The complete mitochondrial genome of Ogmocotyle ailuri: gene content, composition and rearrangement and phylogenetic implications.

Trematodes of the genus Ogmocotyle are intestinal flukes that can infect a variety of definitive hosts, resulting in significant economic losses worldwide. However, there are few studies on molecular data of these trematodes. In this study, the mitochondrial (mt) genome of Ogmocotyle ailuri isolated from red panda (Ailurus fulgens) was determined and compared with those from Pronocephalata to investigate the mt genome content, genetic distance, gene rearrangements and phylogeny. The complete mt genome of O. ailuri is a typical closed circular molecule of 14 642 base pairs, comprising 12 protein-coding genes (PCGs), 22 transfer RNA genes, 2 ribosomal RNA genes and 2 non-coding regions. All genes are transcribed in the same direction. In addition, 23 intergenic spacers and 2 locations with gene overlaps were determined. Sequence identities and sliding window analysis indicated that cox1 is the most conserved gene among 12 PCGs in O. ailuri mt genome. The sequenced mt genomes of the 48 Plagiorchiida trematodes showed 5 types of gene arrangement based on all mt genome genes, with the gene arrangement of O. ailuri being type I. Phylogenetic analysis using concatenated amino acid sequences of 12 PCGs revealed that O. ailuri was closer to Ogmocotyle sikae than to Notocotylus intestinalis. These data enhance the Ogmocotyle mt genome database and provide molecular resources for further studies of Pronocephalata taxonomy, population genetics and systematics.

Survey of the Prosthogonimus spp. metacercariae infection in the second intermediate host dragonfly in Heilongjiang Province, China.

Prosthogonimiasis poses a threat to the reproductive system of poultry and wild birds, which are the definitive hosts of the parasite causing this disease. However, the parasite infection of the second intermediate host (dragonfly), the primary vector of this pathogen, is rarely reported. In this study, the prevalence of Prosthogonimus infection in dragonflies was investigated from June 2019 to October 2022 in Heilongjiang Province, northeast China. The species of metacercariae isolated from dragonfly were identified by morphological characteristics, molecular biology techniques, and animal infection experiments. The results showed that 11 species of dragonflies and one damselfly were identified and among six of the dragonflies infected by Prosthogonimus metacercariae, Sympetrum depressiusculum (28.53%) had the highest infection rate among all positive dragonflies, followed by Sympetrum vulgatum (27.86%) and Sympetrum frequens (20.99%), which are preferred hosts, and the total prevalence was 20.39% (2061/10,110) in Heilongjiang Province. Three species of Prosthogoniumus metacercariae were isolated, including Prosthogonimus cuneatus, Prosthogonimus pullucidus, and Prosthogonimus sp., among which P. cuneatus was the dominant species in dragonflies in Heilongjiang Province. This is the first report on the prevalence of Prosthogonimus in dragonflies in China, which provides baseline data for the control of prosthogonimiasis in Heilongjiang Province and a reference for the prevention of prosthogonimiasis in other areas of China.

Prevalence of Theileria in cattle in China: A systematic review and meta-analysis.

Theileria, one of the causative agents of blood protozoan, has brought a huge economic loss to the cattle industry worldwide. However, the epidemiology of Theileria in Chinese cattle has not been systematically investigated. This comprehensive review aimed at investigating the prevalence of Theileria infection in cattle in China. A total of 48 published papers on Theileria infection in cattle in China (including data from 21,366 animals) from inception to October 8, 2021 met the inclusion standard after searching in five databases (Technology Periodical Database, Wan Fang Database, China National Knowledge Infrastructure, PubMed, and ScienceDirect). The pooled prevalence of Theileria in cattle in China was 32.4% identified by using a random effects model. The prevalence in Northeastern China (45.3%) was higher than that in other regions. In the sex subgroup, the prevalence of Theileria was higher in females (48.9%) than that in males (45.8%). The prevalence of Theileria was higher in cattle of free range (34.4%) compared with that of intensive farming (22.3%). The prevalence prior to 2013 (36.1%) was higher than that after 2013 (33.6%). Among three cattle species, dairy cows had the lowest prevalence (21.5%). The prevalence of Theileria (T.) annulata (22.2%) and T. sergenti (26.2%) was higher than other species of Theileria (T. buffeli: 17.5%, T. luwenshuni: 0.9%, T. orientalis: 15.5%, T. ovis: 0.21%, T. sinensis: 20.2%, T. uilenbergi: 6.2%, Others: 0.9%). We also analyzed the impact of different geographic factor subgroups (longitude, latitude, precipitation, temperature, humidity, and altitude) on the prevalence of Theileria in cattle. Among them, climatic factors of longitude, latitude, precipitation, humidity, temperature were associated with the prevalence of Theileria. These analyses suggested that Theileria was common in cattle in China. Targeted prevention programs based on geographic and climatic conditions in different areas may play an important role in reducing Theileria infection among cattle.

Molecular detection of Cryptosporidium spp. and Enterocytozoon bieneusi in Longjiang Wagyu cattle in Northeastern China.

Cryptosporidium spp. and Enterocytozoon bieneusi are two important zoonotic pathogens that can cause diarrhea and other gastrointestinal illnesses in humans and animals. However, the prevalence and genotype of the parasites in Longjiang Wagyu cattle in Heilongjiang Province, Northeast China have not been reported. In the present study, a total of 423 fecal samples of Longjiang Wagyu cattle collected from different farms in Heilongjiang Province, Northeast China, were examined for Cryptosporidium spp. and E. bieneusi using nested PCR. The overall infection rates for Cryptosporidium spp. and E. bieneusi were 6.38% (n = 27) and 7.09% (n = 30), respectively. The prevalence in different age groups ranged from 3.80% (95% confidence interval (CI) 1.01-6.59) to 8.36% (95% CI 4.83-11.90) for Cryptosporidium spp. and 5.97% (95% CI 2.52-9.43) to 7.94% (95% CI 4.49-11.40) for E. bieneusi. By analyzing the DNA sequences of the small subunit (SSU) rRNA gene, two Cryptosporidium species were detected in this study, namely C. parvum (n = 25) and C. ryanae (n = 2). The IIdA20G1 subtype was further identified by using the 60-kDa glycoprotein (gp60) gene of C. parvum. E. bieneusi was identified using three known sequences through the analysis of internal transcribed spacer (ITS) sequences: J (n = 23), I (n = 5), and BEB4 (n = 2), and all belonged to group 2. The results indicated that some of the Cryptosporidium species and E. bieneusi genotypes identified in Longjiang Wagyu cattle in the study areas might have zoonotic potential.

Prevalence of Clonorchis sinensis infection in fish in South-East Asia: A systematic review and meta-analysis.

Clonorchis sinensis, an important fish-borne zoonotic trematode, is widely distributed in South-East Asia, especially in China. Infections from human and animal reservoir hosts occur due to the consumption of raw or undercooked fish with C. sinensis metacercariae. This study aimed to evaluate the prevalence of C. sinensis metacercariae in fish in South-East Asia via systematic review and meta-analysis. We searched PubMed, ScienceDirect, China National Knowledge Infrastructure, Wanfang and Chongqing VIP databases for studies published between 1976 and 2020 that are related to the prevalence of C. sinensis metacercariae in fish. Studies were screened with keywords based on inclusion and exclusion criteria. Seventy-one eligible articles were identified, covering three countries: China, Korea and Vietnam. The pooled prevalence of C. sinensis metacercariae in fish from South-East Asia was 30.5%, with 35.1% in China, 29.7% in Korea and 8.4% in Vietnam. In subgroup analyses of climate, season, water source and publication date, the highest prevalence was identified in the Dwb climate type (43.3%), summer (70.2%), river (34.5%) and pre-2001 publications (38.9%), respectively. In comparison, the lowest prevalence was found in the Dfa climate type (14.5%), winter (19.5%), lake (8.0%) and post-2001 publications (23.8%). Meta-regression results indicated that country (p = .009), the published time (p = .035) and water source subgroups (p = .003) may be the source of heterogeneity. Overall, our study indicates that a high prevalence of C. sinensis infections occurs in fish in China, Korea and Vietnam, illuminating a significant public health concern in these countries.

Proteomic analysis of Fasciola gigantica excretory and secretory products (FgESPs) interacting with buffalo serum of different infection periods by shotgun LC-MS/MS.

Fasciolosis, caused by Fasciola hepatica and Fasciola gigantica, is an important zoonotic disease in the world. It affects livestock, especially for sheep and cattle, causing major economic loss due to morbidity and mortality. Although the excretory and secretory products (ESPs) of F. hepatica have been relatively well studied, little is known about the interaction between the ESP and host, and the mechanism of the key proteins involved in interaction. In this study, buffaloes were infected by Fasciola gigantica, and infection serum was collected at three different periods (42dpi, 70dpi, and 98dpi). The interaction proteins were pulled down with three different period serum by Co-IP assay, respectively, and then identified by LC-MS/MS analysis. A number of proteins were identified; some of them related to the biological function of the parasite, while most of them the functions were unknown. For the annotated proteins, 13, 5, and 7 proteins were pulled down by the infected serum in 42dpi, 70dpi, and 98dpi, respectively, and 18 proteins could be detected in all three periods. Among them, 13 belong to the cathepsin family, 4 proteins related to glutathione S-transferase, and 3 proteins are calcium-binding protein; other proteins related to catalytic activity and cellular process. This study could provide new insights into the central role played by ESPs in the protection of F. gigantica from the host immune response. At the same time, our research provided material for further studies about the interaction between F. gigantica and host.

De novo assembly and functional annotations of the transcriptome of Metorchis orientalis (trematoda: Opisthorchiidae).

Metorchis orientalis is a neglected zoonotic parasite, living in the gallbladder and bile duct of poultry and some mammals as well as humans. In spite of its economic and medical importance, the information known about the transcriptome and genome of M. orientalis is limited. In this study, we performed de novo sequencing, transcriptome assembly and functional annotations of the adult M. orientalis, obtained about 77.4 million high-quality clean reads, among which the length of the transcript contigs ranged from 100 to 11,249 nt with mean length of 373 nt and N50 length of 919 nt. We then assembled 31,943 unigenes, of which 20,009 (62.6%) were annotated by BLASTn and BLASTx searches against the available database. Among these unigenes, 19,795 (62.0%), 3407 (10.7%), 10,620 (33.2%) of them had significant similarity in the NR, NT and Swiss-Prot databases, respectively; 5744 (18.0%) and 4678 (14.6%) unigenes were assigned to GO and COG, respectively; and 9099 (28.5%) unigenes were identified and mapped onto 256 pathways in the KEGG Pathway database. Furthermore, we found that 98 (1.08%) unigenes were related to bile secretion and 5 (0.05%) to primary bile acid biosynthesis pathways category. The characterization of these transcriptomic data has implications for the better understanding of the biology of M. orientalis, and will facilitate the development of intervention agents for this and other pathogenic flukes of human and animal health significance.

Complete mitochondrial genome of parasitic nematode Cylicocyclus nassatus and comparative analyses with Cylicocyclus insigne.

Cylicocyclus nassatus is a common and important parasite in the large intestine of equine. In this study, the complete mitochondrial (mt) genome sequence of C. nassatus was determined and comparatively analyzed with Cylicocyclus insigne. The mt genome size of C. nassatus was 13,846 bp, 18 bp longer than that of C. insigne. The circular mt genome includes 12 protein-coding genes, two rRNA genes, 22 tRNA genes and two non-coding regions. All the genes are transcribed in the same direction and gene arrangement is consistent with that of gene arrangement 3 (GA3). The overall sequence difference between the two complete mt genomes was 10.7%. For the 12 protein-coding genes, the comparison between C. nassatus and C. insigne revealed sequence divergence at both the nucleotide (6.3-13.0%) and amino acid (0.8-6.6%) levels. The nucleotide lengths of the 12 protein-coding genes were the same, except for cox3 which was longer in C. insigne. Phylogenetic analysis based on the concatenated amino acid sequence of the 12 protein-coding genes was performed using all the Strongylidae nematodes of the horse available in the GenBank. Phylogenetic analysis showed that C. nassatus and C. insigne clustered together with very high nodal support and the genus Cylicocyclus was closer to the genus Triodontophorus than to genus Strongylus. The mtDNA data determined in this study provides novel genetic markers for further studies on the identification, population genetics and molecular epidemiology of the genus Cylicocyclus.

Complete mitochondrial genomes of Triodontophorus serratus and Triodontophorus nipponicus, and their comparison with Triodontophorus brevicauda.

Triodontophorus serratus and Triodontophorus nipponicus are two of the most common nematodes inhabiting in the large intestine of horse. In the present study, the complete mitochondrial (mt) genome sequences of T. serratus and T. nipponicus have been determined. The mt genomes of T. serratus and T. nipponicus are circular molecules with 13,794 bp and 13,701 bp in size, respectively. These circular mt genomes encode 36 genes, including 12 protein-coding genes, two rRNA genes and 22 tRNA genes. All of these genes are transcribed in the same direction and gene arrangements are consistent with that of gene arrangement 3 (GA3-type). T. serratus and T. nipponicus had two non-coding regions, but T. brevicauda had three. Phylogenetic relationships were reconstructed using concatenated amino acid sequences of the 12 protein-coding genes with three methods, indicating that three species of Triodontophorus clustered together with strong statistical support. However, the genera of Strongylus and Triodontophorus belonged to Strongylinae do not cluster together, and Triodontophorus is more closely related to Cylicocyclus insigne, Cylicocyclus nassatus, Cylicostephanus goldi (Cyathostominae) than to Strongylus. The findings from the present study provide useful genetic markers for studying the molecular ecology, systematics, and population genetics of Triodontophorus in equine.

Metacercaria Infection Status of Fishborne Zoonotic Trematodes, Except for Clonorchis sinensis in Fish from the Heilongjiang Province, China.

To investigate the metacercarial infections of fishborne zoonotic trematodes (FZT), a total of 6815 freshwater fish (in representing 13 species of 5 families) were collected from Songhua river (n = 2636), Nenjiang river (n = 1935), Mudanjiang river (n = 301), and other lakes or ponds (n = 1943) in 36 representative regions in Heilongjiang Province, China, from August 2012 to December 2015. Metacercariae of four FZT species, that is, Clonorchis sinensis, Metorchis orientalis, Isthmiophora hortensis, and Metagonimus yokogawai, metacercariae were detected in the examination by the artificial digestion method. As the partial data for C. sinensis were previously reported, the remaining three FZT species are to be treated in this study. The overall prevalence of M. orientalis, I. hortensis, and M. yokogawai, metacercariae was 10.54%, 0.28%, and 1.35%, respectively. Metacercariae of M. orientalis were detected in seven fish species, that is, Pseudorasbora parva, Hemiculter leucisculus, Saurogobio dabryi, Rhynchocypris lagowskii, Carassius auratus, Rhodeus ocellatus and Perccottus glehnii. Their prevalences were the highest in false dace, P. parva (26.81%), and in fish from Songhua river (17.94%). Metacercariae of I. hortensis were detected in only one fish species, Misgurnus anguillicaudatus, from Nenjiang river only. Metacercariae of M. yokogawai were detected in three fish species, that is, P. parva, H. leucisculus and S. dabryi. Their prevalences were the highest in sharpbelly, H. leucisculus (6.05%), and in fish from Mudanjiang river (5.65%). This study first demonstrated the existence of M. orientalis, I. hortensis, and M. yokogawai in freshwater fish from Heilongjiang Province, posing a major public health concern. Eight fish species, namely M. anguillicaudatus, P. parva, H. leucisculus, S. dabryi, R. lagowskii, C. auratus, R. ocellatus, and P. glehnii, cannot be eaten raw. Moreover, the findings of this study not only extended the second intermediate host range of FZT, but also improve the information of the distribution of FZT in China.

Complete mitochondrial genomes of Dermacentor silvarum and comparative analyses with another hard tick Dermacentor nitens.

Ticks are obligate blood-sucking ectoparasites of a wide range of vertebrates. They can transmit a range of pathogens that cause economic losses to livestock production as well as human disease. In the present study, the complete mitochondrial (mt) genome of Dermacentor silvarum was determined. The mt genome is 14,945 bp in length contains 37 genes, including 13 are protein-coding genes (cox1-3, nad1-6, nad4L, cytb, atp6 and atp8), two ribosomal RNA genes and 22 transfer RNA genes. The nucleotide composition of the D. silvarum mt genome was A + T biased at 78.78%; T was the most abundant nucleotide and G the least abundant. The mt genome of D. silvarum was 106 bp longer than that of Dermacentor nitens and the arrangements of two genomes were identical. For the 13 protein-coding genes, comparison between D. silvarum and D. nitens revealed sequence divergence at both the nucleotide (15.46-35.14%) and amino acid (6.05-48.98%) levels. Among them, cox1 was the most conserved gene, while atp8 was the least conserved. The lengths of the 13 protein-coding genes were the same or similar, except for cytb which was significantly longer in D. silvarum than in D. nitens. The mtDNA contained a variable repeat region consisting of a "similar to nad1" motif that was repeated three times, and the "Tick-box" motifs were also found. The overall difference between the nucleotide sequences of the two complete mt genomes was 21.4%. The mtDNA data presented in this study provide a rich resource for further studies on the phylogenetics, population genetics, and molecular epidemiology of ticks.

The complete mitochondrial genome of Gasterophilus intestinalis, the first representative of the family Gasterophilidae.

Gasterophilus spp. (Diptera: Gasterophilidae) has a worldwide distribution; however, no complete mitochondrial (mt) genome data is available for Diptera which has greatly impeded population genetics, phylogenetics, and systematics studies in Gasterophilidae. Mt genome is known to provide genetic markers for investigations in these areas, but complete mt genomic datasets have been lacking for many Gasterophilidae species. Herein, we present the complete mt genome of the third-stage larvae (L3) of Gasterophilus intestinalis from the stomach wall of naturally infected horses in Heilongjiang province (HLJ) and Xinjiang Uygur Autonomous Region (XJ), China. The complete mt genome of G. intestinalis was 15,687 bp (HLJ) and 15,660 bp (XJ) in length and consists of 37 genes, including 13 genes for proteins, 22 genes for tRNA, and 2 genes for rRNA. The gene arrangement is the same as those of Oestroidae species. Phylogenetic analyses using concatenated amino acid sequences of 12 protein-coding genes by Bayesian inference (BI) and maximum likelihood (ML), suggested that the families Gasterophilidae and Oestroidae were more closely related than to Tachinidae. The mt genome of G. intestinalis represents the first mt genome of any member of the family Gasterophilidae. These data provide novel mtDNA markers for studying the molecular epidemiology and population genetics of the G. intestinalis and its congeners.

The complete mitochondrial genome of rabbit pinworm Passalurus ambiguus: genome characterization and phylogenetic analysis.

Passalurus ambiguus (Nematda: Oxyuridae) is a common pinworm which parasitizes in the caecum and colon of rabbits. Despite its significance as a pathogen, the epidemiology, genetics, systematics, and biology of this pinworm remain poorly understood. In the present study, we sequenced the complete mitochondrial (mt) genome of P. ambiguus. The circular mt genome is 14,023 bp in size and encodes of 36 genes, including 12 protein-coding, two ribosomal RNA, and 22 transfer RNA genes. The mt gene order of P. ambiguus is the same as that of Wellcomia siamensis, but distinct from that of Enterobius vermicularis. Phylogenetic analyses based on concatenated amino acid sequences of 12 protein-coding genes by Bayesian inference (BI) showed that P. ambiguus was more closely related to W. siamensis than to E. vermicularis. This mt genome provides novel genetic markers for studying the molecular epidemiology, population genetics, systematics of pinworm of animals and humans, and should have implications for the diagnosis, prevention, and control of passaluriasis in rabbits and other animals.

Complete Mitochondrial Genome of Echinostoma hortense (Digenea: Echinostomatidae).

Echinostoma hortense (Digenea: Echinostomatidae) is one of the intestinal flukes with medical importance in humans. However, the mitochondrial (mt) genome of this fluke has not been known yet. The present study has determined the complete mt genome sequences of E. hortense and assessed the phylogenetic relationships with other digenean species for which the complete mt genome sequences are available in GenBank using concatenated amino acid sequences inferred from 12 protein-coding genes. The mt genome of E. hortense contained 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 non-coding region. The length of the mt genome of E. hortense was 14,994 bp, which was somewhat smaller than those of other trematode species. Phylogenetic analyses based on concatenated nucleotide sequence datasets for all 12 protein-coding genes using maximum parsimony (MP) method showed that E. hortense and Hypoderaeum conoideum gathered together, and they were closer to each other than to Fasciolidae and other echinostomatid trematodes. The availability of the complete mt genome sequences of E. hortense provides important genetic markers for diagnostics, population genetics, and evolutionary studies of digeneans.

The complete mitochondrial genomes of five Eimeria species infecting domestic rabbits.

Rabbit coccidiosis caused by members of the genus Eimeria can cause enormous economic impact worldwide, but the genetics, epidemiology and biology of these parasites remain poorly understood. In the present study, we sequenced and annotated the complete mitochondrial (mt) genomes of five Eimeria species that commonly infect the domestic rabbits. The complete mt genomes of Eimeria intestinalis, Eimeria flavescens, Eimeria media, Eimeria vejdovskyi and Eimeria irresidua were 6261bp, 6258bp, 6168bp, 6254bp, 6259bp in length, respectively. All of the mt genomes consist of 3 genes for proteins (cytb, cox1, and cox3), 14 gene fragments for the large subunit (LSU) rRNA and 11 gene fragments for the small subunit (SSU) rRNA, but no transfer RNA (tRNA) genes. The gene order of the mt genomes is similar to that of Plasmodium, but distinct from Haemosporida and Theileria. Phylogenetic analyses based on full nucleotide sequences using Bayesian analysis revealed that the monophyly of the Eimeria of rabbits was strongly statistically supported with a Bayesian posterior probabilities. These data provide novel mtDNA markers for studying the population genetics and molecular epidemiology of the Eimeria species, and should have implications for the molecular diagnosis, prevention and control of coccidiosis in rabbits.

The complete mitochondrial genome of Oxyuris equi: Comparison with other closely related species and phylogenetic implications.

The equine pinworm Oxyuris equi (Nematoda: Oxyuridomorpha) is the most common horse nematode, has a worldwide distribution, and causes major economic losses. In the present study, the complete O. equi mitochondrial (mt) genome was sequenced, and the mt genome structure and organization were compared with those of other closely related pinworm species, Enterobius vermicularis and Wellcomia siamensis. The O. equi mt genome is a 13,641-bp circular DNA molecule that encodes 36 genes (12 protein-coding genes, 22 tRNAs, and two rRNAs) and one non-coding region, which is slightly shorter than that of E. vermicularis and W. siamensis. The O. equi mt gene arrangement was consistent with that of GA13-type E. vermicularis but it differs from GA12-type W. siamensis. Phylogenetic analyses using concatenated amino acid sequences of the 12 protein-coding genes with three different computational algorithms (maximum parsimony, maximum likelihood, and Bayesian inference) revealed that there were two distinct clades in Chromadorea nematodes that reflected infraorder. Spiruromorpha formed one clade, whereas Rhabditomorpha, Ascaridomorpha, and Oxyuridomorpha formed another clade. O. equi, E. vermicularis, and W. siamensis represent distinct but closely related species, which indicated that Oxyuridomorpha is paraphyletic. Sequencing the O. equi mt genome provides novel genetic markers for studying the molecular epidemiology and population genetics of pinworms.

The complete mitochondrial genome of Strongylus equinus (Chromadorea: Strongylidae): Comparison with other closely related species and phylogenetic analyses.

The roundworms of genus Strongylus are the common parasitic nematodes in the large intestine of equine, causing significant economic losses to the livestock industries. In spite of its importance, the genetic data and epidemiology of this parasite are not entirely understood. In the present study, the complete S. equinus mitochondrial (mt) genome was determined. The length of S. equinus mt genome DNA sequence is 14,545 bp, containing 36 genes, of which 12 code for protein, 22 for transfer RNA, and two for ribosomal RNA, but lacks atp8 gene. All 36 genes are encoded in the same direction which is consistent with all other Chromadorea nematode mtDNAs published to date. Phylogenetic analysis based on concatenated amino acid sequence data of all 12 protein-coding genes showed that there were two large branches in the Strongyloidea nematodes, and S. equinus is genetically closer to S. vulgaris than to Cylicocyclus insignis in Strongylidae. This new mt genome provides a source of genetic markers for the molecular phylogeny and population genetics of equine strongyles.

Characterization of the complete nuclear ribosomal DNA sequences of Paramphistomum cervi.

Sequences of the complete nuclear ribosomal DNA (rDNA) gene from five individual Paramphistomum cervi were determined for the first time. The five complete rDNA sequences, which included the 18S rDNA, the internal transcribed spacer 1 (ITS1), the 5.8S rDNA, the internal transcribed spacer 2 (ITS2), the 28S rDNA, and the intergenic spacer (IGS) regions, had a length range of 8,493-10,221 bp. The lengths of the investigated 18S, ITS1, 5.8S, ITS2, and 28S rDNA sequences, which were 1,994 bp, 1,293 bp, 157 bp, 286 bp, and 4,186 bp, respectively, did not vary. However, the IGS rDNA sequences had a length range of 577-2,305 bp. The 5.8S and ITS-2 rDNA sequences had 100% identity among the five investigated samples, while the identities among the IGS had a range of 53.7-99.8%. A comparative analysis revealed that different types and numbers of repeats were found within each ITS1 and IGS region, which may be related to the length polymorphism of IGS. The phylogenetic position of P. cervi in Paramphistomatidae was analyzed based on the 18S rDNA sequences. These results will aid in studying the intra- and interspecific variation of the Paramphistomatidae and the systematics and phylogenetics of Digenea.

[Immunoreactivity and immunogenicity analysis of the recombinant cathepsin L-like protease of Fasciola hepatica in SD rats].

OBJECTIVE: To analyze the immunoreactivity of recombinant cathepsin L-like proteases (CatL) protein of Fasciola hepatica and its immunogenicity in SD rats. METHODS: The E. coli BL21(DE3) cells harbouring recombinant plasmid pET30a-FhCatL were inoculated in LB medium, and the protein expression was induced with IPTG. The recombinant protein FhCatL was analyzed by SDS-PAGE and the immunoreactivity was identified by Western blotting with sera from Fasciola hepatica-infected goat as the primary antibody. Twenty SD rats were randomly divided into immunized group and adjuvant control group. SD rats in immunized group were injected subcutaneously with 200 microg of purified FhCatL protein. All the rats received three immunizations at 3-week intervals. The adjuvant control group with 10 SD rats received only adjuvants emulsified with the same amount of PBS. Serum samples were collected at the day before the second and final immunization, 3, 6, and 9 weeks after the final immunization. The IgG antibody of rats' sera was examined by indirect ELISA and spleen lymphocyte proliferation (SLP) was tested by methyl thiazolyl tetrazolium (MTT). RESULTS: The molecular weight of purified FhCatL was about Mr 42,000. The recombinant FhCatL was recognized by pool sera of goats naturally infected with F. hepatica. The titer of specific antibody IgG in SD rats induced by the recombinant protein against CatL protein was significantly higher than that of the control, and the antibody titer reached the peak at three weeks after the final immunization (1 : 102,400). The stimulation index of splenocytes in immunized group was 2.176 +/- 0.047, which was significantly higher than that of the control (1.171 +/- 0.032) (P<0.05). CONCLUSION: The recombinant FhCatL protein bears stronger immnoreactivity and immunogenicity.

Global prevalence of liver disease in human and domestic animals caused by Fasciola: A systematic review and meta-analysis.

Background: Liver disease caused by Fasciola is a significant zoonotic and parasitic disease with substantial economic impacts on humans and animals. Many studies have looked at the prevalence of fasciolis worldwide, yet the overall prevalence and risk factors in cattle, ruminants, and humans remains unknown. Methods: We conducted a systematic review and meta-analysis to estimate the global prevalence and risk factors of fascioliasis in humans and domestic ruminants. With this aim, we searched PubMed, ScienceDirect, Web of Science, and Scopus from inception to 8 December 2022 for studies reporting the prevalence of fascioliasis in humans or domestic ruminants post-2000. We then used random effects models to describe the prevalence of fascioliasis; trim-and-fill analysis and Egger's test to assess publication bias; and meta-regression and sensitivity analyses to examine the risk factors for prevalence and heterogeneity. Results: We retrieved 4422 articles, with 371 being included in the analysis, as they concerned fascioliasis in humans and ruminants globally. The pooled prevalence of bovine fasciolosis was 17%, while ovine fasciolosis and human fascioliasis had pooled prevalences of 13% and 5%, respectively. We also conducted subgroup analyses by continents, countries, Fasciola species, sampling years, altitude, rainfall, temperature, humidity, age, sex, feeding mode, and residence. Here, altitude and age emerged as risk factors associated with an increased prevalence of fascioliasis. Both the trim-and-fill analysis and Egger's test confirmed the presence of publication bias, while the sensitivity analysis showed that the omission of any single study did not significantly influence the combined pooled prevalence. Conclusions: Fascioliasis is a widely prevalent zoonosis among humans and livestock worldwide. Strategies targeting risk factors such as altitude and age are urgently needed for prevention and control of this disease, which will consequently reduce Fasciola infection. Additionally, given the inadequacy or absence of data in some countries, greater attention should be paid to Fasciola infection, with further epidemiological studies focussing on improving data quality.