<Emphasis Type="Italic">Gynichthys diakidnus</Emphasis> n. g., n. sp. (Digenea: Cryptogonimidae) from the grunt <Emphasis Type="Italic">Plectorhinchus gibbosus</Emphasis> (Lacépède, 1802) (Perciformes: Haemulidae) off the Great Barrier Reef,Australia

Gynichthys diakidnus n. g., n. sp. (Digenea: Cryptogonimidae) is described from the fish Plectorhinchus gibbosus (Lacépède) (Perciformes: Haemulidae) off Heron and Lizard Islands on the Great Barrier Reef, Australia. The monotypic Gynichthys n. g. is distinguished from all other cryptogonimid genera by the combination of a fusiform body, the lack of oral spines, a forebody that occupies approximately half or more of the body length, a deeply lobed ovary, opposite to slightly oblique testes, a seminal vesicle that is confined mainly in the forebody and the presence of multiple gonotyls in the form of two small slightly muscular pores or pseudosucker-like structures in the mid-line well anterior to the ventral sucker. Bayesian inference analysis of LSU rDNA data revealed that G. diakidnus n. sp. grouped relatively distant to species of the cryptogonimid genus Oligogonotylus Watson, 1976, which also have multiple gonotyls, suggesting that the presence of multiple gonotyls is homoplasious and has thus at least evolved twice in the family. The secondary structure of the internal transcribed spacer 2 (ITS2) rDNA region was inferred for G. diakidnus using minimum free energy and homology modelling algorithms. A four helix model was inferred with helices I and IV being relatively short (<30 nucleotides) and helix three being the longest; this structure is homologous with that observed for other digeneans and eukaryotes in general.     

<Emphasis Type="Italic">Sphingomonas rosea</Emphasis> sp. nov. and <Emphasis Type="Italic">Sphingomonas swuensis</Emphasis> sp. nov., rosy colored <Emphasis Type="Italic">β</Emphasis>-glucosidase-producing bacteria isolated from soil

Two strains PB196^T and PB62^T of Gram-negative, non-motile, and non-spore-forming bacteria, were isolated from soil in South Korea and characterized to determine their taxonomic positions. 16S rRNA gene sequence analysis showed that the two strains belonged to the genus Sphingomonas. The highest degree of sequence similarity of strain PB196^T was found with PB62^T (98.9%), Sphingomonas humi PB323^T (98.9%), Sphingomonas kaistensis PB56^T (98.2%), and Sphingomonas astaxanthinifaciens TDMA-17^T (98.0%). The highest degree of sequence similarity of strain PB62^T was found with Sphingomonas humi PB323^T (98.8%), Sphingomonas astaxanthinifaciens TDMA-17^T (98.2%), and Sphingomonas kaistensis PB56^T (98.1%). Chemotaxonomic data revealed that they possessed ubiquinone-10 (Q-10) as common in the genus Sphingomonas, that the predominant fatty acids were summed feature 7 (C_18:1 ω7c/ω9t/ω12t), summed feature 4 (C_16:1 ω7c/C_15:0 iso 2OH), C_16:0, and C_17:1 ω6c, and that they contained sphingoglycolipid, phosphatidylglycerol (PG), and phosphatidyle-thanolamine (PE) in common but they showed difference for diphosphatidylglycerol (DPG). Based on these data, PB196^T (=KCTC 12339^T =JCM 16604^T) and PB62^T (=KCTC 12336^T =JCM 16605^T =KEMB 9004-005^T) should be classified as type strains of two novel species, for which the names Sphingomonas rosea sp. nov. and Sphingomonas swuensis sp. nov. are proposed, respectively.     

Bioinformatics Characterization of Potential New Beta-Glucuronidase from <Emphasis Type="Italic">Streptococcus</Emphasis> <Emphasis Type="Italic">equi</Emphasis> subsp. <Emphasis Type="Italic">zooepidemicus</Emphasis>

Recently, the gene coding for a new beta-glucuronidase enzyme has been identified and cloned from Streptococcus equi subsp. zooepidemicus. This is another report of a beta-glucuronidase gene cloned from bacterial species. The ORF Finder analysis of a sequenced DNA (EMBL, AJ890474) revealed a presence of 1,785 bp large ORF potentially coding for a 594 aa protein. Three protein families in (Pfam) domains were identified using the Conserved Domain Database (CDD) analysis: Pfam 02836, glycosyl hydrolases family 2, triose phosphate isomerase (TIM) barrel domain; Pfam 02837, glycosyl hydrolases family 2, sugar binding domain; and Pfam 00703, glycosyl hydrolases family 2, immunoglobulin-like beta-sandwich domain. To gain more insight into the enzymatic activity, the domains were used to generate a bootstrapped unrooted distance tree using ClustalX. The calculated distances for two domains, TIM barrel domain, and sugar-binding domain were comparable and exhibited similarity pattern based on function and thus being in accordance with recently published works confirming beta-glucuronidase activity of the enzyme. The calculated distances and the tree arrangement in the case of centrally positioned immonoglobulin-like beta-sandwich domain were somewhat higher when compared to other two domains but clustering with other beta-glucuronidases was rather clear. Nine proteins, including beta-glucuronidases, beta-galactosidase, and mannosidase were selected for multiple alignment and subsequent distance tree creation.     

Description of “<Emphasis Type="Italic">Desulfotomaculum nigrificans</Emphasis> subsp. <Emphasis Type="Italic">salinus</Emphasis>” as a New Species, <Emphasis Type="Italic">Desulfotomaculum salinum</Emphasis> sp. nov.

This study focused on the physiological, chemotaxonomic, and genotypic characteristics of two thermophilic spore-forming sulfate-reducing bacterial strains, 435^T and 781, of which the former has previously been assigned to the subspecies “Desulfotomaculum nigrificans subsp. salinus”. Both strains reduced sulfate with the resulting production of H₂S on media supplemented with H₂ + CO₂, formate, lactate, pyruvate, malate, fumarate, succinate, methanol, ethanol, propanol, butanol, butyrate, valerate, or palmitate. Lactate oxidation resulted in acetate accumulation; butyrate was oxidized completely, with acetate as an intermediate product. Growth on acetate was slow and weak. Sulfate, sulfite, thiosulfate, and elemental sulfur, but not nitrate, served as electron acceptors for growth with lactate. The bacteria performed dismutation of thiosulfate to sulfate and hydrogen sulfide. In the absence of sulfate, pyruvate but not lactate was fermented. Cytochromes of b and c types were present. The temperature and pH optima for both strains were 60–6°C and pH 7.0. Bacteria grew at 0 to 4.5–6.0% NaCl in the medium, with the optimum being at 0.5–1.0%. Phylogenetic analysis based on a comparison of incomplete 16S rRNA sequences revealed that both strains belonged to the C cluster of the genus Desulfotomaculum, exhibiting 95.5–98.3% homology with the previously described species. The level of DNA–DNA hybridization of strains 435^T and 781 with each other was 97%, while that with closely related species D. kuznetsovii 17^T was 51–52%. Based on the phenotypic and genotypic properties of strains 435^T and 781, it is suggested that they be assigned to a new species: Desulfotomaculum salinum sp. nov., comb. nov. (type strain 435^T = VKM B 1492^T).     

New family of pectinase genes <Emphasis Type="Italic">PGU1</Emphasis>b–<Emphasis Type="Italic">PGU3b</Emphasis> of the pectinolytic yeast <Emphasis Type="Italic">Saccharomyces bayanus</Emphasis> var. <Emphasis Type="Italic">uvarum</Emphasis>

Using yeast genome databases and literature data, we have conducted a phylogenetic analysis of pectinase PGU genes from Saccharomyces strains assigned to the biological species S. arboricola, S. bayanus (var. uvarum), S. cariocanus, S. cerevisiae, S. kudriavzevii, S. mikatae, S. paradoxus, and hybrid taxon S. pastorianus (syn. S. carlsbergensis). Single PGU genes were observed in all Saccharomyces species, except S. bayanus. The superfamily of divergent PGU genes has been documented in S. bayanus var. uvarum for the first time. Chromosomal localization of new PGU1b, PGU2b, and PGU3b genes in the yeast S. bayanus var. uvarum has been determined by molecular karyotyping and Southern hybridization.     

Mating system and seed variation of <Emphasis Type="Italic">Acacia</Emphasis> hybrid (<Emphasis Type="Italic">A. mangium</Emphasis> × <Emphasis Type="Italic">A. auriculiformis</Emphasis>)

The mating system and seed variation of Acacia hybrid (A. mangium × A. auriculiformis) were studied using allozymes and random amplified polymorphic DNA (RAPD) markers, respectively. Multi-locus outcrossing rate estimations indicated that the hybrid was predominantly outcrossed (mean±s.e. t _m = 0.86±0.01). Seed variation was investigated using 35 polymorphic RAPD fragments. An analysis of molecular variance (AMOVA) revealed the highest genetic variation among seeds within a pod (66%–70%), followed by among pods within inflorescence (29%–37%), and the least variation among inflorescences within tree (<1%). In addition, two to four RAPD profiles could be detected among seeds within pod. Therefore, the results suggest that a maximum of four seeds per pod could be sampled for the establishment of a mapping population for further studies.     

Characteristics of β-lactamases and their genes (<Emphasis Type="Italic">blaA</Emphasis> and <Emphasis Type="Italic">blaB</Emphasis>) in <Emphasis Type="Italic">Yersinia intermedia</Emphasis> and <Emphasis Type="Italic">Y. frederiksenii</Emphasis>

Background  

The presence of β-lactamases in Y. enterocolitica has been reported to vary with serovars, biovars and geographical origin of the isolates. An understanding of the β-lactamases in other related species is important for an overall perception of antibiotic resistance in yersiniae. The objective of this work was to study the characteristics of β-lactamases and their genes in strains of Y. intermedia and Y. frederiksenii, isolated from clinical and non-clinical sources in India.     

<Emphasis Type="Italic">Empis</Emphasis> (<Emphasis Type="Italic">Pachymeria</Emphasis>) <Emphasis Type="Italic">roditakisi</Emphasis>—a New Species of Dance Flies (Diptera: Empididae) from Greece

A new species of dance flies (Diptera, Empididae) of the Empis subgenus Pachymeria Stephens, 1829, E. (P.) roditakisi sp. n. from Greece is described and illustrated.     

<Emphasis Type="Italic">Caligus ignotus</Emphasis> n. sp. (Copepoda: Caligidae) parasitic on Poey’s scabbardfish <Emphasis Type="Italic">Evoxymetopon poeyi</Emphasis> (Günther) off Taiwan

A new species of caligid copepod, Caligus ignotus n. sp., is described from Poey’s scabbardfish Evoxymetopon poeyi (Günther) (Trichiuridae) caught off the southeast coast of Taiwan. The parasite was found in the gill-cavity of this benthopelagic fish. The new species is distinguished from its congeners mainly in the structure of the female genital complex (shaped like a guitar) and abdomen (with three sections and gradually broadening from anterior to posterior). This is the first report of a copepod parasite from Poey’s scabbardfish.     

Biological Characterisation of <Emphasis Type="Italic">Haliclona</Emphasis> (?<Emphasis Type="Italic">gellius</Emphasis>) sp.: Sponge and Associated Microorganisms

We have characterised the northern Pacific undescribed sponge Haliclona (?gellius) sp. based on rDNA of the sponge and its associated microorganisms. The sponge is closely related to Amphimedon queenslandica from the Great Barrier Reef as the near-complete 18S rDNA sequences of both sponges were identical. The microbial fingerprint of three specimens harvested at different times and of a transplanted specimen was compared to identify stably associated microorganisms. Most bacterial phyla were detected in each sample, but only a few bacterial species were determined to be stably associated with the sponge. A sponge-specific β- and γ-Proteobacterium were abundant clones and both of them were present in three of the four specimens analysed. In addition, a Planctomycete and a Crenarchaea were detected in all sponge individuals. Both were closely related to operational taxonomic units that have been found in other sponges, but not exclusively in sponges. Interestingly, also a number of clones that are closely related to intracellular symbionts from insects and amoeba were detected.     

Functional expression of the lipase gene Lip2 of <Emphasis Type="Italic">Pleurotus</Emphasis> <Emphasis Type="Italic">sapidus</Emphasis> in <Emphasis Type="Italic">Escherichia</Emphasis> <Emphasis Type="Italic">coli</Emphasis>

The lipase Lip2 of the edible basidiomycete, Pleurotus sapidus, is an extracellular enzyme capable of hydrolysing xanthophyll esters with high efficiency. The gene encoding Lip2 was expressed in Escherichia coli TOP10 using the gene III signal sequence to accumulate proteins in the periplasmatic space. The heterologous expression under control of the araBAD promoter led to the high level production of recombinant protein, mainly as inclusion bodies, but partially in a soluble and active form. A fusion with a C-terminal His tag was used for purification and immunochemical detection of the target protein. This is the first example of a heterologous expression and periplasmatic accumulation of a catalytically active lipase from a basidiomycete fungus.     

Brood cell size of <Emphasis Type="Italic">Apis</Emphasis> <Emphasis Type="Italic">mellifera</Emphasis> modifies the reproductive behavior of <Emphasis Type="Italic">Varroa</Emphasis> <Emphasis Type="Italic">destructor</Emphasis>

We undertook a field study to determine whether comb cell size affects the reproductive behavior of Varroa destructor under natural conditions. We examined the effect of brood cell width on the reproductive behavior of V. destructor in honey bee colonies, under natural conditions. Drone and worker brood combs were sampled from 11 colonies of Apis mellifera. A Pearson correlation test and a Tukey test were used to determine whether mite reproduction rate varied with brood cell width. Generalized additive model analysis showed that infestation rate increased positively and linearly with the width of worker and drone cells. The reproduction rate for viable mother mites was 0.96 viable female descendants per original invading female. No significant correlation was observed between brood cell width and number of offspring of V. destructor. Infertile mother mites were more frequent in narrower brood cells.     

<Emphasis Type="Italic">Sarcocystis menglaensis</Emphasis> n. sp. (Apicomplexa: Sarcocystidae) from Williamson’s mouse deer <Emphasis Type="Italic">Tuagulus williamsoni</Emphasis> (Kloss) (Artiodactyla: Tragulidae)

Williamson’s mouse deer, Tuagulus williamsoni (Kloss), is one of the smallest ungulates among tragulid species found in northern Thailand, and Yunnan Province, China. Here we describe Sarcocystis menglaensis n. sp., infecting two of 14 (14.3%) Williamson’s mouse deer from south-western China. By light microscopy, sarcocysts of S. menglaensis are microscopic, up to 2,170 μm in length, and have a striated sarcocyst wall with 1.5–3.6 μm long palisade-like protrusions. Transmission electron microscopy observations revealed that sarcocyst wall is of “type 10f”, and has numerous villar protrusions folded over the cyst wall. The villar protrusions contained microtubules dispersed throughout the protrusions. Phylogenetic analysis based on 18S rDNA and mitochondrial cox1 gene sequences indicated that S. menglaensis shared a close affinity with species of Sarcocystis Lankester, 1982 from ruminants, which utilise felids as definitive hosts.     

Regeneration of transformed verbena (<Emphasis Type="Italic">Verbena </Emphasis>× <Emphasis Type="Italic">hybrida</Emphasis>) by <Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>

Verbena (Verbena x hybrida), an important floricultural species, was successfully regenerated from stem segments on Murashige and Skoog's basal medium supplemented with thidiazuron and indole-3-acetic acid. A transformation system was developed using cvs. Temari Scarlet, Temari Sakura, Tapien Rose and TP-P2. Agrobacterium tumefaciens strain Agl0 harboring the sGFP gene was infected into stem segments. Transformation efficiency was improved by evaluating and manipulating the age of the plant material, the concentration of kanamycin in the medium during selection, and the length of the culture period in the dark. After 2-3 months of culture on the selection medium, GFP-positive shoots were obtained in all four of the cultivars tested. These shoots were successfully acclimated and set flowers within 2-3 months in a greenhouse. GFP was expressed in all of the organs including the floral parts. Stable genomic transformation was confirmed by Southern blot analysis. No morphological differences were observed between the transformed plants and their host plants.     

In vitro growth response of <Emphasis Type="Italic">Phytophthora cactorum</Emphasis>, <Emphasis Type="Italic">P. nicotianae</Emphasis> and <Emphasis Type="Italic">P.</Emphasis> × <Emphasis Type="Italic">pelgrandis</Emphasis> to antibiotics and fungicides

The reactions of isolates of Phytophthora cactorum, P. nicotianae and P. × pelgrandis to metalaxyl, mancozeb, dimethomorph, streptomycin and chloramphenicol were tested to obtain information about the variability of resistance in these pathogens. Distinct genetic groups showed significant differences in resistance to all tested substances except streptomycin. In response to streptomycin, the growth inhibition rates of distinct groups did not differ significantly. The most remarkable differences were detected in the reactions to chloramphenicol and metalaxyl. Discriminant analysis evaluating the effect of all substances confirmed the differences among the groups, which are in agreement with the differences revealed by earlier DNA analyses.     

Screening interspecific hybrids of<Emphasis Type="Italic"> Populus</Emphasis> (<Emphasis Type="Italic">P. ciliata</Emphasis> ×<Emphasis Type="Italic"> maximowiczii</Emphasis>) using AFLP markers

Hybrids of Populus ciliata × maximowiczii are very vigorous and outperform both the parents in growth performance and yield. Genetic evaluation of 24 of these interspecific hybrids along with the two mother trees (Populus ciliata), and five male-parent (Populus maximowiczii) genotypes was carried out using the AFLP marker assay. Eight AFLP primer combinations detected 428 markers, of which 280 (66%) were polymorphic. Genetic relationships within the samples were evaluated by generating the similarity matrix based on Jaccards coefficient. The phenetic dendrograms, as well as the PCO plots, separated the hybrids and the two parent species into three distinct clusters. The hybrids grouped closer to the P. ciliata (female parent) cluster as compared to the P. maximowiczii (male parent) cluster. The hybrid cluster contained internal groupings, which correlated to some extent with growth performance. The four best performing hybrids (42m1, 65m1, 23m2, Cm2-5-20/91) formed a distinct sub-cluster. Data from a single primer combination was sufficient for distinguishing the hybrids from the parents and assigning paternity. The hybrids showed 22 markers that were absent in P. ciliata but were monomorphically present in all the hybrids, suggesting outcrossing and common paternity. Further, these 22 markers were found in all the P. maximowiczii genotypes confirming it as the male parent. These male-specific markers can be converted to SCAR markers and used for rapid screening of the P.ciliata × maximowiczii hybrids. The primer combination E-AAC × M-CAA was identified as most suitable for ascertaining true hybridity. AFLP proves to be a useful tool for screening of P. ciliata × maximowiczii hybrids at the early stages of development.Communicated by H.F. Liskens     

<Emphasis Type="Italic">Amphibiophilus mooiensis</Emphasis> n. sp. (Nematoda: Amphibiophilidae), a parasite of <Emphasis Type="Italic">Amietia delalandii</Emphasis> (Duméril &amp; Bibron) (Amphibia: Pyxicephalidae) from South Africa

Four species of the genus Amphibiophilus Skrjabin, 1916 from pyxycephalid frogs in southern and central Africa are currently recognised as valid. Several specimens of Amphibiophilus were found in material from the common river frog, Amietia delalandii (Duméril & Bibron) (Amphibia: Pyxicephalidae), collected in Potchefstroom (North-West Province, South Africa). These specimens clearly differ from all previously known species by the shape of the distal end of the spicule, the shape of the gubernaculum and the structure of the synlophe. They are, thus, considered as a new species, Amphibiophilus mooiensis. As all other species in the family Amphibiophilidae Durette-Desset & Chabaud, 1981, A. mooiensis n. sp. possesses a number of archaic characters, such as a buccal capsule with a well-developed dorsal oesophageal tooth, six inner labial papillae, six outer labial papillae and four cephalic papillae. Molecular data (cox1 and ITS-28S rDNA sequences) are provided and host and geographical specificity are discussed.     

Rapid induction of <Emphasis Type="Italic">Agrobacterium</Emphasis> <Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transgenic roots directly from adventitious roots in <Emphasis Type="Italic">Panax</Emphasis> <Emphasis Type="Italic">ginseng</Emphasis>

Root segments from seedlings of Panax ginseng produced adventitious roots directly when cultured on 1/2 MS solid medium lacking NH₄NO₃ and containing 3.0 mg l⁻¹ IBA. Using this adventitious root formation, we developed rapid and efficient transgenic root formation directly from adventitious root segments in P. ginseng. Root segments were co-cultivated with Agrobacterium tumefaciens (GV3101) caring β-glucuronidase (GUS) gene. Putative transgenic adventitious roots were formed directly from root segments on medium with 400 mg l⁻¹ cefotaxime and 50 mg l⁻¹ kanamycin. Kanamycin resistant adventitious roots were selected and proliferated as individual lines by subculturing on medium with 300 mg l⁻¹ cefotaxime and 50 mg l⁻¹ kanamycin at two weeks subculture interval. Frequency of transient and stable expression of GUS gene was enhanced by acetosyringon (50 mg l⁻¹) treatment. Integration of transgene into the plants was confirmed by the X-gluc reaction, PCR and Southern analysis. Production of transgenic plants was achieved via somatic embryogenesis from the embryogenic callus derived from independent lines of adventitious roots. The protocol for rapid induction of transgenic adventitious roots directly from adventitious roots can be applied for a new Agrobacterium tumefaciens-mediated genetic transformation protocol in P. ginseng.     

<Emphasis Type="Italic">Marcetia candolleana</Emphasis> (<Emphasis Type="Italic">Melastomeae</Emphasis> – <Emphasis Type="Italic">Melastomataceae</Emphasis>), a new species from Bahia (Brazil)

Summary   Marcetia candolleana A. K. A. Santos & A. B. Martins, is apparently restricted to Mucugê, Bahia (Brazil), where it occurs in areas of campo rupestre vegetation. This new species is closely related to the sympatric M. mucugensis Wurdack, but can be easily recognised by its semi-prostate to procumbent habit, reddish glandular-hirsute indument, loose and flexuous branches, leaves with inconspicuous reticulation on the abaxial surface, connectives very shortly prolonged below the thecae, style curved towards the apex, not exceeding the anthers, and pendulous fruit.

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Resumo   Marcetia candolleana A. K. A. Santos & A. B. Martins, é aparentemente restrita a Mucugê, Bahia (Brasil), onde ocorre em áreas de campo rupestre. Esta nova espécie é proximamente relacionada à M. mucugensis Wurdack, mas pode ser facilmente reconhecida por seu hábito semi-prostado a procumbente, indumento glandular-hirsuto, vináceo, ramos flexuosos, folhas inconspicuamente reticuladas na face abaxial, conectivos muito curtamente prolongados abaixo das tecas, estilete curvo no ápice, n?o ultrapassando o comprimento das anteras, e fruto pêndulo.