dClostridium difficile infection (CDI) causes nearly half a million cases of diarrhea and colitis in the United States each year. Although the importance of the gut microbiota in C. difficile pathogenesis is well recognized, components of the human gut flora critical for colonization resistance are not known. Culture-independent high-density Roche 454 pyrosequencing was used to survey the distal gut microbiota for 39 individuals with CDI, 36 subjects with C. difficile-negative nosocomial diarrhea (CDN), and 40 healthy control subjects. A total of 526,071 partial 16S rRNA sequence reads of the V1 to V3 regions were aligned with 16S databases, identifying 3,531 bacterial phylotypes from 115 fecal samples. Genomic analysis revealed significant alterations of organism lineages in both the CDI and CDN groups, which were accompanied by marked decreases in microbial diversity and species richness driven primarily by a paucity of phylotypes within the Firmicutes phylum. Normally abundant gut commensal organisms, including the Ruminococcaceae and Lachnospiraceae families and butyrate-producing C2 to C4 anaerobic fermenters, were significantly depleted in the CDI and CDN groups. These data demonstrate associations between the depletion of Ruminococcaceae, Lachnospiraceae, and butyrogenic bacteria in the gut microbiota and nosocomial diarrhea, including C. difficile infection. Mechanistic studies focusing on the functional roles of these organisms in diarrheal diseases and resistance against C. difficile colonization are warranted.

We review the experience at our institution with galactomannan (GM) testing of bronchoalveolar lavage (BAL) fluid in the diagnosis of invasive pulmonary aspergillosis (IPA) among solid-organ transplant recipients. Among 81 patients for whom BAL GM testing was ordered (heart, 24; kidney, 22; liver, 19; lung, 16), there were five cases of proven or probable IPA. All five patients had BAL GM of >2.1 and survived following antifungal therapy. The sensitivity, specificity, and positive and negative predictive values for BAL GM testing at a cutoff of >1.0 were 100%, 90.8%, 41.7%, and 100%, respectively. The sensitivity of BAL GM testing was better than that of conventional tests such as serum GM or BAL cytology and culture. Moreover, a positive BAL GM test diagnosed IPA several days to 4 weeks before other methods for three patients. Twelve patients had BAL GM of >0.5 but no evidence of IPA. Among these, lung transplant recipients accounted for 41.7% (5/12) of the false-positive results, reflecting frequent colonization of airways in this population. Excluding lung transplants, the specificity and positive predictive value for other solid-organ transplants increased to 92.9% and 62.5%, respectively (cutoff, >1.0). In conclusion, BAL GM testing facilitated more-rapid diagnoses of IPA and the institution of antifungal therapy among non-lung solid-organ transplant recipients and helped to rule out IPA.

We compared the FilmArray RP (Idaho Technology, Inc., Salt Lake City, UT) and the xTAG RVP (Luminex Corporation, Toronto, Canada) multiplex respiratory virus PCR methods for the detection of respiratory viruses in a set of 200 patient specimens frozen at ؊70°C after standard viral culture and antigen detection methods were done. Both systems detected between 40 to 50% more viruses than traditional methods, primarily rhinoviruses and human metapneumovirus. The FilmArray RP detected significantly more total viruses either alone or as part of mixed infections than the xTAG RVP, as well as an additional 21.6% more respiratory syncytial viruses. The xTAG RVP requires 5 to 6 h with 2.5 to 3 h of hands-on time, while the FilmArray RP takes about an hour with 3 to 5 min of hands-on time, making it much easier to perform.Multiplex reverse transcriptase respiratory virus PCR has been shown to be more sensitive than standard respiratory virus culture, direct fluorescent-antigen, and direct enzymelinked immunosorbent assay (ELISA) antigen detection methods (1, 2, 7-9, 13-14, 16, 20). Viral culture is labor-intensive, detects some viruses (e.g., rhinovirus and coronavirus) poorly, and requires 3 to 5 days to detect most agents. Consequently, results are generally not available early in the clinical decisionmaking process. Direct fluorescent-antibody assay (DFA) and chromatographic immunoassays are rapid enough to support real-time clinical decisions, but DFA is highly labor-intensive and chromatographic immunoassays are relatively insensitive. The FilmArray RP multiplex respiratory virus panel uses a pouch system that contains all reagents for the identification of 18 respiratory viruses and 3 bacterial respiratory pathogens in about 1 h after inoculation of a patient sample, obviating both labor and turnaround time (TAT) issues. We compared the performance of the FilmArray RP with that of the FDAcleared Luminex xTAG RVP multiplex panel by using 200 retrospective clinical respiratory virus culture samples.

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clinical microbiology laboratory in a 36-month period. A total of 46% of the cultures were positive; the most common organisms isolated were coagulase-negative staphylococci (36.4%), Pseudomonas aeruginosa (13.9%), enterococci (10.0%), yeasts (9.2%), Staphylococcus aureus (5.8%), and Enterobacter species (4.4%). The frequencies of positive blood cultures within 48 h prior to a positive catheter culture result were as follows: Candida albicans (68.4%), S. aureus (60%), Enterobacter cloacae (42.9%), Staphylococcus epidermidis (32.1%), P. aeruginosa (27.7%), and enterococci (23.3%). The sonication method allowed quantification of the number of CFU removed from a catheter for between 102 and 107 CFU. For catheter cultures in which :102 CFU grew, a linear regression equation could be calculated: (risk of positive blood culture for the same organism) = 14

Conventional methods for the identification of gastrointestinal pathogens are time-consuming and expensive and have limited sensitivity. The aim of this study was to determine the clinical impact of a comprehensive molecular test, the BioFire FilmArray gastrointestinal (GI) panel, which tests for many of the most common agents of infectious diarrhea in approximately 1 h. Patients with stool cultures submitted were tested on the GI panel ( = 241) and were compared with control patients ( = 594) from the year prior. The most common organisms detected by the GI panel were enteropathogenic (EPEC, = 21), norovirus ( = 21), rotavirus ( = 15), sapovirus ( = 9), and ( = 8). Patients tested on the GI panel had an average of 0.58 other infectious stool tests compared with 3.02 in the control group ( = 0.0001). The numbers of days on antibiotic(s) per patient were 1.73 in the cases and 2.12 in the controls ( = 0.06). Patients with the GI panel had 0.18 abdomen and/or pelvic imaging studies per patient compared with 0.39 ( = 0.0002) in the controls. The average length of time from stool culture collection to discharge was 3.4 days in the GI panel group versus 3.9 days in the controls ( = 0.04). The overall health care cost could have decreased by $293.61 per patient tested. The GI panel improved patient care by rapidly identifying a broad range of pathogens which may not have otherwise been detected, reducing the need for other diagnostic tests, reducing unnecessary use of antibiotics, and leading to a reduction in hospital length of stay.

We previously observed marked synergy between daptomycin and both rifampin and ampicillin against vancomycin-resistant enterococci (VRE). Because the synergy between daptomycin and ampicillin was observed for 100% of VRE strains with high-level ampicillin resistance (ampicillin MIC of >128 g/ml), we looked for synergy between daptomycin and other ␤-lactams against 18 strains of methicillin-resistant Staphylococcus aureus (MRSA) by employing a time-kill method using Mueller-Hinton broth supplemented to 50 mg of Ca 2؉ /liter. All strains were resistant to oxacillin (16 of 18 strains were resistant at drug concentrations of >256 g/ml), and all strains were susceptible to daptomycin (the MIC at which 90% of the tested isolates were inhibited was 1 g/ml). Daptomycin was tested at concentrations of 2, 1, 0.5, 0.25, 0.125, and 0.0625 g/ml alone or in combination with oxacillin at a fixed concentration of 32 g/ml. Synergy was found for all 18 strains with daptomycin at one-half the MIC in combination with 32 g of oxacillin/ml, and synergy was found for 11 of 18 strains (61%) with daptomycin at one-fourth the MIC or less in combination with oxacillin. At 24 h, the daptomycin-oxacillin combination with daptomycin at one-half the MIC showed bactericidal activity against all 18 strains, and the combination with one-fourth the daptomycin MIC showed bactericidal activity against 9 of 18 strains. We also used a novel screening method to look for synergy between daptomycin and other ␤-lactams. In this approach, daptomycin was incorporated into Ca Daptomycin is a novel lipopeptide antibiotic with bactericidal activity against a wide range of clinically important grampositive bacteria, including vancomycin-resistant enterococci (VRE) and methicillin-resistant Staphylococcus aureus (MRSA). In a recent study, we found synergy between daptomycin and both rifampin and ampicillin against VRE (12a). In that work, a screening technique was used to test daptomycin with 18 other antibiotics, and promising combinations were further tested by time-kill studies. In brief, daptomycin was added to Ca 2ϩ -supplemented Mueller-Hinton broth at subinhibitory concentrations, and the Etest-determined MIC of individual antibiotics on agar containing daptomycin was compared with the MIC on agar without daptomycin. When MRSA strains were studied with this method, combinations of daptomycin and ␤-lactams were found to warrant further study. In this report, we present comparative results for several daptomycin-␤-lactam combinations by the screening method and confirmative time-kill studies using combinations of daptomycin and oxacillin or ampicillin-sulbactam for 18 strains of MRSA. MATERIALS AND METHODSEighteen randomly selected strains of MRSA were obtained from the clinical microbiology laboratory at Shands Hospital at the University of Florida, Gainesville, between January and March 2002. All isolates were tested for relatedness by pulsed-field gel electrophoresis at the Mayo Medical Laboratories, Rochester, Minn. Five isolates were different from ea...

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