New insights into the DNA extraction and PCR amplification of minute ascomycetes in the genus Laboulbenia (Pezizomycotina, Laboulbeniales).

Molecular studies of fungi within the order Laboulbeniales (Ascomycota, Pezizomycotina) have been hampered for years because of their minute size, inability to grow in axenic culture, and lack of reliable and cost-efficient DNA extraction protocols. In particular, the genus Laboulbenia is notorious for low success with DNA extraction and polymerase chain reaction (PCR) amplification. This is attributed to the presence of melanin, a molecule known to inhibit PCR, in the cells. We evaluated the efficacy of a standard single cell-based DNA extraction protocol by halving the recommended amount of reagents to reduce the cost per extraction and adding bovine serum albumin (BSA) during the multiple displacement amplification step to reverse the effect of melanin. A total of 196 extractions were made, 111 of which were successful. We found that halving the reagents used in the single cell-based extraction kit did not significantly affect the probability of successful DNA extraction. Using the halved protocol reduces cost and resource consumption. Moreover, there was no significant difference in the probability of successfully extracting DNA based on whether BSA was added or not, suggesting that the amount of melanin present in cells of the thallus has no major inhibitory effect on PCR. We generated 277 sequences from five loci, but amplification of the internal transcribed spacer region, the mitochondrial small subunit rDNA, and protein-coding genes remains challenging. The probability of successfully extracting DNA from Laboulbeniales was also impacted by specimen storage methods, with material preserved in > 95% ethanol yielding higher success rates compared to material stored in 70% ethanol and dried material. We emphasize the importance of proper preservation of material and propose the design of Laboulbeniales-specific primers to overcome the problems of primer mismatches and contaminants. Our new insights apply not only to the genus Laboulbenia; Laboulbeniales generally are understudied, and the vast majority of species remain unsequenced. New and approachable molecular developments will benefit the study of Laboulbeniales, helping to elucidate the true diversity and evolutionary relationships of these peculiar microfungi.

Infection patterns of Harmonia axyridis (Coleoptera: Coccinellidae) by ectoparasitic microfungi and endosymbiotic bacteria.

The invasive alien ladybird Harmonia axyridis (Coleoptera: Coccinellidae) hosts a wide range of natural enemies. Many observations have been done in nature but experimental studies of interactions of multiple enemies on Ha. axyridis are rare. In light of this knowledge gap, we tested whether the host phenotype and presence of bacterial endosymbionts Spiroplasma and Wolbachia affected parasitism of Ha. axyridis by the ectoparasitic fungus Hesperomyces harmoniae (Ascomycota: Laboulbeniales). We collected 379 Ha. axyridis in the Czech Republic, processed specimens, including screening for He. harmoniae and a molecular assessment for bacteria, and calculated fecundity and hatchability of females. We found that high hatchability rate (71 %) was conditioned by high fecundity (20 eggs daily or more). The average parasite prevalence of He. harmoniae was 53 %, while the infection rate of Spiroplasma was 73 % in ladybirds that survived in winter conditions. Wolbachia was only present in 2 % of the analyzed ladybirds. Infection by either He. harmoniae or Spiroplasma did not differ among host color morphs. In the novemdecimsignata morph, younger individuals (with orange elytra) were more heavily parasitized compared to old ones (with red elytra). Fecundity and hatchability rate of females were unaffected by infection with either He. harmoniae or Spiroplasma. However, female ladybirds co-infected with He. harmoniae and Spiroplasma had a significantly lower fecundity and hatchability compared to females with only one or no symbiont.

The haustorium as a driving force for speciation in thallus-forming Laboulbeniomycetes.

Laboulbeniomycetes is a class of fungi that have obligate associations with arthropod hosts, either for dispersal (order Pyxidiophorales) or as biotrophic parasites (orders Herpomycetales and Laboulbeniales). Here, we focus on Herpomycetales and Laboulbeniales, which include fungi that form thalli, 3-dimensional, multicellular units of 1000 s of cells. Based on recently published data regarding patterns of speciation, we present the One-Host-One-Parasite model (1H1P) for haustorial thallus-forming Laboulbeniomycetes. We hypothesize that taxa with haustoria, rhizoidal structures that make contact with the host's body cavity, have very strict host specificity. For taxa without haustoria, the microhabitat-as selected by the host-governs host shifting, presence or absence of the fungus, abundance, effective host range, and geographic distribution. We make suggestions for future research including fluorescent labeling of waxy lipids and mass spectrometry. These techniques have the potential to generate the data necessary to evaluate the here proposed 1H1P hypothesis for Herpomycetales and Laboulbeniales.

Laboulbeniomycetes: Intimate Fungal Associates of Arthropods.

Arthropod-fungus interactions involving the Laboulbeniomycetes have been pondered for several hundred years. Early studies of Laboulbeniomycetes faced several uncertainties. Were they parasitic worms, red algal relatives, or fungi? If they were fungi, to which group did they belong? What was the nature of their interactions with their arthropod hosts? The historical misperceptions resulted from the extraordinary morphological features of these oddly constructed ectoparasitic fungi. More recently, molecular phylogenetic studies, in combination with a better understanding of life histories, have clearly placed these fungi among filamentous Ascomycota (subphylum Pezizomycotina). Species discovery and research on the classification of the group continue today as arthropods, and especially insects, are routinely collected and examined for the presence of Laboulbeniomycetes. Newly armed with molecular methods, mycologists are poisedto use Laboulbeniomycetes-insect associations as models for the study of a variety of basic evolutionary and ecological questions involving host-parasite relationships, modes of nutrient intake, population biology, host specificity, biological control, and invasion biology. Collaboration between mycologists and entomologists is essential to successfully advance knowledge of Laboulbeniomycetes and their intimate association with their hosts.

On the Fly: Tritrophic Associations of Bats, Bat Flies, and Fungi.

Parasitism is one of the most diverse and abundant modes of life, and of great ecological and evolutionary importance. Notwithstanding, large groups of parasites remain relatively understudied. One particularly unique form of parasitism is hyperparasitism, where a parasite is parasitized itself. Bats (Chiroptera) may be parasitized by bat flies (Diptera: Hippoboscoidea), obligate blood-sucking parasites, which in turn may be parasitized by hyperparasitic fungi, Laboulbeniales (Ascomycota: Laboulbeniomycetes). In this study, we present the global tritrophic associations among species within these groups and analyze their host specificity patterns. Bats, bat flies, and Laboulbeniales fungi are shown to form complex networks, and sixteen new associations are revealed. Bat flies are highly host-specific compared to Laboulbeniales. We discuss possible future avenues of study with regard to the dispersal of the fungi, abiotic factors influencing the parasite prevalence, and ecomorphology of the bat fly parasites.

The paradoxical rarity of a fruit fly fungus attacking a broad range of hosts.

Understanding the factors that determine the realized and potential distribution of a species requires knowledge of abiotic, physiological, limitations as well as ecological interactions. Fungi of the order Laboulbeniales specialize on arthropods and are typically thought to be highly specialized on a single species or closely related group of species. Because infections are almost exclusively transmitted through direct contact between the hosts, the host ecology, to a large extent, determines the distribution and occurrence of the fungus. We examined ~20,000 fruit flies (Diptera: Dacinae) collected in Malaysia, Sulawesi, Australia, and the Solomon Islands between 2017 and 2019 for fungal infections and found 197 infected flies across eight different Bactrocera species. Morphology and 1,363 bps of small subunit (18S) DNA sequences both support that the infections are from a single polyphagous fungal species Stigmatomyces dacinus-a known ectoparasite of these fruit flies. This leads to the question: why is S. dacinus rare, when its hosts are widespread and abundant? In addition, the hosts are all Bactrocera, a genus with ~480 species, but 37 Bactrocera species found sympatric with the hosts were never infected. Host-selection does not appear to be phylogenetically correlated. These results suggest a hidden complexity in how different, but closely related, host species vary in their susceptibility, which somehow limits the abundance and dispersal capability of the fungus.

Checklist of thallus-forming Laboulbeniomycetes from Belgium and the Netherlands, including Hesperomyces halyziae and Laboulbenia quarantenae spp. nov.

In this paper we present an updated checklist of thallus-forming Laboulbeniomycetes (Ascomycota, Pezizomycotina), that is, the orders Herpomycetales and Laboulbeniales, from Belgium and the Netherlands. Two species are newly described based on morphology, molecular data (ITS, LSU ribosomal DNA) and ecology (host association). These are Hesperomyces halyziae on Halyzia sedecimguttata (Coleoptera, Coccinellidae) from both countries and Laboulbenia quarantenae on Bembidion biguttatum (Coleoptera, Carabidae) from Belgium. In addition, nine new country records are presented. For Belgium: Laboulbenia aubryi on Amara aranea (Coleoptera, Carabidae) and Rhachomyces spinosus on Syntomus foveatus (Coleoptera, Carabidae). For the Netherlands: Chitonomyces melanurus on Laccophilus minutus (Coleoptera, Dytiscidae), Euphoriomyces agathidii on Agathidium laevigatum (Coleoptera, Leiodidae), Laboulbenia fasciculata on Omophron limbatum (Coleoptera, Carabidae), Laboulbenia metableti on Syntomus foveatus and S. truncatellus (Coleoptera, Carabidae), Laboulbenia pseudomasei on Pterostichus melanarius (Coleoptera, Carabidae), Rhachomyces canariensis on Trechus obtusus (Coleoptera, Carabidae), and Stigmatomyces hydrelliae on Hydrellia albilabris (Diptera, Ephydridae). Finally, an identification key to 140 species of thallus-forming Laboulbeniomycetes in Belgium and the Netherlands is provided. Based on the combined data, we are able to identify mutual gaps that need to be filled as well as weigh the impact of chosen strategies (fieldwork, museum collections) and techniques in these neighboring countries. The aim of this work is to serve as a reference for studying Laboulbeniomycetes fungi in Europe.

Laboulbeniomycetes: Evolution, natural history, and Thaxter's final word.

Historically, thallus-forming Laboulbeniomycetes, including the orders Laboulbeniales and Herpomycetales, were set apart because of their distinctive morphology and ecology. Although some biologists correctly interpreted these arthropod ectoparasites as fungi, even ascomycetes, others thought they were worms, red algae, or members of taxa described especially for them. Speculation on the evolution of the group involving red algae, the morphology-based Floridean Hypothesis, persisted deep into the 20th century, in part because valid alternatives were not presented. Although the distinctive features of Laboulbeniales clearly set them apart from other fungi, the difficulty was in the absence of characters grouping them among the fungi. Thaxter considered the Laboulbeniales to be ascomycetes, but he avoided phylogenetic discussions involved in the Floridean Hypothesis all of his life. Eventually, developmental studies of the life history of Pyxidiophora species, hyphal perithecial ascomycetes with 2-celled ascospores, revealed characters connecting Laboulbeniales to other ascomycetes. The distinctive morphological features of Laboulbeniales (absence of mycelium, a thallus developed from 2-celled ascospores by cell divisions in several planes, arthropod parasitism) can be best understood by comparison with Pyxidiophora. The development of a 3-dimensional thallus composed of true parenchyma occurs not only in Laboulbeniales, but also in Pyxidiophora species. The life history of arthropod ectoparasitism of Laboulbeniales as well as mycoparasitism and phoretic dispersal by arthropods of Pyxidiophora species can be explained by Tranzschel's Law, originally applied to rust fungi. Molecular analyses including other arthropod-associated fungi have contributed to a better understanding of an enlarged class, Laboulbeniomycetes, which now includes a clade comprising Chantransiopsis, Tetrameronycha, and Subbaromyces. A two-locus phylogenetic tree highlights evolutionary and life history questions with regard to the placement of Herpomycetales as the first diverging lineage of the Laboulbeniomycetes. The sister group for all the Laboulbeniomycetes remains to be discovered.

Effects of fungal infection on the survival of parasitic bat flies.

BACKGROUND: Parasites are able to alter numerous aspects of their hosts' life history, behaviour and distribution. One central question in parasitology is to determine the degree of impact that parasites have on their hosts. Laboulbeniales (Fungi: Ascomycota) are ectoparasitic fungi of arthropods. Even though these fungi are widely distributed, little is known about their ecology and their possible physiological effects on their hosts. We used a highly specific bat fly-fungi association to assess the effect of these fungal parasites on their dipteran hosts. METHODS: We collected bat flies (Diptera: Nycteribiidae) belonging to two species, Nycteribia schmidlii and Penicillidia conspicua from their bat host Miniopterus schreibersii (Chiroptera: Miniopteridae). We experimentally tested the effect of infection on the lifespan of bat flies. RESULTS: The prevalence of Laboulbeniales fungi was 17.9% in N. schmidlii and 64.8% in P. conspicua. Two fungi species were identified, Arthrorhynchus eucampsipodae and A. nycteribiae, both showing strict host specificity with N. schmidlii and P. conspicua, respectively. We found that fungal infection reduced by half the survival rate of P. conspicua regardless of sex, whereas N. schmidlii was not affected by the infection. Moreover, the intensity of infection showed negative correlation with the lifespan of P. conspicua. CONCLUSIONS: To our knowledge, this is the first indication that fungal infection can alter bat fly survival and thus may play a significant role in the population dynamics of these bat ectoparasites.

Hyperparasitism in caves: Bats, bat flies and ectoparasitic fungus interaction.

Bat flies (Diptera: Nycteribiinae) are highly specialized bloodsucking bat ectoparasites. Some of the ectoparasitic bat flies are themselves parasitized with an ectoparasitic fungus of the genus Arthrorhynchus (Laboulbeniales). Ascospores of the fungus attach to the cuticle of a bat fly and develop a haustorium that penetrates the host cuticle. This interaction defines the fungus as a hyperparasite. Both the fly and the fungus are obligate parasites and this peculiar case of hyperparasitism has remained largely unstudied. We studied the prevalence of Laboulbeniales, genus Arthrorhynchus, in natural populations of bat flies infesting the bat species Miniopterus schreibersii, Myotis bechsteinii, My. blythii, My. daubentonii, My. escalerai and My. myotis in Portuguese caves. Laboulbeniales were found infecting 10 of the 428 screened bat flies (2.3%) in natural populations, with fewer infections in winter. Images obtained with transmission electron microscopy show the fungal haustorium within the bat fly host tissue, from where it extracts nutrition.

Studies on Dimorphomyceteae: I. New species of Nycteromyces and Dimeromyces (Laboulbeniales) on bat flies (Streblidae).

Three new species, Nycteromyces orientalis, Dimeromyces capensis, and D. streblidarum, belonging to the Laboulbeniales (Fungi, Ascomycota) and collected on bat flies (Diptera, Streblidae), are described and illustrated. The new species of Nycteromyces is the second described in this rarely collected genus, with the type species, N. streblidinus, previously collected only three times from Venezuela, Panama, and Honduras. The material for our new species comes from Australia, India, Philippines, and Tanzania, and morphological characters of these new collections require emendation of the original generic diagnosis. The distribution of N. streblidinus is also extended with additional collections from Costa Rica and Peru. We describe two new species of Dimeromyces from streblid hosts, the first report of any members of this large, diverse genus on this host family. Newly described taxa are compared with known species and the genera Nycteromyces and Dimeromyces discussed in relation to other known Laboulbeniales taxa.

Bats, Bat Flies, and Fungi: A Case of Hyperparasitism.

Bats are parasitized by numerous lineages of arthropods, of which bat flies (Diptera, Nycteribiidae and Streblidae) are the most conspicuous. Bat flies themselves can be parasitized by Laboulbeniales, fungal biotrophs of arthropods. This is known as hyperparasitism, a severely understudied phenomenon. Three genera of Laboulbeniales occur on bat flies: Arthrorhynchus on Nycteribiidae, Gloeandromyces and Nycteromyces on Streblidae. In this review we introduce the parasitic partners in this tripartite system and discuss their diversity, ecology, and specificity patterns, alongside some important life history traits. Furthermore, we cover recent advances in the study of the associations between bat flies and Laboulbeniales, which were neglected for decades. Among the most immediate needs for further studies are detailed tripartite field surveys. The vermin only teaze and pinch Their foes superior by an inch So, naturalists observe, a flea Has smaller fleas that on him prey; And these have smaller still to bite 'em, And so proceed ad infinitum. Jonathan Swift (On Poetry: A Rhapsody, 1733).

Laboulbeniales (Fungi: Ascomycota) infection of bat flies (Diptera: Nycteribiidae) from Miniopterus schreibersii across Europe.

BACKGROUND: Bat flies (Diptera: Nycteribiidae and Streblidae) are obligate, blood-sucking ectoparasites of bats with specialized morphology, life-cycle and ecology. Bat flies are occasionally infected by different species of Laboulbeniales (Fungi: Ascomycota), microscopic fungal ectoparasites belonging to three genera: Arthrorynchus spp. are restricted to the Eastern Hemisphere, while species of Gloeandromyces and Nycteromyces occur on Neotropical bat flies. Little is known about the distribution and host specificity of Arthrorynchus spp. on bat flies. In this study, we focused on sampling bat flies from the cave-dwelling bat species Miniopterus schreibersii. Bat and ectoparasite collection took place in Albania, Croatia, Hungary, Italy, Portugal, Slovakia, Spain and Switzerland. Flies were inspected for Laboulbeniales infections. RESULTS: Six hundred sixty seven bat flies of five species were collected: Nycteribia latreillii, N. pedicularia, N. schmidlii, Penicillidia conspicua, and P. dufourii. Laboulbeniales infection was observed on 60 specimens (prevalence = 9%). Two Laboulbeniales species, Arthrorhynchus eucampsipodae and A. nycteribiae, were present on three bat fly species. All observations of A. eucampsipodae were on N. schmidlii, and A. nycteribiae was present on P. conspicua and P dufourii. Arthrorhynchus eucampsipodae is, for the first time, reported from Slovakia and Spain. Arthrorhynchus nycteribiae represents a new country record for Portugal and Slovakia. There were no significant differences among infection rates in different countries. Females of N. schmidlii showed a higher infection rate than males with an observable trend (P = 0.0502). No sex differences in infection rate for P. conspicua and P. dufourii were detected. Finally, thallus density was significantly lower in N. schmidlii compared to P. conspicua and P. dufourii. CONCLUSIONS: With this study, we contribute to the knowledge of the geographical distribution and host specificity of Laboulbeniales fungi associated with ectoparasitic bat flies within Europe. We discuss parasite prevalence and host specificity in the light of our findings and the available literature. Penicillidia conspicua is unambiguously the main host species for A. nycteribiae based on our and previous findings. Differences in parasite intensity and sex-biased infections of the fungi are possible depending on the species.

Molecular phylogeny of the Laboulbeniomycetes (Ascomycota).

A first molecular-based phylogeny is presented for the Laboulbeniomycetes, a group of ascomycete fungi that utilize arthropods for nutrition and/or dispersal. Morphological diversification and life-history evolution has made it difficult to resolve relationships within the group, and to identify close relatives. Here, we infer a preliminary phylogeny based on acquisition of 51 new SSU rDNA sequences, representing a total of 65 taxa. The results of this study demonstrate that Laboulbeniomycetes is monophyletic, and related to Sordariomycetes. The class could be divided into at least 4 or 5 orders, though we refrain from formally giving names to these at this stage. Further evidence for the occurrence of asexuality within the class is provided by the inclusion of the genera Chantransiopsis and Tetrameronycha, both known only as asexual taxa with thalli consisting of linearly superposed cells. The precise placement of the genus Herpomyces (Herpomycetaceae), on cockroaches, remains unresolved in our analysis, but lies outside of the main clade of sexually reproducing Laboulbeniales. There is good support for this latter grouping, comprising taxa that are found on both aquatic and terrestrial hosts. Within this large assemblage, we recognize 5 distinct clades (clades E, F, G, H, I). Relationships among the so-called "aquatic genera" (≡ Ceratomycetaceae + some Euceratomycetaceae and Zodiomyces) are poorly resolved in our analyses, accounting for 3 of these clades (E, F, G), with the remainder of the taxa (largely equivalent to Laboulbeniaceae) split into two major groupings (clades H, I). Across all taxa, antheridial characteristics, features on which the earliest classifications were based, are shown to be homoplastic. On the other hand, features of perithecial development show an overall trend towards reduction, and appear to be phylogenetically informative. Morphological characters are identified that support the dichotomy in the Laboulbeniaceae and subclades within the two major groupings are discussed further in light of information on thallus morphology, development, and host relationships.

A new East African genus of spirostreptid millipedes (Diplopoda, Spirostreptida, Spirostreptidae), with notes on their fungal ectoparasite Rickia gigas.

The new genus Tropostreptus is erected to accommodate seven species which are all endemic in Tanzania: T. hamatus (Demange, 1977) (type species, = Spirostreptus hamatus Demange, 1977), T. sigmatospinus sp. nov., T. microcephalus sp. nov., T. droides sp. nov., T. kipunji spec. nov., T. austerus (Attems, 1950) (= Epistreptus austerus Attems, 1950) and T. severus sp. nov. All but one (T. sigmatospinus) of the species are restricted to the Eastern Arc Mts. in a wide sense, i.e., including Mt. Rungwe. Notes are included on Rickia gigas Santamaria et al., 2016 (Ascomycota: Laboulbeniales), which infects several Tropostreptus species. The typical, long-celled form of R. gigas grows on body parts of both sexes coming into contact during copulation, whereas a short-celled form also ascribed to R. gigas grows on the hind margin of body rings of both sexes. A key to Tropostreptus species is included.

The effects of fungal infection and physiological condition on the locomotory behaviour of the ant Myrmica scabrinodis.

Parasite infection often results in alterations in host behaviour. These changes vary greatly in their magnitude, from slight shifts in the time spent by the host performing a given activity to the appearance of novel behaviours. The effects of parasites can differ with the age and the physiological condition of the host. Rickia wasmannii is an ectoparasitic fungal symbiont in Myrmica ants that covers the whole body surface of the host and reduces its lifespan. The fungus is present in both young and old individuals, making it an optimal subject for the study of age-related parasitic effects. We tested the effect of fungal infection on the locomotory activity of the Myrmica scabrinodis ant in different age categories. The fat content of workers was measured as a proxy for their physiological status. Based on our findings, old workers bore more thalli and were leaner than young individuals, while they tended to move at higher speeds and with a lower degree of meandering. Young individuals covered smaller distances, at slower speeds and with a higher degree of meandering. Contrary to our expectations, the infection intensity of R. wasmannii affected neither the fat content nor the locomotory activity of ant workers. However, the two age classes seem to have different strategies with regards to the relationship between fat content and distance covered. Our results suggest that characteristics of locomotory activity differ between the age classes in many respects, and are also influenced by their physiological status, but parasitism by R. wasmannii does not seem to have a straightforward effect on any of the variables studied.

Fireworks under the microscope: a spectacular new species of Zodiomyces from the Thaxter collection.

A new species of Zodiomyces (Ascomycota, Laboulbeniales) is described, Z. rhizophorus, parasitic on a hydrophilid beetle (Coleoptera, Hydrophilidae) from Trinidad. This species was discovered during the examination of the slides of Laboulbeniales made by Roland Thaxter. It is characterized by numerous long, slender, multicellular and multiseriate outgrowths at the base of the receptacle. Thaxter's outstanding illustrations have set a standard in the field of mycology; we provide a review of the methods he employed in the preparation of these illustrations.

Distribution of the myrmecoparasitic fungus Rickia wasmannii (Ascomycota: Laboulbeniales) across colonies, individuals, and body parts of Myrmica scabrinodis.

The ant species Myrmica scabrinodis plays a markedly important ecological role through much of the humid grasslands of Eurasia. It hosts a species-rich community of pathogens and parasites, including Rickia wasmannii, an enigmatic member of entomoparasitic laboulbenialean fungi. This study provides a descriptive ecology of R. wasmannii by characterizing its prevalence and distribution across several hierarchical levels: colonies, individuals, and anatomic body parts. Infections were restricted to a single ant species, M. scabrinodis, and infected colonies occurred predominantly in wet habitats. Infections tended to be highly prevalent within infected colonies, often reaching 100% sample prevalence among workers. Individual infections exhibited an aggregated distribution typical to host-parasite systems. Workers from the aboveground part of nests (presumably older ones acting as foragers) were more infected than those from the belowground part. Fungal thalli could be found all over the body of the hosts, the head and the abdomen being the most infected parts of the body. The fungi's distribution among host body parts statistically differed between low versus high-intensity infections: the initial dominance of the head decreased with advancing infection. These findings may provide baseline data for future comparative or monitoring studies.

Bringing Laboulbeniales into the 21st century: enhanced techniques for extraction and PCR amplification of DNA from minute ectoparasitic fungi.

Laboulbeniales is one of the most peculiar orders of Ascomycota. These fungi are characterized by an ectoparasitic life-style on arthropods, determinate growth, lack of an asexual stage, high species richness, and intractability to culture. The order Laboulbeniales, sister to Pyxidiophorales, has only recently been assigned a separate class, the Laboulbeniomycetes, based on very few ribosomal DNA sequences. So far, DNA isolations and PCR amplifications have proven difficult. Here, we provide details of isolation techniques and the application of commercially available kits that enable efficient and reliable genetic analyses of these fungi. We provide 43 newly generated Laboulbeniales ribosomal DNA sequences, among which are the first published sequences for species in the genera Gloeandromyces, Herpomyces, Laboulbenia, Monoicomyces, and Polyandromyces. DNA extractions were possible using from 1 to 30 thalli from hosts preserved in ethanol (70-100 %). In two cases, we successfully isolated DNA from thalli on dried insect collections. Laboulbeniales molecular systematics could be substantially enhanced through these improved methods by allowing more complete sampling of both taxa and gene regions.

A new species of the genus euphoriomyces (laboulbeniales ascomycotina) collected in Korea.

One new species of the genus Euphoriomyces was collected on Scaphisoma rufum Achard (Scaphidiidae) in South Korea. Euphoriomyces chaekyui sp. nov. is unique in having a single terminal phialide shaped an elongated cylindrical form on the distal portion of the main axis.