Human-aided dispersal and population bottlenecks facilitate parasitism escape in the most invasive mosquito species.

During biological invasion process, species encounter new environments and partially escape some ecological constraints they faced in their native range, while they face new ones. The Asian tiger mosquito Aedes albopictus is one of the most iconic invasive species introduced in every inhabited continent due to international trade. It has also been shown to be infected by a prevalent yet disregarded microbial entomoparasite Ascogregarina taiwanensis. In this study, we aimed at deciphering the factors that shape the global dynamics of A. taiwanensis infection in natural A. albopictus populations. We showed that A. albopictus populations are highly colonized by several parasite genotypes but recently introduced ones are escaping it. We further performed experiments based on the invasion process to explain such pattern. To that end, we hypothesized that (i) mosquito passive dispersal (i.e. human-aided egg transportation) may affect the parasite infectiveness, (ii) founder effects (i.e. population establishment by a small number of mosquitoes) may influence the parasite dynamics, and (iii) unparasitized mosquitoes are more prompt to found new populations through active flight dispersal. The two first hypotheses were supported as we showed that parasite infection decreases over time when dry eggs are stored and that experimental increase in mosquitoes' density improves the parasite horizontal transmission to larvae. Surprisingly, parasitized mosquitoes tend to be more active than their unparasitized relatives. Finally, this study highlights the importance of global trade as a driver of biological invasion of the most invasive arthropod vector species.

Microbiome analysis in Lascaux Cave in relation to black stain alterations of rock surfaces and collembola.

Anthropization of Palaeolithic caves open for tourism may favour collembola invasion and result in the formation of black stains attributed to pigmented fungi. However, ecological processes underpinning black stain formation are not fully understood. Here, we tested the hypotheses that black stains from the Apse room of Lascaux Cave display a specific microbiota enriched in pigmented fungi, and that collembola thriving on the stains have the potential to consume and disseminate these black fungi. Metabarcoding showed that the microbiota of black stains and neighbouring unstained parts strongly differed, with in black stains a higher prevalence of Ochroconis and other pigmented fungi and the strong regression of Pseudomonas bacteria (whose isolates inhibited in vitro the growth of pigmented fungi). Isotopic analyses indicated that Folsomia candida collembola thriving on stains could feed on black stain in situ and assimilate the pigmented fungi they were fed with in vitro. They could carry these fungi and disseminate them when tested with complex black stains from Lascaux. This shows that black stain formation is linked to the development of pigmented fungi, which coincides with the elimination of antagonistic pseudomonads, and points towards a key role of F. candida collembola in the dynamics of pigmented fungi.

Crosstalk between the microbiota and insect postembryonic development.

Insect sequential development evolves from a simple molt towards complete metamorphosis. Like any multicellular host, insects interact with a complex microbiota. In this review, factors driving the microbiota dynamics were pointed out along their development. Special focus was put on tissue renewal, shift in insect ecology, and microbial interactions. Conversely, how the microbiota modulates its host development through nutrient acquisition, hormonal control, and cellular or tissue differentiation was exemplified. Such modifications might have long-term carry-over effects on insect physiology. Finally, remarkable microbe-driven control of insect behaviors along their life cycle was highlighted. Increasing knowledge of those interactions might offer new insights on how insects respond to their environment as well as perspectives on pest- or vector-control strategies.

Microbiome Analysis of New, Insidious Cave Wall Alterations in the Apse of Lascaux Cave.

Lascaux Cave is a UNESCO site that was closed to the public following wall surface alterations. Most black stains that had formed on wall surface are stable or receding, but a new type of alteration visually quite different (termed dark zones) developed in Lascaux's Apse room in the last 15 years. Here, we tested the hypothesis that dark zones displayed a different microbial community than black stains previously documented in the same room, using metabarcoding (MiSeq sequencing). Indeed, dark zones, black stains and neighboring unstained parts displayed distinct microbial communities. However, similarly to what was observed in black stains, pigmented fungi such as Ochroconis (now Scolecobasidium) were more abundant and the bacteria Pseudomonas less abundant in dark zones than in unstained parts. The collembola Folsomia candida, which can disseminate microorganisms involved in black stain development, was also present on dark zones. Illumina sequencing evidenced Ochroconis (Scolecobasidium) in all collembola samples from dark zones, as in collembola from black stains. This study shows that the microbial properties of dark zones are peculiar, yet dark zones display a number of microbial resemblances with black stains, which suggests a possible role of collembola in promoting these two types of microbial alterations on wall surfaces.

Gregarine parasites are adapted to mosquito winter diapause.

The Asian tiger mosquito Aedes albopictus is one of the most invasive species of mosquito. The prevalence of its apicomplexan gregarine parasite Ascogregarina taiwanensis is high in natural populations across both temperate and tropical regions. However, the parasite's oocysts cannot colonize the insect host during winter, when the mosquito lays diapausing eggs. It is therefore unclear if the parasite can survive outside of its insect host during the cold season in temperate regions. Oocysts stored for 1 month at a low temperature (representative of the temperatures that occur during periods of mosquito diapause) were as infectious as fresh oocysts, but those stored for the same period of time at a higher temperature (representative of the temperatures that occur during periods of mosquito activity) were uninfectious. We therefore suggest that the parasite has evolved traits that maximize its maintenance during periods of mosquito dormancy, while traits that would enable its  long term survival during periods of mosquito activity have not been selected for.

Reciprocal interactions between anthropogenic stressors and insect microbiota.

Insects play many important roles in nature due to their diversity, ecological role, and impact on agriculture or human health. They are directly influenced by environmental changes and in particular anthropic activities that constitute an important driver of change in the environmental characteristics. Insects face numerous anthropogenic stressors and have evolved various detoxication mechanisms to survive and/or resist to these compounds. Recent studies highligted the pressure exerted by xenobiotics on insect life-cycle and the important role of insect-associated bacterial microbiota in the insect responses to environmental changes. Stressor exposure can have various impacts on the composition and structure of insect microbiota that in turn may influence insect biology. Moreover, bacterial communities associated with insects can be directly or indirectly involved in detoxification processes with the selection of certain microorganisms capable of degrading xenobiotics. Further studies are needed to assess the role of insect-associated microbiota as key contributor to the xenobiotic metabolism and thus as a driver for insect adaptation to polluted habitats.

Larval habitat determines the bacterial and fungal microbiota of the mosquito vector Aedes aegypti.

Mosquito larvae are naturally exposed to microbial communities present in a variety of larval development sites. Several earlier studies have highlighted that the larval habitat influences the composition of the larval bacterial microbiota. However, little information is available on their fungal microbiota, i.e. the mycobiota. In this study, we provide the first simultaneous characterization of the bacterial and fungal microbiota in field-collected Aedes aegypti larvae and their respective aquatic habitats. We evaluated whether the microbial communities associated with the breeding site may affect the composition of both the bacterial and fungal communities in Ae. aegypti larvae. Our results show a higher similarity in microbial community structure for both bacteria and fungi between larvae and the water in which larvae develop than between larvae from different breeding sites. This supports the hypothesis that larval habitat is a major factor driving microbial composition in mosquito larvae. Since the microbiota plays an important role in mosquito biology, unravelling the network of interactions that operate between bacteria and fungi is essential to better understand the functioning of the mosquito holobiont.

Alternative developmental and transcriptomic responses to host plant water limitation in a butterfly metapopulation.

Predicting how climate change affects biotic interactions poses a challenge. Plant-insect herbivore interactions are particularly sensitive to climate change, as climate-induced changes in plant quality cascade into the performance of insect herbivores. Whereas the immediate survival of herbivore individuals depends on plastic responses to climate change-induced nutritional stress, long-term population persistence via evolutionary adaptation requires genetic variation for these responses. To assess the prospects for population persistence under climate change, it is therefore crucial to characterize response mechanisms to climate change-induced stressors, and quantify their variability in natural populations. Here, we test developmental and transcriptomic responses to water limitation-induced host plant quality change in a Glanville fritillary butterfly (Melitaea cinxia) metapopulation. We combine nuclear magnetic resonance spectroscopy on the plant metabolome, larval developmental assays and an RNA sequencing analysis of the larval transcriptome. We observed that responses to feeding on water-limited plants, in which amino acids and aromatic compounds are enriched, showed marked variation within the metapopulation, with individuals of some families performing better on control and others on water-limited plants. The transcriptomic responses were concordant with the developmental responses: families exhibiting opposite developmental responses also produced opposite transcriptomic responses (e.g. in growth-associated transcripts). The divergent responses in both larval development and transcriptome are associated with differences between families in amino acid catabolism and storage protein production. The results reveal intrapopulation variability in plasticity, suggesting that the Finnish M. cinxia metapopulation harbours potential for buffering against drought-induced changes in host plant quality.

Complex plant quality-microbiota-population interactions modulate the response of a specialist herbivore to the defence of its host plant.

Many specialist herbivores have evolved strategies to cope with plant defences, with gut microbiota potentially participating to such adaptations.In this study, we assessed whether the history of plant use (population origin) and microbiota may interact with plant defence adaptation.We tested whether microbiota enhance the performance of Melitaea cinxia larvae on their host plant, Plantago lanceolata and increase their ability to cope the defensive compounds, iridoid glycosides (IGs).The gut microbiota were significantly affected by both larval population origin and host plant IG level. Contrary to our prediction, impoverishing the microbiota with antibiotic treatment did not reduce larval performance.As expected for this specialized insect herbivore, sequestration of one of IGs was higher in larvae fed with plants producing higher concentration of IGs. These larvae also showed metabolic signature of intoxication (i.e. decrease in Lysine levels). However, intoxication on highly defended plants was only observed when larvae with a history of poorly defended plants were simultaneously treated with antibiotics.Our results suggest that both adaptation and microbiota contribute to the metabolic response of herbivores to plant defence though complex interactions. Read the free Plain Language Summary for this article on the Journal blog.

De nombreux herbivores spécialistes ont évolué vers des stratégies qui leurs permettent de contourner les défenses de leur plantes hôtes. Le microbiote pourrait potentiellement participer à certaines de ces adaptations.Dans cette étude, nous avons essayé de déterminer si l'adaptation d'un herbivore est influencée par son microbiote et l'historique d'utilisation de sa plante hôte (origine de la population).Nous avons testé en quoi le microbiote contribue à la performance de chenilles Melitaea cinxia sur leur plante hôte Plantago lanceolata ainsi que leur capacité à faire face aux glucosides d'iridoïdes (GI), des molécules de défenses produites par P. lanceolata.Comme attendu, la concentration de GI stockée était plus importante chez les chenilles qui étaient nourries avec des plantes produisant de fortes concentrations de GI. Ces chenilles présentaient par ailleurs des signes d'intoxications (i.e. diminution de la concentration de Lysine). Cependant cela n'était visible que lorsque les chenilles étaient issues de populations qui se nourrissaient historiquement sur des plantes peu défendues et lorsqu'elles étaient simultanément traitées par des antibiotiques.Nos résultats suggèrent donc que des processus complexes d'adaptation couplés à l'activité du microbiote contribuent à la réponse des herbivores aux défenses de leurs plante hôtes.

Diversity and Functions of Yeast Communities Associated with Insects.

Following the concept of the holobiont, insect-microbiota interactions play an important role in insect biology. Many examples of host-associated microorganisms have been reported to drastically influence insect biological processes such as development, physiology, nutrition, survival, immunity, or even vector competence. While a huge number of studies on insect-associated microbiota have focused on bacteria, other microbial partners including fungi have been comparatively neglected. Yeasts, which establish mostly commensal or symbiotic relationships with their host, can dominate the mycobiota of certain insects. This review presents key advances and progress in the research field highlighting the diversity of yeast communities associated with insects, as well as their impact on insect life-history traits, immunity, and behavior.

Microorganisms Associated with Mosquito Oviposition Sites: Implications for Habitat Selection and Insect Life Histories.

Mosquitoes are considered one of the most important threats worldwide due to their ability to vector pathogens. They are responsible for the transmission of major pathogens such as malaria, dengue, zika, or chikungunya. Due to the lack of treatments or prophylaxis against many of the transmitted pathogens and an increasing prevalence of mosquito resistance to insecticides and drugs available, alternative strategies are now being explored. Some of these involve the use of microorganisms as promising agent to limit the fitness of mosquitoes, attract or repel them, and decrease the replication and transmission of pathogenic agents. In recent years, the importance of microorganisms colonizing the habitat of mosquitoes has particularly been investigated since they appeared to play major roles in their development and diseases transmission. In this issue, we will synthesize researches investigating how microorganisms present within water habitats may influence breeding site selection and oviposition strategies of gravid mosquito females. We will also highlight the impact of such microbes on the fate of females' progeny during their immature stages with a specific focus on egg hatching, development rate, and larvae or pupae survival.

Considerations for mosquito microbiome research from the Mosquito Microbiome Consortium.

In the past decade, there has been increasing interest in mosquito microbiome research, leading to large amounts of data on different mosquito species, with various underlying physiological characteristics, and from diverse geographical locations. However, guidelines and standardized methods for conducting mosquito microbiome research are lacking. To streamline methods in mosquito microbiome research and optimize data quality, reproducibility, and comparability, as well as facilitate data curation in a centralized location, we are establishing the Mosquito Microbiome Consortium, a collaborative initiative for the advancement of mosquito microbiome research. Our overall goal is to collectively work on unraveling the role of the mosquito microbiome in mosquito biology, while critically evaluating its potential for mosquito-borne disease control. This perspective serves to introduce the consortium and invite broader participation. It highlights the issues we view as most pressing to the community and proposes guidelines for conducting mosquito microbiome research. We focus on four broad areas in this piece: (1) sampling/experimental design for field, semi-field, or laboratory studies; (2) metadata collection; (3) sample processing, sequencing, and use of appropriate controls; and (4) data handling and analysis. We finally summarize current challenges and highlight future directions in mosquito microbiome research. We hope that this piece will spark discussions around this area of disease vector biology, as well as encourage careful considerations in the design and implementation of mosquito microbiome research. Video Abstract.

The gut bacterial community affects immunity but not metabolism in a specialist herbivorous butterfly.

Plant tissues often lack essential nutritive elements and may contain a range of secondary toxic compounds. As nutritional imbalance in food intake may affect the performances of herbivores, the latter have evolved a variety of physiological mechanisms to cope with the challenges of digesting their plant-based diet. Some of these strategies involve living in association with symbiotic microbes that promote the digestion and detoxification of plant compounds or supply their host with essential nutrients missing from the plant diet. In Lepidoptera, a growing body of evidence has, however, recently challenged the idea that herbivores are nutritionally dependent on their gut microbial community. It is suggested that many of the herbivorous Lepidopteran species may not host a resident microbial community, but rather a transient one, acquired from their environment and diet. Studies directly testing these hypotheses are however scarce and come from an even more limited number of species.By coupling comparative metabarcoding, immune gene expression, and metabolomics analyses with experimental manipulation of the gut microbial community of prediapause larvae of the Glanville fritillary butterfly (Melitaea cinxia, L.), we tested whether the gut microbial community supports early larval growth and survival, or modulates metabolism or immunity during early stages of development.We successfully altered this microbiota through antibiotic treatments and consecutively restored it through fecal transplants from conspecifics. Our study suggests that although the microbiota is involved in the up-regulation of an antimicrobial peptide, it did not affect the life history traits or the metabolism of early instars larvae.This study confirms the poor impact of the microbiota on diverse life history traits of yet another Lepidoptera species. However, it also suggests that potential eco-evolutionary host-symbiont strategies that take place in the gut of herbivorous butterfly hosts might have been disregarded, particularly how the microbiota may affect the host immune system homeostasis.

Who is eating fructose within the Aedes albopictus gut microbiota?

The Asian tiger mosquito Aedes albopictus is a major public health concern because of its invasive success and its ability to transmit pathogens. Given the low availability of treatments against mosquito-borne diseases, vector control remains the most suitable strategy. The methods used thus far are becoming less effective, but recent strategies have emerged from the study of mosquito-associated microorganisms. Although the role of the microbiota in insect biology does not require further proof, much remains to be deciphered in mosquitoes, especially the contribution of the microbiota to host nutrient metabolism. Mosquitoes feed on plant nectar, composed of mostly fructose. We used stable isotope probing to identify bacteria and fungi assimilating fructose within the gut of Ae. albopictus. Mosquitoes were fed a 13 C-labelled fructose solution for 24 h. Differences in the active microbial community according to the sex of mosquitoes were highlighted. The bacterium Lelliottia and the fungi Cladosporium and Aspergillus dominated the active microbiota in males, whereas the bacterium Ampullimonas and the yeast Cyberlindnera were the most active in females. This study is the first to investigate trophic interactions between Ae. albopictus and its microbiota, thus underscoring the importance of the microbial component in nectar feeding in mosquitoes.

The microbiome of the Melitaea cinxia butterfly shows marked variation but is only little explained by the traits of the butterfly or its host plant.

Understanding of the ecological factors that shape intraspecific variation of insect microbiota in natural populations is relatively poor. In Lepidopteran caterpillars, microbiota is assumed to be mainly composed of transient bacterial symbionts acquired from the host plant. We sampled Glanville fritillary (Melitaea cinxia) caterpillars from natural populations to describe their gut microbiome and to identify potential ecological factors that determine its structure. Our results demonstrate high variability of microbiota composition even among caterpillars that shared the same host plant individual and most likely the same genetic background. We observed that the caterpillars harboured microbial classes that varied among individuals and alternated between two distinct communities (one composed of mainly Enterobacteriaceae and another with more variable microbiota community). Even though the general structure of the microbiota was not attributed to the measured ecological factors, we found that phylogenetically similar microbiota showed corresponding responses to the sex and the parasitoid infection of the caterpillar and to those of the host plant's microbial and chemical composition. Our results indicate high among-individual variability in the microbiota of the M. cinxia caterpillar and contradict previous findings that the host plant is the major driver of the microbiota communities of insect herbivores.

Identification of human skin bacteria attractive to the Asian Tiger mosquito.

Aedes albopictus is a vector of arboviruses and filarial nematodes. Originating from Asia, this mosquito has rapidly expanded its geographical distribution and colonized areas across both temperate and tropical regions. Due to the increase in insecticide resistance, the use of environmentally friendly vector control methods is encouraged worldwide. Using methods based on semiochemicals in baited traps are promising for management of mosquito populations. Interestingly, human skin microbiota was shown to generate volatile compounds that attract the mosquito species Anopheles gambiae and Aedes aegypti. Here, we investigated the composition of skin bacteria from different volunteers and the attractive potential of individual isolates to nulliparous Ae. albopictus females. We showed that three out of 16 tested isolates were more attractive and two were more repulsive. We identified dodecenol as being preferentially produced by attractive isolates and 2-methyl-1-butanol (and to a lesser extent 3-methyl-1-butanol) as being overproduced by these isolates compared with the other ones. Those bacterial volatile organic compounds represent promising candidates but further studies are needed to evaluate their potential application for baited traps improvement.

Moderate plant water stress improves larval development, and impacts immunity and gut microbiota of a specialist herbivore.

While host plant drought is generally viewed as a negative phenomenon, its impact on insect herbivores can vary largely depending on the species involved and on the intensity of the drought. Extreme drought killing host plants can clearly reduce herbivore fitness, but the impact of moderate host plant water stress on insect herbivores can vary, and may even be beneficial. The populations of the Finnish Glanville fritillary butterfly (Melitaea cinxia) have faced reduced precipitation in recent years, with impacts even on population dynamics. Whether the negative effects of low precipitation are solely due to extreme desiccation killing the host plant or whether moderate drought reduces plant quality for the larvae remains unknown. We assessed the performance of larvae fed on moderately water-stressed Plantago lanceolata in terms of growth, survival, and immune response, and additionally were interested to assess whether the gut microbial composition of the larvae changed due to modification of the host plant. We found that larvae fed on water-stressed plants had increased growth, with no impact on survival, up-regulated the expression of one candidate immune gene (pelle), and had a more heterogeneous bacterial community and a shifted fungal community in the gut. Most of the measured traits showed considerable variation due to family structure. Our data suggest that in temperate regions moderate host plant water stress can positively shape resource acquisition of this specialized insect herbivore, potentially by increasing nutrient accessibility or concentration. Potentially, the better larval performance may be mediated by a shift of the microbiota on water-stressed plants, calling for further research especially on the understudied gut fungal community.

Short-term impacts of anthropogenic stressors on Aedes albopictus mosquito vector microbiota.

Recent studies have highlighted the potential role of microbiota in the biology of the Aedes albopictus mosquito vector. This species is highly anthropogenic and exhibits marked ecological plasticity, with a resulting high potential to colonize a wide range of habitats-including anthropized areas-under various climatic conditions. We put forward the hypothesis that climate and anthropogenic activities, such as the use of antibiotics in agriculture and human medicine, might affect the mosquito-associated bacterial community. We thus studied the additive impact of a temperature decrease and antibiotic ingestion on the temporal dynamics of Ae. albopictus survival and its associated bacterial communities. The results showed no effects of disturbances on mosquito survival. However, short-term temperature impacts on bacterial diversity were observed, while both the community structure and bacterial diversity were affected by early antibiotic ingestion. The genera Elizabethkingia, Chryseobacterium and Wolbachia, as well as an unclassified member of the Bacteroidales order were particularly affected. Antibiotics negatively impacted Elizabethkingia abundance, while Chryseobacterium was completely eliminated following both disturbances, to the benefit of Wolbachia and the unclassified Bacteroidales species. These results generated fresh insight into the effects of climate and anthropogenic activities such as the use of antibiotics on mosquito microbiota.

Shared larval rearing environment, sex, female size and genetic diversity shape Ae. albopictus bacterial microbiota.

The Asian tiger mosquito Aedes albopictus became of public health concern as it can replicate and transmit viral and filarial pathogens with a strong invasive success over the world. Various strategies have been proposed to reduce mosquito population's vectorial capacity. Among them, symbiotic control of mosquito borne disease offers promising perspectives. Such method is likely to be affected by the dynamics of mosquito-associated symbiotic communities, which might in turn be affected by host genotype and environment. Our previous study suggested a correlation between mosquitoes' origin, genetic diversity and midgut bacterial diversity. To distinguish the impact of those factors, we have been studying the midgut bacterial microbiota of two Ae. albopictus populations from tropical (La Réunion) and temperate (Montpellier) origins under controlled laboratory conditions. the two populations experienced random mating or genetic bottleneck. Microbiota composition did not highlight any variation of the α and ß-diversities in bacterial communities related to host's populations. However, sizes of the mosquitoes were negatively correlated with the bacterial α-diversity of females. Variations in mosquito sex were associated with a shift in the composition of bacterial microbiota. The females' mosquitoes also exhibited changes in the microbiota composition according to their size and after experiencing a reduction of their genetic diversity. These results provide a framework to investigate the impact of population dynamics on the symbiotic communities associated with the tiger mosquito.