Tooth intentional replantation from 1964 to 2023: A bibliometric analysis.

BACKGROUND/AIM: There is no thorough overview of intentional tooth replantation techniques. We performed a bibliometric analysis of the development of intentional tooth replantation. MATERIALS AND METHODS: A comprehensive search of the Web of Science and SCOPUS databases was conducted in February 2023. Original articles and reviews of human studies with "intentional replantation" or synonyms in the titles, abstracts, or keywords were included. A descriptive analysis of bibliographic data, co-occurrence analysis, and coupling of publications was performed. Multivariate analysis was used to explore the bibliometric parameters associated with the citation counts. RESULTS: The study included 171 papers, which were co-authored by 500 individuals affiliated with 217 institutions from 28 countries/regions. The USA contributed the greatest number of publications, followed by China, and Japan. The USA had 694 citations, followed by Japan (210), and Turkey (210). The Journal of Endodontics and Dental Traumatology contributed the most citations. Five directions for future research were identified based on the coupling relationships of publications, including "managing vertical root fractures with adhesive resin using the intentional replantation technique," "intentional replantation for periodontally hopeless or endodontically compromised teeth," "intentional replantation for treating abnormalities of morphological development," "outcomes and prognosis factors of intentional replantation," and "treating root replacement resorption by intentional replantation." Multivariate analysis showed that the publication year, Journal Citation Reports ranking of journals, study design, and disease type were predictors of citation counts. CONCLUSIONS: This bibliometric analysis provides a comprehensive description of the intentional replantation technique. The USA published the greatest volume of papers and generated the most citations. The Journal of Endodontics and Dental Traumatology are considered the most influential. The Journal Citation Reports journal ranking (Q1, Q2), study design (case reports, cohort studies), and disease type (crown root fractures) were associated with the citation counts.

Molecular characterization of TRPA1 and its function in temperature preference in Eriocheir sinensis.

Chinese mitten crab (Eriocheir sinensis) is an economically important aquaculture species, and its growth and development are regulated by temperature, but the molecular mechanisms of the responses to temperature remain unclear. Herein, we identified TRPA1 from E. sinensis, a member of the TRP family of heat receptor potential channels, performed RACE cloning and bioinformatics analysis, and investigated the effect of TRPA1 on temperature responses and molting by real-time PCR and RNA interference (RNAi). The open reading frame of Es-TRPA1 is 3660 bp, and the encoded protein has a molecular weight of 136.91 kDa, and is expressed in embryos and juveniles. RNAi-mediated silencing decreased Es-TRPA1 expression in juvenile crabs, molting rate was decreased, mortality was increased, and crabs avoided cold areas (4 °C) much less than control juvenile crabs. The results suggest that Es-TRPA1 is involved in regulating temperature adaptation and molting processes in E. sinensis. The findings lay a foundation for further exploration of temperature regulation mechanisms in E. sinensis and other crustaceans.

Genome-Wide Identification of Aqp Family Related to Spermatogenesis in Turbot (Scophthalmus maximus).

The development and maturation of sperm entails intricate metabolic processes involving water molecules, amino acids, hormones, and various substances. Among these processes, the role of aquaporins (aqps) in the testis is crucial. Turbot (Scophthalmus maximus) is a significant marine flatfish species in China; however, natural egg laying in females is not feasible under cultured conditions. Consequently, artificial insemination becomes necessary, requiring the retrieval of sperm and eggs through artificial methods. In this study, we combined genomic, transcriptomics, RT-qPCR, computer-assisted sperm analysis (CASA), and immunohistochemistry to investigate the involvement of the aqp family in spermatogenesis in turbot. Through genomic data analysis, we identified 16 aqps genes dispersed across 13 chromosomes, each exhibiting the characteristic major intrinsic protein (MIP) domain associated with AQPs. The results from RNA-seq and RT-qPCR analysis revealed prominent expression of aqp4, 10, and 12 during the proliferative stage, whereas aqp1 showed primary expression during the mature stage. aqp11 displayed high expression levels during both MSII and MSV stages, potentially contributing significantly to the proliferation and maturation of male germ cells. Conversely, aqp8 showed elevated expression levels during the MSIII, MSIII-IV, and MSIV stages, suggesting its direct involvement in spermiogenesis. Immunohistochemical analysis unveiled the predominant localization of AQP1 protein in male germ cells rather than Sertoli cells, specifically concentrated in the head of sperm within cysts. Furthermore, a noteworthy decline in sperm motility was observed when sperm were subjected to treatment with either the AQP1-specific inhibitor (HgCl2) or the AQP1 antibody. However, no direct correlation was found between the expression of Smaqp1 and sperm quality. Overall, these findings provide new insights into the involvement of aqps in teleost spermatogenesis. Moreover, they hold potential for improving techniques related to sperm activation and cryopreservation, offering valuable knowledge for future advancements in this field.

Clinical applications and outcomes of the surgical tooth extrusion technique: A bibliometric analysis from 1982 to 2023.

STATEMENT OF PROBLEM: The surgical extrusion technique has been advocated for severely damaged teeth as well as for intruded and unerupted teeth. However, a quantitative literature analysis is lacking. PURPOSE: The purpose of this bibliometric analysis was to provide a comprehensive overview of surgical extrusion, assess the applications and outcomes, and evaluate topics for future research. MATERIAL AND METHODS: An electronic search was conducted in leading databases in April 2023. Human studies with surgical tooth extrusion or synonyms in titles or abstracts were included. A bibliographic analysis, co-occurrence analysis, and coupling of publications were performed. The analysis of variance was used to assess the characteristics of the subgroups based on indications. The survival rate was evaluated by using the Kaplan-Meier method in the R software program (α=.05). RESULTS: Fifty-one studies from 1982 to 2023 were included. The Journal of Dental Traumatology and the Journal of Prosthetic Dentistry combined published approximately one-third of the papers and acquired the most citations. The burst detection of keywords revealed 4 popular research topics: the Benex extraction system, chlorhexidine for infection control, rigid or semi-rigid splints to decrease the risk of complications, and novel restorative materials to improve esthetics. An overall survival rate of 96.36% was reported among 316 participants with 330 extruded teeth. No significant differences were found in study types, geographic distribution, year of publication, or citation counts among the 4 subgroups (surgical extrusion for subgingival fractures, extensive caries, intruded teeth, and unerupted teeth). A significant difference was observed among the Kaplan-Meier curves of the 4 subgroups (P=.030). CONCLUSIONS: Surgical extrusion may be used to save otherwise nonrestorable teeth, and the procedure has shown promising survival rates when teeth are properly selected and the procedure is well executed. Subgingival fractures and extensive caries near bone level are the predominant indications, as well as intruded and unerupted teeth. Surgical extrusion is a reliable treatment option to facilitate tooth restoration for severely damaged teeth, especially for maxillary anterior teeth.

Prevalence and characteristics of needlestick injuries among dental interns during their first-year clinical training: an observational study.

BACKGROUND: Dental interns are vulnerable to needlestick injuries (NSI). The objectives of this study were to examine the prevalence and characteristics of NSI exposures among dental interns during their first-year clinical training, assess risk factors, and evaluate reporting behaviours. METHODS: An online survey was conducted among dental interns of Class 2011-2017 at Peking University School and Hospital of Stomatology (PKUSS), China. The self-administrated questionnaire consisted of information on demographic profiles, NSI characteristics, and reporting practices. The outcomes were presented by descriptive statistics. A multivariate regression analysis was performed to assess NSI sources using a forward step-wise approach. RESULTS: A total of 407 dental interns completed the survey (response rate 91.9%, 407/443), and 23.8% sustained at least one NSI. The mean number of NSIs per intern was 0.28 during the first clinical year. More occupation exposures occurred from October to December, between 13:00-15:00. Syringe needles were the most frequent sources, followed by dental burs, suture needles, and ultrasonic chips. The risk of peer-inflicted NSIs in the department of Paediatric Dentistry was 12.1 times higher than that in Oral Surgery (OR 12.1, 95% CI: 1.4-101.4). Appropriately 64.9% NSIs occurred when chairside assistants were absent. Compared to working alone, the risk of peer-inflicted NSIs was 32.3 times higher when providing chairside assistance (OR 32.3, 95% CI: 7.2-145.4). The left-hand index finger was the most commonly injured site. About 71.4% of exposures were reported in paperwork. CONCLUSIONS: Dental interns are susceptible to NSIs during their first-year clinical training. Extra attention should be paid to syringe needles, dental burs, suture needles, and ultrasonic chips. The lack of chairside assistance is hazardous regarding NSIs. The training of chairside assistance of the first-year dental interns should be enhanced. First-year dental interns are required to increase their awareness of ignored behaviors related to NSI exposures.

OUTCOMES OF AUTOTRANSPLANTED THIRD MOLARS WITH COMPLETE ROOT FORMATION: A SYSTEMIC REVIEW AND META-ANALYSIS.

OBJECTIVES: Clear evidence is lacking regarding the outcomes of autogenous tooth transplantation (ATT) of third molars with complete root formation. The current review aims to explore the long-term survival and complication rates. METHODS: A comprehensive search was performed in December 2022 of the PubMed, Scopus, Embase, EBSCO, Ovid, Science Direct, and Web of Science databases. The systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines and registered at the International Prospective Register of Systematic Reviews (CRD42022337659). The pooled survival, root resorption, and ankyloses rates were calculated. Subgroup analyses were performed to explore the effects of sample size and 3D techniques. RESULTS: Twelve studies from 5 countries fulfilled the eligibility, with 759 third molars transplanted in 723 patients. Five studies reported 100% survival at 1-year follow-up. After excluding these 5 studies, the pooled survival rate was 93.62% at 1 year. The survival rate of 1 large sample study was significantly higher than that of small ones at 5 years. The complications of studies using 3D techniques were: root resorption 2.06% (95% CI: 0.22, 7.50) and ankyloses 2.81% (95% CI: 0.16, 12.22), compared to those without 3D techniques: root resorption 10.18% (95% CI: 4.50, 17.80) and ankyloses 6.49% (95% CI: 3.45, 10.96). CONCLUSIONS: ATT of third molars with complete root formation is a reliable alternative for replacement of a missing tooth and has a promising survival rate. The use of 3D techniques can reduce complication rates and improve long-term survival.

Reshaping the murine immunoglobulin heavy chain repertoire with bovine DH genes.

Having a limited number of VH segments, cattle rely on uniquely long DH gene segments to generate CDRH3 length variation (3-70 aa) far greater than that in humans or mice. Bovine antibodies with ultralong CDRH3s (>50 aa) possess unusual structures and abilities to bind to special antigens. In this study, we replaced most murine endogenous DH segments with bovine DH genes, generating a mouse line termed B-DH. The use of bovine DH genes significantly increased the length variation of CDRH3 and consequently the Ig heavy chain repertoire in B-DH mice. However, no ultralong CDRH3 was observed in B-DH mice, suggesting that other factors, in addition to long DH genes, are also involved in the formation of ultralong CDRH3. The B-DH mice mounted a normal humoral immune response to various antigens, although the B-cell developmental paradigm was obviously altered compared with wild-type mice. Additionally, B-DH mice are not predisposed to the generation of autoantibodies despite the interspecies DH gene replacement. The B-DH mice reported in this study provide a unique model to answer basic questions regarding the synergistic evolution of DH and VH genes, VDJ recombination and BCR selection in B-cell development.

Microinjection-based CRISPR/Cas9 mutagenesis in the decapoda crustaceans Neocaridina heteropoda and Eriocheir sinensis.

CRISPR/Cas9 technology has been applied to many arthropods. However, application of this technology to crustaceans remains limited because of the unique characteristics of embryos. Our group has developed a microinjection system to introduce the CRISPR/Cas9 system into Neocaridina heteropoda embryos (one-cell stage). Using the developed method, we mutated the target gene Nh-scarlet (N. heteropoda scarlet), which functions in eye development and pigmentation. The results showed that both eye color and shape were altered in individuals in which Nh-scarlet was knocked out. Furthermore, this system was also successfully applied to another decapod crustacean, Eriocheir sinensis. DNA sequencing revealed that the zoeae with red eyes had an edited version of Es-scarlet. This study provides a stable microinjection method for freshwater crustaceans, and will contribute to functional genomics studies in various decapods.

Analysis of the Chinese Alligator TCRα/δ Loci Reveals the Evolutionary Pattern of Atypical TCRδ/TCRµ in Tetrapods.

Atypical TCRδ found in sharks, amphibians, birds, and monotremes and TCRµ found in monotremes and marsupials are TCR chains that use Ig or BCR-like variable domains (VHδ/Vµ) rather than conventional TCR V domains. These unconventional TCR are consistent with a scenario in which TCR and BCR, although having diverged from each other more than 400 million years ago, continue to exchange variable gene segments in generating diversity for Ag recognition. However, the process underlying this exchange and leading to the evolution of these atypical TCR receptor genes remains elusive. In this study, we identified two TCRα/δ gene loci in the Chinese alligator (Alligator sinensis). In total, there were 144 V, 154 Jα, nine Jδ, eight Dδ, two Cα, and five Cδ gene segments in the TCRα/δ loci of the Chinese alligator, representing the most complicated TCRα/δ gene system in both genomic structure and gene content in any tetrapod examined so far. A pool of 32 VHδ genes divided into 18 subfamilies was found to be scattered over the two loci. Phylogenetic analyses revealed that these VHδ genes could be related to bird VHδ genes, VHδ/Vµ genes in platypus or opossum, or alligator VH genes. Based on these findings, a model explaining the evolutionary pattern of atypical TCRδ/TCRµ genes in tetrapods is proposed. This study sheds new light on the evolution of TCR and BCR genes, two of the most essential components of adaptive immunity.

Apocrine Hidrocystoma with IgG4 Plasma Cell Infiltration Presenting as Recurrent Chalazion: A Case Report.

Apocrine hidrocystomas are benign cystic tumors resulting from apocrine sweat glands' proliferation. They typically present as solitary, slow-growing nodules at the head and neck, especially in the periorbital cutaneous region. We present a case of periorbital apocrine hidrocystoma in a 22-year-old woman that was treated as chalazion previously. Besides the hallmark histopathological findings of apocrine hidrocystoma, IgG4 plasma cell infiltration of the cystic wall was also found. The ratio of IgG4-to-IgG-positive plasma cells was high, whereas serum IgG4 was within the standard limit. This is, to date, the only probable IgG4-related ophthalmic disease associated with apocrine hidrocystoma.

FcRn is not the receptor mediating the transfer of serum IgG to colostrum in pigs.

In contrast to humans or rabbits, in which maternal IgG is transmitted to offspring prenatally via the placenta or the yolk sac, large domestic animals such as pigs, cows and sheep transmit IgG exclusively through colostrum feeding after delivery. The extremely high IgG content in colostrum is absorbed by newborns via the small intestine. Although it is widely accepted that the neonatal Fc receptor, FcRn, is the receptor mediating IgG transfer across both the placenta and small intestine, it remains unclear whether FcRn also mediates serum IgG transfer across the mammary barrier to colostrum/milk, especially in large domestic animals. In this study, using a FcRn knockout pig model generated with a CRISPR-Cas9-based approach, we clearly demonstrate that FcRn is not responsible for the IgG transfer from serum to colostrum in pigs, although like in other mammals, it is involved in IgG homeostasis and mediates IgG absorption in the small intestine of newborns.

Identification of Two Nonrearranging IgSF Genes in Chicken Reveals a Novel Family of Putative Remnants of an Antigen Receptor Precursor.

In this study, we identified a pair of nonrearranging VJ-joined Ig superfamily genes, termed putative remnants of an Ag receptor precursor (PRARP) genes, in chicken. Both genes encode a single V-set Ig domain consisting of a canonical J-like segment and a potential immunoreceptor tyrosine-based inhibitory or switch motif in the cytoplasmic region. In vitro experiments showed that both genes were expressed at the cell surface as membrane proteins, and their recombinant products formed a monomer and a disulfide-linked homodimer or a heterodimer. These two genes were mainly expressed in B and T cells and were upregulated in response to stimulation with poly(I:C) in vitro and vaccination in vivo. Orthologs of PRARP have been identified in bony fish, amphibians, reptiles, and other birds, and a V-C1 structure similar to that of Ig or TCR chains was found in all these genes, with the exception of those in avian species, which appear to contain degenerated C1 domains or divergent Ig domains. Phylogenetic analyses suggested that the newly discovered genes do not belong to any known immune receptor family and appear to be a novel gene family. Further elucidation of the functions of PRARP and their origin might provide significant insights into the evolution of the immune system of jawed vertebrates.

MMP-14 regulates innate immune responses to Eriocheir sinensis via tissue degradation.

Matrix metalloproteinases (MMPs) are a cluster of enzymes that degrade the extracellular matrix (ECM) and some intracellular proteins; as such, they play an important role in tissue regeneration, infant growth, animal reproduction, and immunity. Most research into MMPs focuses mainly on their effects on the mammalian immune system. However, it is not clear how MMPs affect immune processes in crustaceans. Here, we cloned the open reading frame (ORF) of Eriocheir sinensis (Chinese mitten crab) MMP-14 (EsMMP-14) to explore the role of MMPs in crustacean innate immune responses. RT-PCR results showed that stimulation of crab with LPS and poly I:C upregulated expression of EsMMP-14 markedly. Besides, following the stimulation of 20-Hydroxyecdysone, the expression level of EsMMP-14 increased robustly, suggesting that EsMMP-14 involved in the molt process of E. sinensis. Hematoxylin and eosin staining of hepatopancreas and intestine revealed that knocking down EsMMP-14 maintained morphology following infection by Bacillus thuringiensis. Moreover, downregulated expression of EsMMP-14 increased the survival rate of infected E. sinensis. These results show that EsMMP-14 plays a role in innate immune responses of E. sinensis and fills a gap in our knowledge about the function of MMPs in crustaceans.

Genomic analysis of a spinal muscular atrophy (SMA) discordant family identifies a novel mutation in TLL2, an activator of growth differentiation factor 8 (myostatin): a case report.

BACKGROUND: Spinal muscular atrophy (SMA) is a rare neuromuscular disorder threating hundreds of thousands of lives worldwide. And the severity of SMA differs among different clinical types, which has been demonstrated to be modified by factors like SMN2, SERF1, NAIP, GTF2H2 and PLS3. However, the severities of many SMA cases, especially the cases within a family, often failed to be explained by these modifiers. Therefore, other modifiers are still waiting to be explored. CASE PRESENTATION: In this study, we presented a rare case of SMA discordant family with a mild SMA male patient and a severe SMA female patient. The two SMA cases fulfilled the diagnostic criteria defined by the International SMA Consortium. With whole exome sequencing, we confirmed the heterozygous deletion of exon7 at SMN1 on the parents' genomes and the homozygous deletions on the two patients' genomes. The MLPA results confirmed the deletions and indicated that all the family members carry two copies of SMN2, SERF1, NAIP and GTF2H2. Further genomic analysis identified compound heterozygous mutations at TLL2 on the male patient's genome, and compound heterozygous mutations at VPS13A and the de novo mutation at AGAP5 on female patient's genome. TLL2 is an activator of myostatin, which negatively regulates the growth of skeletal muscle tissue. Mutation in TLL2 has been proved to increase muscular function in mice model. VPS13A encodes proteins that control the cycling of proteins through the trans-Golgi network to endosomes, lysosomes and the plasma membrane. And AGAP5 was reported to have GTPase activator activity. CONCLUSIONS: We reported a case of SMA discordant family and identified mutations at TLL2, VPS13A and AGAP5 on the patients' genomes. The mutations at TLL2 were predicted to be pathogenic and are likely to alleviate the severity of the male SMA patient. Our finding broadens the spectrum of genetic modifiers of SMA and will contribute to accurate counseling of SMA affected patients and families.

Multifocal musculoskeletal tuberculosis mimicking multiple bone metastases: a case report.

BACKGROUND: The occurrence of non-contiguous, multiple, and remote involvement tuberculous spondylitis is rare. The clinical presentation in patients with multifocal musculoskeletal tuberculosis may closely mimic that in patients with multiple bone metastases, which makes the accurate clinical diagnosis challenging. Herein, we report a multifocal musculoskeletal tuberculosis case that was misdiagnosed for 8 months as multiple bone metastases. CASE PRESENTATION: A 63-year-old male farmer of Chinese Han ethnicity presented to us with pain in left side of the neck, right side of the chest and the back for 10 months without typical tuberculosis symptoms. His past medical history, the CT and fluoroscopy-guided biopsy were negative for tuberculosis. Interferon gamma by T-SPOT was also negative. Radiological findings including CT, MRI and PET-CT suggested that the patient had multiple metastases. Accordingly, the patient was misdiagnosed as having musculoskeletal tumors until a swelling under the right nipple ulcerated. The smear test for acid-fast bacilli and the PCR test for TB-DNA of the pus from the swollen area were both positive, leading to the final correct diagnosis of musculoskeletal tuberculosis. CONCLUSION: The proper diagnosis of musculoskeletal tuberculosis is clinically challenging due to Mycobacterium tuberculosis variants involved and atypical presentations, especially when the lesions are multiple. Our findings indicate that multiple tuberculous spondylitis must be considered in the differential diagnosis of multiple musculoskeletal lesions.

Four carbapenem-resistant gram-negative species carrying distinct carbapenemases in a single patient.

Carbapenem-resistant Escherichia coli, Klebsiella pneumoniae, Enterobacter aerogenes, and Acinetobacter baumannii were isolated from a single patient, each producing different carbapenemases (NDM-1, KPC-2, IMP, and OXA-23, respectively). The NDM-1-producing E. coli strain was preceded by a clonally related carbapenem-susceptible strain a month earlier, suggesting in vivo acquisition of blaNDM-1.

Surgical Reconstruction of Maxillary and Mandibular Defects Using a Printed Titanium Mesh.

PURPOSE: To reconstruct maxillary and mandibular defects with printed titanium mesh using computer-assisted surgery (CAS) for the achievement of structural, esthetic, and functional goals. PATIENTS AND METHODS: The authors designed and implemented this prospective study of patients with maxillary or mandibular defects who underwent reconstruction with printed titanium mesh using CAS. After surgery, the preoperative design and postoperative outcome were evaluated using Geomagic Studio software. RESULTS: The sample was comprised of 2 patients with maxillary defects and 2 with mandibular defects. A satisfactory contour was achieved in all patients. The rate of concordance between the preoperative design and the postoperative outcome was higher than 81 and 94% within 3 mm for the mandibular and maxillary reconstructions, respectively. CONCLUSION: The results of this study suggest that complicated maxillary and mandibular defects can be satisfactorily reconstructed with customized printed titanium meshes using CAS.

Transient receptor potential (TRP) channels in Sebastes schlegelii: Genome-wide identification and ThermoTRP expression analysis under high-temperature.

Transient Receptor Potential (TRP) channels, essential for sensing environmental stimuli, are widely distributed. Among them, thermosensory TRP channels play a crucial role in temperature sensing and regulation. Sebastes schlegelii, a significant aquatic economic species, exhibits sensitivity to temperature across multiple aspects. In this study, we identified 18 SsTRP proteins using whole-genome scanning. Motif analysis revealed motif 2 in all TRP proteins, with conserved motifs in subfamilies. TRP-related domains, anchored repeats, and ion-transmembrane domains were found. Chromosome analysis showed 18 TRP genes on 11 chromosomes and a scaffold. Phylogenetics classified SsTRPs into four subfamilies: TRPM, TRPA, TRPV, and TRPC. In diverse organisms, four monophyletic subfamilies were identified. Additionally, we identified key TRP genes with significantly upregulated transcription levels under short-term (30 min) and long-term (3 days) exposure at 24 °C (optimal elevated temperature) and 27 °C (critical high temperature). We propose that genes upregulated at 30 min may be involved in the primary response process of temperature sensing, while genes upregulated at 3 days may participate in the secondary response process of temperature perception. This study lays the foundation for understanding the regulatory mechanisms of TRPs responses to environmental stimuli in S. schlegelii and other fishes.

Impact of dry eye disease treatment on patient quality of life.

Dry eye disease (DED) is a common multifactorial disease affecting a substantial proportion of the population worldwide. Objective tests and subjective symptoms evaluation are necessary to assess DED. Although various treatments have been introduced, accurately evaluating the efficacy of those treatments is difficult because of the disparity between diagnostic tests and patient-reported symptoms. We reviewed the questionnaires used to evaluate DED and the improvements of quality of life with various treatments. In addition, we highlighted the importance of patient-reported outcomes (PRO) assessments for evaluating the effect of DED treatments. Given that the assessment of DED treatment effectiveness substantially relies on individual ocular experiences, acquiring qualitative PRO data is essential for comprehensive evaluation and optimal treatment management. Clinicians should not only focus on improving objective symptoms but also prioritize the well-being of patients in clinical management.

Innovative approach for high-throughput exploiting sex-specific markers in Japanese parrotfish Oplegnathus fasciatus.

BACKGROUND: The use of sex-specific molecular markers has become a prominent method in enhancing fish production and economic value, as well as providing a foundation for understanding the complex molecular mechanisms involved in fish sex determination. Over the past decades, research on male and female sex identification has predominantly employed molecular biology methodologies such as restriction fragment length polymorphism, random amplification of polymorphic DNA, simple sequence repeat, and amplified fragment length polymorphism. The emergence of high-throughput sequencing technologies, particularly Illumina, has led to the utilization of single nucleotide polymorphism and insertion/deletion variants as significant molecular markers for investigating sex identification in fish. The advancement of sex-controlled breeding encounters numerous challenges, including the inefficiency of current methods, intricate experimental protocols, high costs of development, elevated rates of false positives, marker instability, and cumbersome field-testing procedures. Nevertheless, the emergence and swift progress of PacBio high-throughput sequencing technology, characterized by its long-read output capabilities, offers novel opportunities to overcome these obstacles. FINDINGS: Utilizing male/female assembled genome information in conjunction with short-read sequencing data survey and long-read PacBio sequencing data, a catalog of large-segment (>100 bp) insertion/deletion genetic variants was generated through a genome-wide variant site-scanning approach with bidirectional comparisons. The sequence tagging sites were ranked based on the long-read depth of the insertion/deletion site, with markers exhibiting lower long-read depth being considered more effective for large-segment deletion variants. Subsequently, a catalog of bulk primers and simulated PCR for the male/female variant loci was developed, incorporating primer design for the target region and electronic PCR (e-PCR) technology. The Japanese parrotfish (Oplegnathus fasciatus), belonging to the Oplegnathidae family within the Centrarchiformes order, holds significant economic value as a rocky reef fish indigenous to East Asia. The criteria for rapid identification of male and female differences in Japanese parrotfish were established through agarose gel electrophoresis, which revealed 2 amplified bands for males and 1 amplified band for females. A high-throughput identification catalog of sex-specific markers was then constructed using this method, resulting in the identification of 3,639 (2,786 INS/853 DEL, ♀ as reference) and 3,672 (2,876 INS/833 DEL, ♂ as reference) markers in conjunction with 1,021 and 894 high-quality genetic sex identification markers, respectively. Sixteen differential loci were randomly chosen from the catalog for validation, with 11 of them meeting the criteria for male/female distinctions. The implementation of cost-effective and efficient technological processes would facilitate the rapid advancement of genetic breeding through expediting the high-throughput development of sex genetic markers for various species. CONCLUSIONS: Our study utilized assembled genome information from male and female individuals obtained from PacBio, in addition to data from short-read sequencing data survey and long-read PacBio sequencing data. We extensively employed genome-wide variant site scanning and identification, high-throughput primer design of target regions, and e-PCR batch amplification, along with statistical analysis and ranking of the long-read depth of the variant sites. Through this integrated approach, we successfully compiled a catalog of large insertion/deletion sites (>100 bp) in both male and female Japanese parrotfish.