Isolation and identification of a psychrotrophic Acinetobacter sp. CR9 and characterization of its alkaline lipase

Forty three psychrotrophic bacteria were isolated from soil samples collected from Chandra river in sub-alpine region of western Himalaya, India. Among these, 11 isolates were found positive for lipase production at low temperature. Of 11 isolates, CR9 produced largest zone of clearance on plate assay and was able to produce lipase under wide range of pH. The isolate CR9 was identified as Acinetobacter sp. based on morphological and physiochemical characterization and 16S rRNA gene sequencing analysis. According to 16S rRNA gene sequencing data the closest phylogenetic neighbor for strain CR9 was Acinetobacter lwoffii (98.9%). The partially purified lipase from strain CR9 exhibited maximum activity at temperature 40 degrees C and pH optima at 8.0. Cu(2+), Mo(2+), Mg(2+), Zn(2+), phenylmethanesulfonyl fluoride (PMSF), dithiothreitol (DTT) and beta-mercaptoethanol (2-ME) enhanced the enzyme activity, whereas Ca(2+) and ethylenediaminetetraacetic acid (EDTA) had inhibitory effect. Lipase hydrolyzed wide range of short chain fatty acid esters of p-nitrophenyl. The organism CR9 also hydrolyzed tributyrin, Tween 80, soybean oil, mustard oil and olive oil. The results highlight the relevance of unexplored microbes from cold environments of western Himalaya for the isolation of novel lipase producing bacteria.