Nisin is a peptide bacteriocin, grouped under the category of lantibiotics. It is naturally produced by Lactococcus lactis to eliminate other competing gram-positive bacteria from its vicinity. Moreover under certain conditions it is reported to be effective against a broad range of gram-negative bacteria as well. Thus, it has been widely used as a safe food preservative especially in the dairy industry. Because of its wide-scale consumption, its effect on eukaryotic cells should be of great concern. Here we examine the immunomodulatory efficacy of nisin in vitro. MTT-based cytotoxicity assay demonstrated nisin's cytotoxicity on human T-cell lymphoma Jurkat cells, Molt-4 cells and freshly cultured human lymphocytes at over 200 µM concentration (IC(50) 225 µM). The cell death mechanism induced by nisin in all these lymphocyte types was independent of oligonucleosomal DNA fragmentation, as analyzed by agarose gel electrophoresis and comet assay. Additionally, scanning electron microscope and fluorescence microscopy demonstrated the ability of nisin to activate human PMNs in vitro. Nisin-activated neutrophils extruded intact nuclear chromatin to form NETs, well known for neutralization of virulence factors and extermination of bacterial pathogens. Nisin's presence also elevated neutrophil intracellular superoxide levels, normally produced by activated NADPH oxidase and prerequisite to NET formation. These nisin-induced responses in cellular representatives of two separate branches of human immune system-adaptive and innate-although leading to cell death, did not include DNA fragmentation. From these findings, we propose that nisin might trigger similar AICD mechanisms in lymphocytes and neutrophils, different from conventional apoptosis which involves DNA fragmentation.

The current work was carried out under a screening program targeted at isolation of bioactive Streptomyces species from soil samples. A total of 54 Streptomyces species were isolated from soil samples, out of which 4 isolates were found to be promising. These isolates were identified as Streptomyces spectabilis, Streptomyces purpurascens, Streptomyces coeruleorubidus and Streptomyces lavendofoliae and their sequences have been deposited in the GenBank. The influence of culture conditions including, incubation time, incubation temperature, initial pH and different carbon and nitrogen sources on growth and bioactive compound formation was investigated. Isolate R1, identified as Streptomyces spectabilis, showed maximum bioactive metabolite production with cellobiose and peptone as the carbon and nitrogen sources, on the 5 th day at pH 5 at 30˚C. The optimum conditions for production by isolate R3, identified as Streptomyces purpurascens, were observed to be starch and casein as the carbon and nitrogen sources, pH 7, temperature 30˚C and an incubation period of eight days. For isolate R5, identified as Streptomyces coeruleorubidus, maximal production resulted on the sixth day at pH 6 and temperature of 35˚C with mannitol and JBM. Isolate Y8, identified as Streptomyces lavendofoliae, was found to produce high levels of bioactive metabolites in the medium supplemented with starch and peptone on the 10 th day at pH 7 and at an incubation temperature of 30˚C. The four strains tested here behaved differently, each one requiring specific conditions for maximum growth as well as bioactive metabolite production.

Rice blast resistance gene, Pi54 provides broad-spectrum resistance against different strains of Magnaporthe oryzae. Understanding the cellular localization of Pi54 protein is an essential step towards deciphering its place of interaction with the cognate Avr-gene. In this study, we investigated the subcellular localization of Pi54 with Green Fluorescent Protein (GFP) as a molecular tag through transient and stable expression in onion epidermal cells (Allium cepa) and susceptible japonica cultivar rice Taipei 309 (TP309), respectively. Confocal microscopy based observations of the onion epidermal cells revealed nucleus and cytoplasm specific GFP signals. In the stable transformed rice plants, GFP signal was recorded in the stomata, upper epidermal cells, mesophyll cells, vascular bundle, and walls of bundle sheath and bulliform cells of leaf tissues. These observations were further confirmed by Immunocytochemical studies. Using GFP specific antibodies, it was found that there was sufficient aggregation of GFP::Pi54protein in the cytoplasm of the leaf mesophyll cells and periphery of the epidermal cells. Interestingly, the transgenic lines developed in this study could show a moderate level of resistance to Xanthomonas oryzae and Rhizoctonia solani, the causal agents of the rice bacterial blight and sheath blight diseases, respectively. This study is a first detailed report, which emphasizes the cellular and subcellular distribution of the broad spectrum blast resistance gene Pi54 in rice and the impact of its constitutive expression towards resistance against other fungal and bacterial pathogens of rice.

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