BACKGROUND: The incidence and etiology of hepatocellular carcinoma (HCC) vary widely according to race and geographic regions. The insertional mutagenesis of adeno-associated virus 2 (AAV2) has recently been considered a new viral etiology of HCC. The aim of this study was to investigate the frequency and clinical characteristics of AAV2 in Korean patients with HCC. METHODS: A total of 289 unrelated Korean patients with HCC, including 159 Hepatitis-B-related cases, 16 Hepatitis-C-related cases, and 114 viral serology-negative cases, who underwent surgery at the Samsung Medical Center in Korea from 2009 to 2014 were enrolled in this study. The presence of AAV2 in fresh-frozen tumor tissues was investigated by DNA PCR and Sanger sequencing. The clinical and pathological characteristics of AAV2-associated HCC in these patients were compared with previous findings in French patients. RESULTS: The AAV2 detection rate in Korean patients (2/289) was very low compared with that in French patients (11/193). Similar to the French patients, the Korean patients with AAV2-related HCC showed no signs of liver cirrhosis. The Korean patients were younger than the French patients with the same AAV2-associated HCC; the ages at diagnosis of the two Korean patients were 47 and 39 yr, while the median age of the 11 French patients was 55 yr (range 43-90 yr). CONCLUSIONS: AAV2-associated HCC was very rare in Korean patients with HCC. Despite a limited number of cases, this study is the first to report the clinical characteristics of Korean patients with AAV2-associated HCC. These findings suggest epidemiologic differences in viral hepatocarcinogenesis between Korean and European patients.
Adult ; Asian Continental Ancestry Group ; Capsid Proteins/genetics ; Carcinoma, Hepatocellular/etiology/*pathology/virology ; DNA, Viral/chemistry/genetics/metabolism ; DNA-Binding Proteins/genetics ; Dependovirus/*genetics/isolation & purification/pathogenicity ; Female ; Humans ; Incidence ; Inverted Repeat Sequences/genetics ; Liver Neoplasms/etiology/*pathology/virology ; Male ; Middle Aged ; Parvoviridae Infections/complications/epidemiology ; Polymerase Chain Reaction ; Republic of Korea ; Sequence Analysis, DNA ; Viral Proteins/genetics

To analyze the molecular basis of the variation of the pathogenicity of the influenza B virus, we rescued a recombinant virus with a deletion in the carboxyl terminal of the NS1 protein using reverse genetics based on the parental virus B-S9 of B/Yamagata/16/88. A mutant strain with a deletion of 171 amino acids in the carboxyl terminal of the NS1 protein was named "B-L5". BALB/c mice were inoculated with 3 X 105 EID50 of B-L5 and the parental virus B-S9, respectively. Then, weight changes, survival, and viral titers were documented. During 3 days post-inoculation (dpi) to 7 dpi, the weight of mice infected with B-S9 decreased. However, the weight of mice infected with B-L5 showed weight decreases only at 2 dpi, and quickly recovered at 3 dpi. B-S9 and B-L5 could replicate in the lungs of BALB/c mice. However, viral titers in the lungs of mice infected with B-L5 were 7900-times lower than those of mice infected with B-S9 at 3 dpi. Viral titers in the lungs of mice infected with B-L5 were not detected at 6 dpi. These results showed that, compared with the parent virus B-S9, the mutant virus B-L5 showed lower pathogenicity in BALB/c mice. Our study suggests that deletion of the carboxyl terminal of the NS1 protein decreases the pathogenicity of the influenza B virus. Establishment of a reverse-genetics system for the B influenza virus will provide a platform for studying its pathogenesis, and mechanism of transmission, and for developing live-attenuated influenza B virus vaccines.
Animals ; Body Weight ; Dogs ; Female ; HEK293 Cells ; Humans ; Influenza B virus ; genetics ; pathogenicity ; physiology ; Madin Darby Canine Kidney Cells ; Mice ; Mice, Inbred BALB C ; Sequence Deletion ; Survival Analysis ; Viral Load ; genetics ; Viral Nonstructural Proteins ; chemistry ; genetics ; Virulence

To study the complete genomic sequence, genomic characteristics, and genetic variation of the bovine papillomavirus 2 genotype (BPV-2) Aks-01 strain at the molecular level, genotyping of this strain from the skin samples of cows in southern Xinjiang (China) was first detected by the polymerase chain reaction with FAP59/FAP64 primers. Based on the complete genome of the BPV-2 reference strain, specific primers and sequencing primers were designed, and the complete genome of the Aks-01 strain amplified and sequenced. Sequence analyses showed that genotyping of the Aks-01 strain belonged to BPV-2. The Aks-01 strain had the structural characteristics of BPV-2. The 7944-bp full-length genomic sequence of the Aks-01 strain was compiled using DNAStar™. The sequence of the Aks-01 strain had 98% similarity to the reference strain from GenBank. The Aks-01 strain was most closely related to BPV-1 and BPV-13. BPV-2, BPV-1 and BPV-13 were grouped within the genus Deltapapillomavirus. The Aks-01 strain is the first BPV-2 strain reported in southern Xinjiang.
Amino Acid Sequence ; Animals ; Base Sequence ; Bovine papillomavirus 1 ; genetics ; Cattle ; China ; Evolution, Molecular ; Female ; Genome, Viral ; genetics ; Genomics ; Genotype ; Molecular Sequence Data ; Oncogene Proteins, Viral ; chemistry ; genetics ; metabolism ; Phylogeny ; Sequence Analysis, DNA ; Skin ; virology

The rabies virus (RABV) is an enveloped RNA virus. It mainly damages the central nervous system and causes anencephaly in mammals and humans. There is now compelling evidence that enveloped virions released from infected cells can carry many host proteins, some of which may play an important part in viral replication. Several host proteins have been reported to be incorporated into RABV particles. However, a systematic study to reveal the proteomics of RABV particles has not been conducted. In the present study, after virus culture and purification by sucrose density gradient ultracentrifugation, a proteomics approach was used to analyze the protein composition of purified RABV particles to understand the molecular mechanisms of virus-cell interactions. Fifty host proteins, along with five virus-encoded structural proteins, were identified in purified RABV particles. These proteins could be classified into ten categories according to function: intracellular trafficking (14%), molecular chaperone (12%), cytoskeletal (24%), signal transduction (8%), transcription regulation (12%), calcium ion-binding (6%), enzyme binding (6%), metabolic process (2%), ubiquitin (2%) and other (14%). Of these, four proteins (beta-actin, p-tubulin, Cofilin, Hsc70) were validated by western blotting to be present in purified RABV particles. This novel study of the composition of host proteins in RABV particles may aid investigation of the mechanism of RABV replication.
Animals ; Humans ; Molecular Sequence Data ; Proteomics ; Rabies ; genetics ; metabolism ; virology ; Rabies virus ; chemistry ; genetics ; metabolism ; Viral Proteins ; analysis ; chemistry ; genetics ; metabolism ; Virion ; chemistry ; genetics ; metabolism

To identity the pathogen that causes the mosaic and yellowing symptoms on Atractylodes macrocephala Koidz in Jiangxian, Shanxi province, biological inoculation, sequence-independent amplification (SIA),RT-PCR and other identification methods were used. The results showed that the chlorotic and necrosis symptoms occurred in the indicator plant Chenopodium quinoa after it was infected with the pathogen,and the same symptoms appeared after the reinoculation of healthy Atractylodes macrocephala Koidz; this reflected that the disease was likely to be caused by a virus. The results of SIA and sequencing showed that Broad bean wilt virus 2 (BBWV2) was present in severely mosaic Atractylodes macrocephala Koidz leaves. To further characterize the BBWV2 isolate from Atractylodes macrocephala (BBWV2-Am), the polyprotein partial gene encoded by BBWV2-Am RNA2 was cloned and sequenced. Sequence alignments showed that the nucleotide sequence identity of BBWV2-Am SCP and LCP genes ranged from 79.3% to 87.2% and from 80.1% to 89.2% compared to other BBWV2 strains,respectively; the deduced amino acid sequence similarities of the two gene products ranged from 91.2% to 95.7% and from 89.44 to 95.5%, respectively,compared to those of other BBWV2 strains. Phylogenetic comparisons showed that BBWV2-Am was most likely to be related to BBWV2-Rg,but formed an independent branch. This is the first report of BBWV2 in Atractylodes macrocephala Koidz.
Amino Acid Sequence ; Atractylodes ; virology ; Fabavirus ; chemistry ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Plant Diseases ; virology ; Sequence Analysis ; Viral Proteins ; chemistry ; genetics

In early 2011, the serious outbreak of porcine pseudorabies virus (PRV) infection suddenly recurred in Henan and neighboring Provinces. To investigate the etiology of massive infection with PRV, 16 800 serum samples, 905 porcine epidemic diarrhea virus (PEDV) back-feeding tissues, and 56 PR gene deleted live vaccines were colleted from January 2011 to May 2013 to detect PRV field infection using a PRV gE antibody test kit. The gE and TK genes of 11 new epidemic PRV strains were sequenced by PCR, and their molecular characteristics were analyzed. Moreover, virus titer determination, protective test against PRV, and vaccine potency testing were performed. The results showed that the detection rate of PRV field infection-positive pig farms was 68.06%, and the overall positive rate of PRV field infection in serum was 38.47%; the positive rates in breeding sows, breeding boars, reserve pigs, and commercial pigs were 40.12%, 30.88%, 54.67%, and 26.52%, respectively. The new epidemic strains were in the same evolutionary branch and belonged to the virulent strain group. Compared with the classical PRV strain, the virulence of new epidemic strains changed a little. The length of gE gene was 1 787 bp, and the length of TK gene was 963 bp. The nucleotide homologies of gE and TK genes to Chinese reference strains were 98.2%-99.8% and 98.90%-99.6%, respectively, and the amino acid homologies were 97.1%-99.8% and 97.5%-99.4%, respectively. Commercial vaccine had a 100% protective effect against the new epidemic strains. The positive rate of PRV field infection was 0% in vaccine and 40.44% in back-feeding tissues. The results confirmed that PRV field infection rates were rising sharply among pigs in Henan and neighboring Provinces after 2011. The main virulence genes of new epidemic PRV strains did not change significantly over the years. PR gene deleted live vaccines had no PRV field infection and could completely resist the attack of new strains. The virus carriage of breeding boars and reserve pigs and the serious PRV field infection in PEDV back-feeding tissues were the main causative factors for massive infection with PRV and epidemic outbreak in Henan and neighboring Provinces from 2011 to 2013.
Amino Acid Sequence ; Animal Feed ; analysis ; virology ; Animals ; China ; epidemiology ; Epidemics ; Female ; Herpesvirus 1, Suid ; chemistry ; classification ; genetics ; isolation & purification ; Male ; Molecular Sequence Data ; Phylogeny ; Pseudorabies ; epidemiology ; virology ; Sequence Alignment ; Sequence Homology, Amino Acid ; Sus scrofa ; Swine ; Swine Diseases ; epidemiology ; virology ; Viral Proteins ; chemistry ; genetics

The glycoprotein 3 (GP3) of type II porcine reproductive and respiratory syndrome virus has the characteristic domains of a membrane protein. However, this protein has been reported to be retained in the endoplasmic reticulum (ER) rather than transported to the plasma membrane of the cell. In this study, we performed confocal laser scanning microscopy analysis of variants of GP3 and foundthat the signal sequence of the GP3 led to confinement of GP3 in the ER, while the functional ortransmembrane domain did not affect its localization. Based on these results, we concludedthat the signal sequence of GP3 contains the ER retention signal, which might play an important role in assembly of viral proteins.
Animals ; Cell Line ; Cell Membrane/*metabolism/virology ; Cricetinae ; Endoplasmic Reticulum/*metabolism/virology ; Microscopy, Confocal/veterinary ; Plasmids/genetics/metabolism ; Porcine respiratory and reproductive syndrome virus/*genetics/metabolism ; *Protein Sorting Signals ; Sequence Analysis, Protein/veterinary ; Viral Envelope Proteins/chemistry/*genetics/metabolism

To gain more insights into epidemiologic characteristics and genotype of hantavirus in Apodemus agrarius in Changbai Area. Complete hantavirus S segment sequences were amplified by RT-PCR and sequenced. The phylogenetic trees were constructed for analysis of genetic characters of hantavirus. A total of 58 Apodemus agrarius were trapped in the epidemic areas, and complete hantavirus S segment sequences were obtained from 4 lung samples of these rodents (6. 90%0). Phylogenetic analysis of the four S segment sequences indicated that all viruses isolated from Apodemu sagrarius were closely related to genotype 6 of Hantaan virus (95. 8%-96. 3%, nucleotide identity; 98. 6%-99. 5%, amino acid identity), all of them had a specific S387 different from other genotypes of Hantaan virus.
Animals ; Base Sequence ; China ; epidemiology ; DNA, Complementary ; chemistry ; genetics ; Disease Reservoirs ; virology ; Genotype ; Hantavirus ; classification ; genetics ; isolation & purification ; Hantavirus Infections ; epidemiology ; veterinary ; virology ; Lung ; virology ; Murinae ; virology ; Phylogeny ; RNA, Viral ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Rodent Diseases ; virology ; Sequence Analysis, DNA ; Viral Proteins ; genetics

Classical swine fever virus (CSFV) causes a highly contagious disease among swine that has an important economic impact worldwide. CSFV strain LOM is an attenuated virus of low virulent strain of Miyagi isolated from Japan in 1956. Eight DNA fragments representing the genome of the CSFV strain LOM were obtained by RT-PCR. These were used to determine the complete nucleotide sequence and construct a full-length cDNA clone which was called Flc-LOM. Sequence analysis of the recombinant clone (Flc-LOM) revealed the presence of eight mutations, resulting in two amino acid substitutions, when compared to the parental sequence. RNA transcripts of both LOM and Flc-LOM were directly infectious in PK-15 cells. The rescued Flc-LOM virus grew more slowly than the parental virus, LOM, in the cells. Intramuscular immunization with Flc-LOM was safe and highly immunogenic in pigs; no clinical signs or virus transmission to sentinel animals were observed after 35 days. CSFV-specific neutralizing antibodies were detected 14 days post-infection. After challenge with the virulent CSFV strain SW03, pigs immunized with Flc-LOM were shown to be fully protected. Thus, our newly established infectious clone of CSFV, Flc-LOM, could serve as a vaccine candidate.
Animals ; Antibodies, Viral/blood ; Base Sequence ; Cell Line ; Classical Swine Fever/immunology/*virology ; Classical swine fever virus/*genetics/immunology/pathogenicity ; Cloning, Molecular ; DNA, Complementary/genetics/immunology ; Immunization/methods/standards/veterinary ; Molecular Sequence Data ; Neutralization Tests/veterinary ; RNA, Viral/chemistry/genetics ; Recombinant Proteins/immunology ; Reverse Transcriptase Polymerase Chain Reaction/veterinary ; Sequence Analysis, DNA ; Specific Pathogen-Free Organisms ; Swine ; Virulence

This paper investigated the envelope protein E1/E2 quasispecies genetic characterization of 4 HCV positive sera (Genotype 1b: 274, 366, 383; Genotype 2a: 283) in China. Nucleotide acid was extracted and glycoprotein E1/E2 (191-764aa) coding genes were obtained by RT-PCR, positive clones were randomly selected for sequencing. The phylogenetic relationships and the homology of nucleotide and amino acid were analyzed based on E1/E2 coding genes, and some vital functional regions of E1/E2 were characterized. A total of 43 sequences (274: 10; 283: 12; 366: 13; 383: 8) were obtained showing high genetic heterogeneity in HVR1 and HVR2 regions, while sequences of the neutralizing epitopes, transmembrane domain I, II and N-terminal ectodomain were comparatively conservative. Single base (C) insertion mutation at nt1279 ( E1 region, aa313), resulting in a mutated E1 coding protein (beginning at aa 313) and interruption at N terminus (aa 398) of HVR1 region of E2, was dominant quasispecies sequence(11/12) found in serum 283 . This is the first report on E1/E2 quasispecies in Chinese HCV patients and this novel pattern of insertion mutation provides important information for further study on HCV pathogenesis and immune evasion.
Amino Acid Sequence ; Base Sequence ; DNA Mutational Analysis ; Hepacivirus ; genetics ; pathogenicity ; Hepatitis C ; virology ; Humans ; Molecular Sequence Data ; Mutagenesis, Insertional ; Viral Envelope Proteins ; chemistry ; genetics