AME 45:15-29 (2006)  -  doi:10.3354/ame045015

Characterization of a cold-active bacteriophage on two psychrophilic marine hosts

Llyd E. Wells1,2,*, Jody W. Deming1

1School of Oceanography, University of Washington, Box 357940, Seattle, Washington 98195-7940, USA
2Present address: Center for Northern Studies, Sterling College, PO Box 72, Craftsbury Common, Vermont 05827, USA

ABSTRACT: A cold-active bacteriophage designated 9A was isolated against Colwellia psychrerythraea Strain 34H at near in situ temperature (–1°C) by enrichment of seawater from an Arctic nepheloid layer, using a newly developed isothermal overlay technique. Phage 9A is classified as a Siphoviridae with a genome size of 80 to 90 kb. In addition to 34H, 9A infects C. demingiae ACAM 459T; no other hosts (of 22 tested) were identified. In replete media, 9A formed plaques on 34H from –6 to 4°C and on C. demingiae from –6 to 8°C; the temperature range of plaque formation on 34H could be extended to 8°C by prior host starvation. An indirect plating method and microscopic evaluation also determined phage production at temperatures between –13 and –10°C. At –1°C, 34H had a broader salinity range of plaque formation than C. demingiae: 20 to 50 (but not 65) psu vs. 27 to 34 (but not 50) psu. As monitored by epifluorescence microscopy, phage production by 34H was observed at 1, 10, 100 and 200 atm (all at –1°C), but not at 400 or 600 atm. The 9A–34H system commonly had a low efficiency of plating (EOP; typically ~1%) which varied with culture age. Despite repeated attempts, no meaningful adsorption rate could be determined at –1 or 8°C. This result, the low EOP, and the effect of starvation on plaque formation suggest that fluctuating host phenotypes may play an important role in the dynamics of this system. One-step growth curves (using 34H as host) revealed a longer latent period (4 to 5 vs. 2.5 to 3 h) and greater burst size (55 vs. 5) at –1 than 8°C; at temperatures between –10 and –12°C, the estimated latent period was 5–10 d and the burst size 5. At both –1 and 8°C, rise times were comparable to latent periods. Although the cycle from infection to burst at –1°C required only 10 to 20% of the generation time of 34H at this temperature, the amount of viral DNA synthesized was comparable to the size of the host genome, suggesting very efficient and cold-active virus-encoded enzymes.

KEY WORDS: Virus · Phage 9A · Colwellia psychrerythraea 34H · Colwellia demingiae · Arctic · Adsorption · One-step growth curve · Pressure · Salinity

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