A Single-Cell Approach Reveals Variation in Cellular Phage-Producing Capacities

Abstract

Virus burst size, a component of viral fitness, is the average number of viral particles released from a single infected cell. In this study, we estimated bacteriophage lambda (λ) burst size mean and distribution at different lysis times. To estimate phage λ burst sizes at single-cell level, we employed a lysis-deficient E. coli lysogen, which allowed chemical lysis at desired times after the induction of lytic cycle. Induced cultures of E. coli lysogen were diluted and aliquoted into the wells of 96-well plates. A high dilution rate results in mostly empty wells and minimizes the probability of having multiple cells in wells that do receive cells. Burst size was estimated by titering single-cell lysates obtained after chemical lysis at desired times. Our data shows that the viral burst size initially increases exponentially with the lysis time, and then saturates at longer lysis times. We also demonstrate that cell-to-cell variation or “noise” in lysis timing does not significantly contribute to the burst size noise. The burst size noise remains constant with increasing mean burst size. The most likely explanation for the experimentally observed constant burst size noise is that cell-to-cell differences in burst size originate from differences in cellular capacity to produce phages. The mean burst size measured at different lysis times is positively correlated to cell volume, which may determine the cellular phage production capacity. However, experiments controlling for cell size indicates that there are other factors in addition to cell size that determine this cellular capacity.ARTICLE IMPORTANCEPhages produce offspring by hijacking a cell’s replicative machinery. Previously, it was noted that the variation in the number of phages produced by single infected cells far exceeded cell size variation. It has been hypothesized that this variation is a consequence of variation in the timing of host cell lysis. Here we show that cell-to-cell variation in lysis timing does not significantly contribute to the burst size variation. We suggest that the constant burst size variation across different host lysis times results from cell-to-cell differences in capacity to produce phages. We find that the mean burst size measured at different lysis times is positively correlated to cell volume, which may determine the cellular phage production capacity. However, experiments controlling for cell size indicates that there are other factors in addition to cell size that determine this cellular capacity.

An Optimal Lysis Time Maximizes Bacteriophage Fitness in Quasi-Continuous Culture

“…However, the deletion did not greatly delay lysis because holin-mediated membrane permeabilization reduced protein synthesis, ceasing phage production, and thus eliminating the trade-off that is an important assumption of optimality models. In contrast, our holin mutants with significantly longer lysis times show continued phage production ( 40 ). Our experimental set up was sensitive to dilution due to a constant flow rate, and the strains with lysis times that deviate from the optimum range show reduced phage productivity over subsequent transfers ( Fig.…”

Anaerobiosis, a neglected factor in phage-bacteria interactions

“…Some studies have indicated that burst size can be influenced by the ratio of host cell volume to virus volume ( 71 ). In the case of phages, Choi et al ( 72 ) observed that the burst size of T4 phage is greater in larger host cells, while Kannoly et al ( 73 ) found a positive correlation between burst size and cell volume in lambda phage. In fact, the likelihood of lysogeny in lambda phage increases in small cells during both the stationary and exponential phases ( 74 ), possibly due to the larger viral progeny produced in larger cells.…”

Phenotypic flux: The role of physiology in explaining the conundrum of bacterial persistence amid phage attack

“…The latent period determines the time that a phage spends on linear reproduction within a cell, i.e. the burst size ( Kannoly et al. 2021 ), as opposed to the potential geometric increase attained by infecting new cells.…”

“…In this case, very short latent periods would be disadvantageous as they would lead to few or even no phage progeny released, which could explain the low variance observed in empirical measurements of latent period distributions ( Cahill and Young 2019 ). Except for very short latent periods, however, the variation in burst size is potentially only weakly influenced by variation in latent period ( Kannoly et al. 2021 ).…”