ABSTRACT
Leishmaniasis is a neglected tropical parasitic disease affecting a large number of countries in the world. Early diagnosis of Leishmania infections is essential for therapeutic reasons, as it can decrease morbidity and mortality. L. siamensis and L. martiniquensis are novel Leishmania species recently described in Thailand and Myanmar. The disease is usually found in immunocompromised patients, especially those who have AIDS. Currently, the diagnosis of Leishmania infection in Thailand relies on microscopy, microbial culture, and polymerase chain reaction (PCR). In this study, we established a quantitative PCR (qPCR) method for detection of L. martiniquensis DNA in various types of clinical specimens, including whole blood, buffy coat, saliva, and urine of L. martiniquensis infected patients. The results of the qPCR assay were positive in all saliva samples. The assay is therefore effective to detect L. martiniquensis DNA even in noninvasive specimens, and it could be used for the diagnosis, follow up, and survey of L. martiniquensis infections.
ABSTRACT
The outbreak of the human pandemic influenza A (H1N1) has caused a considerable public concern. The aim of this review was to improve our understanding of this novel virus by analyzing the relationships between its molecular characteristics and pathogenic properties. Results of this analysis indicate that the human pandemic influenza A (H1N1) virus is a new re-assorted virus, which combines genetic materials from the avian flu (H1N1) virus, classical swine flu (H1N1) virus, human flu (H3N2) virus, and Eurasian swine flu (H1N1) virus. Analysis of the sequences for receptor-binding and cleavage sites of hemagglutinin (HA), stalk region of neuraminidase (NA), non-structural protein 1 (NS1), polymerase basic protein 2 (PB2), and polymerase basic protein 1 (PB1) suggested that (i) the human pandemic influenza A (H1N1) virus is a low virulent and low pathogenic virus, (ii) its replication is restricted to the cells of upper respiratory tract, so it does not lead to a systemic infection, (iii) it spreads among humans only, (iv) its replication could be inhibited by oseltamivir, zanamivir, interferon (IFN), and tumor necrosis factor alpha (TNF-alpha). A potential application of amantadine might be complicated by the drug-resistant virus strains.
Subject(s)
Influenza A Virus, H1N1 Subtype/genetics , Influenza A Virus, H1N1 Subtype/pathogenicity , Amino Acid Sequence , Antiviral Agents/pharmacology , Disease Outbreaks , Humans , Influenza A Virus, H1N1 Subtype/chemistry , Influenza A Virus, H1N1 Subtype/isolation & purification , Influenza A virus/chemistry , Influenza A virus/drug effects , Influenza A virus/genetics , Influenza A virus/isolation & purification , Influenza, Human/epidemiology , Influenza, Human/virology , Molecular Sequence Data , Sequence Alignment , Viral Proteins/geneticsSubject(s)
Blood Donors , Parvoviridae Infections/virology , Parvovirus/isolation & purification , Substance Abuse, Intravenous/virology , Female , Genotype , Humans , Male , Molecular Sequence Data , Parvoviridae Infections/blood , Parvovirus/classification , Parvovirus/genetics , Phylogeny , Substance Abuse, Intravenous/blood , ThailandABSTRACT
In a recent study of hepatitis A virus (HAV) in Thailand, viral isolates recovered during several outbreaks of infection that occurred between 2001 and 2005 were genotyped and subjected to phylogenetic analysis. Anti-HAV IgM was detected, by ELISA, in many of the 283 serum samples that were collected from the provinces of Suphanburi, Songkhla, Chiangrai and Lampang: 40 (48.2% of those investigated), 38 (47.5%), 25 (41.0%) and 32 (54.2%), respectively. The HAV RNA in the positive samples was reverse transcribed and amplified, using a nested PCR focussed on the VP1-2A region, before the nucleotides of the VP1-2A region of each HAV-RNA-positive sample were sequenced. All the isolates investigated clustered in subgenotype IA and, hence, are closely related to the strains previously investigated in Thailand. When the genome of one sample from an outbreak in Lampang (LP014) was fully sequenced, the results of genome comparison and phylogenetic analysis again indicated subgenotype 1A, which appears to be the predominant form of HAV circulating throughout Thailand.
Subject(s)
Disease Outbreaks , Hepatitis A Virus, Human/genetics , Hepatitis A virus/genetics , Hepatitis A/virology , Adolescent , Adult , Base Sequence , Genotype , Hepatitis A/epidemiology , Hepatitis A Antibodies/blood , Hepatitis A Virus, Human/immunology , Hepatitis A Virus, Human/isolation & purification , Hepatitis A virus/classification , Humans , Molecular Epidemiology , Molecular Sequence Data , Phylogeny , RNA, Viral/blood , Sequence Analysis, DNA , Seroepidemiologic Studies , Thailand/epidemiologyABSTRACT
AIMS: To develop and validate assays based on real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for rapid detection and strain identification (European and North American strains) of porcine reproductive and respiratory syndrome virus (PRRSV) by using SYBR Green I and TaqMan probe chemistries. METHODS AND RESULTS: This study describes two alternative assays based on real-time RT-PCR for rapid detection and strain identification of PRRSV in comparison with conventional RT-PCR. The first assay utilized SYBR Green I with melting curve analysis; another assay was performed using strain-specific TaqMan probes. Primers were selected from the conserved regions within ORF7 (N) of both strains whereas two TaqMan probes labelled with different fluorescent dyes were specifically designed for each strain. The result of strain identification was confirmed by direct sequencing. Both assays can be used for rapid detection and strain identification of PRRSV with a sensitivity of 10(4) and 10(3) copies microl(-1) for SYBR Green and TaqMan probe, respectively. CONCLUSIONS: Real-time RT-PCR is a powerful method combining rapidity, specificity and efficiency for large-scale screening and strain identification of PRRSV. SIGNIFICANCE AND IMPACT OF THE STUDY: The data indicate that the methods developed are invaluable for detecting low levels of PRRSV infection in swine.
Subject(s)
Bacterial Typing Techniques , Porcine Reproductive and Respiratory Syndrome/virology , Porcine respiratory and reproductive syndrome virus/classification , Porcine respiratory and reproductive syndrome virus/isolation & purification , Reverse Transcriptase Polymerase Chain Reaction/methods , Animals , Benzothiazoles , DNA Primers , Diamines , Organic Chemicals , Porcine Reproductive and Respiratory Syndrome/diagnosis , Porcine respiratory and reproductive syndrome virus/genetics , Quinolines , RNA, Viral/genetics , Sensitivity and Specificity , Sequence Analysis, DNA , SwineABSTRACT
Complete genome sequences of H5N1 viruses derived from a domestic cat "A/Cat/Thailand/KU-02/04" and dog "A/Dog/Thailand/KU-08/04" were comprehensively analyzed and compared with H5N1 isolates obtained during the 2004 and 2005 outbreaks. Phylogenetic analysis of both cat and dog viruses revealed that they are closely related to the H5N1 viruses recovered from avian influenza outbreaks of the same period. Genetic analysis of 8 viral gene segments showed some evidence of virulence in mammalian species. In summary, the H5N1 viruses that infected a domestic cat and dog are highly pathogenic avian influenza viruses that are virulent in mammalian species, potentially indicating transmission of H5N1 viruses from domestic animals to humans.
Subject(s)
Influenza A Virus, H5N1 Subtype/genetics , Influenza A Virus, H5N1 Subtype/isolation & purification , Influenza A Virus, H5N1 Subtype/pathogenicity , Orthomyxoviridae Infections/veterinary , Orthomyxoviridae Infections/virology , Amino Acid Sequence , Animals , Animals, Domestic , Aspartic Acid/chemistry , Cats , Disease Outbreaks/veterinary , Dogs , Gene Deletion , Genes, Viral , Influenza A Virus, H5N1 Subtype/chemistry , Molecular Sequence Data , Orthomyxoviridae Infections/epidemiology , Phylogeny , Sequence Homology, Amino Acid , Thailand/epidemiology , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/genetics , VirulenceABSTRACT
The hemagglutinin (HA) and neuraminidase (NA) genes of eight influenza A virus (H5N1) isolates obtained from various avian species in Thailand in 2003-2004 have been characterized in comparison with the Thai isolate A/Chicken/Nakorn-Pathom/Thailand/CU-K2/04(H5N1). Phylogenetic analyses of both genes revealed that all the eight avian isolates were closely related to the A/Chicken/Nakorn-Pathom/Thailand/CU-K2/ 04(H5N1). The amino acid sequence of the HA cleavage site revealed a common characteristic of a highly pathogenic virus strain. Moreover, a deletion of 20 amino acids in the NA stalk region was detected in all Thai isolates in contrast to the H5N1 strain that had caused outbreaks in eastern Asia in 1996-1997 and 2000-2001.