Experiments were performed to determine the thermal resistance of hepatitis A virus (HAV) in three types of dairy products containing increased amounts of fat content (skim milk, homogenized milk; 3.5% MFG, and table cream; 18% MFG). HAV-inoculated dairy products were introduced into custom-made U-shaped microcapillary tubes that in turn were simultaneously immersed in a waterbath, using custom-made floating boats and a carrying platform. Following exposure to the desired time and temperature combinations, the contents of each of the tubes was retrieved and was tested by plaque assay to determine the reduction in virus titer. Our data indicated that < 0.5 min at 85 degrees C was sufficient to cause a 5-log reduction in HAV titer in all three dairy products, whereas at 80 degrees C, < or = 0.68 min (for skim and homogenized milk), and 1.24 min (for cream) were needed to cause a similar log reduction. Using a nonlinear two-phase negative exponential model (two-compartment model) to analyze the data, it was found that at temperatures of 65, 67, 69, 71, and 75 degrees C, significantly (P < 0.05) higher exposure times were needed to achieve a 1-log reduction in virus titer in cream, as compared to skim and homogenized milk. For example, at 71 degrees C, a significantly (P < 0.05) higher exposure time of 0.52 min (for cream) was needed as compared to < or = 0.18 min (for skim and homogenized milk) to achieve a 1-log reduction in virus titer. A similar trend of inactivation was observed at 73 and 75 degrees C where significantly (P < 0.05) higher exposure times of 0.29 to 0.36 min for cream were needed to cause a 1-log reduction in HAV in cream, as compared to < or = 0.17 min for skim and homogenized milk. This study has provided information on the heat resistance of HAV in skim milk, homogenized milk, and table cream and demonstrated that an increase in fat content appears to play a protective role and contributes to the heat stability of HAV.
Immunomagnetic beads-PCR (IM-PCR), positively-charged virosorb filters (F), or a combination of both methods (F-IM-PCR) were used to capture, concentrate and rapidly detect hepatitis A virus (HAV) in samples of lettuce and strawberries experimentally contaminated. Direct reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of the collected HAV-beads complex showed a detection limit of 0.5 plaque forming units (PFU) of the virus present in 1-ml of wash solution from the produce, which was several hundred-fold more sensitive than that demonstrated by RT-PCR. In separate trials, virus-containing wash solutions from the produce were passed through the filters and the captured virus was eluted with 10 ml volumes of 1% beef extract. Of the 62% filter-captured HAV, an average of 34.8% was eluted by the 1% beef extract. PCR amplification of 2 microl from this eluate failed to produce a clear positive band signal. As little as 10 PFU, present on each piece of the lettuce or strawberry, was detectable by the F-IM-PCR, which was almost 20 times less sensitive than the detection limit of 0.5 PFU by the IM-PCR. However, considering the large volumes (< or =50 ml) used in the F-IM-PCR, the sensitivity of detection could be much greater than that of the IM-PCR, which was restricted to < or =20 ml volumes. These data indicate that the F-IM-PCR method provides the potential for a greater sensitivity of detection than the IM-PCR, since low levels of virus could be detected from large volumes of sample than possible by the IM-PCR method. Although positively-charged filters captured a greater amount of virus than both the IM-PCR and F-IM-PCR methods, direct PCR amplification from beef extract eluates was not successful in detecting HAV from produce.
A simple, rapid and objective infectivity assay based on an in situ enzyme immunoassay (EIA) was developed for the fast-growing and cytopathic cell culture-adapted hepatitis A virus (HAV) strain HM175A.2. Infectivity titration by EIA correlated well with titration by cytopathic effects. The reliability of this assay was demonstrated by close agreement in virus infectivity titers among different assays of the same virus aliquot and between assays of different virus aliquots. HAV infected cell cultures after fixation could be stored for up to 1 week before testing without decline in virus titer.
Hepatitis A virus (HAV) is an important pathogen which has been responsible for many food-borne outbreaks. HAV-excreting food handlers, especially those with poor hygienic practices, can contaminate the foods which they handle. Consumption of such foods without further processing has been known to result in cases of infectious hepatitis. Since quantitative data on virus transfer during contact of hands with foods is not available, we investigated the transfer of HAV from artificially contaminated fingerpads of adult volunteers to pieces of fresh lettuce. Touching the lettuce with artificially contaminated fingerpads for 10 s at a pressure of 0.2 to 0.4 kg/cm(2) resulted in transfer of 9.2% +/- 0.9% of the infectious virus. The pretreatments tested to interrupt virus transfer from contaminated fingerpads included (i) hard-water rinsing and towel drying, (ii) application of a domestic or commercial topical agent followed by water rinsing and towel drying, and (iii) exposure to a hand gel containing 62% ethanol or 75% liquid ethanol without water rinsing or towel drying. When the fingerpads were treated with the topical agents or alcohol before the lettuce was touched, the amount of infectious virus transferred to lettuce was reduced from 9.2% to between 0.3 and 0.6% (depending on the topical agent used), which was a reduction in virus transfer of up to 30-fold. Surprisingly, no virus transfer to lettuce was detected when the fingerpads were rinsed with water alone before the lettuce was touched. However, additional experiments with water rinsing in which smaller volumes of water were used (1 ml instead of 15 ml) showed that the rate of virus transfer to lettuce was 0.3% +/- 0.1%. The variability in virus transfer rates following water rinsing may indicate that the volume of water at least in part influences virus removal from the fingerpads differently, a possibility which should be investigated further. This study provided novel information concerning the rate of virus transfer to foods and a model for investigating the transfer of viral and other food-borne pathogens from contaminated hands to foods, as well as techniques for interrupting such transfer to improve food safety.
110 normal, healthy adults were tested for antibody to hepatitis A (anti-HA) type IgG and 86 (78.2%) were found to be positive. An age-specific prevalence wasfound to be lowest in the lower agegroup and highest in the higher age-group. Out of 24 IgG positive individuals, only one was found to have type IgM. No significant difference in the incidence ofanti-HA type IgG was found between 42 patients in the Urology Unit, General Hospital, Kuala Lumpur and normal individuals (P > 0.1). 15 patients diagnosed as viral hepatitis were investigated for HA V IgG and IgM antibodies. 13 (86.7%) were positive for type IgG. Of this, only five (33%) were positive for the type IgM, suggesting that HA V is the cause of acute viral hepatitis in 33% of cases admitted to hospital as viral hepatitis.