Inactivation of hepatitis A virus and norovirus surrogate in suspension and on food-contact surfaces using pulsed UV light (pulsed light inactivation of food-borne viruses).

This study was conducted to evaluate the inactivation of murine norovirus (MNV-1) and hepatitis A virus (HAV) by pulsed ultraviolet (UV) light. MNV-1 was used as a model for human norovirus. Viral suspensions of about 10(6) PFU/ml were exposed to pulses of UV light for different times and at different distances in a Xenon Steripulse device (model RS-3000C). Inactivation studies were also carried out on 1-cm(2) stainless steel and polyvinyl chloride disks with 10(5) PFU/ml. Inactivation of MNV-1 and HAV at 10.5 cm from the UV source was greater on inert surfaces than in suspension. The presence of organic matter (fetal bovine serum) reduced the effectiveness of pulsed light both in suspension and on surfaces. However, 2-s treatment in the absence of FBS completely inactivated (5 log reduction) the viral load at different distances tested, whether in suspension (MNV-1) or on disks (MNV-1 and HAV). The same treatment in the presence of fetal bovine serum (5%) allowed a reduction of about 3 log. This study showed that short duration pulses represent an excellent alternative for inactivation of food-borne viruses. This technology could be used to inactivate viruses in drinking water or on food-handling surfaces.

Rotaviruses from Canadian farm samples.

Animal rotavirus (RoV) strains detected in Canadian swine and dairy cattle farms were characterized by sequence analysis of viral protein 4 (VP4), VP6, VP7 and non-structural protein 4 segments from 15 RoV strains. Some porcine strains were found to contain a mixture of segments typical of human and animal viruses. One strain represented a novel VP6 genotype "I14", G2-P[27]-I14. Other strains detected in porcine samples represented multiple different segment types. These results illustrate the active evolution of animal RoV strains and underline the need for surveillance of both animal and human strains in public health-monitoring programs.

Real-time molecular beacon NASBA for rapid and sensitive detection of norovirus GII in clinical samples.

To improve the sensitivity and efficiency of the real-time nucleic acid sequence based amplification (NASBA) assay targeting the open reading frame 1-2 (ORF1-ORF2) junction of the norovirus (NoV) genome, a selection of clinical samples were analyzed. The assay results were compared with those of TaqMan and conventional reverse transcription PCR (RT-PCR) and a commercial enzyme-linked immunoassay (ELISA) for the specific detection of GII NoV in 96 fecal samples. Based on end-point dilution, the two real-time assays had similar sensitivities (0.01 particle detectable units), two log10 cycles greater than that of conventional RT-PCR. GII NoV was detected in 88.54% of the samples by real-time NASBA, in 86.46% by TaqMan RT-PCR, in 81.25% by conventional RT-PCR, and in 65.7% by ELISA. The two real-time assays were in agreement for 88.5% of the samples. These results demonstrate that real-time NASBA with a molecular beacon probe is highly sensitive, accurate, and specific for NoV detection in clinical samples. Applying this technique to samples with complex matrix and low viral loads, such as food and environmental samples, could be useful for the detection of NoVs and will improve the prevention of NoV outbreaks.

Evaluation of the persistence of infectious human noroviruses on food surfaces by using real-time nucleic acid sequence-based amplification.

Noroviruses (NoV) are the major cause of nonbacterial gastroenteritis. However, there is no published study to ascertain their survival on foodstuffs which are directly related to human health risk. In the present study, we developed a rapid, simple, and sensitive real-time nucleic acid sequence-based amplification (NASBA) combined with an enzymatic treatment for distinguishing infectious from noninfectious human NoV. The developed method was validated using spiked ready-to-eat food samples. When feline calicivirus (FCV) was used as a NoV surrogate in the preliminary assays, it appeared more sensitive to heat inactivation and enzymatic pretreatment than the human NoV. This suggests that FCV may not be an ideal model for studying NoV. Our results reveal clearly that the developed enzymatic pretreatment/real-time NASBA combination successfully distinguished the infectious from heat-inactivated NoV. Moreover, we demonstrate that NoV survived for at least 10 days on refrigerated ready-to-eat foods, such as lettuce and turkey. However, the survival rate was higher on turkey than on lettuce, probably because of their different surface natures. The approach developed in this study may be suitable for more in-depth studies of the persistence and inactivation of human NoV and may be applied to other nonculturable RNA viruses. Moreover, the evaluation of infectious NoV survival provided valuable information concerning its persistence on ready-to-eat food.

Phylogenetic analysis of norovirus isolates involved in some Canadian gastroenteritis outbreaks in 2004 and 2005.

Noroviruses are recognized as the most common cause of nonbacterial gastroenteritis worldwide. In this study, we investigated the molecular epidemiology of noroviral isolates in Canada from 2004 to 2005 by sequencing the RNA polymerase gene and capsid N-terminal/shell (N/S) domain. Norovirus genogroups I and II were thus found to have co-circulated in Canada during the studied period, with a higher incidence of genogroup II (95.7%). The GII-4 or Lordsdale subgroup was the predominant genotype, suggesting that norovirus genogroup II is the major cause of viral gastroenteritis in Canada, as it is in many other countries. Phylogenetic analyses of the RNA polymerase gene and the capsid N/S domain indicated different genotypes for 2 strains, suggesting probable genetic recombination. Sequencing of the norovirus polymerase gene may reflect actual classification but should be supported by sequence information obtained from the capsid gene.