While the ResPlex II and MultiCode-PLx offer a broader virus detection range and greater ease of use, the xTAG PRV showed increased sensitivity to common viral targets represented in the assays, and also had the ability to differentiate human from non-human influenza A H1.

Clostridium difficile is the most important cause of nosocomial diarrhea. Several laboratory techniques are available to detect C. difficile toxins or the genes that encode them in fecal samples. We evaluated the Xpert C. difficile and Xpert C. difficile/Epi (Cepheid, CA) that detect the toxin B gene (tcdB) and tcdB, cdt, and a deletion in tcdC associated with the 027/NAP1/BI strain, respectively, by real-time PCR, and the Illumigene C. difficile (Meridian Bioscience, Inc.) that detects the toxin A gene (tcdA) by loop-mediated isothermal amplification in stool specimens. Toxigenic culture was used as the reference method for discrepant stool specimens. Two hundred prospective and fifty retrospective diarrheal stool specimens were tested simultaneously by the cell cytotoxin neutralization assay (CCNA) and the Xpert C. difficile, Xpert C. difficile/Epi, and Illumigene C. difficile assays. Of the 200 prospective stools tested, 10.5% (n ‫؍‬ 23) were determined to be positive by CCNA, 17.5% (n ‫؍‬ 35) were determined to be positive by Illumigene C. difficile, and 21.5% (n ‫؍‬ 43) were determined to be positive by Xpert C. difficile and Xpert C. difficile/ Epi. Of the 50 retrospective stools, previously determined to be positive by CCNA, 94% (n ‫؍‬ 47) were determined to be positive by Illumigene C. difficile and 100% (n ‫؍‬ 50) were determined to be positive by Xpert C. difficile and Xpert C. difficile/Epi. Of the 11 discrepant results (i.e., negative by Illumigene C. difficile but positive by Xpert C. difficile and Xpert C. difficile/Epi), all were determined to be positive by the toxigenic culture. A total of 21% of the isolates were presumptively identified by the Xpert C. difficile/Epi as the 027/NAP1/BI strain. The Xpert C. difficile and Xpert C. difficile/Epi assays were the most sensitive, rapid, and easy-to use assays for the detection of toxigenic C. difficile in stool specimens.

Sepsis-related laminitis (SRL) is a common complication in the septic/endotoxemic critically-ill equine patient, in which lamellar injury and failure commonly lead to crippling distal displacement of the distal phalanx. Similar to organ injury in human sepsis, lamellar injury in SRL has been associated with inflammatory events, including the influx of leukocytes into the lamellar tissue and markedly increased expression of a wide array of inflammatory mediators at the onset of Obel grade 1 (OG1) laminitis. The only treatment reported both clinically and experimentally to protect the lamellae in SRL, local hypothermia ("cryotherapy"), has been demonstrated to effectively inhibit lamellar expression of multiple inflammatory mediators when initiated at the time of administration of a carbohydrate overload in experimental models of SRL. However, the effect of hypothermia on leukocyte influx into affected tissue has not been assessed. We hypothesized that cryotherapy inhibits leukocyte emigration into the digital lamellae in SRL. Immunohistochemical staining using leukocyte markers MAC387 (marker of neutrophils, activated monocytes) and CD163 (monocyte/macrophage-specific marker) was performed on archived lamellar tissue samples from an experimental model of SRL in which one forelimb was maintained at ambient temperature (AMB) and one forelimb was immersed in ice water (ICE) immediately following enteral oligofructose administration (10g/kg, n=14 horses). Lamellae were harvested at 24h post-oligofructose administration (DEV, n=7) or at the onset of OG1 laminitis (OG1, n=7). Both MAC387-positive and CD163-positive cells were counted by a single blinded investigator on images [n=10 (40× fields/digit for MAC387 and 20x fields/digit for CD163)] obtained using Aperio microscopy imaging analysis software. Data were assessed for normality and analyzed with a paired t-test and one-way ANOVA with significance set at p<0.05. MAC387-positive cells were present in low numbers in the lamellar tissue and were decreased in the hypothermic limbs (vs. AMB limbs, p<0.05) in the OG1 group; no change in CD163-positive cell numbers was noted across the conditions of the model. This study demonstrated that hypothermia of the distal limbs instituted early in the disease process in the horse at risk of SRL significantly attenuates the increase of MAC387-positive leukocytes in the digital lamellae, but has minimal effect on increases in lamellar concentrations of the major leukocyte cell type present in that tissue, CD163-positive mononuclear cells.

hThe Verigene Clostridium difficile Nucleic Acid test (Verigene CDF test) (Nanosphere, Northbrook, IL) is a multiplex qualitative PCR assay that utilizes a nanoparticle-based array hybridization method to detect C. difficile tcdA and tcdB in fecal specimens. In addition, the assay detects binary toxin gene sequences and the single base pair deletion at nucleotide 117 (⌬ 117) in tcdC to provide a presumptive identification of the epidemic strain 027/NAP1/BI (referred to here as ribotype 027). This study compared the Verigene CDF test with anaerobic direct and enriched toxigenic culture on stool specimens from symptomatic patients among five geographically diverse laboratories within the United States. The Verigene CDF test was performed according to the manufacturer's instructions, and the reference methods performed by a central laboratory included direct culture onto cycloserine cefoxitin fructose agar (CCFA) and enriched culture using cycloserine cefoxitin mannitol broth with taurocholate and lysozyme. Recovered isolates were identified as C. difficile using gas liquid chromatography and were tested for toxin using a cell culture cytotoxicity neutralization assay. Strains belonging to ribotype 027 were determined by PCR ribotyping and bidirectional sequencing for ⌬ 117 in tcdC. A total of 1,875 specimens were evaluable. Of these, 275 specimens (14.7%) were culture positive by either direct or enriched culture methods. Compared to direct culture alone, the overall sensitivity, specificity, positive predictive value, and negative predictive value for the Verigene CDF test were 98.7%, 87.5%, 42%, and 99.9%, respectively. Compared to combined direct and enriched culture results, the sensitivity, specificity, positive predictive value, and negative predictive values of the Verigene CDF test were 90.9%, 92.5%, 67.6%, and 98.3%, respectively. Of the 250 concordantly culture-positive specimens, 59 (23.6%) were flagged as "hypervirulent"; 53 were confirmed as ribotype 027, and all 59 possessed ⌬ 117 in tcdC. Time to results was approximately 2.5 h per specimen. The Verigene CDF test is a novel nucleic acid microarray that reliably detects both C. difficile toxins A and B in unformed stool specimens and appears to adequately identify ribotype 027 isolates. The optimum testing method for the diagnosis of Clostridium difficile is still a matter of debate. A variety of testing methods are available and include enzyme immunoassays (EIAs) for detection of toxins A and B, cell culture cytotoxicity neutralization assays (CCNAs), anaerobic toxigenic culture, detection of glutamate dehydrogenase, and nucleic acid amplification tests (NAATs). Although EIAs provide same-day results and are relatively inexpensive and easy to perform, there is general agreement in the United States and in some European countries that EIAs should not be performed as stand-alone tests due to their lack of sensitivity (1, 2). CCNAs and anaerobic toxigenic culture do not provide timely results and have never been standardized. Laboratories are beginning...