Byron J. Adams

Cross-biome metagenomic analyses of soil microbial communities and their functional attributes

For centuries ecologists have studied how the diversity and functional traits of plant and animal communities vary across biomes. In contrast, we have only just begun exploring similar questions for soil microbial communities despite soil microbes being the dominant engines of biogeochemical cycles and a major pool of living biomass in terrestrial ecosystems. We used metagenomic sequencing to compare the composition and functional attributes of 16 soil microbial communities collected from cold deserts, hot deserts, forests, grasslands, and tundra. Those communities found in plant-free cold desert soils typically had the lowest levels of functional diversity (diversity of protein-coding gene categories) and the lowest levels of phylogenetic and taxonomic diversity. Across all soils, functional beta diversity was strongly correlated with taxonomic and phylogenetic beta diversity; the desert microbial communities were clearly distinct from the nondesert communities regardless of the metric used. The desert communities had higher relative abundances of genes associated with osmoregulation and dormancy, but lower relative abundances of genes associated with nutrient cycling and the catabolism of plant-derived organic compounds. Antibiotic resistance genes were consistently threefold less abundant in the desert soils than in the nondesert soils, suggesting that abiotic conditions, not competitive interactions, are more important in shaping the desert microbial communities. As the most comprehensive survey of soil taxonomic, phylogenetic, and functional diversity to date, this study demonstrates that metagenomic approaches can be used to build a predictive understanding of how microbial diversity and function vary across terrestrial biomes.

Co-variation in soil biodiversity and biogeochemistry in northern and southern Victoria Land, Antarctica

Data from six sites in Victoria Land (72–77°S) investigating co-variation in soil communities (microbial and invertebrate) with biogeochemical properties showthe influence of soil properties on habitat suitability varied among local landscapes as well as across climate gradients. Species richness of metazoan invertebrates (Nematoda, Tardigrada and Rotifera) was similar to previous descriptions in this region, though identification of three cryptic nematode species of Eudorylaimus through DNA analysis contributed to the understanding of controls over habitat preferences for individual species. Denaturing Gradient Gel Electrophoresis profiles revealed unexpectedly high diversity of bacteria. Distribution of distinct bacterial communities was associated with specific sites in northern and southern Victoria Land, as was the distribution of nematode and tardigrade species. Variation in soil metazoan communities was related to differences in soil organic matter, while bacterial diversity and community structure were not strongly correlated with any single soil property. There were no apparent correlations between metazoan and bacterial diversity, suggesting that controls over distribution and habitat suitability are different for bacterial and metazoan communities. Our results imply that top-down controls over bacterial diversity mediated by their metazoan consumers are not significant determinants of bacterial community structure and biomass in these ecosystems.

An Entomopathogenic Nematode by Any Other Name

Among the diversity of insect-parasitic nematodes, entomopathogenic nematodes (EPNs) are distinct, cooperating with insect-pathogenic bacteria to kill insect hosts. EPNs have adapted specific mechanisms to associate with and transmit bacteria to insect hosts. New discoveries have expanded this guild of nematodes and refine our understanding of the nature and evolution of insect–nematode associations. Here, we clarify the meaning of “entomopathogenic” in nematology and argue that EPNs must rapidly kill their hosts with the aid of bacterial partners and must pass on the associated bacteria to future generations.

MOLECULAR AND MORPHOMETRIC EVIDENCE FOR SEPARATE SPECIES OF UNCINARIA (NEMATODA: ANCYLOSTOMATIDAE) IN CALIFORNIA SEA LIONS AND NORTHERN FUR SEALS: HYPOTHESIS TESTING SUPPLANTS VERIFICATION

California sea lions (Zalophus californianus) and northern fur seals (Callorhinus ursinus) are each believed to host distinct hookworm species (Uncinaria spp.). However, a recent morphometric analysis suggested that a single species parasitizes multiple pinniped hosts, and that the observed differences are host-induced. To explore the systematics of these hookworms and test these competing hypotheses, we obtained nucleotide sequences of nuclear ribosomal DNA (D2/D3 28S, D18/D19 28S, and internal transcribed spacer [ITS] regions) from 20 individual hookworms parasitizing California sea lion and northern fur seal pups where their breeding grounds are sympatric. Five individuals from an allopatric population of California sea lions were also sampled for ITS-1 and D18/D19 28S sequences. The 28S D2/D3 sequences showed no diagnostic differences among hookworms sampled from individual sea lions and fur seals, whereas the 28S D18/D19 sequences had one derived (apomorphic) character demarcating hookworms from northern fur seals. ITS sequences were variable for 7 characters, with 4 derived (apomorphic) states in ITS-1 demarcating hookworms from California sea lions. Multivariate analysis of morphometric data also revealed significant differences between nematodes representing these 2 host-associated lineages. These results indicate that these hookworms represent 2 species that are not distributed indiscriminately between these host species, but instead exhibit host fidelity, evolving independently with each respective host species. This evolutionary approach to analyzing sequence data for species delimitation is contrasted with similarity-based methods that have been applied to numerous diagnostic studies of nematode parasites.

Comparing Phylogenetic Codivergence between Polyomaviruses and Their Hosts

ABSTRACT Seventy-two full genomes corresponding to nine mammalian (67 strains) and two avian (5 strains) polyomavirus species were analyzed using maximum likelihood and Bayesian methods of phylogenetic inference. Our fully resolved and well-supported (bootstrap proportions > 90%; posterior probabilities = 1.0) trees separate the bird polyomaviruses (avian polyomavirus and goose hemorrhagic polyomavirus) from the mammalian polyomaviruses, which supports the idea of spitting the genus into two subgenera. Such a split is also consistent with the different viral life strategies of each group. Simian (simian virus 40, simian agent 12 [Sa12], and lymphotropic polyomavirus) and rodent (hamster polyomavirus, mouse polyomavirus, and murine pneumotropic polyomavirus [MPtV]) polyomaviruses did not form monophyletic groups. Using our best hypothesis of polyomavirus evolutionary relationships and established host phylogenies, we performed a cophylogenetic reconciliation analysis of codivergence. Our analyses generated six optimal cophylogenetic scenarios of coevolution, including 12 codivergence events (P< 0.01), suggesting that Polyomaviridae coevolved with their avian and mammal hosts. As individual lineages, our analyses showed evidence of host switching in four terminal branches leading to MPtV, bovine polyomavirus, Sa12, and BK virus, suggesting a combination of vertical and horizontal transfer in the evolutionary history of the polyomaviruses.

Phylogeny of Meloidogyne spp. based on 18S rDNA and the intergenic region of mitochondrial DNA sequences

The 18S rDNA of 19 populations of Meloidogyne spp. was amplified and directly sequenced. The region of mitochondrial DNA, located in the 3′ portion of the gene that codes for cytochrome oxidase subunit II (COII) through a portion of the 16S rRNA (lRNA) gene, from 16 of these populations was cloned and sequenced. Heteroplasmic sequences were identified from both rDNA and mtDNA regions for several taxa. Several sequences sampled from nominal taxa differed from previously published accounts. Phylogenetic trees based on alignments of these sequences were constructed using distance, parsimony and likelihood optimality criteria. For 18S rDNA data, three main clades were identified. One well supported clade (86–91% bootstrap) included the most common and widely disseminated species, e.g.,M. arenaria, M. javanica and M. incognita, some recently described or redescribed species (M. floridensis, M. paranaensis, and M. ethiopica) plus numerous unidentified isolates. All mitotic parthenogenetic species, except for M. oryzae, were included in this clade. Other, less well supported clades included the amphimictic and facultative meiotic species M. hapla, M. microtyla, M. maritima and M. duytsi. One such clade comprised three meiotic parthenogens (M. exigua, M. graminicola and M. chitwoodi) and M. oryzae. This clade was moderately supported (77% bootstrap) but the relationships within this clade were poor. For mitochondrial DNA data, only the species in clade I from rDNA analysis, and M. hapla were analysed. These species formed a well supported clade (100% bootstrap) to the exclusion of M. mayaguensis and M. hapla. The addition of taxa and mtDNA data to publicly available records improved the discrimination sensitivity of species and atypical, non-identified, isolates.

Desiccation survival in an Antarctic nematode: molecular analysis using expressed sequenced tags

BackgroundNematodes are the dominant soil animals in Antarctic Dry Valleys and are capable of surviving desiccation and freezing in an anhydrobiotic state. Genes induced by desiccation stress have been successfully enumerated in nematodes; however we have little knowledge of gene regulation by Antarctic nematodes which can survive multiple environmental stresses. To address this problem we investigated the genetic responses of a nematode species, Plectus murrayi, that is capable of tolerating Antarctic environmental extremes, in particular desiccation and freezing. In this study, we provide the first insight into the desiccation induced transcriptome of an Antarctic nematode through cDNA library construction and suppressive subtractive hybridization.ResultsWe obtained 2,486 expressed sequence tags (ESTs) from 2,586 clones derived from the cDNA library of desiccated P. murrayi. The 2,486 ESTs formed 1,387 putative unique transcripts of which 523 (38%) had matches in the model-nematode Caenorhabditis elegans, 107 (7%) in nematodes other than C. elegans, 153 (11%) in non-nematode organisms and 605 (44%) had no significant match to any sequences in the current databases. The 1,387 unique transcripts were functionally classified by using Gene Ontology (GO) hierarchy and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. The results indicate that the transcriptome contains a group of transcripts from diverse functional areas. The subtractive library of desiccated nematodes showed 80 transcripts differentially expressed during desiccation stress, of which 28% were metabolism related, 19% were involved in environmental information processing, 28% involved in genetic information processing and 21% were novel transcripts. Expression profiling of 14 selected genes by quantitative Real-time PCR showed 9 genes significantly up-regulated, 3 down-regulated and 2 continuously expressed in response to desiccation.ConclusionThe establishment of a desiccation EST collection for Plectus murrayi, a useful model in assessing the structural, physiological, biochemical and genetic aspects of multiple stress tolerance, is an important step in understanding the genome level response of this nematode to desiccation stress. The type of transcript analysis performed in this study sets the foundation for more detailed functional and genome level analyses of the genes involved in desiccation tolerance in nematodes.

Survey of entomopathogenic nematodes and fungi endemic to pecan orchards of the Southeastern United States and their virulence to the pecan weevil (Coleoptera: Curculionidae).

Abstract The pecan weevil, Curculio caryae (Horn), is a major pest of pecans in the Southeastern United States. Entomopathogenic nematodes and fungi are potential alternatives to chemical insecticides for C. caryae control. Our objective was to survey pecan orchards in the southeastern United States for entomopathogenic nematodes and fungi and determine the virulence of the new isolates to C. caryae larvae. Soil was collected from 105 sites in 21 orchards in Arkansas, Georgia, Louisiana, and Mississippi. Entomopathogens were isolated by exposing soil to C. caryae and greater wax moth larvae, Galleria mellonella, (L.). We isolated entomopathogenic fungi and nematodes from 16 and 6 of the 21 orchards surveyed, respectively. The entomopathogenic fungi included Beauveria bassiana (Balsamo) Vuillemin and Metarhizium anisopliae (Metschnikoff) Sorokin, and nematodes included Heterorhabditis bacteriophora Poinar, Steinernema carpocapsae (Weiser), Steinernema glaseri (Steiner), and Steinernema rarum (Doucet). This is the first report of Steinernema rarum in the United States. Soil characteristics in orchards were analyzed for pH, organic matter, and nutrients; we detected a negative relationship between fungal occurrence and manganese levels in soil and a positive relationship between M. anisopliae occurrence and calcium or magnesium levels. In laboratory assays, virulence of 15 nematode and 22 fungal isolates to C. caryae larvae was tested in small plastic cups containing soil. Results indicated poor susceptibility of the C. caryae larvae to entomopathogenic nematodes. Several fungal isolates that caused significantly higher mortality in C. caryae larvae than other strains (including a commercial strain of B. bassiana) should be investigated further as potential control agents of C. caryae.

Comparative phylogeography of codistributed species of Chilean Liolaemus (Squamata: Tropiduridae) from the central-southern Andean range

In this study, we used a recently developed supertrees method to test for shared phylogeographical signal in partially overlapping geographical ranges of lizards of the genus Liolaemus from the Andean Range in south‐central Chile. We reconstruct mtDNA gene trees for three partially codistributed species (Liolaemus tenuis, L. lemniscatus and L. pictus), and our sampling effort is sufficient to allow statistical tests of shared signal between the combinations L. tenuis–L. pictus, and L. tenuis–L. lemniscatus. For both combinations, standardized maximum agreement subtrees scores showed statistically significant signal for shared pattern in regions of overlap, as evaluated by randomization tests (P < 0.001 and < 0.05, respectively). The matrix representation with parsimony tree obtained from the combination of the three different gene trees revealed concordant phylogeographical associations of all species, and was consistent with the geographical association of intraspecific haploclades with three Chilean bioclimatic zones. A multidimensional scaling analysis of several climate variables showed highly significant differences among these zones, which further suggests that they may have contributed to similar patterns of intraspecific divergence across all three species. In the mesomorphic zone in Central Chile, the species L. tenuis and L. lemniscatus may have codiverged in response to shared orogenic vicariant events, which likely predominated over climatic events associated with cycles of glacial advance and retreat. In the hygromorphic zone in southern Chile, however, glacial cycles likely predominated in structuring the phylogeographical histories of L. tenuis and L. pictus, although important ecological differences between these two caution against broad generalizations at this point.

Phylogenetic analysis identifies the invertebrate pathogen Helicosporidium sp. as a green alga (Chlorophyta).

Historically, the invertebrate pathogens of the genus Helicosporidium were considered to be either protozoa or fungi, but the taxonomic position of this group has not been considered since 1931. Recently, a Helicosporidium sp., isolated from the blackfly Simulium jonesi Stone & Snoddy (Diptera: Simuliidae), has been amplified in the heterologous host Helicoverpa zea. Genomic DNA has been extracted from gradient-purified cysts. The 185, 28S and 5.8S regions of the Helicosporidium rDNA, as well as partial sequences of the actin and beta-tubulin genes, were amplified by PCR and sequenced. Comparative analysis of these nucleotide sequences was performed using neighbour-joining and maximum-parsimony methods. All inferred phylogenetic trees placed Helicosporidium sp. among the green algae (Chlorophyta), and this association was supported by bootstrap and parsimony jackknife values. Phylogenetic analysis focused on the green algae depicted Helicosporidium sp. as a close relative of Prototheca wickerhamii and Prototheca zopfii (Chlorophyta, Trebouxiophyceae), two achlorophylous, pathogenic green algae. On the basis of this phylogenetic analysis, Helicosporidium sp. is clearly neither a protist nor a fungus, but appears to be the first described algal invertebrate pathogen. These conclusions lead us to propose the transfer of the genus Helicosporidium to Chlorophyta, Trebouxiophyceae.