BIOTECHNOLOGICAL STUDIES ON SALINITY-TOLERANT MICROORGANISMS.

Abstract

Halophilic and halo-tolerant bacteria are micro-organisms that living in hyper saline environments. They have many potential in several fields in life such as Industry and agriculture. In this study a total of 17 isolates were obtained from the four saline ponds of Qarun Lake on Nutrient Agar (NA) medium prepared using ponds' water. The 16S rRNA analysis showed that our isolates affiliated with four groups, Firmicutes (47.06% of the isolates) that include family Bacillaceae with six different genera including Bacillus, Halobacillus, Lentibacillus, Oceanobacillus, Thalassobacillus and Virgibacillus, Gammaproteobacteria (35.29% of the isolates) including two genera Halomonas and Salinivibrio, two within Bacteriodetes group (11.76% of the isolates) and one of Alphaproteobacteria (5.88%). Firmicutes seems to be the more dominant group, including microorganisms of various genera, especially Bacillus (37.5%) followed by Halobacillus, Oceanobacillus, Lentibacillus and Virgibacillus (12.5% for each genera). 52.94% of isolates were gram negative, 35.29% were gram positive and 11.76% were gram variable. All isolates were catalase producers and motile. Halotolerance assay observed that only two isolates (11.76 %) were able to grow on the absence of salt and considered as halotolerant, while fifteen isolates (88.24%) were able to grow on NA media with different salt concentrations and considered as halophilic bacteria. Among them, thirteen isolates (76.47%) were able to grow on NA media with 7% NaCL. All isolates grew on NA with 12.5% NaCl; nine iso-lates (52.94%) were able to grow on NA media with 20% salinity, while three isolates (17.65%) appeared on NA media with salt concentration of 22% and considered extremely halophilic bacteria. All isolates were tested for plant growth promoting factors. The results showed that all isolates can produce IAA, while 47.06% of isolates can fix Nitrogen and 47.06 % have ability for phosphate solubilization. Concerning hydrolytic enzymes activi-ties of the isolates, it was found that 58.82% and 76.47% of isolates pro-duced lipase using olive oil and tween80 as a substrate respectively. It was also found that 47.06% of isolates showed ability for esterase production and same percentage for casease production. Gelatinase was excereted by 58.82%, while chitinase was produced by 70.59% of isolates. Five isolates were screened for biosurfactants and other bioactive compounds production. These isolates namly as QSLA1, QSLA7, QSLA14, QSLA16 and QSLA17 which were similar to, Halomonas sp.RS-17, Salinivibrio sp. pr6, Lentibacillus sp.strain BCHS25, Uncultured bacterium clone QAMU23 and Bacillus sp.strain6 in the GenBank database, respectively. Among them two isolates QSLA16 and QSLA17 had NRPs genes encoding for lipopeptide biosurfactants synthetase enzymes and HPLC analysis confirmed presence of different biosurfactants such as, surfactin, fengycin and mycosubtilin. The culture of the two isolates, QSLA1and QSLA7 were analysed by Gas chromatography–mass spectrometry (GC-MS) and showed ability for production of antifungal, antimicrobial and antitumer metabolites such as Desulphosinigrin, Undeca-2,4,6,8,10-pentaenal, 11-(2-furyl)-, oxime and Strychane, 1-acetyl-20à-hydroxy-16-methylene. Accordingly, QSLA1 had in vitro antibacterial activity against Salmonella typhi and Acinetobacter baumannii. Whereas, QSLA16 was able to inhibit the growth of bacterial pathogens Staphylococcus aureus, Klebsiella pneumonia and Salmonella typhi in contaminated salty whey. In this regard, QSLA1 showed in vitro antifungal activity against Alternaria solani and Fusarium oxysporium f.sp. lycopersici, and had in vivo a siginificant effect on the growth of Fusarium oxysporium f.sp. lycopersici infecting tomato seedlings compared to control and therefore can be used among biological control systems under saline conditions