New faunistic data on Diptera (Hexapoda, Insecta) from the Ziarat Juniperus forest ecosystem (Pakistan).

Background: This study presents the first faunistic record and DNA barcoding for some Diptera species recorded from the Juniperus forest ecosystem of Balochistan, Pakistan. DNA barcoding was used to explore species diversity of Dipterans and collections carried out using a Malaise trap between December 2018 to December 2019. This process involved sequencing the 658 bp Cytochrome Oxidase I (COI) gene. New information: Amongst the collected Diptera specimens, nine families were identified, representing 13 genera. These species include Atherigonasoccata (Rondani, 1871), Atherigonavaria (Schiner, 1868), Chironomusdorsalis (Meigen, 1818), Eupeodescorollae (Linnaeus, 1758), Eristalistenax (Linnaeus,1758), Goniaornata (Meigen, 1826), Luciliasericata (Meigen, 1826), Paragusquadrifasciatus (Linnaeus, 1758), Polleniarudis (Fabricius, 1794), Raviniapernix (Thompson, 1869), Sarcophagadux (Thompson, 1869), Trupaneaamoena (Schiner, 1868) and Wohlfahrtiabella (Linnaeus, 1758). The families Syrphidae and Sarcophagidae exhibited the highest representation, each comprising three genera and three species. They were followed by the family Muscidae, which had a single genus and two species. Anthomyiidae, Chironomidae, Calliphoridae, Polleniidae, Tachinidae and Tephritidae were represented by only one genus and one species. A nique Barcode Index Number (BIN) was allotted to Tachinidae (specie i.e Goniaornata). The results indicated that barcoding through cytochrome oxidase I is an effective approach for the accurate identification and genetic studies of Diptera species. This discovery highlights the significant diversity of this insect order in study region. Furthermore, a comprehensive list of other Diptera species remains elusive because of difficulties in distinguishing them, based on morphology and a lack of professional entomological knowledge.

Initial study and phylogenetic analysis of hard ticks (Acari: Ixodidae) in Nantong, China along the route of avian migration.

The growing concern about migratory birds potentially spreading ticks due to global warming has become a significant issue. The city of Nantong in this study is situated along the East Asia-Australasian Flyway (EAAF), with numerous wetlands serving as roosting sites for migratory birds. We conducted an investigation of hard ticks and determined the phylogenetic characteristics of tick species in this city. We utilized three different genes for our study: the mitochondrial cytochrome oxidase subunit 1 (COX1) gene, the second internal transcribed spacer (ITS2), and the mitochondrial small subunit rRNA (12 S rRNA) gene. The predominant tick species were Haemaphysalis flava (H. flava) and Haemaphysalis longicornis (H. longicornis). Additionally, specimens of Haemaphysalis campanulata (H. campanulata) and Rhipicephalus sanguineus (R. sanguineus) were collected. The H. flava specimens in this study showed a close genetic relationship with those from inland provinces of China, as well as South Korea and Japan. Furthermore, samples of H. longicornis exhibited a close genetic relationship with those from South Korea, Japan, Australia, and the USA, as well as specific provinces in China. Furthermore, R. sanguineus specimens captured in Nantong showed genetic similarities with specimens from Egypt, Nigeria, and Argentina.

Optimizing Nucleic Acid Delivery Systems through Barcode Technology.

Conventional biological experiments often focus on in vitro assays because of the inherent limitations when handling multiple variables in vivo, including labor-intensive and time-consuming procedures. Often only a subset of samples demonstrating significant efficacy in the in vitro assays can be evaluated in vivo. Nonetheless, because of the low correlation between the in vitro and in vivo tests, evaluation of the variables under examination in vivo and not solely in vitro is critical. An emerging approach to achieve high-throughput in vivo tests involves using a barcode system consisting of various nucleotide combinations. Unique barcodes for each variant enable the simultaneous testing of multiple entities, eliminating the need for separate individual tests. Subsequently, to identify crucial parameters, samples were collected and analyzed using barcode sequencing. This review explores the development of barcode design and its applications, including the evaluation of nucleic acid delivery systems and the optimization of gene expression in vivo.

Integrated DNA barcoding methods to identify species in the processed fish products from Chinese market.

DNA-based methods are reliable for a precise identification of species in processed products. In this study, we assessed five typical DNA extraction methods from multiple aspects. Full-length and mini-length DNA barcoding were performed to detect the species substitution and mislabeling of 305 processed fish products from the Chinese market covering six processed fish products. The salt extraction method that exhibited the best overall performance was applied. All samples were successfully extracted; however, only 19.3 % of samples could be amplified using the full-DNA barcode primer set, and 90.2 % of samples could be amplified using the newly designed mini-DNA barcode primer sets (401 and 320 bp). Overall, the molecular identification results revealed that 36.4 % (111/305) of the samples were inconsistent with the labels, with commercial fraud observed in all six types of processed fish products. The survey findings provide technical references for effective fish authentication monitoring, offering insights into the seafood safety in markets.

The InBIO Barcoding Initiative Database: DNA barcodes of Iberian Bees.

Background: Bees are important actors in terrestrial ecosystems and are recognised for their prominent role as pollinators. In the Iberian Peninsula, approximately 1,100 bee species are known, with nearly 100 of these species being endemic to the Peninsula. A reference collection of DNA barcodes, based on morphologically identified bee specimens, representing 514 Iberian species, was constructed. The "InBIO Barcoding Initiative Database: DNA Barcodes of Iberian bees" dataset contains records of 1,059 sequenced specimens. The species of this dataset correspond to about 47% of Iberian bee species diversity and 21% of endemic species diversity. For peninsular Portugal only, the corresponding coverage is 71% and 50%. Specimens were collected between 2014 and 2022 and are deposited in the research collection of Thomas Wood (Naturalis Biodiversity Center, The Netherlands), in the FLOWer Lab collection at the University of Coimbra (Portugal), in the Andreia Penado collection at the Natural History and Science Museum of the University of Porto (MHNC-UP) (Portugal) and in the InBIO Barcoding Initiative (IBI) reference collection (Vairão, Portugal). New information: Of the 514 species sequenced, 75 species from five different families are new additions to the Barcode of Life Data System (BOLD) and 112 new BINs were added. Whilst the majority of species were assigned to a single BIN (94.9%), 27 nominal species were assigned to multiple BINs. Although the placement into multiple BINs may simply reflect genetic diversity and variation, it likely also represents currently unrecognised species-level diversity across diverse taxa, such as Amegillaalbigena Lepeletier, 1841, Andrenarussula Lepeletier, 1841, Lasioglossumleucozonium (Schrank, 1781), Nomadafemoralis Morawitz, 1869 and Sphecodesalternatus Smith, 1853. Further species pairs of Colletes, Hylaeus and Nomada were placed into the same BINs, emphasising the need for integrative taxonomy within Iberia and across the Mediterranean Basin more broadly. These data substantially contribute to our understanding of bee genetic diversity and DNA barcodes in Iberia and provide an important baseline for ongoing taxonomic revisions in the West Palaearctic biogeographical region.

Molecular characterization and demographic insights into soybean bud borer (Lepidoptera: Tortricidae) in Brazil.

The soybean bud borer, a soybean pest in Brazil, was initially identified as Crocidosema aporema (Walsingham 1914) (Lepidoptera: Tortricidae). Outbreaks of this species have recently increased, but identification of this pest remains uncertain, and the historical factors associated with its geographic distribution in Brazil are little known. Here, we conducted a species characterization and phylogeographic analysis based on molecular and morphological evidence. Ninety individuals of bud-borers Lepidoptera were collected in different regions of Brazil. We sequenced COI and COII mitochondrial genes and examined wing patterns and male genital morphology. DNA barcoding approach revealed that 10 individuals were Argyrotaenia sphaleropa (Meyrick 1909) (Lepidoptera: Tortricidae) and 80 were a species of the genus Crocidosema Zeller. The morphology of the adult genitalia and wings proved to be insufficient to confirm the identification of Brazilian individuals as C. aporema, a species originally described from a high-elevation site in Costa Rica. Furthermore, the genetic distance between putative C. aporema specimens from Brazil and Costa Rica (ranging from 5.2% to 6.4%) supports the hypothesis that the Brazilian specimens are not referable to C. aporema. Our analysis revealed a single genetic strain (i.e., species) with low genetic diversity on soybean crops. We found no indication that the genetic structure was related to geographic distance among populations or edaphoclimatic regions. The population expansion of the soybean bud borer coincides with the increase in the area of soybean production in Brazil, suggesting that expanded soybean farming has allowed a significant increase in the effective population size of this pest.

Conservation Strategies for Aquilaria sinensis: Insights from DNA Barcoding and ISSR Markers.

The evergreen tree species Aquilaria sinensis holds significant economic importance due to its specific medicinal values and increasing market demand. However, the unrestricted illegal exploitation of its wild population poses a threat to its survival. This study aims to contribute to the conservation efforts of A. sinensis by constructing a library database of DNA barcodes, including two chloroplast genes (psbA-trnH and matK) and two nuclear genes (ITS and ITS2). Additionally, the genetic diversity and structure were estimated using inter-simple sequence repeats (ISSR) markers. Four barcodes of 57 collections gained 194 sequences, and 1371 polymorphic bands (98.63%) were observed using DNA ISSR fingerprinting. The Nei's gene diversity (H) of A. sinensis at the species level is 0.2132, while the Shannon information index (I) is 0.3128. The analysis of molecular variance revealed a large significant proportion of total genetic variations and differentiation among populations (Gst = 0.4219), despite a relatively gene flow (Nm = 0.6853) among populations, which were divided into two groups by cluster analysis. There was a close genetic relationship among populations with distances of 0.0845 to 0.5555. This study provides evidence of the efficacy and dependability of establishing a DNA barcode database and using ISSR markers to assess the extent of genetic diversity A. sinensis. Preserving the genetic resources through the conservation of existing populations offers a valuable proposition. The effective utilization of these resources will be further deliberated in subsequent breeding endeavors, with the potential to breed agarwood commercial lines.

Diversity, Distribution and Host Blood Meal Analysis of Adult Black Flies (Diptera: Simuliidae) from Thailand.

Understanding the factors associated with the species diversity and distribution of insect vectors is critically important for disease epidemiology. Black flies (Diptera: Simuliidae) are significant hematophagous insects, as many species are pests and vectors that transmit pathogens to humans and other animals. Ecological factors associated with black fly species distribution have been extensively examined for the immature stages but are far less well explored for the adult stage. In this study, we collected a total of 7706 adult black fly specimens from various locations in forests, villages and animal shelters in Thailand. The integration of morphology and DNA barcoding revealed 16 black fly taxa, including Simulium yvonneae, a species first found in Vietnam, which is a new record for Thailand. The most abundant species was the Simulium asakoae complex (n = 5739, 74%), followed by S. chumpornense Takaoka and Kuvangkadilok (n = 1232, 16%). The Simulium asakoae complex was dominant in forest (3786 of 4456; 85%) and village (1774 of 2077; 85%) habitats, while S. chumpornense predominated (857 of 1175; 73%) in animal shelter areas. The Simulium asakoae complex and S. nigrogilvum Summers, which are significant pests and vectors in Thailand, occurred at a wide range of elevations, although the latter species was found mainly in high (>1000 m) mountain areas. Simulium chumpornense, S. nodosum Puri and the S. siamense Takaoka and Suzuki complex occurred predominately in low (<800 m)-elevation areas. Simulium furvum Takaoka and Srisuka; S. phurueaense Tangkawanit, Wongpakam and Pramual; and S. nr. phurueaense were only found in high (>1000 m) mountain areas. A host blood meal analysis revealed that the S. asakoae; S. chamlongi Takaoka and Suzuki; S. nigrogilvum; S. chumpornense; and the S. striatum species group were biting humans. This is the first report of the latter two species biting humans. We also found that S. chumpornense was biting turkeys, and S. chamlongi was biting chickens, which are new host blood sources recorded for these species. In addition, we found that the S. feuerborni Edwards complex was biting water buffalo, which is the first report on the biting habits of this species.

DNA barcoding of the genus Gampsocleis (Orthoptera, Tettigoniidae) from China.

DNA barcoding is a useful addition to the traditional morphology-based taxonomy. A ca. 650 bp fragment of the 5' end of mitochondrial cytochrome c oxidase subunit I (hereafter COI-5P) DNA barcoding was sued as a practical tool for Gampsocleis species identification. DNA barcodes from 889 specimens belonging to 8 putative Gampsocleis species was analyzed, including 687 newly generated DNA barcodes. These barcode sequences were clustered/grouped into Operational Taxonomic Units (OTUs) using the criteria of five algorithms, namely Barcode Index Number (BIN) System, Assemble Species by Automatic Partitioning (ASAP), a Java program uses an explicit, determinate algorithm to define Molecular Operational Taxonomic Unit (jMOTU), Generalized Mixed Yule Coalescent (GMYC), and Bayesian implementation of the Poisson Tree Processes model (bPTP). The Taxon ID Tree grouped sequences of morphospecies and almost all MOTUs in distinct nonoverlapping clusters. Both long- and short-winged Gampsocleis species are reciprocally monophyletic in the Taxon ID Tree. In BOLD, 889 barcode sequences are assigned to 17 BINs. The algorithms ASAP, jMOTU, bPTP and GMYC clustered the barcode sequences into 6, 13, 10, and 23 MOTUs, respectively. BIN, ASAP, and bPTP algorithm placed three long-winged species, G. sedakovii, G. sinensis and G. ussuriensis within the same MOTU. All species delimitation algorithms split two short-winged species,G. fletcheri and G. gratiosa into at least two MOTUs each, except for ASAP algorithm. More detailed molecular and morphological integrative studies are required to clarify the status of these MOTUs in the future.

Mitochondrial genome sequence comparisons indicate that the elephant louse Haematomyzus elephantis (Piaget, 1869) contains cryptic species.

The parvorder Rhynchopthirina contains three currently recognised species of lice that parasitize elephants (both African savanna elephant Loxodonta africana and Asian elephant Elephas maximus), desert warthogs (Phacochoerus aethiopicus) and Red River hogs (Potamochoerus porcus), respectively. The Asian elephant lice and the African savanna elephant lice are currently treated as the same species, Haematomyzus elephantis (Piaget, 1869), based on morphology despite the fact that their hosts diverged 8.4 million years ago. In the current study, we sequenced 23 mitochondrial (mt) genes of African savanna elephant lice collected in South Africa and analysed the sequence divergence between African savanna elephant lice and previously sequenced Asian elephant lice. Sequence comparisons revealed >23% divergence for the 23 mt genes as a whole and ~17% divergence for cox1 gene between African savanna and Asian elephant lice, which were far higher than the divergence expected within a species. Furthermore, the mt gene sequence divergences between these lice are 3.76-4.6 times higher than that between their hosts, the African savanna and Asian elephants, which are expected for the co-divergence and co-evolution between lice and their elephant hosts. We conclude that (1) H. elephantis (Piaget, 1869) contains cryptic species and (2) African savanna and Asian elephant lice are different species genetically that may have co-diverged and co-evolved with their hosts.

Construction of multiplexed transcriptome NGS libraries of microdissected tissue samples based on combinational DNA barcode microbeads.

The combination of single-cell RNA sequencing and microdissection techniques that preserves positional information has become a major tool for spatial transcriptome analyses. However, high costs and time requirements, especially for experiments at the single cell scale, make it challenging for this approach to meet the demand for increased throughput. Therefore, we proposed combinational DNA barcode (CDB)-seq as a medium-throughput, multiplexed approach combining Smart-3SEQ and CDB magnetic microbeads for transcriptome analyses of microdissected tissue samples. We conducted a comprehensive comparison of conditions for CDB microbead preparation and related factors and then applied CDB-seq to RNA extracts, fresh frozen (FF) and formalin-fixed paraffin-embedded (FFPE) mouse brain tissue samples. CDB-seq transcriptomic profiles of tens of microdissected samples could be obtained in a simple, cost-effective way, providing a promising method for future spatial transcriptomics.

Bemisia tabaci MEAM1 still remains the dominant species in open field crops in Brazil / Bemisia tabaci MEAM1 ainda é a espécie dominante em cultivos a céu aberto no Brasil

Among Bemisia tabaci species, the invasive MEAM1 and MED species are key agricultural pests for many crops. In Brazil, most part of B. tabaci population outbreaks were associated with MEAM1, which, since 1990s quickly spread across the entire country. Later in 2014, the MED was identified in Brazil, initially more restricted to greenhouses, but suddenly reaching new areas in the South and Southeast open regions. Thus, our objective was to investigate the geographical distribution of MEAM1 and MED on open field crops in Brazil. MEAM1 is still the predominant species on open field crops such as soybean, cotton, and tomato. The sequencing of a cytochrome c oxidase subunit I (COI) gene fragment revealed a single haplotype of MEAM1, suggesting the establishment of a single MEAM1 strain in the country. The haplotypes found for MEAM1 and MED are genetically related to the globally dispersed strains, Jap1 and Mch1, respectively. Continuous monitoring of B. tabaci species is crucial because landscape alterations, climatic changes, and pest management methods may shift the B. tabaci species distribution and dominance in Brazilian crop areas.

Dentre as espécies de Bemisia tabaci, as espécies invasoras MEAM1 e MED se destacam como pragas de grande importância para várias culturas. No Brasil, a maior parte dos surtos populacionais de mosca-branca são associados a presença da espécie MEAM1, que a partir 1990 se espalhou por todo o país. Por outro lado, em 2014 a espécie MED foi identificada no Brasil, inicialmente restrita a casas de vegetação, mas rapidamente se difundindo em novas áreas nas regiões Sul e Sudeste do Brasil. Assim, nosso objetivo foi investigar a distribuição geográfica das espécies MEAM1 e MED em grandes culturas no Brasil. A espécie MEAM1 continua sendo predominante nas monoculturas como algodão, soja e tomate. O sequenciamento de um fragmento do gene citocromo c oxidase subunidade I (COI) revelou a presença de um haplótipo para MEAM1, sugerindo o estabelecimento de apenas uma linhagem no país. Os haplótipos encontrados para MEAM1 e MED são geneticamente relacionados as linhagens globalmente dispersas Jap1 e Mch1, respectivamente. O monitoramento contínuo das espécies de B. tabaci é crucial pois as mudanças na paisagem, mudanças climáticas e métodos de manejo das pragas podem alterar a dominância e a distribuição dessas espécies nas áreas agrícolas do Brasil.

Bemisia tabaci MEAM1 still remains the dominant species in open field crops in Brazil

Abstract Among Bemisia tabaci species, the invasive MEAM1 and MED species are key agricultural pests for many crops. In Brazil, most part of B. tabaci population outbreaks were associated with MEAM1, which, since 1990s quickly spread across the entire country. Later in 2014, the MED was identified in Brazil, initially more restricted to greenhouses, but suddenly reaching new areas in the South and Southeast open regions. Thus, our objective was to investigate the geographical distribution of MEAM1 and MED on open field crops in Brazil. MEAM1 is still the predominant species on open field crops such as soybean, cotton, and tomato. The sequencing of a cytochrome c oxidase subunit I (COI) gene fragment revealed a single haplotype of MEAM1, suggesting the establishment of a single MEAM1 strain in the country. The haplotypes found for MEAM1 and MED are genetically related to the globally dispersed strains, Jap1 and Mch1, respectively. Continuous monitoring of B. tabaci species is crucial because landscape alterations, climatic changes, and pest management methods may shift the B. tabaci species distribution and dominance in Brazilian crop areas.

Resumo Dentre as espécies de Bemisia tabaci, as espécies invasoras MEAM1 e MED se destacam como pragas de grande importância para várias culturas. No Brasil, a maior parte dos surtos populacionais de mosca-branca são associados a presença da espécie MEAM1, que a partir 1990 se espalhou por todo o país. Por outro lado, em 2014 a espécie MED foi identificada no Brasil, inicialmente restrita a casas de vegetação, mas rapidamente se difundindo em novas áreas nas regiões Sul e Sudeste do Brasil. Assim, nosso objetivo foi investigar a distribuição geográfica das espécies MEAM1 e MED em grandes culturas no Brasil. A espécie MEAM1 continua sendo predominante nas monoculturas como algodão, soja e tomate. O sequenciamento de um fragmento do gene citocromo c oxidase subunidade I (COI) revelou a presença de um haplótipo para MEAM1, sugerindo o estabelecimento de apenas uma linhagem no país. Os haplótipos encontrados para MEAM1 e MED são geneticamente relacionados as linhagens globalmente dispersas Jap1 e Mch1, respectivamente. O monitoramento contínuo das espécies de B. tabaci é crucial pois as mudanças na paisagem, mudanças climáticas e métodos de manejo das pragas podem alterar a dominância e a distribuição dessas espécies nas áreas agrícolas do Brasil.

Specific DNA barcodes screening, germplasm resource identification, and genetic diversity analysis of Platycodon grandiflorum / 药学学报

Platycodonis Radix is the dry root of Platycodon grandiflorum of Campanulaceae, which has a variety of pharmacological effects and is a commonly used bulk Chinese medicine. In this study, the chloroplast genome sequences of six P. grandiflorum from different producing areas has been sequenced with Illumina HiSeq X Ten platform. The specific DNA barcodes were screened, and the germplasm resources and genetic diversity were analyzed according to the specific barcodes. The total length of the chloroplast genome of 6 P. grandiflorum samples was 172 260-172 275 bp, and all chloroplast genomes showed a typical circular tetrad structure and encoded 141 genes. The comparative genomics analysis and results of amplification efficiency demonstrated that trnG-UCC and ndhG_ndhF were the potential specific DNA barcodes for identification the germplasm resources of P. grandiflorum. A total of 305 P. grandiflorum samples were collected from 15 production areas in 9 provinces, for which the fragments of trnG-UCC and ndhG_ndhF were amplificated and the sequences were analyzed. The results showed that trnG-UCC and ndhG_ndhF have 5 and 11 mutation sites, respectively, and 5 and 7 haplotypes were identified, respectively. The combined analysis of the two sequences formed 13 haplotypes (named Hap1-Hap13), and Hap4 is the main genotype, followed by Hap1. The unique haplotypes possessed by the three producing areas can be used as DNA molecular tags in this area to distinguish from the germplasm resources of P. grandiflorum from other areas. The haplotype diversity, nucleotide diversity and genetic distance were 0.94, 4.79×10-3 and 0.000 0-0.020 3, respectively, suggesting that the genetic diversity was abundant and intraspecific kinship was relatively close. This study laid a foundation for the identification of P. grandiflorum, the protection and utilization of germplasm resources, and molecular breeding.

Identifying Antigenic Switching by Clonal Cell Barcoding and Nanopore Sequencing in Trypanosoma brucei.

Many organisms alternate the expression of genes from large gene sets or gene families to adapt to environmental cues or immune pressure. The single-celled protozoan pathogen Trypanosoma brucei spp. periodically changes its homogeneous surface coat of variant surface glycoproteins (VSGs) to evade host antibodies during infection. This pathogen expresses one out of ~2,500 VSG genes at a time from telomeric expression sites (ESs) and periodically changes their expression by transcriptional switching or recombination. Attempts to track VSG switching have previously relied on genetic modifications of ES sequences with drug-selectable markers or genes encoding fluorescent proteins. However, genetic modifications of the ESs can interfere with the binding of proteins that control VSG transcription and/or recombination, thus affecting VSG expression and switching. Other approaches include Illumina sequencing of the VSG repertoire, which shows VSGs expressed in the population rather than cell switching; the Illumina short reads often limit the distinction of the large set of VSG genes. Here, we describe a methodology to study antigenic switching without modifications of the ES sequences. Our protocol enables the detection of VSG switching at nucleotide resolution using multiplexed clonal cell barcoding to track cells and nanopore sequencing to identify cell-specific VSG expression. We also developed a computational pipeline that takes DNA sequences and outputs VSGs expressed by cell clones. This protocol can be adapted to study clonal cell expression of large gene families in prokaryotes or eukaryotes. Key features • This protocol enables the analysis of variant surface glycoproteins (VSG) switching in T. brucei without modifying the expression site sequences. • It uses a streamlined computational pipeline that takes fastq DNA sequences and outputs expressed VSG genes by each parasite clone. • The protocol leverages the long reads sequencing capacity of the Oxford nanopore sequencing technology, which enables accurate identification of the expressed VSGs. • The protocol requires approximately eight to nine days to complete.

DNA Barcode library of the endemic-rich avifauna of the oceanic islands of the Gulf of Guinea.

Background: The BioSTP: DNA Barcoding of endemic birds from oceanic islands of the Gulf of Guinea dataset contains records of 155 bird specimens belonging to 56 species in 23 families, representing over 80% of the diversity of the breeding landbird community. All specimens were collected on Príncipe, São Tomé and Annobón Islands between 2002 and 2021 and morphologically identified to species or subspecies level by qualified ornithologists. The dataset includes all endemic species and 3/4 of the extant endemic subspecies of the islands. This dataset is the second release by BioSTP and it greatly increases the knowledge on the DNA barcodes of Gulf of Guinea birds. All DNA extractions are deposited at Associação BIOPOLIS - CIBIO, Research Center in Biodiversity and Genetic Resources. New information: The dataset includes DNA barcodes for all 29 endemic bird species and for 11 of the 15 extant endemic bird subspecies from the oceanic islands of the Gulf of Guinea. This is the first major DNA barcode set of African birds. The three endemic subspecies of Crithagrarufobrunnea, an island endemic with three allopatric populations within the Archipelago, are also represented. Additionally, we obtained DNA barcodes for 16 of the 21 non-endemic landbirds and for one vagrant (Sylviacommunis). In total, forty-one taxa were new additions to the Barcode of Life Data System (BOLD), with another 11 corresponding to under-represented taxa in BOLD. Furthermore, the submitted sequences were found to cluster in 55 Barcode Index Numbers (BINs), 37 of which were new to BOLD. All specimens have their DNA barcodes publicly accessible through BOLD online database and GenBank.

FishDNAIDs: DNA barcoding as a tool in the development and validation in silico and in vitro of detection systems to four species of Characiformes of commercial importance in the Brazilian Amazon.

BACKGROUND: The COI mitochondrial gene has been chosen as the "DNA barcode in animals" and the large quantity of genetic information in public databanks gives solid support for the use of DNA barcoding as a promising tool for the development of a specific molecular detection system. METHODS AND RESULTS: The present study aimed to develop a Specific Molecular Detection System (SMDS: FishDNAIDs) (primers and probe sets) for the following four target species: Prochilodus nigricans, Potamorhina altamazonica, Psectrogaster rutiloides and Triportheus angulatus, in qPCR assays. In silico and in vitro tests (using gDNA) were performed to test these sets. The database generated contained the cytochrome c oxidase subunit I (COI) nucleotide sequence for 183 specimens of Characiformes, distributed in 34 species representing eight families. In silico, primers designed for the target species amplified different species from the same genus, except for P. rutiloides, which amplified only the target species. In the in vitro test, using the SYBRGreentm and TaqMan® fluorescence systems, both sets detected the respective target species (P. nigricans, P. altamazonica, P. rutiloides and T. angulatus). In the qPCR assays using SYBRGreentm, species considered to be related were also detected, in addition to the target species, with the exception of P. amazonica and P. essequibensis (correlated to P. rutiloides). All target species were detected in the qPCR assays using the TaqMan® system; however, with the SMDS PALT, the target species P. altamazonica was detected with low CT values (22.21 ± 0.17) as well as the correlates of P. latior and P. pristigaster, though with high CT values (23.51 ± 0.19 and 30.21 ± 0.95). This assay uniquely identifies known adult tissue samples from all four species. CONCLUSIONS: The primers and probe sets developed can act as powerful tools for detecting the target Characiformes species.

Diversity and Distribution of Mites (ACARI) Revealed by Contamination Survey in Public Genomic Databases.

Acari (mites and ticks) are a biodiverse group of microarthropods within the Arachnida. Because of their diminutive size, mites are often overlooked. We hypothesized that mites, like other closely related microorganisms, could also contaminate public genomic database. Here, using a strategy based on DNA barcodes previously reported, we scanned contaminations related to mites (Acari, exclusive of Ixodida) in Genbank WGS/TSA database. In 22,114 assemblies (17,845 animal and 4269 plant projects), 1717 contigs in 681 assemblies (3.1%) were detected as mite contaminations. Additional taxonomic analysis showed the following: (1) most of the contaminants (1445/1717) were from the specimens of Magnoliopsida, Insecta and Pinopsida; (2) the contamination rates were higher in plant or TSA projects; (3) mite distribution among different classes of hosts varied considerably. Additional phylogenetic analysis of these contaminated contigs further revealed complicated mite-host associations. Overall, we conducted a first systemic survey and analysis of mite contaminations in public genomic database, and these DNA barcode related mite contigs will provide a valuable resource of information for understanding the diversity and phylogeny of mites.

Ensemble learning-based approach for automatic classification of termite mushrooms.

Termite mushrooms are edible fungi that provide significant economic, nutritional, and medicinal value. However, identifying these mushroom species based on morphology and traditional knowledge is ineffective due to their short development time and seasonal nature. This study proposes a novel method for classifying termite mushroom species. The method utilizes Gradient Boosting machine learning techniques and sequence encoding on the Internal Transcribed Spacer (ITS) gene dataset to construct a machine learning model for identifying termite mushroom species. The model is trained using ITS sequences obtained from the National Center for Biotechnology Information (NCBI) and the Barcode of Life Data Systems (BOLD). Ensemble learning techniques are applied to classify termite mushroom species. The proposed model achieves good results on the test dataset, with an accuracy of 0.91 and an average AUCROC value of 0.99. To validate the model, eight ITS sequences collected from termite mushroom samples in An Linh commune, Phu Giao district, Binh Duong province, Vietnam were used as the test data. The results show consistent species identification with predictions from the NCBI BLAST software. The results of species identification were consistent with the NCBI BLAST prediction software. This machine-learning model shows promise as an automatic solution for classifying termite mushroom species. It can help researchers better understand the local growth of these termite mushrooms and develop conservation plans for this rare and valuable plant resource.

Assessing sequence heterogeneity in Chlorellaceae DNA barcode markers for phylogenetic inference.

Phylogenetic inference is an important approach that allows the recovery of the evolutionary history and the origin of the Chlorellaceae species. Despite the species' potential for biofuel feedstock production, their high phenotypic plasticity and similar morphological structures among the species have muddled the taxonomy and identification of the Chlorellaceae species. This study aimed to decipher Chlorellaceae DNA barcode marker heterogeneity by examining the sequence divergence and genomic properties of 18S rRNA, ITS (ITS1-5.8S rRNA-ITS2-28S rRNA), and rbcL from 655 orthologous sequences of 64 species across 31 genera in the Chlorellaceae family. The study assessed the distinct evolutionary properties of the DNA markers that may have caused the discordance between individual trees in the phylogenetic inference using the Robinson-Foulds distance and the Shimodaira-Hasegawa test. Our findings suggest that using the supermatrix approach improves the congruency between trees by reducing stochastic error and increasing the confidence of the inferred Chlorellaceae phylogenetic tree. This study also found that the phylogenies inferred through the supermatrix approach might not always be well supported by all markers. The study highlights that assessing sequence heterogeneity prior to the phylogenetic inference could allow the approach to accommodate sequence evolutionary properties and support species identification from the most congruent phylogeny, which can better represent the evolution of Chlorellaceae species.