Evaluation of the Cepheid Xpert Clostridium difficile Epi assay for diagnosis of Clostridium difficile infection and typing of the NAP1 strain at a cancer hospital.

Clostridium difficile is the most common cause of health care-associated diarrhea. Accurate and rapid diagnosis is essential to improve patient outcome and prevent disease spread. We compared our two-step diagnostic algorithm, an enzyme immunoassay for glutamate dehydrogenase (GDH) followed by the cytotoxin neutralization test (CYT) with a turnaround time of 24 to 48 h, versus the Cepheid Xpert C. difficile Epi assay, a PCR-based assay with a turnaround time of <1 h. In the first phase of the study, only GDH-positive stool samples were tested by both CYT and Xpert PCR. Discordant results were resolved by toxigenic culture. In the second phase, all stool samples were tested by GDH and Xpert PCR. Only GDH-positive stools were further tested by CYT. Genotypic characterization of 45 Xpert PCR-positive stools was performed by sequencing of the tcdC gene and PCR ribotyping. In phase 1, the agreement between the GDH-CYT and the GDH-Xpert PCR was 72%. The sensitivities and specificities of GDH-CYT and GDH-Xpert PCR were 57% and 97% and 100% and 97%, respectively. In phase 2, the agreement between GDH-CYT and Xpert PCR alone was 95%. As in phase 1, sensitivity of the Xpert PCR was higher than that of the GDH-CYT. The correlation between PCR-ribotyping, sequencing, and Xpert PCR for detection of NAP1 strains was excellent (>90%). The excellent sensitivity and specificity and the rapid turnaround time of the Xpert PCR assay as well as its strain-typing capability make it an attractive option for diagnosis of C. difficile infection.

Use of sloppy molecular beacon probes for identification of mycobacterial species.

We report here the use of novel "sloppy" molecular beacon probes in homogeneous PCR screening assays in which thermal denaturation of the resulting probe-amplicon hybrids provides a characteristic set of amplicon melting temperature (T(m)) values that identify which species is present in a sample. Sloppy molecular beacons possess relatively long probe sequences, enabling them to form hybrids with amplicons from many different species despite the presence of mismatched base pairs. By using four sloppy molecular beacons, each possessing a different probe sequence and each labeled with a differently colored fluorophore, four different T(m) values can be determined simultaneously. We tested this technique with 27 different species of mycobacteria and found that each species generates a unique, highly reproducible signature that is unaffected by the initial bacterial DNA concentration. Utilizing this general paradigm, screening assays can be designed for the identification of a wide range of species.

Vancomycin-resistant enterococcus in pediatric oncology patients: An analysis of potential consequences of colonization and infection.

A retrospective analysis of 57 pediatric oncology patients with a baseline positive vancomycin-resistant enterococcus (VRE) culture who subsequently received chemotherapy and/or radiation therapy was performed. The incidence of subsequent VRE infection was calculated using a competing risk analysis accounting for death from non-VRE causes as a competing risk. Ten patients had subsequent VRE infection. The cumulative incidence of subsequent infection was 14% (7-27%, 95% confidence interval) at 1 year and 16% (9-29%, 95% confidence interval) at 2 years. None of the hypothesized risk factors appeared to differ between patients who developed a subsequent infection and those who did not.

Clinical characterization of human metapneumovirus infection among patients with cancer.

BACKGROUND: Human metapneumovirus is a recently discovered RNA virus that typically causes respiratory disease in children. It has been linked to severe lower airway disease in hematopoietic stem cell and solid-organ transplant recipients. hMPV infection in a large population of patients with underlying cancer and varying degrees of immunosuppression has not been reported. We sought to characterize hMPV infection in patients with cancer. METHODS: Review of all cases of hMPV infection from two seasons (2005-6 and 2006-7) detected by DFA and/or real-time PCR at MSKCC, a tertiary cancer center in New York City. RESULTS: Among MSKCC patients with cancer, 51 (2.7%) of 1899 patients were positive for hMPV, including 3.2% with hematologic neoplasm and 1.7% with solid tumors. More children (4.5%) were positive than adults (2.2%). PCR detected twice as many cases as DFA. Cough and fever were common complaints. The longest shedding period was 80 days. 40 patients received radiographic evaluation; of these, 22 showed abnormalities including patchy (11), ground glass (5), and interstitial infiltrates (4). CONCLUSIONS: hMPV causes a nonspecific respiratory illness and was found in more than 2% of all tested persons with cancer. PCR detected substantially more cases than DFA. Unlike previous reports, we observed no fatalities due to hMPV, including 22 HSCT recipients with the infection.

Isolation of avian paramyxovirus 1 from a patient with a lethal case of pneumonia.

An unknown virus was isolated from a lung biopsy sample and multiple other samples from a patient who developed a lethal case of pneumonia following a peripheral blood stem cell transplant. A random PCR-based molecular screening method was used to identify the infectious agent as avian paramyxovirus 1 (APMV-1; a group encompassing Newcastle disease virus), which is a highly contagious poultry pathogen that has only rarely been found in human infections. Immunohistochemical analysis confirmed the presence of APMV-1 antigen in sloughed alveolar cells in lung tissue from autopsy. Sequence from the human isolate showed that it was most closely related to virulent pigeon strains of APMV-1. This is the most completely documented case of a systemic human infection caused by APMV-1 and is the first report of an association between this virus and a fatal disease in a human.

Colonization, bloodstream infection, and mortality caused by vancomycin-resistant enterococcus early after allogeneic hematopoietic stem cell transplant.

Bloodstream infection caused by vancomycin-resistant enterococcus (VRE) is associated with very high mortality among allogeneic hematopoietic stem cell transplant (alloHSCT) recipients. However, it remains unclear whether VRE bloodstream infection directly causes mortality in the early posttransplant period or is simply a marker of poor outcome. To determine the risk factors for VRE bloodstream infection and its effect on outcome, we followed 92 patients screened for stool colonization by VRE upon admission for alloHSCT. Patient records were reviewed to determine outcomes, including mortality and microbiologic failure. Colonization by VRE was extremely common, occurring in 40.2% of patients. VRE bloodstream infection developed in 34.2% of colonized patients by day +35, compared to 1.8% without VRE colonization (P < .01). VRE bloodstream infection was associated with a significant decrement in survival and frequent microbiologic failure, despite treatment with linezolid and/or daptomycin. Five (35.7%) of 14 patients with VRE bloodstream infection had attributable mortality or contributing mortality from the infection. Strain typing by pulsed-field gel electrophoresis identified 9 different VRE strains among the 37 colonized patients and 5 patients with different strains recovered from the stool and the blood. In conclusion, stool screening effectively identified patients at extremely high risk for VRE bloodstream infection. The high mortality of VRE in the early posttransplant period supports the use of empiric antibiotics with activity against VRE during periods of fever and neutropenia in colonized patients.

Prevention of peritransplantation viridans streptococcal bacteremia with early vancomycin administration: a single-center observational cohort study.

BACKGROUND: Viridans streptococcal bacteremia (VSB) after allogeneic hematopoietic stem cell transplantation (HSCT) is associated with substantial mortality. Prevention of this serious complication is therefore a high priority. The objective of this study was to evaluate the effect of early vancomycin administration on rates and outcomes of VSB. METHODS: We analyzed the effect of early vancomycin on the incidence of VSB in a cohort of 430 consecutive HSCTs performed during the period of 1 January 1998 to 30 September 2002. The primary end point was time to diagnosis of VSB. Early vancomycin was defined as >or=2 doses of vancomycin between days -7 through +7 after HSCT or diagnosis of VSB, whichever occurred first. Risk factors for VSB were identified in univariate and multivariate Cox proportional hazard models. RESULTS: The incidence of VSB in the cohort was 7.4%. The incidence of VSB in patients who did not receive early vancomycin was 24.8%, compared with 0.3% in patients who did (P<.001). Additional risk factors were female sex, conditioning with total body irradiation, and diagnosis of chronic myelogenous leukemia. CONCLUSIONS: The attributable mortality rate for VSB in our cohort was 21%. Early vancomycin was associated with decreased VSB (hazard ratio, 0.02; 95% confidence interval, 0.003-0.19) after controlling for age, sex, underlying disease, and transplantation variables. The benefits of vancomycin prophylaxis for the prevention of VSB and associated mortality need to be evaluated in a prospective clinical trial.

Pythium insidiosum pleuropericarditis complicating pneumonia in a child with leukemia.

We describe a 12-year-old boy with acute myeloid leukemia who developed pleuropericarditis while he was neutropenic and was receiving intravenously administered antibiotic and antifungal therapy for pneumonia. A KOH preparation of the purulent material from an extensive diagnostic and therapeutic pleuropericardial drainage procedure revealed multiple irregularly septate hyphae, and cultures yielded the organism Pythium insidiosum. After completing a 12-month course of intravenously administered liposomal amphotericin B (AmBisome; Fujisawa Healthcare) and itraconazole, the patient remained alive, in clinical remission, and symptom free.