A New Generation Microarray for the Simultaneous Detection and Identification of Yersinia pestis and Bacillus anthracis in Food

The use of microarrays as a multiple analytic system has generated increased interest and provided a powerful analytical tool for the simultaneous detection of pathogens in a single experiment. A wide array of applications for this technology has been reported. A low density oligonucleotide microarray was generated from the genetic sequences of Y. pestis and B. anthracis and used to fabricate a microarray chip. The new generation chip, consisting of 2,240 spots in 4 quadrants with the capability of stripping/rehybridization, was designated as "Y-PESTIS/B-ANTHRACIS 4x2K Array." The chip was tested for specificity using DNA from a panel of bacteria that may be potentially present in food. In all, 37 unique Y. pestis-specific and 83 B. anthracis-specific probes were identified. The microarray assay distinguished Y. pestis and B. anthracis from the other bacterial species tested and correctly identified the Y. pestis-specific oligonucleotide probes using DNA extracted from experimentally inoculated milk samples. Using a whole genome amplification method, the assay was able to detect as low as 1 ng genomic DNA as the start sample. The results suggest that oligonucleotide microarray can specifically detect and identify Y. pestis and B. anthracis and may be a potentially useful diagnostic tool for detecting and confirming the organisms in food during a bioterrorism event

A New Generation Microarray for the Simultaneous Detection and Identification of Yersinia pestis and Bacillus anthracis in Food

The use of microarrays as a multiple analytic system has generated increased interest and provided a powerful analytical tool for the simultaneous detection of pathogens in a single experiment. A wide array of applications for this technology has been reported. A low density oligonucleotide microarray was generated from the genetic sequences of Y. pestis and B. anthracis and used to fabricate a microarray chip. The new generation chip, consisting of 2,240 spots in 4 quadrants with the capability of stripping/rehybridization, was designated as “Y-PESTIS/B-ANTHRACIS 4x2K Array.” The chip was tested for specificity using DNA from a panel of bacteria that may be potentially present in food. In all, 37 unique Y. pestis-specific and 83 B. anthracis-specific probes were identified. The microarray assay distinguished Y. pestis and B. anthracis from the other bacterial species tested and correctly identified the Y. pestis-specific oligonucleotide probes using DNA extracted from experimentally inoculated milk samples. Using a whole genome amplification method, the assay was able to detect as low as 1 ng genomic DNA as the start sample. The results suggest that oligonucleotide microarray can specifically detect and identify Y. pestis and B. anthracis and may be a potentially useful diagnostic tool for detecting and confirming the organisms in food during a bioterrorism event

Rapid detection and serovar identification of common Salmonella enterica serovars in Canada using a new pyrosequencing assay

Serotyping of Salmonella enterica subspecies enterica is a critical step for foodborne salmonellosis investigation. We have developed a new assay to identify Salmonella enterica subsp. enterica (S. enterica) serovars based on a triplex polymerase chain reaction (PCR) with pyrosequencing for amplicon confirmation and phylogenetic discrimination of strain. The top 54 most prevalent serovars of S. enterica in Canada were examined with a total of 23 single nucleotide polymorphisms (SNPs) and/or variations (SNVs) located on three genes (fliD, sopE2, and spaO). Seven of the most common serovars, including Newport, Typhi, Javiana, Infantis, Thompson, Heidelberg and Enteritidis are successfully distinguished from the other serovars based on their unique SNP/SNV combinations. The remaining serovars, including Typhimurium, ssp I 4,[5],12:i:-, and Saintpaul are further divided into 47 subgroups that demonstrate the relatedness to phylogenetic classifications of each serovar. This pyrosequencing assay is not only cost-effective, rapid, and user-friendly, but also provides phylogenetic information by analysing 23 selected SNPs. With the additional layer of confidence to the PCR results and the accuracy and speed of pyrosequencing, this novel method would benefit the food industry and provides a tool for rapid outbreak investigation through quick detection and identification of common S. enterica serovars in Canada.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Evaluation of the diagnostic accuracy of CareStart G6PD deficiency Rapid Diagnostic Test (RDT) in a malaria endemic area in Ghana, Africa.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most widespread enzyme defect that can result in red cell breakdown under oxidative stress when exposed to certain medicines including antimalarials. We evaluated the diagnostic accuracy of CareStart G6PD deficiency Rapid Diagnostic Test (RDT) as a point-of-care tool for screening G6PD deficiency.A cross-sectional study was conducted among 206 randomly selected and consented participants from a group with known G6PD deficiency status between February 2013 and June 2013. A maximum of 1.6ml of capillary blood samples were used for G6PD deficiency screening using CareStart G6PD RDT and Trinity qualitative with Trinity quantitative methods as the "gold standard". Samples were also screened for the presence of malaria parasites. Data entry and analysis were done using Microsoft Access 2010 and Stata Software version 12. Kintampo Health Research Centre Institutional Ethics Committee granted ethical approval.The sensitivity (SE) and specificity (SP) of CareStart G6PD deficiency RDT was 100% and 72.1% compared to Trinity quantitative method respectively and was 98.9% and 96.2% compared to Trinity qualitative method. Malaria infection status had no significant (P=0.199) change on the performance of the G6PD RDT test kit compared to the "gold standard".The outcome of this study suggests that the diagnostic performance of the CareStart G6PD deficiency RDT kit was high and it is acceptable at determining the G6PD deficiency status in a high malaria endemic area in Ghana. The RDT kit presents as an attractive tool for point-of-care G6PD deficiency for rapid testing in areas with high temperatures and less expertise. The CareStart G6PD deficiency RDT kit could be used to screen malaria patients before administration of the fixed dose primaquine with artemisinin-based combination therapy

Map of the study communities.

<p>Map of the study communities.</p

An outline comparing some common factors of importance between the methods used for G6PD screening in the study.

<p>*: Volume of blood to estimate Haemoglobin concentration is needed.</p><p>An outline comparing some common factors of importance between the methods used for G6PD screening in the study.</p

Flow chart of profile for study participants’ selection and recruitment.

<p>Flow chart of profile for study participants’ selection and recruitment.</p

Estimates of G6PD enzyme activity levels by malaria parasite status.

<p>Estimates of G6PD enzyme activity levels by malaria parasite status.</p

ROC curve of RDT performance with plotted points of considered enzyme activity.

<p>ROC curve of RDT performance with plotted points of considered enzyme activity.</p