High genetic diversity revealed by the study of TLMV infection in French hemodialysis patients

TT virus-like minivirus (TLMV) was recently discovered as a human circovirus. Little is known about its natural history and molecular epidemiology. A study of TLMV infection is described in a population of French hemodialysis patients. TLMV DNA was tested by seminested PCR system located in the noncoding region in 81 patients divided into seven groups according to the origin of their renal disease. Quantitation of TLMV DNA in serum was carried out. Sequences from 28 patients were compared with 40 sequences retrieved from databases and 53 TLMV sequences cloned from the serum of a single patient. The prevalence of TLMV DNA in hemodialysis patients was 95.1%. In this study, 24 samples (29.6%) presented viral loads of > 125 equivalents of plasmid (Ep)/ml, and only 6 (7.8%) had viral loads of > 125 x 10(2) Ep/ml. A significant correlation (P < 0.029) was found between viral loads of > 125 x 10(2) Ep/ml and the neoplastic origin of end-stage renal disease. Analysis of 53 sequences cloned from a single individual demonstrated high sequence variability, as shown by the genetic distance of 40.2%. This genetic distance is comparable to that between the most divergent sequences of TLMV reported to date (43.5%). These data suggest that TLMV viral load is possibly related to the level of immunocompetence of hemodialysis patients; the genetic diversity of TLMV is extremely high; and co-infection by different strains is possible.     

Mixed infections of adults and children with multiple TTV-like mini virus isolates

Testing of the DNA of TTV-like mini virus (TLMV) was done with serum samples obtained from 184 patients (children and adults) who visited different outpatient clinics at a university hospital in Florianopolis, south of Brazil. TLMV DNA was detected by PCR primers from the non-coding region of the genome. A global TLMV prevalence of 78% was found (94% among children below 11 years). PCR products from three serum samples (patients A-C) were cloned, and the sequences with a length of 201-227 nucleotides were determined for 16-19 clones derived from each of the sera. Among the 16 clones derived from patient C, 15 were identical, and the remaining one had a sequence homology of 99%. In contrast, eight different sequences were obtained among the 19 clones derived from patient A, and 10 distinct sequences were depicted among the 17 clones derived from the serum of patient B. Additionally, 13 clones derived from a saliva sample of patient B were sequenced, and seven different nucleotide sequences obtained. One particular sequence was predominant in both serum (8/17 clones) and saliva (7/13 clones) of patient B. On a phylogenetic tree, sequences derived from patient A (a 6-year-old boy), as well as those derived from patient B (a 24-year-old man), were located in five distinct evolutionary branches, taking a minimum divergence of 5% between branches. This suggested that adults and children are coinfected frequently with several TLMV isolates of different origins.     

High detection rates of TTV-like mini virus sequences in sera from Brazilian blood donors

TT virus (TTV) is an unenveloped virus with a single-stranded, circular DNA genome of 3,818-3,853 nucleotides (nt) that infects humans and non-human primates. Recently, the existence of a novel human virus, TTV-like mini virus (TLMV), that shows a genetic organization similar to that of TTV, but with smaller virion particle and genome, was proposed [Takahashi et al. (2000) Archives of Virology 145:979-993]. To date, no information is available with respect to the prevalence and pathogenicity of TLMV. A sensitive PCR assay was developed by using two oligonucleotide primers (LS2 and LA2) designed from the conserved non-coding region of the TLMV genome. One hundred thirty-seven sera from volunteer Brazilian blood donors were tested and 99 (72%) were TLMV DNA positive. No significant differences were observed between the groups of TLMV positive and negative subjects in relation to sex ratio, seroprevalence of TTV DNA, prevalence of anti-hepatitis A virus antibodies, area of residence, occurrence of daily contact with animals, family income, education level, and level of alanine aminotransferase. The specificity of the PCR assay was demonstrated after cloning of amplification products and determination of the nucleotide sequences (200-228 nt) of clones derived from 23 individuals. When DNAs extracted from TLMV/TTV-coinfected sera were submitted to PCR with LS2 and LA2 primers, the amplification products were derived exclusively from the TLMV genome. A markedly wide range of sequence divergence, even higher than that existent among TTV strains, was noted among TLMV isolates, with a maximum evolutionary distance of 0.80.     

TLMV5＇非编码区基因的克隆与序列分析

目的　调查TTV阴性的非甲～庚型肝炎患者中TTV～like mini virus(TLMV)感染情况,对TLMV5＇非编码区(5＇NCR)部分基因进行分子克隆与序列分析。方法　采用巢式PCR技术检测53例T T V阴性的非甲～庚肝炎患者血清TLMV DNA,对PCR产物进行克隆、测序和分析。结果　53例病例中TLMV DNA阳性37例(69.8％),对其中8株TLMV基因克隆测序,并与Takahashi报道的TLMV序列(GenBank Accession No ab 026930、ab026931)比较,其核苷酸序列同源性在64％～83％之间。结论　在T TV阴性的非甲～庚肝炎患者中存在TLMV感染。TLMV5＇NCR基因变异性较大。TLMV的致病性及其与非甲～庚肝炎的关系尚有待进一步研究。     

病毒性肝炎患者中TLMV DNA检测

[目的]用PCR方法检测临床肝炎患者中TLMV DNA，以初步了解临床肝炎与TLMV的关系。[方法]提取血清中DNA，采用巢式PCR方法，检测血清中TLMV DNA扩增产物。[结果]一般人群、输血员的TLMV DNA阳性率分别为8．33％（6／72）和8．04％（9／112）；急性肝炎和慢性肝炎的TLMV阳性率分别为25．0％（5／20）和19．1％）18／94）。肝炎患者TLMV DNA阳性率与一般人群和输血员比较，二者差异有显著性。[结论]TLMV与临床肝炎呈相关性，TLMV在临床肝炎中的作用有待于深入探讨。     

High prevalence of TT virus-related DNA (90%) and diverse viral genotypes in Norwegian blood donors.

Early estimates of the prevalence of TTV viremia in healthy adults of developed countries were in the order of 1--10 %, while similar estimates in Third World countries were considerably higher. Using three different PCRs, TTV-related DNA was detected in serum from 180 out of the 201 Norwegian blood donors tested, indicating that these viruses are almost universally present in adults. Sequence analysis revealed heterogeneity similar to what is found world-wide. The data suggest that the previous discrepancy in prevalences might be related to a lower serum concentration of virus in developed countries. The high prevalence adds evidence to the benign nature of the virus.     

TLMV的初步流行病学调查

目的克隆中国株TTV样微小病毒(TLMV)的基因序列,进行初步流行病学调查。方法取门诊及住院患者和健康献血员151例,采静脉血5 ml,分离血清制备DNA模板,进行PCR扩增;将PCR产物电泳图在GIS凝胶成像系统上分析。结果提示健康献血员、丙型肝炎患者及慢性乙型肝炎患者体内TLMV含量较高。甲型肝炎、戊型肝炎、未分型肝炎及酒精性肝炎患者体内TLMV含量较低。结论中国人群体内存在TLMV感染。