Immunogenicity and potential protection of DNA vaccine of Leishmania martiniquensis against Leishmania infection in mice.

INTRODUCTION: In Thailand, Leishmania martiniquensis is the predominant species causing cutaneous and visceral leishmaniasis. Its incidence has been increasing among immunocompetent and immunocompromised hosts. We developed a prototype DNA vaccine using a partial consensus sequence of the cysteine protease B (cpb) gene derived from L. martiniquensis from Thai patients. METHODOLOGY: The laboratory inbred strain of albino BALB/c mice were immunized intramuscularly three times at 2-week intervals (weeks 0, 2, and 4) with cpb plasmid DNA (pcDNA_cpb) with or without the adjuvant, monoolein (pcDNA_cpb-MO). Mice were challenged at week 6 with L. martiniquensis promastigotes. Sera were analysed for IgG1, IgG2a, interferon gamma and interleukin 10 (IFN-γ and IL-10, respectively) levels at weeks 0, 4, and 9. Additionally, livers and spleens were also analysed for parasite burden using immunohistochemistry and real-time polymerase chain (qPCR) assays. RESULTS: Three weeks after promastigote challenge, vaccinated mice showed significantly increased levels of IgG2a and IFN-γ while IL-10 level was significantly reduced when compared with those in the control group (p < 0.01). Parasite burden in the livers and spleens of vaccinated mice significantly decreased. In addition, a significant increase in mature granuloma formation in the livers when compared with those of the control group (p < 0.05) was found, indicating increased T-helper cells (Th1)-induced inflammation and destruction of amastigotes. Monoolein produced a booster effect to enhance the mouse Th1 protective immunity. CONCLUSIONS: The prototype DNA vaccine could induce a Th1 immune response that conferred potential protection to the L. martiniquensis promastigote challenge in BALB/c mice.

Antimicrobial Properties of a Potential Probiotic Lactobacillus from Thai Newborn Feces.

BACKGROUND: Probiotics are increasingly used to treat infectious diarrhea and antibiotic-associated diarrhea. Many probiotic bacteria are classified in general such as Lactobacillus and are able to colonize the gastrointestinal tracts of infants. OBJECTIVE: This study was performed to detect antimicrobial substances and activity in 200 Lactobacillus isolates obtained from healthy Thai newborn feces. MATERIAL AND METHOD: Reuterin production was detected by the spot overlay technique and colorimetric assay. Antimicrobial activity was tested by using a well diffusion, agar method. RESULTS: Lactobacillus strain MSMC64-1 produced reuterin and demonstrated potent antimicrobial activity against seven pathogenic indicator strains with very strong inhibitory activities against Salmonella typhi DMST 5784 and methicillin resistant Staphylococcus aureus (MRSA) DMST 20651. There was strong inhibitory activity against methicillin resistant Staphylococcus aureus (MRSA) DMST20654, Vibrio parahaemolyticus DMST 5665 and Shigella dysenteriae DMST 15111. There was moderate to weak inhibitory activities against Vibrio cholerae DMST 2873 and Helicobacter pylori (H40). The Lactobacillus strain MSMC 64-1 showed resistance to acidic pH (pH 2, 3, 4) and tolerance to 1%, 2%, 3%, and 4% bile concentrations. Sequencing of the 16S ribosomal DNA identified the candidate's strain as Lactobacillus reuteri with 98% sequence homology. CONCLUSION: The active isolate could potentially be used as a probiotic to prevent and treat enteric infections.

Anticancer and apoptosis-inducing activities of microbial metabolites

The problems of systemic toxicity and drug resistance in cancer chemotherapy urge the continuing discovery of new anticancer agents. We explored the specific anticancer activity from microbial metabolites to find new lead compound. 394 microbial extracts were evaluated on anti-proliferative activity against 4 cancer cell lines using MTT assay. Of these, 20 samples showed varying degree of cytotoxicity but specifically to the cancer cell lines since the growth of normal cells was not significantly inhibited by 1 mg/ml of each cell extracts. The 4 most potent extracts exhibited strongest growth inhibition to each cancer cell type were selected for further studied. Cell morphological changes such as cell shrinkage, lose of surface contact and blebbing were observed in all treated cancer cells. DNA-binding dye staining demonstrated nuclear condensation and fragmentation. Chromosomal DNA cleavage detected as DNA ladder pattern by gel electrophoresis including activation of cellular caspase-3 activity, a hallmark of apoptosis, were observed in all treated cancer cell lines. These characteristics suggested the mechanism of apoptosis cell death induced by the extracts. No growth inhibition and apoptosis characteristic were detected in normal cells even at high concentration used suggesting the selective cytotoxicity and potential candidates to develop as anticancer agents.