Screening of essential genes for cold growth in Pseudomonas extremaustralis.

PAULA M. TRIBELLI; ESMERALDA SOLAR VENERO; LAURA J. RAIGER IUSTMAN; NANCY I. LOPEZ

Mendoza

Congreso; • XLVIII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB); 2012

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB).

P. extremaustralis is an Antarctic bacterium capable to grow and survive at low temperatures with high stress resistance in association with the accumulation of large amounts of polyhydroxybutyrate (PHB). A mini Tn5 library was constructed to detect essential genes for growth under cold conditions. The resulting collection was screened in Luria-Broth agar plates supplemented with 0.25% sodium octanoate and were incubated at 10C during 7 days; controls were performed at 28C. Five mutants were detected and two of them, C42 and C60, were identified using a PCR based strategy. The sequence analysis indicated that the transposon insertion site for C42 was located in the pyrroloquinoline quinone biosynthesis cluster (pqq), specifically in the pqqB gene and in the wabH gene coding for a glycosyltransferase for C60. At 28C in liquid cultures both clones presented similar final OD600nm compared with the wild type strain and showed PHB accumulation. However, the C42 clone showed a higher growth rate during exponential phase, while the C60 exhibited a long lag phase and high adherence. In contrast, none of the mutants grew at 10C while the wild type strain reached an OD600nm value of approximately 1.The phenotype was not altered by changes in the carbon source. The results describe for the first time the involvement of these two genes in cold survival and adaptability.