Climate, demography, immunology, and virology combine to drive two decades of dengue virus dynamics in Cambodia.

The incidence of dengue virus disease has increased globally across the past half-century, with highest number of cases ever reported in 2019 and again in 2023. We analyzed climatological, epidemiological, and phylogenomic data to investigate drivers of two decades of dengue in Cambodia, an understudied endemic setting. Using epidemiological models fit to a 19-y dataset, we first demonstrate that climate-driven transmission alone is insufficient to explain three epidemics across the time series. We then use wavelet decomposition to highlight enhanced annual and multiannual synchronicity in dengue cycles between provinces in epidemic years, suggesting a role for climate in homogenizing dynamics across space and time. Assuming reported cases correspond to symptomatic secondary infections, we next use an age-structured catalytic model to estimate a declining force of infection for dengue through time, which elevates the mean age of reported cases in Cambodia. Reported cases in >70-y-old individuals in the 2019 epidemic are best explained when also allowing for waning multitypic immunity and repeat symptomatic infections in older patients. We support this work with phylogenetic analysis of 192 dengue virus (DENV) genomes that we sequenced between 2019 and 2022, which document emergence of DENV-2 Cosmopolitan Genotype-II into Cambodia. This lineage demonstrates phylogenetic homogeneity across wide geographic areas, consistent with invasion behavior and in contrast to high phylogenetic diversity exhibited by endemic DENV-1. Finally, we simulate an age-structured, mechanistic model of dengue dynamics to demonstrate how expansion of an antigenically distinct lineage that evades preexisting multitypic immunity effectively reproduces the older-age infections witnessed in our data.

Faster-growing parasites threaten host populations via patch-level population dynamics and higher virulence; a case study in Varroa mites (Mesostigmata: Varroidae) and honey bees (Hymenoptera: Apidae).

Honey bee parasites remain a critical challenge to management and conservation. Because managed honey bees are maintained in colonies kept in apiaries across landscapes, the study of honey bee parasites allows the investigation of spatial principles in parasite ecology and evolution. We used a controlled field experiment to study the relationship between population growth rate and virulence (colony survival) of the parasite Varroa destructor (Anderson and Trueman). We used a nested design of 10 patches (apiaries) of 14 colonies to examine the spatial scale at which Varroa population growth matters for colony survival. We tracked Varroa population size and colony survival across a full year and found that Varroa populations that grow faster in their host colonies during the spring and summer led to larger Varroa populations across the whole apiary (patch) and higher rates of neighboring colony loss. Crucially, this increased colony loss risk manifested at the patch scale, with mortality risk being related to spatial adjacency to colonies with fast-growing Varroa strains rather than with Varroa growth rate in the colony itself. Thus, within-colony population growth predicts whole-apiary virulence, demonstrating the need to consider multiple scales when investigating parasite growth-virulence relationships.

The three Ts of virulence evolution during zoonotic emergence.

There is increasing interest in the role that evolution may play in current and future pandemics, but there is often also considerable confusion about the actual evolutionary predictions. This may be, in part, due to a historical separation of evolutionary and medical fields, but there is a large, somewhat nuanced body of evidence-supported theory on the evolution of infectious disease. In this review, we synthesize this evolutionary theory in order to provide a framework for clearer understanding of the key principles. Specifically, we discuss the selection acting on zoonotic pathogens' transmission rates and virulence at spillover and during emergence. We explain how the direction and strength of selection during epidemics of emerging zoonotic disease can be understood by a three Ts framework: trade-offs, transmission, and time scales. Virulence and transmission rate may trade-off, but transmission rate is likely to be favoured by selection early in emergence, particularly if maladapted zoonotic pathogens have 'no-cost' transmission rate improving mutations available to them. Additionally, the optimal virulence and transmission rates can shift with the time scale of the epidemic. Predicting pathogen evolution, therefore, depends on understanding both the trade-offs of transmission-improving mutations and the time scales of selection.

Persistent effects of management history on honeybee colony virus abundances.

Infectious diseases are a major threat to both managed and wild pollinators. One key question is how the movement or transplantation of honeybee colonies under different management regimes affects honeybee disease epidemiology. We opportunistically examined any persistent effect of colony management history following relocation by characterising the virus abundances of honeybee colonies from three management histories, representing different management histories: feral, low-intensity management, and high-intensity "industrial" management. The colonies had been maintained for one year under the same approximate 'common garden' condition. Colonies in this observational study differed in their virus abundances according to management history, with the feral population history showing qualitatively different viral abundance patterns compared to colonies from the two managed population management histories; for example, higher abundance of sacbrood virus but lower abundances of various paralysis viruses. Colonies from the high-intensity management history exhibited higher viral abundances for all viruses than colonies from the low-intensity management history. Our results provide evidence that management history has persistent impacts on honeybee disease epidemiology, suggesting that apicultural intensification could be majorly impacting on pollinator health, justifying much more substantial investigation.

Knock-on community impacts of a novel vector: spillover of emerging DWV-B from Varroa-infested honeybees to wild bumblebees.

Novel transmission routes can directly impact the evolutionary ecology of infectious diseases, with potentially dramatic effect on host populations and knock-on effects on the wider host community. The invasion of Varroa destructor, an ectoparasitic viral vector in Western honeybees, provides a unique opportunity to examine how a novel vector affects disease epidemiology in a host community. This specialist honeybee mite vectors deformed wing virus (DWV), an important re-emerging honeybee pathogen that also infects wild bumblebees. Comparing island honeybee and wild bumblebee populations with and without V. destructor, we show that V. destructor drives DWV prevalence and titre in honeybees and sympatric bumblebees. Viral genotypes are shared across hosts, with the potentially more virulent DWV-B overtaking DWV-A in prevalence in a current epidemic. This demonstrates disease emergence across a host community driven by the acquisition of a specialist novel transmission route in one host, with dramatic community level knock-on effects.

Coevolution of parasite virulence and host mating strategies.

Parasites are thought to play an important role in sexual selection and the evolution of mating strategies, which in turn are likely to be critical to the transmission and therefore the evolution of parasites. Despite this clear interdependence we have little understanding of parasite-mediated sexual selection in the context of reciprocal parasite evolution. Here we develop a general coevolutionary model between host mate preference and the virulence of a sexually transmitted parasite. We show when the characteristics of both the host and parasite lead to coevolutionarily stable strategies or runaway selection, and when coevolutionary cycling between high and low levels of host mate choosiness and virulence is possible. A prominent argument against parasites being involved in sexual selection is that they should evolve to become less virulent when transmission depends on host mating success. The present study, however, demonstrates that coevolution can maintain stable host mate choosiness and parasite virulence or indeed coevolutionary cycling of both traits. We predict that choosiness should vary inversely with parasite virulence and that both relatively long and short life spans select against choosy behavior in the host. The model also reveals that hosts can evolve different behavioral responses from the same initial conditions, which highlights difficulties in using comparative analysis to detect parasite-mediated sexual selection. Taken as a whole, our results emphasize the importance of viewing parasite-mediated sexual selection in the context of coevolution.

Clonal expansion during Staphylococcus aureus infection dynamics reveals the effect of antibiotic intervention.

To slow the inexorable rise of antibiotic resistance we must understand how drugs impact on pathogenesis and influence the selection of resistant clones. Staphylococcus aureus is an important human pathogen with populations of antibiotic-resistant bacteria in hospitals and the community. Host phagocytes play a crucial role in controlling S. aureus infection, which can lead to a population "bottleneck" whereby clonal expansion of a small fraction of the initial inoculum founds a systemic infection. Such population dynamics may have important consequences on the effect of antibiotic intervention. Low doses of antibiotics have been shown to affect in vitro growth and the generation of resistant mutants over the long term, however whether this has any in vivo relevance is unknown. In this work, the population dynamics of S. aureus pathogenesis were studied in vivo using antibiotic-resistant strains constructed in an isogenic background, coupled with systemic models of infection in both the mouse and zebrafish embryo. Murine experiments revealed unexpected and complex bacterial population kinetics arising from clonal expansion during infection in particular organs. We subsequently elucidated the effect of antibiotic intervention within the host using mixed inocula of resistant and sensitive bacteria. Sub-curative tetracycline doses support the preferential expansion of resistant microorganisms, importantly unrelated to effects on growth rate or de novo resistance acquisition. This novel phenomenon is generic, occurring with methicillin-resistant S. aureus (MRSA) in the presence of ß-lactams and with the unrelated human pathogen Pseudomonas aeruginosa. The selection of resistant clones at low antibiotic levels can result in a rapid increase in their prevalence under conditions that would previously not be thought to favor them. Our results have key implications for the design of effective treatment regimes to limit the spread of antimicrobial resistance, where inappropriate usage leading to resistance may reduce the efficacy of life-saving drugs.

How Important is Vertical Transmission of Dengue Viruses by Mosquitoes (Diptera: Culicidae)?

Vertical transmission of dengue viruses by mosquitoes was discovered at the end of the late 1970s and has been suggested to be a means by which these viruses persist. However, it is unclear how widespread it is in nature, and its importance in the epidemiology of this disease is still debated. Here, we review the literature on vertical transmission and discuss its role in dengue's epidemiology and control. We conclude that given the number of studies that failed to find evidence of vertical transmission, as well as mathematical models and its mechanistic basis, it is unlikely that vertical transmission is important for the epidemiological persistence of dengue viruses. A combination of asymptomatic infection in humans and movement of people are likely to be more important determinants of dengue's persistence. We argue, however, that there may be some need for further research into the prevalence of dengue viruses in desiccated, as well as diapausing, eggs and the role of horizontal transmission through larval cannibalism.

Disease epidemiology in arthropods is altered by the presence of nonprotective symbionts.

Inherited microbial symbionts can modulate host susceptibility to natural enemy attack. A wider range of symbionts influence host population demography without altering individual susceptibility, and it has been suggested that these may modify host disease risk through altering the rate of exposure to natural enemies. We present the first test of this thesis, specifically testing whether male-killing symbionts alter the epidemiology of a sexually transmitted infection (STI) carried by its host. STIs are typically expected to show female-biased epidemics, and we first present a simple model which indicates that male-biased STI epidemics may occur where symbionts create female-biased population sex ratios. We then examined the dynamics of a STI in the ladybird beetle Adalia bipunctata, which is also host to a male-killing bacterium. We present evidence that male-biased epidemics of the STI are observed in natural populations when the male-killer is common. Laboratory experiments did not support a role for differential susceptibility of male and female hosts to the STI, nor a protective role for the symbiont, in creating this bias. We conclude that the range of symbionts likely to alter parasite epidemiology will be much wider than previously envisaged, because it will additionally include those that impact host demography alone.

A privileged intraphagocyte niche is responsible for disseminated infection of Staphylococcus aureus in a zebrafish model.

The innate immune system is the primary defence against the versatile pathogen, Staphylococcus aureus. How this organism is able to avoid immune killing and cause infections is poorly understood. Using an established larval zebrafish infection model, we have shown that overwhelming infection is due to subversion of phagocytes by staphylococci, allowing bacteria to evade killing and found foci of disease. Larval zebrafish coinfected with two S. aureus strains carrying different fluorescent reporter gene fusions (but otherwise isogenic) had bacterial lesions, at the time of host death, containing predominantly one strain. Quantitative data using two marked strains revealed that the strain ratios, during overwhelming infection, were often skewed towards the extremes, with one strain predominating. Infection with passaged bacterial clones revealed the phenomenon not to bedue to adventitious mutations acquired by the pathogen. After infection of the host, all bacteria are internalized by phagocytes and the skewing of population ratios is absolutely dependent on the presence of phagocytes. Mathematical modelling of pathogen population dynamics revealed the data patterns are consistent with the hypothesis that a small number of infected phagocytes serve as an intracellular reservoir for S. aureus, which upon release leads to disseminated infection. Strategies to specifically alter neutrophil/macrophage numbers were used to map the potential subpopulation of phagocytes acting as a pathogen reservoir, revealing neutrophils as the likely 'niche'. Subsequently in a murine sepsis model, S. aureus abscesses in kidneys were also found to be predominantly clonal, therefore likely founded by an individual cell, suggesting a potential mechanism analogous to the zebrafish model with few protected niches. These findings add credence to the argument that S. aureus control regimes should recognize both the intracellular as well as extracellular facets of the S. aureus life cycle.

The effects of simulated rainfall on immature population dynamics of Aedes albopictus and female oviposition.

Larvae of Aedes albopictus Skuse typically inhabit natural and artificial containers. Since these larval habitats are replenished by rainfall, Ae. albopictus may experience increased loss of immature stages in areas with high levels of rainfall. In this study, we investigated the effects of rainfall and container water level on population density, and oviposition activity of Ae. albopictus. In field and laboratory experiments, we found that rainfall resulted in the flushing of breeding habitats. Excess rain negatively impacted larval and pupal retention, especially in small habitats. When filled with water to overflowing, container habitats were significantly repellent to ovipositing females. Taken together, these data suggest that rainfall triggers population loss of Ae. albopictus and related species through a direct detrimental effect (flushing out) and an indirect effect (ovipositional repellency).

The evolution of host protection by vertically transmitted parasites.

Hosts are often infected by a variety of different parasites, leading to competition for hosts and coevolution between parasite species. There is increasing evidence that some vertically transmitted parasitic symbionts may protect their hosts from further infection and that this protection may be an important reason for their persistence in nature. Here, we examine theoretically when protection is likely to evolve and its selective effects on other parasites. Our key result is that protection is most likely to evolve in response to horizontally transmitted parasites that cause a significant reduction in host fecundity. The preponderance of sterilizing horizontally transmitted parasites found in arthropods may therefore explain the evolution of protection seen by their symbionts. We also find that protection is more likely to evolve in response to highly transmissible parasites that cause intermediate, rather than high, virulence (increased death rate when infected). Furthermore, intermediate levels of protection select for faster, more virulent horizontally transmitted parasites, suggesting that protective symbionts may lead to the evolution of more virulent parasites in nature. When we allow for coevolution between the symbiont and the parasite, more protection is likely to evolve in the vertically transmitted symbionts of longer lived hosts. Therefore, if protection is found to be common in nature, it has the potential to be a major selective force on host-parasite interactions.

Cadaver wrapping and arrival performance of adult flies in an oil palm plantation in northern Peninsular Malaysia.

There is accumulating evidence that criminals wrap dead bodies in an attempt to conceal evidence. To anticipate the forensic implications of this phenomenon, we examined whether flies that are naturally associated with cadavers exhibit a delay in attendance or differ in species composition and abundance patterns because of the presence of wrapping material. Wrapped and exposed carcasses of dead monkeys placed in an oil plantation in Kedah, Malaysia, were visited over 50 d. On daily visits to each of the six carcasses, visiting adult flies were sampled using hand nets. Flies of 12 families were encountered. Calliphoridae (Chrysomya rufifacies Macquart and C. megacephala (F.) was the most prevalent family, followed by Sphaeroceridae. Some families tended to be more abundant in WRCs (i.e., Calliphoridae, Muscidae, and Phoridae), whereas others (i.e., Piophilidae, Sepsidae, and Psychodidae) were more prevalent in exposed carcasses. Wrapping delayed the arrival of all fly species encountered, with delays varying from 1 to 13 d depending on species. Wrapping did not affect species composition of flies, but prolong the occurrence of some species. The results of the current study emphasize the need to take into consideration the presence of a wrap when estimating postmortem interval.

Discarded cigarette butts attract females and kill the progeny of Aedes albopictus.

Discarded cigarette butts (DCB) waste occurs worldwide, pollutes landscapes, is unsightly, and results in added debris removal costs. There is, therefore, a great deal of current interest in making use of DCBs in beneficial ways. Despite evidence that DCBs are harmful to water fleas (Daphnia magna), which breed in aquatic environments as do mosquito larvae, their impact on dengue vectors is unknown. We examined whether Aedes albopictus alters its ovipositional responses, larval eclosion, and development in response to presence of DCBs in its habitats. We found oviposition activity in DCB-treated water similar to that of control water and that ovipositional activity in DCB solutions steadily increased over time as those solutions aged to 10 days. Larval eclosion was initially suppressed on day 1 in DCB solution, but increased thereafter to levels similar to control larval eclosion rates. The DCB-water solutions produced significantly higher mortality in both 1st and 2nd instars over control larvae for several days after initial exposure. Mortality rates decreased sharply 3 to 5 days postexposure as DCBs continued to decompose. We found increased survival rates during late development, but daily input of fresh DCBs prevented most young larvae from completing development. Taken together, these observations suggest that decomposing did not deter gravid Ae. albopictus females from ovipositing in treated containers and that DCB solutions had larvicidal effects on early instars. Our results are discussed in the context of DCB use to control container-breeding Ae. albopictus, a competent dengue vector in Asia and other parts of the world.

The effects of moisture on ovipositional responses and larval eclosion of Aedes albopictus.

Moisture plays a major role in the dynamics of mosquito populations, especially those breeding in container habitats. Despite this importance, the role of moisture conditions as they affect oviposition and egg development in Aedes vectors remains largely unexplored. We investigated the effect of exposing gravid female Aedes albopictus mosquitoes and their eggs to different moisture levels (MLs) for various periods on oviposition and hatching. Overall, high-moisture substrates (HMSs; 66% and 72%) provided better environments for egg laying. The timing of initial egg laying was far longer at the lowest substrate moisture level (LSML, 25% and 41.2%) than at HMSs. The numbers of eggs laid were much lower in the drier environments. At LSMLs, gravid females retained increasing numbers of mature eggs until death, and egg retention decreased gradually with increasing ML. The HMSs also provided better environments for larval eclosion. The numbers of eggs hatched were lower at the LSML than the HSML environment. No egg hatching occurred after 1 h exposure to moisture. However, egg hatching occurred by installment, with spontaneous hatching (SH) increasing gradually with increasing ML. High-moisture conditions combined with long exposure (30 h and 48 h) favored SH. These results suggest that Ae. albopictus females can respond to better moisture conditions for increased success of embryonation and larval eclosion. This information may be useful in the colonization of floodwater Aedes species.

The Role of Vector Trait Variation in Vector-Borne Disease Dynamics.

Many important endemic and emerging diseases are transmitted by vectors that are biting arthropods. The functional traits of vectors can affect pathogen transmission rates directly and also through their effect on vector population dynamics. Increasing empirical evidence shows that vector traits vary significantly across individuals, populations, and environmental conditions, and at time scales relevant to disease transmission dynamics. Here, we review empirical evidence for variation in vector traits and how this trait variation is currently incorporated into mathematical models of vector-borne disease transmission. We argue that mechanistically incorporating trait variation into these models, by explicitly capturing its effects on vector fitness and abundance, can improve the reliability of their predictions in a changing world. We provide a conceptual framework for incorporating trait variation into vector-borne disease transmission models, and highlight key empirical and theoretical challenges. This framework provides a means to conceptualize how traits can be incorporated in vector borne disease systems, and identifies key areas in which trait variation can be explored. Determining when and to what extent it is important to incorporate trait variation into vector borne disease models remains an important, outstanding question.

Synergistic China-US Ecological Research is Essential for Global Emerging Infectious Disease Preparedness.

The risk of a zoonotic pandemic disease threatens hundreds of millions of people. Emerging infectious diseases also threaten livestock and wildlife populations around the world and can lead to devastating economic damages. China and the USA-due to their unparalleled resources, widespread engagement in activities driving emerging infectious diseases and national as well as geopolitical imperatives to contribute to global health security-play an essential role in our understanding of pandemic threats. Critical to efforts to mitigate risk is building upon existing investments in global capacity to develop training and research focused on the ecological factors driving infectious disease spillover from animals to humans. International cooperation, particularly between China and the USA, is essential to fully engage the resources and scientific strengths necessary to add this ecological emphasis to the pandemic preparedness strategy. Here, we review the world's current state of emerging infectious disease preparedness, the ecological and evolutionary knowledge needed to anticipate disease emergence, the roles that China and the USA currently play as sources and solutions to mitigating risk, and the next steps needed to better protect the global community from zoonotic disease.

Industrial bees: The impact of apicultural intensification on local disease prevalence.

It is generally thought that the intensification of farming will result in higher disease prevalences, although there is little specific modelling testing this idea. Focussing on honeybees, we build multi-colony models to inform how "apicultural intensification" is predicted to impact honeybee pathogen epidemiology at the apiary scale.We used both agent-based and analytical models to show that three linked aspects of apicultural intensification (increased population sizes, changes in population network structure and increased between-colony transmission) are unlikely to greatly increase disease prevalence in apiaries. Principally this is because even low-intensity apiculture exhibits high disease prevalence.The greatest impacts of apicultural intensification are found for diseases with relatively low R0 (basic reproduction number), however, such diseases cause little overall disease prevalence and, therefore, the impacts of intensification are minor. Furthermore, the smallest impacts of intensification are for diseases with high R0 values, which we argue are typical of important honeybee diseases. Policy Implications: Our findings contradict the idea that apicultural intensification by crowding honeybee colonies in large, dense apiaries leads to notably higher disease prevalences for established honeybee pathogens. More broadly, our work demonstrates the need for informative models of all agricultural systems and management practices in order to understand the implications of management changes on diseases.

The influence of trade-off shape on evolutionary behaviour in classical ecological scenarios.

Trade-off shapes are crucial to evolutionary outcomes. However, due to different ecological feedbacks their implications may depend not only on the trade-off being considered but also the ecological scenario. Here, we apply a novel geometric technique, trade-off and invasion plots (TIPs), to examine in detail how the shape of trade-off relationships affect evolutionary outcomes under a range of classic ecological scenarios including Lotka-Volterra type and host-parasite interactions. We choose models of increasing complexity in order to gain an insight into the features of ecological systems that determine the evolutionary outcomes. In particular we focus on when evolutionary attractors, repellors and branching points occur and how this depends on whether the costs are accelerating (benefits become 'increasingly' costly), decelerating (benefits become 'decreasingly' costly) or constant. In all cases strongly accelerating costs lead to attractors while strongly decelerating ones lead to repellors, but with weaker relationships, this no longer holds. For some systems weakly accelerating costs may lead to repellors and decelerating costs may lead to attractors. In many scenarios it is weakly decelerating costs that lead to branching points, but weakly accelerating and linear costs may also lead to disruptive selection in particular ecological scenarios. Using our models we suggest a classification of ecological interactions, based on three distinct criteria, that can produce one of four fundamental TIPs which allow for different evolutionary behaviour. This provides a baseline theory which may inform the prediction of evolutionary outcomes in similar yet unexplored ecological scenarios. In addition we discuss the implications of our results to a number of specific life-history trade-offs in the classic ecological scenarios represented by our models.

A genotypic trade-off between constitutive resistance to viral infection and host growth rate.

Genotypic trade-offs are fundamental to the understanding of the evolution of life-history traits. In particular, the evolution of optimal host defense and the maintenance of variation in defense against infectious disease is thought to be underpinned by such evolutionary trade-offs. However, empirical demonstrations of these trade-offs that satisfy the strict assumptions made by theoretical models are rare. Additionally, none of these trade-offs have yet been shown to be robustly replicable using a variety of different experimental approaches to rule out confounding issues with particular experimental designs. Here, we use inbred isolines as a novel experimental approach to test whether a trade-off between viral resistance and growth rate in Plodia interpunctella, previously demonstrated by multiple selection experiments, is robust and meets the strict criteria required to underpin theoretical work in this field. Critically, we demonstrate that this trade-off is both genetic and constitutive. This finding helps support the large body of theory that relies on these assumptions, and makes this trade-off for resistance unique in being replicated through multiple experimental approaches and definitively shown to be genetic and constitutive.