INDUCTION OF MULTIPLE PROPHAGES FROM AN ANTARCTIC MARINE BACTERIUM

LOPEZ JOSÉ LUIS; HERNÁNDEZ EDGARDO A.; ARÁN MARTÍN; CORIA SILVIA; MAC CORMACK WP

Ceske Budejovice

Congreso; 6th International Conference on Polar and Alpine Microbiology; 2015

The genomic sequencing of the marine bacterium Bizionia argentinensis (B.a), isolated from Antarctic surfacewater, has promoted the interest to know the functionality of their gene content. As part of this goal, we arestudying the presence of a protein (that we called C24) showing structural homology with a T4 phage tail fiberprotein. As it was previously reported that approximately 70% of sequenced bacterial genomes contain prophagelikestructures, this finding of C24 suggested us the presence of genetic elements of the type of the prophages inthe genome of B.a. Some prophages may contain morons or lysogenic conversion genes that could change thephenotype or enhance the ecological fitness of lysogens. Bacterial viruses represent one of three major mobilegenetic elements and contribute significantly to horizontal gene transfer in bacterial genomes. However little isknown about the characteristics, function and ecological relevance of marine phages. For this reason, in this workwe search for the presence of prophages in a recently sequenced marine Antarctic bacterium.METHODOLOGYInduction of Ba with mitomycin C. The traditional and most common approach to studying prophages or temperatephages is to induce lysogenic bacteria with mitomycin C or UV exposure. We grown B.a overnight in marine brothincubated at 15°C in a rotatory shaker (200 rpm). Two ml of an exponential growth culture of B.a (optical densityat 600 nm [OD600], ca. 1.0) was transferred to 50 ml of fresh marine broth (six replicates containing, 0.2 μg/mlmitomycin C and six without mitomycin C). After 24 hours, cells in both, control and treated groups, were pelletedby centrifugation (7,500g for 10 min) washed twice and resuspended in 50 ml of fresh marine broth. All cultureswere incubated and monitored by OD600 at 8, 12, 24, 30 and 36 hours. Samples were kept at 4°C in the dark.CsCl purification of induced phages. 50ml of induced viral lysate was centrifuged (10,000g, 4°C) and phage particlesin the supernatant were purified using CsCl (as in protocol 8, Chapter 2.51, Maniatis). Phages particles werevisulized by TEM after negative stain.RESULTSFig. 1 shows the effect of Mitomycin C treatment on B.a growth. A clear deleterious effect, compatible with lysisby phages were observed.TEM of the samples confirmed the presence of virused and denoted that at least three different phages wereinduced from mitomycin C treated cultures of Bizionia argentinensis (Fig. 2)DISCUSSIONThe biological effects of the presence of these multiple inducible phages in Bizionia argentinensis are stillunknown, as also the genomic structure of them. Currently we are isolating the phages (lysis plaques formation)and subsequently its complete nucleotide genomic sequences will be done.To our knowledge, this is the first work showing the induction of lysogenic bacteriophages from an Antarcticmarine bacterium. Further studies about the abundance of lysogenic bacterial phages in the Antarcticenvironment and its biological relevance would provide a better view of the bacterial communities? dynamic ofthe coastal Antarctic marine environments.