Bacteriophage Adsorption Rate and Optimal Lysis Time

Shao, Yongping; Wang, Ing-Nang

doi:10.1534/genetics.108.090100

Cited by 186 publications

(205 citation statements)

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“…Optimal lysis time is positively correlated with adsorption rate, and phage strains with higher adsorption rates show faster lysis than those with lower adsorption rates (21). To determine if the adsorption rate provides any contribution to the delay in lysis time observed with the T-lacking mutant, we measured adsorption rates of the wild-type and mutant phages in infection of exponentially growing BL21 at 37°C (Fig.…”

Section: Resultsmentioning

confidence: 99%

Lysis Delay and Burst Shrinkage of Coliphage T7 by Deletion of Terminator Tφ Reversed by Deletion of Early Genes

Nguyen

Kang

2014

J Virol

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Bacteriophage T7 terminator T is a class I intrinsic terminator coding for an RNA hairpin structure immediately followed by oligo(U), which has been extensively studied in terms of its transcription termination mechanism, but little is known about its physiological or regulatory functions. In this study, using a T7 mutant phage, where a 31-bp segment of T was deleted from the genome, we discovered that deletion of T from T7 reduces the phage burst size but delays lysis timing, both of which are disadvantageous for the phage. The burst downsizing could directly result from T deletion-caused upregulation of gene 17.5, coding for holin, among other T downstream genes, because infection of gp17.5-overproducing Escherichia coli by wild-type T7 phage showed similar burst downsizing. However, the lysis delay was not associated with cellular levels of holin or lysozyme or with rates of phage adsorption. Instead, when allowed to evolve spontaneously in five independent adaptation experiments, the Tlacking mutant phage, after 27 or 29 passages, recovered both burst size and lysis time reproducibly by deleting early genes 0.5, 0.6, and 0.7 of class I, among other mutations. Deletion of genes 0.5 to 0.7 from the T-lacking mutant phage decreased expression of several T downstream genes to levels similar to that of the wild-type phage. Accordingly, phage T7 lysis timing is associated with cellular levels of T downstream gene products. This suggests the involvement of unknown factor(s) besides the known lysis proteins, lysozyme and holin, and that T plays a role of optimizing burst size and lysis time during T7 infection. IMPORTANCEBacteriophages are bacterium-infecting viruses. After producing numerous progenies inside bacteria, phages lyse bacteria using their lysis protein(s) to get out and start a new infection cycle. Normally, lysis is tightly controlled to ensure phage progenies are maximally produced and released at an optimal time. Here, we have discovered that phage T7, besides employing its known lysis proteins, additionally uses its transcription terminator T to guarantee the optimal lysis of the E. coli host. T, positioned in the middle of the T7 genome, must be inactivated at least partially to allow for transcription-driven translocation of T7 DNA into hosts and expression of T downstream but promoter-lacking genes. What role is played by T before inactivation? Without T, not only was lysis time delayed but also the number of progenies was reduced in this study. Furthermore, T7 can overcome T deletion by further deleting some genes, highlighting that a phage has multiple strategies for optimizing lysis. Bacteriophage T7 is an obligate lytic Escherichia coli phage that has been extensively studied for more than 60 years (1). T7 RNA polymerase, the only RNA polymerase (gp1) in the T7 genome, is one of the best-characterized RNA polymerases, and its transcription mechanisms have been studied in detail (2-6). In the T7 genome, an intrinsic termination signal for T7 RNA polymerase, terminator T, is positioned ...

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Section: Resultsmentioning

confidence: 99%

Lysis Delay and Burst Shrinkage of Coliphage T7 by Deletion of Terminator Tφ Reversed by Deletion of Early Genes

Nguyen

Kang

2014

J Virol

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“…Data reveal precision in the timing of lysis-individual cells infected by a single virus lyse on average at 65 min, with an SD of 3.5 min, implying a coefficient of variation of ≈ 5% (SI Appendix, section S9). Such precision is expected given the existence of an optimal lysis time (46)(47)(48)(49). Intuitively, if λ lysis is early then there are no viral progeny.…”

Section: Discussionmentioning

confidence: 99%

First-passage time approach to controlling noise in the timing of intracellular events

Ghusinga

Dennehy

Singh

2017

Proc. Natl. Acad. Sci. U.S.A.

111

127

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In the noisy cellular environment, gene products are subject to inherent random fluctuations in copy numbers over time. How cells ensure precision in the timing of key intracellular events despite such stochasticity is an intriguing fundamental problem. We formulate event timing as a first-passage time problem, where an event is triggered when the level of a protein crosses a critical threshold for the first time. Analytical calculations are performed for the first-passage time distribution in stochastic models of gene expression. Derivation of these formulas motivates an interesting question: Is there an optimal feedback strategy to regulate the synthesis of a protein to ensure that an event will occur at a precise time, while minimizing deviations or noise about the mean? Counterintuitively, results show that for a stable long-lived protein, the optimal strategy is to express the protein at a constant rate without any feedback regulation, and any form of feedback (positive, negative, or any combination of them) will always amplify noise in event timing. In contrast, a positive feedback mechanism provides the highest precision in timing for an unstable protein. These theoretical results explain recent experimental observations of single-cell lysis times in bacteriophage λ. Here, lysis of an infected bacterial cell is orchestrated by the expression and accumulation of a stable λ protein up to a threshold, and precision in timing is achieved via feedforward rather than feedback control. Our results have broad implications for diverse cellular processes that rely on precise temporal triggering of events.first-passage time | event timing | stochastic gene expression | feedback control | single cell T iming of events in many cellular processes, such as cell-cycle control (1-3), cell differentiation (4, 5), sporulation (6, 7), apoptosis (8, 9), development (10, 11), temporal order of gene expression (12)(13)(14), and so on, depend on regulatory proteins reaching critical threshold levels. Triggering of these events in single cells is influenced by fluctuations in protein levels that arise naturally due to noise in gene expression (15)(16)(17)(18). Increasing evidence shows considerable cell-to-cell variation in timing of intracellular events among isogenic cells (19-21), and it is unclear how noisy expression generates this variation. Characterization of control strategies that buffer stochasticity in event timing are critically needed to understand reliable functioning of diverse intracellular pathways that rely on precision in timing.Mathematically, noise in the timing of events can be investigated via the first-passage time (FPT) framework, where an event is triggered when a stochastic process (single-cell protein level) crosses a critical threshold for the first time. There is already a rich tradition of using such FPT approaches to study timing of events in biological and physical sciences (22-26). Following this tradition, exact analytical expression for the FPT distribution is computed in experimentally valida...

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“…These studies reported that no hindrance is seen when host concentration is low because of the low fraction of volume they occupy. However, at high host concentrations, the balance shifts and the host density acts as a barrier to diffusion, hindering the process of viral spread in the surrounding medium (32,35,36). Also, as mentioned, antibiotics actually lengthened the productive/plaque enlargement phase (during which the bulk of plaque enlargement occurs), as bacterial lawns take longer to mature under these conditions, thus delaying the onset of lawn maturation and giving more time to progeny virions to add onto the growing plaque during this period.…”

Section: Discussionmentioning

confidence: 99%

Methicillin-Resistant Staphylococcus aureus Phage Plaque Size Enhancement Using Sublethal Concentrations of Antibiotics

Kaur

Harjai

Chhibber

2012

Appl Environ Microbiol

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Phage therapy presents an alternative approach against the emerging methicillin-resistant Staphylococcus aureus (MRSA) threat. Some of the problems encountered during isolation of MRSA phages include the high prevalence of enteric phages in natural sources, nonspecific absorption of viable phage, and the formation of pinpoint or tiny plaques. The phage isolated in this study, MR-5, also formed tiny plaques against its host S. aureus ATCC 43300 (MRSA), making its detection and enumeration difficult. An improved method of increasing the plaque size of MRSA phage by incorporating sublethal concentrations of three different classes of antibiotics (inhibitors of protein synthesis) in the classical double-layer agar (DLA) method was investigated. The ␤-lactam and quinolone antibiotics commonly employed in earlier studies for increasing the plaque size did not show any significant effect on the plaque size of isolated MR-5 phage. Linezolid (oxazolidinone class), tetracycline, and ketolide antibiotics brought significant enhancements (3 times the original size) in the plaque size of MR-5 phage. Prior treatment with these antibiotics resulted in significant reductions in the time of adsorption and the latent period of MR-5 phage. To rule out whether the action of linezolid (which brought the maximum increase in plaque size) was specific for a single phage only, its effect on the plaque size of seven other S. aureus-specific phages was also assessed. Significant enhancements in the plaque size of these phages were observed. These results indicate that this modification can therefore safely be incorporated in the traditional DLA overlay method to search for new MRSA-virulent phages. Staphylococcus aureus is a leading cause of both communityand hospital-acquired infections, including skin and soft tissue infections, wound infections, endocarditis, osteomyelitis, and lifethreatening bacteremia (6, 23). The emergence of high levels of penicillin resistance, followed by the development and spread of strains resistant to the action of nafcillin, oxacillin, methicillin, and now, even vancomycin has made therapy of staphylococcal disease a global challenge (23,24,34). More people in the United States now die from methicillin-resistant S. aureus (MRSA) infection than from AIDS (33). Hence, there is an unmet medical urgency and pressing need to develop novel antibacterials to tackle the staphylococcal menace. Phage therapy can serve as one of the stand-alone approaches for treating staphylococcal infections refractory to the action of commonly deployed antibiotics (6).The first step in investigating the potential of therapy using phage or its products against MRSA infections involves the isolation and selection of a lytic bacteriophage covering a maximum number of MRSA strains, both from community and hospital sources. The difficulty encountered in the isolation of phages specific to staphylococcal strains is partly attributed to the prevalence of enteric phages in sewage samples and partly to the high level of nonspecific adsorpti...

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Bacteriophage Adsorption Rate and Optimal Lysis Time

Cited by 186 publications

References 28 publications

Lysis Delay and Burst Shrinkage of Coliphage T7 by Deletion of Terminator Tφ Reversed by Deletion of Early Genes

Lysis Delay and Burst Shrinkage of Coliphage T7 by Deletion of Terminator Tφ Reversed by Deletion of Early Genes

First-passage time approach to controlling noise in the timing of intracellular events

Methicillin-Resistant Staphylococcus aureus Phage Plaque Size Enhancement Using Sublethal Concentrations of Antibiotics

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