Abstract.
Bacillus sp. N18, which produces a strongly fibrinolytic enzyme, was isolated from soil samples. The gene coding for the fibrinolytic enzyme (N18) was cloned and high level of expression/secretion was achieved using a high copy number plasmid and a triple protease deficient Bacillus subtilis strain. The recombinant enzyme (termed ReN18) was purified by consecutive chromatography with Streamline SP XL and Sephacryl S-100, resulting in a single protein band on IEF gel with an isoelectric point approximately at pH 8.6 and on SDS-PAGE with an apparent molecular weight of 28,000 Da, which is very close to the molecular mass determined by mass spectrometry (27,728 Da). N-terminal sequencing revealed that the first 15 amino acid residues are AQSVPYGISQIKAPA , identical to those deduced from DNA sequence. The purified ReN18 showed a higher affinity for fibrin in comparison with urokinase or plasmin in vitro. ReN18 was also active in vivo in experimental animals using either oral or intravenous route of administration. Furthermore, analysis of plasmin (Plm) activity, plasminogen activator inhibitor (PAI) activity and D-dimer concentration, and residual plasma fibrinogen (Fbg) indicated that the recombinant enzyme was able to cleave directly cross-linked fibrin without concurrently destroying fibrinogen. Results obtained from a brain-thrombus mouse model demonstrated that the recombinant enzyme was able to increase the surviving rate of experimental animals to 82% at a dose of 8,000 U/kg, 77% at 4,000 U/kg, 40% at 2,000 U/kg, respectively, in comparison with 57% at a dose of 8,000 U/kg when urokinase was used. The current study demonstrated that ReN18 could be used as a potent thrombolytic agent.

Key words: Bacillus, Fibrinolytic enzyme, Nattokinase, Plasmin, Thrombolytic agent, Urokinase

^*Author for Correspondence.
Mailing address: School of Life Science, East China Normal University, Shanghai 200062, China
Tel. : 86-21-62233295, Fax.: 86-21-62233754, E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Abstract.
The 1.35-kb fragment of P97/302 infectious bursal disease virus (IBDV) VP2 gene, encompassing the hypervariable region was successfully amplified by the reverse transcriptase/polymerase chain reaction (RT/PCR). This isolate was cloned and sequenced. The sequenced was analyzed and compared to other eight reference IBDV strains. This isolate has amino acid substitutions at the 222(A), 256(I), 294(I) and 299(S) as other reported reference vv (very virulent) strains of UK661, HK46 and OKYM. The amino acid residues at the two hydrophilic regions and the serine-rich heptapeptide region of the P97/302 are the same as vv strains of UK661, HK46 and OKYM. The P97/302 IBDV isolate can be digested with enzymes TaqI, StyI, SspI but not with SacI as previously reported in many others vv strain. Phylogenetic analysis based on the nucleotide sequence of hypervariable region showed that this isolate clustered with the vv strains of UK661, HK46 and OKYM and distinct from other groups of classical, attenuated and variant strains. It was concluded that this P97/302 IBDV isolate was serotype 1, vv IBDV strain.

Key words: Very Virulent Infectious Bursal Disease

^*Author for Correspondence.

Abstract.
The use of polymerase chain reaction (PCR) technique in detection of immunoglobulin heavy (IgH) chain gene rearrangement enables clonal populations of lymphoid cells involved in malignant tumor to be detected hence differentiating malignant from reactive proliferation. The aim of this study was to detect clonal lymphoid populations using IgH chain gene rearrangement in cases of Non-Hodgkin's lymphoma (NHL) observed in Universiti Sains Malaysia Hospital. DNA from 43 fresh-frozen lymph nodes (LN) were extracted from patients suspected of lymphoma and subjected to PCR using specific primers for IgH chain gene rearrangement (V_H/J_H). Final diagnosis showed 21 cases were NHL-B, 2 NHL-T cases, 6 Hodgkin's Disease, 6 reactive hyperplasia, 4 metastatic carcinoma and 4 cases of tuberculosis. Monoclonal populations represented by a specific band at 100-180bp were observed in 18/21 B lineage NHL, 2/2 T lineage NHL and 2/6 Hodgkin's Disease while none was observed in cases of reactive hyperplasia, metastatic carcinoma and tuberculosis. We were able to show that IgH chain gene rearrangement could serve as a specific and sensitive technique in detecting clonal populations especially in B-lineage NHL. Thus, IgH chain gene rearrangement analysis could be a useful adjunct to immunohistochemistry in detection of lineage and clonality in NHL.

Key words: clonality, IgH gene rearrangement, Non-Hodgkin's lymphoma, PCR

^*Author for Correspondence.
Mailing address: Department of Medicine, School of Medical Sciences, Health Campus, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan, Malaysia.
Tel: 609-7651700 ext 1466; Fax: 609-7653370. E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Abstract.
Pulsed-field gel electrophoresis (PFGE) was applied to analyse sporadic and outbreak cases of cholera caused by V. cholerae O1 biotype El Tor from Vietnam (n=16) and V. cholerae O139 from Malaysia (n=12) and from five vaccine strains. NotI-Digestion of chromosomal DNA from these isolates, followed by PFGE produced eleven different profiles among the 16 V. cholerae O1 isolates (Dice coefficient: 0.67-0.97). The V. cholerae O1 Ogawa and Inaba strains were clearly differentiated into distinct clusters. NotI-PFGE analysis among V. cholerae O139 isolates showed a predominant PFGE profile (N12) indicating that the outbreak was probably derived a single clone. Among the five vaccine strains, the PFGE profiles of V. cholerae Inaba Cairo 48 and classical Inaba Cairo 48 were similar to some of the field isolates of 01 Inaba strains in Vietnam. The PFGE analysis showed that the V. cholerae 0139 isolates from Malaysia were indistinguishable as these strains were probably derived from a cholera outbreak and the V. cholerae O1 isolates from Vietnam exhibited limited genetic diversity.

Key words: PFGE, Vibrio cholerae O1 and O139

^*Author for Correspondence.
Mailing address: Microbiology Division, Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603, Kuala Lumpur, Malaysia.
Tel No. 603-79674437  Fax No 603-79674606  Email: This email address is being protected from spambots. You need JavaScript enabled to view it.