We developed a new multiplex PCR assay for detection of Panton-Valentine leukocidin virulence genes and simultaneous discrimination of methicillin-susceptible from -resistant staphylococci. This assay is simple, rapid, and accurate and offers the potential for prompt detection of newly emerging community-associated methicillin-resistant Staphylococcus aureus .

We evaluated blood collected on Nobuto filter-paper (FP) strips for use in detecting Brucella spp. antibodies in caribou. Whole blood (for serum) and blood-saturated FP strips were obtained from 185 killed arctic caribou (Rangifer tarandus groenlandicus). Sample pairs (serum and FP eluates) were simultaneously tested in duplicate using competitive enzyme-linked immunosorbent assay (c-ELISA) and indirect ELISA (i-ELISA) for Brucella spp. Prior work based on isolation of Brucella spp. revealed sensitivity (SE) and specificity (SP) of 100% and 99%, respectively, for both these serum assays in caribou. Infection status of the animals in the current study was unknown but recent sampling had revealed clinical brucellosis and >40% Brucella antibody prevalence in the herd. To assess the performance of FP relative to serum in these assays, serum was used as the putative gold standard. On both assays, the findings for duplicate runs (A and B) were similar. For c-ELISA run A, the FP Brucella prevalence (47%) was lower than serum prevalence (52%), with SE 89% (95% confidence interval [CI]: 82-95%) and SP 99% (97-100%). For i-ELISA run A, serum and FP Brucella prevalence rates were identical (43%), and the SE and SP of FP testing were 100% and 99% (97-100%), respectively. The findings suggest better FP test performance with i-ELISA than with c-ELISA; however, i-ELISA does not distinguish cross-reacting antibodies induced by Brucella vaccination or exposure to certain other Gram-negative pathogens. Results for duplicate FP eluates (prepared using separate FP strips from each animal) were strongly correlated for both protocols (r=0.996 and 0.999 for c-ELISA and i-ELISA, respectively), indicating minimal variability among FPs from any individual caribou. Dried caribou FP blood samples stored for 2 mo at room temperature are comparable with serum for use in Brucella spp. c-ELISA and i-ELISA. Hunter-based FP sampling can facilitate detection of disease exposure in remote regions and under adverse conditions, and can expand wildlife disease surveillance across temporospatial scales.

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Cancer-related, mucin-type carbohydrate epitopes, principally mannose and sialo-syl residues, are expressed on the envelope protein gp 160 of the human immunodeficiency virus (HIV). Anticarbohydrate antibodies directed toward these and other carbohydrate epitopes are known to neutralize HIV-1 infection by cell-free virus. Carbohydrates, however, being T cell-independent antigens, typically elicit diminished immune responses. To overcome this potential draw back, we have examined the ability of peptides that mimic such epitopes to elicit immune responses that cross-react with carbohydrate structures. We report that mouse polyclonal antisera generated against peptides that mimic mucin-related carbohydrate epitopes have anti-HIV-1 activity. Generation of antibodies was not lr-gene restricted, as at least two different strains of mice. Balb/c (H-2d) and C57Bl/6 (H-2b), responded equally to the peptides. The antipeptide sera displayed neutralizing activity against HIV-I/MN and HIV-I/3B viral strains. This neutralization was as good as human anti-HIV sera. These results indicate that peptide mimics of carbohydrates provide a novel strategy for the further development of reagents that elicit immune responses to carbohydrate epitopes associated with many infectious organisms and tumor cells.

ABSTRACT:Filter-paper (FP) blood sampling can facilitate wildlife research and expand disease surveillance. Previous work indicated that Nobuto FP samples from caribou and reindeer (Rangifer tarandus subspecies) had comparable sensitivity and specificity to serum samples ($80% for both) in competitive enzyme-linked immunosorbent assays (cELISAs) for Brucella spp., Neospora caninum, and West Nile virus. The same sensitivity and specificity criteria were met in indirect ELISAs for Brucella spp., bovine herpesvirus type 1 (BHV-1), parainfluenza virus type 3 (PI-3), and bovine respiratory syncytial virus (BRSV), with adjusted FP thresholds used for PI-3 and BRSV. Comparable sensitivity and specificity values to serum were also observed for FP in virus neutralization (VN) assays for bovine viral diarrhea virus types I and II; however, reduced sensitivity is a potential limitation of FP samples in protocols that require undiluted serum (i.e., VN and N. caninum cELISA). We evaluated the performance of FP samples from reindeer and caribou in these nine assays after simulating potential challenges of high-latitude field collections: 1) different durations of storage and 2) different processing/storage regimes involving freezing or drying. Sample pairs (serum and FP) were collected from reindeer and caribou populations in 2007-10 and were tested in duplicate. Comparable performance to serum was defined as sensitivity and specificity $80%. In the storage experiments, FP performance was determined after 2 mo of storage dry at room temperature, and after two longer periods (variable depending on assay; up to 2 yr). After 1 yr, compared to frozen serum stored for the same period, sensitivity was $88% for all but two assays (68% BHV-1; 75% PI-3), and specificity remained .90%. A limited trial evaluated the effect of freezing FP samples as opposed to drying them for storage. There were no observed detrimental effects of freezing on FP sample performance, but rigorous investigation is warranted.