Transcriptional Response of Wolbachia to Dengue Virus Infection in Cells of the Mosquito Aedes aegypti

Aedes aegypti is a vector of several pathogenic viruses, including dengue, Zika, chikungunya, and yellow fever viruses, which are of importance to human health. Wolbachia is an endosymbiotic bacterium currently used in transinfected mosquitoes to suppress replication and transmission of dengue viruses. However, the mechanism of Wolbachia -mediated virus inhibition is not fully understood.

Using Wolbachia to eliminate dengue – will the virus fight back?

Recent field trials have demonstrated that dengue incidence can be substantially reduced by introgressing strains of the endosymbiotic bacterium, Wolbachia into Aedes aegypti mosquito populations. This strategy relies on Wolbachia reducing the susceptibility of Ae. aegypti to disseminated infection by positive-sense RNA viruses like dengue. However, RNA viruses are well known to adapt to antiviral pressures. Here we review the viral infection stages where selection for Wolbachia-resistant virus variants could occur. We also consider the genetic constraints imposed on viruses that alternate between vertebrate and invertebrate hosts, and the likely selection pressures that dengue virus might adapt to in order to be effectively transmitted by Ae. aegypti that carry Wolbachia. Whilst there are hurdles to dengue viruses developing resistance to Wolbachia, we suggest that long-term surveillance for resistant viruses should be an integral component of Wolbachia-introgression biocontrol programs.

Wolbachia infection prevalence as common insects’ endosymbiont in the rural area of Yogyakarta, Indonesia

Kumalawati DA, Supriyati E, Rachman MP. Oktriani R, Kurniasari I, Candrasari DS, Hidayati L, Handayaningsih AE, Probowati VC, Arianto B, Wardana DS, Pramuko NB, Utari A, Tantowijoyo W, Arguni E. 2020. Wolbachia infection prevalence as common insects’ endosymbiont in the rural area of Yogyakarta, Indonesia. Biodiversitas 21: 5608-5614. Control for mosquito-borne diseases such as dengue and chikungunya using vectors is urgently required. The World Mosquito Program, a multinational collaborative research program, is currently studying for the ability of Wolbachia-infected Aedes aegypti to control dengue. Community concerns on Wolbachia's natural existence in their surrounding living area. This study presents the field study of Wolbachia pipientis in insects commonly found in rural daily life. W. pipientis is an endosymbiotic bacterium commonly found in arthropods. Insects were collected from five villages in Sleman and Bantul District, Yogyakarta Province, Indonesia, from July to December 2012 and screened for Wolbachia infection using PCR. One hundred insects, including butterflies, moths, mosquitoes, flies, were collected. The results indicated that 44.9% of identified insect species were positive for Wolbachia pipientis, which support the existing data from other regions on the spread of Wolbachia infection in insects.

Tripartite Symbiosis of an Anaerobic Scuticociliate with Two Hydrogenosome-Associated Endosymbionts, a Holospora-Related Alphaproteobacterium and a Methanogenic Archaeon

ABSTRACT A number of anaerobic ciliates, unicellular eukaryotes, intracellularly possess methanogenic archaea and bacteria as symbiotic partners. Although this tripartite relationship is of interest in terms of the fact that each participant is from a different domain, the difficulty in culture and maintenance of those host species with symbiotic partners has disturbed both ecological and functional studies so far. In this study, we obtained a stable culture of a small anaerobic scuticociliate, strain GW7. By transmission electron microscopic observation and fluorescent in situ hybridization with domain-specific probes, we demonstrate that GW7 possesses both archaeal and bacterial endosymbionts in its cytoplasm. These endosymbionts are in dependently associated with hydrogenosomes, which are organelle producing hydrogen and ATP under anaerobic conditions. Clone library analyses targeting prokaryotic 16S rRNA genes, fluorescent in situ hybridization with endosymbiont-specific probes, and molecular phylogenetic analyses revealed the phylogenetic affiliations and intracellular localizations of these endosymbionts. The endosymbiotic archaeon is a methanogen belonging to the genus Methanoregula (order Methanomicrobiales); a member of this genus has previously been described as the endosymbiont of an anaerobic ciliate from the genus Metopus (class Armophorea), which is only distantly related to strain GW7 (class Oligohymenophorea). The endosymbiotic bacterium belongs to the family Holosporaceae of the class Alphaproteobacteria, which also comprises several endosymbionts of various aerobic ciliates. For this endosymbiotic bacterium, we propose a novel candidate genus and species, “Candidatus Hydrogenosomobacter endosymbioticus.” IMPORTANCE Tripartite symbioses between anaerobic ciliated protists and their intracellular archaeal and bacterial symbionts are not uncommon, but most reports have been based mainly on microscopic observations. Deeper insights into the function, ecology, and evolution of these fascinating symbioses involving partners from all three domains of life have been hampered by the difficulties of culturing anaerobic ciliates in the laboratory and the frequent loss of their prokaryotic partners during long-term cultivation. In the present study, we report the isolation of an anaerobic scuticociliate, strain GW7, which has been stably maintained in our laboratory for more than 3 years without losing either of its endosymbionts. Unexpectedly, molecular characterization of the endosymbionts revealed that the bacterial partner of GW7 is phylogenetically related to intranuclear endosymbionts of aerobic ciliates. This strain will enable future genomic, transcriptomic, and proteomic analyses of the interactions in this tripartite symbiosis and a comparison with endosymbioses in aerobic ciliates.

Changes in Endosymbiont Complexity Drive Host-Level Compensatory Adaptations in Cicadas

ABSTRACTFor insects that depend on one or more bacterial endosymbionts for survival, it is critical that these bacteria are faithfully transmitted between insect generations. Cicadas harbor two essential bacterial endosymbionts, “CandidatusSulcia muelleri” and “CandidatusHodgkinia cicadicola.” In some cicada species,Hodgkiniahas fragmented into multiple distinct but interdependent cellular and genomic lineages that can differ in abundance by more than two orders of magnitude. This complexity presents a potential problem for the host cicada, because low-abundance but essentialHodgkinialineages risk being lost during the symbiont transmission bottleneck from mother to egg. Here we show that all cicada eggs seem to receive the full complement ofHodgkinialineages, and that in cicadas with more complexHodgkiniathis outcome is achieved by increasing the number ofHodgkiniacells transmitted by up to 6-fold. We further show that cicada species with varyingHodgkiniacomplexity do not visibly alter their transmission mechanism at the resolution of cell biological structures. Together these data suggest that a major cicada adaptation to changes in endosymbiont complexity is an increase in the number ofHodgkiniacells transmitted to each egg. We hypothesize that the requirement to increase the symbiont titer is one of the costs associated withHodgkiniafragmentation.IMPORTANCESap-feeding insects critically rely on one or more bacteria or fungi to provide essential nutrients that are not available at sufficient levels in their diets. These microbes are passed between insect generations when the mother places a small packet of microbes into each of her eggs before it is laid. We have previously described an unusual lineage fragmentation process in a nutritional endosymbiotic bacterium of cicadas calledHodgkinia. In some cicadas, a singleHodgkinialineage has split into numerous related lineages, each performing a subset of original function and therefore each required for normal host function. Here we test how this splitting process affects symbiont transmission to eggs. We find that cicadas dramatically increase the titer ofHodgkiniacells passed to each egg in response to lineage fragmentation, and we hypothesize that this increase in bacterial cell count is one of the major costs associated with endosymbiont fragmentation.

Novel Wolbachia strains in Anopheles malaria vectors from Sub-Saharan Africa

Background: Wolbachia, a common insect endosymbiotic bacterium that can influence pathogen transmission and manipulate host reproduction, has historically been considered absent from the Anopheles (An.) genera, but has recently been found in An. gambiae s.l. populations. As there are numerous Anopheles species that have the capacity to transmit malaria, we analysed a range of species to determine Wolbachia prevalence rates, characterise novel Wolbachia strains and determine any correlation between the presence of Plasmodium, Wolbachia and the competing endosymbiotic bacterium Asaia. Methods: Anopheles adult mosquitoes were collected from five malaria-endemic countries: Guinea, Democratic Republic of the Congo (DRC), Ghana, Uganda and Madagascar, between 2013 and 2017. Molecular analysis of samples was undertaken using quantitative PCR, Sanger sequencing, Wolbachia multilocus sequence typing (MLST) and high-throughput amplicon sequencing of the bacterial 16S rRNA gene. Results: Novel Wolbachia strains were discovered in five species: An. coluzzii, An. gambiae s.s., An. arabiensis, An. moucheti and An. species ‘A’, increasing the number of Anopheles species known to be naturally infected. Variable prevalence rates in different locations were observed and novel strains were phylogenetically diverse, clustering with Wolbachia supergroup B strains. We also provide evidence for resident strain variants within An. species ‘A’. Wolbachia is the dominant member of the microbiome in An. moucheti and An. species ‘A’, but present at lower densities in An. coluzzii. Interestingly, no evidence of Wolbachia/Asaia co-infections was seen and Asaia infection densities were also shown to be variable and location dependent. Conclusions: The important discovery of novel Wolbachia strains in Anopheles provides greater insight into the prevalence of resident Wolbachia strains in diverse malaria vectors. Novel Wolbachia strains (particularly high-density strains) are ideal candidate strains for transinfection to create stable infections in other Anopheles mosquito species, which could be used for population replacement or suppression control strategies.