Phylogeny and physiology of candidate phylum ‘Atribacteria' (OP9/JS1) inferred from cultivation-independent genomics

The ‘Atribacteria'' is a candidate phylum in the Bacteria recently proposed to include members of the OP9 and JS1 lineages. OP9 and JS1 are globally distributed, and in some cases abundant, in anaerobic marine sediments, geothermal environments, anaerobic digesters and reactors and petroleum reservoirs. However, the monophyly of OP9 and JS1 has been questioned and their physiology and ecology remain largely enigmatic due to a lack of cultivated representatives. Here cultivation-independent genomic approaches were used to provide a first comprehensive view of the phylogeny, conserved genomic features and metabolic potential of members of this ubiquitous candidate phylum. Previously available and heretofore unpublished OP9 and JS1 single-cell genomic data sets were used as recruitment platforms for the reconstruction of atribacterial metagenome bins from a terephthalate-degrading reactor biofilm and from the monimolimnion of meromictic Sakinaw Lake. The single-cell genomes and metagenome bins together comprise six species- to genus-level groups that represent most major lineages within OP9 and JS1. Phylogenomic analyses of these combined data sets confirmed the monophyly of the ‘Atribacteria'' inclusive of OP9 and JS1. Additional conserved features within the ‘Atribacteria'' were identified, including a gene cluster encoding putative bacterial microcompartments that may be involved in aldehyde and sugar metabolism, energy conservation and carbon storage. Comparative analysis of the metabolic potential inferred from these data sets revealed that members of the ‘Atribacteria'' are likely to be heterotrophic anaerobes that lack respiratory capacity, with some lineages predicted to specialize in either primary fermentation of carbohydrates or secondary fermentation of organic acids, such as propionate.     

The effects of geographic origin and antibiotic treatment on the gut symbiotic communities of Bactrocera oleae populations

The olive fruit fly, Bactrocera oleae (Rossi) (Diptera: Tephritidae), is the major insect pest of olive orchards (Olea europaea L.), causing extensive damages on cultivated olive crops worldwide. Due to its economic importance, it has been the target species for a variety of population control approaches including the sterile insect technique (SIT). However, the inefficiency of the current mass‐rearing techniques impedes the successful application of area‐wide integrated pest management programs with an SIT component. It has been shown that insect mass rearing and quality of sterile insects can be improved by the manipulation of the insect gut microbiota and probiotic applications. In order to exploit the gut bacteria, it is important to investigate the structure of the gut microbial community. In the current study, we characterized the gut bacterial profile of two wild olive fruit fly populations introduced in laboratory conditions using next generation sequencing of two regions of the 16S rRNA gene. We compared the microbiota profiles regarding the geographic origin of the samples. Additionally, we investigated potential changes in the gut bacteria community before and after the first exposure of the wild adult flies to artificial adult diet with and without antibiotics. Various genera – such as Erwinia, Providencia, Enterobacter, and Klebsiella – were detected for the first time in B. oleae. The most dominant species was Candidatus Erwinia dacicola Capuzzo et al. and it was not affected by the antibiotics in the artificial adult diet used in the first generation of laboratory rearing. Geographic origin affected the overall structure of the gut community of the olive fruit fly, but antibiotic treatment in the first generation did not significantly alter the gut microbiota community.     

Phosphate deposition during and after hypokinesia in phosphate supplemented and unsupplemented rats

The objective of this study was to show that prolonged restriction of motor activity (hypokinesia) could reduce phosphate (P) deposition and contribute to P loss with tissue P depletion. To this end, measurements were made of tissue P content, P absorption, plasma P levels, urinary and fecal P excretion of rats during and after hypokinesia (HK) and daily phosphate supplementation. Studies were conducted on male Wistar rats during a pre-hypokinetic period, a hypokinetic period and a post-hypokinetic period. All rats were equally divided into four groups: unsupplemented vivarium control rats (UVCR), unsupplemented hypokinetic rats (UHKR), supplemented vivarium control rats (SVCR) and supplemented hypokinetic rats (SHKR). Bone and muscle P content, plasma intact parathyroid hormone (iPTH) levels, P absorption, plasma P levels and urinary and fecal P excretion did not change in SVCR and UVCR compared with their pre-HK values. During HK, plasma P levels, urinary and fecal P excretion increased significantly (p<0.05) while muscle and bone P content, P absorption and plasma iPTH levels decreased significantly (p<0.05) in SHKR and UHKR compared with their pre-HK values and the values in their respective vivarium controls (SVCR and UVCR). During the initial 9-days of post-HK, plasma, urinary and fecal P levels decreased significantly (p<0.05), and plasma iPTH levels, muscle and bone P levels remained significantly (p<0.05) depressed in hypokinetic rats compared with their pre-HK values and the values in their respective vivarium control rats. By the 15th day, these values approached the control values. During HK and post-HK, changes in P absorption, plasma iPTH levels, and P levels in muscle, bone, plasma, urine and feces were significantly (p<0.05) greater in SHKR than in UHKR. Decreased tissue P content with increased P loss in animals receiving and not receiving P supplementation demonstrates decreased P deposition during HK. Higher P excretion with lower tissue content in SHKR and UHKR demonstrates that P deposition is decreased more with P supplementation than without. Because SHKR with a lower tissue P content showed higher P excretion than UHKR it was concluded that the risk of decreased P deposition with greater tissue P depletion is inversely related to P intake, that is, the higher the P intake the greater the risk for decreased P deposition and the greater tissue P depletion. It was shown that P (regardless of the intensity of its tissue depletion) is lost during HK unless factors contributing to the decreased P deposition are partially or totally reversed. It was concluded that dissociation between (decreased) tissue P content and (increased) P uptake indicates decreased P (absorption and) deposition as the main mechanisms of tissue P depletion during prolonged HK.     

Detection and characterization of Wolbachia infections in laboratory and natural populations of different species of tsetse flies (genus Glossina)

Background

Wolbachia is a genus of endosymbiotic α-Proteobacteria infecting a wide range of arthropods and filarial nematodes. Wolbachia is able to induce reproductive abnormalities such as cytoplasmic incompatibility (CI), thelytokous parthenogenesis, feminization and male killing, thus affecting biology, ecology and evolution of its hosts. The bacterial group has prompted research regarding its potential for the control of agricultural and medical disease vectors, including Glossina spp., which transmits African trypanosomes, the causative agents of sleeping sickness in humans and nagana in animals.

Results

In the present study, we employed a Wolbachia specific 16S rRNA PCR assay to investigate the presence of Wolbachia in six different laboratory stocks as well as in natural populations of nine different Glossina species originating from 10 African countries. Wolbachia was prevalent in Glossina morsitans morsitans, G. morsitans centralis and G. austeni populations. It was also detected in G. brevipalpis, and, for the first time, in G. pallidipes and G. palpalis gambiensis. On the other hand, Wolbachia was not found in G. p. palpalis, G. fuscipes fuscipes and G. tachinoides. Wolbachia infections of different laboratory and natural populations of Glossina species were characterized using 16S rRNA, the wsp (Wolbachia Surface Protein) gene and MLST (Multi Locus Sequence Typing) gene markers. This analysis led to the detection of horizontal gene transfer events, in which Wobachia genes were inserted into the tsetse flies fly nuclear genome.

Conclusions

Wolbachia infections were detected in both laboratory and natural populations of several different Glossina species. The characterization of these Wolbachia strains promises to lead to a deeper insight in tsetse flies-Wolbachia interactions, which is essential for the development and use of Wolbachia-based biological control methods.

     

Genetic differentiation among Greek lake populations of Carassius gibelio and Cyprinus carpio carpio

The genetic structure of the Western Greece lake populations of Carassius gibelio and Cyprinus carpio carpio populations was characterized by using a PCR-based RFLP and sequencing analysis of mitochondrial rDNA genes and regions (16S rDNA, cytochrome b and D-loop). Our analysis was able to detect: (a) two haplotypes in C. c. carpio populations and two haplotypes in C. gibelio populations (b) a high nucleotide divergence between the two species and (c) two genetically distinct C. gibelio populations, one existing in the Amvrakia habitat (AMV1) with a second in Ozeros and Trichonida (OZE1 and TRI1) habitat. The present analysis indicates that genetic diversity observed was limited with a haplotype index between 0.0 and 55.6%, and a nucleotide diversity within and among populations between 0.0 and 1.27%. It also underlines a restricted mtDNA-based evaluation of the phylogenetic relationships among C. gibelio and C. c. carpio populations. In addition, the present study contributed knowledge on the genetic variation and structure of these populations which is absolutely necessary for any efficient fish management and/or conservation programme.     

Bacterial diversity and hydrography of Etoliko,an anoxic semi-enclosed coastal basin in Western Greece

Etoliko, an anoxic semi-enclosed basin, is part of a complex wetland in Western Greece extremely rich in biodiversity. It covers an area of 1,700 ha with an atypical orientation that has been formed tectonically. In order to identify the main factors influencing the bacterial profile at the Etoliko basin, 48 samples were collected, representing seasonal variation at four sampling stations. Physico-chemical analysis of the samples indicates the presence of three layers in the Etoliko basin: (1) low-density surface layer, (2) a layer with a steep density gradient, and (3) dense water below a depth of 20 m. A permanent halocline, whose thickness is varying seasonally, has been identified in the Etoliko basin water column, while the spatiotemporal salinity distribution was highly affected by the basin’s interaction with the nearby Messolonghi lagoon. The anoxic zone extends from 20 m below the surface to the bottom of the Etoliko basin in summer, while the bottom layer was hypoxic during winter. Bacterial populations were analyzed by Automated Ribosomal Intergenic Spacer Analysis (ARISA). Bacterial richness and diversity were calculated and compared across samples. Hierarchical analysis showed that ARISA clustered the surface water samples according to seasonal variation, while sediment and near-to-bottom water samples appear to be stable and to cluster together. Non-metric multidimensional scaling (MDS) indicates that bacterial composition depends on dissolved oxygen and salinity. Increase in salinity of the ecosystem leads to a significant reduction of the microbial diversity.     

Presence of Extensive Wolbachia Symbiont Insertions Discovered in the Genome of Its Host Glossina morsitans morsitans

Tsetse flies (Glossina spp.) are the cyclical vectors of Trypanosoma spp., which are unicellular parasites responsible for multiple diseases, including nagana in livestock and sleeping sickness in humans in Africa. Glossina species, including Glossina morsitans morsitans (Gmm), for which the Whole Genome Sequence (WGS) is now available, have established symbiotic associations with three endosymbionts: Wigglesworthia glossinidia, Sodalis glossinidius and Wolbachia pipientis (Wolbachia). The presence of Wolbachia in both natural and laboratory populations of Glossina species, including the presence of horizontal gene transfer (HGT) events in a laboratory colony of Gmm, has already been shown. We herein report on the draft genome sequence of the cytoplasmic Wolbachia endosymbiont (cytWol) associated with Gmm. By in silico and molecular and cytogenetic analysis, we discovered and validated the presence of multiple insertions of Wolbachia (chrWol) in the host Gmm genome. We identified at least two large insertions of chrWol, 527,507 and 484,123 bp in size, from Gmm WGS data. Southern hybridizations confirmed the presence of Wolbachia insertions in Gmm genome, and FISH revealed multiple insertions located on the two sex chromosomes (X and Y), as well as on the supernumerary B-chromosomes. We compare the chrWol insertions to the cytWol draft genome in an attempt to clarify the evolutionary history of the HGT events. We discuss our findings in light of the evolution of Wolbachia infections in the tsetse fly and their potential impacts on the control of tsetse populations and trypanosomiasis.     

Assessing global range expansion in a cryptic species complex: insights from the red seaweed genus Asparagopsis (Florideophyceae)

The mitochondrial genetic diversity, distribution and invasive potential of multiple cryptic operational taxonomic units (OTUs) of the red invasive seaweed Asparagopsis were assessed by studying introduced Mediterranean and Hawaiian populations. Invasive behavior of each Asparagopsis OTU was inferred from phylogeographic reconstructions, past historical demographic dynamics, recent range expansion assessments and future distributional predictions obtained from demographic models. Genealogical networks resolved Asparagopsis gametophytes and tetrasporophytes into four A. taxiformis and one A. armata cryptic OTUs. Falkenbergia isolates of A. taxiformis L3 were recovered for the first time in the western Mediterranean Sea and represent a new introduction for this area. Neutrality statistics supported past range expansion for A. taxiformis L1 and L2 in Hawaii. On the other hand, extreme geographic expansion and an increase in effective population size were found only for A. taxiformis L2 in the western Mediterranean Sea. Distribution models predicted shifts of the climatically suitable areas and population expansion for A. armata L1 and A. taxiformis L1 and L2. Our integrated study confirms a high invasive risk for A. taxiformis L1 and L2 in temperate and tropical areas. Despite the differences in predictions among modelling approaches, a number of regions were identified as zones with high invasion risk for A. taxiformis L2. Since range shifts are likely climate‐driven phenomena, future invasive behavior cannot be excluded for the rest of the lineages.     

Changes in race-specific virulence in Pseudomonas syringae pv. phaseolicola are associated with a chimeric transposable element and rare deletion events in a plasmid-borne pathogenicity island

Virulence for bean and soybean is determined by effector genes in a plasmid-borne pathogenicity island (PAI) in race 7 strain 1449B of Pseudomonas syringae pv. phaseolicola. One of the effector genes, avrPphF, confers either pathogenicity, virulence, or avirulence depending on the plant host and is absent from races 2, 3, 4, 6, and 8 of this pathogen. Analysis of cosmid clones and comparison of DNA sequences showed that the absence of avrPphF from strain 1448A is due to deletion of a continuous 9.5-kb fragment. The remainder of the PAI is well conserved in strains 1448A and 1449B. The left junction of the deleted region consists of a chimeric transposable element generated from the fusion of homologs of IS1492 from Pseudomonas putida and IS1090 from Ralstonia eutropha. The borders of the deletion were conserved in 66 P. syringae pv. phaseolicola strains isolated in different countries and representing the five races lacking avrPphF. However, six strains isolated in Spain had a 10.5-kb deletion that extended 1 kb further from the right junction. The perfect conservation of the 28-nucleotide right repeat of the IS1090 homolog in the two deletion types and in the other 47 insertions of the IS1090 homolog in the 1448A genome strongly suggests that the avrPphF deletions were mediated by the activity of the chimeric mobile element. Our data strongly support a clonal origin for the races of P. syringae pv. phaseolicola lacking avrPphF.