Phylogenetic Analysis of Algal Symbionts Associated with Four North American Amphibian Egg Masses

Egg masses of the yellow-spotted salamander Ambystoma maculatum form an association with the green alga “Oophila amblystomatis” (Lambert ex Wille), which, in addition to growing within individual egg capsules, has recently been reported to invade embryonic tissues and cells. The binomial O. amblystomatis refers to the algae that occur in A. maculatum egg capsules, but it is unknown whether this population of symbionts constitutes one or several different algal taxa. Moreover, it is unknown whether egg masses across the geographic range of A. maculatum, or other amphibians, associate with one or multiple algal taxa. To address these questions, we conducted a phylogeographic study of algae sampled from egg capsules of A. maculatum, its allopatric congener A. gracile, and two frogs: Lithobates sylvatica and L. aurora. All of these North American amphibians form associations with algae in their egg capsules. We sampled algae from egg capsules of these four amphibians from localities across North America, established representative algal cultures, and amplified and sequenced a region of 18S rDNA for phylogenetic analysis. Our combined analysis shows that symbiotic algae found in egg masses of four North American amphibians are closely related to each other, and form a well-supported clade that also contains three strains of free-living chlamydomonads. We designate this group as the ‘Oophila’ clade, within which the symbiotic algae are further divided into four distinct subclades. Phylogenies of the host amphibians and their algal symbionts are only partially congruent, suggesting that host-switching and co-speciation both play roles in their associations. We also established conditions for isolating and rearing algal symbionts from amphibian egg capsules, which should facilitate further study of these egg mass specialist algae.     

Symbiotic green algae in eggs of Hynobius nigrescens,an amphibian endemic to Japan

The egg capsules of some amphibians' eggs are known to become green colored before hatching. This is due to the increase of green symbionts in the egg capsule surrounding the embryo. The green symbionts in North American amphibian eggs were reported to be unicellular green algae in the Oophila‐clade of Volvocales, Chlorophyceae. However, it remains unclear whether this is also the case in other parts of the world. In this study, we analyzed the green symbionts in green‐colored eggs of Hynobius nigrescens, an amphibian endemic to Japan, obtained from five distinct locations. Microscopic observations revealed that the green symbionts were similar in appearance to Oophila amblystomatis, which was reported in some amphibian eggs in North America, in which non‐motile cells of the algae had thick cell walls with reticulate protuberances. PCR‐DGGE followed by phylogenetic analyses of partial 18S rRNA sequences revealed that the symbionts from the five locations were identical and most likely unialgal in each egg capsule. They formed an independent subclade within the Oophila‐clade, indicating that H. nigrescens has a unique symbiont. Our data are consistent with the previous report on North American amphibian eggs and support the specific symbiotic relationships between Oophila‐clade symbionts and the eggs of amphibians. This is the first report on the specific symbiont‐and‐host association between an Oophila‐clade symbiont and an amphibian outside of North America. We also discuss several possibilities regarding the origin of green symbionts (vertical transmission or invasion) on the basis of the discovery and detailed observation of H. nigrescens eggs without any green symbionts.     

Dynamics of the growth,life history transformation and photosynthetic capacity of Oophila amblystomatis (Chlorophyceae), a green algal symbiont associated with embryos of the northeastern yellow spotted salamander Ambystoma maculatum (Amphibia)

The recent discovery that the unicellular green alga Oophila amblystomatis, invades embryonic tissues and cells of the salamander Ambystoma maculatum prompted us to investigate the growth and life history transformations of the algal symbionts in egg capsules. During embryonic development, symbionts were first detected microscopically as a cohesive population of swimming cells in the vicinity of the blastopore around embryonic stage 17. This population of cells grew and at embryonic stage 25, a fraction of the population began to affix to the inside of the egg capsule. Cells then underwent syngamy, lost flagella, and transformed into non-motile cells. We observed a linear increase in the accumulation of such capsule-associated cells from embryonic stage 25 to 40. The population of zoospores did not grow over this period and showed a declining trend between stage 39 and 40. We verified the population growth by measuring relative chlorophyll a content and also measured quantum yield (QY) of photosystem II (PS II) using pulse amplitude modulated (PAM) fluorometry. The population, but not the cell size, of non-motile capsule membrane-bound cells increased modestly during a one-month period after hatching, and continued to contain high levels of chlorophyll a and photosynthetic capacity. We conclude that O. amblystomatis undergoes a life history transition in egg capsules and speculate that many of these symbionts become zygotes, rather than invading the embryo.     

Free access of published DNA sequences facilitates regular control of (meta-) data quality – an example from shorebird mitogenomes (Aves,Charadriiformes: Charadrius)

Online repositories of DNA sequences are a rich and indispensable source of comparative data for biodiversity research and taxonomic studies. Despite increasingly high data quality of published sequences and associated metadata, particular attention should be paid to taxonomic assignment of DNA sequences, in particular if voucher specimens are not available or cannot be examined. In this study, two nearly identical mitogenomes of two distinctive plover species (Charadrius alexandrinus and Charadrius placidus) were re-analysed and compared with a comprehensive dataset of DNA-barcode sequences (cytochrome-oxidase subunit 1, COI) for 55 shorebird species. Phylogenetic analysis separated the two plover species into two reciprocally monophyletic clades that differed by mean p-distances of 11.5–14.7%; however, the COI sequence from the C. placidus mitogenome was nested in the Kentish Plover clade (C. alexandrinus). A similar mismatch was found for another DNA-barcode sequence from a Charadrius mongolus mitogenome that clustered with one of two clades of Charadrius leschenaultii in the COI tree. These results strongly suggest that, to date, two of seven mitogenomes published for Charadriidae are not representative of the taxon names to which the respective GenBank entries were assigned. Only a few DNA-barcode sequences were associated with outdated taxonomy, while others were suspected to be chimeric sequences. Thus, free access to digital sequence information is a key factor for steady improvement of data quality in online repositories via swarm intelligence of the scientific community.     

Reinstatement of Myelophycus caespitosus (Scytosiphonaceae,Phaeophyceae) from Japan

Reinstatement of Myelophycus caespitosus Kjellman (Ectocarpales s.l., Phaeophyceae) described from western Japan is proposed based on the comparisons of DNA sequences of Japanese Myelophycus specimens, the type specimen of Chordaria simplex and lectotypified specimen of Myelophycus caespitosus. In the genetic analyses using mitochondrial cox1 and cox3, chloroplast atpB, psbA and rbcL DNA sequences, the specimens morphologically referable to M. simplex formed two distinct clades (clade‐1 and clade‐2) supported by high statistical values. Clade‐1 was distributed on the western coast of Japan (Honshu, Shikoku and Kyushu) from the Kii Peninsula to Tsushima and the Pacific coast of Aomori, northeastern Honshu, and clade‐2 on the Pacific coast of central Honshu from Sanriku to the Kii Peninsula. Based on the cox3 DNA sequences and the location of the type locality at Shimoda, Izu Peninsula, clade‐2 was concluded to correspond to true M. simplex. There were no recognizable differences in the representative morphological features (height and diameter of sporophytes and gametophytes, number of cells comprising cortical, subcortical and medullary layers, thickness of plurilocular gametangia, or length and diameter of unilocular zoidangia) between the specimens included in the two taxa. Analyses of the specimens of the two taxa, including sympatric populations on the Kii Peninsula, using a nuclear genetic marker ocm3 did not suggest any genetic exchanges between the two taxa. On the other hand, cox3 gene sequence of the voucher specimen of Myelophycus caespitosus collected from Goto, Kyushu, western Japan housed in the UPS herbarium was included in clade‐1. In conclusion, we propose the reinstatement of M. caespitosus and to lectotypify the specimen in UPS. Korean Myelophycus specimens reported from Cheju Island and Wando were considered to belong to M. caespitosus based on the reported rbcL sequences.     

MITOCHONDRIAL DNA IN THE OOGAMOCHLAMYS CLADE (CHLOROPHYCEAE): HIGH GC CONTENT AND UNIQUE GENOME ARCHITECTURE FOR GREEN ALGAE1

Most mitochondrial genomes in the green algal phylum Chlorophyta are AT‐rich, circular‐mapping DNA molecules. However, mitochondrial genomes from the Reinhardtii clade of the Chlorophyceae lineage are linear and sometimes fragmented into subgenomic forms. Moreover, Polytomella capuana, from the Reinhardtii clade, has an elevated GC content (57.2%). In the present study, we examined mitochondrial genome conformation and GC bias in the Oogamochlamys clade of the Chlorophyceae, which phylogenetic data suggest is closely related to the Reinhardtii clade. Total DNA from selected Oogamochlamys taxa, including four Lobochlamys culleus (H. Ettl) Pröschold, B. Marin, U. G. Schlöss. et Melkonian strains, Lobochlamys segnis (H. Ettl) Pröschold, B. Marin, U. G. Schlöss. et Melkonian, and Oogamochlamys gigantea (O. Dill) Pröschold, B. Marin, U. G. Schlöss. et Melkonian, was subjected to Southern blot analyses with cob and cox1 probes, and the results suggest that the mitochondrial genome of these taxa is represented by multiple‐sized linear DNA fragments with overlapping homologies. On the basis of these data, we propose that linear mitochondrial DNA with a propensity to become fragmented arose in an ancestor common to the Reinhardtii and Oogamochlamys clades or even earlier in the evolutionary history of the Chlorophyceae. Analyses of partial cob and cox1 sequences from these Oogamochlamys taxa revealed an unusually high GC content (49.9%–65.1%) and provided evidence for the accumulation of cob and cox1 pseudogenes and truncated sequences in the mitochondrial genome of all L. culleus strains examined.     

Molecular determination of kleptoplast origins from the sea slug Plakobranchus ocellatus (Sacoglossa,Gastropoda) reveals cryptic bryopsidalean (Chlorophyta) diversity in the Hawaiian Islands

The sacoglossan sea slug species complex Plakobranchus ocellatus is a common algivore throughout the tropical Pacific, including the Hawaiian Islands. Plakobranchus ocellatus is kleptoplastic—it sequesters and retains algal chloroplasts—a characteristic that can be exploited to molecularly characterize diminutive bryopsidalean algae that are typically difficult to locate, collect, and identify. Previous DNA barcode analyses of both P. ocellatus and its kleptoplasts have been conducted primarily in the western Pacific and have only minimally sampled the most eastern populations in the Hawaiian Islands. Using two chloroplast markers, rbcL and tufA, kleptoplast samples from an Oahu population of P. ocellatus were amplified and cloned to identify their algal sources. Plakobranchus ocellatus sequester chloroplasts from up to 11 bryopsidalean algal species, all but one being diminutive in thallus size. Notably, eight of the detected algal species were new records to the Hawaiian Islands. A sequestration preference study demonstrated that the O‘ahu population of P. ocellatus preferentially sequesters chloroplasts from diminutive, epilithic taxa. Using coxI barcoding of P. ocellatus, we showed the O‘ahu population to be part of a clade that includes sequences from the neighboring island Maui, Australia, and the Philippines. The use of P. ocellatus as a novel sampling tool allows the exploration of the green algal community diversity and composition at a fine scale.     

First record of Caulerpa lentillifera J. Agardh (Bryopsidales,Chlorophyta) from China

ABSTRACT

An edible green algal species Caulerpa lentillifera J. Agardh is reported from China for the first time. The species was collected from the southwest of Hainan Island and morphologically identified to be C. lentillifera based on the grape-like branches arising from cylindrical stolons. Phylogenetic analysis using tufA and rbcL DNA sequences also confirmed the monophyly of C. lentillifera-microphysa clade.     

Australian Laurencia majuscula (Rhodophyta,Rhodomelaceae) and the Brazilian Laurencia dendroidea are conspecific

Morphological and molecular studies have been undertaken on two species of the red algal genus Laurencia J.V.Lamouroux: Laurencia majuscula (Harvey) A.H.S. Lucas and Laurencia dendroidea J.Agardh, both from their type localities. The phylogenetic position of these species was inferred by analysis of the chloroplast‐encoded rbcL gene sequences from 24 taxa. In all phylogenetic analyses, the Australian Laurencia majuscula and the Brazilian L. dendroidea formed a well‐supported monophyletic clade within the Laurencia sensu stricto. This clade was divided into two subclades corresponding to each geographical region; however, the genetic divergence between Australian L. majuscula and Brazilian L. dendroidea was only 0–1.35%. Examination of the type specimens and sequences of freshly collected samples of both Laurencia majuscula and L. dendroidea show the two to be conspecific despite their disjunct type localities.     

Systematic studies of the antarctic species of the phyllophoraceae (Gigartinales,Rhodophyta) based on rbcL sequence analysis

The taxonomic placement of four antarctic species of the marine red algal family Phyllophoraceae (Gigartinales) is assessed within a preliminary molecular phylogeny of the family based on direct sequence analysis of the chloroplast gene rbcL. Parsimony analysis of rbcL sequences indicates that Gymnogongrus antarcticus and Gymnogongrus turquetii cluster in a clade consisting predominantly of southern hemisphere species currently placed in Gymnogongrus and Ahnfeltiopsis, whereas Phyllophora ahnfeltioides and Phyllophora antarctica cluster in a separate clade that is widely divergent from the northern hemisphere Phyllophora clade. Results from molecular and morphological data challenge the current taxonomic concept that type of life history is a phylogenetically valid criterion for recognition of genera in the Phyllophoraceae.     

Ulocladium systematics revisited: phylogeny and taxonomic status

The genus Ulocladium represents phaeodictyosporic Hyphomycetes that produce conidia that are essentially obovoid in shape. Previous molecular studies that included Ulocladium and related taxa in Alternaria, Embellisia, and Stemphylium revealed a conflict between morphology and phylogeny, and Ulocladium was supported as polyphyletic with a paraphyletic core group. Moreover, the genus consistently resolved within a larger Alternaria/Ulocladium clade, resulting in paraphyly of Alternaria and questions as to the taxonomic status of Ulocladium. In the present study, 13 Ulocladium species and three genetic loci were included for a more comprehensive systematic analysis of the genus than had previously been conducted. Total genomic DNA was extracted from representative taxa and sequences were determined for the nuclear internal transcribed spacer region, including the 5.8S rDNA gene, and the protein-coding genes glyceraldehyde-3-phosphate dehydrogenase and Alt a1. Subsequent phylogenetic analyses based on maximum parsimony and Bayesian methods included related Alternaria, Embellisia, and Stemphylium spp. Results supported previous findings of polyphyletic and paraphyletic relationships of Ulocladium among other taxa. Ten Ulocladium species clustered into a core Ulocladium clade and all taxa possessed the key diagnostic feature of Ulocladium, namely, conidia essentially obovoid in shape. However, A. cheiranthi and E. indefessa also clustered within this group with high bootstrap support but did not posses this diagnostic feature. This paraphyletic clade resolved basal to the core Alternaria clade with high bootstrap support, unlike previous studies in which its position was imbedded within the primary Alternaria clade. Thus, the status of the genus as an independent lineage and a unique taxon is strongly supported. As previously reported, U. alternariae and U. oudemansii, which posses the key conidium characteristics of Ulocladium, clustered as a separate clade sister to the core Ulocladium clade. Further studies are necessary to determine if these taxa represent an independent lineage or share a common ancestor with other Ulocladium species. Obovoid conidia were poorly represented in the isolate of U. lanuginosum that was included in these analyses (the only U. lanuginosum isolate currently available), and the isolate resolved as A. radicina based upon all three loci sequenced. Based upon these data and the origin of the isolate, which was originally deposited as A. malvae, a reassessment of its identity is supported.     

MOLECULAR EVOLUTION OF GLUTAMINE SYNTHETASE II AND III IN THE CHROMALVEOLATES1

Glutamine synthetase (GS) is encoded by three distinct gene families (GSI, GSII, and GSIII) that are broadly distributed among the three domains of life. Previous studies established that GSII and GSIII isoenzymes were expressed in diatoms; however, less is known about the distribution and evolution of the gene families in other chromalveolate lineages. Thus, GSII cDNA sequences were isolated from three cryptophytes (Guillardia theta D. R. A. Hill et Wetherbee, Cryptomonas phaseolus Skuja, and Pyrenomonas helgolandii Santore), and GSIII was sequenced from G. theta. Red algal GSII sequences were obtained from Bangia atropurpurea (Mertens ex Roth) C. Agardh; Compsopogon caeruleus (Balbis ex C. Agardh) Mont.; Flintiella sanguinaria F. D. Ott and Porphyridium aerugineum Geitler; Rhodella violacea (Kornmann) Wehrmeyer and Dixoniella grisea (Geitler) J. L. Scott, S. T. Broadwater, B. D. Saunders, J. P. Thomas et P. W. Gabrielson; and Stylonema alsidii (Zanardini) K. M. Drew. In Bayesian inference and maximum‐likelihood (ML) phylogenetic analyses, chromalveolate GSII sequences formed a weakly supported clade that nested among sequences from glaucophytes, red algae, green algae, and plants. Red algal GSII sequences formed two distinct clades. The largest clade contained representatives from the Cyanidiophytina and Rhodophytina and grouped with plants and green algae. The smaller clade (C. caeruleus, Porphyra yezoensis, and S. alsidii) nested within the chromalveolates, although its placement was unresolved. Chromalveolate GSIII sequences formed a well‐supported clade in Bayesian and ML phylogenies, and mitochondrial transit peptides were identified in many of the sequences. There was strong support for a stramenopile‐haptophyte‐cryptophyte GSIII clade in which the cryptophyte sequence diverged from the deepest node. Overall, the evolutionary history of the GS gene families within the algae is complex with evidence for the presence of orthologous and paralogous sequences, ancient and recent gene duplications, gene losses and replacements, and the potential for both endosymbiotic and lateral gene transfers.     

Phylogeny of Oedogoniales (Chlorophyceae, Chlorophyta) inferred from 18S rDNA sequences with emphasis on the relationships in the genus Oedogonium based on ITS-2 sequences

The phylogeny of Oedogoniales was investigated by using nuclear 18S rDNA sequences. Results showed that the genus Oedocladium, as a separated clade, was clustered within the clade of Oedogonium; whereas the genus Bulbochaete was in a comparatively divergent position to the other two genera. The relationship among the species of Oedogonium was discussed, focusing on ITS-2 phylogeny analyzed combining with some morphological characteristics. Our results showed that all the dioecious nannandrous taxa involved in this study were resolved into one clade, while all the monocious taxa were clustered into another clade as a sister group to the former. The report also suggests that the dioecious macrandrous taxa form a paraphyly and could be more basally situated than the dioecious nannandrous and the monoecious taxa by means of molecular phylogeny and morphotype investigations.     

进境德国云杉原木中夹带栎树猝死病菌的检疫鉴定

[目的] 检测自德国进境云杉原木夹杂的水青冈植物叶片上是否携带栎树猝死病菌。[方法] 采用磁珠法提取水青冈植物叶片DNA,根据巢式PCR方法和DNA序列测定分析方法,进行栎树猝死病菌的检测。[结果] 采用巢式PCR方法,能够扩增出约280 bp的特异性条带;DNA序列测定表明,该DNA序列与GenBank中多个栎树猝死病菌分离物DNA序列相似性达99%,位于同一个发育分支。[结论] 样品中检出栎树猝死病菌,这是我国口岸首次从进境德国云杉原木中截获栎树猝死病菌。     

BIOGEOGRAPHIC ANALYSIS OF THE GLOBALLY DISTRIBUTED HARMFUL ALGAL BLOOM SPECIES ALEXANDRIUM MINUTUM (DINOPHYCEAE) BASED ON rRNA GENE SEQUENCES AND MICROSATELLITE MARKERS1

The toxic dinoflagellate Alexandrium minutum Halim is one of three species that comprise the “minutum” species complex. This complex is notable due to its role in the etiology of paralytic shellfish poisoning (PSP). Recent increases in PSP incidence and the geographic expansion of toxin‐producing Alexandrium dinoflagellates have prompted the intensive examination of genetic relationships among globally distributed strains to address questions regarding their present distribution and reasons for their apparent increase. The biogeography of A. minutum was studied using large subunit ribosomal DNA gene (LSU rRNA) and internal transcribed spacer (ITS) sequences and genotypic data from 12 microsatellite loci. rRNA gene and ITS sequencing data distinguished between two clades, herein termed the “Global” and the “Pacific”; however, little to no resolution was seen within each clade. Genotypic data from 12 microsatellite loci provided additional information regarding genetic relationships within the Global clade, but it was not possible to amplify DNA from the Pacific clade using these markers. With the exception of isolates from Italy and Spain, strains generally clustered according to origin, revealing geographic structuring within the Global clade. Additionally, no evidence supported the separation of A. lusitanicum and A. minutum as different species. With the use of microsatellites, it is now possible to initiate studies on the origin, history, and genetic heterogeneity of A. minutum that were not previously possible using only rRNA gene sequence data. This study demonstrates the power of combining a marker with intermediate resolution (rRNA sequences) with finer‐scale markers (microsatellites) to examine intraspecies variability among globally distributed isolates and represents the first effort to employ this technique in A. minutum.     

Phylogenetic relationship of Clauseneae (Rutaceae) inferred from plastid and nuclear DNA data and taxonomic implication for some major taxa

Using sequences of the nuclear ribosomal ITS region as well as the chloroplast DNA trnL‐trnF and atpB‐rbcL regions, this study aims to provide further insight into the phylogenetic relationships within Clauseneae and its relationship to Citreae (Rutaceae). Using maximum likelihood (ML) and Bayesian inference (BI), we reconstructed the phylogeny of Clauseneae based on trnL‐F, atpB‐rbcL and ITS sequences. Our data matrix contained 91 accessions, representing 72 species and varieties from 31 genera and two outgroups, including new and extensive sampling of species and varieties representing four genera in the tribe Clauseneae. In the subfamily Aurantioideae, six major clades were resolved with strong support: 1) Micromelum clade: Micromelum; 2) Glycosmis clade: Glycosmis; 3) Bergera clade: Murraya sect. Bergera; 4) Clausena clade: Clausena; 5) Murraya clade: Murraya sect. Murraya + Merrillia; 6) Citreae clade: Citreae. Micromelum, Glycosmis, Clausena and Merrillia were confirmed as monophyletic. In contrast, Murraya s.l. was reconstructed as polyphyletic. Murraya sect. Bergera clustered with Clausena while Murraya sect. Murraya and Merrillia together formed a clade that is sister to the tribe Citreae. All members from Citreae were clustered into a natural group. The genus Micromelum was found to be primitive in this subfamily and more close to Glycosmis. Based on the phylogeny and morphological characters, we discuss the taxonomy of some members of Clauseneae and conclude that the current tribal and generic classification need further revision.     

Occurrence of 16SrIV Subgroup A Phytoplasmas in Roystonea regia and Acrocomia mexicana Palms with Lethal Yellowing‐like Syndromes in Yucatán,Mexico

The lethal yellowing (LY) disease and LY‐type syndromes affecting several palm species are associated with 16SrIV phytoplasmas in the Americas. In Mexico, palms of the species Roystonea regia and the native Acrocomia mexicana were found to exhibit LY‐type symptoms, including leaf decay, starting with mature leaves, necrosis and atrophy of inflorescences. DNA extracts obtained from these palms could be amplified by nested‐PCR using phytoplasma‐universal primer pair P1/P7 followed by LY‐group‐specific primer pair LY16Sr/LY16Sf. Blast analysis of the sequences obtained revealed an identity of 100% for R. regia and 99.27% for A. mexicana with 16SrIV‐A strain associated with LY in Florida, USA (Acc. AF498309 ). Computer‐simulated RFLP analysis showed that the patterns for the phytoplasma DNA of the two palm species were highly similar to that for 16SrIV subgroup A strain. A neighbour‐joining tree was constructed, and the sequences of the two palm species clustered in the same clade of group 16SrIV subgroup A. The results therefore support that LY‐type syndromes observed in palms of R. regia and A. mexicana in the Yucatan region of Mexico are associated with 16SrIV subgroup A phytoplasmas.     

Genomic characterization of closely related species in the Rumoiensis clade infers ecogenomic signatures to non-marine environments

Members of the family Vibrionaceae are generally found in marine and brackish environments, playing important roles in nutrient cycling. The Rumoiensis clade is an unconventional group in the genus Vibrio, currently comprising six species from different origins including two species isolated from non-marine environments. In this study, we performed comparative genome analysis of all six species in the clade using their complete genome sequences. We found that two non-marine species, Vibrio casei and Vibrio gangliei, lacked the genes responsible for algal polysaccharide degradation, while a number of glycoside hydrolase genes were enriched in these two species. Expansion of insertion sequences was observed in V. casei and Vibrio rumoiensis, which suggests ongoing genomic changes associated with niche adaptations. The genes responsible for the metabolism of glucosylglycerate, a compound known to play a role as compatible solutes under nitrogen limitation, were conserved across the clade. These characteristics, along with genes encoding species-specific functions, may reflect the habit expansion which has led to the current distribution of Rumoiensis clade species. Genome analysis of all species in a single clade give us valuable insights into the genomic background of the Rumoiensis clade species and emphasize the genomic diversity and versatility of Vibrionaceae.     

Systematics of Cladophora spp. (Chlorophyta) from North Carolina,USA, based upon morphology and DNA sequence data with a description of Cladophora subtilissima sp. nov.

Identification of Cladophora species is challenging due to conservation of gross morphology, few discrete autapomorphies, and environmental influences on morphology. Twelve species of marine Cladophora were reported from North Carolina waters. Cladophora specimens were collected from inshore and offshore marine waters for DNA sequence and morphological analyses. The nuclear‐encoded rRNA internal transcribed spacer regions (ITS) were sequenced for 105 specimens and used in molecular assisted identification. The ITS1 and ITS2 region was highly variable, and sequences were sorted into ITS Sets of Alignable Sequences (SASs). Sequencing of short hyper‐variable ITS1 sections from Cladophora type specimens was used to positively identify species represented by SASs when the types were made available. Secondary structures for the ITS1 locus were also predicted for each specimen and compared to predicted structures from Cladophora sequences available in GenBank. Nine ITS SASs were identified and representative specimens chosen for phylogenetic analyses of 18S and 28S rRNA gene sequences to reveal relationships with other Cladophora species. Phylogenetic analyses indicated that marine Cladophorales were polyphyletic and separated into two clades, the Cladophora clade and the “Siphonocladales” clade. Morphological analyses were performed to assess the consistency of character states within species, and complement the DNA sequence analyses. These analyses revealed intra‐ and interspecific character state variation, and that combined molecular and morphological analyses were required for the identification of species. One new report, Cladophora dotyana, and one new species Cladophora subtilissima sp. nov., were revealed, and increased the biodiversity of North Carolina marine Cladophora to 14 species.     

Analysis of Facultative Lithotroph Distribution and Diversity on Volcanic Deposits by Use of the Large Subunit of Ribulose 1,5-Bisphosphate Carboxylase/Oxygenase

A 492- to 495-bp fragment of the gene coding for the large subunit of the form I ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) (rbcL) was amplified by PCR from facultatively lithotrophic aerobic CO-oxidizing bacteria, colorless and purple sulfide-oxidizing microbial mats, and genomic DNA extracts from tephra and ash deposits from Kilauea volcano, for which atmospheric CO and hydrogen have been previously documented as important substrates. PCR products from the mats and volcanic sites were used to construct rbcL clone libraries. Phylogenetic analyses showed that the rbcL sequences from all isolates clustered with form IC rbcL sequences derived from facultative lithotrophs. In contrast, the microbial mat clone sequences clustered with sequences from obligate lithotrophs representative of form IA rbcL. Clone sequences from volcanic sites fell within the form IC clade, suggesting that these sites were dominated by facultative lithotrophs, an observation consistent with biogeochemical patterns at the sites. Based on phylogenetic and statistical analyses, clone libraries differed significantly among volcanic sites, indicating that they support distinct lithotrophic assemblages. Although some of the clone sequences were similar to known rbcL sequences, most were novel. Based on nucleotide diversity and average pairwise difference, a forested site and an 1894 lava flow were found to support the most diverse and least diverse lithotrophic populations, respectively. These indices of diversity were not correlated with rates of atmospheric CO and hydrogen uptake but were correlated with estimates of respiration and microbial biomass.