Biology subjects

Novakova, E.

Publications and source records attributed to Novakova, E..

At least 19 recordsLinked to original sources

bioRxiv2026

Cerebral organoids recapitulate interneuron chain migration in the human cortex

The human brain is incomplete at birth, with streams of late-born interneurons migrating into the postnatal cortex. This process is absent in rodents, limiting mechanistic investigation to post-mortem tissue. Here, we show that human cerebral organoids cultured for over six months recapitulate postnatal interneuron migration, enabling its mechanistic and genetic dissection. Live imaging and mathematical modeling revealed that interneurons intrinsically migrate in chains, while astrocyte interactions promote stream organization. Spatial and single-cell transcriptomics identified a distinct molecular signature of human postnatal migratory streams enriched for neurodevelopmental disorder-associated genes. Perturbation of the autism-associated genes CNTN5 and FGF14 differentially disrupted chain organization and migration. Thus, long-term cerebral organoids enable mechanistic analysis of previously inaccessible aspects of postnatal human brain development and their contributions to neurodevelopmental disease.

developmental biology↗

bioRxiv2026

Caught in transition: facultative intracellularity and genome evolution of Symbiopectobacterium in Rhodnius species

Blood-feeding insects typically depend on obligate intracellular bacterial symbionts that provide essential B vitamins absent from vertebrate blood. In contrast, kissing bugs (Triatominae) have long been considered atypical because they rely primarily on extracellular gut-associated bacteria. Recent reports of the genus Symbiopectobacterium in Rhodnius species raise questions about the diversity and evolution of symbiosis in these insects. Here, we investigate the distribution, genome evolution, and tissue localization of Symbiopectobacterium in the genus Rhodnius. Using comparative metagenomics, phylogenomics, fluorescence in situ hybridization, transmission electron microscopy, and hemolymph screening, we characterize a Symbiopectobacterium genome from Rhodnius prolixus and assess its occurrence across publicly available datasets representing multiple Rhodnius species. The R. prolixus strain possesses a large, highly dynamic genome enriched in mobile genetic elements, pseudogenes, and remnants of secretion systems, while retaining biosynthetic pathways for several B vitamins. Comparative analysis revealed variation in genome reduction among Rhodnius-associated strains, suggesting ongoing and potentially independent transitions toward host-restricted symbiosis. Localization analyses detected Symbiopectobacterium intracellularly within posterior midgut epithelial cells and occasionally in the hemolymph, consistent with a facultative intracellular lifestyle. However, no bacteriomes or stable intracellular structures were observed. Together, these findings indicate that Symbiopectobacterium represents an intermediate stage in the transition from environmentally associated bacteria to obligate intracellular mutualists in Triatominae.

microbiology↗

bioRxiv2026

Highly contiguous genomes of Rhodnius prolixus and Triatoma rubida reveal the molecular basis of haematophagy evolution in Triatominae

Insects of the subfamily Triatominae, commonly known as kissing bugs, are obligate blood-feeding vectors of Trypanosoma cruzi, the causative agent of Chagas disease. Rhodnius prolixus is among the most epidemiologically important vectors in Latin America, whereas Triatoma rubida frequently invades homes and is a potential vector in the southern United States and northern Mexico. Triatomines likely evolved from predatory reduviid assassin bugs through a transition from feeding on arthropods associated with vertebrate hosts to feeding directly on vertebrate blood. To investigate the genomic basis of this ecological and dietary shift, we generated highly contiguous, near chromosome-level genome assemblies and structural gene annotations for R. prolixus and T. rubida. The new R. prolixus assembly improves scaffold N50 more than 40-fold over the current reference genome, from 1.1 to 43.9 Mb, while reducing assembly gaps by several orders of magnitude. Both assemblies exceed 97% BUSCO completeness. Comparative analyses with representative hemipteran genomes revealed expansions of gene families associated with chemosensation and metabolism, including detoxification, protein degradation, and digestion, together with signatures of positive selection in genes involved in digestive and sensory functions. These assemblies represent the most contiguous and complete genomic resources available for Triatominae and provide a robust foundation for investigating vector biology, host adaptation, and the evolutionary origins of blood feeding within Reduviidae. Interpretive summaryKissing bugs are insects that are known for feeding on blood. They can spread a disease called Chagas disease because they transmit a parasite called Trypanosoma cruzi. To understand how kissing bugs evolved and which genes facilitate blood feeding of vertebrates, a collaboration between scientists at USDA-ARS, University of Georgia, and University of South Bohemia sequenced the genome of two kissing bugs: Rhodnius prolixus and Triatoma rubida. By comparing the genes with those of other insects in the order Hemiptera, scientists discovered that kissing bugs have more genes involved with detecting environmental chemical stimuli and metabolism as well as positive selection for genes involved with digestion and sensory-related proteins. These genome assemblies will help scientists learn more about how these insects evolved, and this research is important for understanding insect feeding biology which can be used to develop methods to control the kissing bugs and the spread of Chagas disease.

evolutionary biology↗

bioRxiv2025

Generalist vs specialist strategy shapes microbiomes in blood feeding parasite Polyplax serrata.

Insects live in association with bacterial communities, collectively referred to as the microbiome. Microbiome composition varies widely across insect taxa and is shaped by multiple factors, including host phylogeny, environmental conditions, geographic distribution, and nutritional ecology. One hypothesis is that microbiome composition may also reflect whether the host adopts a generalist or specialist ecological strategy. We tested this hypothesis using the sucking louse Polyplax serrata, which offers several advantages as a model system. First, as permanent ectoparasites, lice inhabit a relatively stable and simplified environment, thereby minimizing potential confounding variables. Second, within P. serrata, two closely related lineages have been identified: one restricted to a single rodent host (Apodemus flavicollis), and the other exploiting two hosts (A. flavicollis and A. sylvaticus). We analyzed and compared microbiome structure in these two lineages using 16S rRNA gene amplicon sequencing. While alpha diversity did not differ between the lineages, beta diversity differed significantly, particularly in pairwise dissimilarities among individual samples. These results suggest that in P. serrata, host specialization strategy influences microbiome diversity, with the "generalist" lineage harboring more heterogeneous communities. This finding extends previous observations on ecological divergence between the two lineages, showing that closely related cryptic species with highly similar genomes, living sympatrically in the same environment, can rapidly evolve distinct life strategies that, in turn, shape both their genetic structure and their microbiomes.

evolutionary biology↗

bioRxiv2025

Viral metagenome characterization reveals species-specific virome profiles in Triatominae populations from the southern United States

Kissing bugs (Triatominae) are hematophagous insects and the principal vectors of Trypanosoma cruzi, the causative agent of Chagas disease. While their bacterial microbiomes have received considerable attention, the diversity of viruses associated with these insects remains poorly understood. To address this gap, we investigated the metavirome of five Triatominae species from the southern United States (Triatoma rubida, T. sanguisuga, T. gerstaeckeri, T. indictiva, and Hospesneotomae protracta), sampled in Texas, New Mexico, and Arizona. We sequenced gut and reproductive tissues from 23 field-collected specimens and assembled 41 viral operational taxonomic units (vOTUs), 40 of which are novel and together constitute 13 viral families, including Chuviridae, Arenaviridae, Orthomyxoviridae, Partitiviridae, Solemoviridae, Circoviridae, Rhabdoviridae, Microviridae, Xinmoviridae, Astroviridae, Narnaviridae, Tombusviridae, and the order Elliovirales. The vOTUs composition and abundance analysis examined variables including host species, sex, tissue type, blood meal, and T. cruzi infection status, showing that metavirome diversity varied significantly among host species. Our findings demonstrate a species-specific metavirome and the presence of virus taxa linked to insects, plants, and vertebrates, highlighting the complex ecological interactions between viruses and triatomines. This study uncovers a diverse and largely novel set of metaviromes within North American Triatominae, providing a foundation for future research on virus-vector interactions.

ecology↗

bioRxiv2025

Dynamic but constrained: Repeated acquisitions of nutritional symbionts in bed bugs (Heteroptera: Cimicidae) from a narrow taxonomic pool.

BackgroundBed bugs (Heteroptera: Cimicidae), like other strictly blood-feeding insects, harbor obligate bacterial symbionts that supply essential nutrients (especially B vitamins) lacking in their blood diet. The primary obligate symbionts are transovarially transmitted Wolbachia, notable for having horizontally acquired biotin operon. In addition to these symbionts with the confirmed nutritional role, bacteria from the genera Symbiopectobacterium and Tisiphia have also been found to be transovarially transmitted in bed bugs. Their significance to the host fitness remains unclear; they are considered facultative, and thus likely non-essential. However, this understanding of bed bugs symbiosis is based exclusively on studies of the human-associated common bed bug (Cimex lectularius) and few related Cimicinae species. Virtually nothing is known about the diversity, origins, or metabolic roles of symbionts across the more than hundred described bed bug species. ResultsUsing amplicon and metagenomic analyzes, we identified five different bacterial genera as potential bed bug symbionts: Wolbachia, Symbiopectobacterium, Sodalis, Serratia, and Tisiphia. The distribution of these bacteria among 13 bed bug species indicated at least 16 independent origins of symbiosis (some samples harboring multiple symbiont strains). A comparison of host and symbiont phylogenies suggested that some of these origins were followed by cospeciation. Not all bed bugs, however, harbored Wolbachia. In the subfamily Cacodminae, Symbiopectobacterium (previously known as facultative symbiont) was detected as the sole symbiont, suggesting its role as an essential, obligate symbiont. Analysis of 23 obtained genome drafts revealed considerable differences in their size and gene content, indicating that these bacteria have reached different stages in the evolution towards obligate symbiosis. As is common in symbiotic bacteria, the analyzed genomes have lost many biosynthetic capacities. A comparison of B-vitamin synthesis pathways showed that only two, riboflavin and lipoic acid, were preserved across all symbionts. ConclusionsAn overview of symbiosis across a broad phylogenetic span of bed bugs reveals that this insect family has undergone remarkably dynamic evolution, characterized by multiple independent symbiont acquisitions, episodes of cospeciations, and frequent co-occurrence of multiple symbionts within individual hosts. Interestingly, although these symbionts were acquired through multiple independent events, the majority belong to just three bacterial genera (Wolbachia, Symbiopectobacterium, and Sodalis), suggesting an unknown mechanisms underlying bed bug-symbiont specificity. Two aspects of this study warrant further investigation. First, the finding of Symbiopectobacterium as the sole obligate symbiont in Cacodminae suggests that expanding the taxonomic sampling may reveal an even more complex structure of symbiosis within bed bugs. Second, the inconsistencies and uncertainties in evaluating the functionality of the biotin synthesis indicate that further research will be necessary to better understand the evolution of this B vitamin pathway in symbiotic bacteria.

evolutionary biology↗

bioRxiv2025

Field Evidence of Trypanosoma cruzi Infection, Diverse Host Use and Invasion of Human Dwellings by the Chagas Disease Vector in Florida, USA

Background: Triatomine bugs (Hemiptera, Reduviidae, Triatominae) are blood-sucking vectors of the protozoan parasite, Trypanosoma cruzi, which causes Chagas disease, a significant source of human morbidity and mortality in the Americas. Autochthonous transmission of Chagas disease in the United States is considered rare, despite evidence of Triatoma species harboring T. cruzi, invading homes, and biting occupants. In the southeastern United States, Triatoma sanguisuga is considered common, yet its distribution, host use, and T. cruzi infection are practically unknown in this region. Methodology/Principle Findings: Using field sampling and community science programs from 2013 to 2023, we collected triatomines from peridomestic and domestic settings, identified them to species, analyzed for bloodmeals, and screened for T. cruzi infection and Discrete Typing Units (DTUs) TcI - TcVI utilizing molecular techniques. Triatoma sanguisuga (n=310) were collected from 23 counties throughout the state, particularly in northern and central Florida. More than one third (34.6%) of T. sanguisuga were found inside a human dwelling, and 39.2% were collected by community members. T. cruzi infection was observed in 29.5% (88/298) of tested triatomines, with infection found in 12 of the 23 counties where triatomines had been collected. DTU-typing was successful for 47 of the T. cruzi-positive triatomines: 74.5% were infected with DTU TcI, 21.3% were infected with DTU TcIV, and 4.3% were co-infected with TcI and TcIV. Bloodmeal analysis of 144 T. sanguisuga found broad host use, including mammals (60%), ectothermic vertebrates (37%), and cockroaches (2.5%). Human blood meals contributed nearly a quarter (23%) of bloodmeals, indicating significant vector-human contact. Conclusion/Significance: Our field data from Florida demonstrate that T. sanguisuga is present near, and sometimes in, human dwellings, feeds upon humans, and is infected with multiple DTUs of T. cruzi. This indicates that the environment in the southeastern United States is suitable for autochthonous transmission of Chagas disease or that the human risk for T. cruzi infection is possible. The roles of ectotherms in T. sanguisuga and T. cruzi ecology also warrant further investigation. Author SummaryTriatomines, which can spread Chagas disease to humans and other animals, are found throughout many U.S. states. Little is known about triatomines in Florida - our study aimed to learn more about where triatomines can be found in Florida, how often they are infected with the parasite that causes Chagas disease, and which animals triatomines feed on in Florida, including humans. From 2013-2023, utilizing our own field work and community science programs, we collected 310 triatomines from various regions in Florida. Both adult and immature triatomines were found, and about one-third of the triatomines were found inside human homes (35%). About 30% of the triatomines tested were infected with the Chagas disease parasite, Trypanosoma cruzi. Blood-fed triatomines mostly fed on mammals (60%, including humans), amphibians (27%), and reptiles (11%), but not on birds. Nearly a quarter of bugs (23%) had fed on human blood. Our research raises concerns for possible transmission of Chagas disease to humans from triatomines in Florida.

ecology↗

bioRxiv2024

Fur microbiome as a putative source of symbiotic bacteria in sucking lice

Symbiosis between insects and bacteria has been established countless times. While it is well known that the symbionts originated from a variety of different bacterial taxa, it is usually difficult to determine their environmental source and a route of their acquisition by the host. In this study, we address this question using a model of Neisseriaceae symbionts in rodent lice. These bacteria established their symbiosis independently with different louse taxa (Polyplax, Hoplopleura, Neohaematopinus), most likely from the same environmental source. We first applied amplicon analysis to screen for candidate source bacterium in the louse environment, that is, three species of rodents (Microtus arvalis, Clethrionomys glareolus, and Apodemus flavicollis). The screened samples included rodent fur, skin, spleen, and ectoparasites sampled from the rodents. The amplicon analysis revealed a Neisseriaceae bacterium, closely related to the known louse symbionts. We assembled genome drafts of this environmental bacterium from all three rodent hosts. The sizes of the three drafts converged to a remarkably small size of approximately 1.4 Mbp, which is even smaller than the genomes of the related symbionts. Based on these findings, we propose a hypothetical scenario of the genome evolution during the transition of a free-living bacterium to the member of the rodent fur-associated microbiome and subsequently to the facultative and obligate louse symbionts.

ecology↗

bioRxiv2023

Arsenophonus symbiosis with louse flies: multiple origins, coevolutionary dynamics, and metabolic significance

Arsenophonus is a widespread insect symbiont with life strategies that vary from parasitism to obligate mutualism. In insects living exclusively on vertebrate blood, mutualistic Arsenophonus strains are presumed to provide B vitamins missing in the insect host diet. Hippoboscidae, obligate blood-feeders related to tsetse flies, were previously suggested to have acquired Arsenophonus symbionts in several independent events. Based on comparative genomic analyzes of eleven Hippoboscidae-associated strains, nine of them newly assembled, we reveal a wide range of their genomic characteristics and phylogenetic affiliations. Phylogenetic patterns and genomic traits split the strains into two different types. Seven strains display characteristics of obligate mutualists with significantly reduced genomes and long phylogenetic branches. The remaining four genomes cluster on short branches, and their genomes resemble those of free-living bacteria or facultative symbionts. Both phylogenetic positions and genomic traits indicate that evolutionary history of the Hippoboscidae-Arsenophonus associations is a mixture of short-term coevolutions with at least four independent origins. The comparative approach to a reconstruction of B vitamin pathways across the available Arsenophonus genomes produced two kinds of patterns. On one hand, it indicated the different importance of individual B vitamins in the host-symbiont interaction. While some (riboflavin, pantothenate, folate) seem to be synthesized by all Hippoboscidae-associated obligate symbionts, pathways for others (thiamine, nicotinamide, cobalamin) are mostly missing. On the other hand, the broad comparison produced patterns which can serve as bases for further assessments of the pathways completeness and functionality. ImportanceInsects that live exclusively on vertebrate blood utilize symbiotic bacteria as a source of essential compounds, e.g. B vitamins. In louse flies, the most frequent symbiont originated in genus Arsenophonus, known from a wide range of insects. Here, we analyze genomic traits, phylogenetic origins, and metabolic capacities of eleven Arsenophonus strains associated with louse flies. We show that in louse flies Arsenophonus established symbiosis in at least four independent events, reaching different stages of symbiogenesis. This allowed for comparative genomic analysis, including convergence of metabolic capacities. The significance of the results is two-fold. First, based on a comparison of independently originated Arsenophonus symbioses, it determines the importance of individual B vitamins for the insect host. This expands our theoretical insight into insect-bacteria symbiosis. The second outcome is of methodological significance. We show that the comparative approach reveals artifacts that would be difficult to identify based on a single-genome analysis.

evolutionary biology↗

bioRxiv2023

Genome dynamics across the evolutionary transition to endosymbiosis

Endosymbiosis - where a microbe lives and replicates within a host - is an important contributor to organismal function that has accelerated evolutionary innovations and catalysed the evolution of complex life. The evolutionary processes associated with transitions to endosymbiosis, however, are poorly understood. Here, we use comparative genomics of the genus Arsenophonus to reveal the complex processes that occur on evolution of an endosymbiotic lifestyle. We compared the genomes of 38 strains spanning diverse lifestyles from environmentally acquired infections to obligate inter-dependent endosymbionts. We observed recent endosymbionts had larger genome sizes than closely related environmentally acquired strains, consistent with evolutionary innovation and rapid gain of new function. Increased genome size was a consequence of prophage and plasmid acquisition including a cargo of type III effectors, and concomitant loss of CRISPR-Cas genome defence systems enabling mobile genetic element expansion. Persistent endosymbiosis was also associated with loss of type VI secretion, likely reflecting reduced microbe-microbe competition. Thereafter, the transition to stable endosymbiosis and vertical inheritance was associated with the expected relaxation of purifying selection, pseudogenisation of genes and reduction of metabolism, leading to genome reduction. However, reduced %GC that is typically considered a progressive linear process was observed only in obligate interdependent endosymbionts. We argue that a combination of the need for rapid horizontal gene transfer-mediated evolutionary innovation together with reduced phage predation in endosymbiotic niches drives loss of genome defence systems and rapid genome expansion upon adoption of endosymbiosis. These remodelling processes precede the reductive evolution traditionally associated with adaptation to endosymbiosis.

genomics↗

bioRxiv2023

Division of labour within psyllids: Metagenomics reveals an ancient dual endosymbiosis with metabolic complementarity in the genus Cacopsylla

Hemipteran insects are well-known for their ancient associations with beneficial bacterial endosymbionts, particularly nutritional symbionts providing the host with essential nutrients such as amino acids or vitamins lacking from the hosts diet. Thereby, these primary endosymbionts enable the exploitation of nutrient-poor food sources such as plant sap or vertebrate blood. In turn, the strictly host-associated lifestyle strongly impacts the genome evolution of the endosymbionts, resulting in small and degraded genomes. Over time, even the essential nutritional functions can be compromised, leading to the complementation or replacement of an ancient endosymbiont by another, more functionally versatile, bacterium. Herein, we provide evidence for a dual primary endosymbiosis in several psyllid species. Using metagenome sequencing, we produced the complete genome sequences of both the primary endosymbiont Candidatus Carsonella ruddii and an as yet uncharacterized Enterobacteriaceae bacterium from four species of the genus Cacopsylla. The latter represents a new psyllid-associated endosymbiont clade for which we propose the name Candidatus Psyllophila symbiotica. Fluorescent in situ hybridisation confirmed the co-localization of both endosymbionts in the bacteriome. The metabolic repertoire of Psyllophila is highly conserved across host species and complements the tryptophan biosynthesis pathway that is incomplete in the co-occurring Carsonella. Unlike co-primary endosymbionts in other insects, the genome of Psyllophila is almost as small as the one of Carsonella, indicating an ancient co-obligate endosymbiosis rather than a recent association to rescue a degrading primary endosymbiont. IMPORTANCEHeritable beneficial bacterial endosymbionts have been crucial for the evolutionary success of numerous insects, enabling the exploitation of nutritionally limited food sources such as vertebrate blood and plant sap. Herein, we describe a previously unknown dual endosymbiosis in the psyllid genus Cacospylla, consisting in the primary endosymbiont Candidatus Carsonella ruddii and a co-occurring Enterobacteriaceae bacterium for which we propose the name Candidatus Psyllophila symbiotica. Its localization within the bacteriome and its small genome size confirm that Psyllophila is a co-primary endosymbiont widespread within the genus Cacopsylla. Despite its highly eroded genome, Psyllophila complements the tryptophan biosynthesis pathway that is incomplete in the co-occurring Carsonella. Moreover, the genome of Psyllophila is almost as small as the one of Carsonella, indicating an ancient dual endosymbiosis rather than a recent acquisition of a new symbiont. Hence, our results shed light on the dynamic interactions of psyllids and their endosymbionts over evolutionary time.

evolutionary biology↗

bioRxiv2023

Microbiomes of blood feeding triatomines in the context of their predatory relatives and the environment

AbstractThe importance of gut microbiomes has become generally recognized in vector biology. This study addresses microbiome signatures in North American Triatoma species of public health significance (vectors of Trypanosoma cruzi) linked to their blood feeding strategy and the natural habitat. To place the Triatoma associated microbiomes within a complex evolutionary and ecological context, we sampled sympatric Triatoma populations, related predatory reduviids, unrelated ticks, and environmental material from vertebrate nests where these arthropods reside. Along with five Triatoma species, we have characterized microbiomes of five reduviids (Stenolemoides arizonensis, Ploiaria hirticornis, Zelus longipes, and two Reduvius species), a single soft tick species, Ornithodoros turicata, and environmental microbiomes from selected sites in Arizona, Texas, Florida and Georgia. The microbiomes of predatory reduviids lack a shared core microbiota. Like in triatomines, microbiome dissimilarities among species corelate with dominance of a single bacterial taxa. These include Rickettsia, Lactobacillus, Candidatus Midichloria, and Zymobacter, which are often accompanied by known symbiotic genera, i.e., Wolbachia, Candidatus Lariskella, Asaia, Gilliamella, and Burkholderia. We have further identified compositional convergence of analyzed microbiomes in respect to the host phylogenetic distance in both blood feeding and predatory reduviids. While microbiomes of two reduviid species from Emesinae family reflect their close relationship, the microbiomes of all Triatoma species repeatedly form a distinct monophyletic cluster highlighting their phylosymbiosis. Furthermore, based on environmental microbiome profiles and blood meal analysis, we propose three epidemiologically relevant and mutually interrelated bacterial sources for Triatoma microbiomes, i.e., host abiotic environment, host skin microbiome, and pathogens circulating in host blood. ImportanceThis study places microbiomes of blood feeding North American Triatoma vectors (Reduviidae) into a broader evolutionary and ecological context provided by related predatory assassin bugs (Reduviidae), another unrelated vector species (soft tick Ornithodor turicata), and the environment these arthropods cohabit. For both vectors, microbiome analyses suggest three interrelated sources of bacteria, i.e., microbiome of vertebrate nests as their natural habitat, vertebrate skin microbiome, and pathobiome circulating in vertebrate blood. Despite an apparent influx of environment-associated bacteria into the arthropod microbiomes, Triatoma microbiomes retain their specificity, forming a distinct cluster that significantly differ from both predatory relatives and ecologically comparable ticks. Similarly, within the related predatory Reduviidae, we found the host phylogenetic distance to underlie microbiome similarities.

ecology↗

bioRxiv2021

Extremely reduced supergroup F Wolbachia: transition to obligate insect symbionts

Wolbachia are widely distributed symbionts among invertebrates that manifest by a broad spectrum of lifestyles from parasitism to mutualism. Wolbachia Supergroup F is considered a particularly interesting group which gave rise to symbionts of both arthropods and nematodes, and some of its members are obligate mutualists. Further investigations on evolutionary transitions in symbiosis have been hampered by a lack of genomic data for Supergroup F members. In this study, we present genomic data for five new supergroup F Wolbachia strains associated with four chewing lice species. These new strains in different evolutionary stages show genomic characteristics well-illustrating the evolutionary trajectory which symbiotic bacteria experience during their transition to mutualism. Three of the strains have not yet progressed with the transition, the other two show typical signs of ongoing gene deactivation and removal (genome size, coding density, low number of pseudogenes). Particularly, wMeur1, a symbiont fixed in all Menacanthus eurysternus populations across four continents, possesses a highly reduced genome of 733,850 bp with a horizontally acquired capacity for pantothenate synthesis. Comparing with other strains showed wMeur1 genome as the smallest currently known among all Wolbachia and the first example of Wolbachia which has completed genomic streamlining known from the gammaproteobacterial obligate symbionts.

evolutionary biology↗

bioRxiv2021

Microbiome structure of a wild Drosophila community along tropical elevational gradients and comparison to laboratory lines

Variation along environmental gradients in host-associated microbial communities is not well understood, compared to free-living microbial communities. Because elevational gradients may serve as natural proxies for climate change, understanding patterns along these gradients can inform our understanding of the threats hosts and their symbiotic microbes face in a warming world. In this study, we analysed bacterial microbiomes from pupae and adults of four Drosophila species native to Australian tropical rainforests. We sampled wild individuals at high and low elevation along two mountain gradients, to determine natural diversity patterns. Further, we sampled laboratory-reared individuals from isofemale lines established from the same localities, to see if any natural patterns are retained in the lab. In both environments, we controlled for diet to help elucidate other deterministic patterns of microbiome composition. We found small but significant differences in Drosophila bacterial community composition across elevation, with some notable taxonomic differences between different Drosophila species and sites. Further, we found that field-collected fly pupae had significantly richer microbiomes than laboratory-reared pupae. We also found similar microbiome composition in both types of provided diet, suggesting that the significant differences found among Drosophila microbiomes are the product of surrounding environments with different bacterial species pools, possibly bound to elevational differences in temperature. Our results suggest that comparative studies between lab and field specimens help reveal the true variability in microbiome communities that can exist within a single species. ImportanceBacteria form microbial communities inside most higher-level organisms, but we know little about how the microbiome varies along environmental gradients and between natural host populations and laboratory colonies. To explore such effects on insect-associated microbiomes, we studied gut microbiome in four Drosophila species over two mountain gradients in tropical Australia. We also compared these data to individuals kept in the laboratory to understand how different settings changed microbiome communities. We found that field sampled individuals had significantly higher microbiome diversity than those from the lab. In wild Drosophila populations, elevation explains a small but significant amount of the variation in their microbial communities. Our study highlights the importance of environmental bacterial sources for Drosophila microbiome composition across elevational gradients, and shows how comparative studies help reveal the true flexibility in microbiome communities that can exist within a species.

ecology↗

bioRxiv2021

Development and validation of an LC-MS/MS method for determination of B vitamins and some its derivatives in whole blood

Obligate symbiotic bacteria associated with the insects feeding exclusively on vertebrate blood are supposed to complement B vitamins presumably lacking in their diet, vertebrate blood. Recent genomic analyses revealed considerable differences in biosynthetic capacities across different symbionts, indicating that levels of B vitamins vary across different vertebrate hosts. However, a rigorous determination of B vitamins content in blood of various vertebrates has not yet been approached. A reliable analytical method focused on B vitamin complex in blood and hemolymph can provide valuable informative background and understanding of general principles of insect symbiosis. In this work chromatographic separation of a mixture of eight B vitamins (B1 - thiamine, B2 - riboflavin, B3 - niacin, B5 - pantothenic acid, B6 - pyridoxine, B7 - biotin, B9 - folic acid and B12 - cyanocobalamine), four B vitamin derivatives (B3 - niacinamide, B6 - pyridoxal-5-phosphate, B6 - 4-pyridoxic acid and B9 - tetrahydrofolic acid) and 3 stable isotope labelled internal standards (B3 - niacin-13C6, B5 - pantothenic acid (di-{beta}-alanine-13C6,15N2) and B7 - biotin-(ring-6,6-d2)) on C30 column was developed. Detection was carried out using dual-pressure linear ion trap mass spectrometer in FullScan MS/MS and SIM mode. Matching internal standards with analytes was done according to the results of linearity, accuracy and precision. Except for vitamin B9 (tetrahydrofolic acid) instrument quantitation limits of all analytes were ranging from 0.42 to 5.0 g/L, correlation coefficients from 0.9997 to 1.0000 and QC coefficients from 0.53 to 3.2 %. Optimization of whole blood sample preparation step was focused especially on evaluation of two types of protein-precipitation agents: trichloroacetic acid and zinc sulphate in methanol. Samples of whole blood prepared in six independent replicates were spiked at 10 g/L and 100 g/L level. The best results were obtained for zinc sulphate in methanol, but only nine analytes (B1 - thiamine, B2 - riboflavin, B3 - niacin, B3 - niacinamide, B5 - pantothenic acid, B6 - pyridoxine, B6 - 4-pyridoxic acid, B7 - biotin and B12 - cyanocobalamine) were successfully validated. Accuracy of the procedure using this protein-precipitating agent was ranging from 89 to 120 %, precision from 0.5 to 13 % and process efficiency from 65 to 108 %. HighlightsO_LILC-MS/MS method for quantitation of eight B vitamins and four B vitamin derivatives was developed. C_LIO_LIDeproteinization agents trichloroacetic acid and ZnSO4/methanol were tested for protein precipitation of whole blood. C_LIO_LIAccuracy, precision and process efficiency were evaluated. C_LIO_LISuccessful method validation for seven B vitamins and two B vitamin derivatives in whole blood. C_LI

biochemistry↗

bioRxiv2020

Ontogeny, species identity and environment dominate microbiome dynamics in wild populations of kissing bugs (Triatominae)

BackgroundKissing bugs (Triatominae) are blood-feeding insects best known as the vectors of Trypanosoma cruzi, the causative agent of Chagas disease. Considering the high epidemiological relevance of these vectors, their biology and bacterial symbiosis remains surprisingly understudied. While previous investigations revealed generally low individual complexity but high among-individual variability of the triatomine microbiomes, any consistent microbiome determinants have not yet been identified across multiple Triatominae species. MethodsTo obtain a more comprehensive view of triatomine microbiomes, we investigated the host-microbiome relationship of five Triatoma species sampled from white-throated woodrat (Neotoma albigula) nests in multiple locations across the USA. We applied optimized 16S rRNA gene metabarcoding with a novel 18S rRNA gene blocking primer to a set of 170 T. cruzi negative individuals across all six instars. ResultsTriatomine gut microbiome composition is strongly influenced by three principal factors: ontogeny, species identity, and the environment. The microbiomes are characterised by significant loss in bacterial diversity throughout ontogenetic development. First instars possess the highest bacterial diversity while adult microbiomes are routinely dominated by a single taxon. Primarily, the bacterial genus Dietzia dominates late-stage nymphs and adults of T. rubida, T. protracta, and T. lecticularia, but is not present in the phylogenetically more distant T. gerstaeckeri and T. sanguisuga. Species-specific microbiome composition, particularly pronounced in early instars, is further modulated by locality-specific effects. In addition, pathogenic bacteria of the genus Bartonella, acquired from the vertebrate hosts, are an abundant component of Triatoma microbiomes. ConclusionOur study is the first to demonstrate deterministic patterns in microbiome composition among all life stages and multiple Triatoma species. We hypothesize that triatomine microbiome assemblages are produced by species- and life stage-dependent uptake of environmental bacteria and multiple indirect transmission strategies that promote bacterial transfer between individuals. Altogether, our study highlights the complexity of Triatominae symbiosis with bacteria and warrant further investigation to understand microbiome function in these important vectors.

ecology↗

bioRxiv2019

A new symbiotic lineage related to Neisseria and Snodgrassella arises from the dynamic and diverse microbiomes in sucking lice

Phylogenetic diversity of symbiotic bacteria in sucking lice suggests that lice have experienced a complex history of symbiont acquisition, loss, and replacement during their evolution. By combining metagenomics and amplicon screening across several populations of two louse genera (Polyplax and Hoplopleura) we describe a novel louse symbiont lineage related to Neisseria and Snodgrassella, and show its independent origin within dynamic lice microbiomes. While the genomes of these symbionts are highly similar in both lice genera, their respective distributions and status within lice microbiomes indicate that they have different functions and history. In Hoplopleura acanthopus, the Neisseria-related bacterium is a dominant obligate symbiont universally present across several hosts populations, and seems to be replacing a presumably older and more degenerated obligate symbiont. In contrast, the Polyplax microbiomes are dominated by the obligate symbiont Legionella polyplacis, with the Neisseria-related bacterium co-occurring only in some samples and with much lower abundance.

evolutionary biology↗

bioRxiv2018

Microbiomes of North American Triatominae: the grounds for Chagas disease epidemiology

AbstarctInsect microbiomes influence many fundamental host traits, including functions of practical significance such as their capacity as vectors to transmit parasites and pathogens. The knowledge on the diversity and development of the gut microbiomes in various blood feeding insects is thus crucial not only for theoretical purposes, but also for the development of better disease control strategies. In Triatominae (Heteroptera: Reduviidae), the blood feeding vectors of Chagas disease in South America and parts of North America, the investigation of the microbiomes is in its infancy. The few studies done on microbiomes of South American Triatominae species indicate a relatively low taxonomic diversity and a high host specificity. We designed a comparative survey to serve several purposes: I) to obtain a better insight into the overall microbiome diversity in different species, II) to check the long term stability of the interspecific differences, III) to describe the ontogenetic changes of the microbiome, and IV) to determine the potential correlation between microbiome composition and presence in the vector gut of Trypanosoma cruzi, the causative agent of Chagas disease. Using 16S amplicons of two abundant species from the southern US, and four laboratory reared colonies, we showed that the microbiome composition is determined by host species, rather than locality or environment. The OTUs (Operational Taxonomic Units) determination confirms a low microbiome diversity, with 12-17 main OTUs detected in wild populations of T. sanguisuga and T. protracta. Among the dominant bacterial taxa are Acinetobacter and Proteiniphilum but also the symbiotic bacterium Arsenophonus triatominarum, previously believed to only live intracellularly. The possibility of ontogenic microbiome changes was evaluated in all six developmental stages and faeces of the laboratory reared model Rhodnius prolixus. We detected considerable changes along the hosts ontogeny, including clear trends in the abundance variation of the three dominant bacteria, namely Enterococcus, Acinetobacter and Arsenophonus. Finally, we screened the samples for the presence of T. cruzi. Comparing the parasite presence with the microbiome composition, we assessed the possible significance of the latter in the epidemiology of the disease. Particularly, we found a trend towards more diverse microbiomes in T. cruzi positive T. protracta specimens.

microbiology↗