Bacteria and archaea face continual onslaughts of rapidly diversifying viruses and plasmids. Many prokaryotes maintain adaptive immune systems known as clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated genes (Cas). CRISPR-Cas systems are genomic sensors that serially acquire viral and plasmid DNA fragments (spacers) that are utilized to target and cleave matching viral and plasmid DNA in subsequent genomic invasions, offering critical immunological memory. Only 50% of sequenced bacteria possess CRISPR-Cas immunity, in contrast to over 90% of sequenced archaea. To probe why half of bacteria lack CRISPR-Cas immunity, we combined comparative genomics and mathematical modeling. Analysis of hundreds of diverse prokaryotic genomes shows that CRISPR-Cas systems are substantially more prevalent in thermophiles than in mesophiles. With sequenced bacteria disproportionately mesophilic and sequenced archaea mostly thermophilic, the presence of CRISPR-Cas appears to depend more on environmental temperature than on bacterial-archaeal taxonomy. Mutation rates are typically severalfold higher in mesophilic prokaryotes than in thermophilic prokaryotes. To quantitatively test whether accelerated viral mutation leads microbes to lose CRISPR-Cas systems, we developed a stochastic model of virus-CRISPR coevolution. The model competes CRISPR-Cas-positive (CRISPR-Cas+) prokaryotes against CRISPR-Cas-negative (CRISPR-Cas−) prokaryotes, continually weighing the antiviral benefits conferred by CRISPR-Cas immunity against its fitness costs. Tracking this cost-benefit analysis across parameter space reveals viral mutation rate thresholds beyond which CRISPR-Cas cannot provide sufficient immunity and is purged from host populations. These results offer a simple, testable viral diversity hypothesis to explain why mesophilic bacteria disproportionately lack CRISPR-Cas immunity. More generally, fundamental limits on the adaptability of biological sensors (Lamarckian evolution) are predicted.

Well-studied innate immune systems exist throughout bacteria and archaea, but a more recently discovered genomic locus may offer prokaryotes surprising immunological adaptability. Mediated by a cassette-like genomic locus termed Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), the microbial adaptive immune system differs from its eukaryotic immune analogues by incorporating new immunities unidirectionally. CRISPR thus stores genomically recoverable timelines of virus-host coevolution in natural organisms refractory to laboratory cultivation. Here we combined a population genetic mathematical model of CRISPR-virus coevolution with six years of metagenomic sequencing to link the recoverable genomic dynamics of CRISPR loci to the unknown population dynamics of virus and host in natural communities. Metagenomic reconstructions in an acid-mine drainage system document CRISPR loci conserving ancestral immune elements to the base-pair across thousands of microbial generations. This ‘trailer-end conservation’ occurs despite rapid viral mutation and despite rapid prokaryotic genomic deletion. The trailer-ends of many reconstructed CRISPR loci are also largely identical across a population. ‘Trailer-end clonality’ occurs despite predictions of host immunological diversity due to negative frequency dependent selection (kill the winner dynamics). Statistical clustering and model simulations explain this lack of diversity by capturing rapid selective sweeps by highly immune CRISPR lineages. Potentially explaining ‘trailer-end conservation,’ we record the first example of a viral bloom overwhelming a CRISPR system. The polyclonal viruses bloom even though they share sequences previously targeted by host CRISPR loci. Simulations show how increasing random genomic deletions in CRISPR loci purges immunological controls on long-lived viral sequences, allowing polyclonal viruses to bloom and depressing host fitness. Our results thus link documented patterns of genomic conservation in CRISPR loci to an evolutionary advantage against persistent viruses. By maintaining old immunities, selection may be tuning CRISPR-mediated immunity against viruses reemerging from lysogeny or migration.

SUMMARY HIV latency is the chief obstacle to eradicating HIV but is widely believed to be an evolutionary accident providing no lentiviral fitness advantage. However, findings of latency being “hardwired” into HIV’s gene-regulatory circuitry appear inconsistent with latency being an evolutionary accident, given HIV’s rapid mutation rate. Here, we propose that latency is an evolutionary “bet-hedging” strategy whose frequency has been optimized to maximize lentiviral transmission by reducing viral extinction during mucosal infections. The model quantitatively fits the available patient data, matches observations of high-frequency latency establishment in cell culture and primates, and generates two counterintuitive but testable predictions. The first prediction is that conventional CD8-depletion experiments in SIV-infected macaques increase latent cells more than viremia. The second prediction is that strains engineered to have higher replicative fitness—via reduced latency—will exhibit lower infectivity in animal-model mucosal inoculations. Therapeutically, the theory predicts treatment approaches that may substantially enhance “activate-and-kill” HIV-cure strategies.

Classic studies proposed that stochastic variability ("noise") can drive biological fate switching, enhancing evolutionary success. Now, Ho et al. report that HIV's reactivation from dormant (latently infected) patient cells-the major barrier to an HIV cure-is inherently stochastic. Eradicating an incompletely inducible (probabilistic) viral phenotype will require inventive approaches.

The rapid evolution of RNA-encoded viruses such as HIV presents a major barrier to infectious disease control using conventional pharmaceuticals and vaccines. Previously, it was proposed that defective interfering particles could be developed to indefinitely control the HIV/AIDS pandemic; in individual patients, these engineered molecular parasites were further predicted to be refractory to HIV’s mutational escape (i.e., be ‘resistance-proof’). However, an outstanding question has been whether these engineered interfering particles—termed Therapeutic Interfering Particles (TIPs)—would remain resistance-proof at the population-scale, where TIP-resistant HIV mutants may transmit more efficiently by reaching higher viral loads in the TIP-treated subpopulation. Here, we develop a multi-scale model to test whether TIPs will maintain indefinite control of HIV at the population-scale, as HIV (‘unilaterally’) evolves toward TIP resistance by limiting the production of viral proteins available for TIPs to parasitize. Model results capture the existence of two intrinsic evolutionary tradeoffs that collectively prevent the spread of TIP-resistant HIV mutants in a population. First, despite their increased transmission rates in TIP-treated sub-populations, unilateral TIP-resistant mutants are shown to have reduced transmission rates in TIP-untreated sub-populations. Second, these TIP-resistant mutants are shown to have reduced growth rates (i.e., replicative fitness) in both TIP-treated and TIP-untreated individuals. As a result of these tradeoffs, the model finds that TIP-susceptible HIV strains continually outcompete TIP-resistant HIV mutants at both patient and population scales when TIPs are engineered to express >3-fold more genomic RNA than HIV expresses. Thus, the results provide design constraints for engineering population-scale therapies that may be refractory to the acquisition of antiviral resistance.

In 50% of progressing HIV-1 patients, CXCR4-tropic (X4) virus emerges late in infection, often overtaking CCR5-tropic (R5) virus as the dominant viral strain. This “phenotypic switch” is strongly associated with rapidly declining CD4+ T cell counts and AIDS onset, yet its causes remain unknown. Here, we analyze a mathematical model for the mechanism of X4 emergence in late-stage HIV infection and use this analysis to evaluate the utility of a promising new class of antiretroviral drugs—CCR5 inhibitors—in dual R5, X4 infection. The model shows that the R5-to-X4 switch occurs as CD4+ T cell activation levels increase above a threshold and as CD4+ T cell counts decrease below a threshold during late-stage HIV infection. Importantly, the model also shows that highly active antiretroviral therapy (HAART) can inhibit X4 emergence but that monotherapy with CCR5 blockers can accelerate X4 onset and immunodeficiency if X4 infection of memory CD4+ T cells occurs at a high rate. Fortunately, when CXCR4 blockers or HAART are used in conjunction with CCR5 blockers, this risk of accelerated immunodeficiency is eliminated. The results suggest that CCR5 blockers will be more effective when used in combination with CXCR4 blockers and caution against CCR5 blockers in the absence of an effective HAART regimen or during HAART failure.

Historically landmark experiments showed that capsule switching is critical for Streptococcus pneumonia survival. Further studies demonstrated that capsule ‘transformation’ occurs via DNA uptake. In this issue of Cell Host and Microbe, Bikard et al. (2012) show that CRISPR-Cas systems inhibit DNA uptake, selecting for the outgrowth of CRISPR-defective pneumococci.

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