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Several nucleotide analogues have been approved for use in treating hepatitis B virus (HBV) infection. Long-term exposure to therapy leads to the emergence of mutations within the HBV DNA polymerase gene, resulting in drug resistance, a major factor contributing to therapy failure. Chronic HBV patients from the Khyber Pakhtunkhwa province, Pakistan, who had completed 6 months of therapy participated in this study. Samples were collected from 60 patients. In this study, the entire reverse transcriptase domain of the HBV polymerase gene was amplified using nested polymerase chain reaction and sequenced. Drug-resistant mutations were detected in nine (22.5%) patients. All of these patients had lamivudine-resistant mutations (rtM204V + L180M), while seven individuals (17.5%) had both lamivudine- plus entecavir-resistant mutations (L180M + M204V + S202G). N236T, a mutation that gives rise to tenofovir and adefovir resistance, was observed in two (5%) patients. T184A, a partial drug-resistant mutation to entecavir, was found in five (12.5%) patients. Furthermore, other genotypic variants (100%) and vaccine escape mutations (5%) were additionally observed. Moreover, pN459Y (35%), pN131D (20%), pL231S (20%), pP130Q (17.5%), pS189Q (12.5%), pP161S (5%), pH160P (2.5%), pT322S (2.5%), and pA223S (2.5%) mutations in the polymerase gene, as well as sA166V (17.5%), sQ181K (12.5%), sV184R (7.5%), sA17E (5%), sP153S/K (5%), sW156C (5%), sC76Y (2.5%), and S132F (2.5%) mutations in the small surface gene, were identified for the first time in this study. Phylogenetic analysis showed that genotype D was predominant amongst the HBV carriers. Subtype D1 was found in most patients, while two patients were subtype D9. These novel findings may contribute to the body of knowledge and have clinical significance for treating and curing HBV infections in Pakistan. [ABSTRACT FROM AUTHOR]

Several nucleotide analogues have been approved for use in treating hepatitis B virus (HBV) infection. Long-term exposure to therapy leads to the emergence of mutations within the HBV DNA polymerase gene, resulting in drug resistance, a major factor contributing to therapy failure. Chronic HBV patients from the Khyber Pakhtunkhwa province, Pakistan, who had completed 6 months of therapy participated in this study. Samples were collected from 60 patients. In this study, the entire reverse transcriptase domain of the HBV polymerase gene was amplified using nested polymerase chain reaction and sequenced. Drug-resistant mutations were detected in nine (22.5%) patients. All of these patients had lamivudine-resistant mutations (rtM204V + L180M), while seven individuals (17.5%) had both lamivudine- plus entecavir-resistant mutations (L180M + M204V + S202G). N236T, a mutation that gives rise to tenofovir and adefovir resistance, was observed in two (5%) patients. T184A, a partial drug-resistant mutation to entecavir, was found in five (12.5%) patients. Furthermore, other genotypic variants (100%) and vaccine escape mutations (5%) were additionally observed. Moreover, pN459Y (35%), pN131D (20%), pL231S (20%), pP130Q (17.5%), pS189Q (12.5%), pP161S (5%), pH160P (2.5%), pT322S (2.5%), and pA223S (2.5%) mutations in the polymerase gene, as well as sA166V (17.5%), sQ181K (12.5%), sV184R (7.5%), sA17E (5%), sP153S/K (5%), sW156C (5%), sC76Y (2.5%), and S132F (2.5%) mutations in the small surface gene, were identified for the first time in this study. Phylogenetic analysis showed that genotype D was predominant amongst the HBV carriers. Subtype D1 was found in most patients, while two patients were subtype D9. These novel findings may contribute to the body of knowledge and have clinical significance for treating and curing HBV infections in Pakistan.

Astroviruses have been associated with enteric and extra-intestinal disorders in many animal species, including chickens. Here, we describe the detection and characterisation of chicken astrovirus (CAstV) in broilers and its seroprevalence in broiler breeder flocks. Based on PCR protocol, viral confirmation was carried out on clinical tissue samples from broiler chickens suffering from uneven growth and poor performance. The tissues were molecularly detected for CAstV with differential diagnostic testing against the Newcastle disease virus, infectious bronchitis virus, avian nephritis virus, avian rotavirus, fowl adenovirus and avian reovirus. Polymerase gene-based phylogenetic analyses of the twenty samples detected positive for CAstV indicate they belong to Group I and are related to strains from the US, UK, India and Poland. From these 20 samples, CastV could be isolated from 3 samples upon inoculation in 5-day-old specific-pathogenfree (SPF) embryonated chicken eggs (ECE); virus-infected embryos showed dwarfing, haemorrhages, oedema and gelatinous lesions at harvest. The enzymelinked immunosorbent assay (ELISA) results revealed a high prevalence of antibodies against CAstV amongst the broiler breeder flocks tested. It is the first study that describes the detection and prevalence of CAstV in broiler chickens and broiler breeder flocks in Malaysia. [ABSTRACT FROM AUTHOR]

Barley yellow dwarf virus (BYDV) causes a significant yield reduction and crop loss in cereal and grasses worldwide. Studies revealed that barley yellow dwarf virus is present in most regions of Iran. Wheat and barley plants showing the infection symptoms were collected from cereal fields in Hamedan Province during the springs of 2020 and 2021. Samples were tested for BYDV infection by Indirect ELISA. Positive reactions to BYDV antibody in most samples were detected. After total RNA extraction, RT-PCR was performed to amplify polymerase gene using specific BYDV-PAV primers. BYDV-PAV is the most prevalent species of BYDVs in Iran. Results confirmed BYDV infection. For molecular and phylogenetic comparison, two samples from Hamedan and Asadabad were selected and their polymerase genes were sequenced. The sequencing results revealed that Hamedan isolate had maximum homology with Kahnouj, Karaj, Yazd and Arak isolates (with 95% - 96% identity). The isolates in this group proposed as a distinct species due to their low similarity with other isolates in GenBank. Asadabad isolate had maximum homology with Estonia and America BYDV-PAS with more than 92% homology. The results of phylogenetic analysis revealed that Iranian isolates of BYDV clustered in three distinct groups which shows the presence of a high diversity in the polymerase gene of this virus. This study is the first report of BYDV-PAS species in Iran. [ABSTRACT FROM AUTHOR]

Copyright of Viral Hepatitis Journal / Viral Hepatit Dergisi is the property of Galenos Yayinevi Tic. LTD. STI and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

A case-control study was performed to assess prescence of inclusion body hepatitis (IBH) caused by fowl adenoviruses (FAdVs) at Kafr EL-Shiekh Governorate, Egypt, during spring, 2017. The case group consisted of 100 liver and spleen samples collected from 10 broiler chickens flocks (10 samples from each flock) suspected to be infected with IBH depending on clinical manefestations and necropsy examination. Controls were randamly selected from chickens without clinical sings or evidence of the disease on postmortem examination. Molecular screening of the disease disease in collected samples based on the DNA polymerase gene of FAdVs was carried out. Furthermore, the DNA polymerase gene sequence was determined and analyzed with published reference sequences on GeneBank. Respectively, enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) were used to confirm existence of co-infection with chicken infectious anemia virus (CIAV) and/or infectious bursal disease virus (IBDV in flocks involved in the study. Using PCR, FAdV genome was detected in seven flocks in the case group and one in the control group. FAdV identified in this study revealed close genetic relationship with FAdVs-D previously identified in UK and Canada, suggesting potential virus transmission from these countries. All tested serum samples from diseased chickens were positive for CIAV infection via ELISA while none of the collected bursa of Fabricius samples tested IBDV positive by RT-PCR. Therefore, results obtained from the current study highlighted the importance of implementation of control measures against FAdV and CIAV in Egyptian poultry flocks. This study opens the door for future work toward specific identification of FAdV serotypes circulating in Egyptian poultry farms and molecular characterization of the virus based on hexon gene or full genome sequencing for better understanding of genetic diversity among FAdVs in Egypt at higher reolution. [ABSTRACT FROM AUTHOR]

The reliable identification and classification of infectious diseases is critical for understanding their biology and controlling their impact. Recent advances in sequencing technology have allowed insight into the remarkable diversity of the virosphere, of which a large component remains undiscovered. For these emerging or undescribed viruses, the process of classifying unknown sequences is heavily reliant on existing nucleotide sequence information in public databases. However, due to the enormous diversity of viruses, and past focus on the most prevalent and impactful virus types, databases are often incomplete. Picobirnaviridae is a dsRNA virus family with broad host and geographic range, but with relatively little sequence information in public databases. The family contains one genus, Picobirnavirus, which may be associated with gastric illness in humans and animals. Little further information is available due in part to difficulties in identification. Here, we investigate diversity both within the genus Picobirnavirus and among other dsRNA virus types using a combined phylogenetic and functional (protein structure homology-modelling) approach. Our results show that diversity within picobirnavirus exceeds that seen between many other dsRNA genera. Furthermore, we find that commonly used practices employed to classify picobirnavirus, such as analysis of short fragments and trimming of sequences, can influence phylogenetic conclusions. The degree of phylogenetic and functional divergence among picobirnavirus sequences in our study suggests an enormous undiscovered diversity, which contributes to the undescribed “viral dark matter„ component of metagenomic studies.

Copyright of Agriscientia is the property of Revista AgriScientia and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

We generated reassorted PR8 viruses containing six different combinations of avian influenza virus (AIV) polymerase genes from A/chicken/Korea/01310/2001 (H9N2) (01310) and A/chicken/Korea/KBNP-0028/2000 (H9N2) (0028) to examine the effects of the AIV polymerase genes PB1, PB2, and PA on replication efficiency in different host cells and pathogenicity in mice. The virus titers of the reassorted viruses possessing 01310 [rPR8-PB2(01310)] and 0028 [rPR8-PB2(0028)] PB2 genes were significantly higher than those of the others except the rPR8 virus in embryonated chicken eggs at 37°C, and those of avian polymerase reassorted viruses were significantly less than rPR8 in MDCK cells at 32 and 37°C. rPR8-PB2(01310), rPR8-PB2(0028), and rPR8-PA(0028) caused no body weight loss in BALB/c mice but rPR8-PA(01310), rPR8-PB1(01310), and rPR8-PB1(0028) caused mortality and significantly different body weight loss compared to those in the mock treatment. In contrast to rPR8-PB2(0028) and rPR8-PA(0028), rPR8-PB2(01310) was not isolated from infected mice, and rPR8-PB1(0028) was less pathogenic than rPR8-PB1(01310). We determined the amino acid residues that were specific to the less pathogenic polymerases. A comparison with those of pandemic 2009 H1N1, human fatal H5N1 and H7N9, and pathogenic AIVs to mice without adaptation revealed that they possessed the mammalian pathogenic constellation of polymerases. Thus, the novel polymerase genes and amino acid residues may be useful to understand the host-barrier overcome of AIVs in mice and to develop safer and efficacious vaccines. [ABSTRACT FROM AUTHOR]

Background Continuous long-term treatment is recommended to reduce the hepatitis B virus (HBV) viral load. However, as a consequence, resistance mutations can emerge and be transmitted to other individuals. The polymerase (POL) gene overlaps the surface (S) gene. Thus, during treatment, mutations in the POL gene may lead to changes in hepatitis B surface antigen (HBsAg). The purpose of this study was to evaluate the frequency of lamivudine and vaccine escape mutations in HBsAg-positive blood donors from the city of Santos and in untreated HBV mono-infected patients from the city of São Paulo, Brazil. Methods HBV DNA was extracted from 80 serum samples, of which 61 were from volunteer blood donors and 19 were from untreated HBV patients. A fragment of the POL/S genes containing 593 base pairs was amplified using nested PCR. Thirty four were PCR-positive and sequencing was performed using an ABI Prism 3130 Genetic Analyzer. Alignments and mutation mapping were performed using BioEdit software. Results HBV DNA from 21 blood donors and 13 untreated patient samples were characterized using nucleotide sequencing PCR products from the POL/S genes. We were able to detect one sample with the resistance mutation to lamivudine rtM204V + rtL180M (2.94%), which was found in a volunteer blood donor that has never used antiviral drugs. The other samples showed only compensatory mutations, such as rtL80F (5.88%), rtL80V (2.94%), rtL82V + rtV207L (2.94%), rtT128P (5.88%), rtT128N/S (2.94%) and rtS219A (5.88%). We found modifications in the S gene in 14 of the 34 samples (41.16%). The mutations detected were as follows: sM133L + sI195T (2.94%), sI195M (2.94%), sP120T (2.94%), sY100S/F (2.94%), sY100C (17.64%), sI/T126P + sQ129P (2.94%), sM198I + sF183C (2.94%) and sS210R (5.88%). Conclusions Our results suggest the transmission of lamivudine-resistant forms. Thus, the evaluation of HBV-infected subjects for lamivudine resistance would improve treatment regime. Moreover, the mutations in the S gene may impair HBsAg antigenicity and contribute to HBsAg failure detection and vaccine escape. [ABSTRACT FROM AUTHOR]

Abstract: Identification of the full repertoire of hepatitis B virus (HBV) peptides that are presented to CD8+ T cells by common HLA class I alleles will be useful for designing immunotherapies for chronic hepatitis B. One hundred and seventy five cloned sequences containing the pre-S/S and P open reading frames (ORF) of the HBV were obtained from serum HBV-DNA of HBeAg-positive (n =4) and HBeAg-negative (inactive healthy carriers (IHC), n =16) Tongan subjects with an inactive chronic HBV infection. In addition, 34 and 32 sequences were obtained 5.2±1.4 (mean±SD) years apart from eight subjects. PAML was used to identify codons in the pre-S/S and P ORFs that were under positive selection pressure (ω >1). The number of non-synonymous substitutions in these codons was compared in IHC who were homozygous for either HLA-B∗4001 (n =9) or HLA-B*5602 (n =7), and who were either positive (n =6) or negative (n =10) for HLA-A*02. 34 codons in the pre-S/S and 11 codons in the P ORFs were under positive selection pressure. There was a higher number of non-synonymous substitutions in these codons in HBeAg-negative versus HBeAg-positive subjects in the P (p =0.02) but not the pre-S/S (p =0.64) ORF. There was no association between any HLA class I allele and non-synonymous substitutions in these codons. There was no increase in positive selection pressure on the pre-S/S and P ORFs with time. In conclusion, we could not find HLA class I-restricted selection pressure on any pre-S/S or P ORF amino acid; raising the possibility that peptide-based immunotherapies for chronic hepatitis B may not require peptides from these ORFs. [Copyright &y& Elsevier]

Hepatitis B virus (HBV) affects more than 350 million people worldwide and is a leading cause of morbidity and mortality in developing countries like Pakistan. Lamivudine has potential to inhibit hepatitis B virus (HBV) replication but long term lamivudine treatment results in mutations in YMDD region of HBV, making this therapy ineffective. In this study, we have optimized a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) based protocol to detect two mutations in HBV DNA polymerase gene (at codon 528 and 552) in chronic hepatitis patients, without any prior lamivudine treatment. HBV genome was extracted and tested by PCR-RFLP for detection of mutations in polymerase gene. Variations in HBV genome were not detected in enrolled patients confirming that lamivudine can be used to treat chronic Hepatitis B in these patients. Several studies have reported the natural occurrence of mutation in YMDD motif of polymerase gene in chronic hepatitis B patients, not treated with lamivudine, but these mutants were not detected in Pakistani lamivudine-untreated chronic hepatitis B patients. [ABSTRACT FROM AUTHOR]

Abstract: Nairobi sheep disease (NSD) virus, the prototype tick-borne virus of the genus Nairovirus, family Bunyaviridae is associated with acute hemorrhagic gastroenteritis in sheep and goats in East and Central Africa. The closely related Ganjam virus found in India is associated with febrile illness in humans and disease in livestock. The complete S, M and L segment sequences of Ganjam and NSD virus and partial sequence analysis of Ganjam viral RNA genome S, M and L segments encoding regions (396bp, 701bp and 425bp) of the viral nucleocapsid (N), glycoprotein precursor (GPC) and L polymerase (L) proteins, respectively, was carried out for multiple Ganjam virus isolates obtained from 1954 to 2002 and from various regions of India. M segments of NSD and Ganjam virus encode a large ORF for the glycoprotein precursor (GPC), (1627 and 1624 amino acids in length, respectively) and their L segments encode a very large L polymerase (3991 amino acids). The complete S, M and L segments of NSD and Ganjam viruses were more closely related to one another than to other characterized nairoviruses, and no evidence of reassortment was found. However, the NSD and Ganjam virus complete M segment differed by 22.90% and 14.70%, for nucleotide and amino acid respectively, and the complete L segment nucleotide and protein differing by 9.90% and 2.70%, respectively among themselves. Ganjam and NSD virus, complete S segment differed by 9.40–10.40% and 3.2–4.10 for nucleotide and proteins while among Ganjam viruses 0.0–6.20% and 0.0–1.4%, variation was found for nucleotide and amino acids. Ganjam virus isolates differed by up to 17% and 11% at the nucleotide level for the partial S and L gene fragments, respectively, with less variation observed at the deduced amino acid level (10.5 and 2%, S and L, respectively). However, the virus partial M gene fragment (which encodes the hypervariable mucin-like domain) of these viruses differed by as much as 56% at the nucleotide level. Phylogenetic analysis of partial sequence differences suggests considerable mixing and movement of Ganjam virus strains within India, with no clear relationship between genetic lineages and virus geographic origin or year of isolation. Surprisingly, NSD virus does not represent a distinct lineage, but appears as a variant with other Ganjam virus among NSD virus group. [Copyright &y& Elsevier]

The hepatitis B virus (HBV) has been classified into eight genotypes (A-H) based on intergenotypic divergence of at least 8% in the complete nucleotide sequence or more than 4% in the S gene. To facilitate the investigation of the relationship between the efficacy of drug treatment and the mutation with specific genotype of HBV, we have established a new genotyping strategy based on a fragment of the HBV DNA polymerase gene. Pairwise sequence and phylogenetic analyses were performed using CLUSTAL V (DNASTAR) on the eight (A-H) standard full-length nucleotide sequences of HBV DNA from GenBank (NCBI) and the corresponding semi-nested PCR products from the HBV DNA polymerase gene. The differences in the semi-nested PCR fragments of the polymerase genes among genotypes A through F were greater than 4%, which is consistent with the intergenotypic divergence of at least 4% in HBV DNA S gene sequences. Genotyping using the semi-nested PCR products of the DNA polymerase genes revealed that only genotypes B, C, and D were present in the 50 cases, from Shenyang, China, with a distribution of 11 cases (22%), 25 cases (50%), and 14 cases (28%) respectively. These results demonstrate that our new genotyping method utilizing a fragment of the HBV DNA polymerase gene is valid and can be employed as a general genotyping strategy in areas with prevalent HBV genotypes A through F. In Shenyang, China, genotypes C, B, and D were identified with this new genotyping method, and genotype C was demonstrated to be the dominant genotype. [ABSTRACT FROM AUTHOR]

The complete nucleotide sequence of an Iranian isolate of Barley yellow striate mosaic virus (BYSMV) L gene comprising 6171 nucleotides was determined using the random polymerase chain reaction followed by filling of gaps by the use of specific primers. The deduced L protein sequence of BYSMV showed similarities with the L proteins of other plant rhabdoviruses and contained polymerase module motifs characteristic of RNA-dependent RNA polymerases of negative-strand RNA viruses. Pairwise and multiple alignments and phylogenetic analysis of BYSMV L protein revealed that it was more closely related to cytorhabdoviruses. These results revealed that, on the basis of polymerase gene, the Iranian isolate of BYSMV and Northern cereal mosaic virus (NCMV) appeared to be the most closely related plant rhabdoviruses sequenced to date. Interestingly, the amino acid sequence identity of BYSMV/NCMV (61.3%), shared more than twice the amino acid sequence identity compared with the next two most similar cytorabdoviruses, Lettuce necrotic yellows virus (28.8%) and Lettuce yellow mottle virus (28.2%). In this paper, we discuss the similarities and differences of BYSMV with other rhabdoviruses which support the classification of BYSMV as a distinct Cytorhabdovirus. This is the first report of BYSMV genome sequences. [ABSTRACT FROM AUTHOR]

Abstract: Background: Noroviruses (NoVs) are one of the major causal agents of acute gastroenteritis among different age groups. Some of the recent studies reveal that NoV genome is highly prone to mutation and recombination which often leads to emergence of new strains. Objectives: To explore the genetic diversity of human Caliciviruses (HuCVs) among diarrhoeic children in Kolkata. Study design: The HuCVs were detected by reverse transcription-polymerase chain reaction (RT-PCR) of the partial RNA dependent RNA polymerase gene (RdRp) and capsid gene and confirmed by sequencing. The sequences were analyzed and the recombination point was detected. Results: Faecal specimens of children (n =111) visiting outpatient department of Dr B. C. Roy Memorial Hospital for Children with acute gastroenteritis were studied: 22 cases were HuCV positive with 21 NoVs. Of these, 12 NoV cases (54.5%) were GII.4 and six cases showed 99% identity with the new variant Japanese strain Hu/NoV/GII.4/OC07138/JP. Three novel NoV GII inter-genotype recombinant strains V1628/IND, V1656/IND and V1737/IND were also detected. The RdRp region of V1628 showed 96% identity with Pont de Roide 673/FRN whereas capsid region resembled GII.7/Osaka F140/JPN strain (98%); the strain V1656 showed 98% identity with RdRp region of GII.4/Monastir 375/TUN but capsid region resembled GII.8/Leverkusen 267/DE (91%); the strain V1737 showed 88% identity with RdRp of GII.5/Minato 6/N1/6/JPN whereas capsid region resembled the GII.12/Gifu 96/JPN (93%). During characterization of Caliciviruses two strains of NoV GII.b and one strain of each NoV GI.1/V1622/06/IND, GI.3/V1707/07/IND, GII.3/V1668/IND, GII.16/V1729/IND, Sapovirus GII.1/V1716/IND were also detected. Conclusions: The emergence of new variant of GII.4/2007, three novel NoV GII inter-genotype recombinant strains and various other NoVs, indicates the remarkable genetic diversity of the HuCVs as diarrhoeagenic viruses circulating in Kolkata, India. [Copyright &y& Elsevier]

Summary: The mutation of YMDD motif of hepatitis B virus (HBV) polymerase gene is the most frequent cause in HBV resistant to lamivudine. The aim of the study was to investigate variation features of HBV polymerase gene in chronic hepatitis B (CHB) patients before and after lamivudine treatment. From the serum samples of five CHB patients before and after 12 months of lamivudine treatment, HBV polymerase gene was amplificated and positive DNA fragments were cloned into JM105 competent cell. Twenty positive clones of every sample were checked with mismatched polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and YMDD variants were sequenced. Among five patients after 12 months of lamivudine treatment, M552I mutations in two patients with HBV DNA rebounding and D553G mutation in one non-responder were detected except two patients with negative HBV DNA consecutively. In summary, D553G mutation is probably one of the reasons that caused non-responders during lamivudine treatment. The mutations of YMDD motif occurred after lamivudine treatment are caused by the induction of lamivudine. [Copyright &y& Elsevier]

Abstract: Background: Little is known about the HIV-1 drug resistance mutations in Ghana. Objectives: To determine the background protease (PR) and reverse transcriptase (RT) mutations of HIV-1 from treatment naïve patients in Ghana. Study design: Twenty-five plasma samples randomly selected were analyzed for drug resistance mutations. The molecular phylogeny and recombinant patterns of the polymerase gene of HIV-1 were also analysed. Results: No major drug-resistance mutations were seen in protease or reverse transcriptase genes. The L10I, L10V, V11I and E35G minor mutations were seen in four patients, while the V179E was observed in a patient with subtype G. An insertion of lysine was found at codon 36 of the protease gene of one patient. The predominant subtype was the CRF02_AG strain (n =22), but 3 (13.6%) of these were recombinants with HIV-1 subtype K and/or A1. Two patients harboured unclassified/complex strains with D/CRF01_AE and G/CRFAG_02 subtypes for the PR and RT, respectively, using the Stanford Database. Viral loads (VL) ranged from 2290 to >1,500,000c/ml (mean=339,065c/ml). Conclusions: Treatment naïve patients in Ghana before scale-up may have minor but not major PR mutations and high viral loads. The clinical effects of minor mutations/polymorphisms in the PR and RT genes and recombinants need to be investigated. [Copyright &y& Elsevier]

Background Lymphocryptovirus (LCV) is found in various non-human primates. As a herpesvirus naturally infecting gibbons it is closely related to human Epstein–Barr virus (EBV) with which it shares considerable genetic, biological and epidemiologic features. Methods We collected blood samples from 70 gibbons (51 Hylobates lar, 18 Hylobates pileatus and 1 Hylobates agilis) for further separation into serum and peripheral blood mononuclear cells (PBMC). Results Only 13 of 70 (18.6%) sera were serologically positive for human EBV IgG but 64 of 70 (91.4%) PBMCs yielded the partial LCV DNA polymerase gene by semi-nested PCR, which we subjected to direct sequencing. All sequences showed 84% nucleic acid and 91% amino acid identity to human EBV. Phylogenetic tree analysis demonstrated gibbon LCVs clustered separately from other gammaherpesvirinae but closely related to LCV of other species. Conclusions Based on LCV DNA detection, we discovered a high prevalence of LCV infection among gibbons. Further characterization of non-human primate LCV might thus provide new insight into both evolution and pathogenicity of gammaherpesvirinae. [ABSTRACT FROM AUTHOR]

Abstract: A real-time based amplification assay with molecular beacons was used to detect and quantify PCR amplicons to discriminate between the newly described Lamivudine-resistant YSDD variant, a known YIDD variant and wild-type Hepatitis B virus (HBV) DNA in the YMDD region of the polymerase gene. Using this assay, we retrospectively analysed samples from two HBV chronically infected Asian twin sisters, starting 9 weeks before therapy, during and between two periods of treatment with Lamivudine. In order to analyse more accurately the dynamics of variant DNA loads during and after therapy, this real time assay was compared to three other mutation analysis techniques, restriction fragment length polymorphism (RFLP), InnoLipa HBV-DR assay and direct sequence analysis. With this technique, new information on the dynamics of variants during and after therapy was obtained. [Copyright &y& Elsevier]

Grapevine leafroll-associated virus 3 (GLRaV-3) is an important pathogen of grapevine in the Czech Republic. A Czech isolate was studied at the molecular level in order to extend data on molecular variability of the virus from the central European region. Three conserved regions (genes encoding the RNA-dependent RNA polymerase, the heat shock protein 70 and the 55 kDa protein) were amplified using reverse transcriptase-polymerase chain reaction, sequenced and compared with related available sequences. On the sequenced regions, the Czech isolate revealed more than 99% nucleotide and amino acid identity with the published full-length sequence of a North American GLRaV-3 isolate and a similarly high identity with available partial GLRaV-3 sequences of isolates from different parts of the world. The phylogenetic position of the new and already characterized GLRaV-3 isolates within the Ampelovirus genus is discussed. [ABSTRACT FROM AUTHOR]

We have analysed the dynamics of HBV variants related to Lamivudine resistance in 22 chronically infected patients during and after the end of Lamivudine therapy. Thirteen patients had a confirmed methionine mutation in the YMDD region of the reverse transcriptase domain determined by sequence analysis. They responded to therapy having a mean reduction of HBV DNA of 4.55 log (range 2.93–8.91). Nine patients partly responded to therapy, with a small decline of HBV DNA (mean reduction of 3.39 log, range 1.72–5.12) and no indication for an YMDD variant. Samples were re-analysed with a HBV Drug Resistance Line Probe assay (InnoLipa HBV-DR), which detected as low as 10% of a variant HBV population. With this assay, changes in the YMDD region were detected with a mean of 2 weeks (range -8 to 10) earlier than by an increase of HBV DNA levels. Increase of ALT was observed with a mean of 31 weeks (range 29–51) later than the methionine changes in the YMDD motif. Indications for a methionine into valine change could be determined in only one of the partial responders. An unexpected observation was the predominant presence of variant virus populations after end of therapy. In ten patients, we detected the wild type virus with a mean of 14 weeks after end of therapy (range 1–42 weeks). In three patients, no variant virus population could be detected at 25, 36 and 37 weeks, respectively, after cessation of treatment. This observation is important for the inclusion of the so-called naive patients in clinical trials. [Copyright &y& Elsevier]