Mosquito Microbiome Research Articles

Dietary L-3,4-dihydroxyphenylalanine (L-DOPA) augments cuticular melanization in Anopheles mosquitos while reducing their lifespan and malaria parasite burden.

L-3,4-dihydroxyphenylalanine (L-DOPA), a naturally occurring tyrosine derivative, is prevalent in environments that include mosquito habitats, potentially serving as part of their diet. Given its role as a precursor for melanin synthesis we investigated the effect of dietary L-DOPA on mosquito physiology and immunity to Plasmodium falciparum and Cryptococcus neoformans infection. Dietary L-DOPA was incorporated into mosquito melanin via a non-canonical pathway and had profound transcriptional effects that were associated with enhanced immunity, increased pigmentation, and reduced lifespan. Increased melanization resulted in an enhanced capacity to absorb electromagnetic radiation that affected mosquito temperatures. Bacteria in the mosquito microbiome were sources of dopamine, which is a substrate for melanization. Our results illustrate how an environmentally abundant amino acid analogue can affect mosquito physiology and suggest its potential usefulness as an environmentally friendly vector control agent to reduce malaria transmission, warranting further research and field studies.

Abiotic factors shape mosquito microbiomes that enhance host development.

Metazoans rely on interactions with microorganisms through multiple life stages. For example, developmental trajectories of mosquitoes can vary depending on the microorganisms available during their aquatic larval phase. However, the role that the local environment plays in shaping such host-microbe dynamics and the consequences for the host organism remain inadequately understood. Here, we examine the influence of abiotic factors, locally available bacteria, and their interactions on the development and associated microbiota of the mosquito Aedes albopictus. Our findings reveal that leaf detritus infused into the larval habitat water, sourced from native Hawaiian tree "ōhi'a lehua Metrosideros polymorpha, invasive strawberry guava Psidium cattleyanum, or a pure water control, displayed a more substantial influence than either temperature variations or simulated microbial dispersal regimes on bacterial community composition in adult mosquitoes. However, specific bacteria exhibited divergent patterns within mosquitoes across detrital infusions which did not align with their abundance in the larval habitat. Specifically, we observed a higher relative abundance of a Chryseobacterium sp. strain in mosquitoes from the strawberry guava infusion than the pure water control, whereas the opposite trend was observed for a Pseudomonas sp. strain. In a follow-up experiment, we manipulated the presence of these two bacterial strains and found larval developmental success was enhanced by including the Chryseobacterium sp. strain in the strawberry guava infusion and the Pseudomonas sp. strain in the pure water control. Collectively, these data suggest that interactions between abiotic factors and microbes of the larval environment can help shape mosquito populations" success.

Characterization of the microbiome of Aedes albopictus populations in different habitats from Spain and São Tomé

The mosquito microbiome significantly influences vector competence, including in Aedes albopictus, a globally invasive vector. Describing the microbiome and Wolbachia strains of Ae. albopictus from different regions can guide area-specific control strategies. Mosquito samples from Spain and São Tomé were analyzed using 16S rRNA gene sequencing and metagenomic sequencing. Wolbachia infection patterns were observed by sex and population. Female mosquitoes were blood-fed, a factor considered in analyzing their microbiota. Results revealed a dominance of dual Wolbachia infections, strains A and B, in the microbiome of both populations of Ae. albopictus, especially among females. Both populations shared a core microbiome, although 5 and 9 other genera were only present in Spain and São Tomé populations, respectively. Genera like Pelomonas and Nevskia were identified for the first time in Aedes mosquitoes. This study is the first to describe the Ae. albopictus bacteriome in Spain and São Tomé, offering insights for the development of targeted mosquito control strategies. Understanding the specific microbiome composition can help in designing more effective interventions, such as microbiome manipulation and Wolbachia-based approaches, to reduce vector competence and transmission potential of these mosquitoes.

Mosquitoes reared in distinct insectaries within an institution in close spatial proximity possess significantly divergent microbiomes.

The microbiome affects important aspects of mosquito biology and differences in microbial composition can affect the outcomes of laboratory studies. To determine how the biotic and abiotic conditions in an insectary affect the composition of the bacterial microbiome of mosquitoes we reared mosquitoes from a single cohort of eggs from one genetically homogeneous inbred Aedes aegypti colony, which were split into three batches, and transferred to each of three different insectaries located within the Liverpool School of Tropical Medicine. Using three replicate trays per insectary, we assessed and compared the bacterial microbiome composition as mosquitoes developed from these eggs. We also characterised the microbiome of the mosquitoes' food sources, measured environmental conditions over time in each climate-controlled insectary, and recorded development and survival of mosquitoes. While mosquito development was overall similar between all three insectaries, we saw differences in microbiome composition between mosquitoes from each insectary. Furthermore, bacterial input via food sources, potentially followed by selective pressure of temperature stability and range, did affect the microbiome composition. At both adult and larval stages, specific members of the mosquito microbiome were associated with particular insectaries; and the insectary with less stable and cooler conditions resulted in slower pupation rate and higher diversity of the larval microbiome. Tray and cage effects were also seen in all insectaries, with different bacterial taxa implicated between insectaries. These results highlight the necessity of considering the variability and effects of different microbiome composition even in experiments carried out in a laboratory environment starting with eggs from one batch; and highlights the impact of even minor inconsistencies in rearing conditions due to variation of temperature and humidity.

The dynamics of the midgut microbiome in Aedes aegypti during digestion reveal putative symbionts.

Blood-feeding is crucial for the reproductive cycle of the mosquito Aedes aegypti, as well as for the transmission of arboviruses to hosts. It is postulated that blood meals may influence the mosquito microbiome but shifts in microbial diversity and function during digestion remain elusive. We used whole-genome shotgun metagenomics to monitor the midgut microbiome in 60 individual females of A. aegypti throughout digestion, after 12, 24, and 48 h following blood or sugar meals. Additionally, ten individual larvae were sequenced, showing microbiomes dominated by Microbacterium sp. The high metagenomic coverage allowed for microbial assignments at the species taxonomic level, also providing functional profiling. Females in the post-digestive period and larvae displayed low microbiome diversities. A striking proliferation of Enterobacterales was observed during digestion in blood-fed mosquitoes. The compositional shift was concomitant with enrichment in genes associated with carbohydrate and protein metabolism, as well as virulence factors for antimicrobial resistance and scavenging. The bacterium Elizabethkingia anophelis (Flavobacteriales), a known human pathogen, was the dominant species at the end of blood digestion. Phylogenomics suggests that its association with hematophagous mosquitoes occurred several times. We consider evidence of mutually beneficial host-microbe interactions raised from this association, potentially pivotal for the mosquito's resistance to arbovirus infection. After digestion, the observed shifts in blood-fed females' midguts shifted to a sugar-fed-like microbial profile. This study provides insights into how the microbiome of A. aegypti is modulated to fulfil digestive roles following blood meals, emphasizing proliferation of potential symbionts in response to the dynamic midgut environment.

Mosquito Microbiomes of Rwanda: Characterizing Mosquito Host and Microbial Communities in the Land of a Thousand Hills.

Mosquitoes are a complex nuisance around the world and tropical countries bear the brunt of the burden of mosquito-borne diseases. Rwanda has had success in reducing malaria and some arboviral diseases over the last few years, but still faces challenges to elimination. By building our understanding of in situ mosquito communities in Rwanda at a disturbed, human-occupied site and at a natural, preserved site, we can build our understanding of natural mosquito microbiomes toward the goal of implementing novel microbial control methods. Here, we examined the composition of collected mosquitoes and their microbiomes at two diverse sites using Cytochrome c Oxidase I sequencing and 16S V4 high-throughput sequencing. The majority (36 of 40 species) of mosquitoes captured and characterized in this study are the first-known record of their species for Rwanda but have been characterized in other nations in East Africa. We found significant differences among mosquito genera and among species, but not between mosquito sexes or catch method. Bacteria of interest for arbovirus control, Asaia, Serratia, and Wolbachia, were found in abundance at both sites and varied greatly by species.

Revealing the microbiome diversity and biocontrol potential of field Aedes ssp.: Implications for disease vector management.

The mosquito Aedes spp. holds important relevance for human and animal health, as it serves as a vector for transmitting multiple diseases, including dengue and Zika virus. The microbiome's impact on its host's health and fitness is well known. However, most studies on mosquito microbiomes have been conducted in laboratory settings. We explored the mixed microbial communities within Aedes spp., utilizing the 16S rRNA gene for diversity analysis and shotgun metagenomics for functional genomics. Our samples, which included Ae. aegypti and Ae. albopictus, spanned various developmental stages-eggs, larvae, and adults-gathered from five semiurban areas in Mexico. Our findings revealed a substantial diversity of 8,346 operational taxonomic units (OTUs), representing 967 bacterial genera and 126,366 annotated proteins. The host developmental stage was identified as the primary factor associated with variations in the microbiome composition. Subsequently, we searched for genes and species involved in mosquito biocontrol. Wolbachia accounted for 9.6% of the 16S gene sequences. We observed a high diversity (203 OTUs) of Wolbachia strains commonly associated with mosquitoes, such as wAlb, with a noticeable increase in abundance during the adult stages. Notably, we detected the presence of the cifA and cifB genes, which are associated with Wolbachia's cytoplasmic incompatibility, a biocontrol mechanism. Additionally, we identified 221 OTUs related to Bacillus, including strains linked to B. thuringiensis. Furthermore, we discovered multiple genes encoding insecticidal toxins, such as Cry, Mcf, Vip, and Vpp. Overall, our study contributes to the understanding of mosquito microbiome biodiversity and metabolic capabilities, which are essential for developing effective biocontrol strategies against this disease vector.

Harnessing the power of mosquito gut microbiome in disease control

Mosquito-borne diseases pose an escalating threat to global public health, necessitating more effective strategies for vector control and disease prevention. Recent investigations into the mosquito gut microbiome unveil new avenues for symbiont-based control of mosquito-borne diseases, as it has been revealed that the mosquito gut microbiome has a profound impact on disease transmission. Accumulating evidence demonstrates that the mosquito gut microbiome plays a significant role in modulating infection by generating pathogen-targeting molecules, influencing the midgut physical barrier, shaping the biochemical environment, and regulating immune responses, and in mosquito ecology which affects disease transmission dynamics. Several proof-of-principle studies have explored some microbiome-based control measures, including supplementing the mosquitoes with microorganisms possessing anti-pathogen activities, immunizing hosts against pro-pathogen bacteria, and limiting mosquito larval development using bacteriophages. These strategies present promising efficacy, yet their practical implementation demands further scrutiny and assessment.

Geographical Characteristics of Culex tritaeniorhynchus and Culex orientalis Microbiomes in Korea.

Mosquitoes, the primary vectors of arboviruses, harbor a diverse microbiome that plays a crucial role in their development, immunity, and vector competence. The composition of the mosquito microbiome is heavily influenced by the environment and habitats. Therefore, identifying the relationship between the habitat and the mosquito's microbial community can improve the overall understanding of mosquito biology. However, The microbiome profiles of Culex tritaeniorhynchus and Culex orientalis, known as transmission vectors of the Japanese encephalitis virus, are poorly understood. Using 16S rRNA Illumina sequencing, we hereby investigated the microbial profiles in these two mosquito species collected in several areas in the Republic of Korea. Thirty-six prevalent bacterial families were identified from these mosquito species. The microbial composition variations were primarily influenced by the mosquito collecting sites. Moreover, species biomarkers were identified by utilizing the regional specificity of the mosquito microbiome. Based on the microbiome profiles representing high similarity, Culex orientalis may share an ecological niche with Culex tritaeniorhynchus.

Interkingdom interactions shape the fungal microbiome of mosquitoes

BackgroundThe mosquito microbiome is an important modulator of vector competence and vectoral capacity. Unlike the extensively studied bacterial microbiome, fungal communities in the mosquito microbiome (the mycobiome) remain largely unexplored. To work towards getting an improved understanding of the fungi associated with mosquitoes, we sequenced the mycobiome of three field-collected and laboratory-reared mosquito species (Aedes albopictus, Aedes aegypti, and Culex quinquefasciatus).ResultsOur analysis showed both environment and host species were contributing to the diversity of the fungal microbiome of mosquitoes. When comparing species, Ae. albopictus possessed a higher number of diverse fungal taxa than Cx. quinquefasciatus, while strikingly less than 1% of reads from Ae. aegypti samples were fungal. Fungal reads from Ae. aegypti were < 1% even after inhibiting host amplification using a PNA blocker, indicating that this species lacked a significant fungal microbiome that was amplified using this sequencing approach. Using a mono-association mosquito infection model, we confirmed that mosquito-derived fungal isolates colonize Aedes mosquitoes and support growth and development at comparable rates to their bacterial counterparts. Strikingly, native bacterial taxa isolated from mosquitoes impeded the colonization of symbiotic fungi in Ae. aegypti suggesting interkingdom interactions shape fungal microbiome communities.ConclusionCollectively, this study adds to our understanding of the fungal microbiome of different mosquito species, that these fungal microbes support growth and development, and highlights that microbial interactions underpin fungal colonization of these medically relevent species.

Two promising candidates for paratransgenesis, Elizabethkingia and Asaia, increase in both sexes of Anopheles gambiae mosquitoes after feeding

BackgroundThe male mosquito microbiome may be important for identifying ideal candidates for disease control. Among other criteria, mosquito-associated symbionts that have high localization in both male and female mosquitoes and are transmissible through both vertical and sexual routes are desirable. However, mosquito microbiome studies have mainly been female-focused. In this study, the microbiota of male and female Anopheles gambiae sensu lato (s.l.) were compared to identify shared or unique bacteria.MethodsLate larval instars of Anopheles mosquitoes were collected from the field and raised to adults. Equal numbers of males and females of 1-day-old non-sugar-fed, 4–5-day-old sugar-fed and post-blood-fed females were randomly selected for whole-body analyses of bacteria 16S rRNA.ResultsResults revealed that male and female mosquitoes generally share similar microbiota except when females were blood-fed. Compared to newly emerged unfed mosquitoes, feeding on sugar and/or blood increased variability in microbial composition (⍺-diversity), with a higher disparity among females (39% P = 0.01) than in males (29% P = 0.03). Elizabethkingia meningoseptica and Asaia siamensis were common discriminants between feeding statuses in both males and females. While E. meningoseptica was particularly associated with sugar-fed mosquitoes of both sexes and sustained after blood feeding in females, A. siamensis was also increased in sugar-fed mosquitoes but decreased significantly in blood-fed females (LDA score > 4.0, P < 0.05). Among males, A. siamensis did not differ significantly after sugar meals.ConclusionsResults indicate the opportunities for stable infection in mosquitoes should these species be used in bacteria-mediated disease control. Further studies are recommended to investigate possible host-specific tissue tropism of bacteria species which will inform selection of the most appropriate microbes for effective transmission-blocking strategies.

Effects of seasonality and developed land cover on Culex mosquito abundance and microbiome diversity.

The vectorial capacity of mosquitoes, which influences the dynamics of vector-borne disease transmission, is intricately linked to mosquito abundance and the composition and diversity of their associated microbiomes. However, the influence of environmental factors on mosquito populations and microbiome diversity remains underexplored. Here we examined the effects of seasonality and developed land cover on Culex mosquito abundance and bacterial diversity. Biweekly field sampling of female Culex mosquitoes was conducted using gravid and CDC light traps, spanning summer to autumn across varying developed land cover levels in two urban areas in Central Illinois. Mosquito abundance was assessed by the number of mosquitoes captured per trap night and compared across seasons and developed levels. The mean mosquito abundance for gravid and light traps was 12.96 ± 2.15 and 7.67 ± 1.44, respectively. Notably, higher levels of developed land cover exhibited higher Culex abundance than the low level for light traps, but no significant difference was found between summer and early autumn. In gravid traps, no significant differences were detected across seasons or developed levels. Microbial analysis of the mosquito microbiome revealed that Proteobacteria and Wolbachia, with a mean relative abundance of 80.77 and 52.66% respectively, were identified as the most dominant bacterial phylum and genus. Their relative abundance remained consistent across seasons and developed land cover levels, with negligible variations. Alpha diversity, as measured by observed species, Chao1, Shannon, and Simpson, showed slightly higher values in early-autumn compared to late-summer. A notable pattern of bacterial diversity, as indicated by all four diversity indexes, is evident across varying levels of land development. Significantly, high or intermediate developed levels consistently showed reduced alpha diversity when compared to the lower level. This underscores the pronounced impact of anthropogenic ecological disturbances in shaping mosquito microbiomes. Beta diversity analysis revealed no significant dissimilarities in bacterial community composition across seasons and developed levels, although some separation was noted among different levels of developed land cover. These findings highlight the significant role of environmental factors in shaping mosquito abundance and their associated microbiomes, with potential implications for the vectorial capacity in the transmission of vector-borne diseases.

Aedes aegypti gut transcriptomes respond differently to microbiome transplants from field-caught or laboratory-reared mosquitoes.

The mosquito microbiome is critical for host development and plays a major role in many aspects of mosquito biology. While the microbiome is commonly dominated by a small number of genera, there is considerable variation in composition among mosquito species, life stages, and geography. How the host controls and is affected by this variation is unclear. Using microbiome transplant experiments, we asked whether there were differences in transcriptional responses when mosquitoes of different species were used as microbiome donors. We used microbiomes from four different donor species spanning the phylogenetic breadth of the Culicidae, collected either from the laboratory or the field. We found that when recipients received a microbiome from a donor reared in the laboratory, the response was remarkably similar regardless of donor species. However, when the donor had been collected from the field, many more genes were differentially expressed. We also found that while the transplant procedure did have some effect on the host transcriptome, this is likely to have had a limited effect on mosquito fitness. Overall, our results highlight the possibility that variation in mosquito microbiome communities is associated with variability in host-microbiome interactions and further demonstrate the utility of the microbiome transplantation technique for investigating host-microbe interactions in mosquitoes.

A tangled threesome: understanding arbovirus infection in Aedes spp. and the effect of the mosquito microbiota.

Arboviral infections transmitted by Aedes spp. mosquitoes are a major threat to human health, particularly in tropical regions but are expanding to temperate regions. The ability of Aedes aegypti and Aedes albopictus to transmit multiple arboviruses involves a complex relationship between mosquitoes and the virus, with recent discoveries shedding light on it. Furthermore, this relationship is not solely between mosquitoes and arboviruses, but also involves the mosquito microbiome. Here, we aimed to construct a comprehensive review of the latest information about the arbovirus infection process in A. aegypti and A. albopictus, the source of mosquito microbiota, and its interaction with the arbovirus infection process, in terms of its implications for vectorial competence. First, we summarized studies showing a new mechanism for arbovirus infection at the cellular level, recently described innate immunological pathways, and the mechanism of adaptive response in mosquitoes. Second, we addressed the general sources of the Aedes mosquito microbiota (bacteria, fungi, and viruses) during their life cycle, and the geographical reports of the most common microbiota in adults mosquitoes. How the microbiota interacts directly or indirectly with arbovirus transmission, thereby modifying vectorial competence. We highlight the complexity of this tripartite relationship, influenced by intrinsic and extrinsic conditions at different geographical scales, with many gaps to fill and promising directions for developing strategies to control arbovirus transmission and to gain a better understanding of vectorial competence. The interactions between mosquitoes, arboviruses and their associated microbiota are yet to be investigated in depth.

Wolbachia dominance influences the Culex quinquefasciatus microbiota

Microorganisms present in mosquitoes and their interactions are key factors affecting insect development. Among them, Wolbachia is closely associated with the host and affects several fitness parameters. In this study, the bacterial and fungal microbiota from two laboratory Culex quinquefasciatus isolines (wild type and tetracycline-cured) were characterized by metagenome amplicon sequencing of the ITS2 and 16S rRNA genes at different developmental stages and feeding conditions. We identified 572 bacterial and 61 fungal OTUs. Both isolines presented variable bacterial communities and different trends in the distribution of diversity among the groups. The lowest bacterial richness was detected in sugar-fed adults of the cured isoline, whereas fungal richness was highly reduced in blood-fed mosquitoes. Beta diversity analysis indicated that isolines are an important factor in the differentiation of mosquito bacterial communities. Considering composition, Penicillium was the dominant fungal genus, whereas Wolbachia dominance was inversely related to that of Enterobacteria (mainly Thorsellia and Serratia). This study provides a more complete overview of the mosquito microbiome, emphasizing specific highly abundant components that should be considered in microorganism manipulation approaches to control vector-borne diseases.

Identification and characterization of two CRISPR/Cas systems associated with the mosquito microbiome.

The microbiome profoundly influences many traits in medically relevant vectors such as mosquitoes, and a greater functional understanding of host-microbe interactions may be exploited for novel microbial-based approaches to control mosquito-borne disease. Here, we characterized two novel clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems in Serratia sp. Ag1, which was isolated from the gut of an Anopheles gambiae mosquito. Two distinct CRISPR/Cas systems were identified in Serratia Ag1, CRISPR1 and CRISPR2. Based on cas gene composition, CRISPR1 is classified as a type I-E CRISPR/Cas system and has a single array, CRISPR1. CRISPR2 is a type I-F system with two arrays, CRISPR2.1 and CRISPR2.2. RT-PCR analyses show that all cas genes from both systems are expressed during logarithmic growth in culture media. The direct repeat sequences of CRISPRs 2.1 and 2.2 are identical and found in the arrays of other Serratia spp., including S. marcescens and S. fonticola , whereas CRISPR1 is not. We searched for potential spacer targets and revealed an interesting difference between the two systems: only 9 % of CRISPR1 (type I-E) targets are in phage sequences and 91 % are in plasmid sequences. Conversely, ~66 % of CRISPR2 (type I-F) targets are found within phage genomes. Our results highlight the presence of CRISPR loci in gut-associated bacteria of mosquitoes and indicate interplay between symbionts and invasive mobile genetic elements over evolutionary time.

Microplastic ingestion perturbs the microbiome of Aedes albopictus (Diptera: Culicidae) and Aedes aegypti.

Microplastics (MPs) are common environmental pollutants; however, little is known about their effects after ingestion by insects. Here we fed Aedes (Stegomyia) aegypti (L.) and Aedes (Stegomyia) albopictus (Skuse) mosquito larvae 1 µm polystyrene MPs and examined the impacts of ingestion on adult emergence rates, gut damage, and fungal and bacterial microbiota. Results show that MPs accumulate in the larval guts, resulting in gut damage. However, little impact on adult emergence rates was observed. MPs are also found in adult guts postemergence from the pupal stage, and adults expel MPs in their frass after obtaining sugar meals. Moreover, MPs effects on insect microbiomes need to be better defined. To address this knowledge gap, we investigated the relationship between MP ingestion and the microbial communities in Ae. albopictus and Ae. aegypti. The microbiota composition was altered by the ingestion of increasing concentrations of MPs. Amplicon sequence variants (ASVs) that contributed to differences in the bacterial and fungal microbiota composition between MP treatments were from the genera Elizabethkingia and Aspergillus, respectively. Furthermore, a decrease in the alpha diversity of the fungal and bacterial microbiota was observed in treatments where larvae ingested MPs. These results highlight the potential for the bacterial and fungal constituents in the mosquito microbiome to respond differently to the ingestion of MPs. Based on our findings and the effects of MP ingestion on the mosquito host micro- and mycobiome, MP pollution could impact the vector competence of important mosquito-transmitted viruses and parasites that cause human and animal diseases.

The relationships between microbiome diversity and epidemiology in domestic species of malaria-mediated mosquitoes of Korea

Microbiota in the mosquito plays an important role in their behavior and vector competence. The composition of their microbiome is strongly influenced by the environment, especially their habitat. The microbiome profiles of adult female Anopheles sinensis mosquitoes from malaria hyperendemic and hypoendemic areas in Republic of Korea were compared using 16S rRNA Illumina sequencing. In different epidemiology groups, the alpha and beta diversity analyses were significant. The major bacterial phylum was Proteobacteria. The most abundant species in the microbiome of hyperendemic mosquitoes were the genera Staphylococcus, Erwinia, Serratia, and Pantoea. Notably, a distinct microbiome profile characterized by the dominance of Pseudomonas synxantha was identified in the hypoendemic area, suggesting a potential correlation between the microbiome profiles and the incidence of malaria cases.

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 Mānoa 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.

Recovery of metagenomic data from the Aedes aegypti microbiome using a reproducible snakemake pipeline: MINUUR.

Background: Ongoing research of the mosquito microbiome aims to uncover novel strategies to reduce pathogen transmission. Sequencing costs, especially for metagenomics, are however still significant. A resource that is increasingly used to gain insights into host-associated microbiomes is the large amount of publicly available genomic data based on whole organisms like mosquitoes, which includes sequencing reads of the host-associated microbes and provides the opportunity to gain additional value from these initially host-focused sequencing projects. Methods: To analyse non-host reads from existing genomic data, we developed a snakemake workflow called MINUUR (Microbial INsights Using Unmapped Reads). Within MINUUR, reads derived from the host-associated microbiome were extracted and characterised using taxonomic classifications and metagenome assembly followed by binning and quality assessment. We applied this pipeline to five publicly available Aedes aegypti genomic datasets, consisting of 62 samples with a broad range of sequencing depths. Results: We demonstrate that MINUUR recovers previously identified phyla and genera and is able to extract bacterial metagenome assembled genomes (MAGs) associated to the microbiome. Of these MAGS, 42 are high-quality representatives with >90% completeness and <5% contamination. These MAGs improve the genomic representation of the mosquito microbiome and can be used to facilitate genomic investigation of key genes of interest. Furthermore, we show that samples with a high number of KRAKEN2 assigned reads produce more MAGs. Conclusions: Our metagenomics workflow, MINUUR, was applied to a range of Aedes aegypti genomic samples to characterise microbiome-associated reads. We confirm the presence of key mosquito-associated symbionts that have previously been identified in other studies and recovered high-quality bacterial MAGs. In addition, MINUUR and its associated documentation are freely available on GitHub and provide researchers with a convenient workflow to investigate microbiome data included in the sequencing data for any applicable host genome of interest.