Phylogeography of Asparagopsis taxiformis revisited: Combined mtDNA data provide novel insights into population structure in Japan

Six intraspecific lineages (Lineages 1–6) of Asparagopsis taxiformis have been previously established based on mitochondrial cox2‐cox3 intergenic spacer and a partial cox1 sequences. ‘Lineage 2’ (L2) was suggested to be a recent introduction to the Mediterranean Sea, but its source population has not yet been identified. In order to clarify the nature of northwestern Pacific populations, we performed extensive sampling in Japan (60 individuals from 16 locations) and molecular phylogenetic analyses based on mitochondrial sequences. Sixteen additional individuals, collected from eight locations in the Indo‐Pacific, Caribbean, and Mediterranean regions, were also analyzed. Combined sequence analyses revealed that the Japanese populations only consisted of L2. Out of 19 combined haplotypes identified within L2, two are shared between Japan and the Mediterranean Sea and the Hawaiian Islands, and 12 were identified as endemic to Japan. Genetic analyses of population differentiation suggested that Japanese populations are genetically isolated from the Mediterranean and the Hawaiian populations. A genetic disjunction appears to separate two subpopulations within Japan: one between Toi and Kagoshima and the other between Ojikajima Island and Kagoshima in the Kyushu area.     

FIRST EVIDENCE OF PALYTOXIN ANALOGUES FROM AN OSTREOPSIS MASCARENENSIS (DINOPHYCEAE) BENTHIC BLOOM IN SOUTHWESTERN INDIAN OCEAN1

Benthic dinoflagellates of the genus Ostreopsis Schmidt are common in tropical and subtropical water, and some species produce toxins potentially involved in human intoxication events. A benthic bloom of Ostreopsis mascarenensis Quod was observed near Rodrigues Island during a survey of benthic dinoflagellates in the southwestern Indian Ocean. The morphology of O. mascarenensis was studied by LM and SEM. Preliminary screening of a crude extract of an O. mascarenensis bloom revealed neurotoxicity in mice similar to that induced by palytoxin. After partition of the crude extract, the highest toxicity was retained in the butanol‐soluble fraction, which retained hemolytic activity suggestive of palytoxin analogues. Two new toxins, mascarenotoxin‐A and ‐B, were resolved from this fraction by HPLC coupled to a diode array detector. The closed mass spectrum profile and fragmentation pattern obtained by advanced nano–electrospray ionization quadrupole time‐of‐flight mass spectrometry between purified toxins and a reference palytoxin confirmed the mascarenotoxins as palytoxin analogues. These results were confirmed by tandem mass spectrometry with the identification of specific fragment ion m/z 327. An on‐line liquid chromatography protocol coupled to tandem mass spectrometry was developed for detection of these palytoxin analogues. The present study describes the first purification, chemical, and toxicological characterization of new palytoxin analogues isolated from a benthic bloom of O. mascarenensis. These results suggest that O. mascarenensis, which is largely distributed in the southwestern Indian Ocean, could be a source of palytoxin poisoning in this tropical area.     

New species, observations, and a list of new records of brown algae (Phaeophyceae) from the Hawaiian Islands

Two new species of brown algae (Phaeophyceae), Padina moffittiana Abbott et Huisman, sp. nov. and Cutleria irregularis Abbott et Huisman, sp. nov., are described from the Hawaiian Islands (between 19°04′N, 155°35′W and 28°25′N, 178°20′W). In addition, the new combination Cutleria canariensis is proposed for Aglaozonia canariensis. New observations are presented on Nereia intricata Yamada, a species described 67 years ago and known only from its type specimen. New records of a further 14 species are given for the Hawaiian Islands. These 17 taxa bring the total number of species of brown algae recorded for the Hawaiian Islands to 55, an increase of 28%. Of the 15 new records, two are recent ‘accidental’ introductions: Dictyota flabellata (Collins) Setchell et Gardner and Sargassum muticum (Yendo) Fensholt from California. Six records are notable because of their great distances from previously known collections: Nereia intricata Yamada from the Ryukyu Is., Japan; Discosporangium mesarthrocarpum (Meneghini) Hauck, from the Adriatic, Mediterranean, warm Atlantic and southern Australia; Distromium flabellatum Womersley, Spatoglossum macrodontum J. Agardh, and Sporochnus moorei Harvey, from Australia; Desmarestia ligulata (Lightfoot) Lamouroux from temperate and colder waters in the Pacific and Atlantic. A comparison with some Japanese species of Padina confirms that Padina japonica Yamada should be subsumed with Padina sanctae‐crucis Børgesen, as proposed earlier.     

Biomass allocation,growth, and photosynthesis of genotypes from native and introduced ranges of the tropical shrub<Emphasis Type="Italic"> Clidemia hirta</Emphasis>

We tested the hypothesis that the tropical shrub Clidemia hirta appears more shade tolerant and is more abundant in its introduced than native range because of genetic differences in resource acquisition, allocation, and phenotypic plasticity between native and introduced genotypes. We examined growth, biomass allocation, and photosynthetic parameters of C. hirta grown in a greenhouse from seed collected from four populations in part of its native range (Costa Rica) and four populations in part of its introduced range (Hawaiian Islands). Six-month-old seedlings were placed in high (10.3–13.9 mol m^–2 day^–1) or low (1.4–4.5 mol m^–2 day^–1) light treatments and grown for an additional 6 months. Our study provided little evidence that Hawaiian genotypes of C. hirta differed genetically from Costa Rican genotypes in ways that would contribute to differences in habitat distribution or abundance. Some of the genetic differences that were apparent, such as greater allocation to stems and leaf area relative to whole plant biomass in Costa Rican genotypes and greater allocation to roots in Hawaiian genotypes, were contrary to predictions that genotypes from the introduced range would allocate more biomass to growth and less to storage than those from the native range. Hawaiian and Costa Rican genotypes displayed no significant differences in relative growth rates, maximal photosynthetic rates, or specific leaf areas in either light treatment. In the high light environment, however, Hawaiian genotypes allocated more biomass to reproductive parts than Costa Rican genotypes. Phenotypic plasticity for only 1 of 12 morphological and photosynthetic variables was greater for Hawaiian than Costa Rican genotypes. We conclude that genetic shifts in resource use, resource allocation, or plasticity do not contribute to differences in habitat distribution and abundance between the native and introduced ranges of C. hirta. Electronic Supplementary Material Supplementary material is available in the online version of this article at . Clidemia hirta individuals grown in a common garden in high light for 10 months from Hawaiian (left) or Costa Rica (right) seed sources     

Palytoxin: exploiting a novel skin tumor promoter to explore signal transduction and carcinogenesis

Palytoxin is a novel skin tumor promoter, which has been used to help probe the role of different types of signaling mechanisms in carcinogenesis. The multistage mouse skin model indicates that tumor promotion is an early, prolonged, and reversible phase of carcinogenesis. Understanding the molecular mechanisms underlying tumor promotion is therefore important for developing strategies to prevent and treat cancer. Naturally occurring tumor promoters that bind to specific cellular receptors have proven to be useful tools for investigating important biochemical events in multistage carcinogenesis. For example, the identification of protein kinase C as the receptor for the prototypical skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) (also called phorbol 12-myristate 13-acetate, PMA) provided key evidence that tumor promotion involves the aberrant modulation of signaling cascades that govern cell fate and function. The subsequent discovery that palytoxin, a marine toxin isolated from zoanthids (genus Palythoa), is a potent skin tumor promoter yet does not activate protein kinase C indicated that investigating palytoxin action could help reveal new aspects of tumor promotion. Interestingly, the putative receptor for palytoxin is the Na⁺,K⁺-ATPase. This review focuses on palytoxin-stimulated signaling and how palytoxin has been used to investigate alternate biochemical mechanisms by which important targets in carcinogenesis can be modulated. mitogen-activated protein kinase; dual-specificity phosphatase; prostaglandins; sodium, potassium adenosinetriphosphatase     

Functional role of the N-terminus of Na,K-ATPase α-subunit as an inactivation gate of palytoxin-induced pump channel

The N-terminus of the Na⁺,K⁺-ATPase α-subunit shows some homology to that of Shaker-B K⁺ channels; the latter has been shown to mediate the N-type channel inactivation in a ball-and-chain mechanism. When the Torpedo Na⁺,K⁺-ATPase is expressed in Xenopus oocytes and the pump is transformed into an ion channel with palytoxin (PTX), the channel exhibits a time-dependent inactivation gating at positive potentials. The inactivation gating is eliminated when the N-terminus is truncated by deleting the first 35 amino acids after the initial methionine. The inactivation gating is restored when a synthetic N-terminal peptide is applied to the truncated pumps at the intracellular surface. Truncated pumps generate no electrogenic current and exhibit an altered stoichiometry for active transport. Thus, the N-terminus of the α-subunit appears to act like an inactivation gate and performs a critical step in the Na⁺,K⁺-ATPase pumping function.     

Properties of ion channels in the protoplasts of the Mediterranean seagrass Posidonia oceanica

Posidonia oceanica (L) Delile, a seagrass endemic of the Mediterranean sea, provides food and shelter to marine organisms. As environment contamination and variation in physico‐chemical parameters may compromise the survival of the few Posidonia genotypes living in the Mediterranean, comprehending the molecular mechanisms controlling Posidonia growth and development is increasingly important. In the present study the properties of ion channels in P. oceanica plasma membranes studied by the patch‐clamp technique in protoplasts obtained from the young non‐photosynthetic leaves were investigated. In protoplasts that were presumably originated from sheath cells surrounding the vascular bundles of the leaves, an outward‐rectifying time‐dependent channel with a single channel conductance of 58 ± 2 pS which did not inactivate, was selective for potassium and impermeable to monovalent cations such as Na⁺, Li⁺ and Cs⁺ was identified. In the same protoplasts, an inward‐rectifying channel that has a time‐dependent component with single channel conductance of the order of 10 pS, a marked selectivity for potassium and no permeation to sodium was also identified, as was a third type of channel that did not display any ionic selectivity and was reversibly inhibited by tetraethylammonium and lanthanum. A comparison of Posidonia channel characteristics with channels identified in terrestrial plants and other halophytic plants is included.     

Area and the rapid radiation of Hawaiian Bidens (Asteraceae)

Aim To estimate the rate of adaptive radiation of endemic Hawaiian Bidens and to compare their diversification rates with those of other plants in Hawaii and elsewhere with rapid rates of radiation. Location Hawaii. Methods Fifty‐nine samples representing all 19 Hawaiian species, six Hawaiian subspecies, two Hawaiian hybrids and an additional two Central American and two African Bidens species had their DNA extracted, amplified by polymerase chain reaction and sequenced for four chloroplast and two nuclear loci, resulting in a total of approximately 5400 base pairs per individual. Internal transcribed spacer sequences for additional outgroup taxa, including 13 non‐Hawaiian Bidens, were obtained from GenBank. Phylogenetic relationships were assessed by maximum likelihood and Bayesian inference. The age of the most recent common ancestor and diversification rates of Hawaiian Bidens were estimated using the methods of previously published studies to allow for direct comparison with other studies. Calculations were made on a per‐unit‐area basis. Results We estimate the age of the Hawaiian clade to be 1.3–3.1 million years old, with an estimated diversification rate of 0.3–2.3 species/million years and 4.8 × 10^?5 to 1.3 × 10^?4 species Myr^?1 km^?2. Bidens species are found in Europe, Africa, Asia and North and South America, but the Hawaiian species have greater diversity of growth form, floral morphology, dispersal mode and habitat type than observed in the rest of the genus world‐wide. Despite this diversity, we found little genetic differentiation among the Hawaiian species. This is similar to the results from other molecular studies on Hawaiian plant taxa, including others with great morphological variability (e.g. silverswords, lobeliads and mints). Main conclusions On a per‐unit‐area basis, Hawaiian Bidens have among the highest rates of speciation for plant radiations documented to date. The rapid diversification within such a small area was probably facilitated by the habitat diversity of the Hawaiian Islands and the adaptive loss of dispersal potential. Our findings point to the need to consider the spatial context of diversification – specifically, the relative scale of habitable area, environmental heterogeneity and dispersal ability – to understand the rate and extent of adaptive radiation.     

Palytoxin disrupts cardiac excitation-contraction coupling through interactions with P-type ion pumps

Palytoxin is a coral toxin that seriously impairs heart function, but its effects on excitation-contraction (E-C) coupling have remained elusive. Therefore, we studied the effects of palytoxin on mechanisms involved in atrial E-C coupling. In field-stimulated cat atrial myocytes, palytoxin caused elevation of diastolic intracellular Ca²⁺ concentration ([Ca²⁺]_i), a decrease in [Ca²⁺]_i transient amplitude, Ca²⁺ alternans followed by [Ca²⁺]_i waves, and failures of Ca²⁺ release. The decrease in [Ca²⁺]_i transient amplitude occurred despite high sarcoplasmic reticulum (SR) Ca²⁺ load. In voltage-clamped myocytes, palytoxin induced a current with a linear current-voltage relationship (reversal potential 5 mV) that was blocked by ouabain. Whole cell Ca²⁺ current and ryanodine receptor Ca²⁺ release channel function remained unaffected by the toxin. However, palytoxin significantly reduced Ca²⁺ pumping of isolated SR vesicles. In current-clamped myocytes stimulated at 1 Hz, palytoxin induced a depolarization of the resting membrane potential that was accompanied by delayed afterdepolarizations. No major changes of action potential configuration were observed. The results demonstrate that palytoxin interferes with the function of the sarcolemmal Na⁺-K⁺ pump and the SR Ca²⁺ pump. The suggested mode of palytoxin toxicity in the atrium involves the conversion of Na⁺-K⁺ pumps into nonselective cation channels as a primary event followed by depolarization, Na⁺ accumulation, and Ca²⁺ overload, which, in turn, causes arrhythmogenic [Ca²⁺]_i waves and delayed afterdepolarizations. atrial myocytes; intracellular calcium     

Ostreopsis fattorussoi sp. nov. (Dinophyceae), a new benthic toxic Ostreopsis species from the eastern Mediterranean Sea

The new benthic toxic dinoflagellate, Ostreopsis fattorussoi sp. nov., is described from the Eastern Mediterranean Sea, Lebanon and Cyprus coasts, and is supported by morphological and molecular data. The plate formula, Po, 3′, 7″, 6c, 7s, 5?, 2′′′′, is typical for the Ostreopsis genus. It differs from all other Ostreopsis species in that (i) the curved suture between plates 1′ and 3′ makes them approximately hexagonal, (ii) the 1′ plate lies in the left half of the epitheca and is obliquely orientated leading to a characteristic shape of plate 6″. The round thecal pores are bigger than the other two Mediterranean species (O. cf. ovata and O. cf. siamensis). O. fattorussoi is among the smallest species of the genus (DV: 60.07 ± 5.63 μm, AP: 25.66 ± 2.97 μm, W: 39.81 ± 5.05 μm) along with O. ovata. Phylogenetic analyses based on the LSU and internal transcribed spacer rDNA shows that O. fattorussoi belongs to the Atlantic/Mediterranean Ostreopsis spp. clade separated from the other Ostreopsis species. Ostreopsis fattorussoi produces OVTX‐a and structural isomers OVTX‐d and ‐e, O. cf. ovata is the only other species of this genus known to produce these toxins. The Lebanese O. fattorussoi did not produce the new palytoxin‐like compounds (ovatoxin‐i, ovatoxin‐j₁, ovatoxin‐j₂, and ovatoxin‐k) that were previously found in O. fattorussoi from Cyprus. The toxin content was in the range of 0.28–0.94 pg · cell^?1. On the Lebanon coast, O. fattorussoi was recorded throughout the year 2015 (temperature range 18°C–31.5°C), with peaks in June and August.     

Redefining the dose of the entomopathogenic fungus Metarhizium brunneum (Ascomycota,Hypocreales) to increase Fe bioavailability and promote plant growth in calcareous and sandy soils

Background and aims

Plants differ in their ability to use different nitrogen (N) chemical forms, these differences can be related to their ecology and drive community structure. The capacity to uptake intact organic N has been observed in plants of several ecosystems. However, soil organic N uptake by Mediterranean plants is unknown despite organic N being abundant in Mediterranean ecosystems. We compare the uptake of different N forms in two widespread coexisting Mediterranean forest trees with contrasting ecophysiological characteristics: Quercus ilex and Pinus halepensis.

Methods

To estimate root uptake rate of each N form we used equimolar solutions (1 mM N) of ¹⁵NO₃ ^?, ¹⁵NH₄ ⁺ and ¹⁵N-¹³C glycine.

Results

NH₄ ⁺ and glycine were taken up at a similar rate, but faster than NO₃ ^? in both species. Intact dual labeled glycine was found in both species, demonstrating that both species can absorb intact organic N.

Conclusions

Despite their ecological differences, both species had similar preference for N forms suggesting no fundamental niche complementarity for N uptake. The higher preference for NH₄ ⁺ and glycine over NO₃ ^? possibly reflects adaptation to the differing proportions of N forms in Mediterranean soils.

     

Association and the Adh locus with a lethal factor (l(2) Stm) in Drosophila melanogaster

Keeping Drosophila cultures at 28 C results in elimination of all minor multiple ADH bands, thought to be due to conformational change. Thus in diploid and triploid adults heterozygous for the Adh ^F and Adh ^Salleles, relative staining intensities are found for the three bands which were in conformity with the assumption that both alleles are equally expressed. Among all polymorphic strains derived from natural Central European and Mediterranean populations, the strain ⁺Tüb is unique in that its Adh ^Fallele is closely linked to a new recessive lethal factor, named 1(2)Stm. All Adh ^F 1/Adh^F 1 pupae are unable to emerge, and die. The lethal effect is obvious 50 hr earlier by retarded eye, bristle, and body wall pigmentation. Although all pupae of the phenotype F die, Adh ^F allele frequency scarcely seems to be lowered in this natural population.     

Bioactive Constituents of Juniperus turbinata Guss. from La Maddalena Archipelago

A comprehensive phytochemical study of Juniperus turbinata (Cupressaceae) collected from La Maddalena Archipelago (Sardinia, Italy) is reported. Both the essential oil and the ethanolic extract obtained from the aerial parts were analyzed. The essential oil appears to belong to a new chemotype compared to other Mediterranean juniper accessions, as it was favored by geographic isolation of the isles. It showed a low content of monoterpene hydrocarbons and α‐terpineol, ent‐manoyl oxide, 1,10‐di‐epi‐cubenol as the major constituents. The ethanolic fraction contained mainly diterpenoids. Among these, 15‐formyloxyimbricatolic acid ( 7 ) is a new natural product since it has hitherto been obtained only by synthetic route. The phenolic fraction contained biflavonoids: cupressuflavone ( 9 ), followed by minor amounts of amentoflavone ( 10 ) and hinokiflavone ( 11 ). The essential oil and six purified compounds ( 1 – 4 , 8 and 9 ) were assessed for biological activities, namely antioxidant (assessed by DPPH^·, ABTS^·+ and FRAP methods) and cytotoxic effects towards selected human tumor cell lines (MDA‐MB 231, A375 and HCT116 cells). Compound 3 exhibited higher radical scavenging activity against ABTS^·+ radical than the reference Trolox. Noteworthy, compound 8 showed powerful effects towards tumor cell lines, with IC₅₀ values in the range of 0.060 – 0.201 μm , which make it a promising anticancer drug candidate.     

Testing Tropiduchini Stål 1866 (Hemiptera: Tropiduchidae) monophyly,a young inter‐tropical taxon of mainly insular species: taxonomy,distribution patterns and phylogeny,with the description of a new genus from Papua New Guinea

A morphological phylogenetic analysis of the tribe Tropiduchini (Hemiptera: Tropiduchidae) is provided for 21 genera among the 26 recognized, including the new genus Oechalinella Wang gen.n. , with a new species Oechalinella bifasciata Wang sp.n. Monophyly of the tribe is well supported by synapomorphies derived from the male genitalia. The strict consensus supports two subclades: ((Montrouzierana + Thymbra) + Thaumantia⁺) as sister to the clade (Leptovanua + (Vanua + Varma⁺) + Daradacella⁺). In the first subclade, the clade (Montrouzierana + Thymbra), distributed in New Caledonia and Papua New Guinea, is well supported by three synapomorphies based on the tegmina, while the Thaumantia⁺ group is characterized by the conformation of the anal tube and male gonostyli, and distributed from Malesia to Papuasia and in Central Africa. The second subclade is largely distributed from China to Malesia, Papuasia, Southwestern and the Northwestern Pacific islands. The position of the genus Leptovanua remains uncertain. The Varma⁺ clade is recovered in all analyses. Implied weighting analysis placed the Varma⁺ lineage as sister to all other genera forming a new group (Leptovanua⁺ + ((Montrouzierana + Thymbra) + Thaumantia⁺)). Fennah's definition of Tropiduchini is reviewed according to the phylogenetic results and an identification key to all genera is provided with a referenced synopsis of their distribution. Globally, Tropiduchini exhibits an inter‐tropical latitudinal distribution and a paradoxical bimodal longitudinal distribution pattern in continental Africa, South‐east Asia and the Pacific islands. Distribution data concern nearly 80% of insular taxa. The phylogenetical results suggest that: (i) the tribe originates from continental China; (ii) it evolved mainly from Papuasia by stepping‐stone colonization events, promoting subsequent insular speciation; and (iii) its evolution has mainly been directed by dispersion versus vicariance.     

Investigation of toxin profile of Mediterranean and Atlantic strains of Ostreopsis cf. siamensis (Dinophyceae) by liquid chromatography–high resolution mass spectrometry

Blooms of Ostreopsis spp. once confined to tropical and subtropical areas have recently spread to more temperate regions such as the Mediterranean and the Southern-Atlantic coasts of Europe. However, while O. confronta (cf.) ovata has caused several toxic outbreaks, the presence of O. cf. siamensis has been reported rather occasionally and in very few regions; as a consequence, O. cf. ovata toxin profile has been in-depth studied while poor information exists on toxicity of the Mediterranean and Atlantic O. cf. siamensis. In the present study toxin profile of Mediterranean and Atlantic O. cf. siamensis isolates also phylogenetically related has been studied through liquid chromatography–high resolution mass spectrometry (LC–HRMS) versus a palytoxin standard, a crude extract of O. cf. ovata containing all the ovatoxins so far known (ovatoxin-a to -f), and a Japanese O. siamensis extract which contained ostreocin-d and ostreocin-b. The Mediterranean and Atlantic O. cf. siamensis strains were shown not to produce either ostreocins, which are produced by the Japanese O. siamensis strain, or ovatoxins, which are produced by the Mediterranean O. cf. ovata. Only sub-fg levels of palytoxin on a per cell basis were detected in the Mediterranean strain. This study demonstrates that the Mediterranean and the Atlantic O. cf. siamensis strains are devoid of any appreciable toxicity. Thus, at least in the European area, O. cf. siamensis seems to present a much lower risk to human health than O. cf. ovata.     

The sensitive [SWI+] prion

Yeast prions are heritable protein-based genetic elements which rely on molecular chaperone proteins for stable transmission to cell progeny. Within the past few years, five new prions have been validated and 18 additional putative prions identified in Saccharomyces cerevisiae. The exploration of the physical and biological properties of these “nouveau prions” has begun to reveal the extent of prion diversity in yeast. We recently reported that one such prion, [SWI⁺], differs from the best studied, archetypal prion [PSI⁺] in several significant ways.¹ Notably, [SWI⁺] is highly sensitive to alterations in Hsp70 system chaperone activity and is lost upon growth at elevated temperatures. In that report we briefly noted a correlation amongst prions regarding amino acid composition, seed number and sensitivity to the activity of the Hsp70 chaperone system. Here we extend that analysis and put forth the idea that [SWI⁺] may be representative of a class of asparagine-rich yeast prions which also includes [URE3], [MOT3⁺] and [ISP⁺], distinct from the glutamine-rich prions such as [PSI⁺] and [RNQ⁺]. While much work remains, it is apparent that our understanding of the extent of the diversity of prion characteristics is in its infancy.     

A review of Polynesian Carposina Herrich‐Schäffer (Lepidoptera: Carposinidae), with descriptions of four new species

Hawaiian Carposina represent over 17% of the known world fauna of Carposinidae. In contrast, only two species are known for all of French Polynesia in the South Pacific. Here we describe four new species: two from the Hawaiian Islands, C arposina urbanae sp. nov. and C . gagneorum sp. nov. , and two from the Society Islands, C . longignathosa sp. nov. and C . brevinotata sp. nov. We further recognize another new Hawaiian species too worn to describe. Additionally, we present the first phylogeny for Polynesian Carposina, including 19 taxa, using one mitochondrial and two nuclear gene regions. The Hawaiian Carposina sampled thus far form a monophyletic clade. Lastly, we provide a framework to better understand the diversification and phylogeography of this group, and provide a summary of currently known host plant associations. Diversification appears to have resulted from interplay between host switching and geographic isolation across the Hawaiian Archipelago.     

SEQUENTIAL RADIATIONS AND PATTERNS OF SPECIATION IN THE HAWAIIAN CRICKET GENUS LAUPALA INFERRED FROM DNA SEQUENCES

The tremendous diversity of endemic Hawaiian crickets is thought to have originated primarily through intraisland radiations, in contrast to an interisland mode of diversification in the native Hawaiian Drosophila. The Hawaiian cricket genus Laupala (family Gryllidae) is one of several native genera of flightless crickets found in rain-forest habitat across the Hawaiian archipelago. I examined the phylogenetic relationships among mitochondrial DNA (mtDNA) sequences sampled from 17 species of Laupala, including the 12S ribosomal RNA (rRNA), transfer RNA (RNA)^val and 16S rRNA regions. The distribution of mtDNA variants suggests that species within Laupala are endemic to single islands. The phylogenetic estimate produced from both maximum likelihood and maximum parsimony supports the hypothesis that speciation in Laupala occurred mainly within islands. The inferred biogeographical history suggests that diversification in Laupala began on Kauai, the oldest rain-forested Hawaiian island. Subsequently, colonization to younger islands in the archipelago resulted in a radiation of considerable phylogenetic diversity. Phylogenetic patterns in mtDNA are not congruent with prior systematic or taxonomic hypotheses. Hypotheses that may explain the conflict between the phylogenetic patterns of mtDNA variation and the species taxonomy are discussed.     

Complex interactions among residues within pore region determine the K+ dependence of a KAT1‐type potassium channel AmKAT1

KAT1‐type channels mediate K⁺ influx into guard cells that enables stomatal opening. In this study, a KAT1‐type channel AmKAT1 was cloned from the xerophyte Ammopiptanthus mongolicus. In contrast to most KAT1‐type channels, its activation is strongly dependent on external K⁺ concentration, so it can be used as a model to explore the mechanism for the K⁺‐dependent gating of KAT1‐type channels. Domain swapping between AmKAT1 and KAT1 reveals that the S5–pore–S6 region controls the K⁺ dependence of AmKAT1, and residue substitutions show that multiple residues within the S5–Pore linker and Pore are involved in its K⁺‐dependent gating. Importantly, complex interactions occur among these residues, and it is these interactions that determine its K⁺ dependence. Finally, we analyzed the potential mechanism for the K⁺ dependence of AmKAT1, which could originate from the requirement of K⁺ occupancy in the selectivity filter to maintain its conductive conformation. These results provide new insights into the molecular basis of the K⁺‐dependent gating of KAT1‐type channels.     

Involvement of kil and kor genes in the phenotype of a host-range mutant of RP4

Summary Plasmid pRP761 is a derivative of the promiscuous plasmid RP4, which has a Tn76 insert 1.8 kb from its EcoRI site within the trfB region (Barth 1979). This mutation was pleiotropic, having three effects: the plasmid is unstably maintained in E. coli, it reduces the growth rate of its host and it has suffered a reduction in host-range. We show that pRP761 has reduced expression from both its korA and korB genes and that Tn76 has inserted between them. Fragment exchange experiments showed that this is the only mutant region in pRP761 and is therefore solely responsible for the pleiotropic effects. A spontaneous deletion derivative pRP761-6 has lost Tn76 and its adjacent kilA and korA genes: it has reacquired stability, does not inhibit host growth but is still reduced in its host-range. The provision of cloned korA ⁺ in trans complements the first two phenotypic effects in pRP761 to a large extent, but neither korA ⁺ alone nor korA ⁺ with korB ⁺ complements the host-range reduction in pRP761 or pRP761-6. A possible explanation for these results is that there is a site between korA and korB, affected by the Tn76 insert, that is essential to stable replication of these plasmids in some of their bacterial hosts.