species of which approximately half (52%) were common pathogens with atypical biochemical profiles and the remainder were rarer yeast species. A significant proportion (33%) of sequenced organisms displayed elevated MICs to fluconazole. Our experience supports the use of molecular techniques as an adjunct to conventional methods for the identification of medically important yeasts. Susceptibility testing alone may provide valuable treatment information in situations where phenotypic assessments are inconclusive and molecular or proteomic testing is not readily available.
Medical mycology: official publication of the International Society for Human and Animal Mycology 11/2011; 50(5):458-66. DOI:10.3109/13693786.2011.630683 · 2.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Since the introduction of the Haemophilus influenzae type b vaccine, the incidence of invasive H. influenzae type b disease among children has fallen dramatically, but the effect on invasive H. influenzae disease among adults may be more complex. In this population-based study we examined the epidemiology and outcomes of invasive disease caused by typeable and nontypeable H. influenzae among Utah adults during 1998-2008. The overall incidence increased over the study period from 0.14/100,000 person-years in 1998 to 1.61/100,000 person-years in 2008. The average incidence in persons >65 years old was 2.74/100,000 person-years, accounting for 51% of cases and 67% of deaths. The incidence was highest for nontypeable H. influenzae (0.23/100,000 person-years), followed by H. influenzae type f (0.14/100,000 person-years). The case-fatality rate was 22%. The incidence of invasive H. influenzae in Utah adults appears to be increasing. Invasive H. influenzae infection disproportionately affected the elderly and was associated with a high mortality rate.
[Show abstract][Hide abstract] ABSTRACT: First-line treatment of influenza A 2009 H1N1 relies on neuraminidase inhibitors such as oseltamivir. Resistance conferred by the H275Y neuraminidase gene mutation is concerning and likely to increase.
To characterize oseltamivir resistance in a hospital-based patient population.
All available respiratory specimens positive for influenza A by direct fluorescent antibody, RT-PCR, or culture from patients at the University of Utah 5/09-12/09 were collected. Specimens were confirmed as 2009 H1N1 by the Utah Department of Health. RT-PCR and pyrosequencing were used to test for the H275Y mutation (CDC protocol). PyroMark Q24 AQ software (Qiagen, Valencia, CA, USA) was used to allow for quantitative H275Y mutation analysis. Medical records of patients with resistant virus were reviewed.
We tested 191 influenza A virus-positive samples from 187 unique patients. Fifty (27%) patients were hospitalized. Four patient specimens (2.1%) were found to carry the H275Y mutation. Three patients were hospitalized, representing 6% of inpatient samples tested. Three patients had undergone hematopoietic stem cell transplant in the past year. Two patients died. Their influenza viruses were confirmed to be oseltamivir-resistant at an independent reference laboratory and through the Center for Disease Control and Prevention (CDC). One patient reported no history of prior oseltamivir exposure.
Widespread oseltamivir resistance among 2009 H1N1 remains a potential threat. Rapid techniques, such as pyrosequencing, which has the additional benefit of identifying mixed mutant populations of virus, may play a key role in identifying at-risk individuals and potentially unsuspected cases. Targeted surveillance of immunocompromised patients will be critical toward improving future influenza planning and therapy.
Influenza and Other Respiratory Viruses 07/2010; 4(4):199-204. DOI:10.1111/j.1750-2659.2010.00144.x · 2.20 Impact Factor