2009
DOI: 10.1007/s10096-009-0759-8
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Abstract: The replication of hepatitis A virus (HAV) is via a complementary negative-strand RNA. Each negative strand may serve as a template for the synthesis of many positive strands. The aim of this study was to detect the intermediate replicative (negative strand) of HAV in order to monitor its replication in vitro and in vivo. Real-time polymerase chain reaction (PCR) was standardized to detect the intermediate replicative of HAV in cell culture and liver from non-human primates infected experimentally. HAV primers… Show more

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Cited by 12 publications
(11 citation statements)
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“…1). Viral RNA was reported recently to be present in the liver of cynomolgous monkeys as late as 9 wk after experimental HAV infection (40,41), but its continued presence beyond 46 wk in the chimpanzee (Fig. 1) was unexpected and represents another major difference from acute resolving hepatitis C. In acute resolving HCV infection, viral RNA is typically cleared by 10-20 wk of infection (Fig.…”
Section: Discussionmentioning
confidence: 92%
“…1). Viral RNA was reported recently to be present in the liver of cynomolgous monkeys as late as 9 wk after experimental HAV infection (40,41), but its continued presence beyond 46 wk in the chimpanzee (Fig. 1) was unexpected and represents another major difference from acute resolving hepatitis C. In acute resolving HCV infection, viral RNA is typically cleared by 10-20 wk of infection (Fig.…”
Section: Discussionmentioning
confidence: 92%
“…This lack of strand specificity has been attributed to a combination of factors, including self-priming of the RNA due to secondary hairpin structures (Haddad et al, 2006;Lanford et al, 1994;Lerat et al, 1996;Stahlberg et al, 2004;Timofeeva and Skrypina, 2001;Tuiskunen et al, 2010), false priming of the incorrect strand (Gunji et al, 1994;Lanford et al, 1994;Lin et al, 2002;Sangar and Carroll, 1998) and random priming by contaminating endogenous or exogenous nucleic acids (Gunji et al, 1994;Piche and Schernthaner, 2003;Timofeeva and Skrypina, 2001). Attempts to overcome these problems include performing RT reactions at high temperatures (Haddad et al, 2007;Komurian-Pradel et al, 2004), use of the thermostable RTth enzyme (de Paula et al, 2009;Lanford et al, 1994;Radkowski et al, 2002;Selva et al, 2004), use of tagged RT primers (Peyrefitte et al, 2003;Plaskon et al, 2009;Purcell et al, 2006;Vashist et al, 2012) or a combination of tagged primers and RTth enzyme (Craggs et al, 2001). It was shown that the approach employing tagged primers in combination with high RT temperature greatly improved the specificity of the RT reactions.…”
Section: Introductionmentioning
confidence: 98%
“…It was shown that the approach employing tagged primers in combination with high RT temperature greatly improved the specificity of the RT reactions. So far, quantitation of negative strand RNA for analysis of positive-sense RNA virus replication has been applied for a number of viruses, which include hepatitis A (de Paula et al, 2009;Jiang et al, 2004), hepatitis C (Craggs et al, 2001;Lanford et al, 1994;Lerat et al, 1996;Lin et al, 2002;Takyar et al, 2000), hepatitis E (Williams et al, 2001), GB virus C (Handa and Brown, 2000), dengue (Peyrefitte et al, 2003), O'nyong-nyong and Chikungunya (Plaskon et al, 2009), murine norovirus (Vashist et al, 2012) and foot-and-mouth disease virus (Horsington and Zhang, 2007). Due to the problems associated with false detection of negative strand RNA, conclusions derived from assays not optimized for strandspecific detection of RNA may not be valid (Crovatto et al, 2000;Hsu et al, 1999).…”
Section: Introductionmentioning
confidence: 99%
“…After cleavage by proteases, the single polyprotein of HAV yields three major protein groups. While P1 encodes for capsid proteins VP1-VP4, P2 and P3 encode for non-structural proteins related to viral replication, including proteases and polymerase (8,9).…”
Section: Introductionmentioning
confidence: 99%