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  1. Sasidharan S, Uyub AM
    FEMS Immunol. Med. Microbiol., 2009 Jun;56(1):94-7.
    PMID: 19309485 DOI: 10.1111/j.1574-695X.2009.00554.x
    The present study was aimed at modifying the original formulation of Commercial Eugon agar (CEA) to develop a new H. pylori growth medium. Initial studies were carried out to determine the number of H. pylori colonies recovered on in-house H. pylori agar (IHPA), IHPA without L-cysteine and sodium sulfite (IHPA-NC), IHPA without L-cysteine (IHPA-C), IHPA without sodium sulfite (IHPA-N) and CEA as the control. Significant differences (P < 0.001) in the number of colonies recovered were observed between IHPA-N, IHPA-NC and IHPA-C. Incorporation of sodium sulfite decreased the number of colonies recovered, indicating that sodium sulfite was inhibitory to H. pylori growth. Removal of L-cysteine reduced the number of colonies recovered, suggesting that L-cysteine is necessary for the growth of H. pylori. In the subsequent study, incorporation of K(2)HPO(4) further increased the number of colonies recovered compared with IHPA-N (P < 0.001), and 0.25% (w/v) of K(2)HPO(4) yielded the highest numbers of colonies (P < or = 0.04). Finally, thirty other H. pylori clinical isolates were evaluated for their growth in the IHPAP-N, a new medium consisting of 1.5% (w/v) pepticase, 0.5% (w/v) peptone, 0.4% (w/v) sodium chloride, 0.03% (w/v) L-cysteine, 0.55% (w/v) dextrose, 0.25% (w/v) K(2)HPO(4) and 1.5% (w/v) agar. The number of colonies recovered in IHPAP-N was significantly (P < 0.005) higher than that of CEA. IHPAP-N with 0.25% K(2)HPO(4) and without sodium sulfite were adequate solid media for the growth of H. pylori.
  2. Allwood EM, Logue CA, Hafner GJ, Ketheesan N, Norton RE, Peak IR, et al.
    FEMS Immunol. Med. Microbiol., 2008 Oct;54(1):144-53.
    PMID: 18657105 DOI: 10.1111/j.1574-695X.2008.00464.x
    Burkholderia pseudomallei, the causative agent of melioidosis, is endemic to Southeast Asia and northern Australia. Clinical manifestations of the disease are diverse, ranging from chronic localized infection to acute septicaemia, with death occurring within 24-48 h after the onset of symptoms. Definitive diagnosis of melioidosis involves bacterial culture and identification, with results obtained within 3-4 days. This delayed diagnosis is a major contributing factor to high mortality rates. Rapid diagnosis is vital for successful management of the disease. This study describes the purification and evaluation of three recombinant antigenic proteins, BPSL0972, BipD and OmpA from B. pseudomallei 08, for their potential in the serodiagnosis of melioidosis using an indirect enzyme-linked immunosorbent assay (ELISA) method. The recombinant proteins were evaluated using 74 serum samples from culture-confirmed melioidosis patients from Malaysia, Thailand and Australia. In addition, 62 nonmelioidosis controls consisting of serum samples from clinically suspected melioidosis patients (n=20) and from healthy blood donors from an endemic region (n=18) and a nonendemic region (n=24) were included. The indirect ELISAs using BipD and BPSL0972 as antigens demonstrated poor to moderate sensitivities (42% and 51%, respectively) but good specificity (both 100%). In contrast, the indirect ELISA using OmpA as an antigen achieved 95% sensitivity and 98% specificity. These results highlight the potential for OmpA to be used in the serodiagnosis of melioidosis in an endemic area.
  3. Nathan SA, Qvist R, Puthucheary SD
    FEMS Immunol. Med. Microbiol., 2005 Feb 1;43(2):177-83.
    PMID: 15681148
    The oxidative response of Burkholderia pseudomallei and Escherichia coli infected macrophages from normal and melioidosis subjects was determined by measuring the production of nitric oxide which is one of the reactive nitrogen intermediates, and the activation state of these macrophages was determined by measuring the generation of 8-iso-PGF(2alpha), a bioactive product of free radical induced lipid peroxidation. Macrophages obtained from the melioidosis patients generated significantly lower levels of nitric oxide and 8-iso-PGF(2alpha) compared to macrophages obtained from the normal subjects (P<0.001). The reduced efficiency of the oxygen dependent microbicidal mechanism in macrophages of melioidosis patients may be one of the survival strategies developed by B. pseudomallei to remain viable intracellularly.
  4. Chan SW, Nathan S
    FEMS Immunol. Med. Microbiol., 2005 Jan 1;43(1):37-44.
    PMID: 15607634
    Filamentous phage random peptide libraries were used to identify the epitopes of Burkholderia pseudomallei protease by panning against IgG polyclonal sera that exhibited protease neutralizing properties. The isolated fusion peptides presented a consensus peptide sequence, TKSMALSG, which closely resembles part of the active site sequence, 435GTSMATPHVAG445, of B. pseudomallei serine metalloprotease. By comparing the consensus sequence, TKSMALSG, with the predicted three-dimensional molecular model of B. pseudomallei serine metalloprotease, it appears that the potential antibody binding epitope was buried within the molecule. This active site was conformational whereby one continuous sub-region (SMA) was located between two discontinuous sub-regions, supplied by the flanking residues in the same polypeptide. All phages selected from the biopanning with IgG polyclonal sera showed good binding towards the polyclonal antibodies when compared to the negative control. In addition, these peptide-bearing phages showed competitive inhibition of B. pseudomallei serine metalloprotease binding to the polyclonal IgG.
  5. Ramelah M, Aminuddin A, Alfizah H, Isa MR, Jasmi AY, Tan HJ, et al.
    FEMS Immunol. Med. Microbiol., 2005 May 1;44(2):239-42.
    PMID: 15866222
    Helicobacter pylori infection of a distinct subtype of cagA may lead to different pathological manifestation. The aim of this study is to determine the presence of cagA gene and its variants in H. pylori infection among different ethnic groups and its effect on gastroduodenal diseases. Overall detection of cagA among the 205 clinical isolates of H. pylori was 94%. Variations in size of the 3' region of cagA gene were examined among 192 Malaysian H. pylori cagA-positive strains. Results showed that three cagA variants differing in fragment length of PCR products were detected and designated as type A (621-651bp), type B (732-735bp) and type C (525 bp). Although there was no association between any of the cagA subtypes with peptic ulcer disease (p>0.05), an association between cagA subtypes with a specific ethnic group was observed. Specific-cagA subtype A strains were predominantly isolated from Chinese compared to Malays and Indians (p<0.0005), and cagA subtype B strains were predominantly isolated from Malays and Indians compared to Chinese (p<0.05). The cagA type A strains of H. pylori is commonly found in the Chinese patients who have a higher risk of peptic ulcer disease, thus indicating that it could be used as an important clinical biomarker for a more severe infection.
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