ABSTRACT
OBJECTIVE: The invasion of dengue virus (DENV)-2 Cosmopolitan genotype into the Philippines, where the Asian II genotype previously circulated challenges the principle of dengue serotype-specific immunity. Assessment of antibodies in this population may provide a mechanistic basis for how new genotypes emerge in dengue-endemic areas. METHODS: We evaluated the neutralizing antibody (nAb) and antibody-dependent enhancement (ADE) responses against the two genotypes using archived serum samples collected from 333 patients with confirmed dengue in Metro Manila, Philippines, before, during, and after the introduction of the Cosmopolitan genotype. We quantified nAb titers in baby hamster kidney (BHK-21) cells with or without the Fcγ receptor IIA (FcγRIIA) to detect the capacity of virus-antibody complexes to neutralize or enhance DENV. RESULTS: The nAb potency of the archived serum samples against the two genotypes was greatly affected by the presence of FcγRIIA. We found significant differences in nAb titers between the two genotypes in BHK-21 cells with FcγRIIA (P <0.0001). The archived serum samples were incapable of fully neutralizing the Cosmopolitan genotype, but instead strongly promoted its ADE compared to the Asian II genotype (P <0.0001). CONCLUSION: These results reinforce the role of pre-existing immunity in driving genotype shifts. Our finding that specific genotypes exhibit differing susceptibilities to ADE by cross-reactive antibodies may have implications for dengue vaccine development.
Subject(s)
Dengue Virus , Dengue , Animals , Cricetinae , Humans , Antibodies, Viral , Serogroup , Philippines , Retrospective Studies , Antibodies, Neutralizing , GenotypeABSTRACT
BACKGROUND: Colorectal cancer (CRC) ranks third in terms of incidence and second in mortality worldwide. In CRC, the silencing of mismatch repair genes, including the mutL homolog 1 (hMLH1) has been linked to microsatellite instability (MSI), the lengthening or shortening of microsatellite repeats. Very limited data have been presented so far on the link of hMLH1 methylation and MSI in Southeast Asia populations with sporadic CRC, and on its clinical significance. AIM: To investigate the significance of the MSI status and hMLH1 methylation in CRC Filipino patients. METHODS: Fifty-four sporadic CRC patients with complete clinical data were included in this study. Genomic DNA from CRC tumor biopsies and their normal tissue counterparts were profiled for MSI by high resolution melting (HRM) analysis using the Bethesda Panel of Markers (BAT25, BAT26, D2S123, D5S346, and D17S250). hMLH1 methylation screening was performed using bisulfite conversion and methylation specific polymerase chain reaction. Statistical analysis was conducted to calculate their associations to clinicopathological characteristics and survival relevance (Kaplan-Meier curves and the log-rank test). RESULTS: hMLH1 methylation was observed in 9% and 35% of CRC and normal samples, respectively. Higher incidence of consistently methylated hMLH1 found in both normal and CRC was noticed for relation to location of tumor (P < 0.05). As for MSI status, D2S123 the most common unstable microsatellite and MSI-high (MSI-H) was the most common MSI profile, counted for 46% and 50% of normal and CRC tissues, respectively. The presence of MSI-low (MSI-L) and microsatellite stable (MSS) was 43% and 11% for normal, and 31% and 19% for CRC samples. The mean month of patients' survival was shorter in patients whose normal and tumor tissues had methylated compared to those with unmethylated hMLH1 and with MSI-H compared to those with MSI-L/MSS (P < 0.05). This was supported by significant difference in Kaplan-Meier with log-rank analysis. This data indicated that hMLH1 methylation and high MSI status have prognostic value. CONCLUSION: This study showed the clinical significance of hMLH1 methylation and MSI status in sporadic CRC Filipino patients, especially in the normal part of the tumor.
ABSTRACT
UNLABELLED: Cholera continues to be a global threat, with high rates of morbidity and mortality. In 2011, a cholera outbreak occurred in Palawan, Philippines, affecting more than 500 people, and 20 individuals died. Vibrio cholerae O1 was confirmed as the etiological agent. Source attribution is critical in cholera outbreaks for proper management of the disease, as well as to control spread. In this study, three V. cholerae O1 isolates from a Philippines cholera outbreak were sequenced and their genomes analyzed to determine phylogenetic relatedness to V. cholerae O1 isolates from recent outbreaks of cholera elsewhere. The Philippines V. cholerae O1 isolates were determined to be V. cholerae O1 hybrid El Tor belonging to the seventh-pandemic clade. They clustered tightly, forming a monophyletic clade closely related to V. cholerae O1 hybrid El Tor from Asia and Africa. The isolates possess a unique multilocus variable-number tandem repeat analysis (MLVA) genotype (12-7-9-18-25 and 12-7-10-14-21) and lack SXT. In addition, they possess a novel 15-kb genomic island (GI-119) containing a predicted type I restriction-modification system. The CTXΦ-RS1 array of the Philippines isolates was similar to that of V. cholerae O1 MG116926, a hybrid El Tor strain isolated in Bangladesh in 1991. Overall, the data indicate that the Philippines V. cholerae O1 isolates are unique, differing from recent V. cholerae O1 isolates from Asia, Africa, and Haiti. Furthermore, the results of this study support the hypothesis that the Philippines isolates of V. cholerae O1 are indigenous and exist locally in the aquatic ecosystem of the Philippines. IMPORTANCE: Genetic characterization and phylogenomics analysis of outbreak strains have proven to be critical for probing clonal relatedness to strains isolated in different geographical regions and over time. Recently, extensive genetic analyses of V. cholerae O1 strains isolated in different countries have been done. However, genome sequences of V. cholerae O1 isolates from the Philippines have not been available for epidemiological investigation. In this study, molecular typing and phylogenetic analysis of Vibrio cholerae isolated from both clinical and environmental samples in 2011 confirmed unique genetic features of the Philippines isolates, which are helpful to understand the global epidemiology of cholera.
Subject(s)
Cholera/epidemiology , Cholera/microbiology , Disease Outbreaks , Genes, Bacterial , Vibrio cholerae O1/genetics , Vibrio cholerae O1/isolation & purification , Cluster Analysis , Drug Resistance, Bacterial , Genome, Bacterial , Genotype , Minisatellite Repeats , Molecular Sequence Data , Molecular Typing , Philippines/epidemiology , Phylogeny , Recombination, Genetic , Sequence Analysis, DNA , Sequence Homology , Vibrio cholerae O1/classificationABSTRACT
Hepatitis B virus (HBV) is one of the most prevalent viral infections worldwide. Nearly 400 million individuals are chronic carriers of HBV. The aim of the present study was to determine the frequency of human interleukin 28B (IL28B) variants among treatment naive Filipino patients clinically diagnosed with chronic hepatitis B (CHB), and to compare the IL28B frequency distribution with various ethnic populations. Fifty-seven CHB patients and 43 normal controls were enrolled in this study. Real-time PCR was performed using the TaqMan genotyping assay for IL28B rs12979860. The allelic frequencies among normal controls were 0.94 and 0.06 for the IL28B rs12979860 C and T alleles, respectively. Eighty-eight percent were identified as homozygous for the IL28B C/C genotype and 12% were identified as heterozygous for the IL28B C/T genotype. Among CHB patients, the allelic frequencies were 0.90 for the IL28B C allele and 0.10 for the IL28B T allele. No IL28B T/T genotype was observed between the two groups. No significant difference in the distribution of IL28B genotypes was observed between normal controls and CHB patients. Allelic frequencies of IL28B among Filipinos were similar with other Asian populations but significantly different from Caucasians. The frequency of rs12979860 C>T variants among Filipino CHB patients has not yet been reported. These data provided new insight into the geographical frequency distribution of IL28B variants. Further studies are needed to determine the possible association between IL28B variants and response to pegylated-interferon-α plus ribavirin combination therapy among Filipino patients chronically infected with HBV.
Subject(s)
Asian People/genetics , Gene Frequency , Hepatitis B virus , Hepatitis B, Chronic/genetics , Interleukins/genetics , Polymorphism, Single Nucleotide , Adult , Aged , Alleles , Base Sequence , Case-Control Studies , Craniosynostoses , Female , Genetic Predisposition to Disease , Hepatitis B virus/genetics , Hepatitis B, Chronic/pathology , Hepatitis B, Chronic/virology , Holoprosencephaly , Humans , Interferons , Male , Middle Aged , Philippines , Young AdultABSTRACT
Genome-wide association studies have shown that a non-synonymous single nucleotide polymorphism characterized by a C-to-G change encoding an isoleucine-to-methionine substitution at amino acid position 148 in the human patatin-like phospholipase 3 (PNPLA3) gene was found to be associated with non-alcoholic fatty liver disease (NAFLD) and advanced liver damage. A hospital-based study was conducted to determine the distribution of PNPLA3 genotypes among patients clinically diagnosed and histologically confirmed with NAFLD and among normal controls. We also compared the allelic frequencies of PNPLA3 with different ethnic populations. More importantly, we evaluated the association between PNPLA3 genetic variation and risk of developing NAFLD among Filipinos. Real-time PCR was performed using the Taqman SNP genotyping assay for rs738409. Nucleotide sequencing was performed to confirm the PNPLA3 genotypes. Allelic frequencies among normal controls were 0.83 and 0.17 for the PNPLA3 C and PNPLA3 G alleles, respectively. Calculated frequencies in Hardy Weinberg Equilibrium were 72% for PNPLA3 C/C, 22% for PNPLA3 C/G, and 6% for PNPLA3 G/G genotype. There is a significant difference in the distribution of PNPLA3 genotypes between normal controls and NAFLD patients (p = 0.0172). However, there was no significant association found between PNPLA3 genotypes and risk of developing NAFLD after controlling for possible confounding effects (p = 0.0574). Allelic frequencies of PNPLA3 among Filipinos were statistically different from Hispanics, Japanese, and Han Chinese. In conclusion, genetic variation in PNPLA3 rs738409 C>G seems to be associated with NAFLD among Filipinos. Further studies are needed to replicate our observations in an independent larger population.
ABSTRACT
Chikungunya virus is an alphavirus of the Togaviridae family, which causes a febrile illness with arthralgia in humans. We report here on the complete genome sequence of chikungunya virus strain CHIKV-13-112A isolated from a patient in the Philippines who was suspected to have dengue virus. Phylogenetic analysis revealed that the strain is of the Asian genotype.
ABSTRACT
In 2005, we isolated a new species of virus from mosquitoes in the Philippines. The virion was elliptical in shape and had a short single projection. The virus was named Tanay virus (TANAV) after the locality in which it was found. TANAV genomic RNA was a 9562 nt+poly-A positive strand, and polycistronic. The longest ORF contained putative RNA-dependent RNA polymerase (RdRP); however, conserved short motifs in the RdRP were permuted. TANAV was phylogenetically close to Negevirus, a recently proposed taxon of viruses isolated from haemophagic insects, and to some plant viruses, such as citrus leprosis virus C, hibiscus green spot virus and blueberry necrotic ring blotch virus. In this paper, we describe TANAV and the permuted structure of its RdRP, and discuss its phylogeny together with those of plant viruses and negevirus.
Subject(s)
Culicidae/virology , Insect Viruses/isolation & purification , RNA Viruses/isolation & purification , Viruses, Unclassified/isolation & purification , Amino Acid Sequence , Animals , Culex/virology , Genome, Viral , Insect Viruses/classification , Insect Viruses/genetics , Microscopy, Electron, Transmission , Molecular Sequence Data , Open Reading Frames , Philippines , Phylogeny , Plant Viruses/genetics , RNA Viruses/classification , RNA Viruses/genetics , RNA-Dependent RNA Polymerase/genetics , Sequence Homology, Amino Acid , Viral Proteins/genetics , Virion/ultrastructure , Viruses, Unclassified/classification , Viruses, Unclassified/geneticsABSTRACT
Point mutations and multiple variants across the "a" determinant can destroy the antigenicity and immunogenicity of hepatitis B virus (HBV) leading to false negative assay and vaccine escape. In this study, the presence of surface gene variants of HBV was investigated among patients clinically diagnosed with chronic hepatitis B and positive for HBV DNA from 2002 to 2009. Sequence analysis of the surface gene of HBV showed that 23 (43%) of the 53 isolates had variations. Out of the 23 isolates, 15 (65%) exhibited single or multiple substitutions, which resulted to specific amino acid changes. The remaining 8 (35%) isolates had silent mutations. The amino acid substitution M133T which was associated with failure of HBsAg detection was found in one isolate (7%, 1/15), while the amino acid substitution D144A which was associated with vaccine escape was observed in one isolate (7%, 1/15). No G145R mutation was observed. Of the 15 isolates with identified single or multiple substitutions, 6 (40%) were found to have unique sequences which caused changes in the hydrophobicity profile in the protein. Unique sequence variants at amino acid positions M103I, L109P, S117R, F134I, and S136L found in this study have not yet been reported. These data should be taken into account when developing next generation HBV assays to detect both common and unique variants, and when new HBV vaccines will be designed.
Subject(s)
Hepatitis B Surface Antigens/genetics , Hepatitis B virus/genetics , Hepatitis B virus/isolation & purification , Hepatitis B, Chronic/virology , Amino Acid Substitution , DNA, Viral/chemistry , DNA, Viral/genetics , Diagnostic Errors , Genotype , Hepatitis B Surface Antigens/immunology , Hepatitis B Vaccines/immunology , Hepatitis B virus/classification , Hepatitis B virus/immunology , Hepatitis B, Chronic/diagnosis , Hepatitis B, Chronic/prevention & control , Humans , Molecular Sequence Data , Mutation, Missense , Philippines , Point Mutation , Sequence Analysis, DNAABSTRACT
We report the case of a 76-year old Filipino male who presented with pain, redness, and blurring of vision of the right eye. Corneal scraping was done and sent to the St. Luke's Research and Biotechnology Group for detection and identification of the infectious agent. Morphological detection was performed by allowing the organism from the scraping to grow in 1.5% non-nutrient agar plate with heat-killed E. coli. Trophozoites with acanthopodia and double-walled cysts characteristic of Acanthamoeba were observed within the first and second week of observations, respectively. Molecular identification of the amoebae at the genus level based on the presence of Acanthamoeba-specific amplimer S1, ASA.S1 confirmed the morphological identification. Genotyping through sequence revealed that the organism belonged to T4, which is the genotype commonly present in the eye of keratitis patients.
ABSTRACT
OBJECTIVE: This study aimed to detect the presence of microsatellite (MSI) and loss of heterozygosity (LOH) of the Deleted in Colorectal Cancer (DCC) gene in normal and tumor tissues of Filipino colorectal cancer patients and examine its correlation with age, gender, tumor grade, tumor stage and site of lesion.METHODS: Paired frozen normal and tumor tissues from thirtynine (39) patients with colorectal adenocarcinoma were used by polymerase chain reaction (PCR). Single strand conformation polymorphism - polyacrylamide gel electrophoresis (SSCP - PAGE) was used to determine MSI and restriction fragment length polymorphism (RFLP) was used to study LOH.RESULTS: Based on our data, out of the 39 patients, 10 showed LOH of the DCC gene using the LOH markers VNTR, M2 and M3, while no MSI was detected in the samples using the MSI markers BAT25 and BAT26. Correlation with clinicopathological characteristics showed that there is significance for the site of lesion. The LOH has correclation with tumor samples from the colon but not with those from the rectum.CONCLUSION: Preliminary screening for MSI and LOH of the DCC gene shows that occurrences of colorectal cancer among Filipino patients can be correlated with LOH of the DCC gene with colorectal cancer in a Filipino sample population.
Subject(s)
Humans , Male , Female , Aged , Middle Aged , Adult , Genes, DCC , Polymorphism, Single-Stranded Conformational , Colorectal Neoplasms , Adenocarcinoma , Loss of HeterozygosityABSTRACT
In Myanmar, dengue fever (DF)/dengue hemorrhagic fever (DHF) is one of the leading causes of morbidity and mortality among children. From Pyinmana Hospital in 2004 and Mandalay Children Hospital in 2006, 160 patients diagnosed clinically to have DHF/dengue shock syndrome (DSS) were examined for immunoglobulin M (IgM) and IgG levels. A focus reduction neutralization test was also used to determine primary or secondary dengue virus (DENV) infection. By using IgM-capture ELISA, 139 cases were confirmed as DENV infections. Of these IgM-positives, 94 samples were collected 7-24 days from the onset of illness, to which 13 (14%) and 81 (86%) were determined to be primary and secondary DENV infections, respectively. The 13 primary DENV infection cases were spread among the various severity groups (DHF grade I-IV and DSS) and represented age groups ranging from <1 year of age to 9 years of age. The patients in these primary infection cases showed a remarkably high IgM with a low IgG titer response compared with the secondary infection cases. No significant differences were observed in IgG titers with clinical severity. The data obtained in this study suggest that primary DENV infection cases exist certainly among DHF/DSS cases in Myanmar, and that additional mechanism(s) aside from the antibody-dependent enhancement mechanism could have influenced the clinical severity in DHF/DSS cases.
Subject(s)
Antibodies, Viral/blood , Dengue Virus/immunology , Dengue/immunology , Dengue/pathology , Child , Child, Preschool , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunoglobulin G/blood , Immunoglobulin M/blood , Infant , Male , Myanmar , Neutralization TestsABSTRACT
A hospital-based cross-sectional study was conducted to determine the allelic and genotype frequencies in the genes encoding for catechol-O-methyltransferase and CYP2D6*10 among healthy volunteers and patients clinically diagnosed with cancer pain. PCR-RFLP was used to identify COMT and CYP2D6*10 genotypes. Allelic frequencies among healthy volunteer Filipinos were 0.83 and 0.17 for the COMT Val and COMT Met alleles, respectively. Calculated frequencies in Hardy-Weinberg equilibrium (HWE) were 73% for COMT Val/Val, 26% for COMT Val/Met, and 1% for COMT Met/Met genotype. For CYP2D6*10, allelic frequencies in HWE among volunteers were 0.46 for the C allele and 0.54 for the T allele. Twenty percent were identified as homozygous for the wild-type C/C genotype, 56% were identified as heterozygous for the C/T genotype, and 24% were identified as homozygous for the T/T variant genotype. No significant differences in COMT and CYP2D6*10 allele frequencies between cancer patients and healthy volunteers were noted. Our data demonstrated that the allele frequencies of COMT and CYP2D6*10 in the Filipino healthy volunteers were similar with other Asians but markedly different from Caucasian populations.
ABSTRACT
The glutathione S-transferase (GST) supergene family is made up of four gene families responsible for the biotransformation of drugs and other xenobiotics. Genetic variations in this supergene family influence individual detoxification levels and may contribute to the development of cancer. A hospital-based case-control study was conducted to evaluate the association between GST polymorphism among Filipino patients positive for hepatitis B virus (HBV DNA) and clinically diagnosed as either with chronic active hepatitis, liver cirrhosis, and hepatocellular carcinoma as well as normal individuals negative for HBV infection. Multiplex PCR was used to detect the presence or absence of the GSTT1 and GSTM1 polymorphisms in peripheral blood. DNA sequencing of the S gene region of the virus was used to determine the predominant genotype found among HBV-infected patients. Our results showed that the odds of having a chronic liver disease is only 0.95 (95% CI 0.58-1.57) among those with GSTT1 null genotype compared to those with GSTT1+ genotype. On the other hand, the odds of chronic liver disease is 17.85 times (95% CI 7.34-43.45) for those with GSTM1 null genotype compared to those with GSTM1+ genotype. Using the GSTT1+/GSTM1+ genotype as the reference, both GSTT1+/GSTM1- (OR 16.61; 95% CI 6.69-41.22) and GSTT1-/GSTM1- (OR 11.91; 95% CI 4.48-31.66) genotypes seem to be risk factors for chronic liver disease. From our observations, we conclude that polymorphism in GSTM1 null genotype (OR 17.85; 95% CI 7.34-43.45) seem to be associated with an increased risk of chronic liver disease among Filipinos.
ABSTRACT
BACKGROUND: The mechanisms of thrombocytopenia and platelet phagocytosis in dengue illness are not fully understood. METHODS: A prospective hospital-based study was conducted to examine the relationships between platelet counts, serum thrombopoietin (TPO) levels, and platelet apoptosis and phagocytosis in 81 patients with secondary dengue virus (DV) infections and 38 healthy volunteers. The apoptosis and phagocytosis of cultured platelets after exposure to DV were also examined. RESULTS: Platelet apoptosis, platelet phagocytosis, and serum TPO levels were increased significantly in patients during the acute and early convalescence phases compared with levels observed in patients during the convalescence phase and in healthy volunteers. A significant correlation between platelet apoptosis and platelet phagocytosis was also observed in these patients. Platelet phagocytosis was inhibited significantly by the D89E mutant, which carries a point mutation in the RGD motif of milk fat globule-epidermal growth factor 8, a phosphatidylserine-recognizing bridge molecule. DV-induced platelet apoptosis and increased phagocytosis of DV-induced apoptotic platelets was confirmed using in vitro assays. CONCLUSIONS: Our data suggest an increased phagocytosis of DV-induced apoptotic platelets by macrophages via a phosphatidylserine-recognizing pathway in secondary DV infection. Accelerated platelet clearance, however, was overcome by TPO-induced enhanced thrombopoiesis in these patients. CLINICAL TRIALS REGISTRATION: UMIN000004835.
Subject(s)
Apoptosis/physiology , Blood Platelets/cytology , Dengue/pathology , Macrophages/physiology , Adult , Blood Platelets/physiology , Case-Control Studies , Humans , Platelet Count , Thrombopoietin , Young AdultABSTRACT
The first shotgun genome sequence of a microbial pathogen from the Philippines is reported. Yersinia enterocolitica subsp. palearctica strain PhRBD_Ye1 is the first Y. enterocolitica strain sequenced from an animal source, swine, which is a natural source of yersiniosis. The closest phylogenetic match is a human clinical isolate from Germany.
Subject(s)
Genome, Bacterial , Yersinia Infections/microbiology , Yersinia enterocolitica/genetics , Gastrointestinal Diseases/epidemiology , Gastrointestinal Diseases/microbiology , Humans , Molecular Sequence Data , Philippines/epidemiology , Yersinia Infections/epidemiologyABSTRACT
A formalin-inactivated West Nile Virus (WNV) vaccine (WN-VAX) derived from the WNV-NY99 strain was tested for its safety, efficacy, dilution limit for complete protection, and cross-neutralization. Safety tests performed with experimental animals, bacteria, or cultured cell lines showed no evidence of short- or long-term adverse effects. WN-VAX also protected 100% of 4-week-old mice against a lethal challenge from the WNV-NY99 strain after two doses of intraperitoneal inoculation-even when the vaccine was diluted to 3.2ng/dose. Moreover, very limited cross-neutralization activity against Japanese encephalitis virus, Dengue virus, Murray Valley encephalitis virus, Yellow fever virus or St. Louis encephalitis virus was observed. Therefore, the WN-VAX satisfies the requirements for human trials planned to be done in Japan.
Subject(s)
Cross Protection , West Nile Fever/prevention & control , West Nile Virus Vaccines/immunology , Animals , Cell Line , Chlorocebus aethiops , Dogs , Female , Formaldehyde/pharmacology , Male , Mice , Mice, Inbred C57BL , Mutagenicity Tests , Neutralization Tests , Rabbits , Rats , Toxicity Tests , Vaccines, Inactivated/immunology , Vero Cells , West Nile Virus Vaccines/administration & dosageABSTRACT
A dengue-3-specific real-time reverse transcriptase-polymerase chain reaction (RT-PCR) was developed using the novel Light Upon eXtension (LUX™) fluorogenic technology. A labeled forward primer and a standard reverse primer that target a conserved region within the non-structural 1 (NS1) gene of dengue 3 strains were designed. The dengue-3-specific assay did not recognize other dengue serotypes and related flaviviruses. Using a tenfold serial dilution of plasmid DNA containing the dengue 3 NS1 gene as standards, the range of dengue virus detection was determined to be from 10(3) to 10(9) copies/ml or from 80 to 8 × 10(7) copies/reaction with an average correlation coefficient of ≥ 0.99. The mean intra-assay coefficient of variation (CV) at 2.01% and the mean inter-assay CV at 2.68% suggest the repeatability of the procedure. Moreover, the fluorogenic assay was evaluated by using clinical specimens and comparing test results with historical data obtained from conventional RT-PCR, which served as the criterion standard. Using patient sera as test samples, the assay demonstrated 95.45% sensitivity and 100% specificity, respectively. These results reveal that the real-time RT-PCR assay may be utilized as a rapid, convenient, and sensitive tool for the detection of dengue 3 in clinical and laboratory specimens.
Subject(s)
Dengue Virus/classification , Dengue Virus/isolation & purification , Dengue/virology , Fluorescent Dyes , Reverse Transcriptase Polymerase Chain Reaction/methods , Viral Nonstructural Proteins/genetics , Dengue Virus/genetics , Humans , Oligonucleotide Probes , RNA, Viral/analysis , Reproducibility of Results , Sensitivity and Specificity , Species SpecificityABSTRACT
Helicobacter pylori CagA protein is considered a major virulence factor associated with gastric cancer. There are two major types of CagA proteins: the Western and East Asian CagA. The East Asian CagA-positive H. pylori infection is more closely associated with gastric cancer. The prevalence of gastric cancer is quite low in the Philippines, although Philippine populations are considered to originate from an East Asia source. This study investigates the characteristics of the cagA gene and CagA protein in Philippine H. pylori strains and compares them with previously characterized reference strains worldwide. The full-length cagA gene was sequenced from 19 Philippine isolates and phylogenetic relationships between the Philippine and 40 reference strains were analyzed. All Philippine strains examined were cagA positive, and 73.7% (14/19) strains were Western CagA-positive. The phylogenetic tree based on the deduced amino acid sequence of CagA indicated that the Philippine strains were classified into the two major groups of CagA protein: the Western and the East Asian group. These findings suggest that the modern Western influence may have resulted in more Western type H. pylori strains in the Philippines. Therefore, H. pylori-infected Filipinos can be considered to be at a low risk of developing gastric cancer.
Subject(s)
Antigens, Bacterial/genetics , Bacterial Proteins/genetics , Genetic Variation , Helicobacter Infections/microbiology , Helicobacter pylori/genetics , Peptic Ulcer/microbiology , Adult , Aged , Cluster Analysis , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Female , Helicobacter Infections/pathology , Helicobacter pylori/isolation & purification , Humans , Male , Middle Aged , Molecular Sequence Data , Peptic Ulcer/pathology , Philippines , Phylogeny , Sequence Analysis, DNA , Sequence Homology, Amino AcidABSTRACT
RATIONALE: Increased aortic stiffness, an important feature of many vascular diseases, eg, aging, hypertension, atherosclerosis, and aortic aneurysms, is assumed because of changes in extracellular matrix (ECM). OBJECTIVE: We tested the hypothesis that the mechanisms also involve intrinsic stiffening of vascular smooth muscle cells (VSMCs). METHODS AND RESULTS: Stiffness was measured in vitro both by atomic force microscopy (AFM) and in a reconstituted tissue model, using VSMCs from aorta of young versus old male monkeys (Macaca fascicularis) (n=7/group), where aortic stiffness increases by 200% in vivo. The apparent elastic modulus was increased (P<0.05) in old (41.7+/-0.5 kPa) versus young (12.8+/-0.3 kPa) VSMCs but not after disassembly of the actin cytoskeleton with cytochalasin D. Stiffness of the VSMCs in the reconstituted tissue model was also higher (P<0.05) in old (23.3+/-3.0 kPa) than in young (13.7+/-2.4 kPa). CONCLUSIONS: These data support the novel concept, not appreciated previously, that increased vascular stiffness with aging is attributable not only to changes in ECM but also to intrinsic changes in VSMCs.
Subject(s)
Aging/pathology , Aortic Diseases/pathology , Muscle, Smooth, Vascular/pathology , Actins/metabolism , Age Factors , Aging/metabolism , Animals , Aorta, Thoracic/pathology , Aortic Diseases/etiology , Cells, Cultured , Cytochalasin D/pharmacology , Cytoskeleton/drug effects , Cytoskeleton/metabolism , Elastic Modulus , Integrin beta1/metabolism , Macaca fascicularis , Male , Microscopy, Atomic Force , Muscle, Smooth, Vascular/drug effects , Muscle, Smooth, Vascular/metabolism , Tubulin/metabolism , Vimentin/metabolismABSTRACT
A rapid and comprehensive protocol, which combines simple purification and liquid chromatography-electrospray ionisation-tandem mass spectrometry (LC-ESI/MS/MS), was developed for identification of arboviruses in infected culture fluid. Using this protocol, various arboviruses were detected including uncommon viruses that were described previously as Banna virus and Yunnan orbivirus. This approach is useful for the rapid screening of viral samples that cannot be identified by conventional gene amplification or immunological methods.