Structure of the seismosensory system of <Emphasis Type="Italic">Psettodes erumei</Emphasis> (Psettodidae,Pleuronectiformes)

The structure of the seismosensory system of Psettodes erumei was studied. General and specific features in comparison with the previously studied representatives of the suborder Pleuronectoidei were revealed. Similarity is manifested in the presence of four main head canals, coronal commissure, interrupted supratemporal commissure transformed into a supratemporal canal, and one trunk canal developed on both sides of the body. The seismosensory system of Psettodes erumei is characterized by several plesiomorphous characters: the presence of a sensory segment in lacrimale of the eyed side and supracleithrum, the absence of disruptions in all sensory canals, and a strong branching of cutaneous canaliculi. Specific features of P. erumei involve the presence of a large number of fontanels on angulo-articulare, connection on the eyed side of CIO sites in lacrimale and infraorbitale, and the underdevelopment of the bony vault of the sensory canal in praeoperculum.     

Thoracic member (pectoral girdle and forelimb) bones of <Emphasis Type="Italic">Mylodon darwinii</Emphasis> Owen (Xenarthra,Mylodontidae) from the Late Pleistocene of Central Argentina and their phylogenetic implications

A nearly complete skeleton, including most of the thoracic member bones of the sloth Mylodon darwinii, have been found in Upper Pleistocene strata from Anisacate River, Argentina. The thoracic member bones resemble their homologues in Glossotherium robustum, Paramylodon harlani, and Mylodonopsis ibseni in the following traits: (1) the olecranon is mediolaterally compressed; (2) the radius has an acute styloid process; (3) the radial diaphysis medial border is straight for two thirds of its length; (4) the radial shaft medial border forms an angle with the medial border of the styloid process. The radius presents a distinctive, mostly proximally facing articular circumference. The unfused epiphyses and feeble muscle attachment ridges indicate a sub-adult ontogenetic stage. Deviation of the olecranon and weak M. teres major origin and insertion, suggest a low fossorial specialization. The structure of the thoracic limb bones does not support climbing habits, because pronation-related features are reduced and the humeral head is not prominent. The structure of the radius suggests graviportal adaptations: the proximal head is mediolaterally expanded and the diaphysis straight. A phylogenetic analysis adding thoracic member characters recovers M. darwinii as part of a clade that includes Glossotherium robustum and Paramylodon harlani, but excludes Lestodon armatus. This contrasts with the results of previous analyses focusing on the head skeleton, highlighting the relevance of sampling postcranial characters in phylogenetic analyses of mylodontine sloths.     

Comparative characteristics of North Pacific and North Atlantic specimens of a rare bathypelagic fish <Emphasis Type="Italic">Thalassobathia pelagica</Emphasis> (Teleostei: Bythitidae) with discussion of its distribution

A detailed morphological comparison is made of the North Pacific specimen of Thalassobathia pelagica with fish from the North Atlantic (the type locality). The Pacific specimen is recognized as conspecific with Atlantic ones. Insignificant differences are considered as age or individual variation. The structure of the seismosensory system of T. pelagica and morphology of this species are described in more detail. The probable distribution of T. pelagica is discussed. A wide tropical area of this species is assumed and a possibility of its presence in all three oceans. The captures of T. pelagica in high latitudes may depend on its transportation with warm currents.     

New species of pelagic eelpouts of the genus <Emphasis Type="Italic">Melanostigma</Emphasis> (Zoarcidae) from Western Antarctica

A new species, Melanostigma olgae sp. n., is described from the Southern Ocean. The type series was obtained along the South Sandwich Islands (Scotia Sea, Western Antarctic) at the depths of 800?850 m. The new species differs from the other representatives of Melanostigma genus by the original combination of the features of the seismosensory system, axial skeleton, and coloration. The homologization of the structural elements (senses) of the head canals of the lateral line has been done for the first time for the Melanostigma. Morphology of M. olgae allows considering this species as the most evolutionary advanced within the representatives of this genus inhabiting the Southern Ocean. The key of the Antarctic species of the genus Melanostigma is provided.     

New deepwater species of the genus <Emphasis Type="Italic">Gobiopsis</Emphasis> (Gobiidae) from the Tropical Western Pacific

A new species, Gobiopsis uranophilus sp. n., is described off the Anambas Islands, South China Sea, from a depth of 73 m. The species is similar to the deepwater Japanese species G. namnas, but it differs in the dorsal orientation of the eyes, total absence of pores of the seismosensory system on the head, number of fin rays, and other characters. The similar patterns of the two species (presence of scales on the sides of the head, absence of the barbel-shaped appendages on the head, and large scales on the body), most likely, are connected with their adaptation to deepwater habitats.     

<Emphasis Type="Italic">Lecanicillium muscarium</Emphasis> and <Emphasis Type="Italic">Adalia bipunctata</Emphasis> combination for the control of black bean aphid<Emphasis Type="Italic">, Aphis fabae</Emphasis>

The predator Adalia bipunctata (Coleoptera: Coccinellidae) and the entomopathogenic fungus Lecanicillium muscarium, have been considered as potential biological control against aphids. While it can be difficult to achieve a high control level of Aphis fabae Scopoli (Hemiptera: Aphididae) using only a single beneficial agent, the research presented here aimed to determine the interaction between L. muscarium and A. bipunctata potential for control against A. fabae. Lecanicillium muscarium was found to cause about 30% mortality in A. bipunctata and with a reduction in feeding by about 15%. However, co-application of A. bipunctata and L. muscarium can cause an addititive effect in reducing aphid populations, resulting in about 90% reduction in aphid populations compared with control treatment. Thus, these two biocontrol agents have the potential to be complementary. This research study demonstrates that it is possible to combine A. bipunctata with L. muscarium to provide a sustainable method for management of A. fabae on broad bean cropping system and that field studies are required.     

Potential Distribution of Fossil Xenarthrans in South America during the Late Pleistocene: co-Occurrence and Provincialism

Species distribution models (SDMs) are helpful for understanding actual and potential biogeographical traits of organisms. These models have recently started to be applied in the study of fossil xenarthrans. SDMs were generated for 15 South American late Pleistocene xenarthrans: eight Cingulata (Glyptodon clavipes, Doedicurus clavicaudatus, Panochthus tuberculatus, Neosclerocalyptus paskoensis, Pampatherium typum, Pampatherium humboldtii, Holmesina paulacoutoi, and Holmesina occidentalis) and seven Folivora (Glossotherium robustum, Lestodon armatus, Mylodon darwinii, Catonyx cuvieri, Catonyx (=Scelidodon) chilensis, Megatherium americanum, and Eremotherium laurillardi). Models were evaluated for three periods: the last interglacial (LIG), the last glacial maximum (LGM), and the Holocene climatic optimum (HCO). Co-occurrence records were studied based on the overlap of the potential distributions and compared with the available biome reconstructions of South America during the LGM to analyze species distribution patterns, ecological requirements, and possible interactions. Our results suggest the existence of provincialization within xenarthran megamammals grouped in at least three bioregions. Northern and southwestern taxa overlap in the Río de la Plata region where also some endemic taxa are found. We observed overlapping potential distributions but separated and continuous realized distributions between closely related xenarthrans suggesting competitive exclusion. A generalized reduction in potential habitats at the end of the Pleistocene was not obvious as some taxa show stable potential areas during HCO when comparing with LGM. Nonetheless, fragmentation of the most suitable areas due to climate variation and the impact of reduction in available land due to sea level changes cannot be ruled out as involved in the extinction.     

Taxonomic position of Afghan vole (Subgenus <Emphasis Type="Italic">Blanfordimys</Emphasis>) by the sequence of the mitochondrial <Emphasis Type="Italic">cytb</Emphasis> gene

It is rather difficult to construct a system of gray voles of the tribe Microtini by a set of morphological and karyological characters because form generation is mosaic at these organization levels. The sequence of the mitochondrial cytochrome b gene was used to study the phylogenetic relationships and taxonomic position of the Central Asian subgenus Blanfordimys. Afghan vole Microtus (Blanfordimys) afghanus and Bucharian vole M. (Blanfordimys) bucharensis clustered with Pamir vole M. (Neodon) juldaschi, which is conventionally assigned to another subgenus. The last two species proved to be significantly closer to each other than either of them was to M. (Blanfordimys) afghanus, which disagrees with the monophyletic origin accepted for Blanfordimys. The genetic distances between the species of the subgenus Blanfordmys and M. juldaschi were comparable with the distances between the sister subgenera Microtus s. str. and Sumeriomys or Pallasiinus and Alexandromys and with the basal divergence of supraspecific clades in the subgenus Terricola. It was assumed that a special Central Asian group of species exists within the tribe Microtini and includes species of the subgenus Blanfordimys and M. juldaschi and that the subgenera Neodon and Blanfordimys should be revised.     

Nucleotide sequence analysis of <Emphasis Type="Italic">S</Emphasis>-locus genes to unify <Emphasis Type="Italic">S</Emphasis> haplotype nomenclature in radish

Radish, belonging to the family Brassicaceae, has a self-incompatibility which is controlled by multiple alleles on the S locus. To employ the self-incompatibility in an F₁ breeding system, identification of S haplotypes is necessary. Since collection of S haplotypes and determination of nucleotide sequences of SLG, SRK, and SCR alleles in cultivated radish have been conducted by different groups independently, the same or similar sequences with different S haplotype names and different sequences with the same S haplotype names have been registered in public databases, resulting in confusion of S haplotype names for researchers and breeders. In the present study, we developed S homozygous lines from radish F₁ hybrid cultivars in Japan and determined the nucleotide sequences of SCR, the S domain and the kinase domain of SRK, and the SLG of a large number of S haplotypes. Comparing these sequences with our previously published sequences, the haplotypes were ordered into 23 different S haplotypes. The sequences of the 23 S haplotypes were compared with S haplotype sequences registered by different groups, and we suggested a unification of these S haplotypes. Furthermore, dot-blot hybridization using SRK allele-specific probes was examined for developing a standard method for S haplotype identification.     

Comparative pan genome analysis of oral <Emphasis Type="Italic">Prevotella</Emphasis> species implicated in periodontitis

Prevotella is part of the oral bacterial community implicated in periodontitis. Pan genome analyses of eight oral Prevotella species, P. dentalis, P. enoeca, P. fusca, P. melaninogenica, P. denticola, P. intermedia 17, P. intermedia 17-2 and P. sp. oral taxon 299 are presented in this study. Analysis of the Prevotella pan genome revealed features such as secretion systems, resistance to oxidative stress and clustered regularly interspaced short palindromic repeat (CRISPR)-Cas systems that enable the bacteria to adapt to the oral environment. We identified the presence of type VI secretion system (T6SS) in P. fusca and P. intermedia strains. For some VgrG and Hcp proteins which were not part of the core T6SS loci, we used gene neighborhood analysis and identified putative effector proteins and putative polyimmunity loci in P. fusca and polymorphic toxin systems in P. intermedia strains. Earlier studies have identified the presence of Por secretion system (PorSS) in P. gingivalis, P. melaninogenica and P. intermedia. We noted the presence of their homologs in six other oral Prevotella studied here. We suggest that in Prevotella, PorSS is used to secrete cysteine proteases such as interpain and C-terminal domain containing proteins with a “Por_secre_tail” domain. We identified subtype I-B CRISPR-Cas system in P. enoeca. Putative CRISPR-Cas system subtypes for 37 oral Prevotella and 30 non-oral Prevotella species were also predicted. Further, we performed a BLASTp search of the Prevotella proteins which are also conserved in the red-complex pathogens, against the human proteome to identify potential broad-spectrum drug targets. In summary, the use of a pan genome approach enabled identification of secretion systems and defense mechanisms in Prevotella that confer adaptation to the oral cavity.     

Isolating and confirming the <Emphasis Type="Italic">MuDR</Emphasis>-inserted flanking sequences of maize

MuDR exhibits the highest transposition activity and insertional mutagenesis frequency in Mutator (Mu) family. If we isolate the MuDR-insertion-specific flanking sequences (MuDRFs), it will be crucial for using Mu element-mediated mutants. The MuDR-TAIL-PCR system was constructed and optimized using a combination of MuDR-TIR-nested specific primers and 12 arbitrary degenerate (AD) primers, modified reaction system and procedure and mutant DNA templates of 87 genotypes from M₂ or М_2:3 families created by crossing the W22::Mu line (active MuDR donor parent) from the UniformMu population with the Zong31 (Z31) line (recipient parent). Here 129 different MuDRFs were acquired by MuDR-TAIL-PCR, accounting for 86.60% of the total mutant-specific agarose gel bands. In addition, we confirmed the authenticity of the non-redundant flanking sequence amplifications. The amplified non-redundant flanking sequences accounted for 65.12% of the total MuDRFs, and 88.00% of the non-redundant MuDRFs were inserted inside the genes. These results show that the MuDR-TAIL-PCR system that we developed can be used for specifically isolating MuDRFs.     

Molecular phylogeny of pectinase genes <Emphasis Type="Italic">PGU</Emphasis> in the yeast genus <Emphasis Type="Italic">Saccharomyces</Emphasis>

Using yeast genome databases and literature data, phylogenetic analysis of pectinase PGU genes from 112 Saccharomyces strains assigned to the biological species S. arboricola, S. bayanus (var. uvarum), S. cariocanus, S. cerevisiae, S. kudriavzevii, S. mikatae, S. paradoxus, and the hybrid taxon S. pastorianus (syn. S. carlsbergensis) was carried out. A superfamily of divergent PGU genes was found. Natural interspecies transfer of the PGU gene both from S. cerevisiae to S. bayanus and from S. paradoxus to S. cerevisiae may, however, occur. Within the Saccharomyces species, identity of the PGU nucleotide sequences was 98.8–100% for S. cerevisiae, 86.1–95.7% for S. bayanus (var. uvarum), 94–98.3% for S. kudriavzevii, and 96.8–100% for S. paradoxus/S. cariocanus. For the first time, a family of polymeric PGU1b, PGU2b, PGU3b and PGU4b genes is documented for the yeast S. bayanus var. uvarum, a variety important for winemaking.     

Integrated gene mapping and synteny studies give insights into the evolution of a sex proto-chromosome in <Emphasis Type="Italic">Solea senegalensis</Emphasis>

The evolution of genes related to sex and reproduction in fish shows high plasticity and, to date, the sex determination system has only been identified in a few species. Solea senegalensis has 42 chromosomes and an XX/XY chromosome system for sex determination, while related species show the ZZ/ZW system. Next-generation sequencing (NGS), multi-color fluorescence in situ hybridization (mFISH) techniques, and bioinformatics analysis have been carried out, with the objective of revealing new information about sex determination and reproduction in S. senegalensis. To that end, several bacterial artificial chromosome (BAC) clones that contain candidate genes involved in such processes (dmrt1, dmrt2, dmrt3, dmrt4, sox3, sox6, sox8, sox9, lh, cyp19a1a, amh, vasa, aqp3, and nanos3) were analyzed and compared with the same region in other related species. Synteny studies showed that the co-localization of dmrt1-dmrt2-drmt3 in the largest metacentric chromosome of S. senegalensis is coincident with that found in the Z chromosome of Cynoglossus semilaevis, which would potentially make this a sex proto-chromosome. Phylogenetic studies show the close proximity of S. senegalensis to Oryzias latipes, a species with an XX/XY system and a sex master gene. Comparative mapping provides evidence of the preferential association of these candidate genes in particular chromosome pairs. By using the NGS and mFISH techniques, it has been possible to obtain an integrated genetic map, which shows that 15 out of 21 chromosome pairs of S. senegalensis have at least one BAC clone. This result is important for distinguishing those chromosome pairs of S. senegalensis that are similar in shape and size. The mFISH analysis shows the following co-localizations in the same chromosomes: dmrt1-dmrt2-dmrt3, dmrt4-sox9-thrb, aqp3-sox8, cyp19a1a-fshb, igsf9b-sox3, and lysg-sox6.     

Usefulness of the Non-conventional <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> Model to Assess <Emphasis Type="Italic">Candida</Emphasis> Virulence

Invasive candidiasis is caused mainly by Candida albicans, but other Candida species have increasing etiologies. These species show different virulence and susceptibility levels to antifungal drugs. The aims of this study were to evaluate the usefulness of the non-conventional model Caenorhabditis elegans to assess the in vivo virulence of seven different Candida species and to compare the virulence in vivo with the in vitro production of proteinases and phospholipases, hemolytic activity and biofilm development capacity. One culture collection strain of each of seven Candida species (C. albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida metapsilosis, Candida orthopsilosis and Candida parapsilosis) was studied. A double mutant C. elegans AU37 strain (glp-4;sek-1) was infected with Candida by ingestion, and the analysis of nematode survival was performed in liquid medium every 24 h until 120 h. Candida establishes a persistent lethal infection in the C. elegans intestinal tract. C. albicans and C. krusei were the most pathogenic species, whereas C. dubliniensis infection showed the lowest mortality. C. albicans was the only species with phospholipase activity, was the greatest producer of aspartyl proteinase and had a higher hemolytic activity. C. albicans and C. krusei caused higher mortality than the rest of the Candida species studied in the C. elegans model of candidiasis.     

Transmission and characterization of <Emphasis Type="Italic">bla</Emphasis><Subscript>NDM-1</Subscript> in <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> at a teaching hospital in Yunnan,China

Background

In recent years, New Delhi metallo-beta-lactamases 1 (bla _NDM-1) has been reported with increasing frequency and become prevalent. The present study was undertaken to investigate the epidemiological dissemination of the bla _NDM-1 gene in Enterobacter cloacae isolates at a teaching hospital in Yunnan, China.

Methods

Antimicrobial susceptibility testing was performed using VITEK 2 system and E test gradient strips. The presence of integrons and insertion sequence common region 1 were examined by PCR and sequencing. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Conjugation experiments and Southern blot hybridization were performed to determine the transferability of plasmids.

Results

Ten E. cloacae isolates and their Escherichia coli transconjugants were exhibited similar resistant patterns to carbapenems, cephalosporins and penicillins. 8 (80%) of E. cloacae isolates carried class 1 integron and 1 (12.5%) carried class 2 integron. Integron variable regions harbored the genes which encoded resistance to aminoglycosides (aadA1, aadA2, aadA5, aadB, aac(6′)-Ib-cr), sulfamethoxazole/trimethoprim (dfrA17, dfrA12, dfrA15) and Streptozotocin (sat2). Six E. cloacae isolates belonged to ST74 and exhibited highly similar PFGE patterns. Each isolate shared an identical plasmid with ~33.3 kb size that carried the bla _NDM-1 gene, except T3 strain, of which the bla _NDM-1 gene was located on a ~50 kb plasmid.

Conclusions

Our findings suggested that plasmid was able to contribute to the dissemination of bla _NDM-1. Hence, more attention should be devoted to monitor the dissemination of the bla _NDM-1 gene due to its horizontal transfer via plasmid. In addition, nosocomial surveillance system should actively monitor the potential endemic clone of ST74 to prevent their further spread.

     

<Emphasis Type="Italic">PiggyBac</Emphasis> transposon-mediated mutagenesis and application in yeast <Emphasis Type="Italic">Komagataella phaffii</Emphasis>

Objective

Around one-fourth of the Komagataella phaffii genes encode hypothetical proteins with unknown functions. However, lack of powerful tools for genetic screening in K. phaffii significantly limits the functional analysis of these unknown genes. Transposon mutagenesis has been utilized as an insertional mutagenesis tool in many other organisms and would be extremely valuable if it could be applied in K. phaffii.

Results

In this study, we investigated in K. phaffii the transposition activity and efficiency of piggyBac (PB) transposon, a DNA transposon from the cabbage looper moth Trichoplusia ni through the integrated-plasmid system. We also designed a binary-plasmid system which could generate stable mutants. Finally we evaluated the quality of this mutagenesis system by a simple screening for functional genes involved in K. phaffii carbon catabolite repression.

Conclusions

Our results demonstrate that PB-mediated mutagenesis could be a feasible and useful tool for functional gene screening in K. phaffii.

     

<Emphasis Type="Italic">R</Emphasis>-acetoin accumulation and dissimilation in <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis>

Klebsiella pneumoniae is a 2,3-butanediol producer, and R-acetoin is an intermediate of 2,3-butanediol production. R-acetoin accumulation and dissimilation in K. pneumoniae was studied here. A budC mutant, which has lost 2,3-butanediol dehydrogenase activity, accumulated high levels of R-acetoin in culture broth. However, after glucose was exhausted, the accumulated R-acetoin could be reused by the cells as a carbon source. Acetoin dehydrogenase enzyme system, encoded by acoABCD, was responsible for R-acetoin dissimilation. acoABCD mutants lost the ability to grow on acetoin as the sole carbon source, and the acetoin accumulated could not be dissimilated. However, in the presence of another carbon source, the acetoin accumulated in broth of acoABCD mutants was converted to 2,3-butanediol. Parameters of R-acetoin production by budC mutants were optimized in batch culture. Aerobic culture and mildly acidic conditions (pH 6–6.5) favored R-acetoin accumulation. At the optimized conditions, in fed-batch fermentation, 62.3 g/L R-acetoin was produced by budC and acoABCD double mutant in 57 h culture, with an optical purity of 98.0 %, and a substrate conversion ratio of 28.7 %.