Extensive geographic mosaicism in avian influenza viruses from gulls in the northern hemisphere

Due to limited interaction of migratory birds between Eurasia and America, two independent avian influenza virus (AIV) gene pools have evolved. There is evidence of low frequency reassortment between these regions, which has major implications in global AIV dynamics. Indeed, all currently circulating lineages of the PB1 and PA segments in North America are of Eurasian origin. Large-scale analyses of intercontinental reassortment have shown that viruses isolated from Charadriiformes (gulls, terns, and shorebirds) are the major contributor of these outsider events. To clarify the role of gulls in AIV dynamics, specifically in movement of genes between geographic regions, we have sequenced six gull AIV isolated in Alaska and analyzed these along with 142 other available gull virus sequences. Basic investigations of host species and the locations and times of isolation reveal biases in the available sequence information. Despite these biases, our analyses reveal a high frequency of geographic reassortment in gull viruses isolated in America. This intercontinental gene mixing is not found in the viruses isolated from gulls in Eurasia. This study demonstrates that gulls are important as vectors for geographically reassorted viruses, particularly in America, and that more surveillance effort should be placed on this group of birds.     

Sequence analysis of marine virus communities reveals that groups of related algal viruses are widely distributed in nature

Algal-virus-specific PCR primers were used to amplify DNA polymerase (pol) gene fragments from geographically isolated natural virus communities. Natural algal virus communities were obtained from coastal sites in the Pacific Ocean in British Columbia, Canada, and the Southern Ocean near the Antarctic peninsula. Genetic fingerprints of algal virus communities were generated using denaturing gradient gel electrophoresis (DGGE). Sequencing efforts recovered 33 sequences from the gradient gel. Of the 33 sequences examined, 25 encoded a conserved amino acid motif indicating that the sequences were pol gene fragments. Furthermore, the 25 pol sequences were related to pol gene fragments from known algal viruses. In addition, similar virus sequences (>98% sequence identity) were recovered from British Columbia and Antarctica. Results from this study demonstrate that DGGE with degenerate primers can be used to qualitatively fingerprint and assess genetic diversity in specific subsets of natural virus communities and that closely related viruses occur in distant geographic locations. DGGE is a powerful tool for genetically fingerprinting natural virus communities and may be used to examine how specific components of virus communities respond to experimental manipulations.     

1968−2014年中国甲型H3N2流感病毒PB1基因进化分析

【目的】分析季节性H3N2流感病毒PB1基因序列的变异情况,揭示H3N2流感病毒PB1基因的分子特征与进化趋势。【方法】对1968?2014年中国地区82株人H3N2毒株、2012?2014年江苏省分离的81株甲型H3N2流感病毒、6株SIV和4株AIV H3N2亚型PB1、PB1-F2基因进行分子进化分析。【结果】1968?2014年中国H3N2流感毒株PB1核苷酸和氨基酸相似性分别为90.91%?100%和96.91%?100%。系统进化树分析,1968?2014年共173株H3N2流感病毒总体上分为4个分支,2002?2014年分离毒株位于第IV分支上,1968?1994年分离毒株位于第II和III分支;猪源H3N2亚型分布于第I、II、IV分支上;分子特征显示PB1氨基酸52、113、179、216、576、586、619、621、709位在2002年以后发生适应性改变,替换了原来的氨基酸;PB1-F2基因编码截断型蛋白长度有52、34、25、24、11 aa (猪源),PB1-F2蛋白毒力关键位点上未出现高致病性特征突变。【结论】自1968年起H3N2亚型PB1基因变异逐步趋于稳定,且PB1-F2截断型毒株正逐渐成为一类新的进化特征,但PB1基因与其他亚型之间发生重配以及关键毒力位点的变异仍应是监测的重点。     

Wide geographic distribution of bacteriophages that lyse the same indigenous freshwater isolate (Sphingomonas sp. strain B18)

An indigenous freshwater bacterium (Sphingomonas sp. strain B18) from Lake Plubetasee (Schleswig-Holstein, Germany) was used to isolate 44 phages from 13 very different freshwater and brackish habitats in distant geographic areas. This bacterial strain was very sensitive to a broad spectrum of phages from different aquatic environments. Phages isolated from geographically distant aquatic habitats, but also those from the same sample, were diverse with respect to morphology and restriction pattern. Some phages were widely distributed, while different types coexisted in the same sample. It was concluded that phages could be a major factor in shaping the structure of bacterial communities and maintaining a high bacterial diversity.     

Genetic diversity in marine algal virus communities as revealed by sequence analysis of DNA polymerase genes.

Algal-virus-specific PCR primers were used to amplify DNA polymerase gene (pol) fragments (683 to 689 bp) from the virus-sized fraction (0.02 to 0.2 microns) concentrated from inshore and offshore water samples collected from the Gulf of Mexico. Algal-virus-like DNA pol genes were detected in five samples collected from the surface and deep chlorophyll maximum. PCR products from an offshore station were cloned, and the genetic diversity of 33 fragments was examined by restriction fragment length polymorphism and sequence analysis. The five different genotypes or operational taxonomic units (OTUs) that were identified on the basis of restriction fragment length polymorphism banding patterns were present in different relative abundances (9 to 34%). One clone from each OTU was sequenced, and phylogenetic analysis showed that all of the OTUs fell within the family Phycodnaviridae. Four of the OTUs fell within a group of viruses (MpV) which infect the photosynthetic picoplankter Micromonas pusilla. The genetic diversity among these genotypes was as large as that previously found for MpV isolates from different oceans. The remaining genotype formed its own clade between viruses which infect M. pusilla and Chrysochromulina brevifilum. These results imply that marine virus communities contain a diverse assemblage of MpV-like viruses, as well as other unknown members of the Phycodnaviridae.     

Evolutionary analysis of the influenza A virus M gene with comparison of the M1 and M2 proteins

Phylogenetic analysis of 42 membrane protein (M) genes of influenza A viruses from a variety of hosts and geographic locations showed that these genes have evolved into at least four major host-related lineages: (i) A/Equine/prague/56, which has the most divergent M gene; (ii) a lineage containing only H13 gull viruses; (iii) a lineage containing both human and classical swine viruses; and (iv) an avian lineage subdivided into North American avian viruses (including recent equine viruses) and Old World avian viruses (including avianlike swine strains). The M gene evolutionary tree differs from those published for other influenza virus genes (e.g., PB1, PB2, PA, and NP) but shows the most similarity to the NP gene phylogeny. Separate analyses of the M1 and M2 genes and their products revealed very different patterns of evolution. Compared with other influenza virus genes (e.g., PB2 and NP), the M1 and M2 genes are evolving relatively slowly, especially the M1 gene. The M1 and M2 gene products, which are encoded in different but partially overlapping reading frames, revealed that the M1 protein is evolving very slowly in all lineages, whereas the M2 protein shows significant evolution in human and swine lineages but virtually none in avian lineages. The evolutionary rates of the M1 proteins were much lower than those of M2 proteins and other internal proteins of influenza viruses (e.g., PB2 and NP), while M2 proteins showed less rapid evolution compared with other surface proteins (e.g., H3HA). Our results also indicate that for influenza A viruses, the evolution of one protein of a bicistronic gene can affect the evolution of the other protein.(ABSTRACT TRUNCATED AT 400 WORDS)     

RAPD analysis of Fusarium Isolates Causing “Mango Malformation” Disease in Pakistan

Mango Malformation (MM) disease is a major constraint to mango production. A total of 20 Fusarium isolates from MM-affected mango plants were collected from 14 locations in Pakistan and assessed for genetic diversity using the random amplified polymorphic DNA (RAPD) technique. A total of 393 fragments were amplified after screening with 50 random primers. The amplifications with 45 primers identified scoreable polymorphisms among the isolates. A genetic similarity matrix based on Nei and Li’s index determined coefficients ranging from 46.46% to 92.51%. These coefficients were used to construct a dendrogram using the UPGMA algorithm. The isolates grouped into two main clusters, comprising 13 and 7 isolates respectively, at a genetic relatedness of 52%. Within the clusters, Fusarium isolates were not necessarily related either by geographic origin or by the mango cultivar from which they were isolated. RAPD proved a reproducible and tractable means of differentiating Fusarium isolates. These findings also suggest that some infections originate not from adjacent plants within an orchard but from geographically distant areas; indicating that most probably infection occurs in nurseries prior to plants being transported around the country for subsequent cultivation, and that improved plant hygiene could significantly curb MM infection and spread.     

Wide Geographic Distribution of Bacteriophages That Lyse the Same Indigenous Freshwater Isolate (Sphingomonas sp. Strain B18)

An indigenous freshwater bacterium (Sphingomonas sp. strain B18) from Lake Pluβsee (Schleswig-Holstein, Germany) was used to isolate 44 phages from 13 very different freshwater and brackish habitats in distant geographic areas. This bacterial strain was very sensitive to a broad spectrum of phages from different aquatic environments. Phages isolated from geographically distant aquatic habitats, but also those from the same sample, were diverse with respect to morphology and restriction pattern. Some phages were widely distributed, while different types coexisted in the same sample. It was concluded that phages could be a major factor in shaping the structure of bacterial communities and maintaining a high bacterial diversity.     

Sequence Analysis of Marine Virus Communities Reveals that Groups of Related Algal Viruses Are Widely Distributed in Nature

Algal-virus-specific PCR primers were used to amplify DNA polymerase (pol) gene fragments from geographically isolated natural virus communities. Natural algal virus communities were obtained from coastal sites in the Pacific Ocean in British Columbia, Canada, and the Southern Ocean near the Antarctic peninsula. Genetic fingerprints of algal virus communities were generated using denaturing gradient gel electrophoresis (DGGE). Sequencing efforts recovered 33 sequences from the gradient gel. Of the 33 sequences examined, 25 encoded a conserved amino acid motif indicating that the sequences were pol gene fragments. Furthermore, the 25 pol sequences were related to pol gene fragments from known algal viruses. In addition, similar virus sequences (>98% sequence identity) were recovered from British Columbia and Antarctica. Results from this study demonstrate that DGGE with degenerate primers can be used to qualitatively fingerprint and assess genetic diversity in specific subsets of natural virus communities and that closely related viruses occur in distant geographic locations. DGGE is a powerful tool for genetically fingerprinting natural virus communities and may be used to examine how specific components of virus communities respond to experimental manipulations.     

Extensive long-distance pollen dispersal in a fragmented landscape maintains genetic diversity in white spruce

Conifers are among the most genetically diverse plants but show the lowest levels of genetic differentiation, even among geographically distant populations. High gene flow among populations may be one of the most important factors in maintaining these genetic patterns. Here, we provide empirical evidence for extensive pollen-mediated gene dispersal between natural stands of a widespread northern temperate/boreal conifer, Picea glauca. We used 6 polymorphic allozyme loci to quantify the proportion of seeds sired by pollen originating from different sources in a landscape fragmented by agriculture in North Central Ontario, Canada. In 7 stands, a small proportion of seeds were sired by self-pollen or neighboring trees but 87.1% (+/-1.7% standard error [SE]) of seeds were sired by pollen from at least 250 to 3000 m away. In 4 single isolated trees, self-fertilization rates were low and more than 96% (+/-1.3% SE) of seeds were sired by immigrant pollen. The average minimum pollen dispersal distance in outcrossed matings was 619 m. These results provide strong evidence that extensive long-distance pollen dispersal plays a primary role in maintaining low genetic differentiation among natural populations of P. glauca and helps maintain genetic diversity and minimize inbreeding in small stands in a fragmented landscape.     

Phylogeographic signal variation in mitochondrial DNA among geographically isolated grassland butterflies

Aim We surveyed mitochondrial DNA (mtDNA) sequence variation among regionally isolated populations of 10 grassland‐associated butterfly species to determine: (1) the utility of phylogeographic comparisons among multiple species for assessing recent evolutionary patterns, and (2) the respective roles of isolation attributable to range disjunction versus isolation attributable solely to geographic distance in establishing divergence patterns. Location The Peace River grasslands of northern Alberta and British Columbia, Canada, which are isolated by 300+ km from similar communities to the south. Methods We sequenced mtDNA (1420 bp of cytochrome c oxidase subunit I) from five grassland‐restricted butterfly species that have geographically disjunct populations and from five ecologically broader species that have more continuous distributions across the same regions. Using analysis of molecular variance (AMOVA), Mantel and partial Mantel tests, and haplotype networks, we compared population structure within and between species in order to assess the validity of single‐species phylogeographic characterizations. We then contrasted variance components between disjunct and continuously distributed species to assess whether divergences were correlated more with disjunction or with geographic distance. Results Single‐species analyses varied substantially within both the disjunct and the continuous groups. One species in each of these groups had mtDNA with unusually deep intraspecific mitochondrial lineage divergences. On the whole, however, the five species with disjunct ranges exhibited greater divergence between geographically distant populations than did the five species with continuous distributions. Comparison of variance components between disjunct and continuous species indicated that isolation attributable only to geographic distance was responsible for up to half of the total sequence variation between disjunct populations of grassland butterflies. Main conclusions Our findings show that single‐species phylogeographic analyses of post‐Pleistocene butterfly distributions are inadequate for characterizing regional biogeographic divergence histories. However, comparison of mtDNA sequence divergences between groups of disjunct and continuously distributed species can allow isolation attributable to range interruption to be quantitatively distinguished from isolation attributable solely to gene flow attenuation over the same geographic area.     

Genetic characterization of an avian H4N6 influenza virus isolated from the Izumi plain,Japan

An influenza A virus of H4N6 subtype was isolated from the Izumi plain, Japan, in 2013. Genetic analyses revealed that two viral genes (M and NS gene segments) of this isolate were genetically distinct from those of the H4N6 virus isolated from the same place in 2012. Furthermore, three viral genes (PB2, PB1 and M gene segments) of this isolate share high similarity with those of the North American isolates of 2014. These results suggest a high frequency of genetic reassortment of avian influenza viruses in Asian waterfowl and intercontinental movements of avian influenza viruses via migratory waterfowl.

     

ISOZYME ANALYSIS OF MATING POPULATIONS OF GONIUM PECTORALE (CHLOROPHYTA)1

Intraspecific variation among 36 strains of the freshwater alga Gonium pectorale Müller (Chlorophyceae) isolated from three geographically different locations in Tibet, Nepal, and Japan was investigated by isozyme analysis. Variation in isozyme patterns of eight enzyme systems (malate dehydrogenase, glutamate dehydrogenase, tetrazolium oxidase, lactate dehydrogenase, octanol dehydrogenase, xanthine dehydrogenase, phosphoglucomutase, and malic enzyme) of axenic and clonal cultures was revealed by polyacrylamide gel electrophoresis. Unweighted average linkage clustering, based on Jaccard's similarity coefficient, illustrated the high similarity between most strains from Nepal and all strains from the Ryukyu Islands (Japan). However, there was relatively low similarity between strains from Tibet and those from Nepal and Japan. Strains from the Ryukyu Islands (Japan) grouped into two clusters, and most Nepalese strains formed a single cluster, but Tibetan strains were heterogenous.     

Comparative study of the aerobic, heterotrophic bacterial flora of Chesapeake Bay and Tokyo Bay.

A comparative study of the bacterial flora of the water of Chesapeake Bay and Tokyo Bay was undertaken to assess similarities and differences between the autochthonous flora of the two geographical sites and to test the hypothesis that, given similarities in environmental parameters, similar bacterial populations will be found, despite extreme geographic distance between locations. A total of 195 aerobic, heterotrophic bacterial strains isolated from Chesapeake Bay and Tokyo Bay water were examined for 115 biochemical, cultural, morphological, nutritional, and physiological characters. The data were analyzed by the methods of numerical taxonomy. From sorted similarity matrices, 77% of the isolates could be grouped into 30 phena and presumptively identified as Acinetobacter-Moraxella, Caulobacter, coryneforms, Pseudomonas, and Vibrio spp. Vibrio and Acinetobacter species were found to be common in the estuarine waters of Chesapeake Bay, whereas Acinetobacter-Moraxella and Caulobacter predominated in Tokyo Bay waters, at the sites sampled in the study.     

Comparative study of the aerobic, heterotrophic bacterial flora of Chesapeake Bay and Tokyo Bay.

A comparative study of the bacterial flora of the water of Chesapeake Bay and Tokyo Bay was undertaken to assess similarities and differences between the autochthonous flora of the two geographical sites and to test the hypothesis that, given similarities in environmental parameters, similar bacterial populations will be found, despite extreme geographic distance between locations. A total of 195 aerobic, heterotrophic bacterial strains isolated from Chesapeake Bay and Tokyo Bay water were examined for 115 biochemical, cultural, morphological, nutritional, and physiological characters. The data were analyzed by the methods of numerical taxonomy. From sorted similarity matrices, 77% of the isolates could be grouped into 30 phena and presumptively identified as Acinetobacter-Moraxella, Caulobacter, coryneforms, Pseudomonas, and Vibrio spp. Vibrio and Acinetobacter species were found to be common in the estuarine waters of Chesapeake Bay, whereas Acinetobacter-Moraxella and Caulobacter predominated in Tokyo Bay waters, at the sites sampled in the study.     

Mineralogy Drives Bacterial Biogeography of Hydrothermally Inactive Seafloor Sulfide Deposits

Mid-ocean ridge hydrothermal venting creates sulfide deposits containing gradients in mineralogy, fluid chemistry, and temperature. Even when hydrothermal circulation ceases, sulfides are known to host microbial communities. The relationship between mineralogy and microbial community composition in low-temperature, rock-hosted systems has not been resolved at any spatial scale, local or global. To examine the hypothesis that geochemistry of seafloor deposits is a dominant parameter driving environmental pressure for bacterial communities at low-temperature, the shared community membership, richness, and structure was measured using 16S rRNA gene sequences. The focus of the study was on hydrothermally inactive seafloor deposits from multiple locations within one deposit (e.g., single extinct chimney), within one vent field (intra-vent field), and among globally distributed vent fields from three ocean basins (inter-vent field). Distinct mineral substrates, such as hydrothermally inactive sulfides versus basalts, host different communities at low temperature in spite of close geographic proximity and contact with the same hydrothermally influenced deep-sea water. Furthermore, bacterial communities inhabiting hydrothermally inactive sulfide deposits from geographically distant locations cluster together in community cladograms to the exclusion of other deep-sea substrates and settings. From this study, we conclude that at low temperature, mineralogy was a more important variable determining microbial community composition than geographic factors. Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the supplemental file.     

Detection and characterization of hepatitis A virus and Norovirus in estuarine water samples using ultrafiltration – RT-PCR integrated methods

Aims:  Waterborne outbreaks of hepatitis A and Norovirus disease have been reported and associated with contaminated water supply in various countries. However, in Mexico, there are no studies that report HAV and NV presence in water. This study reports the application of ultrafiltration and RT-nested PCR methods to concentrate and identify these viruses.
Methods and Results:  Forty estuarine water samples were collected from the Huizache Caimanero Lagunary Complex. Samples were concentrated by ultrafiltration system (UFS) and RT-nested PCR was performed for HAV and NV identification. These viruses were found in 80% and 70% of the samples collected respectively and both were present in 57·5%. The DNA sequences analysis showed that 21 estuarine water samples were associated with HAV and 13 with NV. Faecal coliforms were isolated in 48·57% of the samples, while Escherichia coli were found in 34·28%.
Conclusions:  DNA sequencing showed that the genotype IB for HAV and GII for NV were predominant in México. No significant relationships were detected between indicators and viruses ( P  < 0·05).
Significance and Impact of the Study:  This study shows that the UFS is adequate for viral concentration. This is the first study analysing the genetic sequence of HAV and NV isolated from Mexican estuarine water.     

Intraspecific variability in leaf traits strongly affects alder leaf decomposition in a stream

This study assessed the intraspecific variability of senescent leaves of alder (Alnus glutinosa Gaertn.) and the effects of this variability on leaf decomposition in streams. Leaves were collected at five geographically distant locations in Europe. We analyzed 10 batches of leaf samples for seven quantitative leaf traits as well as leaf decomposition rate in coarse and fine mesh bags exposed in a single stream. The geographic origin of leaf samples largely explained the observed variation in litter quality and decomposition rate. Phosphorus (0.034–0.187%) and lignin (3.9–18.7%) concentrations in leaves varied widely. Together, these two traits accurately predicted leaf decomposition rate (r²=84.1%). Intraspecific variation in leaf decomposition rate was within a range similar to that reported for interspecific variation among co-occurring riparian plant species in Europe. Our study demonstrates extensive intraspecific variability in leaf traits on a continental scale, which can have enormous effects on major ecosystem processes such as leaf decomposition.     

Enrichment and identification of polycyclic aromatic compound-degrading bacteria enriched from sediment samples

The degradation of polycyclic aromatic compounds (PACs) has been widely studied. Knowledge of the degradation of PACs by microbial populations can be utilized in the remediation of contaminated sites. To isolate and identify PAC-degrading bacteria for potential use in future bioremediation programmes, we established a series of PAC enrichments under the same experimental conditions from a single sediment sample taken from a highly polluted estuarine site. Enrichment cultures were established using the pollutants: anthracene, phenanthrene and dibenzothiophene as a sole carbon source. The shift in microbial community structure on each of these carbon sources was monitored by analysis of a time series of samples from each culture using 16S rRNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Significantly, our findings demonstrate that shifts in the constituent species within each degradative community are directly attributable to enrichment with different PACs. Subsequently, we characterized the microorganisms comprising the degradative communities within each enrichment using 16S rRNA sequence data. Our findings demonstrate that the ability to degrade PACs is present in five divisions of the Proteobacteria and Actinobacteria. By determining the precise identity of the PAC-degrading bacterial species isolated from a single sediment sample, and by comparing our findings with previously published research, we demonstrate how bacteria with similar PAC degrading capabilities and 16S rRNA signatures are found in similarly polluted environments in geographically very distant locations, e.g., China, Italy, Japan and Hawaii. Such a finding suggests that geographical barriers do not limit the distribution of key PAC-degrading bacteria; this finding is in accordance with the Baas-Becking hypothesis “everything is everywhere; the environment selects” and may have significant consequences for the global distribution of PAC-degrading bacteria and their use in bioremediation.