Environmental and geographical factors influence the occurrence and abundance of the southern house mosquito, Culex quinquefasciatus, in Hawai'i.

Hawaiian honeycreepers, a group of endemic Hawaiian forest birds, are being threatened by avian malaria, a non-native disease that is driving honeycreepers populations to extinction. Avian malaria is caused by the parasite Plasmodium relictum, which is transmitted by the invasive mosquito Culex quinquefasciatus. Environmental and geographical factors play an important role in shaping mosquito-borne disease transmission dynamics through their influence on the distribution and abundance of mosquitoes. We assessed the effects of environmental (temperature, precipitation), geographic (site, elevation, distance to anthropogenic features), and trap type (CDC light trap, CDC gravid trap) factors on mosquito occurrence and abundance. Occurrence was analyzed using classification and regression tree models (CART) and generalized linear models (GLM); abundance (count data) was analyzed using generalized linear mixed models (GLMMs). Models predicted highest mosquito occurrence at mid-elevation sites and between July and November. Occurrence increased with temperature and precipitation up to 580 mm. For abundance, the best model was a zero-inflated negative-binomial model that indicated higher abundance of mosquitoes at mid-elevation sites and peak abundance between August and October. Estimation of occurrence and abundance as well as understanding the factors that influence them are key for mosquito control, which may reduce the risk of forest bird extinction.

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.

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.

Changing climate and the altitudinal range of avian malaria in the Hawaiian Islands - an ongoing conservation crisis on the island of Kaua'i.

Transmission of avian malaria in the Hawaiian Islands varies across altitudinal gradients and is greatest at elevations below 1500 m where both temperature and moisture are favorable for the sole mosquito vector, Culex quinquefasciatus, and extrinsic sporogonic development of the parasite, Plasmodium relictum. Potential consequences of global warming on this system have been recognized for over a decade with concerns that increases in mean temperatures could lead to expansion of malaria into habitats where cool temperatures currently limit transmission to highly susceptible endemic forest birds. Recent declines in two endangered species on the island of Kaua'i, the 'Akikiki (Oreomystis bairdi) and 'Akeke'e (Loxops caeruleirostris), and retreat of more common native honeycreepers to the last remaining high elevation habitat on the Alaka'i Plateau suggest that predicted changes in disease transmission may be occurring. We compared prevalence of malarial infections in forest birds that were sampled at three locations on the Plateau during 1994-1997 and again during 2007-2013, and also evaluated changes in the occurrence of mosquito larvae in available aquatic habitats during the same time periods. Prevalence of infection increased significantly at the lower (1100 m, 10.3% to 28.2%), middle (1250 m, 8.4% to 12.2%), and upper ends of the Plateau (1350 m, 2.0% to 19.3%). A concurrent increase in detections of Culex larvae in aquatic habitats associated with stream margins indicates that populations of the vector are also increasing. These increases are at least in part due to local transmission because overall prevalence in Kaua'i 'Elepaio (Chasiempis sclateri), a sedentary native species, has increased from 17.2% to 27.0%. Increasing mean air temperatures, declining precipitation, and changes in streamflow that have taken place over the past 20 years are creating environmental conditions throughout major portions of the Alaka'i Plateau that support increased transmission of avian malaria.

Modeling future conservation of Hawaiian honeycreepers by mosquito management and translocation of disease-tolerant Amakihi.

Avian malaria is an important cause of the decline of endemic Hawaiian honeycreepers. Because of the complexity of this disease system we used a computer model of avian malaria in forest birds to evaluate how two proposed conservation strategies: 1) reduction of habitat for mosquito larvae and 2) establishment of a low-elevation, malaria-tolerant honeycreeper (Hawaii Amakihi) to mid-elevation forests would affect native Hawaiian honeycreeper populations. We evaluated these approaches in mid-elevation forests, where malaria transmission is seasonal and control strategies are more likely to work. Our model suggests the potential benefit of larval habitat reduction depends on the level of malaria transmission, abundance of larval cavities, and the ability to substantially reduce these cavities. Permanent reduction in larval habitat of >80% may be needed to control abundance of infectious mosquitoes and benefit bird populations. Establishment of malaria-tolerant Amakihi in mid-elevation forests increases Amakihi abundance, creates a larger disease reservoir, and increases the abundance of infectious mosquitoes which may negatively impact other honeycreepers. For mid-elevation sites where bird populations are severely affected by avian malaria, malaria-tolerant Amakihi had little impact on other honeycreepers. Both management strategies may benefit native Hawaiian honeycreepers, but benefits depend on specific forest characteristics, the amount of reduction in larval habitat that can be achieved, and how malaria transmission is affected by temperature.

Analysis of the trap gene provides evidence for the role of elevation and vector abundance in the genetic diversity of Plasmodium relictum in Hawaii.

BACKGROUND: The avian disease system in Hawaii offers an ideal opportunity to investigate host-pathogen interactions in a natural setting. Previous studies have recognized only a single mitochondrial lineage of avian malaria (Plasmodium relictum) in the Hawaiian Islands, but cloning and sequencing of nuclear genes suggest a higher degree of genetic diversity. METHODS: In order to evaluate genetic diversity of P. relictum at the population level and further understand host-parasite interactions, a modified single-base extension (SBE) method was used to explore spatial and temporal distribution patterns of single nucleotide polymorphisms (SNPs) in the thrombospondin-related anonymous protein (trap) gene of P. relictum infections from 121 hatch-year amakihi (Hemignathus virens) on the east side of Hawaii Island. RESULTS: Rare alleles and mixed infections were documented at three of eight SNP loci; this is the first documentation of genetically diverse infections of P. relictum at the population level in Hawaii. Logistic regression revealed that the likelihood of infection with a rare allele increased at low-elevation, but decreased as mosquito capture rates increased. The inverse relationship between vector capture rates and probability of infection with a rare allele is unexpected given current theories of epidemiology developed in human malarias. CONCLUSIONS: The results of this study suggest that pathogen diversity in Hawaii may be driven by a complex interaction of factors including transmission rates, host immune pressures, and parasite-parasite competition.

Ecology and conservation biology of avian malaria.

Avian malaria is a worldwide mosquito-borne disease caused by Plasmodium parasites. These parasites occur in many avian species but primarily affect passerine birds that have not evolved with the parasite. Host pathogenicity, fitness, and population impacts are poorly understood. In contrast to continental species, introduced avian malaria poses a substantial threat to naive birds on Hawaii, the Galapagos, and other archipelagoes. In Hawaii, transmission is maintained by susceptible native birds, competence and abundance of mosquitoes, and a disease reservoir of chronically infected native birds. Although vector habitat and avian communities determine the geographic distribution of disease, climate drives transmission patterns ranging from continuous high infection in warm lowland forests, seasonal infection in midelevation forests, and disease-free refugia in cool high-elevation forests. Global warming is expected to increase the occurrence, distribution, and intensity of avian malaria across this elevational gradient and threaten high-elevation refugia, which is the key to survival of many susceptible Hawaiian birds. Increased temperatures may have already increased global avian malaria prevalence and contributed to an emergence of disease in New Zealand.

Taqman real-time PCR detects Avipoxvirus DNA in blood of Hawai'i 'amakihi (Hemignathus virens).

BACKGROUND: Avipoxvirus sp. is a significant threat to endemic bird populations on several groups of islands worldwide, including Hawai'i, the Galapagos Islands, and the Canary Islands. Accurate identification and genotyping of Avipoxvirus is critical to the study of this disease and how it interacts with other pathogens, but currently available methods rely on invasive sampling of pox-like lesions and may be especially harmful in smaller birds. METHODOLOGY/PRINCIPAL FINDINGS: Here, we present a nested TaqMan Real-Time PCR for the detection of the Avipoxvirus 4b core protein gene in archived blood samples from Hawaiian birds. The method was successful in amplifying Avipoxvirus DNA from packed blood cells of one of seven Hawaiian honeycreepers with confirmed Avipoxvirus infections and 13 of 28 Hawai'i 'amakihi (Hemignathus virens) with suspected Avipoxvirus infections based on the presence of pox-like lesions. Mixed genotype infections have not previously been documented in Hawai'i but were observed in two individuals in this study. CONCLUSIONS/SIGNIFICANCE: We anticipate that this method will be applicable to other closely related strains of Avipoxvirus and will become an important and useful tool in global studies of the epidemiology of Avipoxvirus.

Thermal constraints to the sporogonic development and altitudinal distribution of avian malaria Plasmodium relictum in Hawai'i.

More than half of the Hawaiian honeycreepers (Drepanidinae) known from historical records are now extinct. Introduced mosquito-borne disease, in particular the avian malaria Plasmodium relictum , has been incriminated as a leading cause of extinction during the 20th century and a major limiting factor in the recovery of remaining species populations. Today, most native Hawaiian bird species reach their highest densities and diversity in high elevation (>1,800 m above sea level) forests. We determined the thermal requirements for sporogonic development of P. relictum in the natural vector, Culex quinquefasciatus , and assessed the current distribution of native bird species in light of this information. Sporogonic development was completed at constant laboratory and mean field temperatures between 30 and 17 C, but development, prevalence, and intensity decreased significantly below 21 C. Using a degree-day (DD) model, we estimated a minimum threshold temperature of 12.97 C and a thermal requirement of 86.2 DD as necessary to complete development. Predicted (adiabatic lapse-rate) and observed summer threshold isotherm (13 C) correspond to the elevation of high forest refuges on the islands of Maui and Hawai'i. Our data support the hypothesis that avian malaria currently restricts the altitudinal distribution of Hawaiian honeycreeper populations and provide an ecological explanation for the absence of disease at high elevation.

Experimental infection of Hawai'i 'Amakihi (hemignathus virens) with West Nile virus and competence of a co-occurring vector, culex quinquefasciatus: potential impacts on endemic Hawaiian avifauna.

Introduced mosquito-borne avian disease is a major limiting factor in the recovery and restoration of native Hawaiian forest birds. Annual epizootics of avian pox (Avipoxvirus) and avian malaria (Plasmodium relictum) likely led to the extinction of some species and continue to impact populations of susceptible Hawaiian honeycreepers (Drepanidinae). The introduction of a novel pathogen, such as West Nile virus (WNV), could result in further population declines and extinctions. During September and October 2004, we infected Hawai'i' Amakihi (Hemignathus virens) with a North American isolate of WNV by needle inoculation and mosquito bite to observe susceptibility, mortality, and illness in this endemic passerine, and to determine the vector competence of the co-occurring, introduced mosquito Culex quinquefasciatus. All experimentally infected Hawai'i ;Amakihi became viremic, with a mean titer >10(5) plaque-forming units (PFU)/ml, and they experienced clinical signs ranging from anorexia and lethargy to ataxia. The fatality rate among needle-inoculated Hawai'i' Amakihi (n=16) was 31.3%, but mortality in free-ranging birds is likely to increase due to predation, starvation, thermal stress, and concomitant infections of avian malaria and pox. Surviving Hawai'i' Amakihi seem to clear WNV from the peripheral blood by 7-10 days postinfection (DPI), and neutralizing antibodies were detected from 9 to 46 DPI. In transmission trials, Hawaiian Cx. quinquefasciatus proved to be a competent vector and Hawai'i Amakihi an adequate amplification host of WNV, suggesting that epizootic WNV could readily become an additional limiting factor of some native Hawaiian bird populations.

Knemidokoptic mange in Hawai'i' Amakihi (Hemignathus virens) on the island of Hawai'i.

Lesions resembling knemidokoptic mange on the feet and tarsometatarsi of two Hawai'i' Amakihi (Hemignathus virens) were observed while the researchers were mist-netting wild passerines at Manuka Natural Area Reserve on the island of Hawai'i between 14 June 2007 and 19 June 2007. During subsequent mist-netting from September 2007 through February 2008, we found 26% (7/27) of the Hawai'i' Amakihi caught were similarly affected. Microscopic examination of skin scrapings from lesions of affected individuals revealed Knemidokoptes jamaicensis (Acari: Knemidokoptidae). This is the first report of Knemidokoptes spp. found in wild passerines in Hawai'i. No other wild passerines (n=573) have been found with knemidokoptic mange during our islandwide study of Hawai'i' Amakihi.

Larval habitat for the avian malaria vector Culex quinquefasciatus (Diptera: Culicidae) in altered mid-elevation mesic-dry forests in Hawai'i.

Effective management of avian malaria (Plasmodium relictum) in Hawai'i's endemic honeycreepers (Drepanidinae) requires the identification and subsequent reduction or treatment of larval habitat for the mosquito vector, Culex quinquefasciatus (Diptera: Culicidae). We conducted ground surveys, treehole surveys, and helicopter aerial surveys from 2001-2003 to identify all potential larval mosquito habitat within two 100+ ha mesic-dry forest study sites in Hawai'i Volcanoes National Park, Hawai'i; 'Ainahou Ranch and Mauna Loa Strip Road. At 'Ainahou Ranch, anthropogenic sites (43%) were more likely to contain mosquitoes than naturally occurring (8%) sites. Larvae of Cx. quinquefasciatus were predominately found in anthropogenic sites while Aedes albopictus larvae occurred less frequently in both anthropogenic sites and naturally-occurring sites. Additionally, moderate-size (~ 20-22,000 liters) anthropogenic potential larval habitat had >50% probability of mosquito presence compared to larger- and smaller-volume habitat (<50%). Less than 20% of trees surveyed at 'Ainahou Ranch had treeholes and few mosquito larvae were detected. Aerial surveys at 'Ainahou Ranch detected 56% (95% CI: 42-68%) of the potential larval habitat identified in ground surveys. At Mauna Loa Strip Road, Cx. quinquefasciatus larvae were only found in the rock holes of small intermittent stream drainages that made up 20% (5 of 25) of the total potential larval habitat. The volume of the potential larval habitat did not influence the probability of mosquito occurrence at Mauna Loa Strip Road. Our results suggest that Cx. quinquefasciatus abundance, and subsequently avian malaria, may be controlled by larval habitat reduction in the mesic-dry landscapes of Hawai'i where anthropogenic sources predominate.

Prevalence and distribution of pox-like lesions, avian malaria, and mosquito vectors in Kipahulu Valley, Haleakala National Park, Hawai'i, USA.

We determined prevalence and altitudinal distribution of introduced avian malarial infections (Plasmodium relictum) and pox-like lesions (Avipoxvirus) in forest birds from Kipahulu Valley, Haleakala National Park, on the island of Maui, and we identified primary larval habitat for the mosquito vector of this disease. This intensively managed wilderness area and scientific reserve is one of the most pristine areas of native forest remaining in the state of Hawai'i, and it will become increasingly important as a site for restoration and recovery of endangered forest birds. Overall prevalence of malarial infections in the valley was 8% (11/133) in native species and 4% (4/101) in nonnative passerines; prevalence was lower than reported for comparable elevations and habitats elsewhere in the state. Infections occurred primarily in 'Apapane (Himatione sanguinea) and Hawai'i 'Amakihi (Hemignathus virens) at elevations below 1,400 m. Pox-like lesions were detected in only two Hawai'i 'Amakihi (2%; 2/94) at elevations below 950 m. We did not detect malaria or pox in birds caught at 1,400 m in upper reaches of the valley. Adult mosquitoes (Culex quinquefasciatus) were captured at four sites at elevations of 640, 760, 915, and 975 m, respectively. Culex quinquefasciatus larvae were found only in rock holes along intermittent tributaries of the two largest streams in the valley, but not in standing surface water, pig wallows, ground pools, tree cavities, and tree fern cavities. Mosquito populations in the valley are low, and they are probably influenced by periods of high rainfall that flush stream systems.

Landscape factors influencing the spatial distribution and abundance of mosquito vector Culex quinquefasciatus (Diptera: Culicidae) in a mixed residential-agricultural community in Hawai'i.

Mosquito-borne avian diseases, principally avian malaria (Plasmodium relictum Grassi and Feletti) and avian pox (Avipoxvirus sp.) have been implicated as the key limiting factor associated with recent declines of endemic avifauna in the Hawaiian Island archipelago. We present data on the relative abundance, infection status, and spatial distribution of the primary mosquito vector Culex quinquefasciatus Say (Diptera: Culicidae) across a mixed, residential-agricultural community adjacent to Hawai'i Volcanoes National Park on Hawai'i Island. We modeled the effect of agriculture and forest fragmentation in determining relative abundance of adult Cx. quinquefasciatus in Volcano Village, and we implement our statistical model in a geographic information system to generate a probability of mosquito capture prediction surface for the study area. Our model was based on biweekly captures of adult mosquitoes from 20 locations within Volcano Village from October 2001 to April 2003. We used mixed effects logistic regression to model the probability of capturing a mosquito, and we developed a set of 17 competing models a priori to specifically evaluate the effect of agriculture and fragmentation (i.e., residential landscapes) at two spatial scales. In total, 2,126 mosquitoes were captured in CO2-baited traps with an average probability of 0.27 (SE = 0.10) of capturing one or more mosquitoes per trap night. Twelve percent of mosquitoes captured were infected with P. relictum. Our data indicate that agricultural lands and forest fragmentation significantly increase the probability of mosquito capture. The prediction surface identified areas along the Hawai'i Volcanoes National Park boundary that may have high relative abundance of the vector. Our data document the potential of avian malaria transmission in residential-agricultural landscapes and support the need for vector management that extends beyond reserve boundaries and considers a reserve's spatial position in a highly heterogeneous landscape.

Host population persistence in the face of introduced vector-borne diseases: Hawaii amakihi and avian malaria.

The past quarter century has seen an unprecedented increase in the number of new and emerging infectious diseases throughout the world, with serious implications for human and wildlife populations. We examined host persistence in the face of introduced vector-borne diseases in Hawaii, where introduced avian malaria and introduced vectors have had a negative impact on most populations of Hawaiian forest birds for nearly a century. We studied birds, parasites, and vectors in nine study areas from 0 to 1,800 m on Mauna Loa Volcano, Hawaii from January to October, 2002. Contrary to predictions of prior work, we found that Hawaii amakihi (Hemignathus virens), a native species susceptible to malaria, comprised from 24.5% to 51.9% of the avian community at three low-elevation forests (55-270 m). Amakihi were more abundant at low elevations than at disease-free high elevations, and were resident and breeding there. Infection rates were 24-40% by microscopy and 55-83% by serology, with most infected individuals experiencing low-intensity, chronic infections. Mosquito trapping and diagnostics provided strong evidence for year-round local transmission. Moreover, we present evidence that Hawaii amakihi have increased in low elevation habitats on southeastern Hawaii Island over the past decade. The recent emergent phenomenon of recovering amakihi populations at low elevations, despite extremely high prevalence of avian malaria, suggests that ecological or evolutionary processes acting on hosts or parasites have allowed this species to recolonize low-elevation habitats. A better understanding of the mechanisms allowing coexistence of hosts and parasites may ultimately lead to tools for mitigating disease impacts on wildlife and human populations.

Comparative susceptibility of introduced forest-dwelling mosquitoes in Hawai'i to avian malaria, Plasmodium relictum.

To identify potential vectors of avian malaria in Hawaiian native forests, the innate susceptibility of Aedes albopictus, Wyeomyia mitchellii, and Culex quinquefasciatus from 3 geographical sites along an altitudinal gradient was evaluated using local isolates of Plasmodium relictum. Mosquitoes were dissected 5-8 and 9-13 days postinfective blood meal and microscopically examined for oocysts and salivary-gland sporozoites. Sporogony was completed in all 3 species, but prevalence between species varied significantly. Oocysts were detected in 1-2% and sporozoites in 1-7% of Aedes albopictus that fed on infected ducklings. Wyeomyia mitchellii was slightly more susceptible, with 7-19% and 7% infected with oocysts and sporozoites, respectively. In both species, the median oocyst number was 5 or below. This is only the second Wyeomyia species reported to support development of a malarial parasite. Conversely, Culex quinquefasciatus from all 3 sites proved very susceptible. Prevalence of oocysts and sporozoites consistently exceeded 70%, regardless of gametocytemia or origin of the P. relictum isolate. In trials for which a maximum 200 oocysts were recorded, the median number of oocysts ranged from 144 to 200. It was concluded that Culex quinquefasciatus is the primary vector of avian malaria in Hawai'i.