Central venous catheter needleless connectors (NCs) have been shown to develop microbial contamination.A protocol was developed for the collection, processing, and examination of NCs to detect and measure biofilms on these devices. Sixty-three percent of 24 NCs collected from a bone marrow transplant center contained biofilms comprised primarily of coagulase-negative staphylococci.Intravenous (i.v.) access lines (6, 7) and needleless connectors (NCs) (3, 4) have been demonstrated to be a risk factor for blood stream infection (BSI). Patients who require long-term i.v. access, such as bone marrow transplant patients, are at even greater risk for BSI. To deliver i.v. fluids (e.g., medication, blood products, or nutrients), tubing must be connected to i.v. catheters that enter the patient's bloodstream. Until recently, such connections have been made using beveled, hollow-bore needles that pierce an elastic membrane on a catheter end cap. Because of the potential for needle-stick injuries and health care worker exposure to bloodborne pathogens, many institutions have recently adopted the use of NCs. Though safer for health care workers, the potential for NCs to increase BSI risk to patients has been documented in outbreaks of nosocomial BSI (3,4). In October 1998, the Centers for Disease Control and Prevention (CDC) was asked to investigate a BSI outbreak at a bone marrow transplant center in which NCs were involved. As part of this investigation, CDC assessed the ability of NCs to harbor biofilms that could act as a reservoir for BSI pathogens. It is well established that biofilms may develop on intravascular devices, including central venous catheters (CVCs) (1,2,8). Though contamination of NCs by various organisms has been observed (3, 4), the occurrence of biofilms on these devices has, to our knowledge, not been documented. The objectives of this study were (i) to develop a standardized protocol that could be used to collect, ship, and process NCs for biofilm contamination and (ii) to determine whether biofilms could develop on these devices and what organisms were the primary colonizers. Hickman NCs were collected from patients with long-term CVCs in a single bone marrow transplant center in which an outbreak of BSIs had occurred.Collection and shipment of NCs. Female-female luer couplings (no. 06359-42; Cole Parmer, Niles, Ill.) were autoclaved and then used to connect two 5-ml syringes. One of the syringes contained 5-ml of phosphate-buffered saline (PBS; pH 7.2; Life Technologies, Grand Island, N.Y.). Syringe pairs were placed into zip-lock bags and shipped on ice packs to the bone marrow transplant center for the collection of NCs. By using an aseptic technique, the NCs were removed from the patient's CVC and placed into an unused sterile Petri dish and transported to the laboratory. After the two syringes were separated, the luer coupling remained on one syringe. The smaller end of the NC was then wiped with a sterile alcohol pledget and connected to the syringe without the luer coupling. The other end o...

.7% of all samples tested, and no significant differences in sensitivity or specificity were noted between HSCT and non-HSCT patients. IAs, while not as sensitive as direct fecal CBA, produce reasonable predictive values, especially when both antigen and toxin are detected. They also offer significant advantages over CBA in terms of turnaround time and ease of use.

Objective: Not much is known about the zoonotic transmission of methicillin-resistant Staphylococcus aureus (MRSA) in companion animals in the United States. We report the rate of prevalence of S. aureus and MRSA recovered from clinical samples of animals requiring treatment at veterinary clinics throughout the upper midwestern and northeastern United States. Design:We compared phenotypes, genotypes, and virulence profiles of the MRSA isolates identified in companion animals, such as cats, dogs, horses, and pigs, with typical human nosocomial and community-associated MRSA (CA-MRSA) genotypes to assess implied zoonotic transmission or zooanthroponosis. Five hundred thirty-three coagulase-positive staphylococci (CPS) isolates recovered between 2006 and 2008 from a variety of animal-source samples were screened for S. aureus by S. aureus-specific 16S rDNA primers and were screened for methicillin-resistance. All MRSA isolates were genotyped by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and spa typing. They were also screened for common staphylococcal enterotoxin and adhesion genes by multiplex and singleplex PCR.Results: Among the 533 CPS isolates recovered, 66 (12.4%) were determined to be S. aureus and 24 (4.5%) were MRSA. The percent of animals that were positive for S. aureus were as follows: 6.6% (32 of 487) dogs, 39.6% (19 of 48) cats, 83.3% (10 of 12) horses, and 100% of pigs, rabbits, hamsters and rats. Notably, 36.4% of all S. aureus identified were MRSA. Methicillin-resistant S. aureus was present in clinical samples from 12 of 487 dogs (2.5%), 6 of 48 cats (12.5%), 5 of 12 horses (42%), and 1 of 2 pigs (50%). The 24 MRSA isolates resolved into 4 PFGE clones: USA100 (50%), USA300 (16.7%), USA500 (20.8%) and USA800 (12.5%) and 6 sequence types (ST5, ST8, ST105, ST830, and ST986) or 2 clonal complexes, CC5 and CC8. Five major virulence profiles (clusters A to E) were observed in these MRSA isolates. Genotypic and virulence profiles of cats and dogs were more similar to each other than to those of horses. A Panton-Valentine leukocidin positive isolate with ST8:USA300 background was identified in a pig causing skin and soft infection. Conclusion:The presence of human MRSA clones in these animals suggests possible reverse zoonotic transmission. This study reports the first case of a USA300 genotype in a pig. Presence of multiple virulence profiles within a MRSA genotype in these animals suggests the potential of emergence of new MRSA clones by gaining or losing additional virulence genes.

e Enteric illness affects millions of individuals annually in the United States and results in >50,000 hospitalizations. The rapid and accurate identification of bacterial pathogens associated with gastroenteritis can aid acute patient management decisions, including the use of antibiotic therapy and infection control. This study compared the ProGastro SSCS multiplex real-time PCR assay (Gen-Probe Prodesse, San Diego, CA) to culture for the identification of Campylobacter spp. (Campylobacter jejuni and Campylobacter coli), Salmonella spp., and Shigella spp. and to broth enrichment followed by an FDA-cleared enzyme immunoassay (EIA) for the identification of Shiga toxin-producing Escherichia coli (STEC) isolates in stool specimens. Stool samples submitted in preservatives for routine culture and EIA were prospectively enrolled and tested at four clinical centers. Discrepancies between the ProGastro SSCS assay and culture or EIA were resolved using bidirectional sequencing. The overall prevalence of the pathogens as detected by culture was 5.6% (1.8% Campylobacter, 1.8% Salmonella, 1.3% Shigella, and 0.8% STEC). When results based on the ProGastro SSCS assay and bidirectional sequencing were applied, the overall prevalence increased to 8.3% (2.3% Campylobacter, 2.6% Salmonella, 1.8% Shigella, and 1.6% STEC). Following resolution of the discrepant results, the sensitivity of the ProGastro SSCS assay was 100% for all pathogens, and the specificities ranged from 99.4% to 100%. The sensitivity of culture compared to sequence-confirmed ProGastro SSCS results ranged from 52.9% to 76.9%, with the specificities ranging from 99.9% to 100%. Overall, these results suggest that the ProGastro SSCS assay is highly sensitive and specific in a clinical setting.

cBlastomyces spp. antigen testing was evaluated over a 10-year period in an area where blastomycosis is endemic. Antigen testing was less sensitive than previously reported, but serial urine testing was useful in monitoring disease resolution or progression. Culture and cytopathology remain the gold standard for diagnosis and exclusion of this infection. Blastomycosis is a serious and potentially fatal fungal infection. Blastomycosis may be asymptomatic or it can present as isolated pulmonary disease, disseminated disease (pulmonary and extrapulmonary infection), or extrapulmonary disease with manifestations in the central nervous system, bone, skin, or other locations (1-11).Culture and cytopathology are the gold standard for the diagnosis of blastomycosis. However, a variety of other diagnostic tests, including antigen testing, antibody testing, and PCR, are commercially available (8,(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). Given that identification from culture may not be evident for 2 to 4 weeks and that it often requires invasive procedures to obtain specimens, there is great interest in using antigen enzyme immunoassays (EIA) for rapid, noninvasive diagnosis and monitoring of disease progression or resolution (2,4,8,19).We evaluated the use of Blastomyces urine, serum, and bronchoalveolar lavage (BAL) fluid antigen assays for the diagnosis of blastomycosis and the effects of treatment on the clearance of antigenuria at the Marshfield Clinic from 1995 to 2015. Marshfield Clinic is located in Wisconsin where blastomycosis is endemic (1,3,6,9,(23)(24)(25)(26)(27)(28). Research protocols were approved by the Marshfield Clinic Institutional Review Board. Waiver of informed consent was obtained.Patients were included if blastomycosis was confirmed by culture or cytopathology and if urine, serum, or BAL fluid antigen EIA was completed at the time of diagnosis or within 30 days of starting antifungal medication. Blastomycosis was confirmed by culture or cytopathology at Marshfield Labs using conventional techniques. Commercially available blastomycosis antigen EIA was performed at the MiraVista Diagnostics reference laboratory at the time of specimen collection using qualitative EIA prior to March 2011 or quantitative EIA thereafter. EIA results are available within 24 h of sample submission (13,14,16). All specimens were obtained as part of routine clinical evaluations. Retrospective chart review was completed for all patients.Data were abstracted into Excel 2010, and statistical analysis was completed using SAS 9.3. Categorical data were compared using the 2 test or the Fisher exact test. Continuous variables were compared using Wilcoxon rank sum, Kruskal-Wallis, or analysis of variance (ANOVA). Correlation for serial urine antigen testing was measured using Spearman's coefficient. Significance was defined as a P of Ͻ0.05.Patients with quantitative antigen EIA results were reported as negative, positive below the limit of quantification (Ͻ0.2 ng/ml), positive and quantifiable (0.2 to 14.7 ng/ml), or positiv...