Mitochondrial Genome Sequence Research Articles

Complete mitochondrial genome and the phylogenetic position of a new Annelida species belonging to the genus Syllis

The family Syllidae is the most taxonomically complex of the phylum Annelida. Although the gene order in the phylum Annelida's mitochondrial DNA (mtDNA) is well conserved, several exceptional cases have been reported. In this study, we describe the mitochondrial genome of a Syllis sp. that is 17,092 bp in length and contains 13 mitochondrial protein-coding genes (PCGs), 23 transfer RNA (tRNA) genes, including two tRNA-M, two ribosomal RNA (rRNA) genes, and a putative control region between tRNA-W and tRNA-G distinguished by a single short noncoding region. However, the gene order is not similar to those of other species in the family Syllidae. Phylogenetic analyses of 18S rRNA and 13 PCGs sequences demonstrated that this worm was clustered with other Syllis species in the family Syllidae. This is the first study to reveal the complete mitochondrial genome sequence of a previously unidentified Syllis sp. improving our understanding of the molecular biological characteristics of the poorly known genus Syllis.

Phylogenetic Analysis of Russian Native Sheep Breeds Based on mtDNA Sequences.

Eurasia is represented by all climatic zones and various environments. A unique breed variety of farm animals has been developed in Russia, whose territory covers a large area of the continent. A total of 69 local breeds and types of dairy, wool, and meat sheep (Ovis aries) are maintained here. However, the genetic diversity and maternal origin of these local breeds have not been comprehensively investigated. In this study, we describe the diversity and phylogeny of Russian sheep breeds inhabiting different geographical regions based on the analysis of complete sequences of mitochondrial genomes (mtDNA). Complete mtDNA sequences of the studied sheep were obtained using next-generation sequencing technology (NGS). All investigated geographical groups of sheep were characterized by high haplotype (Hd = 0.9992) and nucleotide diversity (π = 0.00378). Analysis of the AMOVA results showed that genetic diversity was majorly determined by within-population differences (77.87%). We identified 128 haplotypes in all studied sheep. Haplotypes belonged to the following haplogroups: B (64.8%), A (28.9%), C (5.5%), and D (0.8%). Haplogroup B was predominant in the western part of Russia. A high level of mtDNA polymorphism in the studied groups of local sheep indicates the presence of a significant reserve of unique genotypes in Russia, which is to be explored.

Diagnostic Yield of Investigations in Symmetric Optic Neuropathy.

Symmetric optic neuropathy (SON) is commonly seen in neuro-ophthalmic practice and is often discovered incidentally. Although multiple investigations might be performed to discover the underlying cause, they are not always indicated. The aim of this study was to report a clinically reasonable and cost-effective approach to investigating patients with SON. SON was defined as bilateral optic neuropathy with normal and/or symmetrically decreased central visual acuity, absence of relative afferent pupillary defect, presence of symmetric optic disc pallor, symmetric thinning of peripapillary retinal nerve fiber layer on optical coherence tomography, and absence of other identifiable causes of optic neuropathy. Records of all patients diagnosed with SON seen at a tertiary university-affiliated neuro-ophthalmology practice from 2016 to 2022 were reviewed to identify the yield of various investigations. Clinical data from the initial and last follow-up visit were obtained. Subgroup analysis was performed to ascertain whether diagnostic yield is higher in patients with severe visual loss (central acuity worse than 20/40) compared with those with mild visual loss (acuity 20/40 or better). One hundred thirty-six patients met inclusion criteria. Testing for OPA1 and OPA2 mutations had the highest diagnostic yield (16.0%), followed by mitochondrial genome sequencing (13.6%), serum vitamin B12 (6.1%), and serum folate (1.6%). MRI brain was performed in 54.4% of patients and had a diagnostic yield of only 5%. Both patients who had abnormal MRI had symptoms of demyelination at presentation. Patients were followed for a mean of 15.0 (SD 21.3) months. The most frequently identified etiologies of SON were Leber hereditary optic neuropathy (8.1%), alcohol/tobacco amblyopia (7.4%), vitamin B12 deficiency (5.9%), and dominant optic atrophy (2.9%). Patients with severe visual impairment were more likely to have a final diagnosis compared with those with milder visual impairment (63.9% vs 12.0%, P < 0.001). The diagnostic yield of investigating SON in patients with preserved visual function, normal diet, and absence of other neurological symptoms is very low. It is reasonable to observe patients with SON with mild visual impairment, reserving costly investigations for those with the visual acuity worse than 20/40 or progressive course.

Mitochondrial Genome Sequencing and Analysis of the Invasive Microstegium vimineum: A Resource for Systematics, Invasion History, and Management

Next article No AccessMitochondrial genome sequencing and analysis of the invasive Microstegium vimineum: a resource for systematics, invasion history, and managementCraig F. Barrett, Dhanushya Ramachandran, Chih-Hui Chen, Cameron W. Corbett, Cynthia D. Huebner, Brandon T. Sinn, Wen-Bin Yu, and Kenji SuetsuguCraig F. Barrett Search for more articles by this author , Dhanushya Ramachandran Search for more articles by this author , Chih-Hui Chen Search for more articles by this author , Cameron W. Corbett Search for more articles by this author , Cynthia D. Huebner Search for more articles by this author , Brandon T. Sinn Search for more articles by this author , Wen-Bin Yu Search for more articles by this author , and Kenji Suetsugu Search for more articles by this author PDFPDF PLUS Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinkedInRedditEmail SectionsMoreDetailsFiguresReferencesCited by International Journal of Plant Sciences Just Accepted Article DOIhttps://doi.org/10.1086/726005 HistoryAccepted May 01, 2023 © 2023 The University of Chicago. All Rights reserved.PDF download Crossref reports no articles citing this article.

Complete mitochondrial genome of Agrostis stolonifera: insights into structure, Codon usage, repeats, and RNA editing.

Plants possess mitochondrial genomes that are large and complex compared to animals. Despite their size, plant mitochondrial genomes do not contain significantly more genes than their animal counterparts. Studies into the sequence and structure of plant mitochondrial genomes heavily imply that the main mechanism driving replication of plant mtDNA, and offer valuable insights into plant evolution, energy production, and environmental adaptation. This study presents the first comprehensive analysis of Agrostis stolonifera's mitochondrial genome, characterized by a branched structure comprising three contiguous chromosomes, totaling 560,800bp with a GC content of 44.07%. Annotations reveal 33 unique protein-coding genes (PCGs), 19 tRNA genes, and 3 rRNA genes. The predominant codons for alanine and glutamine are GCU and CAA, respectively, while cysteine and phenylalanine exhibit weaker codon usage biases. The mitogenome contains 73, 34, and 23 simple sequence repeats (SSRs) on chromosomes 1, 2, and 3, respectively. Chromosome 1 exhibits the most frequent A-repeat monomeric SSR, whereas chromosome 2 displays the most common U-repeat monomeric SSR. DNA transformation analysis identifies 48 homologous fragments between the mitogenome and chloroplast genome, representing 3.41% of the mitogenome's total length. The PREP suite detects 460C-U RNA editing events across 33 mitochondrial PCGs, with the highest count in the ccmFn gene and the lowest in the rps7 gene. Phylogenetic analysis confirms A. stolonifera's placement within the Pooideae subfamily, showing a close relationship to Lolium perenne, consistent with the APG IV classification system. Numerous homologous co-linear blocks are observed in A. stolonifera's mitogenomes and those of related species, while certain regions lack homology. The unique features and complexities of the A. stolonifera mitochondrial genome, along with its similarities and differences to related species, provide valuable insights into plant evolution, energy production, and environmental adaptation. The findings from this study significantly contribute to the growing body of knowledge on plant mitochondrial genomes and their role in plant biology.

Maternally inherited non-syndromic hearing loss is linked with a novel mitochondrial ND6 gene mutation.

Maternally inherited non-syndromic hearing loss is linked with mitochondrial DNA mutations. This investigation demonstrates the features of a Chinese pedigree suffering from maternally inherited non-syndromic hearing loss. Biochemical characterizations included the measurements ofprotein synthesis levels, membrane potential, and the synthesis of reactive oxygen species (ROS) and adenosine triphosphate (ATP) using cybrid cell lines derived from an affected matrilineal subject and control subject. Non-congenital early or late-onset/development hearing impairment has been observed in 4 of 9 in a family (matrilineal), with different degrees of hearing impairment, ranging from normal to severe. A pedigree's whole mitochondrial genome sequence analysis revealed the homoplasmic m.14502T > C (I58V) mutation at ND6's isoleucine location-58, and specific mitocchondrial DNA polymorphisms set haplogroups M10 were highly conserved. In vitro models indicated that m.14502T > C mutation-derived respiratory deficiency decreases ND6 protein synthesis, mitochondrial membrane potential, and ATP synthesis. These mitochondrial dysregulations enhance the generation of ROS in the mutant cells. Identifying nuclear modifiers is essential for elucidating hearing loss's pathogenesis and furnishing novel therapeutic interventions. The m.14502T > C mutation should be considered an inherited risk factor that can help diagnose. The data of this investigation help counsel families of individuals with hearing loss.

A new subgenus, Australixodes n. subgen. (Acari: Ixodidae), for the kiwi tick, Ixodes anatis Chilton, 1904, and validation of the subgenus Coxixodes Schulze, 1941 with a phylogeny of 16 of the 22 subgenera of Ixodes Latreille, 1795 from entire mitochondrial genome sequences.

A new subgenus, Australixodes n. subgen., is described for the kiwi tick, Ixodes anatis Chilton, 1904. The subgenus Coxixodes Schulze, 1941 is validated for the platypus tick, Ixodes (Coxixodes) ornithorhynchi Lucas, 1846, sister group of the subgenus Endopalpiger Schulze, 1935. A phylogeny from mitochondrial genomes of 16 of the 22 subgenera of Ixodes (32 spp.) indicates, for the first time, the relationships of the subgenera of Ixodes Latreille, 1795, the largest genus of ticks.

Genetic epidemiology of late blight in Australia using ancient DNA

Late blight caused by Phytophthora infestans is the most destructive disease of potatoes worldwide and is most notable as the cause of the Irish potato famine of the mid 1840’s. Whole mitochondrial genome sequences were generated from 44 Australian, two UK and one US isolates of P. infestans stored as either herbarium specimens or as recent samples on FTA cards, spanning the period 1873–2019. Mitochondrial (mt) genome sequence comparison confirmed that over the past c. 110 years, an old A1 strain of P. infestans has been present in Australia. There was evidence, however, that P. infestans had been introduced on multiple separate occasions in the early 1900s prior to the introduction of quarantine measures. The 44 Australian samples included six infected specimens of the Australian native kangaroo apple (Solanum aviculare) collected in 1911. The P. infestans mt genome sequences from these isolates clustered with P. infestans from infected potatoes collected in Victoria (1910, 1911, 1986, 1989), South Australia (1909, 2003) and Tasmania (1910, 2019), supporting the hypothesis that Australia has one old strain that has survived since the first arrival of the pathogen into the country. The study highlights the success of Australia’s potato biosecurity setting and supports the need for ongoing surveillance and biosecurity measures to prevent the introduction of the newer more aggressive strains of the pathogen.

Indian Red Jungle fowl reveals a genetic relationship with South East Asian Red Jungle fowl and Indian native chicken breeds as evidenced through whole mitochondrial genome sequences.

Background: Native chickens are dispersed in a wide geographical range and have hereditary assets that are kept by farmers for various purposes. Mitochondrial DNA (mtDNA) is a widely utilized marker in molecular studies because of its quick advancement, matrilineal legacy, and simple molecular structure. Method and Results: We performed NGS sequencing to investigate mitochondrial genomes and to evaluate the hereditary connections, diversity, and measure of gene stream estimation in Indian native chicken breeds and Red Jungle fowl. The chicken breeds were genotyped using the D-loop region and 23 haplotypes were identified. When compared to Indian native breeds, more haplotypes were identified in the NADH dehydrogenase subunits, Cytochrome c oxidase, Cytochrome b, ATP synthase subunit 6, and Ribosomal RNA genes. The phylogenetic examination indicated that the analyzed chicken breeds were divided into six significant clades, namely A, B, C, D, E, and F, of which the F clade indicated the domestication of chicken breeds in India. Additionally, our work affirmed that the Indian Red Jungle Fowl is the origin for both reference Red Jungle Fowl as well as all Indian breeds, which is reflected in the dendrogram as well as network analysis based on the whole mtDNA and D-loop region. Indian Red Jungle Fowl is distributed as an outgroup, suggesting that this ancestry was reciprocally monophyletic. Conclusion: The mtDNA sequences of Indian native chickens provided novel insights into adaptation mechanisms and the significance of important mtDNA variations in understanding the maternal lineages of native birds.

Assembly and analysis of the mitochondrial genome of Prunella vulgaris.

Prunella vulgaris (Lamiaceae) is widely distributed in Eurasia. Former studies have demonstrated that P. vulgaris has a wide range of pharmacological effects. Nevertheless, no complete P. vulgaris mitochondrial genome has been reported, which limits further understanding of the biology of P. vulgaris. Here, we assembled the first complete mitochondrial genome of P. vulgaris using a hybrid assembly strategy based on sequencing data from both Nanopore and Illumina platforms. Then, the mitochondrial genome of P. vulgaris was analyzed comprehensively in terms of gene content, codon preference, intercellular gene transfer, phylogeny, and RNA editing. The mitochondrial genome of P. vulgaris has two circular structures. It has a total length of 297, 777 bp, a GC content of 43.92%, and 29 unique protein-coding genes (PCGs). There are 76 simple sequence repeats (SSRs) in the mitochondrial genome, of which tetrameric accounts for a large percentage (43.4%). A comparative analysis between the mitochondrial and chloroplast genomes revealed that 36 homologous fragments exist in them, with a total length of 28, 895 bp. The phylogenetic analysis showed that P. vulgaris belongs to the Lamiales family Lamiaceae and P. vulgaris is closely related to Salvia miltiorrhiza. In addition, the mitochondrial genome sequences of seven species of Lamiaceae are unconservative in their alignments and undergo frequent genome reorganization. This work reports for the first time the complete mitochondrial genome of P. vulgaris, which provides useful genetic information for further Prunella studies.

Clinical and genetic analysis of essential hypertension with CYB gene m.15024G>A mutation.

To explore the role of mitochondrial CYB 15024G>A mutation in the development of essential hypertension. Mitochondrial genome sequences of hypertensive patients were obtained from previous studies. Clinical and genetic data of a hypertensive patient with mitochondrial CYB 15024G>A mutation and its pedigree were analyzed. Lymphocytes derived from patient and family members were transformed into immortalized lymphoblastoid cell lines, and the levels of adenosine triphosphate (ATP), mitochondrial membrane potential and intracellular reactive oxygen species (ROS) were detected. The penetrance of this essential hypertension family was 42.9%, and the age of onset was 46-68 years old. Mitochondrial genome sequencing results showed that all maternal members carried a highly conserved mitochondrial CYB 15024G>A mutation. This mutation could affect the free energy of mitochondrial CYB for secondary and tertiary structure and protein folding, thereby changing its structural stability and the structure of the electron transfer function area around the mutation site. Compared with the control, the cell line carrying the mitochondrial CYB 15024G>A mutation showed significantly decreased levels of mitochondrial CYB, ATP and mitochondrial membrane potential, and increased levels of ROS (P<0.01). Mitochondrial CYB 15024G>A mutation may affect the structure of respiratory chain subunits and mitochondrial function, leading to cell dysfunction, which suggests that the mutation may play a synergistic role in essential hypertension.

Genetic variation and population structure of brachiopods, Lingula anatina Lamarck, 1801 in the Northern Aceh shore, Indonesia

Abstract. Octavina C, Ramadhaniaty M, Daulay RE, Dewiyanti I, Ulfah M. 2023. Genetic variation and population structure of brachiopods, Lingula anatina Lamarck, 1801 in the Northern Aceh shore. Biodiversitas 24: 3951-3959. Lingula anatina is one of the brachiopods recognized as one of the most primitive groups with a fossil record from the early Cambrian period. Although L. anatina has been targeted as an object of biological studies, the information about the genetic variation within species is still limited. The purpose of this research is to identify the genetic variation and population structure of the species L. anatina found in the Aceh northern shore based on the mtDNA Cytochrome Oxidase I (COI) gene sequence. In this study, L. anatina specimens collected from five populations on Aceh's northern shore, namely Krueng Raya, Syiah Kuala, Ulee Lheue, and Ujong Pancu, with complete mitochondrial genome sequences were analyzed and compared with genebank sequences. The total nucleotide base of L. anatina is 657 bp, with 18 haplotypes. Based on the results of a comparison of the distribution of L. anatina in Asia shows that the Japanese population has the highest population structure value compared to the other five populations. The Fst value obtained from this study is 0.7, so it is classified as high and structured. Phylogenetic analysis showed that the correlation between geographic location and geographic distance affects the genetic distance between populations.

The complete mitochondrial genome and description of a new cryptic Brazilian species of Metopiellus Raffray (Coleoptera: Staphylinidae: Pselaphinae).

Metopiellus Raffray, 1908 is a genus of South American rove beetles typically found in tropical humid forests. Here we describe a new cryptic species from Eastern Amazon, in northern Brazil, Metopiellus crypticus Asenjo sp. nov., and its major morphologic diagnostic features, which were photographed and illustrated. In addition, we bring the complete mitochondrial genome sequence of M. crypticus sp. nov., and its position within the phylogenetic context of the family, including previously available mitogenomes of Staphylinidae species.

The First Complete Mitochondrial Genome of the Genus Pachycondyla (Formicidae, Ponerinae) and Insights into the Phylogeny of Ants.

Ants are the standout group among eusocial insects in terms of their exceptional species richness and ecological dominance. The phylogenetic relationships among the group remain elusive. Mitochondrial genome sequences, as a kind of molecular marker, have been widely utilized in the phylogenetic analysis of insects. However, the number of ant mitogenomes published is still very limited. In this study, we utilized next-generation sequencing to determine the complete mitogenome of Pachycondyla annamita (Formicidae, Ponerinae). This is the first mitogenome from the genus Pachycondyla. Two gene rearrangements were identified in the mitogenome, the transposition of trnQ and trnM and the transposition of trnV and rrnS. The secondary structures of tRNAs were predicted. The tRNA genes trnR and trnS1 lacked the dihydrouridine (DHU) arm, and the trnE lacked the TΨC (T) arm. Phylogenetic analyses of the mitochondrial protein-coding genes under maximum likelihood (ML) and Bayesian inference (BI) criteria resulted in conflicting hypotheses. BI analysis using amino acid data with the site-heterogeneous mixture model produced a tree topology congruent with previous studies. The Formicidae was subdivided into two main clades, namely the "poneroid" clade and the "formicoid" clade. A sister group relationship between Myrmicinae and Formicinae was recovered within the "formicoid" clade.

Clinical Reasoning: A 17-Year-Old Girl With Progressive Cognitive Impairment.

A 17-year-old girl presented with a long history of cognitive impairment, personality and behavioral changes, dysarthria, and paroxysmal lower-extremity weakness. She was initially suspected of having mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes because of stroke-like symptoms, such as episodic lower-extremity weakness, as well as abnormal brain MRI findings of generalized cerebral atrophy, extensive high-intensity lesions in the cortex and subcortical white matter on fluid-attenuated inversion recovery images, decreased N-acetyl aspartate/creatine ratio, and a lactate peak in the focal area on spectrum images. However, there were no relatives with similar presentations in the family of the patient. The whole mitochondrial genome and whole-exome sequencing did not suggest pathogenic mutations, and no abnormalities were found in the blood or CSF lactate levels. In this case, we detail the clinical manifestations, diagnostic workup, and imaging findings. This case highlights the importance of assessing cognitive function and the relevant differential diagnoses in an adolescent with cognitive impairment.

Mitochondrial genome sequence characteristics and phylogenetic analysis of Schizothorax argentatus

Schizothorax argentatus that only distributes in the Ili River basin in Xinjiang is one of the rare and endangered species of schizothorax in China, thus has high scientific and economic values. In this study, the complete mitochondrial genome sequence of S. argenteus with a length of 16 580 bp was obtained by high-throughput sequencing. The gene compositions and arrangement were similar to those of typical vertebrates. It contained 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and a non-coding region (D-loop). The nucleotide compositions were A (30.25%), G (17.28%), C (27.20%), and T (25.27%), respectively, showing obvious AT bias and anti-G bias. Among the tRNA genes, only tRNA-Ser(GCU) could not form a typical cloverleaf structure due to the lack of dihydrouracil arm. The AT-skew and GC-skew values of the ND6 gene were fluctuating the most, suggesting that the gene may experience different selection and mutation pressures from other genes. The mitochondrial control region of S. argenteus contained three different domains, i.e., termination sequence region (ETAS), central conserved region (CSB-F, CSB-E, CSB-D, and CSB-B), and conserved sequence region (CSB1, CSB2, and CSB3). The conserved sequence fragment TT (AT) nGTG, which was ubiquitous in Cypriniformes, was identified at about 50 bp downstream CSB3. Phylogenetic relationships based on the complete mitochondrial genome sequence of 28 Schizothorax species showed that S. argenteus had differentiated earlier and had a distant relationship with other species, which may be closely related to the geographical location and the hydrological environment where it lives.

Assessing Onchocerca volvulus Intensity of Infection and Genetic Diversity Using Mitochondrial Genome Sequencing of Single Microfilariae Obtained before and after Ivermectin Treatment.

Onchocerciasis is a neglected tropical disease targeted for elimination using ivermectin mass administration. Ivermectin kills the microfilariae and temporarily arrests microfilariae production by the macrofilariae. We genotyped 436 microfilariae from 10 people each in Ituri, Democratic Republic of the Congo (DRC), and Maridi County, South Sudan, collected before and 4-5 months after ivermectin treatment. Population genetic analyses identified 52 and 103 mitochondrial DNA haplotypes among the microfilariae from DRC and South Sudan, respectively, with few haplotypes shared between people. The percentage of genotype-based correct assignment to person within DRC was ~88% and within South Sudan ~64%. Rarefaction and extrapolation analysis showed that the genetic diversity in DRC, and even more so in South Sudan, was captured incompletely. The results indicate that the per-person adult worm burden is likely higher in South Sudan than DRC. Analyses of haplotype data from a subsample (n = 4) did not discriminate genetically between pre- and post-treatment microfilariae, confirming that post-treatment microfilariae are not the result of new infections. With appropriate sampling, mitochondrial haplotype analysis could help monitor changes in the number of macrofilariae in a population as a result of treatment, identify cases of potential treatment failure, and detect new infections as an indicator of continuing transmission.

Comparison of Mitochondrial Genome Sequences between Two Palaemon Species of the Family Palaemonidae (Decapoda: Caridea): Gene Rearrangement and Phylogenetic Implications.

To further understand the origin and evolution of Palaemonidae (Decapoda: Caridea), we determined the mitochondrial genome sequence of Palaemon macrodactylus and Palaemon tenuidactylus. The entire mitochondrial genome sequences of these two Palaemon species encompassed 37 typical genes, including 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (rRNAs), and 22 transfer RNA genes (tRNAs), and a control region (CR). The lengths of their mitochondrial genomes were 15,744 bp (P. macrodactylus) and 15,735 bp (P. tenuidactylus), respectively. We analyzed their genomic features and structural functions. In comparison with the ancestral Decapoda, these two newly sequenced Palaemon species exhibited a translocation event, where the gene order was trnK-trnD instead of trnD-trnK. Based on phylogenetic analysis constructed from 13 PCGs, the 12 families from Caridea can be divided into four major clades. Furthermore, it was revealed that Alpheidae and Palaemonidae formed sister groups, supporting the monophyly of various families within Caridea. These findings highlight the significant gene rearrangements within Palaemonidae and provide valuable evidence for the phylogenetic relationships within Caridea.

Mitochondrial DNA sequencing of Kehilan and Hamdani horses from Saudi Arabia

The Arabian horse breed is well known for its purity and played a key role in the genetic improvement of other horses worldwide. The mitochondrial genome plays a vital role in maternal inheritance and it’s helpful to evaluate its genetic diversity and conservation. It has higher mutation rates than nuclear DNA in vertebrates and therefore reveals phylogenetic relationships and haplotypes. In this study, the mitochondrial genome mutations in two Saudi horse strains, Kehilan and Hamdani demonstrated various changes in the gene and amino acid levels and included two other Saudi horses (Hadban and Seglawi) from the previous study for phylogenetic comparison. The whole mitochondrial genome sequencing resulted in intra and inter mtDNA variations between the studied horses. Interestingly, the Hamdani horse has nucleotide substitutions similar to those of the Hadban horse, which is reflected in the phylogenetic tree as a significantly close relationship. This type of study provides a better understanding of mitogenome structure and conservation of livestock species genetic data.

An advanced approach for rapid visual identification of Liposcelis bostrychophila (Psocoptera: Liposcelididae) based on CRISPR/Cas12a combined with RPA.

Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae) is a booklouse pest that is a threat to commodity storage security worldwide. Accurate and sensitive methods of L. bostrychophila on-site identification are essential prerequisites for its effective management. Evidence suggests that L. bostrychophila contains 3 intraspecific biotypes that are morphologically indistinguishable but can be discriminated at the level of mitochondrial genome organization and sequences. The traditional molecular identification methods, such as DNA barcoding and PCR-RFLP, are instrumentally demanding and time-consuming, limiting the application of the identification in the field. Therefore, this study developed a new CRISPR/Cas12a-based visual nucleic acid system based on the mitochondrial gene coding for NADH dehydrogenase subunit 2 (nad2), combined with recombinase polymerase amplification (RPA) to accurately identify L. bostrychophila from 4 other common stored-product booklice, and also differentiate 3 biotypes of this species at the same time. The entire identification process could be completed at 37 °C within 20 min with high sensitivity. The system could stably detect at least 1 ng/μl of DNA template. The green fluorescence signal produced by the trans-cleaving of the single-stranded DNA reporter could be observed by the naked eye under blue light. Additionally, the suggested system combined with the crude DNA extraction method to extract DNA rapidly, enabled identification of all developmental stages of L. bostrychophila. With crude DNA, this novel diagnostic system successfully identified an unknown booklouse by holding the reaction tubes in the hand, thus can be considered as an accurate, rapid, highly sensitive, and instrument-flexible method for on-site visual identification of L. bostrychophila.