Inferring infection hazard in wildlife populations by linking data across individual and population scales.

Our ability to infer unobservable disease-dynamic processes such as force of infection (infection hazard for susceptible hosts) has transformed our understanding of disease transmission mechanisms and capacity to predict disease dynamics. Conventional methods for inferring FOI estimate a time-averaged value and are based on population-level processes. Because many pathogens exhibit epidemic cycling and FOI is the result of processes acting across the scales of individuals and populations, a flexible framework that extends to epidemic dynamics and links within-host processes to FOI is needed. Specifically, within-host antibody kinetics in wildlife hosts can be short-lived and produce patterns that are repeatable across individuals, suggesting individual-level antibody concentrations could be used to infer time since infection and hence FOI. Using simulations and case studies (influenza A in lesser snow geese and Yersinia pestis in coyotes), we argue that with careful experimental and surveillance design, the population-level FOI signal can be recovered from individual-level antibody kinetics, despite substantial individual-level variation. In addition to improving inference, the cross-scale quantitative antibody approach we describe can reveal insights into drivers of individual-based variation in disease response, and the role of poorly understood processes such as secondary infections, in population-level dynamics of disease.

Temporal patterns of chronic wasting disease prion excretion in three cervid species.

Chronic wasting disease (CWD) is the only naturally occurring transmissible spongiform encephalopathy affecting free-ranging wildlife populations. Transmission of CWD occurs by direct contact or through contaminated environments; however, little is known about the temporal patterns of CWD prion excretion and shedding in wild cervids. We tested the urine and faeces of three species of captive cervids (elk, mule and white-tailed deer) at 6, 12, 18 and 24 months after oral inoculation to evaluate the temporal, species- and genotype-specific factors affecting the excretion of CWD prions. Although none of the animals exhibited clinical signs of CWD during the study, we determined that all three cervid species were excreting CWD prions by 6 months post-inoculation. Faecal samples were consistently positive for CWD prions for all three cervid species (88 %), and were more likely to be positive than urine samples (28 %). Cervids with genotypes encoding for the prion protein (PRNP) that were considered to be more susceptible to CWD were more likely to excrete CWD prions (94 %) than cervids with genotypes considered to be less susceptible (64 %). All cervids with CWD prions in their urine also had positive faeces (n=5), but the converse was not true. Our study is the first to demonstrate CWD prion excretion in urine by asymptomatic elk and mule deer. Our results indicate that the excretion of CWD prions in faeces and, to a lesser extent, urine may provide an important avenue for depositing prions in the environment.

Chronic wasting disease in white-tailed deer: infection, mortality, and implications for heterogeneous transmission.

Chronic wasting disease (CWD) is a fatal neurodegenerative disease affecting free-ranging and captive cervids that now occurs in 24 U.S. states and two Canadian provinces. Despite the potential threat of CWD to deer populations, little is known about the rates of infection and mortality caused by this disease. We used epidemiological models to estimate the force of infection and disease-associated mortality for white-tailed deer in the Wisconsin and Illinois CWD outbreaks. Models were based on age-prevalence data corrected for bias in aging deer using the tooth wear and replacement method. Both male and female deer in the Illinois outbreak had higher corrected age-specific prevalence with slightly higher female infection than deer in the Wisconsin outbreak. Corrected ages produced more complex models with different infection and mortality parameters than those based on apparent prevalence. We found that adult male deer have a more than threefold higher risk of CWD infection than female deer. Males also had higher disease mortality than female deer. As a result, CWD prevalence was twofold higher in adult males than females. We also evaluated the potential impacts of alternative contact structures on transmission dynamics in Wisconsin deer. Results suggested that transmission of CWD among male deer during the nonbreeding season may be a potential mechanism for producing higher rates of infection and prevalence characteristically found in males. However, alternatives based on high environmental transmission and transmission from females to males during the breeding season may also play a role.

Will a warmer and wetter future cause extinction of native Hawaiian forest birds?

Isolation of the Hawaiian archipelago produced a highly endemic and unique avifauna. Avian malaria (Plasmodium relictum), an introduced mosquito-borne pathogen, is a primary cause of extinctions and declines of these endemic honeycreepers. Our research assesses how global climate change will affect future malaria risk and native bird populations. We used an epidemiological model to evaluate future bird-mosquito-malaria dynamics in response to alternative climate projections from the Coupled Model Intercomparison Project. Climate changes during the second half of the century accelerate malaria transmission and cause a dramatic decline in bird abundance. Different temperature and precipitation patterns produce divergent trajectories where native birds persist with low malaria infection under a warmer and dryer projection (RCP4.5), but suffer high malaria infection and severe reductions under hot and dry (RCP8.5) or warm and wet (A1B) futures. We conclude that future global climate change will cause significant decreases in the abundance and diversity of remaining Hawaiian bird communities. Because these effects appear unlikely before mid-century, natural resource managers have time to implement conservation strategies to protect this unique avifauna from further decimation. Similar climatic drivers for avian and human malaria suggest that mitigation strategies for Hawai'i have broad application to human health.

The dynamics of avian influenza in Lesser Snow Geese: implications for annual and migratory infection patterns.

Wild water birds are the natural reservoir for low-pathogenic avian influenza viruses (AIV). However, our ability to investigate the epizootiology of AIV in these migratory populations is challenging and, despite intensive worldwide surveillance, remains poorly understood. We conducted a cross-sectional, retrospective analysis in Pacific Flyway Lesser Snow Geese, Chen caerulescens, to investigate AIV serology and infection patterns. We collected nearly 3000 sera samples from Snow Geese at two breeding colonies in Russia and Canada during 1993-1996 and swab samples from >4000 birds at wintering and migration areas in the United States during 2006-2011. We found seroprevalence and annual seroconversion varied considerably among years. Seroconversion and infection rates also differed between Snow Goose breeding colonies and wintering areas, suggesting that AIV exposure in this gregarious waterfowl species is likely occurring during several phases (migration, wintering, and potentially breeding areas) of the annual cycle. We estimated AIV antibody persistence was longer (14 months) in female geese compared to males (6 months). This relatively long period of AIV antibody persistence suggests that subtype-specific serology may be an effective tool for detection of exposure to subtypes associated with highly pathogenic AIV. Our study provides further evidence of high seroprevalence in Arctic goose populations, and estimates of annual AIV seroconversion and antibody persistence for North American waterfowl. We suggest future AIV studies include serology to help elucidate the epizootiological dynamics of AIV in wild bird populations.

The walk is never random: subtle landscape effects shape gene flow in a continuous white-tailed deer population in the Midwestern United States.

One of the pervasive challenges in landscape genetics is detecting gene flow patterns within continuous populations of highly mobile wildlife. Understanding population genetic structure within a continuous population can give insights into social structure, movement across the landscape and contact between populations, which influence ecological interactions, reproductive dynamics or pathogen transmission. We investigated the genetic structure of a large population of deer spanning the area of Wisconsin and Illinois, USA, affected by chronic wasting disease. We combined multiscale investigation, landscape genetic techniques and spatial statistical modelling to address the complex questions of landscape factors influencing population structure. We sampled over 2000 deer and used spatial autocorrelation and a spatial principal components analysis to describe the population genetic structure. We evaluated landscape effects on this pattern using a spatial autoregressive model within a model selection framework to test alternative hypotheses about gene flow. We found high levels of genetic connectivity, with gradients of variation across the large continuous population of white-tailed deer. At the fine scale, spatial clustering of related animals was correlated with the amount and arrangement of forested habitat. At the broader scale, impediments to dispersal were important to shaping genetic connectivity within the population. We found significant barrier effects of individual state and interstate highways and rivers. Our results offer an important understanding of deer biology and movement that will help inform the management of this species in an area where overabundance and disease spread are primary concerns.

Emerging prion disease drives host selection in a wildlife population.

Infectious diseases are increasingly recognized as an important force driving population dynamics, conservation biology, and natural selection in wildlife populations. Infectious agents have been implicated in the decline of small or endangered populations and may act to constrain population size, distribution, growth rates, or migration patterns. Further, diseases may provide selective pressures that shape the genetic diversity of populations or species. Thus, understanding disease dynamics and selective pressures from pathogens is crucial to understanding population processes, managing wildlife diseases, and conserving biological diversity. There is ample evidence that variation in the prion protein gene (PRNP) impacts host susceptibility to prion diseases. Still, little is known about how genetic differences might influence natural selection within wildlife populations. Here we link genetic variation with differential susceptibility of white-tailed deer to chronic wasting disease (CWD), with implications for fitness and disease-driven genetic selection. We developed a single nucleotide polymorphism (SNP) assay to efficiently genotype deer at the locus of interest (in the 96th codon of the PRNP gene). Then, using a Bayesian modeling approach, we found that the more susceptible genotype had over four times greater risk of CWD infection; and, once infected, deer with the resistant genotype survived 49% longer (8.25 more months). We used these epidemiological parameters in a multi-stage population matrix model to evaluate relative fitness based on genotype-specific population growth rates. The differences in disease infection and mortality rates allowed genetically resistant deer to achieve higher population growth and obtain a long-term fitness advantage, which translated into a selection coefficient of over 1% favoring the CWD-resistant genotype. This selective pressure suggests that the resistant allele could become dominant in the population within an evolutionarily short time frame. Our work provides a rare example of a quantifiable disease-driven selection process in a wildlife population, demonstrating the potential for infectious diseases to alter host populations. This will have direct bearing on the epidemiology, dynamics, and future trends in CWD transmission and spread. Understanding genotype-specific epidemiology will improve predictive models and inform management strategies for CWD-affected cervid populations.

Potential Effects of Environmental Conditions on Prairie Dog Flea Development and Implications for Sylvatic Plague Epizootics.

Fleas are common ectoparasites of vertebrates worldwide and vectors of many pathogens causing disease, such as sylvatic plague in prairie dog colonies. Development of fleas is regulated by environmental conditions, especially temperature and relative humidity. Development rates are typically slower at low temperatures and faster at high temperatures, which are bounded by lower and upper thresholds where development is reduced. Prairie dogs and their associated fleas (mostly Oropsylla spp) live in burrows that moderate outside environmental conditions, remaining cooler in summer and warmer in winter. We found burrow microclimates were characterized by stable daily temperatures and high relative humidity, with temperatures increasing from spring through summer. We previously showed temperature increases corresponded with increasing off-host flea abundance. To evaluate how changes in temperature could affect future prairie dog flea development and abundance, we used development rates of O. montana (a species related to prairie dog fleas), determined how prairie dog burrow microclimates are affected by ambient weather, and combined these results to develop a predictive model. Our model predicts burrow temperatures and flea development rates will increase during the twenty-first century, potentially leading to higher flea abundance and an increased probability of plague epizootics if Y. pestis is present.

In vitro detection and quantification of botulinum neurotoxin type e activity in avian blood.

Botulinum neurotoxin serotype E (BoNT/E) outbreaks in the Great Lakes region cause large annual avian mortality events, with an estimated 17,000 bird deaths reported in 2007 alone. During an outbreak investigation, blood collected from bird carcasses is tested for the presence of BoNT/E using the mouse lethality assay. While sensitive, this method is labor-intensive and low throughput and can take up to 7 days to complete. We developed a rapid and sensitive in vitro assay, the BoTest Matrix E assay, that combines immunoprecipitation with high-affinity endopeptidase activity detection by Förster resonance energy transfer (FRET) to rapidly quantify BoNT/E activity in avian blood with detection limits comparable to those of the mouse lethality assay. On the basis of the analysis of archived blood samples (n = 87) collected from bird carcasses during avian mortality investigations, BoTest Matrix E detected picomolar quantities of BoNT/E following a 2-h incubation and femtomolar quantities of BoNT/E following extended incubation (24 h) with 100% diagnostic specificity and 91% diagnostic sensitivity.

Diversity and distribution of white-tailed deer mtDNA lineages in chronic wasting disease (CWD) outbreak areas in southern Wisconsin, USA.

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy affecting North American cervids. Because it is uniformly fatal, the disease is a major concern in the management of white-tailed deer populations. Management programs to control CWD require improved knowledge of deer interaction, movement, and population connectivity that could influence disease transmission and spread. Genetic methods were employed to evaluate connectivity among populations in the CWD management zone of southern Wisconsin. A 576-base-pair region of the mitochondrial DNA of 359 white-tailed deer from 12 sample populations was analyzed. Fifty-eight variable sites were detected within the sequence, defining 43 haplotypes. While most sample populations displayed similar levels of haplotype diversity, individual haplotypes were clustered on the landscape. Spatial clusters of different haplotypes were apparent in distinct ecoregions surrounding CWD outbreak areas. The spatial distribution of mtDNA haplotypes suggests that clustering of the deer matrilineal groups and population connectivity are associated with broad-scale geographic landscape features. These landscape characteristics may also influence the contact rates between groups and therefore the potential spread of CWD; this may be especially true of local disease spread between female social groups. Our results suggest that optimal CWD management needs to be tailored to fit gender-specific dispersal behaviors and regional differences in deer population connectivity. This information will help wildlife managers design surveillance and monitoring efforts based on population interactions and potential deer movement among CWD-affected and unaffected areas.

Impacts of environmental conditions on fleas in black-tailed prairie dog burrows.

Sylvatic plague, caused by the bacterium Yersinia pestis and transmitted by fleas, occurs in prairie dogs of the western United States. Outbreaks can devastate prairie dog communities, often causing nearly 100% mortality. Three competent flea vectors, prairie dog specialists Oropsylla hirsuta and O. tuberculata, and generalist Pulex simulans, are found on prairie dogs and in their burrows. Fleas are affected by climate, which varies across the range of black-tailed prairie dogs (Cynomys ludovicianus), but these effects may be ameliorated somewhat due to the burrowing habits of prairie dogs. Our goal was to assess how temperature and precipitation affect off-host flea abundance and whether relative flea abundance varied across the range of black-tailed prairie dogs. Flea abundance was measured by swabbing 300 prairie dog burrows at six widely distributed sites in early and late summer of 2016 and 2017. Relative abundance of flea species varied among sites and sampling sessions. Flea abundance and prevalence increased with monthly mean high temperature and declined with higher winter precipitation. Predicted climate change in North America will likely influence flea abundance and distribution, thereby impacting plague dynamics in prairie dog colonies.

Spatial and temporal patterns of chronic wasting disease: fine-scale mapping of a wildlife epidemic in Wisconsin.

Emerging infectious diseases threaten wildlife populations and human health. Understanding the spatial distributions of these new diseases is important for disease management and policy makers; however, the data are complicated by heterogeneities across host classes, sampling variance, sampling biases, and the space-time epidemic process. Ignoring these issues can lead to false conclusions or obscure important patterns in the data, such as spatial variation in disease prevalence. Here, we applied hierarchical Bayesian disease mapping methods to account for risk factors and to estimate spatial and temporal patterns of infection by chronic wasting disease (CWD) in white-tailed deer (Odocoileus virginianus) of Wisconsin, U.S.A. We found significant heterogeneities for infection due to age, sex, and spatial location. Infection probability increased with age for all young deer, increased with age faster for young males, and then declined for some older animals, as expected from disease-associated mortality and age-related changes in infection risk. We found that disease prevalence was clustered in a central location, as expected under a simple spatial epidemic process where disease prevalence should increase with time and expand spatially. However, we could not detect any consistent temporal or spatiotemporal trends in CWD prevalence. Estimates of the temporal trend indicated that prevalence may have decreased or increased with nearly equal posterior probability, and the model without temporal or spatiotemporal effects was nearly equivalent to models with these effects based on deviance information criteria. For maximum interpretability of the role of location as a disease risk factor, we used the technique of direct standardization for prevalence mapping, which we develop and describe. These mapping results allow disease management actions to be employed with reference to the estimated spatial distribution of the disease and to those host classes most at risk. Future wildlife epidemiology studies should employ hierarchical Bayesian methods to smooth estimated quantities across space and time, account for heterogeneities, and then report disease rates based on an appropriate standardization.

Surveillance for transmissible spongiform encephalopathy in scavengers of white-tailed deer carcasses in the chronic wasting disease area of Wisconsin.

Chronic wasting disease (CWD), a class of neurodegenerative transmissible spongiform encephalopathies (TSE) occurring in cervids, is found in a number of states and provinces across North America. Misfolded prions, the infectious agents of CWD, are deposited in the environment via carcass remains and excreta, and pose a threat of cross-species transmission. In this study tissues were tested from 812 representative mammalian scavengers, collected in the CWD-affected area of Wisconsin, for TSE infection using the IDEXX HerdChek enzyme-linked immunosorbent assay (ELISA). Only four of the collected mammals tested positive using the ELISA, but these were negative when tested by Western blot. While our sample sizes permitted high probabilities of detecting TSE assuming 1% population prevalence in several common scavengers (93%, 87%, and 87% for raccoons, opossums, and coyotes, respectively), insufficient sample sizes for other species precluded similar conclusions. One cannot rule out successful cross-species TSE transmission to scavengers, but the results suggest that such transmission is not frequent in the CWD-affected area of Wisconsin. The need for further surveillance of scavenger species, especially those known to be susceptible to TSE (e.g., cat, American mink, raccoon), is highlighted in both a field and laboratory setting.

Estimating cause-specific mortality rates using recovered carcasses.

Stranding networks, in which carcasses are recovered and sent to diagnostic laboratories for necropsy and determination of cause of death, have been developed to monitor the health of marine mammal and bird populations. These programs typically accumulate comprehensive, long-term datasets on causes of death that can be used to identify important sources of mortality or changes in mortality patterns that lead to management actions. However, the utility of these data in determining cause-specific mortality rates has not been explored. We present a maximum likelihood-based approach that partitions total mortality rate, estimated by independent sources, into cause-specific mortality rates. We also demonstrate how variance estimates are derived for these rates. We present examples of the method using mortality data for California sea otters (Enhydra lutris nereis) and Florida manatees (Trichechus manatus latirostris).

Chronic wasting disease in a Wisconsin white-tailed deer farm.

In September 2002, chronic wasting disease (CWD), a prion disorder of captive and wild cervids, was diagnosed in a white-tailed deer (Odocoileus virginianus) from a captive farm in Wisconsin. The facility was subsequently quarantined, and in January 2006 the remaining 76 deer were depopulated. Sixty animals (79%) were found to be positive by immunohistochemical staining for the abnormal prion protein (PrP(CWD)) in at least one tissue; the prevalence of positive staining was high even in young deer. Although none of the deer displayed clinical signs suggestive of CWD at depopulation, 49 deer had considerable accumulation of the abnormal prion in the medulla at the level of the obex. Extraneural accumulation of the abnormal protein was observed in 59 deer, with accumulation in the retropharyngeal lymph node in 58 of 59 (98%), in the tonsil in 56 of 59 (95%), and in the rectal mucosal lymphoid tissue in 48 of 58 (83%). The retina was positive in 4 deer, all with marked accumulation of prion in the obex. One deer was considered positive for PrP(CWD) in the brain but not in the extraneural tissue, a novel observation in white-tailed deer. The infection rate in captive deer was 20-fold higher than in wild deer. Although weakly related to infection rates in extraneural tissues, prion genotype was strongly linked to progression of prion accumulation in the obex. Antemortem testing by biopsy of recto-anal mucosal-associated lymphoid tissue (or other peripheral lymphoid tissue) may be a useful adjunct to tonsil biopsy for surveillance in captive herds at risk for CWD infection.

Using amplified fragment length polymorphism analysis to differentiate isolates of Pasteurella multocida serotype 1.

Avian cholera, an infectious disease caused by the bacterium Pasteurella multocida, kills thousands of North American wild waterfowl annually. Pasteurella multocida serotype 1 isolates cultured during a laboratory challenge study of Mallards (Anas platyrhynchos) and collected from wild birds and environmental samples during avian cholera outbreaks were characterized using amplified fragment length polymorphism (AFLP) analysis, a whole-genome DNA fingerprinting technique. Comparison of the AFLP profiles of 53 isolates from the laboratory challenge demonstrated that P. multocida underwent genetic changes during a 3-mo period. Analysis of 120 P. multocida serotype 1 isolates collected from wild birds and environmental samples revealed that isolates were distinguishable from one another based on regional and temporal genetic characteristics. Thus, AFLP analysis had the ability to distinguish P. multocida isolates of the same serotype by detecting spatiotemporal genetic changes and provides a tool to advance the study of avian cholera epidemiology. Further application of AFLP technology to the examination of wild bird avian cholera outbreaks may facilitate more effective management of this disease by providing the potential to investigate correlations between virulence and P. multocida genotypes, to identify affiliations between bird species and bacterial genotypes, and to elucidate the role of specific bird species in disease transmission.

Mineral licks as environmental reservoirs of chronic wasting disease prions.

Chronic wasting disease (CWD) is a fatal neurodegenerative disease of deer, elk, moose, and reindeer (cervids) caused by misfolded prion proteins. The disease has been reported across North America and recently discovered in northern Europe. Transmission of CWD in wild cervid populations can occur through environmental routes, but limited ability to detect prions in environmental samples has prevented the identification of potential transmission "hot spots". We establish widespread CWD prion contamination of mineral licks used by free-ranging cervids in an enzootic area in Wisconsin, USA. We show mineral licks can serve as reservoirs of CWD prions and thus facilitate disease transmission. Furthermore, mineral licks attract livestock and other wildlife that also obtain mineral nutrients via soil and water consumption. Exposure to CWD prions at mineral licks provides potential for cross-species transmission to wildlife, domestic animals, and humans. Managing deer use of mineral licks warrants further consideration to help control outbreaks of CWD.

Impact of Sylvatic Plague Vaccine on Non-target Small Rodents in Grassland Ecosystems.

Oral vaccination is an emerging management strategy to reduce the prevalence of high impact infectious diseases within wild animal populations. Plague is a flea-borne zoonosis of rodents that often decimates prairie dog (Cynomys spp.) colonies in the western USA. Recently, an oral sylvatic plague vaccine (SPV) was developed to protect prairie dogs from plague and aid recovery of the endangered black-footed ferret (Mustela nigripes). Although oral vaccination programs are targeted toward specific species, field distribution of vaccine-laden baits can result in vaccine uptake by non-target animals and unintended indirect effects. We assessed the impact of SPV on non-target rodents at paired vaccine and placebo-treated prairie dog colonies in four US states from 2013 to 2015. Bait consumption by non-target rodents was high (70.8%, n = 3113), but anti-plague antibody development on vaccine plots was low (23.7%, n = 266). In addition, no significant differences were noted in combined deer mice (Peromyscus maniculatus) and western harvest mouse (Reithrodontomys megalotis) abundance or community evenness and richness of non-target rodents between vaccine-treated and placebo plots. In our 3-year field study, we could not detect a significant positive or negative effect of SPV application on non-target rodents.

Mycobacterium avium subsp. paratuberculosis in scavenging mammals in Wisconsin.

The presence of Mycobacterium avium subsp. paratuberculosis (MAP) in non-ruminant wildlife has raised questions regarding the role of these species in Johne's disease transmission. In this study we tested 472 tissues from 212 animals of six different species of scavenging mammals. All animals were taken from within a 210-square-mile area in Dane and Iowa counties of south central Wisconsin from September to May in 2003-04 and tested for the presence of MAP. We detected MAP-specific DNA in 81 of 212 (38%) scavenging mammals, in 98 of the 472 (21%) tissues; viable MAP was cultured from one coyote's ileum and lymph node tissue. Despite the low numbers of viable MAP isolated in this study, our data adds to the increasing evidence demonstrating the potential for transmission and infection of MAP in nonruminant species and provides possible evidence of interspecies transmission. The apparently high exposure of nonruminant wildlife provides potential evidence of a spill-over of MAP to wildlife species and raises the question of spillback to domestic and wild ruminants. These results demonstrate the importance of understanding the role of wildlife species in developing management strategies for Johne's disease in domestic livestock.

Mitigating Future Avian Malaria Threats to Hawaiian Forest Birds from Climate Change.

Avian malaria, transmitted by Culex quinquefasciatus mosquitoes in the Hawaiian Islands, has been a primary contributor to population range limitations, declines, and extinctions for many endemic Hawaiian honeycreepers. Avian malaria is strongly influenced by climate; therefore, predicted future changes are expected to expand transmission into higher elevations and intensify and lengthen existing transmission periods at lower elevations, leading to further population declines and potential extinction of highly susceptible honeycreepers in mid- and high-elevation forests. Based on future climate changes and resulting malaria risk, we evaluated the viability of alternative conservation strategies to preserve endemic Hawaiian birds at mid and high elevations through the 21st century. We linked an epidemiological model with three alternative climatic projections from the Coupled Model Intercomparison Project to predict future malaria risk and bird population dynamics for the coming century. Based on climate change predictions, proposed strategies included mosquito population suppression using modified males, release of genetically modified refractory mosquitoes, competition from other introduced mosquitoes that are not competent vectors, evolved malaria-tolerance in native honeycreepers, feral pig control to reduce mosquito larval habitats, and predator control to improve bird demographics. Transmission rates of malaria are predicted to be higher than currently observed and are likely to have larger impacts in high-elevation forests where current low rates of transmission create a refuge for highly-susceptible birds. As a result, several current and proposed conservation strategies will be insufficient to maintain existing forest bird populations. We concluded that mitigating malaria transmission at high elevations should be a primary conservation goal. Conservation strategies that maintain highly susceptible species like Iiwi (Drepanis coccinea) will likely benefit other threatened and endangered Hawai'i species, especially in high-elevation forests. Our results showed that mosquito control strategies offer potential long-term benefits to high elevation Hawaiian honeycreepers. However, combined strategies will likely be needed to preserve endemic birds at mid elevations. Given the delay required to research, develop, evaluate, and improve several of these currently untested conservation strategies we suggest that planning should begin expeditiously.