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Effect of colony morphology variation of Burkholderia pseudomallei on intracellular survival and resistance to antimicrobial environments in human macrophages in vitro
Sarunporn Tandhavanant, Aunchalee Thanwisai, Direk Limmathurotsakul, Sunee Korbsrisate, Nicholas PJ Day, Sharon J Peacock, Narisara Chantratita
BMC Microbiology , 2010, DOI: 10.1186/1471-2180-10-303
Abstract: Morphotype was associated with survival in the presence of H2O2 and antimicrobial peptide LL-37, but not with susceptibility to acid, acidified sodium nitrite, or resistance to lysozyme, lactoferrin, human neutrophil peptide-1 or human beta defensin-2. Incubation under anaerobic conditions was a strong driver for switching of type III to an alternative morphotype. Differences were noted in the survival and replication of the three types following uptake by human macrophages, but marked strain-to strain-variability was observed. Uptake of type III alone was associated with colony morphology switching.Morphotype is associated with phenotypes that alter the ability of B. pseudomallei to survive in adverse environmental conditions.Burkholderia pseudomallei is an environmental Gram-negative bacterium that causes a severe and often fatal disease called melioidosis. This is an important cause of sepsis in south-east Asia and northern Australia, a geographic distribution that mirrors the presence of B. pseudomallei in the environment [1]. Melioidosis may develop following bacterial inoculation or inhalation and occurs most often in people with regular contact with contaminated soil and water [1]. Clinical manifestations of melioidosis are highly variable and range from fulminant septicemia to mild localized infection. The overall mortality rate is 40% in northeast Thailand (rising to 90% in patients with severe sepsis) and 20% in northern Australia [1,2].A major feature of melioidosis is that bacterial eradication is difficult to achieve. Fever clearance time is often prolonged (median 8 days), antimicrobial therapy is required for 12-20 weeks, and relapse occurs in around 10% of patients despite an appropriate course of antimicrobial therapy [3,4]. The basis for persistence in the infected human host is unknown, although several observations made to date may be relevant to the clinical behaviour of this organism [2,5]. B. pseudomallei can resist the action of bactericidal
Isolation and characterization of a novel podovirus which infects burkholderia pseudomallei
Jiraporn Gatedee, Kanyanan Kritsiriwuthinan, Edouard E Galyov, Jinyu Shan, Elena Dubinina, Narin Intarak, Martha RJ Clokie, Sunee Korbsrisate
Virology Journal , 2011, DOI: 10.1186/1743-422x-8-366
Abstract: Burkholderia pseudomallei is a Gram-negative bacillus and the causative agent of melioidosis which is endemic in Southeast Asia and northern Australia [1]. The bacterium is a widely distributed environmental saprophyte commonly found in water and soil in the endemic areas. The bacterium is highly resilient to environmental stress and can survive in distilled water for months or even years [2]. It infects humans via inoculation through skin abrasions or via the inhalation route and causes either acute or chronic melioidosis. Acute infection is often manifested as septicemia, resulting in death within days of exposure. B. pseudomallei is resistant to many antibiotics including third-generation cephalosporins [3]. There is clearly a need for novel antimicrobials. One promising source of antimicrobials may come from bacteriophages which infect and kill specific bacterial cells. They are the most abundant living entities in the world, and play key roles in bacterial population dynamics and evolution [4]. Bacteriophages can be either temperate where they integrate into the host bacterial genomes, or virulent where they infect, multiply, and lyse their hosts without any initial integration period. The ability of such virulent phages to be used therapeutically has been well documented and reviewed [5,6].Prophages have been shown to be abundant in B. pseudomallei genomes [7-9]. A recent study reported the abundance and diversity of prophages in B. pseudomallei, B. thailandensis and B. mallei genomes and suggested how they contribute to the phenotypic diversity in the Burkholderia species complex [8]. Pertinently to our study, the 37 phages identified in the Burkholderia genomes were either myoviruses, Mu-like viruses, or siphoviruses. Although one previous study reported the presence of a putative podovirus in the genome of B. pseudomallei, there are no published genetic studies to support this work [7]. Another recent study isolated six B. pseudomallei myoviruses from soil
Global transcriptional profiling of Burkholderia pseudomallei under salt stress reveals differential effects on the Bsa type III secretion system
Pornpan Pumirat, Jon Cuccui, Richard A Stabler, Joanne M Stevens, Veerachat Muangsombut, Ekapot Singsuksawat, Mark P Stevens, Brendan W Wren, Sunee Korbsrisate
BMC Microbiology , 2010, DOI: 10.1186/1471-2180-10-171
Abstract: Transcriptome analysis of B. pseudomallei under salt stress revealed several genes significantly up-regulated in the presence of 320 mM NaCl including genes associated with the bsa-derived Type III secretion system (T3SS). Microarray data were verified by reverse transcriptase-polymerase chain reactions (RT-PCR). Western blot analysis confirmed the increased expression and secretion of the invasion-associated type III secreted proteins BipD and BopE in B. pseudomallei cultures at 170 and 320 mM NaCl relative to salt-free medium. Furthermore, salt-treated B. pseudomallei exhibited greater invasion efficiency into the lung epithelial cell line A549 in a manner partly dependent on a functional Bsa system.B. pseudomallei responds to salt stress by modulating the transcription of a relatively small set of genes, among which is the bsa locus associated with invasion and virulence. Expression and secretion of Bsa-secreted proteins was elevated in the presence of exogenous salt and the invasion efficiency was enhanced. Our data indicate that salinity has the potential to influence the virulence of B. pseudomallei.Burkholderia pseudomallei is a saprophyte and the causative agent of melioidosis, a human infectious disease endemic in some tropical areas including southeast Asia and northern Australia [1]. Inhalation is a recognized route of infection with this organism and pulmonary disease is common [1,2]. Owing to its aerosol infectivity, the severe course of infection, and the absence of vaccines and fully effective treatments, B. pseudomallei is classified as a hazard category three pathogen and considered a potential biothreat agent [2]. B. pseudomallei, is a Gram negative bacillus found in soil and water over a wide endemic area and mainly infects people who have direct contact with wet soil [1,3]. In Thailand, the highest incidence of melioidosis is in the northeast region, at a rate of approximately 3.6-5.5 per 100,000 human populations annually. Septicaemic presentati
Analysis of the Prevalence, Secretion and Function of a Cell Cycle-Inhibiting Factor in the Melioidosis Pathogen Burkholderia pseudomallei
Pornpan Pumirat, Charles Vander Broek, Niramol Juntawieng, Veerachat Muangsombut, Pattarachai Kiratisin, Kovit Pattanapanyasat, Joanne M. Stevens, Mark P. Stevens, Sunee Korbsrisate
PLOS ONE , 2014, DOI: 10.1371/journal.pone.0096298
Abstract: Enteropathogenic and enterohaemorrhagic Escherichia coli express a cell cycle-inhibiting factor (Cif), that is injected into host cells via a Type III secretion system (T3SS) leading to arrest of cell division, delayed apoptosis and cytoskeletal rearrangements. A homologue of Cif has been identified in Burkholderia pseudomallei (CHBP; Cif homologue in B. pseudomallei; BPSS1385), which shares catalytic activity, but its prevalence, secretion and function are ill-defined. Among 43 available B. pseudomallei genome sequences, 33 genomes (76.7%) harbor the gene encoding CHBP. Western blot analysis using antiserum raised to a synthetic CHBP peptide detected CHBP in 46.6% (7/15) of clinical B. pseudomallei isolates from the endemic area. Secretion of CHBP into bacterial culture supernatant could not be detected under conditions where a known effector (BopE) was secreted in a manner dependent on the Bsa T3SS. In contrast, CHBP could be detected in U937 cells infected with B. pseudomallei by immunofluorescence microscopy and Western blotting in a manner dependent on bsaQ. Unlike E. coli Cif, CHBP was localized within the cytoplasm of B. pseudomallei-infected cells. A B. pseudomallei chbP insertion mutant showed a significant reduction in cytotoxicity and plaque formation compared to the wild-type strain that could be restored by plasmid-mediated trans-complementation. However, there was no defect in actin-based motility or multinucleated giant cell formation by the chbP mutant. The data suggest that the level or timing of CHBP secretion differs from a known Bsa-secreted effector and that CHBP is required for selected virulence-associated phenotypes in vitro.
The Application of NIOSH Lifting Equation to Prevent Musculoskeletal Disorder Risks  [PDF]
Parvena Meepradit, Nipoporn Sunee, Ruephuwan Chantrasa
Journal of Biosciences and Medicines (JBM) , 2015, DOI: 10.4236/jbm.2015.33006

The objective of the study was to reduce musculoskeletal disorder risks by applying the NIOSH lifting equation variables include the horizontal location, the vertical location, the vertical travel distance, the asymmetric, the lifting frequency and the coupling classification. The 17 specific samples from 4W and ZECP division were selected by the weight of box 15.4 - 28.7 pounds. The standardized Nordic questionnaire for the analysis of musculoskeletal symptoms with pain scale from 0 (no pain) to 10 (worst pain) was used to self-report feeling. The ergonomics redesigns trained for the workers included: 1) brought the load closer to the worker by training; 2) raised the height of objects placed to reduce the vertical distance between the origin and destination of the lift; and 3) moved the origin and destination of lift closer together to reduce the angel twist. The new procedures were trained to all participated workers. The result found that the lifting index was safer (<1.0). For successful outcome, be supposed to monitoring is careful the data about a problem of the worker health, give the carefulness in case of specially exceed environment more than the LI advises and should do training continuously.

The Condition-Dependent Transcriptional Landscape of Burkholderia pseudomallei
Wen Fong Ooi equal contributor,Catherine Ong equal contributor,Tannistha Nandi,Jason F. Kreisberg,Hui Hoon Chua,Guangwen Sun,Yahua Chen,Claudia Mueller,Laura Conejero,Majid Eshaghi,Roy Moh Lik Ang,Jianhua Liu,Bruno W. Sobral,Sunee Korbsrisate,Yunn Hwen Gan,Richard W. Titball,Gregory J. Bancroft,Eric Valade,Patrick Tan
PLOS Genetics , 2013, DOI: 10.1371/journal.pgen.1003795
Abstract: Burkholderia pseudomallei (Bp), the causative agent of the often-deadly infectious disease melioidosis, contains one of the largest prokaryotic genomes sequenced to date, at 7.2 Mb with two large circular chromosomes (1 and 2). To comprehensively delineate the Bp transcriptome, we integrated whole-genome tiling array expression data of Bp exposed to >80 diverse physical, chemical, and biological conditions. Our results provide direct experimental support for the strand-specific expression of 5,467 Sanger protein-coding genes, 1,041 operons, and 766 non-coding RNAs. A large proportion of these transcripts displayed condition-dependent expression, consistent with them playing functional roles. The two Bp chromosomes exhibited dramatically different transcriptional landscapes — Chr 1 genes were highly and constitutively expressed, while Chr 2 genes exhibited mosaic expression where distinct subsets were expressed in a strongly condition-dependent manner. We identified dozens of cis-regulatory motifs associated with specific condition-dependent expression programs, and used the condition compendium to elucidate key biological processes associated with two complex pathogen phenotypes — quorum sensing and in vivo infection. Our results demonstrate the utility of a Bp condition-compendium as a community resource for biological discovery. Moreover, the observation that significant portions of the Bp virulence machinery can be activated by specific in vitro cues provides insights into Bp's capacity as an “accidental pathogen”, where genetic pathways used by the bacterium to survive in environmental niches may have also facilitated its ability to colonize human hosts.
Development of criterion weights for preliminary site selection: A pilot project of Supanburi industrial estate
Pichaya Rachdawong,Sunee Apawootichai
Songklanakarin Journal of Science and Technology , 2003,
Abstract: A generalized framework for criterion weights development was proposed for a pilot project of Supanburi industrial estate for light industries. It encompassed selection of criteria, questionnaire design, data collection, and data analysis. Relative or comparative importance matrix of the six factors was created from the summary of the data from expert survey. Pairwise comparison matrix was subsequently computed to reduce the data dimension and to highlight the relative importance of each factor prior to weight extraction. The highest weight was the distance to water bodies, which emphasized the importance given to natural features by experts. The framework should ease and enable adoption of multi-criteria decision-making technique for other cases of preliminary industrial estate site selection. Care must also be exercised to ensure the adequacy of quantities and quality of survey made as well as of data analysis protocol.
Comparative detection of Plasmodium vivax and Plasmodium falciparum DNA in saliva and urine samples from symptomatic malaria patients in a low endemic area
Pattakorn Buppan, Chaturong Putaporntip, Urassaya Pattanawong, Sunee Seethamchai, Somchai Jongwutiwes
Malaria Journal , 2010, DOI: 10.1186/1475-2875-9-72
Abstract: Matched blood, saliva and urine samples were collected from 100 microscopy-positive and 20 microscopy-negative febrile patients who attended a malaria clinic in Tak Province, northwestern Thailand for nested PCR analysis targeting the small subunit ribosomal RNA gene of human malaria. Both P. falciparum and Plasmodium vivax have been known to circulate at a comparable rate in the study area.Comparing with microscopy results, nested PCR of saliva samples had a sensitivity of 74.1% for P. falciparum detection and 84% for P. vivax detection while 44.4% and 34.0% of the corresponding values were observed for urine samples. Both nested PCR results of saliva and urine samples had a specificity of 100% for identification of P. falciparum and P. vivax when compared with nested PCR results from blood. Co-infections of both species were found in four, 26 and 8 patients by microscopy and nested PCR of blood and saliva samples, respectively. Although the positive rates of nested PCR of saliva samples for P. falciparum increased with parasite density, no tendency occurred in results from nested PCR of saliva samples for P. vivax as well as those of urine samples.Saliva and urine samples could be alternative noninvasive sources of DNA for molecular detection of both P. falciparum and P. vivax. Further improvement of the detection method will offer an opportunity to use these samples for diagnosis of malaria.Plasmodium falciparum and Plasmodium vivax are the main causative agents of human malaria accounting for 300-500 million cases and 130-145 million infections per annum, respectively [1,2]. Definite diagnosis of acute malaria infection in routine laboratory practice relies on demonstration of asexual erythrocytic stages from peripheral blood smears under light microscope. Although the method is feasible, economical and practical, a number of symptomatic cases have been left undiagnosed when parasitaemia did not reach the microscopic detection threshold. Hence, a more sensitive
Purification and amino acid sequence of a bacteriocins produced by Lactobacillus salivarius K7 isolated from chicken intestine
Komkhae Pilasombut,Thavajchai Sakpuaram,Worawidh Wajjwalku,Sunee Nitisinprasert
Songklanakarin Journal of Science and Technology , 2006,
Abstract: A bacteriocin-producing strain, Lactobacillus K7, was isolated from a chicken intestine. The inhibitory activity was determined by spot-on-lawn technique. Identification of the strain was performed by morphological, biochemical (API 50 CH kit) and molecular genetic (16S rDNA) basis. Bacteriocin purification processes were carried out by amberlite adsorption, cation exchange and reverse-phase high perform- ance liquid chromatography. N-terminal amino acid sequences were performed by Edman degradation. Molecular mass was determined by electrospray-ionization (ESI) mass spectrometry (MS). Lactobacillus K7 showed inhibitory activity against Lactobacillus sakei subsp. sakei JCM 1157T, Leuconostoc mesenteroides subsp. mesenteroides JCM 6124T and Bacillus coagulans JCM 2257T. This strain was identified as Lb. salivarius. The antimicrobial substance was destroyed by proteolytic enzymes, indicating its proteinaceous structure designated as a bacteriocin type. The purification of bacteriocin by amberlite adsorption, cation exchange, and reverse-phase chromatography resulted in only one single active peak, which was designated FK22. Molecular weight of this fraction was 4331.70 Da. By amino acid sequence, this peptide was homology to Abp 118 beta produced by Lb. salivarius UCC118. In addition, Lb. salivarius UCC118 produced 2-peptide bacteriocin, which was Abp 118 alpha and beta. Based on the partial amino acid sequences of Abp 118 beta, specific primers were designed from nucleotide sequences according to data from GenBank. The result showed that the deduced peptide was high homology to 2-peptide bacteriocin, Abp 118 alpha and beta.
In vitro adhesion assay of lactic acid bacteria, Escherichia coli and Salmonella sp. by microbiological and PCR methods
Sunee Nitisinprasert,Nuntaporn Pungsungworn,Penkhae Wanchaitanawong,Gerard Loiseau
Songklanakarin Journal of Science and Technology , 2006,
Abstract: In vitro adhesion assay using Lactobacillus reuteri KUB-AC5 as a test strain has been studied by applying simple PCR reaction together with image analysis and plate count techniques. Critical factor affecting the PCR method was quality and quantity of DNA. The cell lysis technique was modified to optimize this method. Thus, lysozyme and proteinase K were added to lyse the cells, followed by SDS solution to obtain a complete cell lysis. Only PCR products from total cells (TC) were obtained, with low consistency, but none from cells bound to mucus (BC) at either 0.1 or 0.5 mg/mL concentration. It was hypothesized that the attached cells might not be extracted into the cell suspension. Therefore, 1% SDS solution and 0.1M NaOH were used directly in the extraction. As expected, PCR products were observed when both TC and BC were used as a DNA template. Adhesion appeared at a wide range of 0-45%, with low consistency. Therefore, a simple microbiological method (plate count) was used. The extraction of bound cells into cell suspension was critical in this method. Extraction times of 20, 60, 120 and 150 min were tried. Results showed that maximum cell number was obtained with 120 min extraction. L. reuteri KUB-AC5, L. reuteri KUB-AC16, L. reuteri KUB-AC20, L. salivarius KUB-AC21, L. acidophilus KV-1, Escherichia coli E010, Salmonella sp. S003, E. coli ATCC8739, and S. typhimurium ATCC 13311 exhibited adhesion activity of 21.6%, 0.8%, 5.7%, 1.1%, 23.1%, 10.7%, 10.3%, 4.4% and 3.2%, respectively. Among the 9 types of microorganisms tested L. acidophilus KV-1 and L. reuteri KUB-AC5 showed higher adhesion activity than the others.
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