Author correction: large enriched fragment targeted sequencing (LEFT-SEQ) applied to capture of Wolbachia genomes

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Impact of intestinal parasites on microbiota and cobalamin gene sequences: A pilot study

Background: Approximately 30% of children worldwide are infected with gastrointestinal parasites. Depending on the species, parasites can disrupt intestinal bacterial microbiota affecting essential vitamin biosynthesis. Methods: Stool samples were collected from 37 asymptomatic children from a previous cross-sectional Argentinian study. A multi-parallel real-time quantitative PCR was implemented for Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Trichuris trichiura, Cryptosporidium spp., Entamoeba histolytica and Giardia duodenalis. In addition, whole-genome sequencing analysis was conducted for bacterial microbiota on all samples and analyzed using Livermore Metagenomic Analysis Toolkit and DIAMOND software. Separate analyses were carried out for uninfected, Giardia-only, Giardia + helminth co-infections, and helminth-only groups. Results: For Giardia-only infected children compared to uninfected children, DNA sequencing data showed a decrease in microbiota biodiversity that correlated with increasing Giardia burden and was statistically significant using Shannon's alpha diversity (Giardia-only > 1 fg/μl 2.346; non-infected group 3.253, P = 0.0317). An increase in diversity was observed for helminth-only infections with a decrease in diversity for Giardia + helminth co-infections (P = 0.00178). In Giardia-only infections, microbiome taxonomy changed from Firmicutes towards increasing proportions of Prevotella, with the degree of change related to the intensity of infection compared to uninfected (P = 0.0317). The abundance of Prevotella bacteria was decreased in the helminths-only group but increased for Giardia + helminth co-infections (P = 0.0262). Metagenomic analysis determined cobalamin synthesis was decreased in the Giardia > 1 fg/μl group compared to both the Giardia < 1 fg/μl and the uninfected group (P = 0.0369). Giardia + helminth group also had a decrease in cobalamin CbiM genes from helminth-only infections (P = 0.000754). Conclusion: The study results may provide evidence for an effect of parasitic infections enabling the permissive growth of anaerobic bacteria such as Prevotella, suggesting an altered capacity of vitamin B12 (cobalamin) biosynthesis and potential impact on growth and development in children.Fil: Mejia, Rojelio. Baylor College of Medicine; Estados Unidos. Universidad Nacional de Salta; ArgentinaFil: Damania, Ashish. Baylor College of Medicine; Estados UnidosFil: Jeun, Rebecca. Baylor College of Medicine; Estados UnidosFil: Bryan, Patricia E.. Baylor College of Medicine; Estados UnidosFil: Vargas, Paola. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; ArgentinaFil: Juarez, Marisa del Valle. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; ArgentinaFil: Cajal, Silvana Pamela. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; ArgentinaFil: Nasser, Julio Rubén. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; ArgentinaFil: Krolewiecki, Alejandro Javier. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; ArgentinaFil: Lefoulon, Emilie. New England Biolabs; Estados UnidosFil: Long, Courtney. New England Biolabs; Estados UnidosFil: Drake, Evan. New England Biolabs; Estados UnidosFil: Cimino, Rubén Oscar. Universidad Nacional de Salta. Sede Regional Orán. Instituto de Investigación de Enfermedades Tropicales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta; ArgentinaFil: Slatko, Barton. New England Biolabs; Estados Unido

Diminutive, degraded but dissimilar: <i>Wolbachia</i> genomes from filarial nematodes do not conform to a single paradigm.

Wolbachia are alpha-proteobacteria symbionts infecting a large range of arthropod species and two different families of nematodes. Interestingly, these endosymbionts are able to induce diverse phenotypes in their hosts: they are reproductive parasites within many arthropods, nutritional mutualists within some insects and obligate mutualists within their filarial nematode hosts. Defining Wolbachia 'species' is controversial and so they are commonly classified into 17 different phylogenetic lineages, termed supergroups, named A-F, H-Q and S. However, available genomic data remain limited and not representative of the full Wolbachia diversity; indeed, of the 24 complete genomes and 55 draft genomes of Wolbachia available to date, 84 % belong to supergroups A and B, exclusively composed of Wolbachia from arthropods. For the current study, we took advantage of a recently developed DNA-enrichment method to produce four complete genomes and two draft genomes of Wolbachia from filarial nematodes. Two complete genomes, wCtub and wDcau, are the smallest Wolbachia genomes sequenced to date (863 988 bp and 863 427 bp, respectively), as well as the first genomes representing supergroup J. These genomes confirm the validity of this supergroup, a controversial clade due to weaknesses of the multilocus sequence typing approach. We also produced the first draft Wolbachia genome from a supergroup F filarial nematode representative (wMhie), two genomes from supergroup D (wLsig and wLbra) and the complete genome of wDimm from supergroup C. Our new data confirm the paradigm of smaller Wolbachia genomes from filarial nematodes containing low levels of transposable elements and the absence of intact bacteriophage sequences, unlike many Wolbachia from arthropods, where both are more abundant. However, we observe differences among the Wolbachia genomes from filarial nematodes: no global co-evolutionary pattern, strong synteny between supergroup C and supergroup J Wolbachia, and more transposable elements observed in supergroup D Wolbachia compared to the other supergroups. Metabolic pathway analysis indicates several highly conserved pathways (haem and nucleotide biosynthesis, for example) as opposed to more variable pathways, such as vitamin B biosynthesis, which might be specific to certain host-symbiont associations. Overall, there appears to be no single Wolbachia-filarial nematode pattern of co-evolution or symbiotic relationship

Palaeosymbiosis Revealed by Genomic Fossils of Wolbachia in a Strongyloidean Nematode

Wolbachia are common endosymbionts of terrestrial arthropods, and are also found in nematodes: the animal-parasitic filaria, and the plant-parasite Radopholus similis. Lateral transfer of Wolbachia DNA to the host genome is common. We generated a draft genome sequence for the strongyloidean nematode parasite Dictyocaulus viviparus, the cattle lungworm. In the assembly, we identified nearly 1 Mb of sequence with similarity to Wolbachia. The fragments were unlikely to derive from a live Wolbachia infection: most were short, and the genes were disabled through inactivating mutations. Many fragments were co-assembled with definitively nematode-derived sequence. We found limited evidence of expression of the Wolbachia-derived genes. The D. viviparus Wolbachia genes were most similar to filarial strains and strains from the host promiscuous clade F. We conclude that D. viviparus was infected by Wolbachia in the past, and that clade F-like symbionts may have been the source of filarial Wolbachia infections

Transcriptomic Analysis of Steinernema Nematodes Highlights Metabolic Costs Associated to Xenorhabdus Endosymbiont Association and Rearing Conditions

Entomopathogenic nematodes of the genus Steinernema have a mutualistic relationship with bacteria of the genus Xenorhabdus and together they form an antagonist partnership against their insect hosts. The nematodes (third-stage infective juveniles, or IJs) protect the bacteria from the external environmental stressors and vector them from one insect host to another. Xenorhabdus produce secondary metabolites and antimicrobial compounds inside the insect that protect the cadaver from soil saprobes and scavengers. The bacteria also become the nematodes’ food, allowing them to grow and reproduce. Despite these benefits, it is yet unclear what the potential metabolic costs for Steinernema IJs are relative to the maintenance and vectoring of Xenorhabdus. In this study, we performed a comparative dual RNA-seq analysis of IJs of two nematode-bacteria partnerships: Steinernema carpocapsae-Xenorhabdus nematophila and Steinernema. puntauvense-Xenorhbdus bovienii. For each association, three conditions were studied: (1) IJs reared in the insect (in vivo colonized), (2) colonized IJs reared on liver-kidney agar (in vitro colonized), and (3) IJs depleted by the bacteria reared on liver-kidney agar (in vitro aposymbiotic). Our study revealed the downregulation of numerous genes involved in metabolism pathways, such as carbohydrate, amino acid, and lipid metabolism when IJs were reared in vitro, both colonized and without the symbiont. This downregulation appears to impact the longevity pathway, with the involvement of glycogen and trehalose metabolism, as well as arginine metabolism. Additionally, a differential expression of the venom protein known to be secreted by the nematodes was observed when both Steinernema species were depleted of their symbiotic partners. These results suggest Steinernema IJs may have a mechanism to adapt their virulence in absence of their symbionts. Copyright © 2022 Lefoulon, McMullen and Stock.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Identification of novel prophage regions in Xenorhabdus nematophila genome and gene expression analysis during phage-like particle induction

Background: Entomopathogenic Xenorhabdus bacteria are endosymbionts of Steinernema nematodes and together they form an insecticidal mutualistic association that infects a wide range of insect species. Xenorhabdus produce an arsenal of toxins and secondary metabolites that kill the insect host. In addition, they can induce the production of diverse phage particles. A few studies have focused on one integrated phage responsible for producing a phage tail-like bacteriocin, associated with an antimicrobial activity against other Xenorhabdus species. However, very little is known about the diversity of prophage regions in Xenorhabdus species. Methods: In the present study, we identified several prophage regions in the genome of Xenorhabdus nematophila AN6/1. We performed a preliminary study on the relative expression of genes in these prophage regions. We also investigated some genes (not contained in prophage region) known to be involved in SOS bacterial response (recA and lexA) associated with mitomycin C and UV exposure. Results: We described two integrated prophage regions (designated Xnp3 and Xnp4) not previously described in the genome of Xenorhabdus nematophila AN6/1. The Xnp3 prophage region appears very similar to complete Mu-like bacteriophage. These prophages regions are not unique to X. nematophila species, although they appear less conserved among Xenorhabdus species when compared to the previously described p1 prophage region. Our results showed that mitomycin C exposure induced an up-regulation of recA and lexA suggesting activation of SOS response. In addition, mitomycin C and UV exposure seems to lead to up-regulation of genes in three of the four integrated prophages regions. Copyright 2022 Lefoulon et al.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Differential tissular distribution of Litomosoides sigmodontis microfilariae between microfilaremic and amicrofilaremic mice following experimental infection

Filariases are caused by onchocercid nematodes that are transmitted by arthropod vectors. More than 180 million people are infected worldwide. Mass drug administration has been set up in many endemic areas to control the parasite burden. Although very successful in limiting microfilarial load, transmission has not been completely interrupted in such areas. A proportion of infected patients with lymphatic filariasis or loiasis are known to be amicrofilaremic, as they do not present microfilariae in their bloodstream despite the presence of adult worms. A mirror status also exists in CBA/Ca mice infected with Litomosoides sigmodontis, the well-established model of filariasis. Using this model, the goal of this study was to determine if the kinetics of blood clearance of microfilariae differed between amicrofilaremic CBA/Ca mice and microfilaremic BALB/c mice. For this purpose, a qPCR approach was devised to detect microfilariae in different tissues, after a controlled inoculation of microfilariae. We showed that the rapid clearance of microfilariae from the pleural cavity or from the bloodstream of CBA/Ca mice was associated with a massive accumulation of first stage larvae in the lungs, liver and spleen

Differential tissular distribution of

Filariases are caused by onchocercid nematodes that are transmitted by arthropod vectors. More than 180 million people are infected worldwide. Mass drug administration has been set up in many endemic areas to control the parasite burden. Although very successful in limiting microfilarial load, transmission has not been completely interrupted in such areas. A proportion of infected patients with lymphatic filariasis or loiasis are known to be amicrofilaremic, as they do not present microfilariae in their bloodstream despite the presence of adult worms. A mirror status also exists in CBA/Ca mice infected with Litomosoides sigmodontis, the well-established model of filariasis. Using this model, the goal of this study was to determine if the kinetics of blood clearance of microfilariae differed between amicrofilaremic CBA/Ca mice and microfilaremic BALB/c mice. For this purpose, a qPCR approach was devised to detect microfilariae in different tissues, after a controlled inoculation of microfilariae. We showed that the rapid clearance of microfilariae from the pleural cavity or from the bloodstream of CBA/Ca mice was associated with a massive accumulation of first stage larvae in the lungs, liver and spleen

Pseudoscorpion Wolbachia symbionts: diversity and evidence for a new supergroup S

Background Wolbachia are the most widely spread endosymbiotic bacteria, present in a wide variety of insects and two families of nematodes. As of now, however, relatively little genomic data has been available. The Wolbachia symbiont can be parasitic, as described for many arthropod systems, an obligate mutualist, as in filarial nematodes or a combination of both in some organisms. They are currently classified into 16 monophyletic lineage groups (“supergroups”). Although the nature of these symbioses remains largely unknown, expanded Wolbachia genomic data will contribute to understanding their diverse symbiotic mechanisms and evolution. Results This report focuses on Wolbachia infections in three pseudoscorpion species infected by two distinct groups of Wolbachia strains, based upon multi-locus phylogenies. Geogarypus minor harbours wGmin and Chthonius ischnocheles harbours wCisc, both closely related to supergroup H, while Atemnus politus harbours wApol, a member of a novel supergroup S along with Wolbachia from the pseudoscorpion Cordylochernes scorpioides (wCsco). Wolbachia supergroup S is most closely related to Wolbachia supergroups C and F. Using target enrichment by hybridization with Wolbachia-specific biotinylated probes to capture large fragments of Wolbachia DNA, we produced two draft genomes of wApol. Annotation of wApol highlights presence of a biotin operon, which is incomplete in many sequenced Wolbachia genomes. Conclusions The present study highlights at least two symbiont acquisition events among pseudoscorpion species. Phylogenomic analysis indicates that the Wolbachia from Atemnus politus (wApol), forms a separate supergroup (“S”) with the Wolbachia from Cordylochernes scorpioides (wCsco). Interestingly, the biotin operon, present in wApol, appears to have been horizontally transferred multiple times along Wolbachia evolutionary history.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Large Enriched Fragment Targeted Sequencing (LEFT-SEQ) Applied to Capture of Wolbachia Genomes

Abstract Symbiosis is a major force of evolutionary change, influencing virtually all aspects of biology, from population ecology and evolution to genomics and molecular/biochemical mechanisms of development and reproduction. A remarkable example is Wolbachia endobacteria, present in some parasitic nematodes and many arthropod species. Acquisition of genomic data from diverse Wolbachia clades will aid in the elucidation of the different symbiotic mechanisms(s). However, challenges of de novo assembly of Wolbachia genomes include the presence in the sample of host DNA: nematode/vertebrate or insect. We designed biotinylated probes to capture large fragments of Wolbachia DNA for sequencing using PacBio technology (LEFT-SEQ: Large Enriched Fragment Targeted Sequencing). LEFT-SEQ was used to capture and sequence four Wolbachia genomes: the filarial nematode Brugia malayi, wBm, (21-fold enrichment), Drosophila mauritiana flies (2 isolates), wMau (11-fold enrichment), and Aedes albopictus mosquitoes, wAlbB (200-fold enrichment). LEFT-SEQ resulted in complete genomes for wBm and for wMau. For wBm, 18 single-nucleotide polymorphisms (SNPs), relative to the wBm reference, were identified and confirmed by PCR. A limit of LEFT-SEQ is illustrated by the wAlbB genome, characterized by a very high level of insertion sequences elements (ISs) and DNA repeats, for which only a 20-contig draft assembly was achieved