Ageratina adenophora is a globally invasive perennial herb that poses a significant threat to biodiversity. However, recent studies highlight its potential for ecological utilization, demonstrating antibacterial, anti-inflammatory, and insecticidal properties. Research on its effect on Ascaris suum eggs and the underlying molecular mechanisms remains limited. This study investigated the impact of A. adenophora aqueous extract on A. suum egg development and explored key responsive pathways and genes via RNA sequencing (RNA-Seq). A. suum eggs were treated in vitro with various concentrations (0.10%, 0.25%, 0.50%, 1.00%, 2.50%, 5.00%) of the extract. Embryonation rates decreased dose-dependently, with the 5.00% treatment showing the strongest inhibition (34.24%). Eggs treated with 2.50% and 5.00% extract caused significantly less severe pulmonary lesions and lower larval counts in infected mice, indicating reduced infectivity. Transcriptome analysis of control (WH), 0.10% (EH), and 0.50% (MH) treated eggs identified 281 and 1,083 differentially expressed genes (DEGs) for EH vs. WH and MH vs. WH, respectively. GO and KEGG enrichment analyses revealed that DEGs were primarily involved in transmembrane transport, catalytic activity, carbohydrate metabolic processes, the drug metabolism-cytochrome P450 pathway, and glycolysis/gluconeogenesis. Key genes, including CYP44A1, CYP4V2, CYP4C3, and GST-4, were significantly downregulated. qRT-PCR validated the RNA-Seq results. These findings suggest that A. adenophora extract inhibits A. suum egg development, potentially by disrupting energy metabolism and xenobiotic detoxification pathways. This study provides a theoretical basis for developing A. adenophora-based agents targeting the egg stage of A. suum.

Epinephelus tukula exhibits both rapid growth and strong environmental adaptability, thereby serving as a widely employed broodstock in grouper aquaculture. However, in practical production, the genetic background of E. tukula remains poorly characterized, and the genetic relationships among individuals have not been adequately considered. To date, no studies have reported on the genetic diversity or selection signatures of E. tukula in aquaculture. Based on this, the present study utilized whole-genome sequencing data from 202 E. tukula individuals across multiple farms in different regions of Hainan Province to investigate population structure, genetic diversity, and genomic variations. Ancestral estimation and phylogenetic analysis successfully divided 169 of 202 E. tukula individuals into 10 distinct genetic populations. These populations exhibited low genetic diversity, with nucleotide diversity (π) values ranging from 2.53 to 3.06 × 10− 4. The range of fixation indices (FST) among populations was 0.060–0.153. Linkage disequilibrium (LD) decay rate and runs of homozygosity (ROHs) indicated that certain population likely underwent multiple rounds of artificial breeding, whereas others experienced less artificial breeding. By integrating two methods (XP-EHH and FST), we identified selection signals across these populations. Functional enrichment analysis highlighted 184 development-related genes as potential importance for future breeding efforts. Allele frequency analysis of selected SNPs in these genes across populations revealed that targeted artificial breeding can influence nucleotide diversity in development-related genes. Whole-genome sequencing provided high-quality data for dissecting the genetic structure of E. tukula populations in Hainan Province. Genomic selection sweep identified candidate regions and genes associated with important traits. This study enhances our understanding of the population genetics and selection signatures in E. tukula, and offers valuable insights for germplasm resource management and molecular -assisted breeding in E. Tukula.

BACKGROUND:Eucoleus annulatus is a parasitic nematode that inhabits the upper digestive tract of avian hosts, posing significant threats to avian health and poultry production. However, the gene information and gene expression characteristics underlying its physiological specialization and parasitic adaptation remain poorly understood. METHODS:In this study, we applied an integrated transcriptomic approach to generate a high-quality full-length transcriptome of E. annulatus using PacBio Iso-Seq and to characterize body section-specific gene expression patterns using Illumina RNA sequencing (RNA-Seq). Differentially expressed transcripts (DETs) among its head, middle, and tail sections were identified, and their functional annotations were assessed through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. RESULTS:PacBio Iso-Seq generated 21,951 high-confidence, non-redundant full-length transcript isoforms, among which 6921 were annotated in the Nr, Pfam, COG, KEGG, and GO databases. Comparative RNA-Seq analysis revealed pronounced section-specific transcriptional divergence, with 1570, 1533, and 1600 DETs enriched in the head, middle, and tail sections, respectively. DETs in the head were significantly enriched in pathways related to amino acid metabolism, RNA processing, and ion transport, while the DETs in the middle body section were primarily associated with glycolysis, oxidative phosphorylation, and transcriptional regulation, indicating elevated metabolic and transcriptional activity. DETs in the tail were significantly enriched in processes related to protein degradation, structural maintenance, and stress adaptation, suggesting roles in environmental response and physiological resilience. CONCLUSIONS:This study, for the first time, reports the first full-length transcriptome of E. annulatus and reveals distinct gene expression profiles across different body sections. These findings provide valuable molecular insights into the spatial organization of gene expression in E. annulatus and establish a foundation for studying its biology and host-parasite interactions in the future.

Fusarium verticillioides is a common pathogenic fungus of corn since it causes severe yield losses and produces mycotoxins to threaten the health of both humans and livestock. Although extensive research has characterized specific genetic and environmental factors influencing mycotoxin production, a systematic understanding of the temporal transcriptional dynamics governing its developmental progression remains lacking. This study addresses this critical knowledge gap through a time-series transcriptomic analysis of F. verticillioides at four key cultivation stages (3, 5, 7, and 9 days post-inoculation). Transcriptomic analysis identified 1928, 2818, and 1934 differentially expressed genes (DEGs) in the comparisons of FV3 vs. FV5, FV5 vs. FV7, and FV7 vs. FV9, respectively. Gene Ontology enrichment revealed 76, 106, and 56 significantly enriched terms across these comparisons, with “integral component of membrane” consistently being the most enriched cellular component. Pathway analysis demonstrated “amino acid metabolism” and “carbohydrate metabolism” as the most significantly enriched metabolic pathways. Notably, the fumonisin (FUM) and fusaric acid (FA) biosynthetic gene clusters exhibited coordinated peak expression during the early cultivation, followed by progressive decline. Mfuzz clustering further delineated 12 distinct expression trajectories, highlighting the dynamic transcriptional networks underlying fungal adaptation. This work provided the first comprehensive temporal transcriptome of F. verticillioides, establishing a foundational resource for understanding its stage-specific biology and revealing potential time-sensitive targets for future intervention strategies.

Ticks are hematophagous arthropods that parasitize livestock, humans, and wild animals, posing serious threats to public health worldwide. Diverse pathogens have been discovered in ticks, including viruses, bacteria, protozoa, and filarial nematodes; however, ticks also harbor a variety of fungal microbes, which remain largely unexplored. In this study, we isolated and identified culturable fungi associated with ticks collected on cattle and goats in Yunnan Province, China. Of the 1730 ticks obtained, 1690 were identified as Rhipicephalus microplus, and only 40 as Haemaphysalis longicornis via morphology and 16S rRNA gene analysis. A total of 90 fungal strains were isolated from the ticks and identified by morphological examination and ITS sequencing, representing 25 species across 20 genera, with Fusarium being the most prevalent. Among these isolates, Fusarium verticillioides (19 isolates) poses a threat to grains, animals, and humans; Cladosporium cladosporioides (six isolates) and Sarocladium zeae (one isolate) are primarily pathogenic to plants, while Schizophyllum commune (eight isolates), Rhizopus arrhizus (two isolates), Diaporthe phaseolorum (two isolates), and Purpureocillium lilacinum (two isolates) are recognized animal-associated pathogens. The diversity of isolated fungi with documented pathogenicity suggests that ticks may serve as potential vectors for fungal transmission, raising concerns about their role in cross-kingdom disease spread across agriculture and livestock. Furthermore, fungi with entomopathogenic potential, such as P. lilacinum, offer valuable resources for developing biocontrol agents against ticks. Our findings highlight the dual role of ticks as both potential vectors of phytopathogenic fungi and reservoirs of entomopathogenic biocontrol agents. This necessitates a paradigm shift in tick surveillance, integrating mycological monitoring to mitigate agricultural losses and exploring tick-borne fungi for sustainable vector control strategies.

The porcupinefish (Diodon hystrix), a coral reef teleost, is widely distributed in tropical/subtropical waters of the Pacific, Atlantic, Indian Oceans, and Mediterranean Sea. It shares easily recognizable features with pufferfish, such as body inflation and spines. Additionally, its culinary value makes D. hystrix a highly desirable species in many tropical coastal regions, with considerable market potential. However, lack of a high-quality genome hindered further studies on its reproduction, molecular biology, and genomic improvement. Here, we assembled the chromosome-scale genome using PacBio HiFi, ultra-long reads, and Hi-C. Of the 713.62 Mb genome, 98.63% anchored to 23 chromosomes (scaffold N50: 31.52 Mb) with 39.82% repetitive sequences. The assembled genome achieved a BUSCO completeness score of 97.7%, with 23,171 protein-coding genes predicted, 22,221 of which were functionally annotated. Phylogenetic analysis identified D. hystrix's evolutionary relationships with other species in the Tetraodontiformes. In summary, the high-quality genome of D. hystrix sheds light on valuable insights into genome size evolution, and provides a valuable resource for exploiting genomic study and breeding applications in this species.

Animal mitochondrial DNA (mtDNA) follows the general rule of strict maternal inheritance. The event of hybridization could have an impact on the inherited pattern of mtDNA in hybrid. To investigate the impact of hybridization on mtDNA in different hybrid groupers, the mitogenomes of Cromileptes altivelis x Epinephelus lanceolatus(Hyb1), E. fuscoguttatus x E. polyphekadion (Hyb2), and their parents were determined using nextgeneration sequencing. Genetic diversity and haplotype network were also comparatively analyzed among two hybrid groupers based on the partial cytochrome c oxidase 3 (cox3), trnG and NADH dehydrogenase subunit 3 (nad3) sequences. Our results confirmed that mtDNA of two hybrid groupers were maternal inheritance, showing 99.8 % sequence similarity with their respective female parent. Their mitogenomes presented similar gene architectures. However, the different degrees of nucleotide variation have been detected in two hybrid groupers. Among these two hybrids, Hyb1 presented a higher level of genetic diversity compared with Hyb2. Our observations provide a significant knowledge to understand the effect of hybridization on mtDNA in hybrids.

Diodon hystrix is widely distributed in tropical and subtropical coastal waters. Owing to its delicious taste, the annual catch of this fish in the South China Sea has been on the rise. It is urgent to assess the population status of D. hystrix. Molecular markers are often used to evaluate the population status of the important and rare fishery resource, however, there was still no suitable markers of D. hystrix with overfishing in recent years. This study totally obtained 7.2 Mb clean reads with 94.92 % of Q20 bases and 4179 variants of 2276 unigenes by Illumina sequencing platform. Additionally, 64 amplifiable variants of 36 unigenes were obtained by PCR amplification, and 25 available SNPs showing polymorphism were confirmed from these amplifiable variants. By assessing the index of association, we ultimately identified 14 SNPs after removing 10 SNPs with linkage disequilibrium and 1 SNP with Hardy-Weinberg disequilibrium. Using these 14 SNPs, we can evaluate the wild population of D. hystrix as a sexually reproducing population and speculate that this population was relatively stable (HWE. p = 0.525) with moderate genetic diversity (PIC = 0.328). In conclusion, transcriptomic sequencing can quickly obtain the SNPs as molecular markers which can be used for population genetic diversity. It provides necessary help for the conservation biology and resource evaluation of D. hystrix.

Fusarium verticillioides (F. verticillioides) is an important fungal pathogen known to infect a variety of economically critical crops, particularly maize, causing substantial yield reductions and economic losses worldwide. In addition to its direct damage to agricultural productivity, F. verticillioides threatens public health by producing/secreting potent compounds, including well-known fumonisins (FUMs), which pose significant health threats to both livestock and humans due to their toxicity and carcinogenicity. However, current knowledge of the materials secreted/produced by F. verticillioides, such as secreted proteins and additional secondary metabolites, remains limited. In the present study, we conducted an integrated secretome analysis of F. verticillioides at the exponential growth stage by using proteomic and metabolomic technologies. The results of the present study showed that proteomic analysis identified 185 proteins, including 138 fungus-specific proteins. GO enrichment of these 138 fungus-specific proteins yielded 24 significant terms spanning carbohydrate/polysaccharide and aminoglycan metabolic/catabolic processes, extracellular and membrane-anchored components, and hydrolase/peptidase activities. Meanwhile, KEGG analysis identified starch and sucrose metabolism as the sole significantly enriched pathway. Metabolomic analysis of medium supernatant showed that a total of 2352 metabolites were identified, with 110 unique to the medium supernatant of the fungal group, including fumonisins (A1, B2, B3, B4), fatty acids, and other bioactive compounds. KEGG pathway enrichment highlighted key metabolic pathways, including the TCA cycle, unsaturated fatty acid biosynthesis, and arachidonic acid metabolism. These findings provide new insights into the pathogenic mechanisms of F. verticillioides, suggesting candidates for virulence-associated functions and metabolic adaptations that potentially contribute to its pathogenicity.

Toxoplasma gondii is an obligate intracellular parasite that causes severe illness in infants infected during pregnancy and in immunocompromised individuals. This parasite manipulates host cells through effector proteins that promote its survival and replication. While the phosphatases in the PP2C family have been shown to regulate host immune responses and contribute to the virulence and pathogenicity of various pathogens, the specific biological functions of PPM3H in T. gondii and its role in host–pathogen interactions remain unclear. In this study, we demonstrate that knockout of ppm3h significantly reduces the virulence and pathogenicity of T. gondii. In contrast, that high expression of ppm3h in the less virulent PRU induced by replacing the ppm3h gene elements of RH strain can enhance its pathogenicity, indicating a direct contribution of PPM3H to virulence in expression-independent manner. Furthermore, PPM3H significantly influenced host gene expression, with differentially expressed genes predominantly enriched in immune and inflammatory pathways. Weighted gene co-expression network analysis identified host immune genes, including chemokines such as Cx3cl1 and Ccl22, as co-expressed with ppm3h. Also, ppm3h co-expressed with T. gondii rhoptry genes including rop18, a well-known virulence factor, suggesting a role for PPM3H in coordinating host–pathogen interactions. Our findings establish that PPM3H enhances T. gondii virulence by modulating the host immune and inflammatory responses. PPM3H does not impact parasite gene expression, invasion or replication in vitro, supporting its role as an immune modulator rather than a general fitness factor. This suggests that T. gondii’s pathogenicity arises not only from immune evasion but also from the active induction of host immune and inflammatory responses mediated by PPM3H.

Epinephelus tukula is an economically important aquaculture animal, and a major parent in grouper crossbreeding. To better preserve and exploit E. tukula germplasm resources, a core collection (containing 34 individuals derived from 10 genetic groups) was first constructed based on phenotypic growth traits and whole-genome resequencing (WGS) data. The phenotypic traits of the individuals within the core collection were not significantly different from those in the original collection, suggesting effective representativeness of the core collection. Additionally, we performed genome-wide association study (GWAS) of E. tukula to identify candidate single nucleotide polymorphisms (SNPs) and genes associated with growth traits, to facilitate the improvements in the growth performance of this species. Twenty-six significant SNPs were identified, scattered among multiple chromosomes. Five SNPs were confirmed to be correlated with growth in another new group of 101 individuals. Based on the annotation results, these five SNPs were located in CCDC102A, NTRK2, CTSL, OTOF, and nestin, and were involved in cell development, differentiation and proliferation, glycolytic metabolism, neurological development, and myoblast differentiation. Our findings not only provide an effective basis for the conservation and utilization of E. tukula germplasm resources, but also promote the development of marker-assisted selection of E. tukula.

This study examined the diversity and responses of intestinal microbiota in hybrid grouper (Epinephelus fuscoguttatus ♀ × E. lanceolatus ♂) fed diets with varying levels of fishmeal replaced by black soldier fly larvae (BSFL). The 10% BSFL substitution (BSFL10) group showed the highest levels of trypsin and amylase. Substituting fishmeal with 30% and 50% BSFL weakened the intestinal wall, resulting in vacuoles, sparse striatal boundaries, and fewer villi. Microbiota diversity, measured through Shannon’s index, was higher in the BSFL10 and BSFL50 groups than in the control. 16S rRNA amplicon data revealed the dominance of Firmicutes, Proteobacteria, Bacteroidetes, Spirochaetota, and Verrucomicrobia phyla. The BSFL-replacement groups showed an increase in Proteobacteria, Bacteroidetes, and Spirochaetota compared to the control, but fewer Firmicutes. PICRUSt analysis indicated significant alterations in microbial function, particularly enhanced protein, carbohydrate, lipid, and energy metabolisms in the BSFL-fed group. Substituting 10% fishmeal with BSFL enhanced nutrient metabolism and gut microbiota in juvenile hybrid grouper. Further research is needed to explore factors affecting the efficacy of insect feed as a sustainable aquaculture diet.

The hybrid grouper (Cromileptes altivelas, ♀ × Epinephelus lanceolatus, ♂) is an economically important aquaculture species that exhibits certain growth advantages compared to its female parent, Cromileptes altivelas. However, the current understanding of the molecular mechanisms underlying the growth of hybrid groupers is lacking. Herein, we performed full-length transcriptome sequencing and next-generation sequencing on the hybrid grouper and its parents to identify growth-related genes and comprehensively analyze the regulatory mechanism of growth heterosis in the hybrid grouper. Approximately 44.70, 40.44, and 45.32 Gb of single-molecule real-time sequencing data were generated in C. altivelas (Cal), E. lanceolatus (Ela), and the hybrid (Hyb), which were combined into 204,322 non-redundant isoforms using the PacBio sequencing platform. Differentially expressed genes (DEGs) were identified between Hyb and Cal (3,494, 2,125, and 1,487 in brain, liver, and muscle tissues, respectively) and Hyb and Ela (3,415, 2,351, and 1,675 in brain, liver, and muscle tissues, respectively). Then, 27 DEGs (13 in the brain and 14 in the muscle) related to growth traits were identified using cluster and correlation network analysis. Quantitative RT-PCR validated 15 DEGs consistent with transcriptome sequencing (RNA-seq) trends. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that these 15 genes were mainly involved in regulating the actin cytoskeleton, suggesting that this pathway plays an essential role in fish growth. In addition, we found that the phosphatase and tensin homologue (PTEN) is a key regulator of growth heterosis in Hyb. These results shed light on the regulatory mechanism of growth in the Hyb, which is important for marker-assisted selection programs to improve the growth quality of groupers.

A new species of Moniliformis, M. tupaia n. sp. is described using integrated morphological methods (light and scanning electron microscopy) and molecular techniques (sequencing and analysing the nuclear 18S, ITS, 28S regions and mitochondrial cox1 and cox2 genes), based on specimens collected from the intestine of the northern tree shrew Tupaia belangeri chinensis Anderson (Scandentia: Tupaiidae) in China. Phylogenetic analyses show that M. tupaia n. sp. is a sister to M. moniliformis in the genus Moniliformis, and also challenge the systematic status of Nephridiacanthus major. Moniliformis tupaia n. sp. represents the third Moniliformis species reported from China.

Investigations into the correlation between growth characteristics and DNA methylation levels, along with genetic variations, can provide fundamental insights to enhance growth performance in groupers. The Myostatin (mstn) gene plays a vital role in regulating skeletal muscle development and growth. This study scrutinized the DNA methylation levels of the mstn gene across hybrid groupers (E. fuscoguttatus (♀) × E. polyphekadion (♂)) and their parental species, to evaluate its impact on growth attributes in grouper fish. The nucleotide sequence of the mstn gene was directly sequenced in the hybrid grouper, exhibiting different growth performance to identify the single nucleotide polymorphisms (SNPs) of the mstn gene and explore their correlation with growth characteristics. The findings revealed no significant differences in global DNA methylation levels within muscle tissue among the hybrid grouper and parents. However, significant differences in DNA methylation sites were discovered between the hybrid grouper and E. polyphekadion at sites 824 and 1521 (located at exon 2 and intron 2, respectively), and between E. fuscoguttatus and E. polyphekadion at site 1521. These variations could potentially influence the mRNA expression of the mstn gene. The study also identified that SNP g.1003 T > C in exon 2 of the mstn gene was significantly associated with various growth traits including body weight, total length, body length, head length, caudal peduncle height, and body height (p < 0.01). Specimens with the TT genotype at site 1003 demonstrated superior growth performance compared to those with the TC genotype. Furthermore, microstructural analyses of muscle tissue showed that the average area and diameter of muscle fibers in TT genotype individuals were significantly greater than those in TC genotype individuals. Therefore, this research provides robust evidence linking the DNA methylation level and polymorphisms of the mstn gene with growth traits, which could be beneficial for grouper breeding programs.

Background Hybridization is a useful strategy to produce offspring with more desirable phenotypic characteristics than those of parents. The hybrid grouper derived from the cross of Cromileptes altivelis (♀, 2n = 48) with Epinephelus lanceolatus (♂, 2n = 48) exhibits improved growth compared with its female parent, which makes it valuable to aquaculture. However, the genetic traits of the hybrid grouper are poorly understood. Results The observations showed that the hybrid grouper was diploid (2n = 48) and displayed intermediate morphology with the parent's measurable characteristics. The ribosomal DNA (rDNA) and mitochondria DNA (mtDNA) were characterized at molecular and phylogenetic level. High similarity and low genetic distance of 5S rDNA and mtDNA sequences between the hybrid grouper and C. altivelis showed that the hybrid grouper had a closer genetic relationship with female parents. The reconstructed phylogenetic tree based on COI gene and D-loop region of mtDNA recovered that mtDNA was maternally inherited in the hybrid grouper. Additionally, the DNA methylation level of 5S rDNA intergenic spacers (IGS) sequence was tested in here. The results showed that the DNA methylation status of the hybrid grouper was significantly lower than that of C. altivelis . Conclusion Results of this study provide important data on the genetic characteristics of the hybrid derived from the cross of C. altivelis and E. lanceolatus , and contribute the knowledge of both evolution and marine fish breeding.

Additional file 2: Table S2. The green tea polyphenol-associated targets obtained from the HERB, TCMSP and PharmMapper databases are shown in sheet 1, and predicted herbs targeting MMP1, MMP7, NOS2 and EPHA2 from the HERB database are shown in sheet 2.

Eimeria tenella is an obligate intracellular parasitic protozoan that invades the chicken cecum and causes coccidiosis, which induces acute lesions and weight loss. Elucidating the anticoccidial mechanism of action of green tea polyphenols could aid the development of anticoccidial drugs and resolve the problem of drug resistance in E. tenella. We constructed a model of E. tenella infection in Wuliangshan black-boned chickens, an indigenous breed of Yunnan Province, China, to study the efficacy of green tea polyphenols against the infection. Alterations in gene expression and in the microbial flora in the cecum were analyzed by ribonucleic acid (RNA) sequencing and 16S ribosomal RNA (rRNA) sequencing. Quantitative real-time polymerase chain reaction was used to verify the host gene expression data obtained by RNA sequencing. Network pharmacology and molecular docking were used to clarify the interactions between the component green tea polyphenols and the targeted proteins; potential anticoccidial herbs were also analyzed. Treatment with the green tea polyphenols led to a reduction in the lesion score and weight loss of the chickens induced by E. tenella infection. The expression of matrix metalloproteinase 7 (MMP7), MMP1, nitric oxide synthase 2 and ephrin type-A receptor 2 was significantly altered in the E. tenella infection plus green tea polyphenol-treated group and in the E. tenella infection group compared with the control group; these genes were also predicted targets of tea polyphenols. Furthermore, the tea polyphenol (-)-epigallocatechin gallate acted on most of the targets, and the molecular docking analysis showed that it has good affinity with interferon induced with helicase C domain 1 protein. 16S ribosomal RNA sequencing showed that the green tea polyphenols had a regulatory effect on changes in the fecal microbiota induced by E. tenella infection. In total, 171 herbs were predicted to act on two or three targets in MMP7, MMP1, nitric oxide synthase 2 and ephrin type-A receptor 2. Green tea polyphenols can directly or indirectly regulate host gene expression and alter the growth of microbiota. The results presented here shed light on the mechanism of action of green tea polyphenols against E. tenella infection in chickens, and have implications for the development of novel anticoccidial products.

Hybridization is one of the primary methods used to cultivate farmed grouper species. The hybrid grouper derived from crossing Epinephelus fuscoguttatus (female) and E. polyphekadion (male) exhibits growth superiority over its parents. The genetic characteristics and growth patterns of the hybrid grouper have not yet been defined. This study confirms the ploidy level of the hybrid grouper (2n = 48) using chromosome count analysis and flow cytometry. The 5S rDNA family was used to evaluate genetic diversity. Only one 5S class (similar to 400 bp) was detected in the hybrid grouper, which could be used to distinguish between two different types based on nucleotide sequences, likely representing homologous unit classes from the female and male parental species. Growth patterns of 5-8-month-old hybrid groupers were also monitored. In this phase, a positive allometric growth pattern in body mass with total length was found. Body height and body mass were significantly correlated based on correlation and path coefficient, suggesting that body height could serve as an excellent index to increase body mass. These results aid our understanding of the genetic evolution of the hybrid grouper and inform the development of improved rearing techniques.

Background The Eucotylidae Cohn, 1904 (Superfamily: Microphalloidea), is a family of digeneans parasitic in kidneys of birds as adults. The group is characterized by the high level of morphological similarities among genera and unclear systematic value of morphological characters traditionally used for their differentiation. In the present study, we sequenced the complete or nearly complete mitogenomes (mt genome) of two eucotylids representing the genera Tamerlania ( T. zarudnyi ) and Tanaisia ( Tanaisia sp.). They represent the first sequenced mt genomes of any member of the superfamily Microphalloidea. Methods A comparative mitogenomic analysis of the two newly sequenced eucotylids was conducted for the investigation of mitochondrial gene arrangement, contents and genetic distance. Phylogenetic position of the family Eucotylidae within the order Plagiorchiida was examined using nucleotide sequences of mitochondrial protein-coding genes (PCGs) plus RNAs using maximum likelihood (ML) and Bayesian inference (BI) methods. BI phylogeny based on concatenated amino acids sequences of PCGs was also conducted to determine possible effects of silent mutations. Results The complete mt genome of T. zarudnyi was 16,188 bp and the nearly complete mt genome of Tanaisia sp. was 13,953 bp in length. A long string of additional amino acids (about 123 aa) at the 5′ end of the cox 1 gene in both studied eucotylid mt genomes has resulted in the cox 1 gene of eucotylids being longer than in all previously sequenced digeneans. The rrnL gene was also longer than previously reported in any digenean mitogenome sequenced so far. The TΨC and DHU loops of the tRNAs varied greatly between the two eucotylids while the anticodon loop was highly conserved. Phylogenetic analyses based on mtDNA nucleotide and amino acids sequences (as a separate set) positioned eucotylids as a sister group to all remaining members of the order Plagiorchiida. Both ML and BI phylogenies revealed the paraphyletic nature of the superfamily Gorgoderoidea and the suborder Xiphidiata. Conclusions The average sequence identity, combined nucleotide diversity and Kimura-2 parameter distances between the two eucotylid mitogenomes demonstrated that atp 6, nad 5, nad 4L and nad 6 genes are better markers than the traditionally used cox 1 or nad 1 for the species differentiation and population-level studies of eucotylids because of their higher variability. The position of the Dicrocoeliidae and Eucotylidae outside the clade uniting other xiphidiatan trematodes strengthened the argument for the need for re-evaluation of the taxonomic content of the Xiphidiata. Graphical Abstract