Vibrio cholerae is recognized as a leading human waterborne pathogen. Traditional diagnostic testing for Vibrio is not always reliable, because this bacterium can enter a viable but nonculturable state. Therefore, nucleic acid-based tests have emerged as a useful alternative to traditional enrichment testing. In this article, a TaqMan PCR assay is presented for quantitative detection of V. cholerae in pure cultures, oysters, and synthetic seawater. Primers and probe were designed from the nonclassical hemolysin (hlyA) sequence of V. cholerae strains. This probe was applied to DNA from 60 bacterial strains comprising 21 genera. The TaqMan PCR assay was positive for all of the strains of V. cholerae tested and negative for all other species of Vibrio tested. In addition, none of the other genera tested was amplified with the TaqMan primers and probe used in this study. The results of the TaqMan PCR with raw oysters and spiked with V. cholerae serotypes O1 and O139 were comparable to those of pure cultures. The sensitivity of the assay was in the range of 6 to 8 CFU g ؊1 and 10 CFU ml ؊1 in spiked raw oyster and synthetic seawater samples, respectively. The total assay could be completed in 3 h. Quantification of the Vibrio cells was linear over at least 6 log units. The TaqMan probe and primer set developed in this study can be used as a rapid screening tool for the presence of V. cholerae in oysters and seawater without prior isolation and characterization of the bacteria by traditional microbiological methods.Vibrio cholerae is a waterborne pathogen that causes gastrointestinal disorders with a wide range of clinical manifestations, including vomiting and rice-like diarrhea (24). The association of human illness with consumption of V. choleraecontaminated oysters, seawater, and other shellfish is well documented (29, 37). Consumption of raw oysters correlates strongly with gastrointestinal infections, and several Vibrio species, including strains of Vibrio parahaemolyticus, Vibrio vulnificus, and V. cholerae, have been implicated as the causative agents.Coastal areas with brackish waters and estuarine regions are niches for many Vibrio species, including strains of toxigenic O1 V. cholerae. Epidemic cholera strains are endemic in several regions, including the U.S. gulf coast and Australia, and are occasionally involved in illnesses in these regions (24). Because Vibrio species attach to material suspended in water, shellfish and mollusks that are in these environments can be expected to consume Vibrio during feeding (24).Traditional identification methods currently used are time-consuming and laborious, requiring prolonged incubation and selective enrichment to reduce the growth of background flora. Vibrio cells may also enter a viable but not culturable (VBNC) state, caused by nutrient starvation and physical stress. This may explain the failure of traditional culture techniques to isolate this organism from contaminated water and food samples implicated in food-borne outbreaks (10,27,47). Several investigator...
In the face of concerns over an influenza pandemic, identification of virulent influenza A virus isolates must be obtained quickly for effective responses. Rapid subtype identification, however, is difficult even in well-equipped virology laboratories or is unobtainable in the field under more austere conditions. Here we describe a genome assay and microarray design that can be used to rapidly identify influenza A virus hemagglutinin subtypes 1 through 15 and neuraminidase subtypes 1 through 9. Also described is an array-based enzymatic assay that can be used to sequence portions of both genes or any other sequence of interest.
Propionibacterium thoenii strain P127, which produces the bacteriocin propionicin PLG-1, was grown in a skim milk medium and produced bacteriocin in that medium. No bacteriocin activity was detected in skim milk medium in which strain P127-1, a bacteriocin-negative variant of strain P127, had been grown. Five psychrotrophic spoilage or pathogenic organisms (one strain each of Listeria monocytogenes, Pseudomonas fluorescens, Vibrio parahaemolyticus, Yersinia enterocolitica, and one strain of Corynebacterium sp.) were incubated for 24 h in laboratory medium, nonfermented skim milk, and skim milk that had been fermented by strain P127 or P127-1. Strains were inhibited only in the skim milk fermented by strain P127, as evidenced by loss in numbers of viable cells after 24 h at 10 degrees C and less growth than in other media after 24 h at optimal growth temperatures. Growth of selected strains was delayed or slowed during prolonged incubation (21 d) at 10 degrees C. Propionicin PLG-1 shows promise as a preservative for food products.
Bacterial and viral upper respiratory infections (URI) produce highly variable clinical symptoms that cannot be used to identify the etiologic agent. Proper treatment, however, depends on correct identification of the pathogen involved as antibiotics provide little or no benefit with viral infections. Here we describe a rapid and sensitive genotyping assay and microarray for URI identification using standard amplification and hybridization techniques, with electrochemical detection (ECD) on a semiconductor-based oligonucleotide microarray. The assay was developed to detect four bacterial pathogens (Bordetella pertussis, Streptococcus pyogenes, Chlamydia pneumoniae and Mycoplasma pneumoniae) and 9 viral pathogens (adenovirus 4, coronavirus OC43, 229E and HK, influenza A and B, parainfluinza types 1, 2, and 3 and respiratory syncytial virus. This new platform forms the basis for a fully automated diagnostics system that is very flexible and can be customized to suit different or additional pathogens. Multiple probes on a flexible platform allow one to test probes empirically and then select highly reactive probes for further iterative evaluation. Because ECD uses an enzymatic reaction to create electrical signals that can be read directly from the array, there is no need for image analysis or for expensive and delicate optical scanning equipment. We show assay sensitivity and specificity that are excellent for a multiplexed format.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.