Filtration of environmentally sourced aquatic media impacts laboratory-colonised Aedes albopictus early development and adult bacteriome composition.

Microorganisms form close associations with metazoan hosts forming symbiotic communities, known as microbiomes, that modulate host physiological processes. Mosquitoes are of special interest in exploring microbe-modulated host processes due to their oversized impact on human health. However, most mosquito work is done under controlled laboratory conditions where natural microbiomes are not present and inferences from these studies may not extend to natural populations. Here we attempt to assemble a wild-resembling bacteriome under laboratory conditions in an established laboratory colony of Aedes albopictus using aquatic media from environmentally-exposed and differentially filtered larval habitats. While we did not successfully replicate a wild bacteriome using these filtrations, we show that these manipulations alter the bacteriomes of mosquitoes, generating a unique composition not seen in wild populations collected from and near our source water or in our laboratory colony. We also demonstrate that our filtration regimens impact larval development times, as well as impact adult survival on different carbohydrate diets.

Microplastic pollution differentially affects development of disease-vectoring Aedes and Culex mosquitoes.

Plastic in the form of microplastic particles (MPs) is now recognized as a major pollutant of unknown consequences in aquatic habitats. Mosquitoes, with aquatic eggs, larvae, and pupae, are likely to encounter microplastic, particularly those species that are abundant in close proximity to human development, including those that vector human and animal disease. We examined the effects of polyethylene MPs, the most common microplastic documented in environmental samples, on the development and survival of the mosquitoes Aedes albopictus and Culex quinquefasciatus. In laboratory egg-laying and larval development container environments similar to those used by both species in the field, a mix of 1-53 µm MPs at concentrations of 60, 600, and 6000 MP ml-1 increased early instar larval mortality in both species relative to control treatments. A significant difference was found in the response of each species to microplastic at the lowest microplastic concentration tested, with Cx. quinquefasciatus survival equivalent to that in control conditions but with Ae. albopictus larvae mortality elevated to 37% within 48 h. These results differ from those of previous studies in which larvae were only exposed to MPs during the last aquatic instar stage and from which it was concluded that microplastic was ontogenically transferred without negatively affecting development. Increasing plastic pollutant concentrations could therefore act as selective pressures on aquatic larvae and ultimately influence outcomes of ecological interactions among mosquito vector populations.

Mosquito Microbiome Diversity Varies Along a Landscape-Scale Moisture Gradient.

Microorganisms live in close association with metazoan hosts and form symbiotic microbiotas that modulate host biology. Although the function of host-associated microbiomes may change with composition, hosts within a population can exhibit high turnover in microbiome composition among individuals. However, environmental drivers of this variation are inadequately described. Here, we test the hypothesis that this diversity among the microbiomes of Aedes albopictus (a mosquito disease vector) is associated with the local climate and land-use patterns on the high Pacific island of O 'ahu, Hawai 'i. Our principal finding demonstrates that the relative abundance of several bacterial symbionts in the Ae. albopictus microbiome varies in response to a landscape-scale moisture gradient, resulting in the turnover of the mosquito microbiome composition across the landscape. However, we find no evidence that mosquito microbiome diversity is tied to an index of urbanization. This result has implications toward understanding the assembly of host-associated microbiomes, especially during an era of rampant global climate change.

Local host specialization, host-switching, and dispersal shape the regional distributions of avian haemosporidian parasites.

The drivers of regional parasite distributions are poorly understood, especially in comparison with those of free-living species. For vector-transmitted parasites, in particular, distributions might be influenced by host-switching and by parasite dispersal with primary hosts and vectors. We surveyed haemosporidian blood parasites (Plasmodium and Haemoproteus) of small land birds in eastern North America to characterize a regional parasite community. Distributions of parasite populations generally reflected distributions of their hosts across the region. However, when the interdependence between hosts and parasites was controlled statistically, local host assemblages were related to regional climatic gradients, but parasite assemblages were not. Moreover, because parasite assemblage similarity does not decrease with distance when controlling for host assemblages and climate, parasites evidently disperse readily within the distributions of their hosts. The degree of specialization on hosts varied in some parasite lineages over short periods and small geographic distances independently of the diversity of available hosts and potentially competing parasite lineages. Nonrandom spatial turnover was apparent in parasite lineages infecting one host species that was well-sampled within a single year across its range, plausibly reflecting localized adaptations of hosts and parasites. Overall, populations of avian hosts generally determine the geographic distributions of haemosporidian parasites. However, parasites are not dispersal-limited within their host distributions, and they may switch hosts readily.

Species formation by host shifting in avian malaria parasites.

The malaria parasites (Apicomplexa: Haemosporida) of birds are believed to have diversified across the avian host phylogeny well after the origin of most major host lineages. Although many symbionts with direct transmission codiversify with their hosts, mechanisms of species formation in vector-borne parasites, including the role of host shifting, are poorly understood. Here, we examine the hosts of sister lineages in a phylogeny of 181 putative species of malaria parasites of New World terrestrial birds to determine the role of shifts between host taxa in the formation of new parasite species. We find that host shifting, often across host genera and families, is the rule. Sympatric speciation by host shifting would require local reproductive isolation as a prerequisite to divergent selection, but this mechanism is not supported by the generalized host-biting behavior of most vectors of avian malaria parasites. Instead, the geographic distribution of individual parasite lineages in diverse hosts suggests that species formation is predominantly allopatric and involves host expansion followed by local host-pathogen coevolution and secondary sympatry, resulting in local shifting of parasite lineages across hosts.

Prevalence of avian haemosporidian parasites is positively related to the abundance of host species at multiple sites within a region.

Parasite prevalence is thought to be positively related to host population density owing to enhanced contagion. However, the relationship between prevalence and local abundance of multiple host species is underexplored. We surveyed birds and their haemosporidian parasites (genera Plasmodium and Haemoproteus) at multiple sites across eastern North America to test whether the prevalence of these parasites in a host species at a particular site is related to that host's local abundance. Prevalence was positively related to host abundance within most sites, although the effect was stronger and more consistent for Plasmodium than for Haemoproteus. In contrast, prevalence was not related to variation in the abundance of most individual host species among sites across the region. These results suggest that parasite prevalence partly reflects the relative abundances of host species in local assemblages. However, three nonnative host species had low prevalence despite being relatively abundant at one site, as predicted by the enemy release hypothesis.

Multiple Reversals of Bill Length over 1.7 Million Years in a Hawaiian Bird Lineage.

Evolutionary change has been documented over geological time, but reversals in morphology, from an ancestral state to a derived state and back again, tend to be rare. Multiple reversals along the same lineage are even rarer. We use the chronology of the Hawaiian Islands and an avian example, the Hawaiian honeycreeper 'amakihi (Hemignathus spp.) lineage, which originated on the oldest main island of Kaua'i 1.7 million years ago, to examine the process of sequential reversals in bill length. We document three single and two multiple reversals of bill length on six main islands from oldest to youngest, consistent with the phylogeny of the lineage. Longer bills occur on islands with endemic species, including phylogenetically relevant outgroups, that may compete with or dominate the 'amakihi. On islands without those species, the 'amakihi had shorter bills of similar length. Both types of reversals in morphology in this lineage integrate microevolutionary processes with macroevolution in the adaptive radiation of Hawaiian honeycreepers.

Decreased small mammal and on-host tick abundance in association with invasive red imported fire ants (Solenopsis invicta).

Invasive species may impact pathogen transmission by altering the distributions and interactions among native vertebrate reservoir hosts and arthropod vectors. Here, we examined the direct and indirect effects of the red imported fire ant (Solenopsis invicta) on the native tick, small mammal and pathogen community in southeast Texas. Using a replicated large-scale field manipulation study, we show that small mammals were more abundant on treatment plots where S. invicta populations were experimentally reduced. Our analysis of ticks on small mammal hosts demonstrated a threefold increase in the ticks caught per unit effort on treatment relative to control plots, and elevated tick loads (a 27-fold increase) on one common rodent species. We detected only one known human pathogen (Rickettsia parkeri), present in 1.4% of larvae and 6.7% of nymph on-host Amblyomma maculatum samples but with no significant difference between treatment and control plots. Given that host and vector population dynamics are key drivers of pathogen transmission, the reduced small mammal and tick abundance associated with S. invicta may alter pathogen transmission dynamics over broader spatial scales.

Plasmodium prevalence across avian host species is positively associated with exposure to mosquito vectors.

The prevalence of vector-borne parasites varies greatly across host species, and this heterogeneity has been used to relate infectious disease susceptibility to host species traits. However, a few empirical studies have directly associated vector-borne parasite prevalence with exposure to vectors across hosts. Here, we use DNA sequencing of blood meals to estimate utilization of different avian host species by Culex mosquitoes, and relate utilization by these malaria vectors to avian Plasmodium prevalence. We found that avian host species that are highly utilized as hosts by avian malaria vectors are significantly more likely to have Plasmodium infections. However, the effect was not consistent among individual Plasmodium taxa. Exposure to vector bites may therefore influence the relative number of all avian Plasmodium infections among host species, while other processes, such as parasite competition and host-parasite coevolution, delimit the host distributions of individual Plasmodium species. We demonstrate that links between avian malaria susceptibility and host traits can be conditioned by patterns of exposure to vectors. Linking vector utilization rates to host traits may be a key area of future research to understand mechanisms that produce variation in the prevalence of vector-borne pathogens among host species.

Identification of Avian and Hemoparasite DNA in Blood-Engorged Abdomens of Culex pipiens (Diptera; Culicidae) from a West Nile Virus Epidemic region in Suburban Chicago, Illinois.

Multiple mosquito-borne parasites cocirculate in nature and potentially interact. To understand the community of parasites cocirculating with West Nile virus (WNV), we screened the bloodmeal content of Culex pipiens L. mosquitoes for three common types of hemoparasites. Blood-fed Cx. pipiens were collected from a WNV-epidemic area in suburban Chicago, IL, from May to September 2005 through 2010. DNA was extracted from dissected abdomens and subject to PCR and direct sequencing to identify the vertebrate host. RNA was extracted from the head or thorax and screened for WNV using quantitative reverse transcriptase PCR. Seventy-nine engorged females with avian host origin were screened using PCR and amplicon sequencing for filarioid nematodes, Haemosporida, and trypanosomatids. Filarioid nematodes were identified in 3.8% of the blooded abdomens, Plasmodium sp. in 8.9%, Haemoproteus in 31.6%, and Trypanosoma sp. in 6.3%. The sequences from these hemoparasite lineages were highly similar to sequences from birds in prior studies in suburban Chicago. Overall, 50.6% of blood-fed Culex pipiens contained hemoparasite DNA in their abdomen, presumably from current or prior bloodmeals. Additionally, we detected hemoparasite DNA in the blooded abdomen of three of 10 Cx. pipiens infected with WNV.

Host compatibility rather than vector-host-encounter rate determines the host range of avian Plasmodium parasites.

Blood-feeding arthropod vectors are responsible for transmitting many parasites between vertebrate hosts. While arthropod vectors often feed on limited subsets of potential host species, little is known about the extent to which this influences the distribution of vector-borne parasites in some systems. Here, we test the hypothesis that different vector species structure parasite-host relationships by restricting access of certain parasites to a subset of available hosts. Specifically, we investigate how the feeding patterns of Culex mosquito vectors relate to distributions of avian malaria parasites among hosts in suburban Chicago, IL, USA. We show that Plasmodium lineages, defined by cytochrome b haplotypes, are heterogeneously distributed across avian hosts. However, the feeding patterns of the dominant vectors (Culex restuans and Culex pipiens) are similar across these hosts, and do not explain the distributions of Plasmodium parasites. Phylogenetic similarity of avian hosts predicts similarity in their Plasmodium parasites. This effect was driven primarily by the general association of Plasmodium parasites with particular host superfamilies. Our results suggest that a mosquito-imposed encounter rate does not limit the distribution of avian Plasmodium parasites across hosts. This implies that compatibility between parasites and their avian hosts structure Plasmodium host range.

Aedes albopictus microbiome derives from environmental sources and partitions across distinct host tissues.

The mosquito microbiome consists of a consortium of interacting microorganisms that reside on and within culicid hosts. Mosquitoes acquire most of their microbial diversity from the environment over their life cycle. Once present within the mosquito host, the microbes colonize distinct tissues, and these symbiotic relationships are maintained by immune-related mechanisms, environmental filtering, and trait selection. The processes that govern how environmental microbes assemble across the tissues within mosquitoes remain poorly resolved. We use ecological network analyses to examine how environmental bacteria assemble to form bacteriomes among Aedes albopictus host tissues. Mosquitoes, water, soil, and plant nectar were collected from 20 sites in the Manoa Valley, Oahu. DNA was extracted and associated bacteriomes were inventoried using Earth Microbiome Project protocols. We find that the bacteriomes of A. albopictus tissues were compositional taxonomic subsets of environmental bacteriomes and suggest that the environmental microbiome serves as a source pool that supports mosquito microbiome diversity. Within the mosquito, the microbiomes of the crop, midgut, Malpighian tubules, and ovaries differed in composition. This microbial diversity partitioned among host tissues formed two specialized modules: one in the crop and midgut, and another in the Malpighian tubules and ovaries. The specialized modules may form based on microbe niche preferences and/or selection of mosquito tissues for specific microbes that aid unique biological functions of the tissue types. A strong niche-driven assembly of tissue-specific microbiotas from the environmental species pool suggests that each tissue has specialized associations with microbes, which derive from host-mediated microbe selection.

Stressed snails release Angiostrongylus cantonensis (rat lungworm) larvae in their slime.

This study investigated the influence of stress on release of Angiostrongylus cantonensis larvae from a snail host, Parmarion martensi. We subjected 140 infected, wild-caught P. martensi to three stress-inducing treatments (heat, molluscicide, physical disturbance) and an unstressed control treatment for 24 h, after which larval presence and abundance in the slime were quantified by qPCR targeting the ITS1 region of the parasite's DNA, and compared among treatments. The significance of stress and host infection load on larval release was determined by generalized linear mixed models and permutation tests. The results indicated that stress significantly increased the probability of larval presence in slime and the number of larvae released, and highly infected snails were also more likely to release larvae. Among stressed snails, 13.3% released larvae into slime, the number of larvae present in the slime ranging from 45.5 to 4216. Unstressed controls released no larvae. This study offers a partial explanation for conflicting results from prior studies regarding A. cantonensis presence in snail slime and sheds light on the broader One Health implications. Stress-induced larval release highlights the potential role of slime as a medium for pathogen transmission to accidental, paratenic, definitive and other intermediate hosts. These findings emphasize the importance of considering stress-mediated interactions in host-parasite systems and their implications for zoonotic disease emergence. As stressors continue to escalate because of anthropogenic activities and climate change, understanding the role of stress in pathogen shedding and transmission becomes increasingly important for safeguarding human and wildlife health within the One Health framework.

Host snail species exhibit differential Angiostrongylus cantonensis prevalence and infection intensity across an environmental gradient.

Diverse snail species serve as intermediate hosts of the parasitic nematode Angiostrongylus cantonensis, the etiological agent of human neuroangiostrongyliasis. However, levels of A. cantonensis infection prevalence and intensity vary dramatically among these host species. Factors contributing to this variation are largely unknown. Environmental factors, such as precipitation and temperature, have been correlated with overall A. cantonensis infection levels in a locale, but the influence of environment on infection in individual snail species has not been addressed. We identified levels of A. cantonensis prevalence and intensity in 16 species of snails collected from 29 sites along an environmental gradient on the island of Oahu, Hawaii. The relationship between infection levels of individual species and their environment was evaluated using AIC model selection of Generalized Linear Mixed Models incorporating precipitation, temperature, and vegetation cover at each collection site. Our results indicate that different mechanisms drive parasite prevalence and intensity in the intermediate hosts. Overall, snails from rainy, cool, green sites had higher infection levels than snails from dry, hot sites with less green vegetation. Intensity increased at the same rate along the environmental gradient in all species, though at different levels, while the relation between prevalence and environmental variables depended on species. These results have implications for zoonotic transmission, as human infection is a function of infection in the intermediate hosts, ingestion of which is the main pathway of transmission. The probability of human infection is greater in locations with higher rainfall, lower temperature and more vegetation cover because of higher infection prevalence in the gastropod hosts, but this depends on the host species. Moreover, severity of neuroangiostrongyliasis symptoms is likely to be greater in locations with higher rainfall, lower temperature, and more vegetation because of the higher numbers of infectious larvae (infection intensity) in all infected snail species. This study highlights the variation of infection prevalence and intensity in individual gastropod species, the individualistic nature of interactions between host species and their environment, and the implications for human neuroangiostrongyliasis in different environments.

Wild gut microbiomes reveal individuals, species, and location as drivers of variation in two critically endangered Hawaiian honeycreepers.

BACKGROUND: The gut microbiome of animals is an important component that has strong influence on the health, fitness, and behavior of its host. Most research in the microbiome field has focused on human populations and commercially important species. However, researchers are now considering the link between endangered species conservation and the microbiome. In Hawai'i, several threats (e.g., avian malaria and habitat loss) have caused widespread population declines of Hawaiian honeycreepers (subfamily: Carduelinae). These threats can have a significant effect on the avian gut microbiome and may even lead to disruption of microbial function. However, the gut microbiome of honeycreeper in the wild has yet to be explored. METHODS: We collected 13 and 42 fecal samples, respectively, from two critically endangered honeycreeper species, the 'akikiki (Oreomystis bairdi) and the 'akeke'e (Loxops caeruleirostris). The 16S rRNA gene was sequenced and processed though a MOTHUR-based bioinformatics pipeline. Bacterial ASVs were identified using the DADA2 program and bacterial community analyses, including alpha and beta diversity measures, were conducted using R packages Phyloseq and vegan. RESULTS: A total of 8,958 bacterial ASVs were identified from the fecal samples. Intraspecific differences in the gut microbiome among individual birds explained most of the variation present in the dataset, however differences between species did exist. Both species had distinct microbiomes with minimal overlap in beta diversity. 'Akikiki had a more diverse microbiome compared to 'akeke'e. Additionally, small but stastically significant differences in beta diversity also exist between sampling location and sexes in 'akikiki. CONCLUSION: 'Akikiki and 'akeke'e are currently the focus of captive breeding efforts and plans to translocate the two species to other islands are underway. This baseline knowledge will help inform management decisions for these honeycreeper species in their native habitats, on other islands, and in captivity.

Variable coverage in an Autocidal Gravid Ovitrap intervention impacts efficacy of Aedes aegypti control.

Control of the arboviral disease vector Aedes aegypti has shown variable levels of efficacy around the globe. We evaluated an Autocidal Gravid Ovitrap (AGO) intervention as a stand-alone control tool for population suppression of A. aegypti in US communities bordering Mexico.We conducted a cluster randomized crossover trial with weekly mosquito surveillance of sentinel households from July 2017 to December 2018. The intervention took place from August to December of both years. Multilevel models (generalized linear and additive mixed models) were used to analyse the changes in population abundance of female A. aegypti.We observed that female populations were being suppressed 77% (2018) and four times lower outdoor female abundance when AGO coverage (number of intervention AGO traps that surrounded a sentinel home) was high (2.7 AGOs/house). However, we also observed that areas with low intervention AGO coverage resulted in no difference (2017) or slightly higher abundance compared to the control. These results suggest that coverage rate might play a critical role on how populations of female A. aegypti are being modulated in the field. The lack of larval source habitat reduction and the short duration of the intervention period might have limited the A. aegypti population suppression observed in this study. Synthesis and applications. The mosquito, A. aegypti, is a public health concern in most tropical and subtropical regions. With the rise of insecticide resistance, the evaluation of non-chemical tools has become pivotal in the fight against arboviral disease transmission. Our study shows that the AGO intervention, as a stand-alone control tool, is limited by its coverage in human settlements. Vector control programmes should consider, that if the target coverage rate is not achieved, measures will be ineffective unless coupled with other control approaches. Although our multilevel modelling was focused on A. aegypti and the AGO, the approach can be applied to other mosquito vector species.

Vector-Borne Blood Parasites of the Great-Tailed Grackle (Quiscalus mexicanus) in East-Central Texas, USA.

Great-tailed grackles (Quiscalus mexicanus) have dramatically expanded into North America over the past century. However, little is known about the blood that parasites they support. Here, for the first time, we document an assemblage of trypanosome, haemosporida, and filarial nematodes co-circulating in invasive great-tailed grackles. Between February and July, 2015, 61 individuals were captured in an urban environment of College Station, Texas. Field microscopy and molecular diagnostics indicate that 52% (24/46) were visually infected with filarioid nematodes, 24% (11/46) with avian trypanosomes, and 73% (n = 44/60) with haemosporida parasites, such as Haemoproteus (Parahaemoproteus) and Plasmodium cathemerium. Overall, 87% of great-tailed grackles were infected with blood parasites. Although 50% of individuals hosted parasites from multiple phylum, no patterns of parasite assembly were observed. Results indicate that great-tailed grackles can support a relatively high level of blood parasitism. However, the consequences for avian health remain to be determined.

The Eco-Bio-Social Factors That Modulate Aedes aegypti Abundance in South Texas Border Communities.

Aedes aegypti control requires dedicated resources that are usually scarce, limiting the reach and sustainability of vector control programs. This generates a need to focus on areas at risk of disease transmission and also understand the factors that might modulate local mosquito abundance. We evaluated the eco-bio-social factors that modulate indoor and outdoor relative abundance of female Ae. aegypti in communities of South Texas. We conducted housing quality and Knowledge Attitudes and Practices surveys in households that were part of a weekly mosquito surveillance program in November of 2017 and 2018. Our results showed widespread knowledge of mosquitoes and Zika virus by our participants. However, less than 35% considered them as serious problems in this region. The presence of window-mounted air conditioning units increased the risk of female mosquito relative abundance indoors. An increase in outdoor relative abundance was associated with larger properties and a higher number of children between 6 to 17 years of age. Interestingly, we observed that an increasing number of children <5 years of age modulated both indoor and outdoor relative abundance, with a 52% increase indoors and 30% decrease outdoors. The low perception of mosquito and disease risk highlights engagement needs for vector-borne disease prevention in this region. The identified risk factors can help guide public health officials in their efforts to reduce human and vector contact.

Landscape and demographic determinants of Culex infection with West Nile virus during the 2012 epidemic in Dallas County, TX.

In 2012, the United States experienced one of the largest outbreaks of West Nile virus (WNV)-associated deaths, with the majority occurring in Dallas County (Co.), Texas (TX) and surrounding areas. In this study, logistic mixed models were used to identify associations between the landscape, human population, and WNV-infected Culex quinquefasciatus mosquitoes during the 2012 WNV epidemic in Dallas Co. We found increased probabilities for WNV-positive mosquitoes in north and central Dallas Co. The most significant predictors of the presence of WNV in Cx. quinquefasciatus pools were increased urbanization (based on an index composed of greater population density, lower normalized difference vegetation index, higher coverage of urban land types, and more impervious surfaces), older human populations, and lower elevation. These relationships between the landscape, sociodemographics, and risk of enzootic transmission identified regions of Dallas Co., TX with highest risk of spillover to human disease during the 2012 WNV epidemic.

Species Identity and Size are Associated with Rat Lungworm Infection in Gastropods.

Angiostrongylus cantonensis, the rat lungworm, is an emerging zoonotic pathogen that cycles between definitive rat and intermediate gastropod hosts. Zoonotic infection occurs when humans intentionally or accidentally consume infectious larvae in a gastropod host, and may manifest as neuroangiostrongyliasis, characterized by eosinophilic meningitis, severe neurological impairment, and even death. Thus, the risk of A. cantonensis zoonoses may be related to the distribution of A. cantonensis larvae across gastropod hosts. We screened 16 gastropod species from 14 communities on the island of O'ahu, Hawai'i, USA, to characterize the distribution of A. cantonensis among species and across host size. Prevalence (proportion of the population infected) and infection intensity (density of worms in host tissue) varied among gastropod species. Prevalence also varied with gastropod host size, but this relationship differed among host species. Most host species showed a positive increase in the probability of infection with host size, suggesting that within species relatively larger hosts had higher prevalence. The density of worms in an infected snail was unrelated to host size. These results suggest that variation in A. cantonensis infection is associated with demographic structure and composition of gastropod communities, which could underlie heterogeneity in the risk of human angiostrongyliasis across landscapes.