Putative Species Research Articles

Monostephanostomum mesospinosum (Digenea: Acanthocolpidae) Infecting the Coastal Trevally Carangoides caeruleopinnatus (Carangiformes: Carangidae) from the Marine Water off Saudi Arabia

Background: Carangids, like any other fish, can become infected with a variety of external and internal parasites. Little information on parasite infestations in these fish species is known. This study aimed to provide light on parasites that infect caragnids in Saudi Arabia. Methods: A total of 20 Carangoides caeruleopinnatus specimens were collected from the Red Sea (Saudi Arabia) and then examined for the presence of digenetic trematodes. Parasitic species were isolated and studied morphologically by using light microscopic examination and molecularly via the partial sequencing of 18S rRNA and 28S rRNA genes. Result: Only one parasitic species has been identified. This parasite is morphologically and morphometric compatible with previously Monostephanostomum mesospinosum, identified from Carangoides malabaricus in VSK, BOB. Phylogeny revealed that these putative acanthocolpid species nested well within a clade clustering Monostephanostomum species, which along with morphological data, suggests it is a member of the genus Monostephanostomum. Query sequences showed identities of 97.99% and 97.98% for 18S (EF506762.1.1) and 28S rRNA (EF506763.1) of Monostephanostomum nolani, respectively. This study reflects the third account of this genus as endoparasitic taxa of various carangid fish, as well as providing novel DNA data for this species.

Genetic population structures of common scavenging species near hydrothermal vents in the Okinawa Trough

Deep-sea mining of hydrothermal deposits off the coast of Japan is currently under consideration, and environmental baseline studies of the area are required to understand possible impacts. The aim of this study is to clarify population structures of dominant benthic megafaunal species near hydrothermal vent fields in the Okinawa Trough, using a population genetics approach. We examined dominant deep-sea scavenging species including eels, several amphipods, and a decapod and performed population genetic analyses based on the mitochondrial cytochrome c oxidase subunit I region. Several sites were sampled within Okinawa Trough to examine intra-population diversity while two other locations 1400–2400 km away were chosen for inter-population comparisons. For synaphobranchid eels Simenchelys parasitica and Synaphobranchus kaupii, our results showed significant intra-population diversity but no inter-population genetic differentiation, suggesting strong genetic connectivity and/or large population sizes. In addition, single nucleotide polymorphism analysis also confirmed strong genetic connectivity for Simenchelys parasitica. Among scavenging amphipods, we detected seven putative species using molecular phylogenetic analysis. We evaluated population structures of the most abundant species of amphipods and a decapod species (Nematocarcinus lanceopes). Our results provide basic information on the genetic population structures of benthic megafaunal species near hydrothermal vent fields, which can be used to select candidate species for future connectivity analysis with high-resolution genetic markers and aid understanding of the potential population impacts of environmental disturbances.

A Study in Scarlet: Integrative Taxonomy of the Spider Genus Loureedia (Araneae: Eresidae)

The eresid spider genus Loureedia (Miller et al., 2012) was described a decade ago, despite its type species being described in the mid-19th century, which illuminates the difficulties in obtaining specimens. The genus was initially described as monotypic. Ever since, four other species have been assigned to Loureedia, including three newly discovered ones. Primarily due to the extravagant appearance of the males, stories about the discovery of species of Loureedia have been the subject of relatively wide media coverage over the years, leading to numerous new populations and putative undescribed species being documented by naturalists and citizen scientists. These species, although bearing distinct differences in their coloration patterns, typically vary only slightly in the structure of their copulatory organs, the primary traits used in spider systematics. This highlights an important taxonomic problem: while it is easy to diagnose the genus or recognize the species that belong to it, it is challenging to differentiate the species from one another, particularly when using only a single line of evidence. In this paper, we have tackled this issue using an integrative approach, i.e., a combination of molecular markers (the mitochondrial COI) and traditional morphological characters. The effects of different observational angles on the perceived shape of the conductor are discussed. Except for one species, we obtained DNA data of all members of the genus. Based on these data, the first phylogeny for Loureedia is presented, and two North African species, Loureedia maroccana (Gál et al., 2017) and Loureedia jerbae (El-Hennawy, 2005), are revalidated from synonymy. The distribution records of all described species are mapped.

A note on cetaceans off Kiribati and Tuvalu from a research cruise in October 2010

This paper summarises the results of a sighting survey conducted around the Gilbert Islands (Kiribati) and Tuvalu Islands (Tuvalu) in the central Pacific Ocean between 3 and 17 October 2010. This was the first systematic collection of cetacean sighting data in this region. The main objective of the survey was to investigate the occurrence and distribution of cetaceans around Kiribati and Tuvalu. In addition biopsy samples were obtained to investigate the species identity of Bryde’s-whale-like baleen whales through genetic analyses and to assess feeding ecology of cetaceans in the survey area through the examination of fatty acids. The survey was carried out using a sighting survey vessel, which covered a total of 1,012 n.miles (≈1,875km). A total of 24 schools (640 individuals) of cetaceans was sighted: three schools (five individuals) of Bryde’s-whale-like baleen whales, one school (nine individuals) of sperm whales (Physeter macrocephalus), one school (six individuals) of killer whales (Orcinus orca), one school (14 individuals) of short finned pilot whales (Globicephala macrorhynchus), one school (two individuals) of false killer whales (Pseudorca crassidens), eight schools (483 individuals) of spinner dolphins (Stenella longirostris), and one school (70 individuals) of striped dolphins (Stenella coeruleoalba). Mitochondrial DNA (mtDNA) analyses based on the biopsy samples identified two of the Bryde’s-whale-like whales sighted, as of the putative species Balaenoptera brydei. Compositions of fatty acids of Bryde’s and killer whales are presented. The survey provided new information on the distribution and fatty-acid composition of cetaceans around Kiribati and Tuvalu.

Diversity of viroids infecting grapevines in the South African Vitis germplasm collection.

Seven viroid species and one putative viroid species have been reported to infect grapevine namely, hop stunt viroid (HSVd), grapevine yellow speckle viroid 1 (GYSVd-1), grapevine yellow speckle viroid 2 (GYSVd-2), Australian grapevine viroid (AGVd), Japanese grapevine viroid (JGVd), grapevine latent viroid (GLVd), and citrus exocortis viroid (CEVd), as well as a grapevine hammerhead viroid-like RNA (GHVd), so far. In this study, RNA sequence (RNA-Seq) data, from 229 Vitis accessions from the field-maintained vineyard of the South African Vitis germplasm collection, were analysed to determine the diversity of the viroids present. Five of the seven known grapevine-infecting viroids and one putative grapevine-infecting viroid species were very commonly found, with 214 of the 229 samples containing at least one viroid species. HSVd, GYSVd-1, GYSVd-2, AGVd, and JGVd, as well as GHVd, were identified in the RNA-Seq data of the samples and confirmed using RT-PCR and Sanger sequencing. The HSVd sequences indicated the presence of two variants, with one showing multiple nucleotide insertions. AGVd and GYSVd-2 did not display significant sequence diversity, confirming past international studies. GYSVd-1 occurs as four major variants worldwide and representatives of all four variants were identified in this vineyard. This is the first report on the diversity of viroids infecting grapevine in South Africa and the first report of JGVd outside of Japan and GHVd in South Africa. Further studies are needed to fully assess the population and to identify potentially new viroid species.

Demographic history and species delimitation of threeZamiaspecies (Zamiaceae) in south-eastern Mexico:Z. katzerianais not a product of hybridization

AbstractSpeculations on hybridization among wild cycads are abundant, but empirical evidence of hybridization in cycad evolution is almost non-existent and the evolutionary consequences of hybridization in cycads have not been discussed. Previously, Zamia katzeriana was hypothesized to have a hybrid origin between two sympatric species: Z. splendens and Z. loddigesii. In this study, we aimed to reveal whether such a hypothesis is confirmed. Variation in 12 microsatellite loci, 12 morphological traits and environmental niche of ten populations representing the three species was examined. Gene flow and scenarios of demographic history were tested. Ecological niche models were reconstructed to examine present and past niche availability of the two putative parental species. Correlations between genetic, morphological, environmental and geographical distances were tested. We found that Z. katzeriana diverged from Z. splendens without experiencing admixture with its sympatric congeners. However, gene flow between Z. splendens and Z. loddigesii has occurred in one locality. The niches of the two latter species have overlapped since the Last Glacial Maximum, when admixture probably occurred. However, the admixed populations do not resemble Z. katzeriana. These results confirm that Z. katzeriana did not originate from hybridization between Z. splendens and Z. loddigesii.

Not the Last Piece of the Puzzle: Niphargus Phylogeny in Hungary

The Palaearctic genus Niphargus is a promising model system to understand subterranean fauna genesis in Europe. The Pannonian Plain (mainly covered by Hungary) in Central Europe, once being the area of the Paratethys, is a key area for Niphargus diversification. However, our knowledge on Hungarian species of Niphargus is primarily based on sporadic taxonomical works from the pre-molecular era. Here, we studied 14 localities, covering the eight valid Hungarian species of Niphargus and including nine previously unstudied populations. Based on sequences of three gene fragments, we reconstructed their phylogeny using maximum likelihood and Bayesian approaches. We found that not all Hungarian species of Niphargus are closely related, and even species sampled at the same localities can belong to different clades. Some Hungarian species form monophyletic clades, while others are nested in various non-Hungarian lineages. The new populations are all genetically distinct from the known species. Our results suggest that the Hungarian Niphargus fauna has originated from seven unrelated clades and its diversity is underestimated due to unknown populations and cryptic species. The detection of genetically distinct species of Niphargus from non-carbonate regions calls for further research efforts. The high diversity and the number of putative new species in the N. tatrensis clade warrants further, high-resolution phylogenetic studies.

Global population genetic structure and lineage differentiation of the stable fly, Stomoxys calcitrans.

The bloodsucking fly, Stomoxys calcitrans (Diptera: Muscidae), is a cosmopolitan pest that transmits potential pathogens mechanically. We conduct phylogeographic analyses of S. calcitrans to resolve its global population genetic structure for establishing baseline of molecular studies. Results from mitochondrial gene suggested that the major divergence of S. calcitrans predominantly occurred 0.32-0.47 million years ago (Mya) and the subsequent diversifications took place during 0.13-0.27 Mya. The Ethiopian region was deduced as the most likely origin of S. calcitrans and the Nearctic lineages were considered to have originated from Oriental or Palaearctic regions. Our results further revealed that each biogeographic region of S. calcitrans likely maintains its genetic specialty, and yet, those non-monophyletic relationships were possibly caused by ancestral retention, dispersal with mammals, long-distance migration, and the international livestock industries. Moreover, the three highly diverged Ethiopian lineages may be putative cryptic species that require clarification of their veterinary importance. Unravelling the genetic structure of stable fly and preventing gene flow among biogeographic regions through anthropogenic activities are thus pivotal in livestock industry administration, particularly genetic exchange among differentiated lineages that might lead to the consequence of ecological trait alterations.

Complete genome sequence of marine Roseobacter lineage member Ruegeria sp. YS9 with five plasmids isolated from red algae.

Ruegeria sp. YS9, an aerobic and chemoheterotrophic bacterium belonging to marine Roseobacter lineage, was a putative new species isolated from red algae Eucheuma okamurai in the South China Sea (Beihai, Guangxi province). The complete genome sequence in strain YS9 comprised one circular chromosome with 3,244,635bp and five circular plasmids ranging from 38,085 to 748,160bp, with a total length of 4.30Mb and average GC content of 58.39%. In total, 4129 CDSs, 52 tRNA genes and 9 rRNA genes were obtained. Genomic analysis of strain YS9 revealed that 85 CAZymes were organized in 147 PUL-associated CAZymes involved in polysaccharides metabolism, which were the highest among its two closely related Ruegeria strains. Numerous PULs related to degradation on the cell wall of algae, especially agar, indicated its major player role in the remineralization of algal-derived carbon. Further, the existence of multiple plasmids provided strain YS9 with distinct advantages to facilitate its rapid environmental adaptation, including polysaccharide metabolism, denitrification, resistance to heavy metal stresses such as copper and cobalt, type IV secretion systems and type IV toxin-antitoxin systems, which were obviously different from the two Ruegeria strains. This study provides evidence for polysaccharide metabolic capacity and functions of five plasmids in strain YS9, broadening our understanding of the ecological roles of bacteria in the environment around red algae and the function patterns of plasmids in marine Roseobacter lineage members for environmental adaptation.

First Report of Powdery Mildew Caused by Golovinomyces sonchicola on Sonchus wightianus in China.

Sonchus wightianus DC. is a perennial plant of the Asteraceae family, which is native to Nepal (Veena et al. 2021). In China, S. wightianus is distributed throughout Hainan Province and is commonly used for edible and medicinal purposes. During the spring of 2021 and 2022, powdery mildew symptoms were observed on 70% of S. wightianus plants on the Hainan Medical University campus (19° 58' 53″ N; 110° 19' 47″ E) in Haikou, Hainan Province, China. Powdery mildew colonies covered the leaf surfaces and stems of affected plants, causing discoloration and defoliation. Mycelia were superficial and hyphal appressoria were nipple-shaped, single or in pairs. Conidiophores (n =30) were unbranched, 81 to 209 × 15 to 19 µm. Foot cells (n =30) were cylindrical, straight or curved at the base, 14 to 78 µm long, followed by one to three shorter cells, and formed conidia in chains. Conidia (n =100) were ellipsoid to doliiform-cylindrical, 27 to 39 ×12 to 19 m (length/width ratio = 1.5 to 2.6), lacked fibrosin bodies, and produced germ tubes at their termini. TheEudoidium-type of conidial germination with a short germ tube was observed. Based on these morphological characteristics, the pathogen resembledGolovinomyces sp. (Braun and Cook 2012). The teleomorph was not observed. A specimen was deposited in the Hainan Medical University Plant Pathology Herbarium as HMSW-21. To confirm the genus identification and ascertain a putative species, genomic DNA was extracted from mycelium, conidiophores, and conidia using a fungal DNA kit (Omega Bio-Tek, USA). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1/ITS4 (White et al. 1990) and sequenced directly. The resulting 607 bp sequence was deposited in GenBank (accession no. ON365772). A BLASTn search in GenBank of this sequence showed ≥99% similarity with the ITS sequences of G. sonchicola isolates from Mexico (MW425872), Slovenia (MK318522), and the United Kingdom (KY660772). Additionally, the 28S rDNA region was amplified using the primer pairs NL1 and NL4 (O´Donnell 1993; accession no. ON365785). This region shared 99% similarity with published G. sonchicola isolates (OM265397 and AB077624) as well. To confirm pathogenicity, five healthy potted plants of S. wightianus were inoculated by gently pressing a powdery mildew-infected leaf onto 15 young leaves. Five non-inoculated plants served as controls. All plants were maintained in a greenhouse at 24 to 30°C, 70% relative humidity, with a 16-h photoperiod. After 7 days, inoculated leaves showed powdery mildew symptoms whereas no symptoms were observed on control plants. The fungal colonies observed on inoculated plants were morphologically identical to those found on the originally infected leaves collected from Hainan Province. Based on the morphological characteristics and molecular identification, the fungus was identified as G. sonchicola. In different countries and regions, G. sonchicolahas been previously reported on some Sonchus species, including S. oleraceus from Mexico (Beltrán-Peña et al. 2021) and Slovenia (Jakše et al. 2019), and S. asper in the United Kingdom. To our knowledge, this is the first record of G. sonchicola infecting S. wightianus in China. We are concerned that the pathogen could become a threat to the widespread planting of S. wightianus in the future.

Putative Mitoviruses without In-Frame UGA(W) Codons: Evolutionary Implications.

Mitoviruses are small vertically transmitted RNA viruses found in fungi, plants and animals. Taxonomically, a total of 105 species and 4 genera have been formally recognized by ICTV, and recently, 18 new putative species have been included in a new proposed genus. Transcriptomic and metatranscriptomic studies are a major source of countless new virus-like sequences that are continually being added to open databases and these may be good sources for identifying new putative mitoviruses. The search for mitovirus-like sequences in the NCBI databases resulted in the discovery of more than one hundred new putative mitoviruses, with important implications for taxonomy and also for the evolutionary scenario. Here, we propose the inclusion of four new putative members to the genus Kvaramitovirus, and the existence of a new large basally divergent lineage composed of 144 members that lack internal UGA codons (subfamily "Arkeomitovirinae"), a feature not shared by the vast majority of mitoviruses. Finally, a taxonomic categorization proposal and a detailed description of the evolutionary history of mitoviruses were carried out. This in silico study supports the hypothesis of the existence of a basally divergent lineage that could have had an impact on the early evolutionary history of mitoviruses.

The Expanding Menagerie of Prunus-Infecting Luteoviruses.

Members of the genus Luteovirus are responsible for economically destructive plant diseases worldwide. Over the past few years, three luteoviruses infecting Prunus trees have been characterized. However, the biological properties, prevalence, and genetic diversity of those viruses have not yet been studied. High-throughput sequencing of samples of various wild, cultivated, and ornamental Prunus species enabled the identification of four novel species in the genus Luteovirus for which we obtained complete or nearly complete genomes. Additionally, we identified another new putative species recovered from Sequence Read Archive data. Furthermore, we conducted a survey on peach-infecting luteoviruses in eight European countries. Analyses of 350 leaf samples collected from germplasm, production orchards, and private gardens showed that peach-associated luteovirus (PaLV), nectarine stem pitting-associated virus (NSPaV), and a novel luteovirus, peach-associated luteovirus 2 (PaLV2), are present in all countries; the most prevalent virus was NSPaV, followed by PaLV. The genetic diversity of these viruses was also analyzed. Moreover, the biological indexing on GF305 peach indicator plants demonstrated that PaLV and PaLV2, like NSPaV, are transmitted by graft at relatively low rates. No clear viral symptoms have been observed in either graft-inoculated GF305 indicators or different peach tree varieties observed in an orchard. The data generated during this study provide a broader overview of the genetic diversity, geographical distribution, and prevalence of peach-infecting luteoviruses and suggest that these viruses are likely asymptomatic in peach under most circumstances.

A review of bats of the genus Pseudoromicia (Chiroptera: Vespertilionidae) with the description of a new species

The Cameroon Volcanic Line, which divides the Congo Basin fauna from the West African fauna, is a known area of high endemism for various taxa, but the region’s bat fauna has received little attention. We review variation in morphological and molecular (mitochondrial Cytochrome b) characters in the Tropical African vespertilionid bat genus Pseudoromicia. Assessment of this variation indicates the existence of a new species of Pseudoromicia, from the Mbam Minkom Massif in the Centre Region of Cameroon. The new species is diagnosable by sequence data and is morphologically similar to its putative sister taxon, P. kityoi, from Uganda. Although we suggest that it be assigned to the IUCN category of Data Deficient, there should be some concern as to the conservation status of this species: the Mbam Minkom Massif ecosystem is threatened due to lack of legal conservation frameworks and exposure to increasing human pressure. The new species is in a clade with P. roseveari and P. kityoi. These three species may be relicts of a single widespread species originating in the West African “white-winged” group of Pseudoromicia that then spread east across the tropical moist broadleaf forest into East Africa, and are now restricted to a few upland rainforest patches in West Africa (P. roseveari), in outliers of the Cameroon Volcanic Line region (Pseudoromicia sp.), and the Lake Victoria area (P. kityoi). The smaller, white-winged species are hypothesized to be ancestral, with one extant putative species (P. rendalli) also extending into East and southern Africa (Van Cakenberghe & Happold, 2013). The larger, dark-winged taxa likely dispersed to East Africa and subsequently back to West Africa (e.g., Pseudoromicia sp., P. roseveari). Our data illustrate the potential importance of the Dahomey Gap and climatic changes in the evolution of this group of species. http://zoobank.org/urn:lsid:zoobank.org:pub:30929BAA-4043-422D-B209-16CAB826043E

Re-evaluating Homoploid Reticulate Evolution in Helianthus Sunflowers.

Sunflowers of the genus Helianthus are models for hybridization research and contain three of the best-studied examples of homoploid hybrid speciation. To understand a broader picture of hybridization within the annual sunflowers, we used whole-genome resequencing to conduct a phylogenomic analysis and test for gene flow between lineages. We find that all annual sunflower species tested have evidence of admixture, suggesting hybridization was common during the radiation of the genus. Support for the major species tree decreases with increasing recombination rate, consistent with hybridization and introgression contributing to discordant topologies. Admixture graphs found hybridization to be associated with the origins of the three putative hybrid species (Helianthus anomalus, Helianthus deserticola, and Helianthus paradoxus). However, the hybridization events are more ancient than suggested by previous work. Furthermore, H. anomalus and H. deserticola appear to have arisen from a single hybridization event involving an unexpected donor, rather than through multiple independent events as previously proposed. This means our results are consistent with, but not definitive proof of, two ancient independent homoploid hybrid speciation events in the genus. Using a broader data set that covers the whole Helianthus genus, including perennial species, we find that signals of introgression span the genus and beyond, suggesting highly divergent introgression and/or the sorting of ancient haplotypes. Thus, Helianthus can be viewed as a syngameon in which largely reproductively isolated species are linked together by occasional or frequent gene flow.

Taxonomic Revision of Tribe Aleurocanthini Takahashi 1954 stat. rev. Using Consortium Gene Analysis (Mito-Nuclear-Primary Endosymbiont) with the First Evidence for Mitochondrial Recombination in Whitefly (Hemiptera: Aleyrodidae)

The discovery of the Aleurocanthus cf. Aleurocanthus spiniferus (Tea spiny whitefly spiniferus morphotype; previously found in Tokyo) for the first time in Shizuoka Prefecture raised the possibility that this species had invaded Honshu Island, Japan. Unlike the allied species, Aleurocanthus spiniferus and A. camelliae, which have been intercepted from China to Japan, the origin of the current species remains unclear. Despite the status of this species as a minor pest on the ornamental plant, Eurya japonica, the cryptic diversity among the black spiny whitefly is fascinating to be elucidated, specifically how the primary endosymbiont of whiteflies, Portiera aleyrodidarum, coevolved and contributed to the classification of whiteflies. The current study examines the taxonomic status of five species of whiteflies, i.e., A. spiniferus (Quaintance), Aleurocanthus aff. A. camelliae, Aleurocanthus cf. A. spiniferus, A. camelliae Kanmiya and Kasai, Aleurotrachelus camelliae Kuwana, and A. ishigakiensis Takahashi. Using consortium molecular typing targeting mitochondrial DNA (COI and 16S of mitoribosome), the nuclear gene (ITS1), and the ribosomal gene of Portiera, the phylogenetic clustering analysis has been conducted and revealed that the genus Aleurotrachelus sensu lato was clustered together with Aleurocanthini Takahashi, 1954 stat. rev. and reinstated Crenidorsum ishigakiensis comb. nov. due to crescent-shaped scallops being clearly defined. The current study also unveiled several putative species in the A. spiniferus species complex, molecularly. In addition, the recombination event was not detected in Portiera but has been detected in the mtCOI genes of the A. spiniferus cryptic species and the A. woglumi sequences deposited in the NCBI database. The mitochondrial recombination gives an insight into the speciation process among this species complex.

Contribution to the knowledge of genome size variation in Calendula L. (Asteraceae) with special focus on the SW Mediterranean region

Calendula is one of the most taxonomically complex genera within the Asteraceae family due to hybridization, polyploidization events, and production of a highly variable morphology of the achenes. Considering the complexity of Calendula, this study was conducted to extend the understanding of the relationships between SW Mediterranean taxa, and assess the relationships between genome size and chromosome number, ploidy level and life cycle in 77 populations covering 14 taxa. Genome size estimations are provided for the first time for five species and for three putative new undescribed species. Mean 2C values differed up to 6-fold among different euploid species (from 1.37 pg in diploid to 8.26 pg in octoploid populations). 1Cx-values varied 2.07-fold (between 0.68 pg and 1.41 pg). Mean 1Cx genome size revealed significant differences between different ploidy levels. Genome size variation is a significant factor for explaining the relationships within Calendula, and individuals not fitting the current classification were found and should be analysed in detail in future studies.

Old Methods, New Insights: Reviewing Concepts on the Ecology of Trypanosomatids and Bodo sp. by Improving Conventional Diagnostic Tools.

Mixed infections by different Trypanosoma species or genotypes are a common and puzzling phenomenon. Therefore, it is critical to refine the diagnostic techniques and to understand to what extent these methods detect trypanosomes. We aimed to develop an accessible strategy to enhance the sensitivity of the hemoculture, as well as to understand the limitations of the hemoculture and the blood clot as a source of parasitic DNA. We investigated trypanosomatid infections in 472 bats by molecular characterization (18S rDNA gene) of the DNA obtained from the blood clot and, innovatively, from three hemoculture sample types: the amplified flagellates ("isolate"), the pellet of the culture harvested in its very initial growth stage ("first aliquot"), and the pellet of non-grown cultures with failure of amplification ("sediment"). We compared (a) the characterization of the flagellates obtained by first aliquots and isolates; and (b) the performance of the hemoculture and blood clot for trypanosomatid detection. We observed: (i) a putative new species of Bodo in Artibeus lituratus; (ii) the potential of Trypanosoma cruzi selection in the hemoculture; (iii) that the first aliquots and sediments overcome the selective pressure of the hemoculture; and (iv) that the blood clot technique performs better than the hemoculture. However, combining these methods enhances the detection of single and mixed infections.

Unravelling the diversity of Anaplasma species circulating in selected African wildlife hosts by targeted 16S microbiome analysis

Organisms in the genus Anaplasma are obligate intracellular alphaproteobacteria. Bovine anaplasmosis, predominantly caused by Anaplasma marginale, is the most prevalent tick-borne disease (TBD) of cattle worldwide. Other Anaplasma species are known to cause disease; these include A. ovis, A. platys in dogs, A. capra in goats and humans, and A. phagocytophilum in humans. The rapid advancement of next-generation sequencing technologies has led to the discovery of many novel sequences ascribed to the genus Anaplasma, with over 20 putative new species being proposed since the last formal organization of the genus. Most 16S rRNA gene surveys for Anaplasma were conducted on cattle and to a lesser extent on rodents, dogs, and ticks. Little is known about the occurrence, diversity, or impact of Anaplasma species circulating in wildlife species. Therefore, we conducted a 16S rRNA gene survey with the goal of identifying Anaplasma species in a variety of wildlife species in the Kruger National Park and neighbouring game reserves, using an unbiased 16S rRNA gene microbiome approach. An Anaplasma/Ehrlichia-group specific quantitative real-time PCR (qPCR) assay revealed the presence of Anaplasma and/or Ehrlichia species in 70.0% (21/30) of African buffalo, 86.7% (26/30) of impala, 36.7% (11/30) of greater kudu, 3.2% (1/31) of African wild dog, 40.6% (13/32) of Burchell's zebra, 43.3% (13/30) of warthog, 22.6% (7/31) of spotted hyena, 40.0% (12/30) of leopard, 17.6% (6/34) of lion, 16.7% (5/30) of African elephant and 8.6% (3/35) of white rhinoceros samples. Microbiome sequencing data from the qPCR positive samples revealed four 16S rRNA sequences identical to previously published Anaplasma sequences, as well as nine novel Anaplasma 16S genotypes. Our results reveal a greater diversity of putative Anaplasma species circulating in wildlife than currently classified within the genus. Our findings highlight a potential expansion of the Anaplasma host range and the need for more genetic information from other important genes or genome sequencing of putative novel species for correct classification and further assessment of their occurrence in wildlife, livestock and companion animals.

Morphological and Genetic Assessment of Sympatric Lenok Species (Brachymystax spp.) in the Onon River, Mongolia

The genus Brachymystax is represented by three putative species that disjointedly inhabit the major river basins of central and northeast Eurasia. Brachymystax lenok and B. tumensis have been regularly misidentified throughout their range, including across sympatric populations in the upper Onon River of Mongolia. This research has focused on Mongolia’s understudied lenok populations by conducting detailed morphometric measurements and genetic analyses to confirm key metrics for accurately distinguishing between these cryptic species. While there are slight differences in colouration and patterns including size and density of their defining black spots, four key metrics were found to be significantly different between the species including gill rakers counts, snout length, lateral line scale counts and body depth. Brachymystax tumensis was also typically heavier than B. lenok at the same length. These results were supported by genetic analyses which clustered the two species into separate clades and displayed unique haplotypes for the Mongolian populations. Initial investigations of diets and gonad maturation indicated an ecological and spatial separation of these sympatric species. The current study is important for these recreationally targeted species which require further research and species-specific management measures to ensure their threatened populations are conserved in Mongolia for the future.

We don’t know the half of it: morphological and molecular evidence reveal dramatic underestimation of diversity in a key pollinator group (Nemestrinidae)

Nemestrinidae (tangle-veined flies) are important pollinators of numerous southern African plant species. Despite their known ecological importance, the family has received little taxonomic attention in recent years and the systematics of the group is poorly understood. In this study we aimed to assess the phylogenetic relationships and species diversity among three southern African nemestrinid genera from the Nemestrininae subfamily: Prosoeca, Moegistorhynchus and Stenobasipteron, with a specific focus on the largest among these, Prosoeca. We reconstructed a molecular phylogeny using both mitochondrial and nuclear (COI, 16S rRNA, 28S rRNA and CAD) DNA sequence data. Both morphology and molecular species delimitation methods (Automatic Barcode Gap Discovery and the Bayesian Poisson Tree Process) were used to estimate species diversity. The topology from the combined analysis places a monophyletic Moegistorhynchus as the sister group to a paraphyletic Prosoeca with Stenobasipteron nested inside Prosoeca. In all three genera, almost half of the putative species sampled did not match the concept of described species based on morphology. Analysis of phylogenetic diversity showed that undescribed putative species make a substantial contribution to the overall phylogenetic diversity among the sampled species. Comparisons among biogeographic regions suggested that diversity is concentrated in multiple biodiversity hotspots and biomes, particularly in Fynbos and Grassland biomes. The numerous undescribed species and paraphyly of Prosoeca both emphasise the need for increased taxonomic attention for this ecologically important group of flies in particular, and for southern African insect taxa in general.