Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis. This study was conducted to clarify the molecular characteristics of BLVs obtained from a specific region in Korea. Proviral BLVs were detected in anti-BLV antibody-positive blood samples by PCR. Env and gag fragments were sequenced and compared to previously published reference sequences. Analysis of the env gene sequence revealed that the YI strain was highly similar to genotype 1, including United States and Japanese strains. The gag gene sequence had the highest degree of similarity with a Japanese strain.
Animals ; Asian Continental Ancestry Group ; Cattle* ; Enzootic Bovine Leukosis ; Genes, env ; Genes, gag* ; Genotype ; Humans ; Korea ; Leukemia Virus, Bovine* ; Polymerase Chain Reaction ; United States

BACKGROUND: Xenotropic murine leukemia virus-related virus (XMRV) has been detected in peripheral blood mononuclear cells (PBMNs), therefore, it has been regarded as being infectious and transmittable by transfusion. Thus, we attempted to detect XMRV in blood samples in order to confirm the absence of XMRV from blood donors. METHODS: We achieved 165 blood donors and four chronic fatigue syndrome (CFS) patients. We performed real-time polymerase chain reaction using the LightCycler 480 (Roche, Penzberg, Germany) for the gag and env genes of the XMRV genome. DNA was extracted from peripheral blood samples. We used Uracil-N-Glycosylase in order to prevent contamination and DNA extracted from mouse embryonic fibroblasts (MEF) for amplification control. RESULTS: No XMRV was detected in any of the blood donors in both the gag and env genes. In four CFS patients, amplification was not detected in the gag gene. In two of four CFS patients, amplifications were detected and the melting temperature was in agreement with that of MEF control in the env gene. CONCLUSION: Although XMRV was not present in blood samples from blood donors, this is the first report on XMRV in Korean blood donors. We confirmed the absence of XMRV in Korean blood donors, the same as studies reported in other countries.
Animals ; Blood Donors ; DNA ; Fatigue Syndrome, Chronic ; Fibroblasts ; Freezing ; Genes, env ; Genes, gag ; Genome ; Humans ; Mice ; Real-Time Polymerase Chain Reaction ; Xenotropic murine leukemia virus-related virus

Xenotransplantation, as a potential solution to the shortage of human organs, is associated with a number of concerns including immunologic rejection and xenogenic infection. While the pigs are considered the most suitable organ source for xenotransplantation, there is a potential public health risk due to zoonosis. Among the known porcine zoonotic microbes, Porcine Endogenous Retrovirus (PERV) is the most considerable virus. PERV belongs to the Gammaretrovirus and has been divided into three groups (A, B, and C). To characterize the gag of PERVs, we isolated the genomic DNAs from three pig breeds (Birkshire, Duroc, and Yorkshire) and two types of SPF miniature pigs. About 1.5 kb fragments covering full length of gag were amplified and cloned into T-vector. A total of 38 clones were obtained and sequenced. Nucleotide sequences were analyzed and phylogenetic trees were constructed from the nucleotide and deduced amino acids. PERV-A, -B and -C were present in the proportion of 47, 19 and 34%, respectively. Regardless of origin or subgroups, gag clones showed highly homology in nucleotide and deduced amino acid sequences. Deduced amino acids sequence alignments showed typical conserve sequences, Cys-His box and processing sites. Among analyzed clones, about 28% of isolates had the correct open reading frame. To test the functional expression of Gag protein, gag was subcloned into expression vector and confirmed its expression in HeLa cell. This research provides the fundamental information about molecular characteristics of gag gene and functional Gag protein related xenotropic PERVs.
Amino Acid Sequence ; Amino Acids ; Base Sequence ; Clone Cells ; DNA ; Endogenous Retroviruses* ; Gammaretrovirus ; Gene Products, gag ; Genes, gag* ; HeLa Cells ; Humans ; Korea* ; Open Reading Frames ; Public Health ; Sequence Alignment ; Swine* ; Transplantation, Heterologous

OBJECTIVETo assess the infectivity of porcine endogenous retrovirus (PERV) via in vitro infection of human embryonic kidney cell line HEK-293.

METHODSPERV particles were detected by immunoelectron microscopy. PERV DNA and mRNA were studied in HEK-293 24 hours after the infection using polymerase chain reaction and reverse transcriptase-PCR respectively. The PERV types were also analyzed. PERV-gag protein was observed by confocal microscopy.

RESULTSRetroviral particles were round under electron microscope. PERV-gag pol gene and gag protein were detected and expressed in the infected HEK-293 cells. The types of PERV were PERV-A and PERV-B. PERV-gag protein was also identified in the cytoplasm of infected cells by confocal microscopy.

CONCLUSIONSPERV is able to infect HEK-293 cell line in vitro; types of PERV-gag protein is also expressed as a result. Further studies are thus necessary in order to evaluate the possibility of xenozoonoses in pig-to-human xenotransplantation.

Animals ; Cell Line ; DNA, Viral ; analysis ; Embryo, Mammalian ; Endogenous Retroviruses ; isolation & purification ; pathogenicity ; Gene Amplification ; Gene Products, gag ; biosynthesis ; genetics ; Genes, gag ; Humans ; Kidney ; metabolism ; virology ; RNA, Messenger ; biosynthesis ; genetics ; Swine

OBJECTIVETo study the epidemic status of human immunodeficiency virus type 1 (HIV-1) subtypes in Shenzhen and to study their transmission source and routes.

METHODSHIV-1 env and gag genes were amplified by nested PCR from uncultured peripheral blood mononuclear cells (PBMCs) obtained from 122 HIV-1 carriers confirmed in Shenzhen. The C2-V3 region (about 450 bp) of HIV-1 env and P17/ P24 region were sequenced.

RESULTSAmong 122 samples, 6 HIV-1 strains including 3 circulating recombinant forms (CRFs) of CRF01_AE, CRF08_BC, CRF07_BC and 3 subtypes of B', B, C were found in Shenzhen, and the percentages were 45.1% (55/122) for CRF01_AE, 31.1% (38/122) for CRF08_BC, 6.6% (8/122) for CRF07_BC, 14.8% (18/122) for B' subtype, 1.6% (2/122) for B subtype, and 0.8% (1/122) for C subtype. The intragroup genetic distances were (4.455 +/- 1.478)%, (2.997 +/- 1.345)%, (4.380 +/- 2.024)%, (5.186 +/- 2.487)%, and (4.869 +/- 2.638)%, respectively. In comparison with the sequence of respective international strains 01AE. TH. 90. CM240, 97CNGX-9F, CN. 97. C54A, B. US. 83. JRFL, and RLA2, the genetic distances were (5. 228 +/- 0.823)%, (3.634 +/- 1.073)%, (4.233 +/- 1.119)%, (4.950 +/- 2.564)%, and (5.795 +/- 2.198)%, respectively. The major subtypes found in injection drug users (IDUs) were CRF07_BC, CRF08_BC, and CRF01_AE strains. CRF01_AE and B' strains were epidemic mainly in sexual workers.

CONCLUSIONThere are 3 HIV-1 subtypes (B', B, C) and 3 CRFs (CRF01_AE, CRF08_BC, CRF07_BC) epidemics in Shenzhen. The predominant subtypes varies among different transmission routes. While CRF01_AE is predominant among sexual workers, CRF08_BC and CRF01_AE are major subtypes among IDU population.

Adolescent ; Adult ; China ; epidemiology ; Female ; Genes, env ; genetics ; Genes, gag ; genetics ; Genes, pol ; genetics ; HIV Infections ; epidemiology ; HIV-1 ; genetics ; Humans ; Male ; Middle Aged ; Molecular Epidemiology ; Polymerase Chain Reaction

PCR implication using the primers for gag, pol and rev genes in BLV (bovine leukemia virus) proviral DNA and syncytium assay were carried out for the Korean native goats experimentally infected with bovine leukemia virus to investigate pathogenesis of BLV in the goats, and to establish a model animal for BLV infection. The oligonucleotide primers used in PCR revealed very high specificity, The minimal amount of FLK-BLV cellular chromosomal DNA to detect the integrated BLV proviral DNA was 10 ng. The peripheral blood lymphocytes from the goat infected with BLV were examined at regular intervals by PCR amplification and syncytium assay. Pol or gag genes were detected in none of three infected goats at the 1st week post-infection (p.i.). At the 4th week p.i., one of three goats showed the amplified gag gene. Thereafter detection rates for the genes were increased, indicating that the BLV proviral genes were integrated in all of the lymphocytes from three goats, at the 16th weeks p.i., when it was evident in syncytium assay that the lymphocytes from all of three goats were infested with infective BLV. Investigating the tissues from the necropsied goats at the 8th month p.i., the amplified BLV proviral genes and infective BLV were detected in all of the peripheral lymphocytes from three infected-goats. Among various tissues examined, the amplified BLV proviral genes were observed in spleen and superficial cervical, mandibular and retropharyngeal lymph nodes, and the infective BLV, in superficial cervical and mandibular lymph nodes. It was assumed that the Korean native goat was quite susceptible to BLV infection, indicating that the goat could be a good model animal for BLV.
Animals ; Cattle ; Deltaretrovirus Infections ; DNA Primers ; DNA* ; Enzootic Bovine Leukosis* ; Genes, gag ; Genes, rev ; Giant Cells ; Goats* ; Leukemia ; Leukemia Virus, Bovine* ; Lymph Nodes ; Lymphocytes ; Polymerase Chain Reaction* ; Sensitivity and Specificity ; Spleen

The late stages of the HIV-1 replication cycle are important to the overall replication cycle. During the late stages, HIV-1 replication undergoes the processes of assembly, release, and maturation, resulting in the production of a mature virus particle capable of infecting a new target cell. The structural protein Gag and its related gene (protein) play a central role in these pathways. The different regions of Gag worked in concert to drive production of a mature infectious particle through protein-protein, protein-RNA and protein-lipid interactions. The designed drug aimed directly at these stages can efficiently block the maturation and infectivity of HIV-1. In this article, the role of structural protein Gag and related gene (protein) in late stages of the HIV-1 replication cycle and related inhibitors is reviewed.
Amphotericin B ; analogs & derivatives ; chemistry ; pharmacology ; Anti-HIV Agents ; chemistry ; pharmacology ; Benzeneacetamides ; chemistry ; pharmacology ; Furans ; chemistry ; pharmacology ; Genes, gag ; HIV-1 ; drug effects ; physiology ; Humans ; Phenylurea Compounds ; chemistry ; pharmacology ; Piperidines ; chemistry ; pharmacology ; Succinates ; chemistry ; pharmacology ; Sulfur Compounds ; chemistry ; pharmacology ; Triterpenes ; chemistry ; pharmacology ; Virus Assembly ; drug effects ; Virus Release ; drug effects ; Virus Replication ; drug effects ; physiology ; gag Gene Products, Human Immunodeficiency Virus ; metabolism ; physiology

STUDY DESIGN: A molecular biological study of intercostal muscles and intervertebral disc cells of Korean scoliosis patients. OBJECTIVES: To study the pathological results of intercostal muscles and molecular biological activity of intervertebral disc cells of the scoliotic major curve in Korean patients. SUMMARY OF LITERATURE REVIEW : The cause of idiopathic scoliosis has been investigated in terms of many parameters. Although, molecular biological studies of intercostal muscles and intervertebral disc cells have been performed in foreign countries, few studies have been conducted in Korea. MATERIALS AND METHODS: Ten patients, one male and nine female, who underwent thoracoscopic surgery were reviewed. The age range was 13 to 23 years old. Intercostal muscles were taken from the portal site of the major curve (1x1 cm sized). Ten tissues were stained with H/E and ATPase immunohistochemical staining. An appropriate amount of intervertebral disc was taken from the major curve of three scoliotic patients and each concentration of collagen type I, II, GAG gene and proteoglycan synthesis activity was measured. The results were compared with those of grade 0 and grade II degenerative change on each MRI. RESULTS: The intercostal muscle of scoliotic patients showed 60.4+/-8.4% in type I muscle fiber and 39.6+/-8.8% in type II-A. These results were not different from those of previous studies. The size of muscle fiber was 48-65 microns, which was slightly smaller than the absolute value, but the difference was not statistically significant. The amount of produced proteoglycans was slightly higher in the intervertebral disc cells of scoliotic patients, the total amount of collagen was significantly lower and there was a difference in the production of type II collagen. CONCLUSIONS: The intercostal muscles were not affected by the muscle of scoliotic patients and there was no molecular biological significant difference between control and scoliotic patients. We can assume that scoliosis was not caused by problems of intervertebral disc or intercostal muscles.
Adenosine Triphosphatases ; Adolescent* ; Collagen ; Collagen Type I ; Collagen Type II ; Female ; Genes, gag ; Humans ; Intercostal Muscles* ; Intervertebral Disc* ; Korea* ; Magnetic Resonance Imaging ; Male ; Molecular Biology ; Proteoglycans ; Scoliosis* ; Thoracoscopy ; Young Adult

OBJECTIVETo study the distribution of human immunodeficiency virus (HIV)-1 genotypes in major prevalent regions of China and to illustrate the relationship between HIV-1 subtypes and mother-to-child transmission in a retrospective cohort.

METHODSHIV-1 gag p17 and env C2-V4 region were amplified by nested-polymerase chain reaction (nPCR) and the sequences were obtained by sequencing gag nPCR products or clones of env gene.

RESULTS60 HIV-1 positive individuals were subject to typing for gag p17 and 69 for env C2-V4 region. Single clade was only found in Henan (subtype B') and Xinjiang (subtype C), and subtypes C and E were demonstrated in Yunnan. These regions represented most of the HIV-1 infections in China. Multiple subtypes (A, B, C, E, etc.) were found in Beijing and Shanghai, where HIV infections were still in low level. The sequences of subtype C were less diversive in Xinjiang (p17: 0.0192 +/- 0.0078, C2-V4: 0.0455 +/- 0.0145) than in Yunnan (p17: 0.0279 +/- 0.0102, C2-V4: 0.0482 +/- 0.0171), but all of them clustered in "C" branch in phylogenetic trees. Trafficking of subtype C from Yunnan to Xinjiang was found but had already been reported by others. Compared to subtype C, subtype E was quite divergent (p17: 0.0473 +/- 0.0105, C2-V4: 0.1114 +/- 0.0112) in Yunnan, but no recombination was found in the C2-V4 region of env gene. Highe divergence of subtype B' was found in Henan and the peripheral provinces (p17: 0.0381 +/- 0.0101, C2-V4: 0.0691 +/- 0.0166), which might be attributed to the early epidemics of HIV-1 in these areas (early 1990's). In maternal-child cohort, subtypes B (7/21), C (11/21), E (1/21) and undefined types (2/21) were identified in non-transmitting HIV-1 positive mothers, while only subtype B (7/11) and C (4/11) appeared in transmitting HIV-1 positive mothers. The rate of transmission was 53.8% (7/13) in mothers infected with subtype B and 30.8% (4/13) in those infected with subtype C, but with no significant difference (P = 0.196). The imbalancing distribution of subtypes might be explained by the fact that transfusion or illegal blood would increased mother-to-child transmission on HIV-1 and most of mothers with clade B were infected by illegal blood transfusion in this cohort. In addition, most of the maternal-child pair's sequences clustered in gag or env phylogenetic trees but only a few did disperse among the unrelated patients because children were older (>/= 4 years).

CONCLUSIONThe characteristics of HIV-1 clade's distribution differed over most parts of China but no difference was demonstrated between subtype B and C in mother-to-child transmission on HIV-1.

Child, Preschool ; China ; epidemiology ; Cohort Studies ; Female ; Gene Products, env ; genetics ; Genes, gag ; genetics ; Genotype ; HIV Infections ; epidemiology ; transmission ; virology ; HIV-1 ; classification ; genetics ; Humans ; Infant ; Infectious Disease Transmission, Vertical ; Male ; Phylogeny ; Retrospective Studies ; Transfusion Reaction

OBJECTIVEThe inactivating mutation of thyrotropin receptor (TSHR) gene results in partial or complete insensitivity of thyrotropin (TSH) and dysfunction of the TSH-TSHR-cAMP cascade. Therefore, it may cause congenital hypothyroidism (CH). Depending on the degree of impairment of TSHR function, patients can present with subclinical hypothyroidism at one extreme of the spectrum, or severe hypothyroidism at the other. This study aimed to understand the relation between inactivating mutations of TSHR gene and Chinese children with CH.

METHODS(1) Seventy-nine Chinese children with CH, including 14 subclinical hypothyroidism patients (8 boys and 6 girls, age 1 - 5.5 years) and 65 hypothyroidism patients (27 boys and 38 girls, age 1.5 - 6 years) were enrolled in this study. Meanwhile, 100 normal children were enrolled as control, 40 were male and 60 were female. The age of the normal children were at a range of 1 - 8 years. (2) Total genomic DNA was extracted from peripheral blood leukocytes of the 79 patients and 100 normal subjects. Exons 1 - 10 of TSHR gene were individually amplified by polymerase chain reaction (PCR) and mutations were detected by direct sequencing.

RESULTS(1) A compound heterozygous missense mutations (Pro52Thr/Val689Gly) and a heterozygous missense mutation (Gly245Ser) were detected in 79 patients. The mutations of Pro52Thr and Gly245Ser were located within the extracellular domain of TSHR, while Val689Gly was located within the intracellular domain of TSHR. In 30 patients the normal cytosine at position 2181 in exon 10 was replaced by a guanine (GAC-->GAG), resulting in the replacement of Glu(727) by Asp. In 47 patients, the normal thymidine at position 561 in exon 7 was replaced by a cytosine (AAT-->AAC). This substitution did not change the amino acid (Asn) at position 187. (2) In 33 normal children the normal cytosine at position 2181 in exon 10 was also replaced by a guanine (GAC-->GAG) and in 50 normal children the normal thymidine at position 561 in exon 7 was replaced by a cytosine (AAT-->AAC).

CONCLUSIONSThree heterozygous missense mutations (Pro52Thr, Gly245Ser, Val689Gly) of TSHR gene were firstly detected in Chinese children with CH. There was a polymorphism in exon 10 at nucleotide 2181 (GAC-->GAG) and in exon 7 at nucleotide 561 (AAT-->AAC) in TSHR gene. The inactivating mutation of TSHR gene is an infrequent pathogeny for CH.

Amino Acid Substitution ; genetics ; Asian Continental Ancestry Group ; Child ; Congenital Hypothyroidism ; genetics ; DNA ; analysis ; Exons ; genetics ; Female ; Gene Silencing ; Genes, gag ; genetics ; Humans ; Hypothyroidism ; genetics ; Male ; Mutation ; Mutation, Missense ; genetics ; Polymorphism, Genetic ; genetics ; Receptors, Thyrotropin ; metabolism ; Thyrotropin ; genetics