RESULTS: A zinc-tolerant probiotic strain TA4, which was isolated from local fermented food, was selected based on the principal component analysis (PCA) with the highest score of probiotic attributes. Based on the 16S rRNA gene analysis, this strain was identified as Lactobacillus plantarum strain TA4, indicating its high resistance to Zn2+ at a maximum tolerable concentration (MTC) value of 500 mM and its capability of producing ZnO NPs. The UV-visible spectroscopy analysis proved the formations of ZnO NPs through the notable absorption peak at 380 nm. It was also found from the dynamic light scattering (DLS) analysis that the Z-average particle size amounted to 124.2 nm with monodisperse ZnO NPs. Studies on scanning electron microscope (SEM), energy-dispersive X-ray (EDX) spectroscopy, and Fourier-transform infrared spectroscopy (FT-IR) revealed that the main mechanisms in ZnO NPs biosynthesis were facilitated by the Zn2+ biosorption ability through the functional groups present on the cell surface of strain TA4.
CONCLUSIONS: The strong ability of zinc-tolerant probiotic of L. plantarum strain TA4 to tolerate high Zn2+ concentration and to produce ZnO NPs highlights the unique properties of these bacteria as a natural microbial cell nanofactory for a more sustainable and eco-friendly practice of ZnO NPs biosynthesis.
METHODS: Postbiotic metabolites (PM) produced by six strains of L. plantarum were determined for their antiproliferative and cytotoxic effects on normal human primary cells, breast, colorectal, cervical, liver and leukemia cancer cell lines via MTT assay, trypan blue exclusion method and BrdU assay. The toxicity of PM was determined for human and various animal red blood cells via haemolytic assay. The cytotoxicity mode was subsequently determined for selected UL4 PM on MCF-7 cells due to its pronounced cytotoxic effect by fluorescent microscopic observation using AO/PI dye reagents and flow cytometric analyses.
RESULTS: UL4 PM exhibited the lowest IC50 value on MCF-7, RG14 PM on HT29 and RG11 and RI11 PM on HL60 cell lines, respectively from MTT assay. Moreover, all tested PM did not cause haemolysis of human, dog, rabbit and chicken red blood cells and demonstrated no cytotoxicity on normal breast MCF-10A cells and primary cultured cells including human peripheral blood mononuclear cells, mice splenocytes and thymocytes. Antiproliferation of MCF-7 and HT-29 cells was potently induced by UL4 and RG 14 PM respectively after 72 h of incubation at the concentration of 30% (v/v). Fluorescent microscopic observation and flow cytometric analyses showed that the pronounced cytotoxic effect of UL4 PM on MCF-7 cells was mediated through apoptosis.
CONCLUSION: In conclusion, PM produced by the six strains of L. plantarum exhibited selective cytotoxic via antiproliferative effect and induction of apoptosis against malignant cancer cells in a strain-specific and cancer cell type-specific manner whilst sparing the normal cells. This reveals the vast potentials of PM from L. plantarum as functional supplement and as an adjunctive treatment for cancer.
RESULTS: Expression of TB antigen-LysM fusion genes was conducted in Escherichia coli, but this resulted in IBs deposition in contrast to the expression of TB antigens only. This suggested that LysM fusion significantly altered solubility of the TB antigens produced in E. coli. The non-denaturing NLS technique was used and optimized to successfully solubilize and purify ~ 55% of the recombinant cell wall-anchoring TB antigen from the IBs. Functionality of the recovered protein was analyzed via immunofluorescence microscopy and whole cell ELISA which showed successful and stable cell wall binding to L. plantarum (up to 5 days).
CONCLUSION: The presented NLS purification strategy enables an efficient and rapid method for obtaining higher yields of soluble cell wall-anchoring Mycobacterium tuberculosis antigens-LysM fusion proteins from IBs in E. coli.