On shortcomings of using mtDNA sequence divergence for the systematics of hares (genus Lepus): An example from cape hares

Despite the well-known fact that evolutionary patterns of single genes or sequences are not necessarily paralleled by organismic evolution, using mitochondrial sequence divergence for inferring phylogenetic relationships among hare species is not uncommon. Earlier studies reported interspecific introgression in the genus Lepus that may slur systematic conclusions drawn exclusively from mtDNA data. We examined pairwise divergence in partial mtHV-1 sequences and nuclear gene pools based on microsatellite and allozyme loci separately in a South and North African population of cape hares, Lepus capensis, and did not find significant correspondence on the individual level within either population. Also, the former population had a significantly higher level of mtHV-1 sequence divergence compared to the latter, but levels of individual nuclear gene pool divergence did not differ significantly between the two populations. Hence, the absence of correspondence between mtHV-1 sequence divergence and nuclear gene pool divergence among individuals within a population might be independent of the population-specific level of differentiation among individuals. Our results complement earlier findings in hares, and strongly recommend to include nuclear gene pool evidence for systematic inferences within this genus, even under the absence of mitochondrial introgression.     

Phylogenetic investigations of Sordariaceae based on multiple gene sequences and morphology

The family Sordariaceae incorporates a number of fungi that are excellent model organisms for various biological, biochemical, ecological, genetic and evolutionary studies. To determine the evolutionary relationships within this group and their respective phylogenetic placements, multiple-gene sequences (partial nuclear 28S ribosomal DNA, nuclear ITS ribosomal DNA and partial nuclear β-tubulin) were analysed using maximum parsimony and Bayesian analyses. Analyses of different gene datasets were performed individually and then combined to generate phylogenies. We report that Sordariaceae, with the exclusion Apodus and Diplogelasinospora, is a monophyletic group. Apodus and Diplogelasinospora are related to Lasiosphaeriaceae. Multiple gene analyses suggest that the spore sheath is not a phylogenetically significant character to segregate Asordaria from Sordaria. Smooth-spored Sordaria species (including so-called Asordaria species) constitute a natural group. Asordaria is therefore congeneric with Sordaria. Anixiella species nested among Gelasinospora species, providing further evidence that non-ostiolate ascomata have evolved from ostiolate ascomata on several independent occasions. This study agrees with previous studies that show heterothallic Neurospora species to be monophyletic, but that homothallic ones may have a multiple origins. Although Gelasinospora and Neurospora are closely related and not resolved as monophyletic groups, there is insufficient evidence to place currently accepted Gelasinospora and Neurospora species into the same genus.     

Open field host selection and behavior by tamarisk beetles (Diorhabda spp.) (Coleoptera: Chrysomelidae) in biological control of exotic saltcedars (Tamarix spp.) and risks to non-target athel (T. aphylla) and native Frankenia spp.

Biological control of invasive saltcedars (Tamarix spp.) in the western U.S. by exotic tamarisk leaf beetles, Diorhabda spp., first released in 2001 after 15 years of development, has been successful. In Texas, beetles from Crete, Greece were first released in 2004 and are providing control. However, adults alight, feed and oviposit on athel (Tamarix aphylla), an evergreen tree used for shade and as a windbreak in the southwestern U.S. and México, and occasionally feed on native Frankenia spp. plants. The ability of tamarisk beetles to establish on these potential field hosts was investigated in the field. In no-choice tests in bagged branches, beetle species from Crete and Sfax, Tunisia produced 30–45% as many egg masses and 40–60% as many larvae on athel as on saltcedar. In uncaged choice tests in south Texas, adult, egg mass and larval densities were 10-fold higher on saltcedar than on adjacent athel trees after 2 weeks, and damage by the beetles was 2- to 10-fold greater on saltcedar. At a site near Big Spring, in west-central Texas, adults, egg masses and 1st and 2nd instar larvae were 2- to 8-fold more abundant on saltcedar than on athel planted within a mature saltcedar stand being defoliated by Crete beetles, and beetles were 200-fold or less abundant or not found at all on Frankenia. At a site near Lovelock, Nevada, damage by beetles of a species collected from Fukang, China was 12–78% higher on saltcedar than on athel planted among mature saltcedar trees undergoing defoliation. The results demonstrate that 50–90% reduced oviposition on athel and beetle dispersal patterns within resident saltcedar limit the ability of Diorhabda spp. to establish populations and have impact on athel in the field.     

Phylogenetic relationships of Nemania plumbea sp. nov. and related taxa based on ribosomal ITS and RPB2 sequences

During a survey fungal diversity of xylariaceous fungi in Thailand, a new Nemania species, N. plumbea, was identified. Nemania plumbea is characterized by soft-textured grey stromata on a persistent mat of white hyphae, pale brown ascospores with a short germ-slit on the more convex side. It also produces a Geniculosporium-like anamorph in culture. In order to evaluate its phylogenetic relationships among related species and genera, ITS-5.8S rDNA and RPB2 were analysed separately and simultaneously. Results from the phylogenetic analyses indicate that there is close phylogenetic association between N. plumbea and N. aenea. A preliminary account into the natural grouping of Xylariaceae based on ITS-5.8S rDNA and RPB2 sequences is also discussed.     

Molecular evidence of conspecificity of South African hares conventionally considered Lepus capensis L., 1758

Conventionally, Lepus capensis is considered to range across large parts of Africa, the Middle East, Central and Far East Asia. However, a recent morphological study restricts cape hares tentatively to a small range in the Western Cape Region of South Africa and groups all other L. capensis-type hares from South Africa into a new species: L. centralis. Here, we studied molecular relationships among L. capensis-type hares from South Africa. Phenotypically and morphologically the individuals matched either the newly described L. capensis or L. centralis. We examined 66 hares for allelic variation at 13 microsatellite loci and for sequence variation of the hypervariable domain 1 of the mitochondrial control region. All tree-generating analyses of the currently obtained sequences and all South African cape hare sequences downloaded from GenBank revealed monophyly when compared to sequences of various other Lepus species. A network analysis indicated close evolutionary relationships between hares of the “L. capensis-phenotype” and the “L. centralis-phenotype” (according to Palacios et al. 2008) from the southwest of the Western Cape, relative to their pronounced evolutionary divergence from all other more central, northern, and north-eastern L. capensis-type hares. F-statistics, a Bayesian admixture STRUCTURE model, as well as a principal coordinate analysis of microsatellite data indicated close genetic relationships among all South African L. capensis-type hares studied presently. A coalescence model-based migration analysis for microsatellite alleles indicated gene flow between most of the considered subspecies of cape hare, including L. capensis capensis and L. capensis centralis, theoretically sufficient to balance stochastic drift effects. Concordantly, AMOVA models revealed only little effects of partitioning microsatellite variation into the two suggested morpho-species “L. capensis” and “L. centralis”. Under an “Interbreeding Species Concept” (e.g. a strict or relaxed Biological Species Concept), the current molecular data demonstrate conspecificity of the two proposed morpho-species “L. capensis” and “L. centralis”. Based on the present molecular data the differentiation of subspecies of cape hares from southern Africa is discussed.     

Biology of Lilioceris spp. (Coleoptera: Chrysomelidae) and their parasitoids in Europe

The biology and parasitoid complex of the lily leaf beetle (LLB), Lilioceris lilii Scopoli, and two congeneric species were investigated in Europe, as part of a biological control program against the LLB in North America. Eggs, larvae, and adults of L. lilii were collected in several countries in Europe, on both cultivated and wild Lilium spp., and reared in the laboratory and under natural conditions. Parasitoids were obtained and their biologies were studied. Similar investigations were made in Switzerland on two closely related species Lilioceris tibialis (Villa) found on wild Lilium spp., and Lilioceris merdigera (L.) on several other Liliaceae. The three species are strictly univoltine. Adults overwinter and lay eggs on leaves in early spring. The three beetle species have four instars, which were characterized by their head capsule width. Pupation occurs in a cocoon in the soil. Adults emerge in late summer and start feeding before reaching overwintering sites. Egg and larval parasitoids were obtained. Eggs of L. lilii and L. merdigera were parasitized by the mymarid Anaphes sp., a multivoltine species that needs alternate hosts for overwintering. Larvae were heavily attacked by several parasitoids, among which the most abundant were three ichneumonids, Lemophagus pulcher (Szepligeti), L. errabundus (Gravenhorst), and Diaparsis jucunda (Holmgren), and the eulophid Tetrastichus setifer Thomson. All four parasitoid species were found in the three beetles and in most European regions, but strong variations were observed in their relative abundance among hosts and geographic regions. Three of the four main larval parasitoids are strictly univoltine, whereas L. pulcher has a partial second generation. Lemophagus spp. are frequently parasitized by the ichneumonid hyperparasitoid Mesochorus lilioceriphilus Schwenke. Further details of the biology of the parasitoids are described, and their potential as biological control agents is discussed.     

Several pseudomonads, associated with the cultivated mushrooms Agaricus bisporus or Pleurotus sp., are hemolytic

Pseudomonas tolaasii, causing brown blotch disease on cultivated mushrooms, and yielding a white line precipitate towards P. “reactans”, has been shown to induce lysis of erythrocytes. Some Finnish strains isolated from diseased mushroom fruit bodies, although harboring the typical features of P. tolaasii, proved to be distinct, and have been allocated to a nov. sp. P. costantinii. We examined in these study whether all brown blotch causing agents were hemolytic. The induction of erythrocytes lysis seemed to be a rather common feature of mushroom associated-pseudomonads, especially for strains involved in the production of a white-line-in agar.     

Hyperparasitism by Mesochorus spp. (Hymenoptera: Ichneumonidae) in Peristenus sp. (Hymenoptera: Braconidae) and development of PCR primers for hyperparasitoid detection

Peristenus sp. pupae collected from Lygus spp. nymphs in 2001 and 2002 were over-wintered in the laboratory. In both years, more than 30% of adults emerging from over-wintering pupae were identified as ichneumonid hyperparasitoids, Mesochorus curvulus Thomson and Meschorus sp. (Hymenoptera: Ichneumonidae). At the end of the over-wintering period, Peristenus sp. males emerged first followed by Peristenus sp. females and finally Mesochorus spp. The male:female ratio in emerging Peristenus sp. adults was skewed towards males. The Internal Transcribed Spacer (ITS) region and the cytochrome oxidase I (COI) gene from Mesochorus spp. were sequenced. ITS sequences were used to develop PCR primers to detect Mesochorus spp. hyperparasitism in the primary host, Lygus spp. PCR analysis of field-collected Lygus spp. nymphs gave similar estimates of Mesochorus spp. hyperparasitism to the rearing protocols (25–28%). Sequence analysis of COI and ITS regions and subsequent restriction endonuclease analysis of ITS PCR products from Mesochorus spp. indicate the presence of two genotypes in the population. The possibility that these two genotypes represent separate or cyrptic species is discussed.     

Comparative toxicity of Pfiesteria spp., prolonging toxicity of P. piscicida in culture and evaluation of toxin(s) stability

Toxicity of Pfiesteria piscicida (strain CAAE #2200) in the presence of fish (juvenile hybrid tilapia, Oreochromis sp., total length 3–6 cm) has been maintained in the laboratory for 19 months by serial transfer of toxic cells using a modified maintenance protocol. Toxicity was re-induced when toxin-producing P. piscicida cells were separated from fish and cultured on algal prey for 50 days and then re-introduced to new tanks containing fish. We confirmed toxicity in a strain of P. shumwayae (strain CAAE #101272). Toxicity to fish was demonstrated in culture filtrates (0.2 μm) derived from cultures of both Pfiesteria spp., however, it was markedly reduced in comparison to unfiltered water. Filtrates retained toxic activity when stored at −20 °C for up to 6 months. Toxicity to fish was retained when filtrates were held at room temperature for 48 h, at 70 °C for 30 min or at 88–92 °C for 2 h. P. piscicida killed all finfish species tested. Grass shrimp (Paleomonetes pugio; adult 2–3 cm), blue crab (Callinectes sapidus; juvenile 4–7 cm) and brine shrimp (Artemia sp.; 18–24 h post-hatch) were unaffected by concentrations of toxin(s) that killed juvenile tilapia in 4–24 h. Ichthyotoxic activity of filtrates from fish-killing cultures and stability of the toxic activity were similar among P. piscicida and P. shumwayae. These results confirm previously reported observations on toxicity of P. piscicidaand P. shumwayae to finfish. We have maintained toxicity in the laboratory for longer periods than have previously been routinely achieved, and we have demonstrated that the toxic activity is heat stable. In contrast to previous studies with other toxic P. piscicida strains, we did not observe toxic activity to blue crabs or other crustaceans.     

Stictis s. lat. (Ostropales, Ascomycota) in northern Scandinavia, with a key and notes on morphological variation in relation to lifestyle

The closely related genera Carestiella, Schizoxylon and Stictis (Stictidaceae, Ostropales, Ascomycota) are revised in northern Scandinavia, and a revised phylogenetic hypothesis, including all seven species of this group in the area, is presented. A key to the species of Stictis s. lat. (including Carestiella, Schizoxylon and Stictis) in Northern Scandinavia is presented. S. albescens and S. confusum are described as new to science, and the new name S. brunnescens is introduced for S. mollis ssp. populorum, when treated as a distinct species. C. socia and S. albescens are the only representatives of their genera in northern Scandinavia, but a total of five species of Stictis are present in the area (S. brunnescens, S. confusum, S. mollis, S. populorum and S. radiata). Substantial within-species variation in ascoma morphology and wall pigmentation is revealed and discussed, and found to correlate with fungal lifestyles; optional lichenization apparently affects several details in fungal anatomy and morphology.     

Ecology and impact of Allorhogas sp. (Hymenoptera: Braconidae) and Apion sp. (Coleoptera: Curculionoidea) on fruits of Miconia calvescens DC (Melastomataceae) in Brazil

Two fruit-feeding insects, a gall wasp, Allorhogas sp. (Hymenoptera: Braconidae), and a beetle, Apion sp. (Coleoptera: Curculionoidea), were evaluated in their native habitat in Brazil as potential biological control agents of Miconia calvescens DC (Melastomataceae). Allorhogas sp. occurred at two out of three field sites with native populations of M. calvescens, and Apion sp. occurred at all three sites. Both species exhibited aggregated distributions among M. calvescens trees sampled at each site. Allorhogas sp. infested 9.0% and 3.8% of fruits at each of two sites. The number of larvae and pupae of Allorhogas sp. and/or an unidentified parasitoid (Hymenopetera: Eulophidae: Tetrastichinae) ranged from one to five per infested fruit. Fruits infested with Allorhogas sp. were 20% larger and had 79% fewer seeds than healthy fruits. Although adults of Apion sp. were found on leaves and inflorescences of M. calvescens at all three sites, larvae and pupae were found in fruits at only one site, where a maximum of 1.4% of fruits were infested. Fruits infested by Apion sp. contained only one larva or pupa, and were 15% smaller and had 62% fewer seeds than healthy fruits. While a variety of apionids have been used for biological control in the past, this is the first time a braconid wasp has been considered for biological control of a weed.     

Cylindrocarpon pauciseptatum sp. nov., with notes on Cylindrocarpon species with wide, predominantly 3-septate macroconidia

A Cylindrocarpon species with up to 10 μm wide, straight and predominantly 3-septate macroconidia, subglobose to ovoidal microconidia and chlamydospores, is described as Cyl. pauciseptatum. It is most similar to Cyl. austrodestructans but no chlamydospores and microconidia are formed in the latter. Similar macroconidia also occur in Cyl. theobromicola, which forms oval to ellipsoidal microconidia at least sparsely and has slightly curved macroconidia, and Cyl. destructans var. crassum, which forms abundant 1-celled microconidia. DNA sequence data of the internal transcribed spacer regions 1 and 2 plus the 5.8S rDNA and the partial beta-tubulin gene were used for phylogenetic inferences. Cylindrocarpon pauciseptatum and Cyl. macrodidymum are monophyletic and are closely related to other species of Cylindrocarpon sensu stricto including members of the Cyl. destructans (teleomorph, Neonectria radicicola) species complex, which accommodates Cyl. liriodendri (teleomorph, Neon. liriodendri), Cyl. destructans var. crassum and Cyl. austrodestructans (teleomorph, Neonectria austroradicicola comb. nov.). Cylindrocarpon theobromicola is distantly related to species of Cylindrocarpon sensu stricto or Neonectria sensu stricto. It clustered among cylindrocarpon-like species with curved macroconidia, of which some belong to the Neon. mammoidea group. Relatively voluminous cells in sporodochial conidiophores of Cyl. theobromicola resembled those described for Campylocarpon, which is closely related to members of the Neon. mammoidea group including Cyl. theobromicola. Cylindrocarpon pauciseptatum has been isolated from roots of Vitis spp. in South-eastern Europe (Slovenia) as well as New Zealand, where it also occurs on roots of Erica melanthera.     

The potential of Ulocladium botrytis for biological control of Orobanche spp.

A strain of Ulocladium botrytis isolated from diseased Orobanche crenata shoots caused disease on the parasitic weed in pathogenicity tests. The potential of the fungus to be developed as a mycoherbicide for Orobanche spp. was further investigated. Although the fungus significantly decreased O. crenata germination in vitro by 80%, it did not generally lead to a decreased number of O. crenata shoots or tubercles in inoculated root chambers or pots. However, the number of diseased or dead tubercles and underground shoots was significantly increased compared to the noninoculated treatments. Postemergence inoculation of O. crenata shoots with a conidial suspension resulted in the death of almost all inoculated plants 14 days after application under greenhouse conditions. In preliminary host-range studies, the pathogen caused disease on Orobanche cumana on sunflower whereas on Orobanche aegyptiaca shoots parasitizing tomato only minimal disease symptoms could be detected after postemergence inoculation. Based on the results of our investigations, we conclude that Ulocladium botrytis has only a limited potential to be used as a biocontrol agent against Orobanche spp.     

Biological control of Lygus lineolaris by Peristenus spp. in strawberry

Peristenus digoneutis Loan (Hymenoptera: Braconidae) was introduced to the US for biological control of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois) (Hemiptera: Miridae), and has since spread through much of the northeast. The purpose of this study was to determine if P. digoneutis and a native congener, Peristenus pallipes (Curtis), parasitize L. lineolaris in strawberry (where it is a key pest), and what factors relate to parasitism levels. During 1997–1999 we monitored parasitism on 17 strawberry farms in 14 counties in eastern and western New York State. We found that in eastern NY (where P. digoneutis has been established since the early 1990s), overall mean parasitism was 19.7% (ranging from 0 to 70%), mostly by P. digoneutis. Mean parasitism was significantly lower (12.3%, ranging from 0 to 58%) in western NY (where P. digoneutis was first recorded in 1999), and was mostly by P. pallipes. P. pallipes parasitism was significantly lower in eastern than western NY, suggesting the potential for competitive interaction with P. digoneutis. The insecticide regime of a farm was an important factor influencing parasitism rate, which was 5- to 6.5-fold higher on organic or casually sprayed farms than on intensely treated farms, though pest density under these three regimes was not significantly different. L. lineolaris density, and parasitism rate in nearby alfalfa and abandoned fields were also significant factors for parasitism in strawberry.     

Effects of different Beauveria bassiana isolates on field populations of Lygus lineolaris in pigweed (Amaranthus spp.)

The tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), is a pest of various fruit, vegetable, fiber, and seed crops; including cotton. Lygus spp. populations often build on alternate host plants before moving to cotton, and in the midsouthern U.S. wild host plants, such as pigweed (Amaranthus spp.), play a major role in L. lineolaris population development. Three isolates of the entomopathogenic fungus Beauveria bassiana (Balsamo) were evaluated for L. lineolaris control in redroot pigweed (Amaranthus retroflexus L.): one from L. lineolaris in Mississippi (TPB3); one from Lygus hesperus (Knight) in California (WTPB2); and one commercial isolate from Mycotrol^® (GHA). Fungal applications resulted in moderate to high mycosis in adults (33 to 80%) and moderate mycosis in nymphs (36 to 53%) that were collected from field plots at 2 days post-treatment and incubated under laboratory conditions. Although TPB3 was previously found to be more pathogenic in laboratory bioassays, there was not a consistent separation of this isolate from the other two isolates in field trials. Where differences in adult mycosis or mortality were observed, TPB3 was the most pathogenic. However, in one field trial 7 day mortality for nymphs treated with GHA was higher than those treated with TPB3 or WTPB2. Infection rates at 2, 7, and 14 days post-treatment from caged and non-caged adults suggested that movement of adults among plots occurred, which could have masked some treatment effects. Fungal treatments did not significantly reduce populations relative to controls. This may have been caused by delayed mortality rates under field conditions and/or difficulties with estimating population change under field conditions characteristic of wild host plant populations (e.g., heterogeneous populations, adult movement, and small plot size). Further work evaluating time–dose–mortality over dynamic temperatures, spring and fall field trials on this and other wild hosts, and improved methods for estimating populations on wild hosts are needed.     

A comparative study on the antimutagenic properties of aqueous extracts of Aspalathus linearis (rooibos), different Cyclopia spp. (honeybush) and Camellia sinensis teas

Antimutagenic activity of aqueous extracts of the South African herbal teas, Aspalathus linearis (rooibos) and Cyclopia spp. (honeybush) was compared with that of Camellia sinensis (black, oolong and green) teas in the Salmonella mutagenicity assay using aflatoxin B₁ (AFB₁) and 2-acetylaminofluorene (2-AAF) as mutagens. The present study presents the first investigation on antimutagenic properties of C. subternata, C. genistoides and C. sessiliflora. The herbal teas demonstrated protection against both mutagens in the presence of metabolic activation, with the exception of “unfermented” (green/unoxidised) C. genistoides against 2-AAF, which either protected or enhanced mutagenesis depending on the concentration. Antimutagenic activity of “fermented” (oxidised) rooibos was significantly (P < 0.05) less than that of Camellia sinensis teas against AFB₁, while for 2-AAF it was less (P < 0.05) than that of black tea and similar (P > 0.05) to that of oolong and green teas. Antimutagenic activity of unfermented C. intermedia and C. subternata exhibited a similar protection as fermented rooibos against AFB₁. Against 2-AAF, fermented rooibos exhibited similar protective properties than unfermented C. intermedia and C. sessiliflora. Unfermented rooibos was less effective than the C. sinensis teas and fermented rooibos, but had similar (P > 0.05) antimutagenicity to that of fermented C. sessiliflora against AFB₁ and fermented C. subternata against 2-AAF. Fermented C. intermedia and C. genistoides exhibited the lowest protective effect against 2-AAF, while fermented C. intermedia exhibited the lowest protection when utilising AFB₁ as mutagen. Aspalathin and mangiferin, major polyphenols in rooibos and Cyclopia spp., respectively, exhibited weak to moderate protective effects when compared to the major green tea catechin, (−)epigallocatechin gallate (EGCG). Antimutagenic activity of selected herbal tea phenolic compounds indicated that they contribute towards (i) observed antimutagenic activity of the aqueous extracts against both mutagens and (ii) enhancement of the mutagenicity of 2-AAF by unfermented C. genistoides. Antimutagenic activity of the South African herbal teas was mutagen-specific, affected by fermentation and plant material, presumably due to changes and variation in phenolic composition.     

基于叶绿体DNA片段的槭属鸡爪槭组系统进化和生物地理学研究

东亚地区经历过复杂的地质事件及气候变化,针对东亚地区进行专科专属的植物区系及地理演化研究具有重要的意义。基于3个叶绿体片段（psbA-trnH、rpl16和trnL-trnF）,对主要分布于东亚地区的鸡爪槭组（section Palmata Pax）18个分类群的分化时间、祖先地理分布、物种扩散路径等进行了分析。叶绿体片段总长度为2,232bp,其中变异位点341个（15.2%）,有效信息位点231个（10.3%）;18个鸡爪槭组内物种（包含亚种）被高度支持为单系群（99/95/1.0）,且组内物种间的支持度相对较高;鸡爪槭组内出现物种分化的时间多集中于中新世时期,组内物种的多样性速率保持相对稳定;根据BioGeoBears分析得出,东亚大陆局部地区是该组物种的祖先分布区,且现今分布于岛屿的物种分布在不同分支。通过讨论与分析得出：我国西南地区可能是鸡爪槭组植物的起源地;东海陆桥隐没与抬升,直接影响着鸡爪槭组物种的迁移与扩散;分布于亚热带亚洲大陆的鸡爪槭组物种,存在多次往同纬度高海拔地区扩散的现象。该研究有助于对东亚地区植物生物地理历史的进一步了解。