Large-Scale Genomic Analysis Suggests a Neutral Punctuated Dynamics of Transposable Elements in Bacterial Genomes

Insertion sequences (IS) are the simplest and most abundant form of transposable DNA found in bacterial genomes. When present in multiple copies, it is thought that they can promote genomic plasticity and genetic exchange, thus being a major force of evolutionary change. The main processes that determine IS content in genomes are, though, a matter of debate. In this work, we take advantage of the large amount of genomic data currently available and study the abundance distributions of 33 IS families in 1811 bacterial chromosomes. This allows us to test simple models of IS dynamics and estimate their key parameters by means of a maximum likelihood approach. We evaluate the roles played by duplication, lateral gene transfer, deletion and purifying selection. We find that the observed IS abundances are compatible with a neutral scenario where IS proliferation is controlled by deletions instead of purifying selection. Even if there may be some cases driven by selection, neutral behavior dominates over large evolutionary scales. According to this view, IS and hosts tend to coexist in a dynamic equilibrium state for most of the time. Our approach also allows for a detection of recent IS expansions, and supports the hypothesis that rapid expansions constitute transient events—punctuations—during which the state of coexistence of IS and host becomes perturbated.     

Variation in the OC Locus of Acinetobacter baumannii Genomes Predicts Extensive Structural Diversity in the Lipooligosaccharide

Lipooligosaccharide (LOS) is a complex surface structure that is linked to many pathogenic properties of Acinetobacter baumannii. In A. baumannii, the genes responsible for the synthesis of the outer core (OC) component of the LOS are located between ilvE and aspS. The content of the OC locus is usually variable within a species, and examination of 6 complete and 227 draft A. baumannii genome sequences available in GenBank non-redundant and Whole Genome Shotgun databases revealed nine distinct new types, OCL4-OCL12, in addition to the three known ones. The twelve gene clusters fell into two distinct groups, designated Group A and Group B, based on similarities in the genes present. OCL6 (Group B) was unique in that it included genes for the synthesis of L-Rhamnosep. Genetic exchange of the different configurations between strains has occurred as some OC forms were found in several different sequence types (STs). OCL1 (Group A) was the most widely distributed being present in 18 STs, and OCL6 was found in 16 STs. Variation within clones was also observed, with more than one OC locus type found in the two globally disseminated clones, GC1 and GC2, that include the majority of multiply antibiotic resistant isolates. OCL1 was the most abundant gene cluster in both GC1 and GC2 genomes but GC1 isolates also carried OCL2, OCL3 or OCL5, and OCL3 was also present in GC2. As replacement of the OC locus in the major global clones indicates the presence of sub-lineages, a PCR typing scheme was developed to rapidly distinguish Group A and Group B types, and to distinguish the specific forms found in GC1 and GC2 isolates.     

The Effect and Relative Importance of Neutral Genetic Diversity for Predicting Parasitism Varies across Parasite Taxa

Understanding factors that determine heterogeneity in levels of parasitism across individuals is a major challenge in disease ecology. It is known that genetic makeup plays an important role in infection likelihood, but the mechanism remains unclear as does its relative importance when compared to other factors. We analyzed relationships between genetic diversity and macroparasites in outbred, free-ranging populations of raccoons (Procyon lotor). We measured heterozygosity at 14 microsatellite loci and modeled the effects of both multi-locus and single-locus heterozygosity on parasitism using an information theoretic approach and including non-genetic factors that are known to influence the likelihood of parasitism. The association of genetic diversity and parasitism, as well as the relative importance of genetic diversity, differed by parasitic group. Endoparasite species richness was better predicted by a model that included genetic diversity, with the more heterozygous hosts harboring fewer endoparasite species. Genetic diversity was also important in predicting abundance of replete ticks (Dermacentor variabilis). This association fit a curvilinear trend, with hosts that had either high or low levels of heterozygosity harboring fewer parasites than those with intermediate levels. In contrast, genetic diversity was not important in predicting abundance of non-replete ticks and lice (Trichodectes octomaculatus). No strong single-locus effects were observed for either endoparasites or replete ticks. Our results suggest that in outbred populations multi-locus diversity might be important for coping with parasitism. The differences in the relationships between heterozygosity and parasitism for the different parasites suggest that the role of genetic diversity varies with parasite-mediated selective pressures.     

Genetic Diversity and Abundance of Flavobacterial Proteorhodopsin in China Seas

Proteorhodopsin (PR) genes related to Flavobacteria were found to be highly diverse in the East and South China seas and displayed a distinct geographic pattern, which appeared to reflect cold versus warm adaptation when Global Oceanic Sampling database metagenomic data were included. Flavobacterial PR genes were more abundant offshore than nearshore, implying that inheritance of the PR gene could be important for Flavobacteria living in the oligotrophic environment.     

Distribution and Genetic Diversity of Bacteriocin Gene Clusters in Rumen Microbial Genomes

Some species of ruminal bacteria are known to produce antimicrobial peptides, but the screening procedures have mostly been based on in vitro assays using standardized methods. Recent sequencing efforts have made available the genome sequences of hundreds of ruminal microorganisms. In this work, we performed genome mining of the complete and partial genome sequences of 224 ruminal bacteria and 5 ruminal archaea to determine the distribution and diversity of bacteriocin gene clusters. A total of 46 bacteriocin gene clusters were identified in 33 strains of ruminal bacteria. Twenty gene clusters were related to lanthipeptide biosynthesis, while 11 gene clusters were associated with sactipeptide production, 7 gene clusters were associated with class II bacteriocin production, and 8 gene clusters were associated with class III bacteriocin production. The frequency of strains whose genomes encode putative antimicrobial peptide precursors was 14.4%. Clusters related to the production of sactipeptides were identified for the first time among ruminal bacteria. BLAST analysis indicated that the majority of the gene clusters (88%) encoding putative lanthipeptides contained all the essential genes required for lanthipeptide biosynthesis. Most strains of Streptococcus (66.6%) harbored complete lanthipeptide gene clusters, in addition to an open reading frame encoding a putative class II bacteriocin. Albusin B-like proteins were found in 100% of the Ruminococcus albus strains screened in this study. The in silico analysis provided evidence of novel biosynthetic gene clusters in bacterial species not previously related to bacteriocin production, suggesting that the rumen microbiota represents an underexplored source of antimicrobial peptides.     

Shifts in Abundance and Diversity of Mobile Genetic Elements after the Introduction of Diverse Pesticides into an On-Farm Biopurification System over the Course of a Year

Biopurification systems (BPS) are used on farms to control pollution by treating pesticide-contaminated water. It is assumed that mobile genetic elements (MGEs) carrying genes coding for enzymes involved in degradation might contribute to the degradation of pesticides. Therefore, the composition and shifts of MGEs, in particular, of IncP-1 plasmids carried by BPS bacterial communities exposed to various pesticides, were monitored over the course of an agricultural season. PCR amplification of total community DNA using primers targeting genes specific to different plasmid groups combined with Southern blot hybridization indicated a high abundance of plasmids belonging to IncP-1, IncP-7, IncP-9, IncQ, and IncW, while IncU and IncN plasmids were less abundant or not detected. Furthermore, the integrase genes of class 1 and 2 integrons (intI1, intI2) and genes encoding resistance to sulfonamides (sul1, sul2) and streptomycin (aadA) were detected and seasonality was revealed. Amplicon pyrosequencing of the IncP-1 trfA gene coding for the replication initiation protein revealed high IncP-1 plasmid diversity and an increase in the abundance of IncP-1β and a decrease in the abundance of IncP-1ε over time. The data of the chemical analysis showed increasing concentrations of various pesticides over the course of the agricultural season. As an increase in the relative abundances of bacteria carrying IncP-1β plasmids also occurred, this might point to a role of these plasmids in the degradation of many different pesticides.     

近缘物种的遗传多样性及其亲缘关系的取样策略研究

取样策略的问题目前大多停留于对单一物种的种质资源研究.然而,对野生近缘植物取样策略的研究,不仅有利于准确快捷地阐明物种间的系统发育关系,而且对于了解遗传多样性分布状况,制定野外考察、材料收集取样及保护策略均有重要的理论指导意义.以巴山松及其近缘种为例,利用cpSSR(叶绿体微卫星)和AFLP2种分子标记对其取样策略和统计方法进行分析,揭示居群取样个体数和基因位点数与遗传多样性的关系以及基因(引物)和系统树构建方法对亲缘关系确定的影响.研究表明:(1)居群取样个体数和基因位点数差异对遗传多样性影响并不显著,但当居群取样个体数为30个左右,单引物基因位点数为60个以上,总位点数为480个以上时,所得遗传多样性及亲缘关系较为准确可靠.(2)较多数目的基因(引物)对于得到较为准确可靠的亲缘关系分析是十分必要的.(3)系统树构建方法对近缘种亲缘关系的确定有一定的影响.     

Application of Sequence-Specific Labeled 16S rRNA Gene Oligonucleotide Probes for Genetic Profiling of Cyanobacterial Abundance and Diversity by Array Hybridization

DNA sequence information for the small-subunit rRNA gene (16S rDNA) obtained from cyanobacterial cultures was used to investigate the presence of cyanobacteria and their abundance in natural habitats. Eight planktonic communities developing in lakes characterized by relatively low algal biomass (mesotrophic) and in lakes with correspondingly high biomass (eutrophic) were selected for the study. The organismal compositions of the water samples were analyzed genetically, using multiplex sequence-specific labeling of oligonucleotide probes targeted to 16S rDNA and subsequent hybridization of the labeled probes to their respective complements spotted onto a solid support (DNA array). Ten probes were established to determine the relative abundances of the discernible cyanobacteria encountered in the selected lakes. The probes were generally specific for their targets, as determined through analyses of clone cultures. Reproducible abundance profiles were established for the lakes investigated in the subsequent analyses of natural cyanobacterial communities. The results from the genetic analyses were then compared with information obtained from standard hydrobiological and hydrochemical analyses. Qualitatively, there were relatively good correlations among the groups of organisms (Nostoc, Microcystis, and Planktothrix species) found in the different lakes. The levels of correlation were lower for the quantitative data. This may, however, be due to differences in sample processing technique. The conclusions from these comparisons are that the genetic abundance profiles may provide a foundation for separating and quantifying genetically distinct groups of cyanobacteria in their natural habitats.     

Conservation and Periodicity of DNA Bend Sites in Eukaryotic Genomes

DNA bend sites appear every 680 bp on average in the human -and ß-globin gene regions. Although most of theirmolecular nature has not been unraveled, a potential bend coresequence A₂N₈A₂N₈A₂ (A/A/A) and its complementary T₂N₈T₂N₈T₂(T/T/T) appeared preferentially either in or very close to mostof the bend sites, whereas other combinations of A₂ and T₂ dinucleotides,A/T/T + A/A/T, T/T/A + T/A/A and A/T/A + T/A/T, did not. Thedistances between any two of the core sequences in the entireß-globin locus showed a strong bias to a length of701–800 bp and multiples thereof, suggesting that thereis periodicity throughout the locus. This bias was not foundfor other combinations of A₂ and T₂. Again, this periodicitywas identified in many eukaryotic genes, whereas the tendencywas absent in mRNAs and prokaryotic as well as viral genomes.     

Genetic Diversity of Eukaryotic Microorganisms in Lake Taihu,a Large Shallow Subtropical Lake in China

We investigated the genetic diversity of eukaryotic microorganisms (0.8-20 mum) by sequencing cloned 18S rRNA genes in six genetic libraries constructed from six locations in Lake Taihu, a large shallow subtropical lake in China. Genetic libraries of eukaryotic ribosomal RNA were screened by restriction fragment length polymorphism (RFLP) analysis, and one clone representative of each RFLP pattern was partially sequenced. A total of 528 clones were clustered into 165 RFLP patterns and finally into 131 operational taxonomic unit (OTUs). Phylogenetic analysis revealed that each library included many unique OTUs, as well as members of distantly related phylogenetic groups. A majority of the clones were from alveolates, stramenopiles, cercozoa, cryptophytes, chlorophytes, and fungi, with members of choanoflagellida, euglenida, centroheliozoa, ancyromonadidae, ichthyosporea, and kathablepharid representing a minor fraction of the library. Six OTUs (15 clones) were not related to any known eukaryotic group. Canonical correspondence analysis suggested that the differences in eukaryotic microorganism community composition of in the six regions were partially related to trophic status, sediment resuspension, and top-down regulation by metazooplankton.     

Abundance and Genetic Diversity of nifH Gene Sequences in Anthropogenically Affected Brazilian Mangrove Sediments

Although mangroves represent ecosystems of global importance, the genetic diversity and abundance of functional genes that are key to their functioning scarcely have been explored. Here, we present a survey based on the nifH gene across transects of sediments of two mangrove systems located along the coast line of São Paulo state (Brazil) which differed by degree of disturbance, i.e., an oil-spill-affected and an unaffected mangrove. The diazotrophic communities were assessed by denaturing gradient gel electrophoresis (DGGE), quantitative PCR (qPCR), and clone libraries. The nifH gene abundance was similar across the two mangrove sediment systems, as evidenced by qPCR. However, the nifH-based PCR-DGGE profiles revealed clear differences between the mangroves. Moreover, shifts in the nifH gene diversities were noted along the land-sea transect within the previously oiled mangrove. The nifH gene diversity depicted the presence of nitrogen-fixing bacteria affiliated with a wide range of taxa, encompassing members of the Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Firmicutes, and also a group of anaerobic sulfate-reducing bacteria. We also detected a unique mangrove-specific cluster of sequences denoted Mgv-nifH. Our results indicate that nitrogen-fixing bacterial guilds can be partially endemic to mangroves, and these communities are modulated by oil contamination, which has important implications for conservation strategies.     

High Genetic Diversity and Novelty in Eukaryotic Plankton Assemblages Inhabiting Saline Lakes in the Qaidam Basin

Saline lakes are intriguing ecosystems harboring extremely productive microbial communities in spite of their extreme environmental conditions. We performed a comprehensive analysis of the genetic diversity (18S rRNA gene) of the planktonic microbial eukaryotes (nano- and picoeukaryotes) in six different inland saline lakes located in the Qaidam Basin. The novelty level are high, with about 11.23% of the whole dataset showing <90% identity to any previously reported sequence in GenBank. At least 4 operational taxonomic units (OTUs) in mesosaline lakes, while up to eighteen OTUs in hypersaline lakes show very low CCM and CEM scores, indicating that these sequences are highly distantly related to any existing sequence. Most of the 18S rRNA gene sequence reads obtained in investigated mesosaline lakes is closely related to Holozoa group (48.13%), whereas Stramenopiles (26.65%) and Alveolates (10.84%) are the next most common groups. Hypersaline lakes in the Qaidam Basin are also dominated by Holozoa group, accounting for 26.65% of the total number of sequence reads. Notably, Chlorophyta group are only found in high abundance in Lake Gasikule (28.00%), whereas less represented in other hypersaline lakes such as Gahai (0.50%) and Xiaochaidan (1.15%). Further analysis show that the compositions of planktonic eukaryotic assemblages are also most variable between different sampling sites in the same lake. Out of the parameters, four show significant correlation to this CCA: altitude, calcium, sodium and potassium concentrations. Overall, this study shows important gaps in the current knowledge about planktonic microbial eukaryotes inhabiting Qaidam Basin (hyper) saline water bodies. The identified diversity and novelty patterns among eukaryotic plankton assemblages in saline lake are of great importance for understanding and interpreting their ecology and evolution.     

Vulture Genomes Reveal Molecular Adaptations Underlying Obligate Scavenging and Low Levels of Genetic Diversity

Obligate scavenging on the dead and decaying animal matter is a rare dietary specialization that in extant vertebrates is restricted to vultures. These birds perform essential ecological services, yet many vulture species have undergone recent steep population declines and are now endangered. To test for molecular adaptations underlying obligate scavenging in vultures, and to assess whether genomic features might have contributed to their population declines, we generated high-quality genomes of the Himalayan and bearded vultures, representing both independent origins of scavenging within the Accipitridae, alongside a sister taxon, the upland buzzard. By comparing our data to published sequences from other birds, we show that the evolution of obligate scavenging in vultures has been accompanied by widespread positive selection acting on genes underlying gastric acid production, and immunity. Moreover, we find evidence of parallel molecular evolution, with amino acid replacements shared among divergent lineages of these scavengers. Our genome-wide screens also reveal that both the Himalayan and bearded vultures exhibit low levels of genetic diversity, equating to around a half of the mean genetic diversity of other bird genomes examined. However, demographic reconstructions indicate that population declines began at around the Last Glacial Maximum, predating the well-documented dramatic declines of the past three decades. Taken together, our genomic analyses imply that vultures harbor unique adaptations for processing carrion, but that modern populations are genetically depauperate and thus especially vulnerable to further genetic erosion through anthropogenic activities.     

Broad Concordance in the Spatial Distribution of Adaptive and Neutral Genetic Variation across an Elevational Gradient in Deer Mice

When species are continuously distributed across environmental gradients, the relative strength of selection and gene flow shape spatial patterns of genetic variation, potentially leading to variable levels of differentiation across loci. Determining whether adaptive genetic variation tends to be structured differently than neutral variation along environmental gradients is an open and important question in evolutionary genetics. We performed exome-wide population genomic analysis on deer mice sampled along an elevational gradient of nearly 4,000 m of vertical relief. Using a combination of selection scans, genotype−environment associations, and geographic cline analyses, we found that a large proportion of the exome has experienced a history of altitude-related selection. Elevational clines for nearly 30% of these putatively adaptive loci were shifted significantly up- or downslope of clines for loci that did not bear similar signatures of selection. Many of these selection targets can be plausibly linked to known phenotypic differences between highland and lowland deer mice, although the vast majority of these candidates have not been reported in other studies of highland taxa. Together, these results suggest new hypotheses about the genetic basis of physiological adaptation to high altitude, and the spatial distribution of adaptive genetic variation along environmental gradients.     

RNA Regulatory Elements in the Genomes of Simple Retroviruses

The RNA genomes of simple retroviruses encode three genes (gag, pol,andenv) which are required for replication. In addition, there are at least three well-definedcis-acting structures which regulate important aspects of the viral life cycle. The packaging signal at the 5′ end of the RNA tags the genomic RNA for specific encapsidation into assembling virus. Since viral Env proteins are translated from spliced mRNAs,cis-acting splicing signals ensure that the proper ratio of spliced and unspliced viral RNAs is present in the infected cell. Finally,cis-acting elements at the 3′ end of the genome promote the export of unspliced RNAs from the nucleus for translation and encapsidation.     

Asymmetry Indices for Analysis and Prediction of Replication Origins in Eukaryotic Genomes

DNA replication was recently shown to induce the formation of compositional skews in the genomes of the yeasts Saccharomyces cerevisiae and Kluyveromyces lactis. In this work, I have characterized further GC and TA skew variations in the vicinity of S. cerevisiae replication origins and termination sites, and defined asymmetry indices for origin analysis and prediction. The presence of skew jumps at some termination sites in the S. cerevisiae genome was established. The majority of S. cerevisiae replication origins are marked by an oriented consensus sequence called ACS, but no evidence could be found for asymmetric origin firing that would be linked to ACS orientation. Asymmetry indices related to GC and TA skews were defined, and a global asymmetry index I_GC,TA was described. I_GC,TA was found to strongly correlate with origin efficiency in S. cerevisiae and to allow the determination of sets of intergenes significantly enriched in origin loci. The generalized use of asymmetry indices for origin prediction in naive genomes implies the determination of the direction of the skews, i.e. the identification of which strand, leading or lagging, is enriched in G and which one is enriched in T. Recent work indicates that in Candida albicans and in several related species, centromeres contain early and efficient replication origins. It has been proposed that the skew jumps observed at these positions would reflect the activity of these origins, thus allowing to determine the direction of the skews in these genomes. However, I show here that the skew jumps at C. albicans centromeres are not related to replication and that replication-associated GC and TA skews in C. albicans have in fact the opposite directions of what was proposed.     

Genetic Diversity of Eukaryotic Plankton Assemblages in Eastern Tibetan Lakes Differing by their Salinity and Altitude

Eukaryotic plankton assemblages in 11 high-mountain lakes located at altitudes of 2,817 to 5,134 m and over a total area of ca. one million square kilometers on the Eastern Tibet Plateau, spanning a salinity gradient from 0.2 (freshwater) to 187.1 g l⁻¹ (hypersaline), were investigated by cultivation independent methods. Two 18S rRNA gene-based fingerprint approaches, i.e., the terminal restriction fragment length polymorphism and denaturing gradient gel electrophoresis (DGGE) with subsequent band sequencing were applied. Samples of the same lake type (e.g., freshwater) generally shared more of the same bands or T-RFs than samples of different types (e.g., freshwater versus saline). However, a certain number of bands or T-RFs among the samples within each lake were distinct, indicating the potential presence of significant genetic diversity within each lake. PCA indicated that the most significant environmental gradient among the investigated lakes was salinity. The observed molecular profiles could be further explained (17–24%) by ion percentage of chloride, carbonate and bicarbonate, and sulfate, which were also covaried with change of altitude and latitude. Sequence analysis of selected major DGGE bands revealed many sequences (largely protist) that are not related to any known cultures but to uncultured eukaryotic picoplankton and unidentified eukaryotes. One fourth of the retrieved sequences showed ≤97% similarity to the closest sequences in the GenBank. Sequences related to well-known heterotrophic nanoflagellates were not retrieved from the DGGE gels. Several groups of eukaryotic plankton, which were found worldwide and detected in low land lakes, were also detected in habitats located above 4,400 m, suggesting a cosmopolitan distribution of these phylotypes. Collectively, our study suggests that there was a high beta-diversity of eukaryotic plankton assemblages in the investigated Tibetan lakes shaped by multiple geographic and environmental factors.     

Genetic Diversity of Grasspea and Its Relative Species Revealed by SSR Markers

The study of genetic diversity between Lathyrus sativus L. and its relative species may yield fundamental insights into evolutionary history and provide options to meet the challenge of climate changes. 30 SSR loci were employed to assess the genetic diversity and population structure of 283 individuals from wild and domesticated populations from Africa, Europe, Asia and ICARDA. The allele number per loci ranged from 3 to 14. The average gene diversity index and average polymorphism information content (PIC) was 0.5340 and 0.4817, respectively. A model based population structure analysis divided the germplasm resources into three subgroups: the relative species, the grasspea from Asia, and the grasspea from Europe and Africa. The UPGMA dendrogram and PCA cluster also demonstrated that Asian group was convincingly separated from the other group. The AMOVA result showed that the cultivated species was quite distinct from its relative species, however a low level of differentiation was revealed among their geographic origins. In all, these results provided a molecular basis for understanding genetic diversity of L. sativus and its relatives.     

Abundance and Diversity of Archaea in Heavy-Metal-Contaminated Soils

The impact of heavy-metal contamination on archaean communities was studied in soils amended with sewage sludge contaminated with heavy metals to varying extents. Fluorescent in situ hybridization showed a decrease in the percentage of Archaea from 1.3% ± 0.3% of 4′,6-diamidino-2-phenylindole-stained cells in untreated soil to below the detection limit in soils amended with heavy metals. A comparison of the archaean communities of the different plots by denaturing gradient gel electrophoresis revealed differences in the structure of the archaean communities in soils with increasing heavy-metal contamination. Analysis of cloned 16S ribosomal DNA showed close similarities to a unique and globally distributed lineage of the kingdom Crenarchaeota that is phylogenetically distinct from currently characterized crenarchaeotal species.