Comparative study on gut microbiota in three Anura frogs from a mountain stream

Composition and diversity in gut microbiota are impacted by a wide variety of factors. The similarity of gut microbiota in related or sympatric species has been gaining recent traction. Here, 16S rRNA gene sequencing technology was employed to study the gut microbiota of three sympatric frog species, namely Odorrana tormota, O. graminea, and Amolops wuyiensis. In these three frog species, the most abundant phylum was Proteobacteria, followed by Bacteroidetes, Verrucomicrobia, and Firmicutes. The most abundant family was Burkholderiaceae in three species. The most dominant genera were Burkholderia, Caballeronia, and Paraburkholderia with the highest relative abundance in O. tormota, O. graminea, and A. wuyiensis, respectively. No differences were observed in alpha diversity indexes among the three frog species. However, bacterial similarity of gut microbiota was significantly different between O. tormota and A. wuyiensis and between O. graminea and A. wuyiensis. Metabolism‐related gene function was predominantly enriched in the gut microbiota of the three evaluated frog species. From these findings, that the relative abundance of the gut microbiota and predicted gene functions differed in three species, we conclude that there were significant differences in the gut microbiota of the three species. Similar alpha diversity and interspecific bacterial similarity in the gut might be related to bacterial transmission among the three Anura frogs evaluated in this study.     

Structure of mitochondrial DNA control region and molecular phylogenetic relationship among three flounders of genus Pleuronectes

Structure of mitochondrial DNA control region about three flounders – Pleuronectes yokohama, Pleuronectes schrenki and Pleuronectes herzensteini – were reported. The TAS, cTAS, CSB-A to CSB-F and CSB-1 to CSB-3 were detected in these three flounders. The results indicated that the structures of these parts were different from most fishes. All the mtDNA control region sequences of the three founders have tandem repeat sequences in the downstream of CSB-3, which is different from most vertebrates. According to the structure of the mtDNA control region, P. yokohama was more similar with P. schrenki and P. herzensteini was much different from the other two species. In addition, three segments such as control region, Cytb and COI are used to analyze the phylogenic relationships of the three species. The genetic distances and phylogenetic tree results support the classification by traditional morphology. It is not clear if P. yokohama and P. schrenki belong to the same species, and this should be accepted with caution.     

The effect of parasitism by the mermithid Romanomermis culicivorax on the dry weight and hemolymph soluble protein content of three species of mosquitoes

The dry weight, hemolymph soluble protein composition, and content of three species of mosquitoes, Culex pipiens, Aedes taeniorhynchus, and Anopheles quadrimaculatus were examined to determine the effects of parasitism by the mermithid nematode Romanomermis culicivorax. The dry weights of infected fourth-stage larvae of all three species were significantly lower than controls. The differences in weight found between infected early and late C. pipiens and A. quadrimaculatus larvae were attributed to the weight of the parasite itself. This difference was not noticeable in A. taeniorhynchus larvae. Hemolymph proteins were severely depleted in all three mosquito species during parasitism by R. culicivorax. Analysis of protein composition by PAGE showed that these depletions were accompanied by a reduction in the number of proteins. Differences between protein composition concentrations were evident between early and late fourthstage control larvae of C. pipiens and A. quadrimaculatus. The concentration of some low-molecular-weight proteins (below 68,000) remained constant between infected and control samples of all three mosquito species.     

Morpho-anatomical affinities and evolutionary relationships of three paleoendemic podocarp genera based on seed cone traits

Background and AimsThe three relict genera Pherosphaera, Microcachrys and Saxegothaea in Podocarpaceae produce quite distinct seed cone types in comparison with other genera and do not form a clade along with Acmopyle. The detailed seed cone morpho-anatomy of these three relict genera and affinities with other podocarps are poorly known. This study aims to understand the seed cone morpho-anatomy and affinities among these three disjunct relict genera and with other podocarps.MethodsWe comparatively analysed the seed cone morpho-anatomical traits of the three podocarps genera and used ancestral state reconstruction to understand the evolution of these traits.Key ResultsWe described the seed cone morpho-anatomical structures of the three relict genera in detail. The three genera produce aggregated multiovulate cones. Both Microcachrys and Saxegothaea have an asymmetrical free cup-like epimatium. Both species of Pherosphaera lack an epimatium. The ancestral state reconstruction implies that the presence of an epimatium is an ancestral trait in podocarps and is independently lost in Pherosphaera and Phyllocladus. The seed cones are fleshy in Microcachrys and non-fleshy in Saxegothaea and Pherosphaera. The seed cone macrofossils of both extinct and living podocarps also show the presence of an epimatium and fleshiness in podocarps.ConclusionsAltogether, the morpho-anatomy suggests that Pherosphaera, Microcachrys and Saxegothaea present affinities with each other and other podocarps, but the reconstruction of the ancestral seed cone in Podcarpaceae is quite complex due to multiple convergent evolutions of several structures. These structures (e.g. epimatium, aril and receptaculum) are of low taxonomic value but of great evolutionary and ecological significance, and are responsive adaptations to ever-changing environmental conditions.     

Wood anatomy of Phragmotheca (Bombacaceae)

The wood anatomy ofPhragmotheca (Bombacaceae) is described based on three species and three specimens. The wood features ofPhragmotheca, Matisia, andQuararibea are very similar and differ only in the size of the intervascular pits and vessel diameters. Based on wood anatomical characteristics, these three genera form a distinct and homogeneous group within both the tribe Quararibeae and the family Bombacaceae.     

Complete mitochondrial genomes of three sea basses Lateolabrax (Perciformes,Lateolabracidae) species: Genome description and phylogenetic considerations

Genus Lateolabrax consists of three species, Japanese sea bass Lateolabrax japonicus, spotted sea bass Lateolabrax maculatus and blackfin sea bass Lateolabrax latus. The complete mitochondrial DNA (mtDNA) of the three sea basses were amplified and sequenced to characterize and discuss their phylogenetic relationships. The length of mitogenomes was 16,593 bp, 16,479 bp and 16,600 bp, respectively, and all of them consisted of 13 protein-coding genes, 2 ribosomal RNA (rRNA), 22 transfer RNA (tRNA) and a control region, which are typical for mtDNA of vertebrate. Most genes were encoded on the H-strand, except for the ND6 and eight tRNA genes encoding on the L-strand. A significant variation among the three species was detected in length of the control region. Phylogenetic relationship among the three species was constructed based on the datasets, including the 12 protein-coding genes (except ND6 gene), 22 tRNA and 2 rRNA sequences. The results supported the sister taxon between L. japonicus and L. maculatus. The genetic resources reported here are useful for further studies in taxonomy and phylogeny of the three sea basses and related species.     

Guaianolides from Centaurea canariensis

The reinvestigation of Centaurea canariensis var. subexpinnata provided, along with three known compounds, three new sesquiterpene lactones of the guaiane series.     

Molecular identification and relative abundance of cryptic Lophodermium species in natural populations of Scots pine,Pinus sylvestris L.

The multi-locus phylogenetic species recognition approach and population genetic analysis of Amplified Fragment Length Polymorphism (AFLP) markers were used to delineate Lophodermium taxa inhabiting needles of Scots pine (Pinus sylvestris) in native pinewoods within Scotland. These analyses revealed three major lineages corresponding to the morphological species Lophodermium seditiosum and Lophodermium conigenum, fruiting on broken branches, and Lophodermium pinastri, fruiting on naturally fallen needles. Within L. pinastri three well supported sister clades were found representing cryptic taxa designated L. pinastri I, L. pinastri II, and L. pinastri III. Significant differences in mean growth rate in culture were found among the cryptic taxa. Taxon-specific primers based on ITS sequences were designed and used to classify over 500 Lophodermium isolates, derived from fallen needles of P. sylvestris in three Scottish and one French pinewood site, into the three L. pinastri cryptic taxa. Highly significant differences in the relative abundance of the three taxa were found among the Scottish pinewood sites, and between the French and all of the Scottish sites.     

Clostridium botulinum Strain Af84 Contains Three Neurotoxin Gene Clusters: Bont/A2, bont/F4 and bont/F5

Sanger and shotgun sequencing of Clostridium botulinum strain Af84 type Af and its botulinum neurotoxin gene (bont) clusters identified the presence of three bont gene clusters rather than the expected two. The three toxin gene clusters consisted of bont subtypes A2, F4 and F5. The bont/A2 and bont/F4 gene clusters were located within the chromosome (the latter in a novel location), while the bont/F5 toxin gene cluster was located within a large 246 kb plasmid. These findings are the first identification of a C. botulinum strain that contains three botulinum neurotoxin gene clusters.     

Wood anatomy of obolinga (Mimosaceae)

The wood anatomy ofObolinga zanonii Barneby (tribe Ingeae) is described based on three specimens representing three trees. The wood ofObolinga is very similar to that ofCojoba, a segregate of thePithecellobium complex. They differ only in the color and fluorescence of water and ethanol extracts.     

Cereal crop volatile organic compound induction after mechanical injury, beetle herbivory (Oulema spp.), or fungal infection (Fusarium spp.)

Herbivory, mechanical injury or pathogen infestation to vegetative tissues can induce volatile organic compounds (VOCs) production, which can provide defensive functions to injured and uninjured plants. In our studies with ‘McNeal’ wheat, ‘Otana’ oat, and ‘Harrington’ barley, plants that were mechanically injured, attacked by either of two Oulema spp. (melanopus or cyanella) beetles, or infected by one of the three Fusarium spp. (graminearum, avenaceum, or culmorum), had significant VOC induction compared to undamaged plants. Mechanical injury to the main stem or one leaf caused the induction of one green leaf volatile (GLV) - (Z)-3-hexenol, and three terpenes (β-linalool, β-caryophyllene, and α-pinene) with all three grasses; wheat and barley also showed β-linalool oxide induction. The blend of induced VOCs after Fusarium spp. infestation or Oulema spp. herbivory was dominated by GLVs ((Z)-3-hexenal, (E)-2-hexenal, (E)-2-hexenol, (Z)-3-hexenyl acetate, and 1-hexenyl acetate) and β-linalool and β-caryophyllene; beetle herbivory also induced (E)-β-farnesene. Different ratios of individual VOCs were induced between the two Oulema spp. for each cereal grass and different ratios across the three cereals for each beetle species. Also, different ratios of individual VOCs were induced between the three Fusarium spp. for each cereal grass and different ratios across the three cereals for each fungal pathogen species. Our results are preliminary since we could not simultaneously measure VOC induction from controls with each of the ten different injury treatments for each of the three cereals. However, the comparison of mechanical injury, insect herbivory, and fungal infection has not been previously examined with VOC responses from three different plant species within the same family. Also, our work suggests large qualitative and quantitative overlap of VOC induction from plants of all three cereals having beetle herbivory injury when compared to infection injury from necrotrophic fungal pathogens.     

Fine root dynamic characteristics and effect on plantation’s carbon sequestration of three Salix shrub plantations in Tibetan Plateau alpine sandy land

Desertification land in Gonghe Basin of Tibetan Plateau, China accounts for 91.9% of the total land area. Vegetation restoration and reconstruction with desert shrubs in degraded ecosystem are effective ways to prevent and control desertification. However, the evaluation studies of fine root dynamic characteristics of desert shrubs and their contribution to carbon sequestration of plantation are limited. To gain a better understanding of vegetation restoration, the vertical distribution of fine root biomass, fine root decomposition, fine root turnover was investigated, as well as their coupling effect on carbon sequestration of plantation in three desert vegetation. The results estimated that the total decomposition time of fine roots of Salix cheilophila (S. cheilophila), Salix psammophila (S. psammophila), and Salix microstachya (S. microstachya) are 39.00, 27.99 and 35.95 years. Biomass carbon density for three Salix plantations ranged from 1.42 to 2.39 t/hm², which showed that three Salix plantations in alpine sandy land are an important carbon pool. In addition, fine root biomass carbon density for the three shrub plantations varied significantly. Fine root biomass carbon density for S. psammophila reached the largest among the three plantations, which was 1.48 t/hm², accounting for the ratio of 62% of the plantation total biomass carbon density. The results indicated that the root system of S. psammophila, especially the fine roots, was very developed, which was conducive to soil water transportation and carbon sequestration. Therefore, S. psammophila might be a better species for carbon sequestration of plantation in alpine sandy areas. The carbon input from the fine roots of the three shrub plantations through decomposition and turnover into the plantations accounts for 11.5% to 15.5% of total carbon sequestration of plantations. Therefore, the fine roots dynamics must be considered for long‐term carbon pool estimations in three Salix plantations, otherwise the total carbon sequestration of plantations would be underestimated.     

Two new species of Chrysomelobia Regenfuss, 1968 (Acariformes: Podapolipidae) from Paropsis charybdis Stål (Coleoptera: Chrysomelidae)

Two new species of Chrysomelobia Regenfuss, 1968, C. alleni n. sp. and C. intrusus n. sp., are described from Tasmanian specimens of the eucalyptus leaf beetle Paropsis charybdis Stål. This beetle is now known to host three species of Chrysomelobia, the other being Chrysomelobia pagurus Seeman, 2008, which is recorded from Tasmania for the first time. Thus, the three species of Paropsis Olivier known to have podapolipid mites each have three mite species from three separate lineages of Chrysomelobia. Collections of P. charybdis in New Zealand (n = 150), where it is an invasive pest species, failed to locate any infested beetles, suggesting that these populations were established by uninfested beetles. The prospect of using these mites as biocontrol agents is discussed.     

pH dependence of the oxidation-reduction potential of cytochrome c2

The pH dependence of the spectra and of the oxidation-reduction potential of three cytochromes c₂, from Rhodopseudomonas capsulata, Rhodopseudomonas sphaeroides and Rhodomicrobium vannielii, were studied. A single alkaline pK was observed for the spectral changes in all three ferricytochromes. In Rps. capsulata cytochrome c₂ this spectroscopic pK corresponds to the pK observed in the dependence of oxidation-reduction potential on pH. For the other two cytochromes the oxidation-reduction potential showed a complex dependency on pH which can be fitted to theoretical curves involving three ionizations. The third ionization corresponds to the ionization observed in the spectroscopic studies but the first two occur without changes in the visible spectra.The possible structural bases for these ionizations are discussed.     

GUS expression in sweet oranges (Citrus sinensis L. Osbeck) driven by three different phloem-specific promoters

Huanglongbing (HLB) is associated with Candidatus Liberibacter spp., endogenous, sieve tube-restricted bacteria that are transmitted by citrus psyllid insect vectors. Transgenic expression in the phloem of specific genes that might affect Ca. Liberibacter spp. growth and development may be an adequate strategy to improve citrus resistance to HLB. To study specific phloem gene expression in citrus, we developed three different binary vector constructs with expression cassettes bearing the β-glucuronidase (GUS) reporter gene (uidA) under the control of one of the three different promoters: Citrus phloem protein 2 (CsPP2), Arabidopsis thaliana phloem protein 2 (AtPP2), and Arabidopsis thaliana sucrose transporter 2 (AtSUC2). Transgenic lines of ‘Hamlin’, ‘Pera’, and ‘Valencia’ sweet oranges [Citrus sinensis (L.) Osbeck] were produced via Agrobacterium tumefaciens transformation. The epicotyl segments collected from in vitro germinated seedlings were used as explants. The gene nptII, which confers resistance to the antibiotic kanamycin, was used for selection. The transformation efficiency was expressed as the number of GUS-positive shoots over the total number of explants and varied from 1.54 to 6.08?% among the three cultivars and three constructs studied. Several lines of the three sweet orange cultivars analyzed using PCR and Southern blot analysis were genetically transformed with the three constructs evaluated. The histological GUS activity in the leaves indicates that the uidA gene was preferentially expressed in the phloem, which suggests that the use of the three promoters might be adequate for producing HLB-resistant transgenic sweet oranges. The results reported here conclusively demonstrate the preferential expression of GUS in the phloem driven by two heterologous and one homologous gene promoters. Key message The results reported here conclusively demonstrate the preferential expression of GUS in the phloem driven by two heterologous and one homologous gene promoters.     

Nutritional and bioactive properties of three edible species of green algae,genus Caulerpa (Caulerpaceae)

The green algae genus Caulerpa is coenocytic, and the thallus consists of only one cell with many nuclei. It is widely distributed in the tropical seas. In the Southeast Asian waters, there are at least ten known species. Three species, particularly Caulerpa racemosa var. clavifera f. macrophysa (Kützing) Weber-van Bosse, C. racemosa var. laetevirens (Montagne) Weber-van Bosse, and Caulerpa lentillifera J. Agardh are widely consumed. The proximate analysis and secondary metabolite composition of these three species were determined to describe their lipid and nutritional values. Glycolipids and phospholipids were the major lipid classes, with significant levels of triacylglycerol. Polyunsaturated fatty acids (PUFA) were the major fatty acids of all the three species. Typical n-3 and n-6 PUFA such as 18:3n-3, 18:4n-3, 20:5n-3, 18;2n-6, and 20:4n-6 were found in significant amount in all these three species. All three species contained a red-pigmented secondary metabolite determined as caulerpin. All three extracts exhibited potent antimicrobial activity against human food pathogenic bacteria and anti-inflammatory activity against the murine macrophage cell line, RAW 264.7.     

Discovery of Lower Famennian Stromatoporoidea in the Kuznetsk Basin

Stromatoporoidea recorded for the first time from the Lower Famennian of the Kuznetsk Basin are described. The assemblage studied includes three new species, Actinostroma quasifenestratum sp. nov., Kyklopora tomiensis sp. nov., and Coenostroma rarum sp. nov., and three previously known species of different genera.     

Internal Spatiotemporal Population Dynamics of Infection with Three Wolbachia Strains in the Adzuki Bean Beetle, Callosobruchus chinensis (Coleoptera: Bruchidae)

The adzuki bean beetle, Callosobruchus chinensis, is infected with three distinct lineages of endosymbiotic bacteria belonging to the genus Wolbachia, which were designated wBruCon, wBruOri, and wBruAus. In an attempt to understand the mechanisms underlying the infection with these three organisms, the spatiotemporal infection dynamics of the three Wolbachia strains was investigated in detail by using a quantitative PCR technique. During the development of C. chinensis, the wBruCon, wBruOri, and wBruAus infection levels consistently increased but the growth patterns were different. The levels of infection plateaued at the pupal stage at approximately 3 × 10⁸, 2 × 10⁸, and 5 × 10⁷ wsp copy equivalents per insect for wBruCon, wBruOri, and wBruAus, respectively. At the whole-insect level, the population densities of the three Wolbachia types did not show remarkable differences between adult males and females. At the tissue level, however, the total densities and relative levels of the three Wolbachia types varied significantly when different tissues and organs were compared and when the same tissues derived from males and females were compared. The histological data obtained by in situ hybridization and electron microscopy were concordant with the results of quantitative PCR analyses. Based on the histological data and the peculiar Wolbachia composition commonly found in nurse tissues and oocytes, we suggest that the Wolbachia strains are vertically transmitted to oocytes not directly, but by way of nurse tissue. On the basis of our results, we discuss interactions among the three coinfecting Wolbachia types, reproductive strategies of Wolbachia, and factors involved in the different cytoplasmic incompatibility phenotypes.