Development of a Gaussia luciferase immunoprecipitation assay for detecting Schistosoma japonicum infection.

Timely and accurate diagnosis of Schistosoma infection is important to adopt effective strategies for schistosomiasis control. Previously, we demonstrated that Schistosoma japonicum can secret extracellular vesicles and their cargos may serve as a novel type of biomarkers for diagnosing schistosomiasis. Here, we developed a Gaussia luciferase immunoprecipitation assay combined with S. japonicum extracellular vesicle (SjEV) protein to evaluate its potential for diagnosing schistosomiasis. A saposin-like protein (SjSLP) identified from SjEVs was fused to the Gaussia luciferase as the diagnostic antigen. The developed method showed good capability for detecting S. japonicum infection in mice and human patients. We also observed that the method could detect Schistosoma infection in mice as early as 7 days of post-infection, which showed better sensitivity than that of indirect ELISA method. Overall, the developed method showed a good potential for detecting Schistosoma infection particularly for early stage, which may provide an alternative strategy for identify Schistosoma infection for disease control.

Study on genetic characteristics of Cryptosporidium isolates and first report of C. parvum IIdA24G2 subtype in dairy cattle in China.

Cryptosporidium is an important gastrointestinal parasite that can cause mild to severe diarrhea in various vertebrates, including humans and domestic animals. Infection is prevalent in dairy cattle, particularly calves, resulting in diarrhea and increased mortality with significant production losses. However, the prevalence and identity of Cryptosporidium spp. in cattle in Heilongjiang Province is still poorly known. Our study aimed to investigate the prevalence and species and subtype distribution of Cryptosporidium in cattle in the region. In addition, we evaluated the zoonotic potential of Cryptosporidium isolates and assessed possible transmission routes and health effects of this organism. We collected 909 fecal samples from five different farms in Heilongjiang Province between August and September 2022. The samples underwent Cryptosporidium detection by nested PCR and small subunit (SSU) rRNA gene sequence analysis. Four Cryptosporidium species were identified, including C. parvum, C. bovis, C. ryanae, and C. andersoni, with an overall prevalence of 4.4% (40/909). Based on sequence analysis of the 60 kDa glycoprotein gene of C. parvum and C. bovis, three subtypes of C. parvum were identified, namely two previously known subtypes (IIdA19G1 and IIdA20G1), and one novel subtype (IIdA24G2). Two distinct subtype families were identified in C. bovis (XXVId and XXVIe). The high diversity of Cryptosporidium in dairy cattle and the emergence of a novel subtype of C. parvum in Heilongjiang Province suggest that dairy cattle may serve as a significant source of zoonotic cryptosporidiosis infection in this region.

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.

[Quality Control of Appliance Products of Medical Devices for Human Assisted Reproductive Technology].

Infertility has become one of the global problems, with the continuous improvement and penetration of human assisted reproductive technology, appliance products of medical devices for assisted reproductive technology, such as aspiration needles, micromanipulator and embryo transfer catheter, have been widely used. In order to ensure the safety and effectiveness of such products, it is necessary to establish an appropriate quality control system. This paper mainly discusses the quality control points of human assisted reproductive medical devices from the aspects of raw materials, product design, production process, key performance, packaging, preservation and transportation, so as to provide technical reference for the research and development, production and supervision of such products.

[Safety of Alginate-based Gastric Mucosal Protective Adhesive and Feasibility Study of Its Use as Submucosal Injection].

OBJECTIVE: To study the safety of alginate based gastric mucosal protective adhesive and its feasibility as a submucosal injection. METHODS: The feasibility of using alginate-based gastric mucosal protective gel as submucosal injection was evaluated by in vitro gastric mucosal uplift test in pigs and in vivo gastric mucosal uplift test in rats. The safety of alginate based gastric mucosa protective adhesive was evaluated by cytotoxicity test, acute toxicity test and oral mucosa stimulation test according to GB/T 16886 series standard of biological evaluation of medical devices. RESULTS: After injection of different concentrations of alginate base mucosal protective adhesive solution, the uplift height was significantly higher than that of normal saline (P<0.05). Gastric mucosal protection glue has no cytotoxic oral mucosal irritation or acute toxicity. CONCLUSIONS: Gastric mucosa protector is a promising new medical device product with feasibility and good biocompatibility as submucosal uplift injection agent.

[Quality Control and Safety Evaluation of Culture Medium for Human Assisted Reproduction].

OBJECTIVE: In this paper, the key points of quality control and safety evaluation of human assisted reproductive medium were summarized to provide reference for the establishment of relevant standards and quality control in the future. METHODS: Through literature research, the key factors of quality control and risk control of human assisted reproductive medium were summarized, and the problems in clinical transformation were discussed. RESULTS: It is very important for the development of human assisted reproduction technology to study the active ingredients and their harmful degradation products and drugs in the culture medium of assisted reproduction. CONCLUSIONS: At present, the biggest challenge is to effectively control the quality of the culture medium for human assisted reproduction, establish corresponding inspection methods and quality standards for the key components, ensure the safety and effectiveness during the product shelf life, and thus improve the success rate of human assisted reproduction technology.

[Research Method of Fatigue Performance of Surgically Implanted Heart Valve Stent].

The aim of this study was to evaluate the fatigue property in vitro of a domestic surgically implanted heart valve stent (SIHVS). The domestic SIHVS was used as a testing sample, and a protocol was developed according to the standards GB 12279-2008 and ISO 5840-2:2015. A dynamic fatigue testing machine was used to apply for a 0.365 mm cyclic radial displacement at the end of the SIHVS. The SIHVS were observed after 400 million cycles by a scanning electron microscope. The observed results show that the structure of SIHVS is intact. There are no fracture, no obvious wear, and no loosening of the wire and casing. In the tested stents, no structural distortion, no cracks and obvious pits were observed. This study demonstrated that SIHVS structure meets the expected fatigue performance requirements.

[Lung Examination in Systemic Toxicitytest of Medical Devices].

The lung is an important organ in systemic toxicity test of medical devices and is significant in safety evaluation. Based on the authors' understanding of medical devices, this study provides a brief analysis of the lung examination and common problems in systemic toxicity, so as to provide references for the pre-clinical safety evaluation of medical devices. It should be noted that a reasonable risk assessment should be made after comprehensive assessment for specific medical device products.

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.

[The Embryo Toxicity Research of Manufacturing Water of Medical Device of in Vitro Fertilization Technology].

Water is an important component in liquid medical device products for human assisted reproductive technology. Water traits, conductivity, microbial limits, total organic carbon, easy oxides, heavy metal content, bacterial endotoxin and other indicators have an important impact on sperm, egg and embryo development in vitro, so for such products, the quality of water control is extremely important. The production water for producing such products is generally prepared by MilliQ purification system. In this research, we used four different types of water to fabricate the IVF liquids. It included deionized reverse osmosis water, ultra purified water and ultra purified water without endotoxin or nucleic acid, and compared with tap water. The in vitro rat embryo test system was used to study the embryotoxicity of this four different culture liquid production waters. From the result, the group of the super purified water without endotoxin and nucleic acid has the best result of the embryo formation rate, the number of total cell number and the inner cell number. This study proved the importance of removing endotoxin and nucleic acid from the water used for the preparation of the liquid products for assisted reproduction, and provided the basis for the selection of water quality for the liquid products for assisted reproduction.

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.

[Research Progress of Skin-substitute and Its Effectiveness Evaluation].

Skin-substitute has developed rapidly and it is essential to evaluate its effectiveness before clinical use.This paper reviews the development of skin-substitute,methods to evaluate the effectiveness of skin-substitute and relevant standards.This paper highlights the in vitro and in vivo means of preclinical effectiveness evaluation and provides theoretical basis for preclinical safety evaluation of skin-substitute.

Backscattered electron imaging and electron backscattered diffraction in the study of bacterial attachment to titanium alloy structure.

The application of secondary electron (SE) imaging, backscattered electron imaging (BSE) and electron backscattered diffraction (EBSD) was investigated in this work to study the bacterial adhesion and proliferation on a commercially pure titanium (cp Ti) and a Ti6Al4V alloy (Ti 64) with respect to substrate microstructure and chemical composition. Adherence of Gram-positive Staphylococcus epidermidis 11047 and Streptococcus sanguinis GW2, and Gram-negative Serratia sp. NCIMB 40259 and Escherichia coli 10418 was compared on cp Ti, Ti 64, pure aluminium (Al) and vanadium (V). The substrate microstructure and the bacterial distribution on these metals were characterised using SE, BSE and EBSD imaging. It was observed that titanium alloy-phase structure, grain boundaries and grain orientation did not influence bacterial adherence or proliferation at microscale. Adherence of all four strains was similar on cp Ti and Ti 64 surfaces whilst inhibited on pure Al. This work establishes a nondestructive and straight-forward statistical method to analyse the relationship between microbial distribution and metal alloy structure.

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.

Tissue Fluid Enzyme-Linked Immunosorbant Assay for Piglets Experimentally Infected with Toxoplasma gondii and Survey on Local and Imported Pork in Korean Retail Meat Markets.

To investigate the prevalence of Toxoplasma gondii in pork on the market in Korea, an in-house enzyme-linked immunosorbent assay for tissue fluid (CAU-tf-ELISA) was developed using a soluble extract of T. gondii RH strain tachyzoites. As the standard positive controls, the piglets were experimentally infected with T. gondii: Group A (1,000 cysts-containing bradyzoites), Group B (500 cysts-containing bradyzoites) and Group C (1.0×103 or 1.0×104 tachyzoites). The CAU-tf-ELISA demonstrated infection intensity-dependent positivity toward tissue fluids with average cut-off value 0.15: 100% for Group A, 93.8% for Group B and 40.6% for Group C. When tissue-specific cut-off values 0.066-0.199 were applied, CAU-tf-ELISA showed 96.7% sensitivity, 100% specificity, 100% positive and 90.0% negative predictive values. When compared with the same tissue fluids, performance of CAU-tf-ELISA was better than that of a commercial ELISA kit. Of the 583 Korea domestic pork samples tested, anti-T. gondii antibodies were detected from 9.1% of whole samples and 37.9% from skirt meat highest among pork parts. In the 386 imported frozen pork samples, 1.8% (skirt meat and shoulder blade) were positive for anti-T. gondii antibodies. In Korea, prevalence of anti-T. gondii antibodies in the pork on retail markets appeared high, suggesting that regulations on pig farming and facilities are necessary to supply safe pork on the tables.

[Bioresorbable Vascular Scaffold: A Focused Review on Development and Preclinical Studies].

Bioresorbable vascular scaffolds(BVS) are new treatment strategies of percutaneous coronary intervention. They have been introduced to overcome limitations of bare metal stents (BMS) and drug-eluting stents(DES), since they provide temporary scaffolding and then disappear, liberate the treated vessel from cage. In this article, we review the current status and problems of BVS, various tests required before gaining regulatory approval for clinical use.

[Cell Count of Mouse Blastocyst on Pre-clinical Evaluation of Safety of Medical Devices in Assisted Reproductive Technologies].

Various types of medical devices used in assisted reproductive technologies (ART) should be detected for their safety by strict biological assays. Mouse embryo assay(MEA)has been recognized as one of the most important and standardized methods with the threshold more than 80% of blastocyst formation rate (BR) after 96 h culture of fertilized eggs. The disadvantage using BR for embryonic quality control has been concerned as it is ubiquitously dependent of embryonic morphology and the detailed data including molecular and genetic information is obviously missing and incomplete. This leads to the urgent requirement for more sensitive and efficient assessments for the quality control of ART. This study evaluated the reliability of an immunofluorescent MEA by counting total cell and differential number of the cells in the inner cell mass (ICM) and trophectoderm (TE) in the blastocyst. This method improved the traditional MEA, provided a sensitive and powerful platform to assess embryonic developmental viability and should be suggested as a standard assay to be globally used for the quality control of medical devices and pre-clinical procedures in ART.