Azithromycin resistance in Escherichia coli and Salmonella from food-producing animals and meat in Europe.

OBJECTIVES: To characterize the genetic basis of azithromycin resistance in Escherichia coli and Salmonella collected within the EU harmonized antimicrobial resistance (AMR) surveillance programme in 2014-18 and the Danish AMR surveillance programme in 2016-19. METHODS: WGS data of 1007 E. coli [165 azithromycin resistant (MIC > 16 mg/L)] and 269 Salmonella [29 azithromycin resistant (MIC > 16 mg/L)] were screened for acquired macrolide resistance genes and mutations in rplDV, 23S rRNA and acrB genes using ResFinder v4.0, AMRFinder Plus and custom scripts. Genotype-phenotype concordance was determined for all isolates. Transferability of mef(C)-mph(G)-carrying plasmids was assessed by conjugation experiments. RESULTS: mph(A), mph(B), mef(B), erm(B) and mef(C)-mph(G) were detected in E. coli and Salmonella, whereas erm(C), erm(42), ere(A) and mph(E)-msr(E) were detected in E. coli only. The presence of macrolide resistance genes, alone or in combination, was concordant with the azithromycin-resistant phenotype in 69% of isolates. Distinct mph(A) operon structures were observed in azithromycin-susceptible (n = 50) and -resistant (n = 136) isolates. mef(C)-mph(G) were detected in porcine and bovine E. coli and in porcine Salmonella enterica serovar Derby and Salmonella enterica 1,4, [5],12:i:-, flanked downstream by ISCR2 or TnAs1 and associated with IncIγ and IncFII plasmids. CONCLUSIONS: Diverse azithromycin resistance genes were detected in E. coli and Salmonella from food-producing animals and meat in Europe. Azithromycin resistance genes mef(C)-mph(G) and erm(42) appear to be emerging primarily in porcine E. coli isolates. The identification of distinct mph(A) operon structures in susceptible and resistant isolates increases the predictive power of WGS-based methods for in silico detection of azithromycin resistance in Enterobacterales.

Global survey of hgcA-carrying genomes in marine and freshwater sediments: Insights into mercury methylation processes.

Mercury (Hg) methylation is a microbially mediated process that produces methylmercury (MeHg), a bioaccumulative neurotoxin. A highly conserved gene pair, hgcAB, is required for Hg methylation, which provides a basis for identifying Hg methylators and evaluating their genomic composition. In this study, we conducted a large-scale omics analysis in which 281 metagenomic freshwater and marine sediment samples from 46 geographic locations across the globe were queried. Specific objectives were to examine the prevalence of Hg methylators, to identify horizontal gene transfer (HGT) events involving hgcAB within Hg methylator communities, and to identify associations between hgcAB and microbial biochemical functions/genes. Hg methylators from the phyla Desulfobacterota and Bacteroidota were dominant in both freshwater and marine sediments while Firmicutes and methanogens belonging to Euryarchaeota were identified only in freshwater sediments. Novel Hg methylators were found in the Phycisphaerae and Planctomycetia classes within the phylum Planctomycetota, including potential hgcA-carrying anammox metagenome-assembled genomes (MAGs) from Candidatus Brocadiia. HGT of hgcA and hgcB were identified in both freshwater and marine methylator communities. Spearman's correlation analysis of methylator genomes suggested that in addition to sulfide, thiosulfate, sulfite, and ammonia may be important parameters for Hg methylation processes in sediments. Overall, our results indicated that the biochemical drivers of Hg methylation vary between marine and freshwater sites, lending insight into the influence of environmental perturbances, such as a changing climate, on Hg methylation processes.

Divergent endophytic viromes and phage genome repertoires among banana (Musa) species.

Introduction: Viruses generally cause disease, but some viruses may be beneficial as resident regulators of their hosts or host microbiomes. Plant-associated viruses can help plants survive by increasing stress tolerance or regulating endophytic communities. The goal of this study was to characterize endophytic virus communities in banana and plantain (Musa spp.) genotypes, including cultivated and wild species, to assess virome repertoires and detect novel viruses. Methods: DNA viral communities were characterized by shotgun sequencing of an enriched endosphere extract from leaves and roots or corm of 7 distinct Musa genotypes (M. balbisiana, Thai Black, M. textilis, M. sikkimensis, Dwarf Cavendish, Williams Hybrid, and FHIA-25 Hybrid). Results: Results showed abundant virus-like contigs up to 108,191 bp long with higher relative abundance in leaves than roots. Analyses predicted 733 phage species in 51 families, with little overlap in phage communities among plants. Phage diversity was higher in roots and in diploid wild hosts. Ackermanniviridae and Rhizobium phage were generally the most abundant taxa. A Rhizobium RR1-like phage related to a phage of an endophytic tumor-causing rhizobium was found, bearing a holin gene and a partial Shiga-like toxin gene, raising interest in its potential to regulate endophytic Rhizobiaceae. Klebsiella phages were of interest for possible protection against Fusarium wilt, and other phages were predicted with potential to regulate Erwinia, Pectobacterium, and Ralstonia-associated diseases. Although abundant phage-containing contigs were functionally annotated, revealing 1,038 predicted viral protein domains, gene repertoires showed high divergence from database sequences, suggesting novel phages in these banana cultivars. Plant DNA viruses included 56 species of Badnavirus and 26 additional non-Musa plant viruses with distributions that suggested a mixture of resident and transient plant DNA viruses in these samples. Discussion: Together, the disparate viral communities in these plants from a shared environment suggest hosts drive the composition of these virus communities. This study forms a first step in understanding the endophytic virome in this globally important food crop, which is currently threatened by fungal, bacterial, and viral diseases.

Metapangenomics of wild and cultivated banana microbiome reveals a plethora of host-associated protective functions.

BACKGROUND: Microbiomes are critical to plants, promoting growth, elevating stress tolerance, and expanding the plant's metabolic repertoire with novel defense pathways. However, generally microbiomes within plant tissues, which intimately interact with their hosts, remain poorly characterized. These endospheres have become a focus in banana (Musa spp.)-an important plant for study of microbiome-based disease protection. Banana is important to global food security, while also being critically threatened by pandemic diseases. Domestication and clonal propagation are thought to have depleted protective microbiomes, whereas wild relatives may hold promise for new microbiome-based biological controls. The goal was to compare metapangenomes enriched from 7 Musa genotypes, including wild and cultivated varieties grown in sympatry, to assess the host associations with root and leaf endosphere functional profiles. RESULTS: Density gradients successfully generated culture-free microbial enrichment, dominated by bacteria, with all together 24,325 species or strains distinguished, and 1.7 million metagenomic scaffolds harboring 559,108 predicted gene clusters. About 20% of sequence reads did not match any taxon databases and ~ 62% of gene clusters could not be annotated to function. Most taxa and gene clusters were unshared between Musa genotypes. Root and corm tissues had significantly richer endosphere communities that were significantly different from leaf communities. Agrobacterium and Rhizobium were the most abundant in all samples while Chitinophagia and Actinomycetia were more abundant in roots and Flavobacteria in leaves. At the bacterial strain level, there were > 2000 taxa unique to each of M. acuminata (AAA genotype) and M. balbisiana (B-genotype), with the latter 'wild' relatives having richer taxa and functions. Gene ontology functional enrichment showed core beneficial functions aligned with those of other plants but also many specialized prospective beneficial functions not reported previously. Some gene clusters with plant-protective functions showed signatures of phylosymbiosis, suggesting long-standing associations or heritable microbiomes in Musa. CONCLUSIONS: Metapangenomics revealed key taxa and protective functions that appeared to be driven by genotype, perhaps contributing to host resistance differences. The recovery of rich novel taxa and gene clusters provides a baseline dataset for future experiments in planta or in vivo bacterization or engineering of wild host endophytes.

Discovery of Early-Branching Wolbachia Reveals Functional Enrichment on Horizontally Transferred Genes.

Wolbachia is a widespread endosymbiont of insects and filarial nematodes that profoundly influences host biology. Wolbachia has also been reported in rhizosphere hosts, where its diversity and function remain poorly characterized. The discovery that plant-parasitic nematodes (PPNs) host Wolbachia strains with unknown roles is of interest evolutionarily, ecologically, and for agriculture as a potential target for developing new biological controls. The goal of this study was to screen communities for PPN endosymbionts and analyze genes and genomic patterns that might indicate their role. Genome assemblies revealed 1 out of 16 sampled sites had nematode communities hosting a Wolbachia strain, designated wTex, that has highly diverged as one of the early supergroup L strains. Genome features, gene repertoires, and absence of known genes for cytoplasmic incompatibility, riboflavin, biotin, and other biosynthetic functions placed wTex between mutualist C + D strains and reproductive parasite A + B strains. Functional terms enriched in group L included protoporphyrinogen IX, thiamine, lysine, fatty acid, and cellular amino acid biosynthesis, while dN/dS analysis suggested the strongest purifying selection on arginine and lysine metabolism, and vitamin B6, heme, and zinc ion binding, suggesting these as candidate roles in PPN Wolbachia. Higher dN/dS pathways between group L, wPni from aphids, wFol from springtails, and wCfeT from cat fleas suggested distinct functional changes characterizing these early Wolbachia host transitions. PPN Wolbachia had several putative horizontally transferred genes, including a lysine biosynthesis operon like that of the mitochondrial symbiont Midichloria, a spirochete-like thiamine synthesis operon shared only with wCfeT, an ATP/ADP carrier important in Rickettsia, and a eukaryote-like gene that may mediate plant systemic acquired resistance through the lysine-to-pipecolic acid system. The Discovery of group L-like variants from global rhizosphere databases suggests diverse PPN Wolbachia strains remain to be discovered. These findings support the hypothesis of plant-specialization as key to shaping early Wolbachia evolution and present new functional hypotheses, demonstrating promise for future genomics-based rhizosphere screens.

Lysis-Hi-C as a method to study polymicrobial communities and eDNA.

Microbes interact in natural communities in a spatially structured manner, particularly in biofilms and polymicrobial infections. While next generation sequencing approaches provide powerful insights into diversity, metabolic capacity, and mutational profiles of these communities, they generally fail to recover in situ spatial proximity between distinct genotypes in the interactome. Hi-C is a promising method that has assisted in analysing complex microbiomes, by creating chromatin cross-links in cells, that aid in identifying adjacent DNA, to improve de novo assembly. This study explored a modified Hi-C approach involving an initial lysis phase prior to DNA cross-linking, to test whether adjacent cell chromatin can be cross-linked, anticipating that this could provide a new avenue for study of spatial-mutational dynamics in structured microbial communities. An artificial polymicrobial mixture of Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli was lysed for 1-18 h, then prepared for Hi-C. A murine biofilm infection model was treated with sonication, mechanical lysis, or chemical lysis before Hi-C. Bioinformatic analyses of resulting Hi-C interspecies chromatin links showed that while microbial species differed from one another, generally lysis significantly increased links between species and increased the distance of Hi-C links within species, while also increasing novel plasmid-chromosome links. The success of this modified lysis-Hi-C protocol in creating extracellular DNA links is a promising first step toward a new lysis-Hi-C based method to recover genotypic microgeography in polymicrobial communities, with potential future applications in diseases with localized resistance, such as cystic fibrosis lung infections and chronic diabetic ulcers.

Co-Occurrence of Viruses, Plant Pathogens, and Symbionts in an Underexplored Hemipteran Clade.

Interactions between insect symbionts and plant pathogens are dynamic and complex, sometimes involving direct antagonism or synergy and sometimes involving ecological and evolutionary leaps, as insect symbionts transmit through plant tissues or plant pathogens transition to become insect symbionts. Hemipterans such as aphids, whiteflies, psyllids, leafhoppers, and planthoppers are well-studied plant pests that host diverse symbionts and vector plant pathogens. The related hemipteran treehoppers (family Membracidae) are less well-studied but offer a potentially new and diverse array of symbionts and plant pathogenic interactions through their distinct woody plant hosts and ecological interactions with diverse tending hymenopteran taxa. To explore membracid symbiont-pathogen diversity and co-occurrence, this study performed shotgun metagenomic sequencing on 20 samples (16 species) of treehopper, and characterized putative symbionts and pathogens using a combination of rapid blast database searches and phylogenetic analysis of assembled scaffolds and correlation analysis. Among the 8.7 billion base pairs of scaffolds assembled were matches to 9 potential plant pathogens, 12 potential primary and secondary insect endosymbionts, numerous bacteriophages, and other viruses, entomopathogens, and fungi. Notable discoveries include a divergent Brenneria plant pathogen-like organism, several bee-like Bombella and Asaia strains, novel strains of Arsenophonus-like and Sodalis-like symbionts, Ralstonia sp. and Ralstonia-type phages, Serratia sp., and APSE-type phages and bracoviruses. There were several short Phytoplasma and Spiroplasma matches, but there was no indication of plant viruses in these data. Clusters of positively correlated microbes such as yeast-like symbionts and Ralstonia, viruses and Serratia, and APSE phage with parasitoid-type bracoviruses suggest directions for future analyses. Together, results indicate membracids offer a rich palette for future study of symbiont-plant pathogen interactions.

Draft Genome Assemblies of Two Campylobacter novaezeelandiae and Four Unclassified Thermophilic Campylobacter Isolates from Canadian Agricultural Surface Water.

This report presents the draft genome sequences of two Campylobacter novaezeelandiae and four unclassified Campylobacter isolates from Canadian agricultural surface water. Phylogenomic analysis revealed that the six isolates formed unique clades, closely related to the disease-causing species C. jejuni, C. coli, and C. hepaticus.

Essential Amino Acid Enrichment and Positive Selection Highlight Endosymbiont's Role in a Global Virus-Vectoring Pest.

Host-associated microbes display remarkable convergence in genome repertoire resulting from selection to supplement missing host functions. Nutritional supplementation has been proposed in the verrucomicrobial endosymbiont Xiphinematobacter sp., which lives within a globally widespread group of plant-parasitic nematodes that vector damaging nepoviruses to plants. Only one genome sequence has been published from this symbiont, leaving unanswered questions about its diversity, host range, role, and selective pressures within its hosts. Because its hosts are exceptionally resistant to culturing, this symbiont is best studied through advanced genomic approaches. To analyze the role of Xiphinematobacter sp. in its host, sequencing was performed on nematode communities, and then genomes were extracted for comparative genomics, gene ontology enrichment tests, polymorphism analysis, de Bruijn-based genome-wide association studies, and tests of pathway- and site-specific selection on genes predicted play a role in the symbiosis. Results showed a closely clustered set of Xiphinematobacter isolates with reduced genomes of ∼917 kbp, for which a new species was proposed. Symbionts shared only 2.3% of genes with outgroup Verrucomicrobia, but comparative analyses showed high conservation of all 10 essential amino acid (EAA) biosynthesis pathways plus several vitamin pathways. These findings were supported by gene ontology enrichment tests and high polymorphisms in these pathways compared with background. Genome-wide association analysis confirmed high between-species fixation of alleles with significant functional enrichment for EAA and thiamine synthesis. Strong positive selection was detected on sites within these pathways, despite several being under increased purifying selection. Together, these results suggest that supplementation of EAAs missing in the host diet may drive this widespread symbiosis.IMPORTANCE Xiphinematobacter spp. are distinctly evolved intracellular symbionts in the phylum Verrucomicrobia, which includes the important human gut-associated microbe Akkermansia muciniphila and many highly abundant free-living soil microbes. Like Akkermansia sp., Xiphinematobacter sp. is obligately associated with the gut of its hosts, which in this case consists of a group of plant-parasitic nematodes that are among the top 10 most destructive species to global agriculture, by vectoring plant viruses. This study examined the hypothesis that the key to this symbiont's stable evolutionary association with its host is through provisioning nutrients that its host cannot make that may be lacking in the nematode's plant phloem diet, such as essential amino acids and several vitamins. The significance of our research is in demonstrating, using population genomics, the signatures of selective pressure on these hypothesized roles to ultimately learn how this independently evolved symbiont functionally mirrors symbionts of phloem-feeding insects.

Variable Abundance and Distribution of Wolbachia and Cardinium Endosymbionts in Plant-Parasitic Nematode Field Populations.

The bacterial endosymbiont Wolbachia interacts with different invertebrate hosts, engaging in diverse symbiotic relationships. Wolbachia is often a reproductive parasite in arthropods, but an obligate mutualist in filarial nematodes. Wolbachia was recently discovered in plant-parasitic nematodes, and, is thus far known in just two genera Pratylenchus and Radopholus, yet the symbiont's function remains unknown. The occurrence of Wolbachia in these economically important plant pests offers an unexplored biocontrol strategy. However, development of Wolbachia-based biocontrol requires an improved understanding of symbiont-host functional interactions and the symbiont's prevalence among nematode field populations. This study used a molecular-genetic approach to assess the prevalence of a Wolbachia lineage (wPpe) in 32 field populations of Pratylenchus penetrans. Populations were examined from eight different plant species in Washington, Oregon, and California. Nematodes were also screened for the endosymbiotic bacterium Cardinium (cPpe) that was recently shown to co-infect P. penetrans. Results identified wPpe in 9/32 and cPpe in 1/32 of P. penetrans field populations analyzed. No co-infection was observed in field populations. Wolbachia was detected in nematodes from 4/8 plant-hosts examined (raspberry, strawberry, clover, and lily), and in all three states surveyed. Cardinium was detected in nematodes from mint in Washington. In the wPpe-infected P. penetrans populations collected from raspberry, the prevalence of wPpe infection ranged from 11 to 58%. This pattern is unlike that in filarial nematodes where Wolbachia is an obligate mutualist and occurs in 100% of the host. Further analysis of wPpe-infected populations revealed female-skewed sex ratios (up to 96%), with the degree of skew positively correlating with wPpe prevalence. Uninfected nematode populations had approximately equal numbers of males and females. Comparisons of 54 wPpe 16S ribosomal RNA sequences revealed high similarity across the geographic isolates, with 45 of 54 isolates being identical at this locus. The complete absence of wPpe among some populations and low prevalence in others suggest that this endosymbiont is not an obligate mutualist of P. penetrans. The observed sex ratio bias in wPpe-infected nematode populations is similar to that observed in arthropods where Wolbachia acts as a reproductive manipulator, raising the question of a similar role in plant-parasitic nematodes.

Comparative Genomics of Wolbachia-Cardinium Dual Endosymbiosis in a Plant-Parasitic Nematode.

Wolbachia and Cardinium are among the most important and widespread of all endosymbionts, occurring in nematodes and more than half of insect and arachnid species, sometimes as coinfections. These symbionts are of significant interest as potential biocontrol agents due to their abilities to cause major effects on host biology and reproduction through cytoplasmic incompatibility, sex ratio distortion, or obligate mutualism. The ecological and metabolic effects of coinfections are not well understood. This study examined a Wolbachia-Cardinium coinfection in the plant-parasitic nematode (PPN), Pratylenchus penetrans, producing the first detailed study of such a coinfection using fluorescence in situ hybridization (FISH), polymerase chain reaction (PCR), and comparative genomic analysis. Results from FISH and single-nematode PCR showed 123/127 individuals in a focal population carried Cardinium (denoted strain cPpe), and 48% were coinfected with Wolbachia strain wPpe. Both endosymbionts showed dispersed tissue distribution with highest densities in the anterior intestinal walls and gonads. Phylogenomic analyses confirmed an early place of cPpe and long distance from a sister strain in another PPN, Heterodera glycines, supporting a long history of both Cardinium and Wolbachia in PPNs. The genome of cPpe was 1.36 Mbp with 35.8% GC content, 1,131 predicted genes, 41% having no known function, and missing biotin and lipoate synthetic capacity and a plasmid present in other strains, despite having a slightly larger genome compared to other sequenced Cardinium. The larger genome revealed expansions of gene families likely involved in host-cellular interactions. More than 2% of the genes of cPpe and wPpe were identified as candidate horizontally transferred genes, with some of these from eukaryotes, including nematodes. A model of the possible Wolbachia-Cardinium interaction is proposed with possible complementation in function for pathways such as methionine and fatty acid biosynthesis and biotin transport.

Endosymbionts of Plant-Parasitic Nematodes.

Some of the most agriculturally important plant-parasitic nematodes (PPNs) harbor endosymbionts. Extensive work in other systems has shown that endosymbionts can have major effects on host virulence and biology. This review highlights the discovery, development, and diversity of PPN endosymbionts, incorporating inferences from genomic data. Cardinium, reported from five PPN hosts to date, is characterized by its presence in the esophageal glands and other tissues, with a discontinuous distribution across populations, and genomic data suggestive of horizontal gene exchange. Xiphinematobacter occurs in at least 27 species of dagger nematode in the ovaries and gut epithelial cells, where genomic data suggest it may serve in nutritional supplementation. Wolbachia, reported in just three PPNs, appears to have an ancient history in the Pratylenchidae and displays broad tissue distribution and genomic features intermediate between parasitic and reproductive groups. Finally, a model is described that integrates these insights to explain patterns of endosymbiont replacement.

The Draft Genome of Globodera ellingtonae.

Globodera ellingtonae is a newly described potato cyst nematode (PCN) found in Idaho, Oregon, and Argentina. Here, we present a genome assembly for G. ellingtonae, a relative of the quarantine nematodes G. pallida and G. rostochiensis, produced using data from Illumina and Pacific Biosciences DNA sequencing technologies.

Genomic evidence for plant-parasitic nematodes as the earliest Wolbachia hosts.

Wolbachia, one of the most widespread endosymbionts, is a target for biological control of mosquito-borne diseases (malaria and dengue virus), and antibiotic elimination of infectious filarial nematodes. We sequenced and analyzed the genome of a new Wolbachia strain (wPpe) in the plant-parasitic nematode Pratylenchus penetrans. Phylogenomic analyses placed wPpe as the earliest diverging Wolbachia, suggesting two evolutionary invasions into nematodes. The next branches comprised strains in sap-feeding insects, suggesting Wolbachia may have first evolved as a nutritional mutualist. Genome size, protein content, %GC, and repetitive DNA allied wPpe with mutualistic Wolbachia, whereas gene repertoire analyses placed it between parasite (A, B) and mutualist (C, D, F) groups. Conservation of iron metabolism genes across Wolbachia suggests iron homeostasis as a potential factor in its success. This study enhances our understanding of this globally pandemic endosymbiont, highlighting genetic patterns associated with host changes. Combined with future work on this strain, these genomic data could help provide potential new targets for plant-parasitic nematode control.

The mitochondrial genome of Globodera ellingtonae is composed of two circles with segregated gene content and differential copy numbers.

BACKGROUND: The evolution of animal mitochondrial (mt) genomes has resulted in a highly conserved structure: a single compact circular chromosome approximately 14 to 20 kb long. Within the last two decades exceptions to this conserved structure, such as the division of the genome into multiple chromosomes, have been reported in a diverse set of metazoans. We report on the two circle multipartite mt genome of a newly described cyst nematode, Globodera ellingtonae. RESULTS: The G. ellingtonae mt genome was found to be comprised of two circles, each larger than any other multipartite circular mt chromosome yet reported, and both were larger than the single mt circle of the model nematode Caenorhabditis elegans. The genetic content of the genome was disproportionately divided between the two circles, although they shared a ~6.5 kb non-coding region. The 17.8 kb circle (mtDNA-I) contained ten protein-coding genes and two tRNA genes, whereas the 14.4 kb circle (mtDNA-II) contained two protein-coding genes, 20 tRNA genes and both rRNA genes. Perhaps correlated with this division of genetic content, the copy number of mtDNA-II was more than four-fold that of mtDNA-I in individual nematodes. The difference in copy number increased between second-stage and fourth-stage juveniles. CONCLUSIONS: The segregation of gene types to different mt circles in G. ellingtonae could provide benefit by localizing gene functional types to independent transcriptional units. This is the first report of both two-circle and several-circle mt genomes within a single genus. The differential copy number associated with this multipartite mt organization could provide a model system for deconstructing mechanisms regulating mtDNA copy number both in somatic cells and during germline development.

Comparative Genomics of a Plant-Parasitic Nematode Endosymbiont Suggest a Role in Nutritional Symbiosis.

Bacterial mutualists can modulate the biochemical capacity of animals. Highly coevolved nutritional mutualists do this by synthesizing nutrients missing from the host's diet. Genomics tools have advanced the study of these partnerships. Here we examined the endosymbiont Xiphinematobacter (phylum Verrucomicrobia) from the dagger nematode Xiphinema americanum, a migratory ectoparasite of numerous crops that also vectors nepovirus. Previously, this endosymbiont was identified in the gut, ovaries, and eggs, but its role was unknown. We explored the potential role of this symbiont using fluorescence in situ hybridization, genome sequencing, and comparative functional genomics. We report the first genome of an intracellular Verrucomicrobium and the first exclusively intracellular non-Wolbachia nematode symbiont. Results revealed that Xiphinematobacter had a small 0.916-Mb genome with only 817 predicted proteins, resembling genomes of other mutualist endosymbionts. Compared with free-living relatives, conserved proteins were shorter on average, and there was large-scale loss of regulatory pathways. Despite massive gene loss, more genes were retained for biosynthesis of amino acids predicted to be essential to the host. Gene ontology enrichment tests showed enrichment for biosynthesis of arginine, histidine, and aromatic amino acids, as well as thiamine and coenzyme A, diverging from the profiles of relatives Akkermansia muciniphilia (in the human colon), Methylacidiphilum infernorum, and the mutualist Wolbachia from filarial nematodes. Together, these features and the location in the gut suggest that Xiphinematobacter functions as a nutritional mutualist, supplementing essential nutrients that are depleted in the nematode diet. This pattern points to evolutionary convergence with endosymbionts found in sap-feeding insects.

Population genomics of a symbiont in the early stages of a pest invasion.

Invasive species often depend on microbial symbionts, but few studies have examined the evolutionary dynamics of symbionts during the early stages of an invasion. The insect Megacopta cribraria and its bacterial nutritional symbiont Candidatus Ishikawaella capsulata invaded the southeastern US in 2009. While M. cribraria was initially discovered on wild kudzu plants, it was found as a pest on soybeans within 1 year of infestation. Because prior research suggests Ishikawaella confers the pest status--that is, the ability to thrive on soybeans--in some Megacopta species, we performed a genomic study on Ishikawaella from US. Megacopta cribraria populations to understand the role of the symbiont in driving host plant preferences. We included Ishikawaella samples collected in the first days of the invasion in 2009 and from 23 locations across the insect's 2011 US range. The 0.75 Mb symbiont genome revealed only 47 fixed differences from the pest-conferring Ishikawaella in Japan, with only one amino acid change in a nutrition-provisioning gene. This similarity, along with a lack of fixed substitutions in the US symbiont population, indicates that Ishikawella likely arrived in the US capable of being a soybean pest. Analyses of allele frequency changes between 2009 and 2011 uncover signatures of both positive and negative selection and suggest that symbionts on soybeans and kudzu experience differential selection for genes related to nutrient provisioning. Our data reveal the evolutionary trajectory of an important insect-bacteria symbiosis in the early stages of an invasion, highlighting the role microbial symbionts may play in the spread of invasive species.

Description of five new Loma (Microsporidia) species in pacific fishes with redesignation of the type species Loma morhua Morrison & Sprague, 1981, based on morphological and molecular species-boundaries tests.

Five new species of Loma were described from five Pacific fishes using light-microscopic and ultrastructural features along with phylogenetic analysis of the gene sequences of ribosomal RNA (rRNA) and elongation factor 1-alpha. Morphological data revealed both qualitative and quantitative differences in developmental stages and timing, vesicles, xenoma features, and spore sizes with statistical support that differentiated Loma pacificodae n. sp. in Pacific cod, Loma wallae n. sp. in walleye pollock, Loma kenti n. sp. in Pacific tomcod, Loma lingcodae n. sp. in lingcod, and Loma richardi n. sp. in sablefish from each other and other species in the genus. Phylogenetic analyses combined with monophyly tests supported species designations, but with low resolution in two cases perhaps due to rRNA paralogs or recent speciation. Loma branchialis in haddock was shown to be separate from Loma morhua in Atlantic cod, thereby making L. morhua, and not L. branchialis, the type species. A species from brook trout was shown to be a separate species from Loma salmonae, not a variant strain selected in the laboratory. By comparison with gadid host phylogeny, these Loma species appear to have coevolved with their hosts, first colonizing the Pacific basin about 12 million years ago.

Low genetic variation in the salmon and trout parasite Loma salmonae (Microsporidia) supports marine transmission and clarifies species boundaries.

Loma salmonae is a microsporidian parasite prevalent in wild and farmed salmon species of the genus Oncorhynchus. This study compared ribosomal RNA (rDNA) and elongation factor-1 alpha (EF-1alpha) gene sequences to look for variation that may provide a basis for distinguishing populations. Specimens were collected from laboratory, captive (sea netpen farm and freshwater hatchery) and wild populations of fish. The host range included rainbow trout O. mykiss, Pacific salmon Oncorhynchus spp. and brook trout Salvelinus fontinalis from British Columbia, Prince Edward Island, Canada, from California, Colorado, Idaho, U.S.A. and from Chile. Both loci suggested that a variant in S. fontinalis (named 'SV') was a separate species. This was supported by the absence of similar variants in the source material (isolated from laboratory-held O. tshawytscha) and high divergence (1.4 to 2.3% in the rDNA and EF-1alpha) from L. salmonae in the type host and locality (0. mykiss in California). L. salmonae from freshwater and anadromous Oncorhynchus spp. were distinguished, providing a basis on which to evaluate possible sources of infection and suggesting geographic boundaries are important. Higher genetic variation occurred among samples of freshwater origin and from a sea netpen farm in Chile, suggesting these environments may present greater population diversity. Invariance in rDNA sequence across 17 samples from anadromous salmon in rivers, lakes, ocean, farms and hatcheries supports the hypothesis that marine transmission occurs and effectively prevents population substructuring caused by freshwater transmission.