Delineating species for conservation using mitochondrial sequence data: the taxonomic status of two problematic <Emphasis Type="Italic">Bombus</Emphasis> species (Hymenoptera: Apidae)

Across Western Europe and North America, many bumblebee species are currently undergoing drastic declines in their abundance and ranges, primarily as a result of habitat fragmentation. In contrast, a smaller number of species are seemingly unaffected by this and remain common. The UK Biodiversity Action Plan-designated Bombus ruderatus belongs to the former group while B. hortorum belongs to the latter. These two species are sympatric and remarkably similar in morphology. There are no diagnostic characters for workers and male genitalia are illustrated with the same diagram in standard keys. Isolated records of putative B. ruderatus occur amongst a mass of records for B. hortorum. This raises two important issues: first, are B. ruderatus and B. hortorum ‘good’ species? Second, if they are, can the uncertainty over their identification be resolved? We present COII and cytochrome b mtDNA sequence data from these and other Bombus species. Molecular data and coat colour characters are in concordance and confirm that B. ruderatus and B. hortorum should be regarded as separate species (although coat colour alone is an unreliable diagnostic character for many individuals). Confirmation of the specific status of B. ruderatus allows the work on the conservation of this species to continue.     

Flooding events and rising water temperatures increase the significance of the reed pathogen <Emphasis Type="Italic">Pythium phragmitis</Emphasis> as a contributing factor in the decline of <Emphasis Type="Italic">Phragmites australis</Emphasis>

Pythium species are economically significant soilborne plant pathogens with worldwide distribution, causing seedling damping-off or root rot diseases. Pythium phragmitis is a newly described pathogen of common reed (Phragmites australis), widespread in the reed-belt of Lake Constance, Germany. It is highly aggressive towards reed leaves and seedlings, but obviously does not affect roots. In the context of ‘reed decline’ phenomena, P. phragmitis infection of reed inundated during flooding events may be of particular significance. We could show that flooding itself is not necessarily detrimental for reed plants. In the presence of the pathogen, however, most submerged leaves and plants were killed within several weeks. Clipped plants did not show regrowth in the Pythium infested treatments. Significant losses in assimilating leaf area of reeds could, thus, be the result of Pythium infection rather than of flooding alone. Therefore, we suggest that the combination of extended flooding and the presence of P. phragmitis might considerably contribute to ‘reed decline’ at Lake Constance. In parallel, we could show that pathogenicity and spread of this species are considerably favoured by rising temperatures. Since an increase in average water temperature has been found for Lake Constance, we propose that P. phragmitis could be an important factor in the dieback of reed stands likely to be promoted by predicted climate change phenomena.     

Taxonomy and pathogenicity of <Emphasis Type="Italic">Leptographium</Emphasis> species associated with <Emphasis Type="Italic">Ips subelongatus</Emphasis> infestations of <Emphasis Type="Italic">Larix</Emphasis> spp. in northern China,including two new species

The larch bark beetle (Ips subelongatus), which occurs in larch plantations over a vast area of eastern Asia, infects both dying and fallen trees. When its population reaches a high density, the beetle may also infect healthy trees, resulting in tree decline and, eventually, death. Leptographium spp., in both their sexual and asexual states, are mainly associated with conifer-infesting bark beetles; some species are important tree pathogens. The aims of this study were to identify the Leptographium spp. associated with I. subelongatus infestations of Larix spp. in northern China and to examine their pathogenicity towards the tree. Morphological studies and phylogenetic approaches based on multilocus DNA sequence data (ITS2- partial r28S, partial β-tubulin, and EF-1α gene regions) showed that three Leptographium species occur in association with I. subelongatus in the areas investigated: Leptographium taigense, which is recorded in China for the first time, and two new species, namely L. innermongolicum sp. nov. and L. zhangii sp. nov. Leptographium innermongolicum is closely related to L. taigense, whereas L. zhangii belongs to the Grosmannia piceaperda species complex. The pathogenicity of these Leptographium species towards mature Larix spp. was tested by stem inoculation in forests. All inoculations only resulted in small lesions on the inner bark; therefore, the three Leptographium species were not considered to be pathogenic.     

Contrasting Brood-Sex Ratio Flexibility in Two Opiine (Hymenoptera: Braconidae) Parasitoids of Tephritid (Diptera) Fruit Flies

Mass-rearing of fruit fly parasitoids for augmentative release would be more economical if production could be biased towards females. If sex ratios are ever to be manipulated under rearing conditions it is important to determine if, then understand why, sex ratio flexibility exists. Unequal brood-sex ratios are common in parasitic Hymenoptera and two reasons are generally advanced for such cases: 1) avoidance of Local Mate Competition (LMC) predicts that the number of females exploiting a host-patch can influence the optimal sex ratio of their offspring; and 2) one sex is more likely to develop under a particular set of physical/competitive conditions. We hypothesized that LMC is more often encountered in relatively uncommon species with coarse-grained distributions. As a result, isolated females of such species would be more likely to expect future LMC to be high and to bias offspring sex ratios towards females. We proposed that the opiine braconid Utetes anastrephae is such a coarse-grained species and compared its responses to differences to LMC (isolated females and those paired with a conspecific competitor) with those of another opiine, Diachasmimorpha longicaudata believed to have a finer-grained distribution. Adult sex ratios were mutable in D. longicaudata and U. anastrephae and, as predicted by avoidance of LMC, males were relatively more abundant among the paired females in both species. However, superparasitized hosts yielded relatively more daughters, perhaps because female larvae are superior intrinsic competitors. Contrary to prediction, there was no suggestion that U. anastrephae was more likely to than D. longicaudata to avoid LMC. While these results did not reveal any species differences in sex-allocation patterns or suggest any modifications to the present practice of fruit fly mass-rearing for augmentative release they did provide new information regarding U. anastrephae, a widespread natural enemy of Anastrepha spp. about which relatively little is known.     

New name for Central Europaean oak formerly labelled as <Emphasis Type="Italic">Quercus dalechampii</Emphasis>

Quercus dalechampii Ten. is an oak species recognised by numerous authors, but it has an inconsistent taxonomical use depending on any particular author’s concept. Typification of Q. dalechampii in 2012 made this name inapplicable for Central European populations traditionally treated as Q. dalechampii. A new name, Q. banatus, based on Quercus aurea Wierzb. ex Kotschy, is here designated for a taxon equivalent to the present use of “Q. dalechampii” in Czech Republic, Slovakia, Hungary and other countries (at least in part) belonging to the Central European region. Notes on selected oak names related to Q. aurea Wierzb. ex Kotschy are appended.     

Taxonomic and functional diversity in <Emphasis Type="Italic">Calogaya</Emphasis> (lichenised Ascomycota) in dry continental Asia

The genus Calogaya (Teloschistaceae, Xanthorioideae) was established to accommodate mainly epilithic lichens with lobate thalli, previously regarded as the “Caloplaca saxicola group.” Data supporting the recognition of this new genus came from European lichens, and although the genus is soundly based, we have found in Asia numerous epiphytic lineages and lineages with reduced, non-lobate thallus in dry continental areas. The taxonomic and functional diversity of Calogaya is distinctly higher in steppe and desert areas of Asia than in the less arid regions of Europe. We sampled 238 specimens, mostly from arid regions of north-western China, Iran, southern Siberia and Turkey. Three nuclear DNA loci were analysed separately and jointly by Bayesian inference, maximum likelihood and *BEAST approaches. Delimitations of 28 putative species were tested by BP&P multispecies coalescent model with joint analysis of species delimitation and species-tree estimation. Finally, we recognised 22 taxonomic units: 16 are at species rank, 3 are treated as subspecies and 3 are complexes, treated here as a single entity, but in reality probably including more than one species. Calogaya altynis, C. biatorina subsp. asiatica, C. decipiens subsp. esorediata, C. haloxylonis, C. orientalis, C. xanthoriella and C. xinjiangis are newly described. Caloplaca zoroasteriorum is combined into Calogaya, and Calogaya persica is reduced to a subspecies. The taxonomic status of Calogaya saxicola is unclear, and the name is employed here “sensu lato” for several non-monophyletic epilithic lineages with short-lobed thalli. Calogaya biatorina and C. ferrugineoides are the two other heterogeneous taxonomic units probably including more species.     

Environmental risk assessment of the egg parasitoid <Emphasis Type="Italic">Anaphes inexpectatus</Emphasis> for classical biological control of the <Emphasis Type="Italic">Eucalyptus</Emphasis> snout beetle, <Emphasis Type="Italic">Gonipterus platensis</Emphasis>

Classical biological control is a valuable tool against invasive pests, but concerns about non-target effects requires risk assessment studies. Potential non-target effects of Anaphes inexpectatus Huber and Prinsloo (Hymenoptera: Mymaridae) were assessed for a classical biological control programme against the Eucalyptus snout beetle, Gonipterus platensis (Marelli) (Coleoptera: Curculionidae). No-choice tests were conducted with 17 non-target species to assess host specificity, including 11 curculionids. In behavioural observations, A. inexpectatus showed no interest in any of the non-target species, but two weevil species were parasitised within five days of exposure, although at significantly lower rates than G. platensis. In choice tests, only one non-target, Hypera postica (Gyllenhal) (Coleoptera: Curculionidae), was parasitised, at a rate of 0.6%, while 50.0% of G. platensis eggs were parasitised. Based on the host specificity test results and the potential host fauna found in the target area, the likelihood of non-target effects resulting from the release of A. inexpectatus is considered to be negligible.     

Evidence for genetic erosion of a California native tree, <Emphasis Type="Italic">Platanus racemosa</Emphasis>, via recent,ongoing introgressive hybridization with an introduced ornamental species

When non-native, genetically diverse species are introduced, hybridization with native congeners may erode the genetic composition of local species, perhaps even resulting in extinction. While such events may lead to adverse consequences at the community and ecosystem level, few studies exist on ecologically important tree species. In the genus Platanus, introgressive hybridization is widespread, and one common ornamental species, introduced to California during the late 19th century, is itself a hybrid. Our microsatellite analysis of more than 400 Platanus trees from north-central California reveals a complex pattern of invasion and hybridization in an age-structured population. By using size as a proxy for age, we have demonstrated that the Platanus population of north-central California has recently gained genetic diversity and effective population size. Principal coordinate analysis (PCoA) and genetic admixture analysis (STRUCTURE) both reveal a strong differentiation of genotypes into two main genetic clusters, with a large number of admixed genotypes. One of the genetic clusters identified is heavily biased towards younger trees, including samples from locations with relatively recently planted ornamental trees likely to be P. × hispanica (formerly known as P. × acerifolia). We conclude that the two genetic clusters correspond to the native P. racemosa and the introduced invasive hybrid species P. × hispanica. Additional hybridization between the invasive ornamental and the native species has occurred in California, and recent hybrid trees are more likely to be younger than trees without admixture. Our findings suggest that the observed increase in genetic diversity among California Platanus is due to rampant ongoing introgression, which may be threatening the continued genetic distinctiveness of the native species. This is cause for concern from a conservation standpoint, due to a direct loss of genetic distinctiveness, and a potential reduction in habitat value of associated species.     

Two new and one known species of <Emphasis Type="Italic">Tergestia</Emphasis> Stossich, 1899 (Trematoda: Fellodistomidae) with novel molecular characterisation for the genus

Combined morphological and molecular analyses are employed to characterise three species of Tergestia Stossich, 1899 (Digenea: Fellodistomidae) from fishes of Moreton Bay, Queensland, Australia. Tergestia clonacantha Manter, 1963 is reported here for the first time from the halfbeak (Beloniformes: Hemiramphidae) species Arrhamphus sclerolepis krefftii (Steindachner), Hyporhamphus australis (Steindachner), H. quoyi (Valenciennes) and H. regularis ardelio (Whitley). Two new species, both infecting trevally (Perciformes: Carangidae) species, are described: T. maryae n. sp. from Alepes apercna Grant and T. henryi n. sp. from Pantolabus radiatus (MacLeay). Complete ITS2 and partial 28S ribosomal DNA data were generated for each of the new taxa. The three species differ from each other by 47–58 base pairs (bp) in the ITS2 rDNA region. Phylogenetic analysis of 28S rDNA supports Tergestia as a reliable generic concept, with our analyses showing that some species of the genus form a well-supported clade to the exclusion of all other fellodistomids for which sequence data are available.     

Revision of <Emphasis Type="Italic">Podocotyloides</Emphasis> Yamaguti, 1934 (Digenea: Opecoelidae), resurrection of <Emphasis Type="Italic">Pedunculacetabulum</Emphasis> Yamaguti, 1934 and the naming of a cryptic opecoelid species

Despite morphological and ecological inconsistencies among species, all plagioporine opecoelids with a pedunculate ventral sucker are currently considered to belong in the genus Podocotyloides Yamaguti, 1934. We revise the genus based on combined morphological and phylogenetic analyses of novel material collected from haemulid fishes in Queensland waters that we interpret to represent species congeneric with the type-species, Pod. petalophallus Yamaguti, 1934, also known from a haemulid, off Japan. Our phylogenetic analysis demonstrates polyphyly of Podocotyloides; prompts us to resurrect Pedunculacetabulum Yamaguti, 1934; and suggests that Pod. brevis Andres & Overstreet, 2013, from a deep-sea congrid in the Caribbean, and Pod. parupenei (Manter, 1963) Pritchard, 1966 and Pod. stenometra Pritchard, 1966, from mullids and chaetodontids, respectively, on the Great Barrier Reef, may each represent a distinct genus awaiting recognition. Our revised concept of Podocotyloides requires a pedunculate ventral sucker, but also a uterine sphincter prior to the genital atrium, a petalloid cirrus appendage, restriction of the vitelline follicles to the hindbody, and for the excretory vesicle to reach to the level of the ventral sucker. Of about 20 nominal species, we recognise just three in Podocotyloides (sensu stricto): Pod. petalophallus, Pod. gracilis (Yamaguti, 1952) Pritchard, 1966 and Pod. magnatestes Aleshkina & Gaevskaya, 1985. We provide new records for Pod. gracilis, and propose two new species of Podocotyloides, Pod. australis n. sp. and Pod. brevivesiculatus n. sp., and one new Pedunculacetabulum species, Ped. inopinipugnus n. sp., all from haemulids. Podocotyloides australis is morphologically indistinguishable from Pod. gracilis, and exploits the same definitive host, but is genetically and biogeographically distinct. It is thus a cryptic species, the first such opecoelid to be formally named.     

Morphogenetic diversity of the ectomycorrhizal genus <Emphasis Type="Italic">Cortinarius</Emphasis> section <Emphasis Type="Italic">Calochroi</Emphasis> in the Iberian Peninsula

The taxonomic and genetic diversity of Cortinarius section Calochroi in one of the most biodiversity-rich regions in Europe, the Iberian Peninsula, was investigated through morphological and phylogenetic methods. This combined methodological approach allowed the identification of 15 known species and one new species, Cortinarius ortegae, which is described here. A dichotomous key is provided for field recognition of Calochroi species inhabiting different Iberian and Balearic Island woodlands. Polymorphism analyses within some studied species showed that the distribution of intraspecific lineages is dependent on geography and, in some cases, tree host. Furthermore, the dating analysis suggested that diversification within Calochroi started in the Pliocene and most of the current genetic diversity originated in the Pleistocene. This temporal scenario supports hypotheses in which climatic oscillations in the Quaternary may have been driving the evolution of this group of ectomycorrhizal fungi.     

Phylogeny and morphology of the <Emphasis Type="Italic">Peziza ammophila</Emphasis> complex (Pezizales,Ascomycota), with description of two new species and a new form

This study was focused on species of Peziza belonging to the “P. ammophila” complex, using both morphological and molecular approaches. Molecular and morphological analyses showed that several taxa are hidden under the name “ammophila” and, as a consequence, two additional species and one form are established: P. hellenica sp. nov., P. oceanica sp. nov. and P. ammophila f. megaspora f. nov. On the basis of the data that we have, these taxa seem to be related to some geographical areas; P. ammophila appears to be prevalent in northern, central and southern Europe, its f. megaspora is distributed in France and in the Netherlands, P. hellenica comes from Greece and P. oceanica from New Zealand. A cryptic species, not yet circumscribed satisfactorily, is described as P. deceptiva ad int. Moreover, the problem concerning the identification of P. ammophila is raised, the species is epitypified with a sequenced collection from Sicily (Italy) and a key to the species of the complex is provided.     

Diversity within the <Emphasis Type="Italic">Hygrophorus agathosmus</Emphasis> group (Basidiomycota,Agaricales) in Northern Europe

The North European species of the Hygrophorus agathosmus group in subsection Tephroleuci were studied. Three new species are identified based on morphology, ecology and sequence data. Two species are associated with Pinus spp. One of these is described here as H. suaveolens, while the other one is only known from one locality in the Nordic countries and seems to have a more South European distribution range. A closely related sister species to H. agathosmus is identified based on ITS sequence data, H. cf. agathosmus. It is confirmed to have an intercontinental distribution range and to be associated with Picea spp. probably on more acidic to neutral soil, whereas H. agathosmus s.s. has a more limited North-East European distribution range and occurs in older and rich Picea abies forests. A neotype for H. agathosmus is here selected from South Sweden. Hygrophorus agathosmus f. albus and H. agathosmus f. aureofloccosus are confirmed as forms. No genetic differences in the ITS region between specimens with grey cap colour and the two forms were observed. A key to the species in Northern Europe is provided.     

Metabolomics as a tool for understanding the evolution of <Emphasis Type="Italic">Tabebuia sensu lato</Emphasis>

Background

Plant systematic studies have changed substantially in the last years, stimulated by new strategies for phylogenetic studies. In this regard, chemistry data has been a useful tool for understanding plant phylogenetic relationships.

Objective

Our aim was to apply metabolomic approaches, followed by multivariate statistical analysis and dereplication of Tabebuia sensu lato species, and compare our results with classifications based on traditional taxonomy and molecular phylogeny. We also evaluated the application of metabolomics as a chemotaxonomic identification tool, as well as to enlighten plant chemical evolution.

Methods

Metabolomic data was generated through a high-resolution mass spectrometry with electrospray ionization of 27 Tabebuia sensu lato specimens from different populations, consisting of 15 Handroanthus (from four species) and 12 Tabebuia sensu stricto (from three species). Chemometric tools, such as principal component analysis and metabolite heatmaps, were used to scrutinize the metabolic changes among species.

Results

Tabebuia and Handroanthus species presented different secondary metabolite storage capacity. The genus Tabebuia revealed higher levels of glycosylated iridoids esterified with a phenylpropanoid moiety, such as specioside, verminoside, and minecoside, while Handroanthus accumulated iridoids linked to a simple phenol, lignans, and verbascoside derivatives.

Conclusion

These results corroborate splitting the Tabebuia s.l., which was supported by profound changes in secondary metabolism, suggesting metabolomics as an excellent tool for understanding species evolution.

     

<Emphasis Type="Italic">Bruguiera hainesii</Emphasis>, a critically endangered mangrove species,is a hybrid between <Emphasis Type="Italic">B. cylindrica</Emphasis> and <Emphasis Type="Italic">B. gymnorhiza</Emphasis> (Rhizophoraceae)

Bruguiera hainesii (Rhizophoraceae) is one of the two Critically Endangered mangrove species listed in the IUCN Red List of Threatened Species. Although the species is vulnerable to extinction, its genetic diversity and the evolutionary relationships with other Bruguiera species are not well understood. Also, intermediate morphological characters imply that the species might be of hybrid origin. To clarify the genetic relationship between B. hainesii and other Bruguiera species, we conducted molecular analyses including all six Bruguiera species using DNA sequences of two nuclear genes (CesA and UNK) and three chloroplast regions (intergenic spacer regions of trnL-trnF, trnS-trnG and atpB-rbcL). For nuclear DNA markers, all nine B. hainesii samples from five populations were heterozygous at both loci, with one allele was shared with B. cylindrica, and the other with B. gymnorhiza. For chloroplast DNA markers, the two haplotypes found in B. hainesii were shared only by B. cylindrica. These results suggested that B. hainesii is a hybrid between B. cylindrica as the maternal parent and B. gymnorhiza as the paternal one. Furthermore, chloroplast DNA haplotypes found in B. hainesii suggest that hybridization has occurred independently in regions where the distribution ranges of the parental species meet. As the IUCN Red List of Threatened Species currently excludes hybrids (except for apomictic plant hybrids), the conservation status of B. hainesii should be reconsidered.     

Circumscription of species in the <Emphasis Type="Italic">Hodophilus foetens</Emphasis> complex (Clavariaceae,Agaricales) in Europe

Four European Hodophilus species with an odour similar to naphthalene, a strong unpleasant odour similar to that of mothballs, are recognized based on sequence and/or morphological data. The traditional concept defines Ho. foetens as the only Hodophilus species with a naphthalene odour in Europe. This name is now assigned to one of the studied species based on morphological examination of the holotype specimen. A recently collected specimen is proposed as the epitype. The other three species with a naphthalene odour are described here as new: Ho. pallidus, Ho. subfoetens and Ho. tenuicystidiatus. They are distinguishable in the field based on a combination of lamellae number and colour of basidiomata. All four species are grouped in the Ho. foetens superclade, one of two superclades, together with the Ho. micaceus superclade, in the genus Hodophilus. All are different species from North American taxa with a naphthalene-like odour recognised in a previous study. The Ho. foetens superclade also includes one species identified as Ho. atropunctus that does not have a distinctive odour. The type collection of Ho. albofloccipes, a recently described European species with a naphthalene odour, is placed together with some collections without a distinctive odour in the Ho. micaceus superclade.