Dynamics of Autotrophic Marine Planktonic Thaumarchaeota in the East China Sea

The ubiquitous and abundant distribution of ammonia-oxidizing Thaumarchaeota in marine environments is now well documented, and their crucial role in the global nitrogen cycle has been highlighted. However, the potential contribution of Thaumarchaeota in the carbon cycle remains poorly understood. Here we present for the first time a seasonal investigation on the shelf region (bathymetry≤200 m) of the East China Sea (ECS) involving analysis of both thaumarchaeal 16S rRNA and autotrophy-related genes (acetyl-CoA carboxylase gene, accA). Quantitative PCR results clearly showed a higher abundance of thaumarchaeal 16S and accA genes in late-autumn (November) than summer (August), whereas the diversity and community structure of autotrophic Thaumarchaeota showed no statistically significant difference between different seasons as revealed by thaumarchaeal accA gene clone libraries. Phylogenetic analysis indicated that shallow ecotypes dominated the autotrophic Thaumarchaeota in the ECS shelf (86.3% of total sequences), while a novel non-marine thaumarchaeal accA lineage was identified in the Changjiang estuary in summer (when freshwater plumes become larger) but not in autumn, implying that Changjiang freshwater discharge played a certain role in transporting terrestrial microorganisms to the ECS. Multivariate statistical analysis indicated that the biogeography of the autotrophic Thaumarchaeota in the shelf water of the ECS was influenced by complex hydrographic conditions. However, an in silico comparative analysis suggested that the diversity and abundance of the autotrophic Thaumarchaeota might be biased by the ‘universal’ thaumarchaeal accA gene primers Cren529F/Cren981R since this primer set is likely to miss some members within particular phylogenetic groups. Collectively, this study improved our understanding of the biogeographic patterns of the autotrophic Thaumarchaeota in temperate coastal waters, and suggested that new accA primers with improved coverage and sensitivity across phylogenetic groups are needed to gain a more thorough understanding of the role of the autotrophic Thaumarchaeota in the global carbon cycle.     

Genomic properties of Marine Group A bacteria indicate a role in the marine sulfur cycle

Marine Group A (MGA) is a deeply branching and uncultivated phylum of bacteria. Although their functional roles remain elusive, MGA subgroups are particularly abundant and diverse in oxygen minimum zones and permanent or seasonally stratified anoxic basins, suggesting metabolic adaptation to oxygen-deficiency. Here, we expand a previous survey of MGA diversity in O₂-deficient waters of the Northeast subarctic Pacific Ocean (NESAP) to include Saanich Inlet (SI), an anoxic fjord with seasonal O₂ gradients and periodic sulfide accumulation. Phylogenetic analysis of small subunit ribosomal RNA (16S rRNA) gene clone libraries recovered five previously described MGA subgroups and defined three novel subgroups (SHBH1141, SHBH391, and SHAN400) in SI. To discern the functional properties of MGA residing along gradients of O₂ in the NESAP and SI, we identified and sequenced to completion 14 fosmids harboring MGA-associated 16S RNA genes from a collection of 46 fosmid libraries sourced from NESAP and SI waters. Comparative analysis of these fosmids, in addition to four publicly available MGA-associated large-insert DNA fragments from Hawaii Ocean Time-series and Monterey Bay, revealed widespread genomic differentiation proximal to the ribosomal RNA operon that did not consistently reflect subgroup partitioning patterns observed in 16S rRNA gene clone libraries. Predicted protein-coding genes associated with adaptation to O₂-deficiency and sulfur-based energy metabolism were detected on multiple fosmids, including polysulfide reductase (psrABC), implicated in dissimilatory polysulfide reduction to hydrogen sulfide and dissimilatory sulfur oxidation. These results posit a potential role for specific MGA subgroups in the marine sulfur cycle.     

Energy Content of Antarctic Mesopelagic Fishes: Implications for the Marine Food Web

For a better understanding of the role of mesopelagic fish in the Southern Ocean food web, the energy and water content of Bathylagus antarcticus, Electrona antarctica and Gymnoscopelus braueri from the Lazarev Sea were investigated. Mean dry weight energy content of B. antarcticus (20.4 kJ g⁻¹) was significantly lower than in E. antarctica and G. braueri (both 29.4 kJ g⁻¹). In E. antarctica, an increase of dry weight energy density with age was evident from 26.9 kJ g⁻¹ in juveniles of less than 1 year of age to 32.0 kJ g⁻¹ in 3-year-old fish. Water content decreased with size in all three species. Abundant high-energy species such as E. antarctica are at a key position in the food web. Due to a marked influence of age on energy content, population structure can be an important variable in estimates of energy fluxes in the Southern Ocean ecosystem.     

Comparative Genomic Hybridization Analysis of Two Predominant Nordic Group I (Proteolytic) Clostridium botulinum Type B Clusters

Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B strains isolated from various sources revealed two homogeneous clusters, clusters BI and BII. The type B strains differed from reference strain ATCC 3502 by 413 coding sequence (CDS) probes, sharing 88% of all the ATCC 3502 genes represented on the microarray. The two Nordic type B clusters differed from each other by their response to 145 CDS probes related mainly to transport and binding, adaptive mechanisms, fatty acid biosynthesis, the cell membranes, bacteriophages, and transposon-related elements. The most prominent differences between the two clusters were related to resistance to toxic compounds frequently found in the environment, such as arsenic and cadmium, reflecting different adaptive responses in the evolution of the two clusters. Other relatively variable CDS groups were related to surface structures and the gram-positive cell wall, suggesting that the two clusters possess different antigenic properties. All the type B strains carried CDSs putatively related to capsule formation, which may play a role in adaptation to different environmental and clinical niches. Sequencing showed that representative strains of the two type B clusters both carried subtype B2 neurotoxin genes. As many of the type B strains studied have been isolated from foods or associated with botulism, it is expected that the two group I C. botulinum type B clusters present a public health hazard in Nordic countries. Knowing the genetic and physiological markers of these clusters will assist in targeting control measures against these pathogens.Clostridium botulinum produces a potent neurotoxin during its growth. The toxin causes a potentially lethal paralytic disease, botulism, in humans and animals. The classical food-borne botulism follows the consumption of toxin-containing food or drink, while infant and adult intestinal botulism results from in vivo spore germination, outgrowth, and toxin production in the gut. Apart from attenuated intestinal microbial population, other factors affecting the colonization of C. botulinum in the intestinal forms of botulism are not known.Based on their physiology and genetic background, C. botulinum strains are divided into groups I to IV (13). Strains of groups I and II are associated with human disease. Group I strains produce neurotoxin serotypes A, B, and/or F, while the group II strains produce type B, E, or F toxin. Physiologically, groups I and II differ markedly from each other as well as from groups III and IV. Genomic analysis of group I and II C. botulinum strains by 16S rrn sequencing (13), ribotyping (10), and amplified fragment length polymorphism (11, 15, 16) is consistent with the divergent physiologies of the two groups (18).Nordic C. botulinum group I strains show a remarkable homogeneity (15, 20, 21, 23). In a large pulsed-field gel electrophoresis (PFGE) analysis, the majority of group I strains isolated from various sources from Finland, Norway, and Denmark formed type B neurotoxin and clustered into two large groups, with the members of each group sharing identical or nearly identical restriction patterns (20, 23). Many of these strains were recovered from honey for human consumption (23), and one strain was related to an infant botulism case (22). Apart from a recent study showing that strains of the two type B clusters, further referred to as clusters BI and BII, differ in their abilities to grow at extreme temperatures (12), the physiological, epidemiological, and genetic markers of the two clusters are not known. An understanding of such traits will assist in designing control measures against these potential food- and environment-borne pathogens.The availability of group I C. botulinum genome sequences has enabled the construction of whole-genome DNA microarrays and a comprehensive genomic analysis of C. botulinum strains (26, 27). In this paper, we describe a comparative genomic hybridization (CGH) analysis of 32 Nordic group I C. botulinum type B cluster BI or BII strains with a DNA microarray based on the protein-coding sequences (CDS) in the ATCC 3502 genome. Strains within each cluster showed no substantial variation. Furthermore, strains belonging to the two clusters differed by their responses to 145 CDS probes, suggesting differential resistance to toxic compounds and a relatively large antigenic variability. Sequencing of botB in a representative cluster BI strain and a representative cluster BII strain revealed subtype B2 neurotoxin genes in both strains.     

Diazotrophic Diversity and Distribution in the Tropical and Subtropical Atlantic Ocean

To understand the structure of marine diazotrophic communities in the tropical and subtropical Atlantic Ocean, the molecular diversity of the nifH gene was studied by nested PCR amplification using degenerate primers, followed by cloning and sequencing. Sequences of nifH genes were amplified from environmental DNA samples collected during three cruises (November-December 2000, March 2002, and October-November 2002) covering an area between 0 to 28.3°N and 56.6 to 18.5°W. A total of 170 unique sequences were recovered from 18 stations and 23 depths. Samples from the November-December 2000 cruise contained both unicellular and filamentous cyanobacterial nifH phylotypes, as well as γ-proteobacterial and cluster III sequences, so far only reported in the Pacific Ocean. In contrast, samples from the March 2002 cruise contained only phylotypes related to the uncultured group A unicellular cyanobacteria. The October-November 2002 cruise contained both filamentous and unicellular cyanobacterial and γ-proteobacterial sequences. Several sequences were identical at the nucleotide level to previously described environmental sequences from the Pacific Ocean, including group A sequences. The data suggest a community shift from filamentous cyanobacteria in surface waters to unicellular cyanobacteria and/or heterotrophic bacteria in deeper waters. With one exception, filamentous cyanobacterial nifH sequences were present within temperatures ranging between 26.5 and 30°C and where nitrate was undetectable. In contrast, nonfilamentous nifH sequences were found throughout a broader temperature range, 15 to 30°C, more often in waters with temperature of <26°C, and were sometimes recovered from waters with detectable nitrate concentrations.     

Phylogenetic Diversity of the Bacillus pumilus Group and the Marine Ecotype Revealed by Multilocus Sequence Analysis

Bacteria closely related to Bacillus pumilus cannot be distinguished from such other species as B. safensis, B. stratosphericus, B. altitudinis and B. aerophilus simply by 16S rRNA gene sequence. In this report, 76 marine strains were subjected to phylogenetic analysis based on 7 housekeeping genes to understand the phylogeny and biogeography in comparison with other origins. A phylogenetic tree based on the 7 housekeeping genes concatenated in the order of gyrB-rpoB-pycA-pyrE-mutL-aroE-trpB was constructed and compared with trees based on the single genes. All these trees exhibited a similar topology structure with small variations. Our 79 strains were divided into 6 groups from A to F; Group A was the largest and contained 49 strains close to B. altitudinis. Additional two large groups were presented by B. safensis and B. pumilus respectively. Among the housekeeping genes, gyrB and pyrE showed comparatively better resolution power and may serve as molecular markers to distinguish these closely related strains. Furthermore, a recombinant phylogenetic tree based on the gyrB gene and containing 73 terrestrial and our isolates was constructed to detect the relationship between marine and other sources. The tree clearly showed that the bacteria of marine origin were clustered together in all the large groups. In contrast, the cluster belonging to B. safensis was mainly composed of bacteria of terrestrial origin. Interestingly, nearly all the marine isolates were at the top of the tree, indicating the possibility of the recent divergence of this bacterial group in marine environments. We conclude that B. altitudinis bacteria are the most widely spread of the B. pumilus group in marine environments. In summary, this report provides the first evidence regarding the systematic evolution of this bacterial group, and knowledge of their phylogenetic diversity will help in the understanding of their ecological role and distribution in marine environments.     

Spatial Distribution of Marine Crenarchaeota Group I in the Vicinity of Deep-Sea Hydrothermal Systems

Distribution profiles of marine crenarchaeota group I in the vicinity of deep-sea hydrothermal systems were mapped with culture-independent molecular techniques. Planktonic samples were obtained from the waters surrounding two geographically and geologically distinct hydrothermal systems, and the abundance of marine crenarchaeota group I was examined by 16S ribosomal DNA clone analysis, quantitative PCR, and whole-cell fluorescence in situ hybridization. A much higher proportion of marine crenarchaeota group I within the microbial community was detected in deep-sea hydrothermal environments than in normal deep and surface seawaters. The highest proportion was always obtained from the ambient seawater adjacent to hydrothermal emissions and chimneys but not from the hydrothermal plumes. These profiles were markedly different from the profiles of epsilon-Proteobacteria, which are abundant in the low temperatures of deep-sea hydrothermal environments.     

Exploration of the Genomic Diversity and Core Genome of the Bifidobacterium adolescentis Phylogenetic Group by Means of a Polyphasic Approach

In the current work, we describe genome diversity and core genome sequences among representatives of three bifidobacterial species, i.e., Bifidobacterium adolescentis, Bifidobacterium catenulatum, and Bifidobacterium pseudocatenulatum, by employing a polyphasic approach involving analysis of 16S rRNA gene and 16S-23S internal transcribed spacer (ITS) sequences, pulsed-field gel electrophoresis (PFGE), and comparative genomic hybridization (CGH) assays.     

Dynamics of Tree Diversity in Undisturbed and Logged Subtropical Rainforest in Australia

In subtropical rainforest in eastern Australia, changes in the diversity of trees were compared under natural conditions and eight silvicultural regimes over 35 years. In the treated plots basal area remaining after logging ranged from 12 to 58 m^{2}/ha. In three control plots richness differed little over this period. In the eight treated plots richness per plot generally declined after intervention and then gradually increased to greater than original diversity. After logging there was a reduction in richness per plot and an increase in species richness per stem in all but the lightest selective treatments. The change in species diversity was related to the intensity of the logging, however the time taken for species richness to return to pre-logging levels was similar in all silvicultural treatments and was not effected by the intensity of treatment. These results suggest that light selective logging in these forests mainly affects dominant species. The return to high diversity after only a short time under all silvicultural regimes suggests that sustainability and the manipulation of species composition for desired management outcomes is possible.     

Construction of a Fibrobacter succinogenes Genomic Map and Demonstration of Diversity at the Genomic Level

The genomic cleavage map of the type strain Fibrobacter succinogenes S85 was constructed. The restriction enzymes AscI, AvrII, FseI, NotI, and SfiI generated DNA fragments of suitable size distribution that could be resolved by pulsed-field gel electrophoresis (PFGE). An average genome size of 3.6 Mb was obtained by summing the total fragment sizes. The linkages between the 15 AscI fragments of the genome were determined by combining two approaches: isolation of linking clones and cross-hybridization of restriction fragments. The genome of F. succinogenes was found to be represented by the single circular DNA molecule. Southern hybridization with specific probes allowed the eight genetic markers to be located on the restriction map. The genome of this bacterium contains at least three rRNA operons. PFGE of the other three strains of F. succinogenes gave estimated genome sizes close to that of the type strain. However, RFLP patterns of these strains generated by AscI digestion are completely different. Pairwise comparison of the genomic fragment distribution between the type strain and the three isolates showed a similarity level in the region of 14.3% to 31.3%. No fragment common to all of these F. succinogenes strains could be detected by PFGE. A marked degree of genomic heterogeneity among members of this species makes genomic RFLP a highly discriminatory and useful molecular typing tool for population studies. Received: 23 October 1996 / Accepted: 31 December 1996     

Genomic and Genetic Diversity within the Pseudomonas fluorescens Complex

The Pseudomonas fluorescens complex includes Pseudomonas strains that have been taxonomically assigned to more than fifty different species, many of which have been described as plant growth-promoting rhizobacteria (PGPR) with potential applications in biocontrol and biofertilization. So far the phylogeny of this complex has been analyzed according to phenotypic traits, 16S rDNA, MLSA and inferred by whole-genome analysis. However, since most of the type strains have not been fully sequenced and new species are frequently described, correlation between taxonomy and phylogenomic analysis is missing. In recent years, the genomes of a large number of strains have been sequenced, showing important genomic heterogeneity and providing information suitable for genomic studies that are important to understand the genomic and genetic diversity shown by strains of this complex. Based on MLSA and several whole-genome sequence-based analyses of 93 sequenced strains, we have divided the P. fluorescens complex into eight phylogenomic groups that agree with previous works based on type strains. Digital DDH (dDDH) identified 69 species and 75 subspecies within the 93 genomes. The eight groups corresponded to clustering with a threshold of 31.8% dDDH, in full agreement with our MLSA. The Average Nucleotide Identity (ANI) approach showed inconsistencies regarding the assignment to species and to the eight groups. The small core genome of 1,334 CDSs and the large pan-genome of 30,848 CDSs, show the large diversity and genetic heterogeneity of the P. fluorescens complex. However, a low number of strains were enough to explain most of the CDSs diversity at core and strain-specific genomic fractions. Finally, the identification and analysis of group-specific genome and the screening for distinctive characters revealed a phylogenomic distribution of traits among the groups that provided insights into biocontrol and bioremediation applications as well as their role as PGPR.     

Genomic Adaptation of the Lactobacillus casei Group

Lactobacillus casei, L. paracasei, and L. rhamnosus form a closely related taxonomic group (Lactobacillus casei group) within the facultatively heterofermentative lactobacilli. Here, we report the complete genome sequences of L. paracasei JCM 8130 and L. casei ATCC 393, and the draft genome sequence of L. paracasei COM0101, all of which were isolated from daily products. Furthermore, we re-annotated the genome of L. rhamnosus ATCC 53103 (also known as L. rhamnosus GG), which we have previously reported. We confirmed that ATCC 393 is distinct from other strains previously described as L. paracasei. The core genome of 10 completely sequenced strains of the L. casei group comprised 1,682 protein-coding genes. Although extensive genome-wide synteny was found among the L. casei group, the genomes of ATCC 53103, JCM 8130, and ATCC 393 contained genomic islands compared with L. paracasei ATCC 334. Several genomic islands, including carbohydrate utilization gene clusters, were found at the same loci in the chromosomes of the L. casei group. The spaCBA pilus gene cluster, which was first identified in GG, was also found in other strains of the L. casei group, but several L. paracasei strains including COM0101 contained truncated spaC gene. ATCC 53103 encoded a higher number of proteins involved in carbohydrate utilization compared with intestinal lactobacilli, and extracellular adhesion proteins, several of which are absent in other strains of the L. casei group. In addition to previously fully sequenced L. rhamnosus and L. paracasei strains, the complete genome sequences of L. casei will provide valuable insights into the evolution of the L. casei group.     

Species of the parasitic genus Duboscquella are members of the enigmatic Marine Alveolate Group I

Small subunit ribosomal RNA gene sequences of Duboscquella spp. infecting the tintinnid ciliate, Favella ehrenbergii, were determined. Two parasites were sampled from different localities. They are morphologically similar to each other and both resemble D. aspida. Nevertheless, two distinct sequences (7.6% divergence) were obtained from them. Phylogenetic trees inferred from maximum likelihood and maximum parsimony revealed that these two Duboscquella spp. sequences are enclosed in an environmental clade named Marine Alveolate Group I. This clade consists of a large number of picoplanktonic organisms known only from environmental samples from various parts of the ocean worldwide, and which therefore lack clear characterization and identification. Here, we provide morphological and genetic characterization of these two Duboscquella genotypes included in this enigmatic clade. Duboscquella spp. produce a large number of small flagellated spores as dispersal agents and the presence of such small cells partially explains why the organisms related to these parasites have been detected within environmental genetic libraries, built from picoplanktonic size fractions of environmental samples. The huge diversity of the Marine Alveolate Group I and the finding that parasites from different marine protists belong to this lineage suggest that parasitism is a widespread and ecologically relevant phenomenon in the marine environment.     

海洋古菌多样性研究进展

海洋古菌是海洋微生物中的一个大的类群,然而绝大多数的古菌不能分离培养.近年来分子生物学的方法广泛地应用于微生物多样性的研究中,研究发现,海洋古菌广泛地生活在各类海域环境中,而不仅仅是生活在极端的环境中.海洋古菌为海洋生态系统中主要的原核细胞成分,在海洋生态系统中的物质与能量循环中扮演着重要角色.主要阐述了生活在海洋不同环境中海洋古菌的多样性,有海洋浮游古菌的多样性、海底环境及海洋沉积物中古菌的多样性、附着或寄共生古菌多样性等的研究状况,以及研究海洋古菌多样性的分子生物学的主要方法.