Burkholderia pseudomallei interferes with inducible nitric oxide synthase (iNOS) production: a possible mechanism of evading macrophage killing

Burkholderia pseudomallei is a causative agent of melioidosis, a life threatening disease which affects humans and animals in tropical and subtropical areas. This bacterium is known to survive and multiply inside cells such as macrophages. The mechanism of host defense against this bacterium is still unknown. In this study, we demonstrated that B. pseudomallei exhibited unique macrophage activation activity compared with Escherichia coli and Salmonella typhi. The mouse macrophage cell line (RAW 264.7) infected with B. pseudomallei at MOI of 0.1:1, 1:1 and 10:1 did not express a detectable level of inducible nitric oxide synthase (iNOS). Moreover, the B. pseudomallei infected cells released TNF-alpha only when they were infected with high MOI (10:1). Unlike the cells infected with B. pseudomallei, the cells infected with E. coli, and S. typhi expressed iNOS even at MOI of 0.1:1. These infected cells also released a significantly higher level of TNF-alpha at the low MOI ratio. The cells that were preactivated with IFN-gamma prior to being infected with B. pseudomallei exhibited an enhanced production of iNOS and TNF-alpha release. The increased macrophage activation activity in the presence of IFN-gamma also correlated with the restriction of the intracellular bacteria survival. Moreover, IFN-gamma also prevented cell fusion and multinucleated cell formation induced by B. pseudomallei, a phenomenon recently described by our group. Altogether, these results indicate that internalization of B. pseudomallei failed to trigger substantial macrophage activation, a phenomenon which could prolong their survival inside the phagocytic cells and facilitate a direct cell to cell spreading of B. pseudomallei to neighboring cells.     

Influence of nutritional and physiological status on behavioral responses of Aedes aegypti (Diptera: Culicidae) to deltamethrin and cypermethrin.

Excito-repellency responses of Aedes aegypti (L.) exposed to deltamethrin and cypermethrin were assessed using an excito-repellency test system. Contact irritancy and non-contact repellency assays compared non-bloodfed (unfed) parous (post-gravid), nulliparous, early blood-fed, late blood-fed, sugar-fed, and unmated female mosquitoes for behavioral responses based on nutritional and physiological conditions at the time of testing. Rates of escape during contact exposure with either compound were most pronounced in parous mosquitoes, followed by unmated mosquitoes, when compared to other conditional states. Significantly higher numbers of parous females also escaped from control chambers compared to other cohorts (P <0.05). Irritability of blood- and sugar-fed mosquitoes was noticeably suppressed. We conclude that nutritional and physiological conditions (including age) of mosquitoes at the time of testing can significantly influence behavioral responses (excito-repellency) to insecticides. The findings indicate that whether due to chronological age, nutrition, physiological state, or innate (circadian) activity patterns, careful consideration must be given to the selection of appropriate conditioned mosquitoes for testing.     

Human papillomavirus genotypes and the p53 codon 72 polymorphism in cervical cancer of Northeastern Thailand

Human papillomavirus (HPV) infection including sub-strain identification was studied in patients with squamous cell cervical cancer (SCCA) in Northeastern Thailand. Subjects were 90 cases of SCCA and 100 healthy controls. Prevalence of high-risk group of HPV infection in the controls and the SCCA patients were 13.0% and 86.7%, respectively. The HPV infection significantly increased the risk for cervical cancer 43.5-fold (95% confidential interval: 17.5-110.6; P <0.00001). Among HPV carrier patients with SCCA (n = 78), HPV-16 was also prominent (70.5%) followed by HPV-18 (23.1%). There was no statistical difference in the subtype distribution between the SCCA and the control groups. There was no significant association between genotype distribution of the p53 codon 72 polymorphism and HPV infection. HPV infection was confirmed as a critical risk factor for cervical cancer development in Northeast Thailand. Since polymorphism of the p53 itself as well as in combination with HPV infection may not be a genetic risk for cervical cancer, much attention should be paid to other risk factors such as sexual behavior and smoking.     

Mitochondrial Changes in β0-Thalassemia/Hb E Disease

The compound β°-thalassemia/Hb E hemoglobinopathy is characterized by an unusually large range of presentation from essentially asymptomatic to a severe transfusion dependent state. While a number of factors are known that moderate presentation, these factors do not account for the full spectrum of presentation. Mitochondria are subcellular organelles that are pivotal in a number of cellular processes including oxidative phosphorylation and apoptosis. A mitochondrial protein enriched proteome was determined and validated from erythroblasts from normal controls and β°-thalassemia/Hb E patients of different severities. Mitochondria were evaluated through the use of mitotracker staining, analysis of relative mitochondrial genome number and evaluation of mitochondrial gene expression in addition to assay of overall cellular redox status through the use of alamarBlue assays. Fifty differentially regulated mitochondrial proteins were identified. Mitotracker staining revealed significant differences in staining between normal control erythroblasts and those from β°-thalassemia/Hb E patients. Differences in relative mitochondria number and gene expression were seen primarily in day 10 cells. Significant differences were seen in redox status as evaluated by alamarBlue staining in newly isolated CD34+ cells. Mitochondria mediate oxidative phosphorylation and apoptosis, both of which are known to be dysregulated in differentiating erythrocytes from β°-thalassemia/Hb E patients. The evidence presented here suggest that there are inherent differences in these cells as early as the erythroid progenitor cell stage, and that maximum deficit is seen coincident with high levels of globin gene expression.     

Irritability and repellency of synthetic pyrethroids on an Aedes aegypti population from Thailand

The main objective of this study was to find the optimal dosage of deltamethrin, cyphenothrin, d‐tetramethrin, and tetramethrin that would elicit repellency and irritability responses of Aedes aegypti. The F1‐F3 generations of field mosquitoes collected from Pu Teuy Village, Sai‐Yok District, Kanchanaburi Province, Thailand, were tested with four pyrethroids to determine the LC₂₅, LC₅₀, and LC₉₉. These concentrations were 0.010%, 0.020%, and 0.055%, respectively, for deltamethrin; 0.113%, 0.167%, and 0.353%, respectively, for cyphenothrin; 2.091%, 2.770%, and 5.114%, respectively, for d‐tetramethrin; and 2.377%, 4.251%, and 10.715%, respectively, for tetramethrin. All dosages were tested in the excito‐repellency system. Survival analysis was used to compare each chamber of the test. It was found that cyphenothrin had a stronger repellent effect than the other pyrethroids, while the contact irritant effect was similar among compounds tested. The LC₅₀ of each pyrethroid was found to be the optimal dose for repelling Ae. aegypti. There was no significant difference in LC₉₉ values for either non‐contact or contact trials for each pyrethroid.     

Phenylene-ethynylene trication as an efficient fluorescent signal transducer in an aptasensor for potassium ion

A tricationic phenylene-ethynylene (N(3+)) fluorophore is investigated as a fluorescent transducer in homogeneous aptasensing system for potassium ion (K(+)) assay in aqueous media. The enhancement of the fluorescent signal of N(3+) by three K(+) aptamers consisting of 12, 15, and 21 nucleotides are observed and used for the determination of N(3+)-aptamer binding affinities. The binding affinities increase with the length of the aptameric oligonucleotides and are proven to be important to the sensitivity and selectivity of the aptasensors. The enhanced fluorescent signal of each N(3+)-aptamer solution is selectively quenched by K(+) due to the ability of K(+) in stabilizing the G-quadruplex structure of the aptamer. Among three aptamers, the 15-base aptamer provides optimal sensitivity and selectivity over other ions such as Li(+), Na(+), NH(4)(+), Mg(2+), Ca(2+) and Sr(2+). The sensing system shows the detection limit of 1 μM of K(+) in clean buffered solution and 30 μM of K(+) in the solution containing 4800-fold excess of Na(+), with wide linear dynamic ranges of micro- to millimolar concentration. This label-free fluorescence aptasensor is conveniently and effectively applicable for analysis of K(+) in urine samples.     

Recent Origin and Cultural Reversion of a Hunter–Gatherer Group

Contemporary hunter–gatherer groups are often thought to serve as models of an ancient lifestyle that was typical of human populations prior to the development of agriculture. Patterns of genetic variation in hunter–gatherer groups such as the !Kung and African Pygmies are consistent with this view, as they exhibit low genetic diversity coupled with high frequencies of divergent mtDNA types not found in surrounding agricultural groups, suggesting long-term isolation and small population sizes. We report here genetic evidence concerning the origins of the Mlabri, an enigmatic hunter–gatherer group from northern Thailand. The Mlabri have no mtDNA diversity, and the genetic diversity at Y-chromosome and autosomal loci are also extraordinarily reduced in the Mlabri. Genetic, linguistic, and cultural data all suggest that the Mlabri were recently founded, 500–800 y ago, from a very small number of individuals. Moreover, the Mlabri appear to have originated from an agricultural group and then adopted a hunting–gathering subsistence mode. This example of cultural reversion from agriculture to a hunting–gathering lifestyle indicates that contemporary hunter–gatherer groups do not necessarily reflect a pre-agricultural lifestyle.