Elyse Poitras

Comparative analysis of different TaqMan real‐time RT‐PCR assays for the detection of swine Hepatitis E virus and integration of Feline calicivirus as internal control

Aims:  The aim of this study was to compare the performance of four TaqMan RT‐PCR assays with a commonly used nested RT‐PCR and to include the Feline calicivirus (FCV) as an internal control.

Hepatitis E virus load in swine organs and tissues at slaughterhouse determined by real-time RT-PCR

Although uncommon in North America, Hepatitis E virus (HEV) has been identified in some industrialized countries in patients without a history of travel to HEV-endemic countries. Its presence is ubiquitous worldwide in swine populations. Zoonotic transmission of swine HEV to non human primates has been achieved experimentally and transmission of HEV after ingestion of contaminated raw or undercooked meat is well documented. In Canada, so far, no HEV outbreak has been documented but HEV genotype 3 strains have been identified in sera and faecal samples of swine origin. The objective of the present study was to determine the viral load of HEV in liver, loin, bladder, hepatic lymph node, bile, tonsil, plasma and faeces samples of 43 pigs at slaughter. Feline calicivirus (FCV) was used as sample process control to validate the RNA extraction process, as a confirmation of the absence of sample inhibitors and as an amplification control. Using FCV/HEV multiplex TaqMan RT-qPCR system, HEV RNA was detected in 14 out of the 43 animals tested. HEV was detected in lymph nodes (11/43), bladder (10/43), liver (9/43), bile (8/43), faeces (6/43), tonsils (3/43), plasma (1/43) samples from infected animals. No HEV-positive loin samples were observed. Viral loads of 10(3) to 10(7) copies/g were estimated in positive liver and bile samples.

Development of a real‐time TaqMan PCR assay for the detection of porcine and bovine Torque teno virus

Aims:  The goal of this study was to develop and to optimize molecular tools to detect the presence of Torque teno virus (TTV) in swine and cattle. A novel real‐time polymerase chain reaction (PCR) using a TaqMan probe was developed to detect both genogroups of TTV strains.

Development and Evaluation of a Multiplexed Real-Time TaqMan RT-PCR Assay with a Sample Process Control for Detection of F-specific RNA Coliphage Genogroups I and IV

There are increasing concerns of zoonotic transmission of some animal enteric viruses, such as calicivirus, hepatitis E virus, and rotavirus, which are closely related to human pathogenic strains. Most enteric viruses are detected by molecular techniques because they cannot be cultured. Surrogates such as F-RNA coliphages are cultivable but few molecular methods exist. Individual real-time TaqMan RT-PCR assays for the replicase gene of F-RNA coliphage genogroups I and IV were developed and multiplexed with a real-time TaqMan RT-PCR assay for feline calicivirus as a sample process control for the simultaneous detection and enumeration of genogroup I and IV F-RNA coliphages. Genogroup IV were successfully detected with the multiplexed assay in 80% of fecal samples that contained F-RNA coliphage levels ≥3.2 log plaque forming units (pfu). F-RNA coliphage were at or below the limit of detection in most fecal samples when levels were ≤4 log pfu/g.

Comparison of different RT‐qPCR assays for the detection of human and bovine group A rotaviruses and characterization by sequences analysis of genes encoding VP4 and VP7 capsid proteins

The aim of this study was to compare the performance of four RT‐qPCR assays for the detection of human and bovine group A rotaviruses and to characterize the positive samples by sequence analysis of VP4 and VP7 genes.

Association of age and gender with Torque teno virus detection in stools from diarrheic and non-diarrheic people

BACKGROUND Torque teno virus (TTV) is a small virus belongs to Anelloviridea family. TTV is a disease orphan virus but it has often been associated with a variety of pathologies and co-infections. TTV was recently identified as an infectious agent that could potentially be involved in cases of acute enteritis. OBJECTIVES To ascertain the presence of TTV in stools from diarrheic and not diarrheic people, and to investigate an association between infection, and patient age and gender. STUDY DESIGN Stool samples from people exhibiting signs of enteritis (954) and from non-diarrheic individuals (76) were collected in the former Chinook Health Region (CHR) in Southwestern Alberta, Canada from May 2008 to April 2009. Viral genetic material was extracted, and detection and quantification of TTV were carried out by real-time PCR. The presence of other viral and bacterial enteric pathogens was also investigated. RESULTS More (P<0.001) diarrheic people (38.8%) tested positive for TTV DNA than non-diarrheic individuals (18.4%). Furthermore, viral load was greater (P<0.001) in stools from diarrheic (2.0×10(7)copies/g) than non-diarrheic (2.0×10(3)copies/g) people. TTV DNA was detected most often in diarrheic individuals that were 0-5 (57.3%) and greater than 81 (59.0%) years of age. Combined across age, the prevalence of TTV was higher among men than women (P=0.003). Co-infections with other enteric pathogens were observed. CONCLUSIONS This study revealed a significant association between TTV prevalence and viral load, and enteritis. Also, TTV prevalence was significantly higher in the very young and elderly suggesting that immunological status is important.