ABSTRACT
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ABSTRACT
The gold standard method for laboratory diagnosis of melioidosis is bacteriological culture. However, this method is time-consuming. Currently, a serological method with high sensitivity and specificity for diagnosis of melioidosis has not been available. Specific antigens for serological diagnosis are highly needed. The aim of this study was to identify immunodominant proteins from B. pseudomallei as antigen for laboratory diagnosis of melioidosis. In this study, a partial Sau3A library of B. pseudomallei genomic DNA was constructed in pUC18 vector. The E. coli clones expressing immunogenic protein of B. pseudomallei were selected by a colony blot assay using pre-absorbed pooled melioidosis sera as the probe. One clone, PHE191, expressing a 40 kDa recombinant protein with high homology to zinc-binding dehydrogenase family protein was chosen for further study. The recombinant protein was purified and evaluated by Western blot analysis for its diagnostic utility using a total of 107 serum samples from culture-proven melioidosis patients, patients with other related diseases and healthy individuals from endemic and non-endemic areas. The results showed that this protein had sensitivity and specificity of 78.9 % and 72.5 %, respectively, whereas indirect hemagglutination assay (IHA) gave sensitivity and specificity of 86.8 % and 66.7 %, respectively. This protein had higher specificity but low sensitivity than IHA. With a higher specificity, this protein is warranted for further study to determine whether its native conformation would provide better diagnostic potential for laboratory diagnosis of melioidosis.
ABSTRACT
Staphylococcus aureus, a well-known human pathogen, causes a variety of infections in human, e.g. superficial skin infection through life-threatening infection. S. aureus is able to produce many enzymes, including exotoxins which lead to tissue inflammation and injury. Moreover, it also plays an importance role in clinical practice by exhibiting resistance phenotype to methicillin. Many virulence determinants are located on mobile genetic elements (MGEs), such as bacteriophages, pathogenicity islands (PAI), and genomic islands, and existed variably in the bacterial population. Then, virulence genes can be used as genetic markers for clinical manipulations, and nosocomial control measurement. The objective of this study was to determine virulence genes associated with those MGEs in S. aureus samples both in methicillin-susceptible S. aureus (MSSA), and methicillin-resistant S. aureus (MRSA). A total of 100 MSSA and MRSA isolates (50 of each) were randomly selected from clinical samples of patients at Srinagarind Hospital during November, 2006 through June, 2007. All isolates were determined for the presence of eta, lukDE, lukSF-PV, tst-1, sak, sea, sec, sel, and sep genes by PCR. In case of MRSA, staphylococcal cassette chromosme mec (SCCmec) types were also determined in order to study the association among the determinants and their allotypes. The results showed that most of S. aureus samples harbored at least one virulence gene, and most of them carried lukDE (90 %), and sak (88 %). High potential virulence genes, eta and lukSF-PV, were detected in 2 and 10 isolates of MSSA only. However, sea gene was detected more frequent in MRSA than MSSA (P \< 0.05). While sec gene was significantly recognized less in MRSA than MSSA isolates (P \< 0.05). The other staphylococcal enterotoxins such as sec, sel and sep were detected in small samples of S. aureus, and none was found to harbor tst-1 gene. The molecular information associated to virulence genes on MGEs may be useful in clinical practice and hospital epidemiology in Srinagarind Hospital, and other tertiary care facilities.