Mitochondrial rRNA Gene Research Articles

Quantifying constraint in the human mitochondrial genome.

Mitochondrial DNA (mtDNA) has an important yet often overlooked role in health and disease. Constraint models quantify the removal of deleterious variation from the population by selection and represent powerful tools for identifying genetic variation that underlies human phenotypes1-4. However, nuclear constraint models are not applicable to mtDNA, owing to its distinct features. Here we describe the development of a mitochondrial genome constraint model and its application to the Genome Aggregation Database (gnomAD), a large-scale population dataset that reports mtDNA variation across 56,434 human participants5. Specifically, we analyse constraint by comparing the observed variation in gnomAD to that expected under neutrality, which was calculated using a mtDNA mutational model and observed maximum heteroplasmy-level data. Our results highlight strong depletion of expected variation, which suggests that many deleterious mtDNA variants remain undetected. To aid their discovery, we compute constraint metrics for every mitochondrial protein, tRNA and rRNA gene, which revealed a range of intolerance to variation. We further characterize the most constrained regions within genes through regional constraint and identify the most constrained sites within the entire mitochondrial genome through local constraint, which showed enrichment of pathogenic variation. Constraint also clustered in three-dimensional structures, which provided insight into functionally important domains and their disease relevance. Notably, we identify constraint at often overlooked sites, including in rRNA and noncoding regions. Last, we demonstrate that these metrics can improve the discovery of deleterious variation that underlies rare and common phenotypes.

Diversity, divergence time, and biogeography of the genus Albatrellus (Agaricomycetes, Russulales)

ABSTRACT The genus Albatrellus is an important group of stipitate terrestrial fungi in the order Russulales. Some species in the genus form ectomycorrhizae, mostly with trees of Pinaceae; some are well-known edible mushrooms. However, its diversity and biogeography are unclear. Taxonomic and phylogenetic studies on Albatrellus were carried out by morphological examination, which included detailed observations of the fruiting body, spore shape and size, and other key features, together with potential hosts. These observations were then compared and analysed using multi-locus molecular phylogenetic analyses, including the internal transcribed spacer regions (ITS), the large subunit nuclear ribosomal RNA gene (nLSU), the translation elongation factor 1-α gene (tef1), the largest subunit of RNA polymerase II (rpb1), the second largest subunit of RNA polymerase II (rpb2), the small subunit mitochondrial rRNA gene (mtSSU), and the small subunit of the nuclear ribosomal RNA gene (nucSSU). The results demonstrated that the species of Albatrellus formed eight clades. Nine new species are described and illustrated, and two new combinations are proposed. A total of 38 species are accepted in Albatrellus worldwide. Of those species, 26, 7, and 8 species are distributed in Asia, Europe, and North America, respectively. The divergence time indicated that the maximum crown age of Albatrellus was approximately 70.5 million years ago, and East Asia and North America are the likely ancestral areas. Dispersal and differentiation to other continents occurred during the late Paleocene and Miocene. Three kinds of dispersal routes are proposed: East Asia and Europe, East Asia and North America, and Europe and North America.

Four new species and a ribosomal phylogeny of Rhabdopleura (Hemichordata: Graptolithina) from New Zealand, with a review and key to all described extant taxa.

All eight extant species ofRhabdopleuradescribed between 1869 and 2018 are provisionally accepted as valid based on a review of the literature and new data on two little-known species from the Azores. Additionally, four new species are described from the New Zealand region, increasing global diversity by 50%, and a dichotomous key to all 12 described species is provided based on morphological criteria. The distinction between colony morphologies based on erect-tube inception is regarded as particularly helpful in initial characterization of species. Erect ringed tubes are either produced directly from the surface of creeping-tubes or indirectly, i.e. a short adherent side branch from a creeping tube is interpolated between the creeping tube and an erect tube; such side branches are blind-ending. These two modes of erect-tube origination are here respectively termeddirectandindirect. Species with indirect erect-tube budding are predominant in the North Atlantic whereas species with direct erect-tube budding dominate in New Zealand waters. The only indirect-erect species from New Zealand, Rhabdopleura chathamica n. sp., was discovered on deepwater coral from 10081075 m, constituting the deepest record of the genus to date. Rhabdopleura emancipata n. sp., collected only in a detached state, constitutes a three-dimensional tangled growth that grew freely into the water columna unique morphology hitherto unknown among extant species. Owing to this growth mode, it provided a substratum for epibionts from several phyla. Rhabdopleura francesca n. sp. and Rhabdopleura decipula n. sp. are morphologically very similar but are distinguishable by their distinct placements in a phylogeny based on 16S mitochondrial and 18S nuclear rRNA genes. Phylogenetic reconstructions based on rRNA and mitochondrial genome data contribute to an updated phylogeny of all Rhabdopleura species sequenced thus far, some of which require more molecular sequences and morphological analyses for taxonomic determination.

Mitochondrial and ribosomal markers in the identification of nematodes of clinical and veterinary importance

BackgroundNematodes of the Ascarididae, Ancylostomatidae and Onchocercidae families are parasites of human and veterinary importance causing infections with high prevalence worldwide. Molecular tools have significantly improved the diagnosis of these helminthiases, but the selection of genetic markers for PCR or metabarcoding purposes is often challenging because of the resolution these may show.MethodsNuclear 18S rRNA, internal transcribed spacers 1 (ITS-1) and 2 (ITS-2), mitochondrial gene cytochrome oxidase 1 (cox1) and mitochondrial rRNA genes 12S and 16S loci were studied for 30 species of the mentioned families. Accordingly, their phylogenetic interspecies resolution, pairwise nucleotide p-distances and sequence availability in GenBank were analyzed.ResultsThe 18S rRNA showed the least interspecies resolution since separate species of the Ascaris, Mansonella, Toxocara or Ancylostoma genus were intermixed in phylogenetic trees as opposed to the ITS-1, ITS-2, cox1, 12S and 16S loci. Moreover, pairwise nucleotide p-distances were significantly different in the 18S compared to the other loci, with an average of 99.1 ± 0.1%, 99.8 ± 0.1% and 98.8 ± 0.9% for the Ascarididae, Ancylostomatidae and Onchocercidae families, respectively. However, ITS-1 and ITS-2 average pairwise nucleotide p-distances in the three families ranged from 72.7% to 87.3%, and the cox1, 12S and 16S ranged from 86.4% to 90.4%. Additionally, 2491 cox1 sequences were retrieved from the 30 analyzed species in GenBank, whereas 212, 1082, 994, 428 and 143 sequences could be obtained from the 18S, ITS-1, ITS-2, 12S and 16S markers, respectively.ConclusionsThe use of the cox1 gene is recommended because of the high interspecies resolution and the large number of sequences available in databases. Importantly, confirmation of the identity of an unknown specimen should always be complemented with the careful morphological examination of worms and the analysis of other markers used for specific parasitic groups.Graphical

Molecular characterization of the camel nasal botfly, Cephalopina titillator (Diptera: Oestridae)

BackgroundLarval stages of the camel nasal botfly, Cephalopina titillator (Diptera: Oestridae), rank among the top obligate endoparasites causing nasopharyngeal myiasis. Such infestations have adverse effects on camel health, reducing their economic value. The current study’s objective is to assess the applicability of PCR using partial regions of the COI mtDNA and 28S rRNA gene sequences to identify, classify, and reveal the phylogenesis of C. titillator.ResultsLarvae of the 2nd and 3rd instars of C. titillator were collected from the El-Bassatin abattoir in Egypt, extracted from the nasopharyngeal passage of the Arabian camel (Camelus dromedarius) during postmortem inspection. Genomic DNA was successfully isolated from 10 samples of 3rd instar larvae of C. titillator and amplified using primer pairs targeting partial mitochondrial COI (916 bp) and ribosomal 28S rRNA (830 bp) gene fragments. Nucleotide sequences from five samples have been sequenced and submitted to GenBank under accession numbers OP482168 to OP482172 for the COI gene and OP482160 to OP482164 for the 28S rRNA gene fragments. The COI gene exhibited 97.3% nucleotide identity across all specimens, while the 28S rRNA gene displayed 99.74% identical nucleotides. Maximum likelihood trees were constructed based on the generalized time-reversible (GTR) model. The resulting COI phylogenetic tree demonstrated that the subfamily Oestrinae does not exhibit monophyly. Additionally, it revealed that C. titillator is a sister group to the subfamily Gasterophilinae. Despite the scarcity of data available for the 28S rRNA gene, the phylogenetic analysis utilizing 28S rRNA revealed one distinct lineage for the Egyptian camel nasal bot fly.ConclusionsMolecular phylogenetic analysis was conducted using molecular markers of distinct origins (both mitochondrial and nuclear) to elucidate the evolutionary relationships within the family Oestridae. This analysis is particularly significant following the inclusion of C. titillator, a first-time discovery in Egypt.

Molecular Analysis of Dihydropteroate Synthase Gene Mutations in Pneumocystis jirovecii Isolates among Bulgarian Patients with Pneumocystis Pneumonia

Pneumocystis jirovecii pneumonia (PCP) is a significant cause of morbidity and mortality in immunocompromised people. The widespread use of trimethoprim-sulfamethoxazole (TMP-SMZ) for the treatment and prophylaxis of opportunistic infections (including PCP) has led to an increased selection of TMP-SMZ-resistant microorganisms. Sulfa/sulfone resistance has been demonstrated to result from specific point mutations in the DHPS gene. This study aims to investigate the presence of DHPS gene mutations among P. jirovecii isolates from Bulgarian patients with PCP. A total of 326 patients were examined via real-time PCR targeting the P. jirovecii mitochondrial large subunit rRNA gene and further at the DHPS locus. P. jirovecii DNA was detected in 50 (15.34%) specimens. A 370 bp DHPS locus fragment was successfully amplified in 21 samples from 19 PCP-positive patients, which was then purified, sequenced, and used for phylogenetic analysis. Based on the sequencing analysis, all (n = 21) P. jirovecii isolates showed DHPS genotype 1 (the wild type, with the nucleotide sequence ACA CGG CCT at codons 55, 56, and 57, respectively). In conclusion, infections caused by P. jirovecii mutants potentially resistant to sulfonamides are still rare events in Bulgaria. DHPS genotype 1 at codons 55 and 57 is the predominant P. jirovecii strain in the country.

New species of the genus Trichosetodes Ulmer, 1915 (Trichoptera, Leptoceridae) from Ratanakiri province, Cambodia, based on morphological and molecular data.

Three new species of Trichosetodes, namely T.carmelaesp. nov., T.katiengensissp. nov. and T.ratanakiriensissp. nov. are described and illustrated by male specimens. The male genitalia of T.carmelaesp. nov. can be distinguished from the other 16 species of the genus found in Southeast Asia by the shape of the phallicata. The phallicata of T.carmelaesp. nov. bears a tuft of long hairs in the middle of the dorsal edge. Trichosetodeskatiengensissp. nov. can be distinguished from the other species in Southeast Asia by the shape of the phallicata which is divided into dorsal and ventral branches in lateral view, and T.ratanakiriensissp. nov. by the characters of the left inferior appendage and the shape of segment IX. The posterior end of the left inferior appendage of T.ratanakiriensissp. nov. is not forked and the ventral and lateral views of the posteroventral lobes of segment IX are rounded. Illustrations of male genitalia of Trichosetodeskampongspeuensis Malicky & Kong, 2020 are provided for comparison. The molecular diversity of new Trichosetodes species was analyzed using the mitochondrial large subunit ribosomal rRNA gene region (16S rRNA). In terms of their genetic divergence, T.ratanakiriensissp. nov. and T.kampongspeuensis exhibited remarkable proximity, with only a 1.4% distance. On the contrary, T.carmelaesp. nov. displayed genetic disparity exceeding 6.3% when compared to both T.ratanakiriensissp. nov. and T.kampongspeuensis.

Replacing mechanical protection with colorful faces-twice: parallel evolution of the non-operculate marine worm-snail genera Thylacodes (Guettard, 1770) and Cayo n. gen. (Gastropoda: Vermetidae).

Vermetid worm-snails are sessile and irregularly coiled marine mollusks common in warmer nearshore and coral reef environments that are subject to high predation pressures by fish. Often cryptic, some have evolved sturdy shells or long columellar muscles allowing quick withdrawal into better protected parts of the shell tube, and most have variously developed opercula that protect and seal the shell aperture trapdoor-like. Members of Thylacodes (previously: Serpulorbis) lack such opercular protection. Its species often show polychromatic head-foot coloration, and some have aposematic coloration likely directed at fish predators. A new polychromatic species, Thylacodes bermudensis n. sp., is described from Bermuda and compared morphologically and by DNA barcode markers to the likewise polychromatic western Atlantic species T. decussatus (Gmelin, 1791). Operculum loss, previously assumed to be an autapomorphy of Thylacodes, is shown to have occurred convergently in a second clade of the family, for which a new genus Cayo n. gen. and four new western Atlantic species are introduced: C. margarita n. sp. (type species; with type locality in the Florida Keys), C. galbinus n. sp., C. refulgens n. sp., and C. brunneimaculatus n. sp. (the last three with type locality in the Belizean reef) (all new taxa authored by Bieler, Collins, Golding & Rawlings). Cayo n. gen. differs from Thylacodes in morphology (e.g., a protoconch that is wider than tall), behavior (including deep shell entrenchment into the substratum), reproductive biology (fewer egg capsules and eggs per female; an obliquely attached egg capsule stalk), and in some species, a luminous, "neon-like", head-foot coloration. Comparative investigation of the eusperm and parasperm ultrastructure also revealed differences, with a laterally flattened eusperm acrosome observed in two species of Cayo n. gen. and a spiral keel on the eusperm nucleus in one, the latter feature currently unique within the family. A molecular phylogenetic analysis based on mitochondrial and nuclear rRNA gene sequences (12SrRNA, trnV, 16SrRNA, 28SrRNA) strongly supports the independent evolution of the two non-operculate lineages of vermetids. Thylacodes forms a sister grouping to a clade comprising Petaloconchus, Eualetes, and Cupolaconcha, whereas Cayo n. gen is strongly allied with the small-operculate species Vermetus triquetrus and V. bieleri. COI barcode markers provide support for the species-level status of the new taxa. Aspects of predator avoidance/deterrence are discussed for these non-operculate vermetids, which appear to involve warning coloration, aggressive behavior when approached by fish, and deployment of mucous feeding nets that have been shown, for one vermetid in a prior study, to contain bioactive metabolites avoided by fish. As such, non-operculate vermetids show characteristics similar to nudibranch slugs for which the evolution of warning coloration and chemical defenses has been explored previously.

Structural analysis of mitochondrial rRNA gene variants identified in patients with deafness.

The last few years have witnessed dramatic advances in our understanding of the structure and function of the mammalian mito-ribosome. At the same time, the first attempts to elucidate the effects of mito-ribosomal fidelity (decoding accuracy) in disease have been made. Hence, the time is right to push an important frontier in our understanding of mitochondrial genetics, that is, the elucidation of the phenotypic effects of mtDNA variants affecting the functioning of the mito-ribosome. Here, we have assessed the structural and functional role of 93 mitochondrial (mt-) rRNA variants thought to be associated with deafness, including those located at non-conserved positions. Our analysis has used the structural description of the human mito-ribosome of the highest quality currently available, together with a new understanding of the phenotypic manifestation of mito-ribosomal-associated variants. Basically, any base change capable of inducing a fidelity phenotype may be considered non-silent. Under this light, out of 92 previously reported mt-rRNA variants thought to be associated with deafness, we found that 49 were potentially non-silent. We also dismissed a large number of reportedly pathogenic mtDNA variants, 41, as polymorphisms. These results drastically update our view on the implication of the primary sequence of mt-rRNA in the etiology of deafness and mitochondrial disease in general. Our data sheds much-needed light on the question of how mt-rRNA variants located at non-conserved positions may lead to mitochondrial disease and, most notably, provide evidence of the effect of haplotype context in the manifestation of some mt-rRNA variants.

Mt-LAF3 is a pseudouridine synthase ortholog required for mitochondrial rRNA and mRNA gene expression in Trypanosoma brucei

Trypanosoma brucei and related kinetoplastid parasites possess unique RNA processing pathways, including in their mitochondria, that regulate metabolism and development. Altering RNA composition or conformation through nucleotide modifications is one such pathway, and modifications including pseudouridine regulate RNA fate and function in many organisms. We surveyed pseudouridine synthase (PUS) orthologs in trypanosomatids, with a particular interest in mitochondrial enzymes due to their potential importance for mitochondrial function and metabolism. Trypanosoma brucei mitochondrial (mt)-LAF3 is an ortholog of human and yeast mitochondrial PUS enzymes, and a mitoribosome assembly factor, but structural studies differ in their conclusion as to whether it has PUS catalytic activity. Here, we generated T. brucei cells that are conditionally null (CN) for mt-LAF3 expression and showed that mt-LAF3 loss is lethal and disrupts mitochondrial membrane potential (ΔΨm). Addition of a mutant gamma ATP synthase allele to the CN cells permitted ΔΨm maintenance and cell survival, allowing us to assess primary effects on mitochondrial RNAs. As expected, these studies showed that loss of mt-LAF3 dramatically decreases levels of mitochondrial 12S and 9S rRNAs. Notably, we also observed decreases in mitochondrial mRNA levels, including differential effects on edited vs. pre-edited mRNAs, indicating that mt-LAF3 is required for mitochondrial rRNA and mRNA processing, including of edited transcripts. To assess the importance of PUS catalytic activity in mt-LAF3 we mutated a conserved aspartate that is necessary for catalysis in other PUS enzymes and showed it is not essential for cell growth, or maintenance of ΔΨm and mitochondrial RNA levels. Together, these results indicate that mt-LAF3 is required for normal expression of mitochondrial mRNAs in addition to rRNAs, but that PUS catalytic activity is not required for these functions. Instead, our work, combined with previous structural studies, suggests that T. brucei mt-LAF3 acts as a mitochondrial RNA-stabilizing scaffold.

Molecular phylogeny and morphology reveal four new species of Thelephora (Thelephorales, Basidiomycota) from subtropical China, closely related to T. ganbajun.

The genus of Thelephora is a group of cosmopolitan ectomycorrhizal fungi with basidiocarps of morphological diversity that has an extremely scarce species reported from the forest ecosystem in China. In this study, phylogenetic analyses of Thelephora species from subtropical China were carried out based on multiple loci including the internal transcribed spacer (ITS) regions, the large subunit of nuclear ribosomal RNA gene (nLSU), and the small subunit of mitochondrial rRNA gene (mtSSU). Maximum likelihood and Bayesian analyses were used to construct the phylogenetic tree. The phylogenetic positions of four new species, Th. aquila, Th. glaucoflora, Th. nebula, and Th. pseudoganbajun, were revealed based on morphological and molecular evidence. Molecular analyses demonstrated that the four new species were closely related to Th. ganbajun and formed a clade with robust support in the phylogenetic tree. Regarding morphology, they share some common morphological characteristics, including flabelliform to imbricate pilei, generative hyphae more or less covered by crystals, and subglobose to irregularly lobed basidiospores (5-8 × 4-7 μm) with tuberculate ornamentation. These new species are described and illustrated and are compared to similar morphological or phylogenetically related species. A key to the new and allied species from China is provided.

Multi-Gene Phylogeny and Taxonomy of the Wood-Rotting Fungal Genus Phlebia sensu lato (Polyporales, Basidiomycota).

Phlebia s.l. (Polyporales, Basidiomycota) accommodates numerous species of wood-inhabiting fungi within the phylum Basidiomycota. The present study employs the morphological and phylogenetic approaches to revise the generic and species classification of Phlebia s.l. and surveys the species diversity. The phylogenetic analyses were performed using multiple gene regions viz. the internal transcribed spacer (ITS), the large subunit nuclear ribosomal RNA gene (nLSU), the translation elongation factor 1-α (tef1), the small subunit of mitochondrial rRNA gene (mtSSU), the glyceraldehyde 3-phosphate dehydrogenase (GAPDH), RNA polymerase II largest subunit (rpb1), and RNA polymerase II second largest subunit (rpb2). We overall recognize twenty genera of Phlebia s.l., including three new genera viz. Ceriporiopsoides, Phlebicolorata, and Pseudophlebia, seven new species viz. Crustodontia rhododendri, Hydnophlebia fissurata, Luteoporia straminea, Merulius sinensis, Mycoaciella brunneospina, Phlebia niveomarginata, and P. poroides and seventeen new combinations viz. Ceriporiopsoides guidella, C. lagerheimii, Hydnophlebia acanthocystis, H. capsica, H. fimbriata, Merulius fuscotuberculatus, M. nantahaliensis, M. tomentopileatus, Mycoacia tuberculata, Mycoaciella uda, Phlebicolorata alboaurantia, Ph. brevispora, Ph. pseudoplacenta, Ph. rosea, Pseudophlebia lindtneri, Ps. semisupina, and Ps. setulosa. Descriptions, illustrations, phylogenetic trees to show the placements, and notes of new taxa are provided.

An Updated Phylogenetic Assessment and Taxonomic Revision of Perenniporia sensu lato (Polyporales, Basidiomycota).

Perenniporia is an important genus of Polyporaceae. In its common acceptation, however, the genus is polyphyletic. In this study, phylogenetic analyses on a set of Perenniporia species and related genera were carried out using DNA sequences of multiple loci, including the internal transcribed spacer (ITS) regions, the large subunit nuclear ribosomal RNA gene (nLSU), the small subunit mitochondrial rRNA gene (mtSSU), the translation elongation factor 1-α gene (TEF1) and the b-tubulin gene (TBB1). Based on morphology and phylogeny, 15 new genera, viz., Aurantioporia, Citrinoporia, Cystidioporia, Dendroporia, Luteoperenniporia, Macroporia, Macrosporia, Minoporus, Neoporia, Niveoporia, Rhizoperenniporia, Tropicoporia, Truncatoporia, Vanderbyliella, and Xanthoperenniporia, are proposed; 2 new species, Luteoperenniporia australiensis and Niveoporia subrusseimarginata, are described; and 37 new combinations are proposed. Illustrated descriptions of the new species are provided. Identification keys to Perenniporia and its related genera and keys to the species of these genera are provided.

Rickettsia spp. in Ticks of South Luangwa Valley, Eastern Province, Zambia

Ticks are important vectors for Rickettsia spp. belonging to the Spotted Fever Group responsible for causing Rickettsiosis worldwide. Rickettsioses pose an underestimated health risk to tourists and local inhabitants. There is evidence of the presence of Rickettsia spp. in Zambia, however there is limited data. A total of 1465 ticks were collected in 20 different locations from dogs and cattle including one cat. Ticks were identified by morphological features or by sequencing of the 16S mitochondrial rRNA gene. Individual ticks were further tested for rickettsiae using a pan-Rickettsia real-time-PCR. Rickettsia species in PCR-positive ticks were identified by sequencing the 23S-5S intergenic spacer region or partial ompA gene, respectively. Seven tick species belonging to three different tick genera were found, namely: Amblyomma variegatum, Rhipicephalus appendiculatus, Rhipicephalus (Boophilus) microplus, Rhipicephalus simus, Rhipicephalus sanguineus, Rhipicephalus zambesiensis and Haemaphysalis elliptica. Out of the 1465 ticks collected, 67 (4.6%) tested positive in the pan-Rickettsia PCR. This study provides detailed data about the presence of Rickettsia species in South Luangwa Valley, Eastern Province, Zambia for the first time. High prevalence of Rickettsia africae in Amblyomma variegatum was found, which indicates the potential risk of infection in the investigated area. Furthermore, to our best knowledge, this is the first time Rickettsia massiliae, a human pathogen causing spotted fever, has been detected in Zambia.

Phylogenetic placement and molecular dating of hammerhead sharks (Sphyrnidae) based on whole mitogenomes

ABSTRACT Due to continued overexploitation and anthropogenic change, hammerhead sharks (Carcharhiniformes: Sphyrnidae) have experienced drastic declines over most of their geographic range. Owing to the K-selected life histories of these sharks, their population resilience and persistence, remain severely strained, further compromising ecosystem stability. Moreover, some species are largely understudied e.g. the cryptic congener, the Carolina hammerhead shark (Sphyrna gilberti), whilst specific regions, such as the South-West Indian Ocean (SWIO), remain relatively devoid of data, risking the eventual extirpation of unique hammerhead shark lineages. The aim of the present study was to verify the phylogenetic placement of the cryptic S. gilberti within the family Sphyrnidae through the inclusion of underrepresented species sequences in order to provide a more comprehensive phylogenetic perspective for understanding historical drivers of Sphyrnidae biodiversity. The present study describes the first complete mitochondrial genome of the cryptic S. gilberti originating from the US Atlantic, as well as the mitogenomes of smooth hammerhead shark (Sphyrna zygaena) and scalloped hammerhead shark (Sphyrna lewini) samples originating from the data deficient South-West Indian Ocean (SWIO). Furthermore, we estimate the phylogenetic interrelationships of the Sphyrnidae family using mitochondrial protein-coding (PCG) and rRNA genes, reaffirming the placement of S. gilberti as a sister lineage to S. lewini. The resulting phylogenetic estimate is further used to evaluate the most likely age of the first occurrence of S. gilberti, corresponding to the Late Miocene to Early Pleiocene Epoch (3.8–10.8 million years ago). Comparative analysis of these Sphyrnids between ocean basins, as well as preliminary divergence estimates for S. lewini and S. gilberti has contributed towards resolving the global hammerhead phylogeny. This has provided unique insights into the evolution of the genus, thereby aiding future efforts directed towards effective conservation and management of hammerhead populations over a larger spatial scale.

Characterization of Colletotrichum Species as Causal Agents of Anthracnose in Strawberry Productive Areas in Tucumán, Argentina

Objective: Anthracnose is a disease caused by Colletotrichum species and is a significant disease in the strawberry crop. This study aims to identify at the level of species Colletotrichum spp. isolates collected from symptomatic strawberry plants in Tucumán, Argentina's second most important strawberry production area. Methods: 45 isolates of Colletotrichum were collected and characterized. Morphological characterization was conducted by analyzing fungal cultural characteristics, the growth rate in potato dextrose agar plates, conidial morphology and size, and sexual state. Phytopathological characterization was carried out by plant-pathogen interactions under controlled conditions in infection chambers. Molecular characterization was performed by polymerase chain reaction analysis of regions of amplified mitochondrial small rRNA genes. Results: Preliminary morphological characterization led to three distinctive groups, based on conidial shape and size, and colony type, colour, and growth, whereas the phytopathological characterization, led to two distinct groups based on the severity of symptoms, virulence, and host specificity. Molecular analyses confirmed the identity of three isolates representing each of the microbiological groups (i.e., isolate F7 of Colletotrichum acutatum (C. acutatum), isolate L9 of C. acutatum, isolate M11 of C. acutatum), demonstrating that they correspond to C. acutatum species. Based on the analysis of ribosomal DNA internal transcribed spacers sequences, and using specific microsatellites, we found a high genetic variability among all C. acutatum sequences. Implications of these results, as well as biological products commercially available and research advancing the field of biological control agents, were also discussed. Conclusion: This is the latest study to report with confidence that the species of Colletotrichum present in Tucumán, Argentina corresponds to C. acutatum, and such species are genetically diverse.

Sensitive and accurate DNA metabarcoding of parasitic helminth mock communities using the mitochondrial rRNA genes

Next-generation sequencing technologies have accelerated the pace of helminth DNA metabarcoding research, enabling species detection in bulk community samples. However, finding suitable genetic markers with robust species-level resolution and primers targeting a broad species range among parasitic helminths are some of the challenges faced. This study aimed to demonstrate the potential use of the mitochondrial 12S and 16S rRNA genes for parasitic helminth (nematodes, trematodes, cestodes) DNA metabarcoding. To demonstrate the robustness of the 12S and 16S rRNA genes for DNA metabarcoding, we determined the proportion of species successfully recovered using mock helminth communities without environment matrix and mock helminth communities artificially spiked with environmental matrices. The environmental matrices are human fecal material, garden soil, tissue, and pond water. Our results revealed the robustness of the mitochondrial rRNA genes, through the high sensitivity of the 12S rRNA gene, and the effectiveness of the 12S and 16S primers targeting platyhelminths. With the mitochondrial rRNA genes, a broad range of parasitc helminths were successfully detected to the species level. The potential of the mitochondrial rRNA genes for helminth DNA metabarcoding was demonstrated, providing a valuable gateway for future helminth DNA metabarcoding applications like helminth detection and biodiversity studies.

Nuclear and Mitochondrial SSU rRNA Genes Reveal Hidden Diversity of Haptophrya Endosymbionts in Freshwater Planarians and Challenge Their Traditional Classification in Astomatia.

Like many other aquatic animals, freshwater planarians have also become partners of symbiotic ciliates from the class Oligohymenophorea. In the present study, we explored the hidden diversity and addressed the questionable systematic position of mouthless obligatory gut endosymbionts of freshwater planarians, using the nuclear and mitochondrial SSU rRNA genes. Although all isolated ciliates morphologically corresponded to a single species, molecular analyses suggested the existence of three genetically distinct entities: Haptophrya planariarum, Haptophrya dugesiarum nov. spec., and Haptophrya schmidtearum nov. spec. The two former species share the same planarian host, which indicates a speciation model involving one duplication event without host switching. Such a diversification pattern was recognized also in astome ciliates inhabiting megascolecid and glossoscolecid earthworms. The present multi-gene phylogenies along with the secondary structure of the mitochondrial 16S rRNA molecule, however, challenge the traditional classification of Haptophrya within the subclass Astomatia. Haptophrya very likely evolved from an orphan scuticociliate lineage by the loss of oral apparatus and by the transformation of the thigmotactic field into an adhesive sucker. Since astomy evolved multiple times independently within the Oligohymenophorea, the loss of cell mouth cannot be used as a sole argument for the assignment of Haptophrya to the Astomatia anymore.

Species Diversity, Molecular Phylogeny, and Ecological Habits of Fomitopsis (Polyporales, Basidiomycota).

Fomitopsis is a worldwide brown-rot fungal genus of Polyporales, which grows on different gymnosperm and angiosperm trees and has important ecological functions and economic values. In this study, species diversity, phylogenetic relationships, and ecological habits of Fomitopsis were investigated. A total of 195 specimens from 24 countries representing 29 species of Fomitopsis were studied. Based on the morphological characters and phylogenetic evidence of DNA sequences including the internal transcribed spacer (ITS) regions, the large subunit of nuclear ribosomal RNA gene (nLSU), the small subunit of nuclear ribosomal RNA gene (nSSU), the small subunit of mitochondrial rRNA gene (mtSSU), the translation elongation factor 1-α gene (TEF), and the second subunit of RNA polymerase II (RPB2), 30 species are accepted in Fomitopsis, including four new species: F. resupinata, F. srilankensis, F. submeliae and F. yimengensis. Illustrated descriptions of the novel species and the geographical locations of the Fomitopsis species are provided.

Diversification of the phytophagous lineages of true bugs (Insecta: Hemiptera: Heteroptera) shortly after that of the flowering plants.

More than 95% of phytophagous true bug (Hemiptera: Heteroptera) species belong to four superfamilies: Miroidea (Cimicomorpha), Pentatomoidea, Coreoidea, and Lygaeoidea (all Pentatomomorpha). These iconic groups of highly diverse, overwhelmingly phytophagous insects include several economically prominent agricultural and silvicultural pest species, though their evolutionary history has not yet been well resolved. In particular, superfamily- and family-level phylogenetic relationships of these four lineages have remained controversial, and the divergence times of some crucial nodes for phytophagous true bugs have hitherto been little known, which hampers a better understanding of the evolutionary processes and patterns of phytophagous insects. In the present study, we used 150 species and concatenated nuclear and mitochondrial protein-coding genes and rRNA genes to infer the phylogenetic relationships within the Terheteroptera (Cimicomorpha + Pentatomomorpha) and estimated their divergence times. Our results support the monophyly of Cimicomorpha, Pentatomomorpha, Miroidea, Pentatomoidea, Pyrrhocoroidea, Coreoidea, and Lygaeoidea. The phylogenetic relationships across phytophagous lineages are largely congruent at deep nodes across the analyses based on different datasets and tree-reconstructing methods with just a few exceptions. Estimated divergence times and ancestral state reconstructions for feeding habit indicate that phytophagous true bugs explosively radiated in the Early Cretaceous-shortly after the angiosperm radiation-with the subsequent diversification of the most speciose clades (Mirinae, Pentatomidae, Coreinae, and Rhyparochromidae) in the Late Cretaceous.