Measuring Viral Life-History Traits

Our method for the detection of selection on virulence is based on the competition of the non-virulent λ wildtype against the virulent λcI857. In order to verify the expected differences in life-history traits between those strains and to obtain rough parameter estimates for the simulations we measured the viral life-history traits virus production (PFU/mL), genome integration rate (% lysogenized) and vertical transmission (CFU/mL). We determined these traits for all constructed viruses (λcI857CFP, λcI857YFP and λCFP, λYFP) by independent life-history assays prior to competition in the chemostat. The life-history traits were measured by the following three independent assays. (1) Virus production (PFU/mL) (Figure S3A) was determined by growing lysogen cultures to OD600 nm = 0.6 at 30°C and shifting them to 35°C and 38°C for 2 h until lysis occurred. From these lysates, viral titers were determined by qPCR on a Roche LightCycler480 (primers F:5′AATGAAGGCAGGAAGTA3′ R:5′GCTTTCCATTCCATCGG3′). Viral titers were calculated from a calibration curve based on CP values of a dilution series of a lysate of λvir of known titer (3×109 pfu). (2) Vertical transmission (CFU/mL) (Figure S3A) was measured by diluting lysogen cultures of λCFP, λYFP and λcI857CFP, λcI857YFP to OD600 nm = 0.07 and growing them for 6 h at 35°C and 38°C in eight replicates each in 96-well plates on a Titramax shaker (Heidolph, Germany) at 900 rpm. Every hour OD600 nm was measured in an Infinity200 microplate reader (Tecan, Austria). OD600 nm values were converted to CFU's by a calibration curve which was obtained by plating. (3) Lysogenization rate (Figure S3B) was determined by challenging non-infected E.coli MG1655 with 10⁸ PFU/mL free virus particles of λCFP, λYFP, λcI857CFP and λcI857YFP for 24 h. After 24 h, the proportion of lysogenized (fluorescent) cells was determined by flow cytometry.