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The apicomplexan Eimeria ovinoidalis is distributed worldwide. It can cause clinical coccidiosis, which is one of the most pathogenic species in sheep, reducing growth rates and resulting in significant economic losses in the industry. Its principal clinical sign is profuse diarrhoea in young animals. In this study, we established a model of E. ovinoidalis infection in lambs to understand its pathogenicity and evaluate the gut microbiota and fecal metabolite profiles. Specifically, we observed a significant shift in the abundance of bacteria and disrupted metabolism in lambs. Especially during the peak period of excrete oocysts, it promoted the reproduction of some harmful bacteria in Proteobacteria and Actinobacteriota, and reduced the abundance of beneficial bacteria such as Lachnospiraceae and Rikenellaceae. In the later stage of the patent period, the abundance of harmful bacteria in the intestine decreased, the abundance of beneficial bacteria which could produce anti-inflammatory substances began to increase, and the abundance and diversity of intestinal flora also tended to parallel with the control group. Coccidia infection could lead to the increase of differential metabolites and metabolic pathways between infected and control group, but the difference decreased with time. During the peak period of excrete oocysts, although the antimicrobial metabolites such as Lividamine were up-regulated, the excess of these metabolites could still induce the production of endotoxin, while Butanoic acid and other anti-inflammatory metabolites decreased significantly. A metabolomics analysis showed that E. ovinoidalis infection altered metabolites and metabolic pathways, with biosynthesis of unsaturated fatty acids, Teichoic acid biosynthesis and Butanoate metabolism as the major disrupted metabolic pathways. Details of the gut microbiota and the metabolome after infection with E. ovinoidalis may aid in the discovery of specific diagnostic markers and help us understand the changes in parasite metabolic pathways., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Cryptosporidium parvum is an important zoonotic pathogen that is studied worldwide. MicroRNAs (miRNAs) act as post-transcriptional regulators and may play a key role in modulating host epithelial responses following Cryptosporidium infection. Our previous study has shown that C. parvum downregulates the expression of miR-181d through the p50-dependent TLRs/NF-κB pathway. However, the mechanism by which miR-181d regulates host cells in response to C. parvum infection remains unclear. The present study found that miR-181d downregulation inhibited cell apoptosis and increased parasite burden in HCT-8 cells after C. parvum infection. Bioinformatics analysis and luciferase reporter assays have shown that BCL2 was a target gene for miR-181d. Moreover, BCL2 overexpression and miR-181d downregulation had similar results. To further investigate the mechanism by which miR-181d regulated HCT-8 cell apoptosis during C. parvum infection, the expression of molecules involved in the intrinsic apoptosis pathway was detected. Bax, caspase-9, and caspase-3 expression was decreased at 4, 8, 12, and 24 hpi and upregulated at 36 and 48 hpi. Interfering with the expression of miR-181d or BCL2 significantly affected the expression of molecules in the intrinsic apoptosis pathway. These data indicated that miR-181d targeted BCL2 to regulate HCT-8 cell apoptosis and parasite burden in response to C. parvum infection via the intrinsic apoptosis pathway. These results allowed us to further understand the regulatory mechanisms of host miRNAs during Cryptosporidium infection, and provided a theoretical foundation for the design and development of anti-cryptosporidiosis drugs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The microRNAs (miRNAs) of their hosts play an important role in regulating both the innate and adaptive immune responses to Cryptosporidium parvum infection. The mechanisms of autophagy and apoptosis are important components of the defense system against C. parvum infection. In this study, we investigate the role of miRNA-199a-3p in regulating MTOR-mediated autophagy and apoptosis in HCT-8 cells induced by C. parvum. The expression of miR-199a-3p increased at 3, 6 and 12 hours postinfection (hpi) but decreased at 24 and 48 hpi. The upregulation of miR-199a-3p promoted autophagy and apoptosis and limited the parasite burden in HCT-8 cells after C. parvum infection. The downregulation of miR-199a-3p inhibited the autophagy and apoptosis induced by C. parvum and enhanced the parasite burden in HCT-8 cells. A luciferase reporter showed that MTOR was a target gene of miR-199a-3p. Suppressed expression of MTOR by small interfering RNA (siRNA) promoted autophagy and apoptosis and limited C. parvum burden in HCT-8 cells. Co-transfection with miR-199a-3p inhibitor or si-mTOR revealed that miR-199a-3p regulates autophagy and apoptosis in HCT-8 cells through MTOR, to resist C. parvum infection. In conclusion, intestinal epithelial cells defend against C. parvum infection by regulating their autophagy and apoptosis through the miR-199a-3p-MTOR axis., (© 2024. The Author(s).)

Blastocystis sp., Enterocytozoon bieneusi, and Giardia duodenalis are three common zoonotic intestinal parasites, and cattle are important hosts of these three intestinal protozoa. In this study, 1632 fecal samples were collected from dairy farms in Heilongjiang Province, China, and screened for Blastocystis sp., E. bieneusi, and G. duodenalis using polymerase chain reaction. Of these, 149 (9.13%) were positive for three zoonotic pathogens, including 104 (6.40%), 22 (1.35%), and 23 (1.41%) for Blastocystis sp., E. bieneusi, and G. duodenalis, respectively. Based on partial SSU rRNA gene sequencing analysis, 104 positive samples of Blastocystis sp. were found, and a total of nine known subtypes were identified, including ST10 (61), ST3 (18), ST14 (6), ST26 (7), ST24 (3), ST25 (2), ST1 (2), ST5 (2), and ST21 (1). Among these, three subtypes (ST1, ST3, and ST5) were recognized as zoonotic subtypes, and two subtypes (ST10 and ST14) were specific to animals. All 23 Giardia duodenalis-positive samples belonged to assemblage E (n = 23) based on sequenced beta-giardin (bg) and triosephosphate isomerase (tpi) genes. Three known genotypes of E. bieneusi, namely J (n = 9), I (n = 6), and BEB4 (n = 7), were identified by sequence analysis of the internal transcriptional spacer region gene. Our study provides basic data for prevention and control in Heilongjiang Province; however, further research is required to better understand the prevalence and public health significance of these pathogens in the Heilongjiang region., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier B.V.)

Cryptosporidium spp. and G. duodenalis often infect humans, cats, and other mammals, causing diarrhea and being responsible for numerous outbreaks of waterborne and foodborne infections worldwide. The rapid increase in the number of pet cats poses a substantial public health risk. However, there were few reports about the infection of Cryptosporidium spp. and G. duodenalis infections in pet cats in Henan Province, central China. Thus, to understand the prevalence and genetic distribution of Cryptosporidium spp. and G. duodenalis in pet cats, and to evaluate the zoonotic potential, possible transmission routes and public health implications of isolates, fecal samples (n = 898) were randomly collected from pet cats in 11 cities in Henan Province, central China. Nested PCR based on the SSU rRNA gene and bg gene was used to the prevalence of Cryptosporidium spp. and G. duodenalis, respectively. The prevalence was 0.8 % (7/898) and 2.0 % (18/898) for Cryptosporidium spp. and G. duodenalis respectively. Additionally, the Cryptosporidium spp. positive isolates were identified as C. parvum subtype IIdA19G1 by gp60 gene. In the present study, the IIdA19G1 subtype was discovered in pet cats for the first time in China, enriching the information on the host type and geographical distribution of Cryptosporidium spp. in China. For G. duodenalis, a total of 18 G. duodenalis positive samples were identified, belonging to four assemblages: a zoonotic assemblage A1 (4/898), three host-specific assemblages C (8/898), D (5/898), and F (1/898). Interestingly, we found that pet cats infected with Cryptosporidium spp. and G. duodenalis are more likely to experience emaciation symptoms compared to the negative group. More importantly, the prevalence of Cryptosporidium spp. and G. duodenalis detected in the present study were low, but the subtype IIdA19G1 of Cryptosporidium spp. and the assemblages A1, C, D, and F of G. duodenalis have the potential for zoonotic transmission. Thus, we should focus on preventing and controlling the risk of cross-species transmission that may occur in pet cats in Henan Province., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Enterocytozoon bieneusi features high genetic diversity among host species and environmental sources and over 500 genotypes in 11 phylogenetic groups have been defined. Here we investigated 291 small rodents in Heilongjiang province, northeast China, for the presence of E. bieneusi by PCR of the ribosomal internal transcribed spacer (ITS). Nine of 60 (15.0 %) gray squirrels from a park in Harbin, 120 of 201 (59.7 %) guinea pigs from a pet shop in Harbin, and two of 30 (6.7 %) peridomestic rats from a pasture in Qiqihar were positive for the parasite. Six known genotypes (EbpB, SCC-1, SCC-2, D, S7 and HLJ-CP1) and two novel genotypes (NESQ1 and NEGP1) were identified by sequence analysis of the ITS, with EbpB, SCC-1, SCC-2 and NESQ1 found in squirrels, D, S7 and NEGP1 in guinea pigs, and EbpB and HLJ-CP1 in rats. Widespread distribution of human-infective Group 10 genotype S7 and Group 1 genotype D in guinea pigs raised our concerns about the importance of pet animals as zoonotic reservoirs of microsporidiosis. Co-occurrence of Group 1 genotypes D and HLJ-CP1 in cancer patients and rodents in Heilongjiang indicated a possibility of zoonotic transmission. The host range of Group 1 genotype EbpB previously considered pig-adapted was extended. A potential variant of genotype S7, namely NESQ1, went into the existing Group 10 in phylogenetic analysis. The other new genotype, NEGP1, was clustered in an undefined clade we proposed as Group 15. With the emerging epidemiologic evidence, the host specificity of existing E. bieneusi genotypes is now being challenged., Competing Interests: Declaration of competing interest The authors “Shuning Jiang, Shui Yu, Yaoyu Feng, Longxian Zhang, Monica Santin, Lihua Xiao, and Wei Li” of this study declare no competing interests. Signed by the authors listed in the manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cyclospora cayetanensis (C. cayetanensis) is a significant pathogen that causes diarrheal illness and causes large foodborne diarrhea outbreaks in the USA and Canada. However, there is currently a lack of published meta-analysis on the prevalence of C. cayetanensis infection in the global population. A real estimation of a disease prevalence should always be done on the basis of studies designed for that purpose. We conducted a comprehensive search of various databases for articles pertaining to the prevalence of C. cayetanensis infection in humans, spanning from the inception of these databases to March 10, 2023. Utilizing a random effects model, we estimated the prevalence of C. cayetanensis infection in humans. Our analysis included a total of 150 datasets sourced from 42 different countries, which were ultimately selected for the final quantitative assessment. The prevalence of C. cayetanensis infection in humans worldwide was estimated to be 3.4 % (5636/166,611). Notably, Africa exhibited the highest prevalence rate at 5.9 % (606/11,068). Further subgroup analysis revealed a significantly higher infection rate in humans residing in low-income countries (7.6 %, 83/921) compared to those in lower-middle-income countries (4.8 %, 3280/48,852), upper-middle-income countries (2.9 %, 2194/99,419), and high-income countries (0.4 %, 79/17,419). The results indicate that the global prevalence of C. cayetanensis infection in humans is relatively low, despite its extensive geographical distribution and children were found to be more susceptible to C. cayetanensis infection compared to those adults. Sensitivity analysis revealed that one study significantly affects the prevalence of C. cayetanensis, which was adjusted to 2.9 % (4017/160,049; 95 % CI: 2.7-3.1 %) by excluding this study. The findings highlight the relatively high prevalence of C. cayetanensis infection in low-income countries and among humans with diarrhea, particularly in Africa. Consequently, routine surveillance for intestinal protozoa is crucial in these regions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The objective of the present study was to develop a real-time PCR (qPCR) technique for the diagnosis of Eimeria spp. in Ovis aries and Capra hircus. The qPCR technique was developed using SYBR Green, resulting in a PCR with high sensitivity, specificity, and reproducibility., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Blastocystis is a parasitic protist that can infect humans and various domestic and wild animals. However, there is limited research on the prevalence of this parasite among rodents, particularly those living in pig farm settings. Therefore, to investigate the occurrence, molecular characterization, and zoonotic potential of Blastocystis among rodents within pig farm environments, we conducted an investigation of 227 rodents and shrews from 34 pig farms located in Henan, Shaanxi, and Shanxi provinces of China using nested PCR of the SSU rRNA gene of Blastocystis . The potential transmission and public health implications were also assessed from a One Health perspective. Blastocystis was detected in 86 (37.9%) fecal samples. The highest infection rate was observed among Ruttus norvegicus (73.7%, 42/58), followed by Ruttus tanezumi (30.1%, 41/136), and Mus musculus (12.0%, 3/25). However, it was not detected among individuals with Apodemus agrarius ( n  = 1) and Crocidura shantungensis ( n  = 7) . Five known zoonotic Blastocystis subtypes (ST1-ST5) were identified, with ST4 (51.2%, 44/86) and ST5 (40.7%, 35/86) being the predominant ones, followed by ST1 (3.5%, 3/86), ST3 (3.5%, 3/86), and ST2 (1.2%, 1/86). ST4 was prevalent among R. norvegicus (83.3%, 35/42), while ST5 dominated R. tanezumi (70.7%, 29/41). Furthermore, ST5 exhibited the widest distribution at pig farm level, accounting for 65.0% (13/20) of Blastocystis -positive pig farms. This investigation presents the first documented Blastocystis infection in R. tanezumi and M. musculus , highlighting the predominant presence of the zoonotic ST5 subtype in rodents for the first time. The results demonstrate that sympatric rodents can serve as natural reservoirs for Blastocystis and play a role in its transmission. These findings provide information on the dynamics of rodent transmission and emphasize the potential public health threat posed by zoonotic Blastocystis subtypes spillover from pig farms., Competing Interests: The authors declare that they have no competing interests., (© 2024 The Authors. Published by Elsevier B.V.)

Background: Cryptosporidium parvum is an apicomplexan zoonotic parasite causing the diarrheal illness cryptosporidiosis in humans and animals. To invade the host intestinal epithelial cells, parasitic proteins expressed on the surface of sporozoites interact with host cells to facilitate the formation of parasitophorous vacuole for the parasite to reside and develop. The gp40 of C. parvum, named Cpgp40 and located on the surface of sporozoites, was proven to participate in the process of host cell invasion., Methods: We utilized the purified Cpgp40 as a bait to obtain host cell proteins interacting with Cpgp40 through the glutathione S-transferase (GST) pull-down method. In vitro analysis, through bimolecular fluorescence complementation assay (BiFC) and coimmunoprecipitation (Co-IP), confirmed the solid interaction between Cpgp40 and ENO1. In addition, by using protein mutation and parasite infection rate analysis, it was demonstrated that ENO1 plays an important role in the C. parvum invasion of HCT-8 cells., Results: To illustrate the functional activity of Cpgp40 interacting with host cells, we identified the alpha-enolase protein (ENO1) from HCT-8 cells, which showed direct interaction with Cpgp40. The mRNA level of ENO1 gene was significantly decreased at 3 and 24 h after C. parvum infection. Antibodies and siRNA specific to ENO1 showed the ability to neutralize C. parvum infection in vitro, which indicated the participation of ENO1 during the parasite invasion of HCT-8 cells. In addition, we further demonstrated that ENO1 protein was involved in the regulation of cytoplasmic matrix of HCT-8 cells during C. parvum invasion. Functional study of the protein mutation illustrated that ENO1 was also required for the endogenous development of C. parvum., Conclusions: In this study, we utilized the purified Cpgp40 as a bait to obtain host cell proteins ENO1 interacting with Cpgp40. Functional studies illustrated that the host cell protein ENO1 was involved in the regulation of tight junction and adherent junction proteins during C. parvum invasion and was required for endogenous development of C. parvum., (© 2024. The Author(s).)

Background: Enterocytozoon bieneusi is a zoonotic pathogen widely distributed in animals and humans. It can cause diarrhea and even death in immunocompromised hosts. Approximately 800 internal transcribed spacer (ITS) genotypes have been identified in E. bieneusi. Farmed foxes and raccoon dogs are closely associated to humans and might be the reservoir of E. bieneusi which is known to have zoonotic potential. However, there are only a few studies about E. bieneusi genotype identification and epidemiological survey in foxes and raccoon dogs in Henan and Hebei province. Thus, the present study investigated the infection rates and genotypes of E. bieneusi in farmed foxes and raccoon dogs in the Henan and Hebei provinces., Result: A total of 704 and 884 fecal specimens were collected from foxes and raccoon dogs, respectively. Nested PCR was conducted based on ITS of ribosomal RNA (rRNA), and then multilocus sequence typing (MLST) was conducted to analyze the genotypes. The result showed that infection rates of E. bieneusi in foxes and raccoon dogs were 18.32% and 5.54%, respectively. Ten E. bieneusi genotypes with zoonotic potential (NCF2, NCF3, D, EbpC, CHN-DC1, SCF2, CHN-F1, Type IV, BEB4, and BEB6) were identified in foxes and raccoon dogs. Totally 178 ITS-positive DNA specimens were identified from foxes and raccoon dogs and these specimens were then subjected to MLST analysis. In the MLST analysis, 12, 2, 7 and 8 genotypes were identified in at the mini-/ micro-satellite loci MS1, MS3, MS4 and MS7, respectively. A total of 14 multilocus genotypes were generated using ClustalX 2.1 software. Overall, the present study evaluated the infection of E. bieneusi in foxes and raccoon dogs in the Henan and Hebei province, and investigated the zoonotic potential of the E. bieneusi in foxes and raccoon dogs., Conclusions: These findings expand the geographic distribution information of E. bieneusi' host in China and was helpful in preventing against the infection of E. bieneusi with zoonotic potential in foxes and raccoon dogs., (© 2024. The Author(s).)

Cryptosporidium is a gastro-intestinal protozoan parasite that has been found to infect both humans and livestock. This study investigated the parasite in 998 fecal samples from Bangladeshi children ( n  = 299) and calves ( n  = 699) to determine its prevalence, genetic variation, and zoonotic importance. The nested PCR and sequencing of the SSU rRNA gene in the samples showed a Cryptosporidium infection rate of 2.3% (7/299) in children and 15.7% (110/699) in calves. Statistical analysis revealed insignificant variations in Cryptosporidium infections among children across age, gender, and study area, while in calves, the infection rate significantly differed based on location and breed. Genotyping of seven human isolates of Cryptosporidium confirmed C. hominis ( n  = 5) and C. parvum ( n  = 2). After characterizing 110 Cryptosporidium isolates from calves, C. andersoni ( n  = 55), C. ryanae ( n  = 29), C. bovis ( n  = 14), C. parvum ( n  = 10), C. ubiquitum (n = 1), and C. occultus (n = 1) were identified. Cryptosporidium hominis and C. parvum -positive samples were further subjected to nested PCR and sequencing of the glycoprotein 60 ( gp60 ) gene for subtyping. Four C. hominis subtypes (IaA19R3, IaA23R3, IbA9G3, and IdA15G1) and one C. parvum subtype (IIdA15G1) were observed. In conclusion, Cryptosporidium was prevalent in calves but less common in children in the study locations, and the presence of zoonotic Cryptosporidium species and subtypes in calves raises concerns regarding zoonotic transmission to humans., Competing Interests: The authors confirm that they have no conflict of interest to declare., (© 2024 The Authors.)

The development of vaccines, which induce effective immune responses while ensuring safety and affordability, remains a substantial challenge. In this study, we proposed a vaccine model of a restructured "head-to-tail" dimer to efficiently stimulate B cell response. We also demonstrate the feasibility of using this model to develop a paramyxovirus vaccine through a low-cost rice endosperm expression system. Crystal structure and small-angle X-ray scattering data showed that the restructured hemagglutinin-neuraminidase (HN) formed tetramers with fully exposed quadruple receptor binding domains and neutralizing epitopes. In comparison with the original HN antigen and three traditional commercial whole virus vaccines, the restructured HN facilitated critical epitope exposure and initiated a faster and more potent immune response. Two-dose immunization with 0.5 μg of the restructured antigen (equivalent to one-127th of a rice grain) and one-dose with 5 μg completely protected chickens against a lethal challenge of the virus. These results demonstrate that the restructured HN from transgenic rice seeds is safe, effective, low-dose useful, and inexpensive. We provide a plant platform and a simple restructured model for highly effective vaccine development., Competing Interests: Competing interests statement:Authors J.O., H.Y., and K.L. were employed by the company Wuhan Healthgen Biotechnology Corp., Wuhan, China. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Enterocytozoon bieneusi is an obligate intracellular microsporidian parasite with a worldwide distribution. As a zoonotic pathogen, E. bieneusi can infect a wide range of wildlife hosts through the fecal-oral route. Although the feces of flying squirrels (Trogopterus xanthipes) are considered a traditional Chinese medicine (as "faeces trogopterori"), no literature is available on E. bieneusi infection in flying squirrels to date. In this study, a total of 340 fresh flying squirrel fecal specimens from two captive populations were collected in Pingdingshan city, China, to detect the prevalence of E. bieneusi and assess their zoonotic potential. By nested PCR amplification of the ITS gene, six specimens tested positive, with positive samples from each farm, with an overall low infection rate of 1.8%. The ITS sequences revealed three genotypes, including known genotype D and two novel genotypes, HNFS01 and HNFS02. Genotype HNFS01 was the most prevalent (4/6, 66.7%). Phylogenetic analysis showed that all genotypes clustered into zoonotic Group 1, with the novel genotypes clustering into different subgroups. To our knowledge, this is the first report of E. bieneusi infection in flying squirrels, suggesting that flying squirrels could act as a potential reservoir and zoonotic threat for E. bieneusi transmission to humans in China., (© X. Liu et al., published by EDP Sciences, 2024.)

To investigate the prevalence and molecular characteristics of Cystoisospora sp. in blue fox (Alopex lagopus), Sheather's sugar floatation method was conducted to detect coccidia in 423 fresh fecal samples randomly collected from blue fox farms from three cities in China. The overall prevalence of coccidia was 1.4% (6/423), and three Cystoisospora sp. (Cystoisospora fennechi, Cystoisospora sp. I and Cystoisospora vulpina) were identified by their morphological characteristics. The 18S ribosomal RNA (rRNA) and cytochrome c oxidase subunit I (COI) locus sequences were sequenced for molecular biological identification, homology comparison, and phylogenetic analysis of Cystoisospora sp. by single-oocyst selection technology and multi-locus-nested PCR amplification. At the 18S rRNA and COI loci, C. vulpina had 99.48% and 99.59% homology, respectively, with Cystoisospora canis and Cystoisospora ohioensis from canines. Phylogenetic analysis indicated that C. vulpina was clustered in a clade with Cystoisospora sp. from Canidae, which the relatives are consistent with the hosts. To our knowledge, this is the first report on molecular identification and evolutionary analysis of C. vulpina at two different loci., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Pestiviruses, including classical swine fever virus, remain a concern for global animal health and are responsible for major economic losses of livestock worldwide. Despite high levels of vaccination, currently available commercial vaccines are limited by safety concerns, moderate efficacy, and required high doses. The development of new vaccines is therefore essential. Vaccine efforts should focus on optimizing antigen presentation to enhance immune responses. Here, we describe a simple herringbone-dimer strategy for efficient vaccine design, using the classical swine fever virus E2 expressed in a rice endosperm as an example. The expression of rE2 protein was identified, with the rE2 antigen accumulating to 480 mg/kg. Immunological assays in mice, rabbits, and pigs showed high antigenicity of rE2. Two immunizations with 284 ng of the rE2 vaccine or one shot with 5.12 μg provided effective protection in pigs without interference from pre-existing antibodies. Crystal structure and small-angle X-ray scattering results confirmed the stable herringbone dimeric conformation, which had two fully exposed duplex receptor binding domains. Our results demonstrated that rice endosperm is a promising platform for precise vaccine design, and this strategy can be universally applied to other Flaviviridae virus vaccines., (© 2023 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Giardia duodenalis is one of the major causes of diarrhea among humans, especially in young children. Statistical analysis revealed that the pooled prevalence of G. duodenalis in humans, dogs, and cats was 9.72% (10,921/112383), 15.60% (7510/48140), and 14.53% (1125/7740), respectively. Unquestionably, the canine-specific assemblages C and D and the feline-specific assemblage F were the dominant genotypes in dogs and cats, respectively. Additionally, the prevalence of zoonotic G. duodenalis assemblages (A and B) in dogs and cats was 23.07% (875/3792) and 41.42% (169/408), respectively, implying that the potential transmission of G. duodenalis from dogs and cats to human infection cannot be ignored. The highest frequency of potentially zoonotic assemblages was found among working dogs (3.55%, 25/705) and the 1-5 age group (22.92%, 11/48). In summary, dogs and cats have a significant role in the zoonotic transmission of G. duodenalis due to their close contact with humans and the higher frequency presence of zoonotic assemblages. Further studies are necessary to explore the presence of G. duodenalis among humans and animals and in environmental samples. Researchers should adopt a one-health approach to gain a deeper understanding of G. duodenalis in dogs and cats and potential transmission routes to humans., Competing Interests: The authors declare that they have no competing interests., (© 2023 The Authors.)

Cryptosporidium is a highly pathogenic water and food-borne zoonotic parasitic protozoan that causes severe diarrhea in humans and animals. Apicomplexan parasites invade host cells via a unique motility process called gliding, which relies on the parasite's microfilaments. Actin depolymerizing factor (ADF) is a fibrous-actin (F-actin) and globular actin (G-actin) binding protein essential for regulating the turnover of microfilaments. However, the role of ADF in Cryptosporidium parvum (C. parvum) remains unknown. In this study, we preliminarily characterized the biological functions of ADF in C. parvum (CpADF). The CpADF was a 135-aa protein encoded by cgd5_2800 gene containing an ADF-H domain. The expression of cgd5_2800 gene peaked at 12 h post-infection, and the CpADF was located in the cytoplasm of oocysts, middle region of sporozoites, and cytoplasm of merozoites. Neutralization efficiency of anti-CpADF serum was approximately 41.30%. Actin sedimentation assay revealed that CpADF depolymerized but did not undergo cosedimentation with F-actin and its ability of F-actin depolymerization was pH independent. These results provide a basis for further investigation of the roles of CpADF in the invasion of C. parvum., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: With the promotion of "One Health," the health of animals and their impact on the environment have become major concerns recently. Widely distributed in China, the whooper swans (Cygnus cygnus) and black swans (Cygnus atratus) are not only important to the ecological environment, but they may also potentially influence public health security. The metagenomic approach was adopted to uncover the impacts of the gut microbiota of swans on host and public health., Results: In this study, the intestinal microbiome and resistome of migratory whooper swans and captive-bred black swans were identified. The results revealed similar gut microbes and functional compositions in whooper and black swans. Interestingly, different bacteria and probiotics were enriched by overwintering whooper swans. We also found that Acinetobacter and Escherichia were significantly enriched in early wintering period swans and that clinically important pathogens were more abundant in black swans. Whooper swans and black swans are potential reservoirs of antibiotic resistance genes (ARGs) and novel ARGs, and the abundance of novel ARGs in whooper swans was significantly higher than that in black swans. Metagenomic assembly-based host tracking revealed that most ARG-carrying contigs originated from Proteobacteria (mainly Gammaproteobacteria)., Conclusions: The results revealed spatiotemporal changes in microbiome and resistome in swans, providing a reference for safeguarding public health security and preventing animal epidemics., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Cryptosporidiosis is a zoonotic disease in humans and animals that is caused by infection with the oocysts of Cryptosporidium. MicroRNAs (miRNAs) are important players in regulating the innate immune response against parasitic infection. Public miRNAs data for studying pathogenic mechanisms of cryptosporidiosis, particularly in natural hosts, are scarce. Here, we compared miRNA profiles of the glandular stomach of C. muris-infected and uninfected BALB/c mice using microarray sequencing. A total of 10 miRNAs (including 3 upregulated and 7 downregulated miRNAs) with significant differential expression (|FC| ≥ 2 and P value < 0.05) were identified in the glandular stomach of BALB/c mice 8 h after infection with C. muris. MiRWalk and miRDB online bioinformatics tools were used to predict the target genes of differentially expressed miRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed to annotate the target genes. GO analysis indicate that gene transcription-related and ion transport-related GO terms were significantly enriched. In addition, the KEGG analyses showed that the target genes were strongly related to diverse types of tumor disease progression and anti-pathogen immunity pathways. In the current study, we firstly report changes in miRNA expression profiles in the glandular stomach of BALB/c mice at the early phase of C. muris invasion. This dysregulation in miRNA expression may contribute to our understanding of cryptosporidiosis pathology. This study provides a new perspective on the miRNA regulatory mechanisms of cryptosporidiosis, which may help in the development of effective control strategies against this pathogen., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Cryptosporidium is a protozoan parasite capable of infecting humans and animals and is a leading cause of diarrheal disease and early childhood mortality. The molecular mechanisms underlying invasive infection and its pathogenesis remain largely unknown. To better understand the molecular mechanism of the interaction between C. parvum and host cells, we profiled the changes of host cells membrane proteins extracted using native membrane protein extraction kit between C. parvum-infected HCT-8 cells and the control group after C. parvum infected 6 h combined with quantitative Tandem Mass Tags (TMT) liquid chromatography-dual mass spectrometry proteomic analysis. Among the 4844 quantifiable proteins identified, the expression levels of 625 were upregulated, and those of 116 were downregulated at 6 h post-infection compared with controls (1.5-fold difference in abundance, p < 0.05). Enrichment analysis of the function, protein domain and Kyoto Encyclopedia of Genes and Genomes pathway of the differentially expressed proteins revealed that the differentially expressed proteins were mainly related to biological functions related to the cytoskeleton and cytoplasmic matrix. We also found that infection with C. parvum may destroy HCT-8 intercellular space adhesion. Six proteins were further verified using quantitative real-time reverse transcription polymerase chain reaction and western blotting. Through systematic analysis of proteomics related to HCT-8 cell membranes infected by C. parvum, we found many host membrane proteins that can serve as potential receptors in C. parvum adhesion or invasion. C. parvum infection destroyed host cell barrier function and caused extensive changes in host cytoskeleton proteins, providing a deeper understanding of the molecules and their functions involved in the host-C. parvum interaction. SIGNIFICANCE: There is a lack of systematic research on the molecular mechanisms underlying the interaction of C. parvum with host cells. Changes of host cell membrane proteins after C. parvum infection may be used to examine the host cell receptors for parasite adhesion and invasion, and how the parasite interacts with these receptors. It is of great significance that host cells undergo membrane fusion to mediate invasion. Through proteomic studies on the host cell membrane after infection with HCT-8 cells by C. parvum, we observed disruption of the host cell cellular barrier function and widespread alteration of host cytoskeletal proteins caused by C. parvum infection, providing a deeper understanding of the molecules and their functions involved in host-C. parvum interaction., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflicts of interest to report regarding the present study., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Coccidiosis caused by Eimeria is one of the most common diseases in domestic rabbits (Oryctolagus cuniculus f. domesticus), with 11 Eimeria species in domestic rabbits recognized internationally. To identify Eimeria species more accurately, a method based on the molecular characteristics of a single oocyst with multiple gene loci was established by combining morphological and molecular biology. The results showed that the total infection rate of Eimeria in domestic rabbits was 44.2 % (152/344). Ten Eimeria species were identified in domestic rabbits based on morphological characteristics, namely Eimeria vejdovskyi (39.5 %, 136/344), E. magna (18.0 %, 62/344), E. perforans (17.4 %, 60/344), E. intestinalis (12.5 %, 43/344), E. media (11.9 %, 41/344), E. coecicola (4.4 %, 15/344), E. irresidua (3.8 %, 13/344), E. exigua (2.6 %, 9/344), E. stiedai (2.3 %, 8/344), and E. piriformis (1.5 %, 5/344). The molecular biological identification of Eimeria in domestic rabbits was conducted through single oocyst selection and nested polymerase chain reaction amplification with multiple gene loci. We obtained the sequences of the 18S rRNA, ITS-1 and COI gene loci of E. magna, E. perforans, E. vejdovskyi, E. media, E. intestinalis, and E. coecicola. The results showed that the molecular biology and morphological identification results of single oocysts were consistent and could be used for the molecular identification of Eimeria at the single oocyst level. This study provides an efficient tool for identification of Eimeria in domestic rabbits and the population genetic study of Eimeria in domestic rabbits., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflicts of interest to report regarding the present study., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Rhipicephalus microplus is a major threat to the cattle industry worldwide. The intensive use of acaricides and repellents has resulted in drug resistance. Hence, effective and eco-friendly pest control alternatives are urgently needed, especially from natural plant resources. In this study, the acaricidal and repellent activities of nine herbs against the larvae and eggs of R. microplus were evaluated. The results showed that ethanol extracts of star anise (Illicium verum), chaulmoogra (Hydnocarpus anthelmintica), motherwart (Leonurus artemisia), mandarin orange peel (citri reticulatae pericarpium, i.e., peel of Citrus reticulata fruit), and stemona (Stemona sessilifolia) had good contact acaricidal activities of 100, 98, 94, 88 and 86%, respectively, whereas star anise and clove (Syzygium aromaticum) had good fumigant acaricidal activities of 98 and 96%, respectively. The hatching inhibition rate of star anise against R. microplus eggs was 100%. All nine herbs had good real-time repellent rates, but only castor bean and star anise had repellent effects after 48 h (81.3 and 79.6%, respectively). This is the first report of the acaricidal and repellent activities of these medicinal herbs against R. microplus. Ethanol extracts of these herbs might be considered as potential alternatives to chemical acaricides for control of R. microplus., (© 2023. The Author(s).)

Giardia is a genus of flagellated protozoan parasites that infect the small intestine of humans and animals, causing the diarrheal illness known as giardiasis. Giardia exhibits significant genetic diversity among its isolates, which can have important implications for disease transmission and clinical presentation. This diversity is influenced by the coevolution of Giardia with its host, resulting in the development of unique genetic assemblages with distinct phenotypic characteristics. Although panmixia has not been observed, some assemblages appear to have a broader host range and exhibit higher transmission rates. Molecular diagnostic methods enable researchers to examine the genetic diversity of Giardia populations, enhancing our understanding of the genetic diversity, population structure, and transmission patterns of this pathogen and providing insights into clinical presentations of giardiasis., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Cryptosporidium is an important zoonotic pathogen that causes diarrhea in humans and animals, and a leading cause of diarrhea morbidity and mortality in children under 5 years old. However, the meta-analysis of Cryptosporidium infection in children in China has not been published. We searched the databases for articles published on the prevalence of Cryptosporidium infection in children in China since the inception of these databases to 31 October 2022. The prevalence of Cryptosporidium infection in children was estimated using a random effects model. The results showed that 111 datasets from 24 provinces were selected for the final quantitative analysis. The estimated pooled Cryptosporidium infection prevalence in children in China was 2.9% (3300/126,381). The highest prevalence rate was in southwestern China (4.8%, 365/7766). Subgroup analysis indicated that the Cryptosporidium infection rate in children aged < 3 years (4.9%, 330/8428) was significantly higher than that in children aged 3-6 years (2.5%, 609/26,080) and >6 years (2.6%, 647/27,586). Six Cryptosporidium species were detected in children in China from the selected studies. C. hominis was the dominant species (77.1%, 145/188) and the proportions of subgenotype IaA14R4 of C. hominis was highest (42.8%, 62/145). The findings suggest that Chinese children is in a low level of Cryptosporidium infection, however, the geographical distribution of the infection is extensive. We suggest that measures should be taken to ensure the healthy growth of Chinese children by improving the water environment, increasing public health facilities, strengthening children's health education, and developing sound Cryptosporidium infection control programs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the study reported in this manuscript., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: A close connection between a protozoan parasite and the balance of the other gut microbes of the host has been demonstrated. The calves may be naturally co-infected with many parasites, and the co-effects of parasites on other intestinal microbes of calves remain unclear. This study aims to preliminarily reveal the relationship between intestinal parasites and other intestinal microbes in calves., Methods: Fecal samples were collected from four calves with bloody diarrhea, four calves with watery diarrhea, and seven normal calves, and the microbial flora of the samples were analyzed by whole-genome sequencing. Protozoal parasites were detected in the metagenome sequences and identified using polymerase chain reaction (PCR)., Results: Cryptosporidium, Eimeria, Giardia, Blastocystis, and Entamoeba were detected by metagenomic analysis, and the identified species were Giardia duodenalis assemblage E, Cryptosporidium bovis, Cryptosporidium ryanae, Eimeria bovis, Eimeria subspherica, Entamoeba bovis, and Blastocystis ST2 and ST10. Metagenomic analysis showed that the intestinal microbes of calves with diarrhea were disordered, especially in calves with bloody diarrhea. Furthermore, different parasites show distinct relationships with the intestinal microecology. Cryptosporidium, Eimeria, and Giardia were negatively correlated with various intestinal bacteria but positively correlated with some fungi. However, Blastocystis and Entamoeba were positively associated with other gut microbes. Twenty-seven biomarkers not only were significantly enriched in bloody diarrhea, watery diarrhea, and normal calves but were also associated with Eimeria, Cryptosporidium, and Giardia. Only Eimeria showed a distinct relationship with seven genera of bacteria, which were significantly enriched in the healthy calves. All 18 genera of fungi were positively correlated with Cryptosporidium, Eimeria, and Giardia, which were also significantly enriched in calves with bloody diarrhea. Functional genes related to parasites and diseases were found mainly in fungi., Conclusions: This study revealed the relationship between intestinal protozoan parasites and the other calf gut microbiome. Different intestinal protozoan parasites have diametrically opposite effects on other gut microecology, which not only affects bacteria in the gut, but also is significantly related to fungi and archaea., (© 2023. The Author(s).)

Background: Cryptosporidium is second only to rotavirus as a cause of moderate-to-severe diarrhea in young children. There are currently no fully effective drug treatments or vaccines for cryptosporidiosis. MicroRNAs (miRNAs) are involved in regulating the innate immune response to Cryptosporidium parvum infection. In this study, we investigated the role and mechanism of miR-3976 in regulating HCT-8 cell apoptosis induced by C. parvum infection., Methods: Expression levels of miR-3976 and C. parvum burden were estimated using real-time quantitative polymerase chain reaction (RT-qPCR) and cell apoptosis was detected by flow cytometry. The interaction between miR-3976 and B-cell lymphoma 2-related protein A1 (BCL2A1) was studied by luciferase reporter assay, RT-qPCR, and western blotting., Results: Expression levels of miR-3976 were decreased at 8 and 12 h post-infection (hpi) but increased at 24 and 48 hpi. Upregulation of miR-3976 promoted cell apoptosis and inhibited the parasite burden in HCT-8 cells after C. parvum infection. Luciferase reporter assay indicated that BCL2A1 was a target gene of miR-3976. Co-transfection with miR-3976 and a BCL2A1 overexpression vector revealed that miR-3976 targeted BCL2A1 and suppressed cell apoptosis and promoted the parasite burden in HCT-8 cells., Conclusions: The present data indicated that miR-3976 regulated cell apoptosis and parasite burden in HCT-8 cells by targeting BCL2A1 following C. parvum infection. Future study should determine the role of miR-3976 in hosts' anti-C. parvum immunity in vivo., (© 2023. The Author(s).)

Blastocystis sp. is a kind of unicellular intestinal commensal which is widely distributed in humans and animals, and frequently found in the people who are in close contact with animals. To investigate the prevalence and evaluate the zoonotic potential of Blastocystis sp. in sheep and goats from Inner Mongolia, China, a total of 1037 samples were collected from them, and subjected to nested PCR amplification based on the small subunit ribosomal RNA (SSU rRNA) gene of Blastocystis sp. The sanger sequencing was used for Blastocystis sp. subtype identification. The results indicated that the average infection rate of Blastocystis sp. was 10.70% [95CI: 8.82%-12.58%] (111/1037), including 11.30% [95CI: 7.96%-14.64%] for sheep (39/345) and 10.40% [95CI: 8.13%-12.67%] for goats (72/692). Five Blastocystis subtypes (ST5, ST10, ST14, ST21 and ST26) were identified in the present study. Among them, ST10 was the most dominant subtype in sheep and goats, accounting for 70.27% (78/111) of the total identified positive samples. This is the first report regarding Blastocystis sp. subtypes ST21 and ST26 in goats in China. This study has provided a detail epidemiological data on the prevalence and subtypes distribution of Blastocystis sp. in sheep and goats in Inner Mongolia, China. Our results indicated that sheep and goats could be reservoir host for multiple Bastocystis subtypes, including the zoonotic subtypes. Further studies among humans, livestock and wild animals are needed to better understand their role in the spread of Blastocystis sp., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Cryptosporidium spp. are significant opportunistic pathogens causing diarrhoea in humans and animals. Pigs are one of the most important potential hosts for Cryptosporidium . We evaluated the prevalence of Cryptosporidium in pigs globally using published information and a random-effects model. In total, 131 datasets from 36 countries were included in the final quantitative analysis. The global prevalence of Cryptosporidium in pigs was 16.3% (8560/64 809; 95% confidence interval [CI] 15.0–17.6%). The highest prevalence of Cryptosporidium in pigs was 40.8% (478/1271) in Africa. Post-weaned pigs had a significantly higher prevalence (25.8%; 2739/11 824) than pre-weaned, fattening and adult pigs. The prevalence of Cryptosporidium was higher in pigs with no diarrhoea (12.2%; 371/3501) than in pigs that had diarrhoea (8.0%; 348/4874). Seven Cryptosporidium species ( Cryptosporidium scrofarum , Cryptosporidium suis , Cryptosporidium parvum , Cryptosporidium muris , Cryptosporidium tyzzeri , Cryptosporidium andersoni and Cryptosporidium struthioni ) were detected in pigs globally. The proportion of C. scrofarum was 34.3% (1491/4351); the proportion of C. suis was 31.8% (1385/4351) and the proportion of C. parvum was 2.3% (98/4351). The influence of different geographic factors (latitude, longitude, mean yearly temperature, mean yearly relative humidity and mean yearly precipitation) on the infection rate of Cryptosporidium in pigs was also analysed. The results indicate that C. suis is the dominant species in pre-weaned pigs, while C. scrofarum is the dominant species in fattening and adult pigs. The findings highlight the role of pigs as possible potential hosts of zoonotic cryptosporidiosis and the need for additional studies on the prevalence, transmission and control of Cryptosporidium in pigs.

Background: Cryptosporidium is a protozoan parasite causing diarrhoea in humans and animals. Although Cryptosporidium has been found in domestic horses (farmed or kept at pasture), there has been only one published study of Cryptosporidium infections in Chinese racehorses, which was restricted to a very small geographical area., Objectives: To investigate the presence of Cryptosporidium spp. in the faeces of racehorses in China and to perform molecular characterisation of the parasite., Study Design: Cross-sectional., Methods: A total of 621 fresh faecal samples were collected for DNA extraction from racehorses at 17 equestrian clubs from 12 provinces of China from December 2016 to May 2018. All the DNA were analysed for the presence of Cryptosporidium species/genotypes and subtypes by PCR amplification of the small subunit ribosomal RNA and 60 kDa glycoprotein genes respectively., Results: PCR analysis revealed that 11 samples (1.8%) were positive for Cryptosporidium spp. Among them seven samples were identified as C. parvum and four were C. hominis. The C. parvum isolates were identified as subtype IIdA14G1 (n = 4) and IIdA15G1 (n = 3), while all C. hominis isolates were identified as subtype IkA18G1 (n = 4)., Main Limitations: A single faecal sample from each horse was used instead of multiple samples that could improve the detection rates of the parasite., Conclusions: Although Cryptosporidium infection rate was relatively low in the investigated racehorses, the presence of zoonotic subtypes IIdA14G and 1IIdA15G1 of C. parvum and IkA18G1 of C. hominis, suggesting that these animals are a potential source of Cryptosporidium in humans., (© 2022 EVJ Ltd.)

Cryptosporidium is a significant zoonotic pathogen that often occurs in dairy cattle. We conducted a systematic review and meta-analysis of the prevalence of Cryptosporidium parvum infection in dairy calves worldwide to help improve global animal husbandry and public policy implementation. Published articles were obtained from PubMed and Web of Science from January 1, 2000 to December 31, 2021. The prevalence of C. parvum infection in dairy calves was estimated using a random effects model, and the sources of heterogeneity were explored using meta-regression. In total, 118 datasets were included in the final quantitative analysis. The results showed that the global prevalence of C. parvum in dairy calves was 21.9% (7755/42,890; 95% confidence interval: 19.9-23.9%). C. parvum infection was high in pre-weaned dairy calves (24.9%, 6706/29,753) and diarrhea dairy calves (33.6%, 1637/6077). In countries with low dairy stocking density (<10 cows/farm), the prevalence of C. parvum in dairy calves was also relatively low (15.2%, 1960/16,584). Three subtype families [IIa (72.2%, 2293/3177), IId (27.4%, 872/3177), and IIl (0.4%, 12/3177)] were detected in dairy calves globally from selected studies. C. parvum IIa was the dominant zoonotic subtype. In the IIa subtype family of C. parvum, the proportions of subtypes from high to low (top nine) were IIaA15G2R1 (32.4%, 742/2293), IIaA18G3R1 (11.8%, 271/2293), IIaA13G2R1 (8.2%, 187/2293), IIaA16G1R1 (6.4%, 147/2293), IIaA20G1R1 (3.5%, 81/2293), IIaA16G3R1 (3.4%, 78/2293), IIaA17G2R1 (2.7%, 62/2293), IIaA18G1R1 (2.5%, 58/2293), and IIaA15G1R1 (2.4%, 56/2293). In the IId subtype family of C. parvum, the proportions of subtypes (top four) were IIdA19G1 (36.0%, 314/872), IIdA15G1 (27.3%, 238/872), IIdA20G1 (16.2%, 141/872), and IIdA14G1 (13.0%, 113/872). Furthermore, IId is commonly found in China (771/872). The study results indicated that the IIa subtype family is globally prevalent, while IId is found in Asia, Europe, and Africa and IIl is only found in Europe. Diarrhea in dairy calves is associated with C. parvum infection and a significantly higher prevalence is observed in diarrheic calves. Age and stock density are two significant risk factors in the prevalence of C. parvum in dairy calves. The prevention and control of this zoonosis in dairy calves should receive greater attention, especially in regions with a high degree of intensive dairy farming., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the study reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Cryptosporidium parvum has gained much attention as a major cause of diarrhea in the world, particularly in those with compromised immune systems. The data currently available on how the immune system recognizes C. parvum are growing rapidly, but we lack data on the interactions among host major histocompatibility complex (MHC) diversity and parasitic T-cell epitopes. To identify antigenic epitopes in a murine model, we performed systematic profiling of H-2K b -restricted peptides by screening the dominant Cryptosporidium antigens. The results revealed that the glycoprotein-derived epitope Gp40/15-SVF9 induced an immunodominant response in C. parvum-recovered C57BL/6 mice, and injection of the cytotoxic-T-lymphocyte (CTL) peptide with the adjuvant activated peptide-specific CD8 + T cells. Notably, the SVF9 epitope was highly conserved across Cryptosporidium hominis, C. parvum, and many other Cryptosporidium species. SVF9 also formed stable peptide-MHC class I (MHC I) complexes with HLA-A*0201, suggesting cross-reactivity between H-2K b and human MHC I specificities. Crystal structure analyses revealed that the interactions of peptide-MHC surface residues of H-2K b and HLA-A*0201 are highly conserved. The hydrogen bonds of H-2K b -SVF9 are similar to those of a dominant epitope presented by HLA-A*0201, which can be recognized by a public human T-cell receptor (TCR). Notably, we found double conformations in position 4 (P4), 5 (P5) of the SVF9 peptide, which showed high flexibility, and multiple peptide conformations generated more molecular surfaces that can potentially be recognized by TCRs. Our findings demonstrate that an immunodominant C. parvum epitope and its homologs from different Cryptosporidium species and subtypes can benefit vaccine development to combat cryptosporidiosis. IMPORTANCE Adaptive immune responses and T lymphocytes have been implicated as important mechanisms of parasite-induced protection. However, the role of CD8 + T lymphocytes in the resolution of C. parvum infection is largely unresolved. Our results revealed that the glycoprotein-derived epitope Gp40/15-SVF9 induced an immunodominant CD8 + T-cell response in C57BL/6 mice. Crystal structure analyses revealed that the interactions of the H-2K b -SVF9 peptide are similar to those of a dominant epitope presented by HLA-A*0201, which can be recognized by human TCRs. In addition, we found double conformations of the SVF9 peptide, which showed high flexibility and multiple peptide conformations that can potentially be recognized by TCRs.

Cryptosporidium is a leading cause of diarrheal disease and mortality in young children worldwide. Cryptosporidium invades small intestinal epithelial cells and forms a unique intracellular niche, a process that may alter gut microbes and their production metabolites. However, the mechanism of interactions between gut microbes, metabolites, and parasites is poorly understood. Here, we first characterized the impacts of Cryptosporidium infection on gut microbiota using a microbiome-to-metabolome association study. BALB/c mice were gavaged with Cryptosporidium muris, and fecal samples were collected at 0, 7, 14, 21, and 28 days postinfection (dpi) to observe changes in the intestinal microbes of the body during parasite infection. The infected group had a significantly increased relative abundance of bacterial taxa, such as Lachnospiraceae and Prevotella ( P < 0.05), associated with the biosynthesis of short-chain fatty acids (SCFAs). Metabolites related to the metabolic pathways, steroid hormone biosynthesis, and biosynthesis of unsaturated fatty acids pathway were upregulated at 7 dpi, indicating that related metabolites in the biosynthesis of unsaturated fatty acids may be essential for C. muris reproduction in vivo . The metabolites involved in metabolic pathways, bile secretion, and primary bile acid biosynthesis were upregulated at 14 dpi, and we speculate that these metabolites may be critical to the growth and development of Cryptosporidium oocysts in the host. Correlation analysis revealed that Firmicutes bacteria are significantly associated with α-linolenic acid metabolism pathways ( P < 0.05). The gut microbiota changes dynamically, and the metabolites involved in fatty acid and bile acid biosynthesis may play important roles during cryptosporidiosis. Details of the gut microbiota and the metabolome after infection with Cryptosporidium may aid in the discovery of specific diagnostic markers and help us understand the changes in parasite metabolic pathways. IMPORTANCE Cryptosporidiosis is a gastrointestinal disease in humans and animals caused by the protozoan parasite Cryptosporidium. Control and treatment of the disease is challenging due to the lack of sensitive diagnostic tools and effective chemotherapy. The dynamics of gut microbiota and metabolites during Cryptosporidium infection may be the key to finding drugs and targets for parasite infection control. Our results indicate that C. muris infection can disrupt gut microbiota and metabolites, resulting in decreased bacterial abundance at the parasitic site. Unsaturated fatty acid pathway biosynthesis-related metabolites are significantly elevated at the patent period. Interestingly, the metabolite pathway that significantly elevated during peak parasite growth was bile acid, the metabolites of which may be important for the circulation of infection of Cryptosporidium oocysts in the host. The enhancing effects of short-chain fatty acid and bile acid metabolism on the growth and development of Cryptosporidium proposed in this study may provide a theoretical basis for future research on novel drugs and vaccines against this intestinal parasite.

Four species of Giardia out of nine have been identified in rodents based on molecular data: G. muris , G. microti , G. cricetidarum , and G. duodenalis . A total of seven G. duodenalis assemblages (A, B, C, D, E, F, G) have been identified in rodents to date. The zoonotic assemblages A and B are responsible for 74.88% (480/641) of the total identified genotypes in rodents by statistic. For sub-assemblage A in humans, AII is responsible for 71.02% (1397/1967) of the identified sub-assemblages, followed by AI with 26.39% (519/1967) and AIII with 1.17% (23/1967), indicating a significantly greater zoonotic potential for G. duodenalis infections in humans originating from animals. For sub-assemblages of type A in rodents, AI was identified in 86.89% (53/61), and AII in 4.92% (3/61). For assemblage B, 60.84% (390/641) were identified in rodents as having zoonotic potential to humans. In environmental samples, the zoonotic assemblages A and B were responsible for 83.81% (533/636) in water samples, 86.96% (140/161) in fresh produce samples, and 100% (8/8) in soil samples. The same zoonotic potential assemblage A or B simultaneously identified in humans, rodents, and environment samples had potential zoonotic transmission between humans and animals via a synanthropic environment. The infections and zoonotic potential for G. duodenalis were higher in farmed rodents and pet rodents than that in zoo, lab, and wild rodents. In conclusion, the role of rodents in zoonotic transmission of giardiasis should be noticed. In addition to rodents, dogs, cats, wild animals, and livestock could be involved in the zoonotic transmission cycle. This study aims to explore the current situation of giardiasis in rodents and seeks to delineate the role of rodents in the zoonotic transmission of giardiasis from the One Health perspective., Competing Interests: The authors declare no competing interests., (© 2023 The Authors. Published by Elsevier B.V.)

Cryptosporidium spp. are zoonotic intestinal parasites that infect fish, birds, reptiles and mammals. Cryptosporidium spp. are common cause of diarrhea. In this study, a total of 1032 fecal samples were collected from the rectums of sheep and goats. The samples were analyzed using nested polymerase chain reaction (nested PCR) based on the small subunit ribosomal RNA (SSU rRNA) gene of Cryptosporidium spp. The average infection rate of Cryptosporidium spp. was 2.23% (n = 23), and three Cryptosporidium species were identified, namely Cryptosporidium ubiquitum (8/23), Cryptosporidium andersoni (5/23) and Cryptosporidium xiaoi (10/23). Subtyping of C. ubiquitum and C. xiaoi was carried out by DNA sequence analysis of the 60-kDa glycoprotein (gp60) gene. Eight C. ubiquitum isolates were identified as zoonotic subtype XIIa. Nine C. xiaoi isolates were identified as subtypes XXIIIc (n = 1), XXIIIf (n = 3) and XXIIIg (n = 5). Subtype XXIIIg was first found in Chinese sheep. C. ubiquitum subtype XIIa was found in both sheep and goats, suggesting that sheep and goats are important sources of C. ubiquitum infections., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Non-biting flies such as the house fly (Musca domestica), the Australian sheep blowfly (Lucilia cuprina) and the oriental latrine fly (Chrysomya megacephala) may carry many parasites. In the present study, we performed a systematic overview of the different species of parasites carried by non-biting flies, as well as of isolation methods, different geographical distribution, seasonality and risk assessment., Methods: A meta-analysis was carried out with the aim to review the global prevalence of parasite transmission in non-biting flies. A total sample size of 28,718 non-biting flies reported in studies worldwide satisfied the predetermined selection criteria and was included in the quantitative analysis., Results: The global prevalence of parasites in non-biting flies was 42.5% (95% confidence interval [CI] 31.9-53.2%; n = 15,888/28,718), with the highest prevalence found for non-biting flies in Africa (58.3%; 95% CI 47.4-69.3%; n = 9144/13,366). A total of 43% (95% CI 32.1-54.4%; n = 7234/15,282) of house flies (M. domestica), the fly species considered to be the most closely associated with humans and animals, were found with parasites. The prevalence of parasites in the intestine of non-biting flies was 37.1% (95% CI 22.7-51.5%; n = 1045/3817), which was significantly higher than the prevalence of parasites isolated from the body surface (35.1%; 95% CI 20.8-49.4%; n = 1199/3649; P < 0.01). Of the 27 reported parasites, a total of 20 known zoonotic parasites were identified, with an infection rate of 38.1% (95% CI 28.2-48.0%; n = 13,572/28,494)., Conclusions: This study provides a theoretical basis for the public health and ecological significance of parasites transmitted by non-biting flies., (© 2023. The Author(s).)

Blastocystis is a usual intestinal protist that always found in humans and various animals . Currently, the prevalence of Blastocystis in the migratory whooper swan ( Cygnus cygnus ) is unknown. In our research, we aimed to determine the occurrence, prevalence, subtype distribution and dynamic transmission mechanisms of Blastocystis in the migratory whooper swan in China. We also assessed the zoonotic potential of Blastocystis isolates, as well as possible routes of transmission and impact of this organism on One Health perspective. Fecal samples ( n  = 770) were collected from whooper swans inhabiting the Sanmenxia Swan Lake National Urban Wetland Park, China. The overall prevalence of Blastocystis was 11.6% (89/770). We identified 9 subtypes of Blastocystis sp., including 5 zoonotic subtypes [ST1 (Cakir et al., 2019 (8)), ST4 (Selma and Karanis, 2011 (4)), ST5 (Stensvold et al., 2009 (1)), ST6 (Fare et al., 2019 (5)) and ST7(58)] and 3 host-specific subtypes [ST10 (Zhao et al., 2018 (7)), ST14 (Tan et al., 2010 (2)), ST23 (Wang et al., 2018 (3)), and ST25 (Stensvold et al., 2009 (1))]. Subtypes ST4, ST5, ST6, ST10, ST14, ST23, and ST25 were first identified in the whooper swan. Among these subtypes, ST23 and ST25 were identified in birds for the first time, indicating that these subtypes are expanding their host range. So far, this is the first research reporting on the prevalence and subtypes distribution of Blastocystis in the migratory whooper swan in China. The findings obtained in this study will provide new insights into the genetic diversity and transmission routes of Blastocystis , and the possible public health concerns posed by this organism., Competing Interests: The authors have no conflicts of interest to declare., (© 2023 The Authors.)

Blastocystis is a gastrointestinal protozoan parasite commonly reported in humans and animals globally, including poultry, and it can cause zoonotic transmission of blastocystosis. However, comprehensive information is not available on the prevalence, subtype distribution and zoonotic potential of Blastocystis in chickens in China. In this study, a total of 1,000 individual fecal samples of free-range broiler chickens of 4 breeds were collected from 43 farms in 5 cities of Guangdong Province and investigated for the occurrence of Blastocystis infection. Blastocystis was determined by nested PCR analysis of the small subunit ribosomal RNA (SSU rRNA) gene. The overall prevalence was 20.1% (201/1,000) in chicken samples and 69.8% (30/43) in screened farms, and considerable variation in prevalence between farms was evident, with a range of 0 to 76.9%. Population differences of Blastocystis in broilers among sites, breeds, and ages were assessed. The highest infection rates were observed in Yangjiang city (35.8%, 38/106), Sanhuang chickens (29.7%, 104/350), and the >80-day-old chicken group (30.5%, 40/131). DNA sequencing and phylogeny analyses identified 2 zoonotic subtypes, ST6 and ST7. A large predominance was observed for ST7, and genetic polymorphisms were confirmed at the intra-ST7 level with the identification of 5 divergent ST7 types. The incidence of both STs varied largely based on the breed, site, farm, and age. This is the first large-scale study to explore the prevalence and genetic characteristics of Blastocystis in chickens in China. The widespread distribution and avian adaptation of both zoonotic subtypes were demonstrated. The findings of this study highlight a potential threat to humans and will provide a better understanding of the epidemiology and public health impact of poultry Blastocystis., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Cyclosporiasis is an emerging disease caused by Cyclospora cayetanensis , which induces protracting and relapsing gastroenteritis and has been linked to huge and complicated travel- and food-related outbreaks worldwide. Cyclosporiasis has become more common in both developing and developed countries as a result of increased global travel and the globalization of the human food supply. It is not just a burden on individual human health but also a worldwide public health problem. As a pathogen of interest, the molecular biological characteristics of C. cayetanensis have advanced significantly over the last few decades. However, only one FDA-approved molecular platform has been commercially used in the investigation of cyclosporiasis outbreaks. More potential molecular markers and genotyping of C. cayetanensis in samples based on the polymorphic region of the whole genomes might differentiate between separate case clusters and would be useful in tracing back investigations, especially during cyclosporiasis outbreak investigations. Considering that there is no effective vaccine for cyclosporosis, epidemiological investigation using effective tools is crucial for controlling cyclosporiasis by source tracking. Therefore, more and more epidemiological investigative studies for human cyclosporiasis should be promoted around the world to get a deeper understanding of its characteristics as well as management. This review focuses on major cyclosporiasis outbreaks and potential molecular markers for tracing back investigations into cyclosporiasis outbreaks.

Background: Cryptosporidium species are zoonotic protozoans that are important causes of diarrhoeal disease in both humans and animals. Non-coding RNAs (ncRNAs) play an important role in the innate immune defense against Cryptosporidium infection, but the underlying molecular mechanisms in the interaction between human ileocecal adenocarcinoma (HCT-8) cells and Cryptosporidium species have not been entirely revealed., Methods: The expression profiles of messenger RNAs (mRNAs), long non-coding RNAs (lncRNAs), microRNAs (miRNAs) and circular RNAs (circRNAs) in the early phase of infection of HCT-8 cells with Cryptosporidium parvum and at 3 and 12 h post infection were analyzed using the RNA-sequencing technique. The biological functions of differentially expressed RNAs (dif-RNAs) were discovered through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. The targeting relationships between three ncRNAs and mRNAs were analyzed using bioinformatics methods, followed by building a competing endogenous RNA (ceRNA) regulatory network centered on miRNAs., Results: After strictly filtering the raw data, our analysis revealed 393 dif-lncRNAs, 69 dif-miRNAs and 115 dif-mRNAs at 3 hpi, and 450 dif-lncRNAs, 129 dif-miRNAs, 117 dif-mRNAs and one dif-circRNA at 12 hpi. Of these, 94 dif-lncRNAs, 24 dif-miRNAs and 22 dif-mRNAs were detected at both post-infection time points. Eleven dif-lncRNAs, seven dif-miRNAs, eight dif-mRNAs and one circRNA were randomly selected and confirmed using the quantitative real-time PCR. Bioinformatics analyses showed that the dif-mRNAs were significantly enriched in nutritional absorption, metabolic processes and metabolism-related pathways, while the dif-lncRNAs were mainly involved in the pathways related to the infection and pathogenicity of C. parvum (e.g. tight junction protein) and immune-related pathways (e.g. cell adhesion molecules). In contrast, dif-miRNAs and dif-circRNA were significantly enriched in apoptosis and apoptosis-related pathways. Among the downregulated RNAs, the miRNAs has-miR-324-3p and hsa-miR-3127-5p appear to be crucial miRNAs which could negatively regulate circRNA, lncRNA and mRNA., Conclusions: The whole transcriptome profiles of HCT-8 cells infected with C. parvum were obtained in this study. The results of the GO and KEGG pathway analyses suggest significant roles for these dif-RNAs during the course of C. parvum infection. A ceRNA regulation network containing miRNA at its center was constructed for the first time, with hsa-miR-324-3p and hsa-miR-3127-5p being the crucial miRNAs. These findings provide novel insights into the responses of human intestinal epithelial cells to C. parvum infection., (© 2022. The Author(s).)

Coccidia are protozoan parasites in the class Conoidasida. To determine the prevalence and molecular characteristics of coccidia species in blue peafowl (Pavo cristatus) in Henan, China, 240 fecal specimens were collected and screened for the presence of Eimeria spp. and Isospora spp. The overall prevalence was 65.0% (156/240), and seven different coccidia species were identified: E. pavonis (51.3%, 123/240), E. arabic (40.0%, 96/240), E. riyadhae (37.1%, 89/240), E. mandali (22.9%, 55/240), E. mayurai (14.2%, 34/240), I. mayuir (10.9%, 26/240), and I. lacazei (8.5%, 21/240). E. arabic and E. riyadhae were detected for the first time in China. Additionally, we provide molecular data of the seven different coccidia species at the 18S and 28S ribosomal RNA and the COI loci. Sequence homology percentages among the five species of Eimeria at the 18S rRNA, 28S rRNA, and COI loci were 96.0%-98.6%, 90.7%-98.2%, and 85.0%-94.9%, respectively, whereas for two species of Isospora the sequence homology percentages were 98.8%, 99.1%, and 95.4% at three corresponding loci. This is the first report of the molecular data of the seven coccidia species in blue peafowl in China., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

This study i dentifies four Eimeria spp. recorded from fecal samples of migratory whooper swans (Cygnus cygnus) in Sanmenxia Swan Lake National Urban Wetland Park in Sanmenxia city in the middle reaches of the Yellow River, China. Eimeria hermani, Eimeria nocens, Eimeria stigmosa, and Eimeria magnalabia were compatible in all characteristic features with their respective original descriptions. In addition to the preliminary morphological identification, this study provides a preliminary genotypic identification of these four Eimeria spp. via sequencing of the 18S rRNA, 28S rRNA, and COI gene loci that are suitable for the genotypic differentiation of these coccidia. This is the first report of molecular data for the four Eimeria spp. in migratory whooper swans. Finally, this study discusses the environmental risks of these coccidia for migratory whooper swans in Sanmenxia Swan Lake National Urban Wetland Park., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Cryptosporidium spp. are important pathogens with some species causing diarrhoea in humans and animals. Sheep are one of the most common potential hosts for various Cryptosporidium spp. The prevalence of Cryptosporidium in sheep globally was evaluated from published information including molecular data via meta-analysis. In total, 126 datasets from 41 countries were included for final quantitative analysis. Sheep aged <3 months had a significantly higher prevalence (27.8%; 3284/11 938) than those at the age of 3–12 and >12 months. The prevalence of Cryptosporidium in sheep with diarrhoea of 35.4% (844/1915) was higher than in sheep that did not show diarrhoea (11.3%; 176/1691). Fourteen Cryptosporidium species/genotypes were detected in sheep globally. The proportion of subgenotype family XIIa of Cryptosporidium ubiquitum was 90.0% (216/240); the proportions of subgenotypes IIdA20G1 and IIaA15G2R1 of Cryptosporidium parvum were 15.4% (62/402) and 19.7% (79/402). The results indicate that C. parvum is the dominant species in Europe while Cryptosporidium xiaoi is the dominant species in Oceania, Asia and Africa and C. ubiquitum is the dominant species in North America and South America. Subgenotype family IIa of C. parvum is particularly widespread among sheep worldwide. The results highlight the role of sheep as a reservoir host for zoonotic cryptosporidia and the need for further study of prevalence, transmission and control of this pathogen in sheep.

This study was conducted to examine the molecular characteristics and assess the zoonotic potential of Enterocytozoon bieneusi in ruminants in northwest China. A total of 1581 fresh fecal samples were collected from eight categories of ruminants. The E. bieneusi was screened and genotyped via nested polymerase chain reaction (PCR) amplification targeting the internal transcribed spacer (ITS) of the small subunit rRNA (ssu rRNA) gene. The result indicated that the average infection rate of E. bieneusi in ruminants was 16.0% (253/1581), with infection rates of E. bieneusi in Mongolian sheep, Mongolian goats, Chifeng cattle, red deer, alpine musk deer, and blue sheep at 21.8% (169/777), 8.2% (46/561), 25.9% (28/108), 13.2% (5/38), 20.0% (4/20), and 6.3% (1/16), respectively. The infections of E. bieneusi varied by different categories. For the different age groups, the infection rates in lambs (29.3%, 108/369) and calves (57.1%, 8/14) were significantly higher than that in ewes (21.1%, 215/1020) and cows (21.3%, 20/94). For the molecular characterization, diverse E. bieneusi ITS genotypes were identified, with a total of 13 genotypes were observed, including 10 known genotypes (BEB6, COS-I, J, CHC8, I, CHG1, BEB4, CHG3, CHS7, and NCF2) and 3 novel genotypes (CNR1 to CNR3). Genotype BEB6 was predominant (59.7%, 151/253). Phylogenetic analysis revealed that most E. bieneusi ITS genotypes clustered into group 2 and only one (NCF2) genotype belonged to group 1. The zoonotic genotypes identified in ruminants in the present study indicated the zoonotic potential of E. bieneusi. In addition, simultaneous identification of genotypes, such as BEB6, COS-I, and BEB4, in the same eco-geographical system indicated some host multiplicity transmission potential of E. bieneusi., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Few studies have molecularly characterized the potential zoonotic protozoa, Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi in sheep and goats in China, therefore total 472 fecal samples were collected from eight provinces and infection rates of three protozoa were determined by PCR analysis of corresponding loci. All PCR positive samples were sequenced to identify the genotype., Results: The overall infection rates for Cryptosporidium, G. duodenalis, and E. bieneusi were 1.9% (9/472), 20.6% (97/472), and 44.5% (210/472), respectively. C. xiaoi (n = 5), C. ubiquitum (n = 3), and C. anderson (n = 1) were identified in goats. 97 G. duodenalis strains were successfully detected, and assembly E (n = 96) and assembly A (n = 1) were identified. Two novel G. duodenalis multilocus genotype (MLGs) were identified, with one belonging to subgroup AI and the other to subgroup E5. Nine known genotype (BEB6, CD6, CHC8, CHG3, CHG5, Peru6, CHG1, CHG2, and COS-I) and four new genotype (CHG26, CHG27, CHG28, and CHS18) were identified in E. bieneusi, with CHG3 dominant in this group., Conclusions: The present results highlight the role of sheep and goats as reservoir hosts for this three gastrointestinal pathogens. In summary, we provided a platform for more detailed research on genotyping or subtyping intestinal pathogens to better understand their risks and modes of transmission., (© 2022. The Author(s).)

The poultry red mite Dermanyssus gallinae is an economically important pest in poultry farms worldwide, but an effective treatment option is lacking. The current study determined the effectiveness of six Chinese herbal medicines [ Syzygium aromaticum (clove), Hibiscus syriacus (Hibiscus), Illicium verum (star anise), Leonurus artemisia (motherwort), Cinnamomum cassia (cinnamon), and Taraxacum sp. (dandelion)] against D. gallinae . Alcohol extracts were prepared via the solvent extraction method and the phenol, flavonoid, and tannin contents were determined. These active components were highest in S. aromaticum and lowest in H. syriacus, I. verum . No tannin content was detected in L. artemisia . All extracts showed contact toxicity against D. gallinae at a test concentration of 1 g/mL, with S. aromaticum and L. artemisia resulting in 100% mortality. S. aromaticum, L. artemisia , and I. verum showed the best efficacy (LC 50 0.159, 0.200, and 0.292 g/mL, respectively). Different combinations of extracts showed an additive effect of I. verum LC 90 + L. artemisia LC 90 . The acaricidal efficacy of this combination was tested against different developmental stages of D. gallinae , being most efficacious against nymphal and larval D. gallinae , with a corrected mortality rate of 100%. However, inhibition of egg hatching was only 53.69%. Taken together, these results highlight I. verum LC 90 + L. artemisia LC 90 as a promising compound with severe contact toxicity against D. gallinae . Given the wide cultivation of these species and their extensive use in foodstuffs and cosmetics as flavors and fragrances, they could be a cheap, readily available ecofriendly alternative to pesticides currently used in poultry farms., Competing Interests: Author QG was employed by Hennan Hemu Animal Pharmaceutical Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Jian, Yuan, Li, Guo, Li, Zhang, Ning, Zhang and Jian.)

Background: Cryptosporidium parvum is a zoonotic parasitic protozoan that can infect a variety of animals and humans and is transmitted between hosts via oocysts. The oocyst wall provides strong protection against hostile environmental factors; however, research is limited concerning the oocyst wall at the proteomic level., Methods: A comprehensive analysis of the proteome of oocyst wall of C. parvum was performed using label-free qualitative high-performance liquid chromatography (HPLC) fractionation and mass spectrometry-based qualitative proteomics technologies. Among the identified proteins, a surface protein (CpSP1) encoded by the C. parvum cgd7&#95;5140 (Cpcgd7&#95;5140) gene was predicted to be located on the surface of the oocyst wall. We preliminarily characterized the sequence and subcellular localization of CpSP1., Results: A total of 798 proteins were identified, accounting for about 20% of the CryptoDB proteome. By using bioinformatic analysis, functional annotation and subcellular localization of the identified proteins were examined for better understanding of the characteristics of the oocyst wall. To verify the localization of CpSP1, an indirect immunofluorescent antibody assay demonstrated that the protein was localized on the surface of the oocyst wall, illustrating the potential usage as a marker for C. parvum detection in vitro., Conclusion: The results provide a global framework about the proteomic composition of the Cryptosporidium oocyst wall, thereby providing a theoretical basis for further study of Cryptosporidium oocyst wall formation as well as the selection of targets for Cryptosporidium detection., (© 2022. The Author(s).)

Cryptosporidium is a common cause of diarrhea in children globally. However, there is limited information on the prevalence and genetic characteristics of Cryptosporidium in children in Xinjiang, China. This study aimed to assess the genetic characteristics and epidemiological status of Cryptosporidium in kindergarten children in Southern Xinjiang, China. A total of 609 fecal samples were collected from kindergartners aged 2-6 years from 11 counties in Southern Xinjiang, China. We used nested PCR amplification of the partial SSU rDNA gene to screen samples for Cryptosporidium spp. Isolates containing Cryptosporidium parvum and C. hominis were further subtyped for a gene encoding a 60-kDa glycoprotein (gp60). We used MEGA7 to construct a phylogenetic tree to study the genetic relationship between the gp60 subtypes of these two species via the Maximum Likelihood method based on the Tamura-Nei model. Only 1.3% (8/609) of asymptomatic children were confirmed to be infected with Cryptosporidium, with a 2.0% (6/299) infection rate in boys and 0.6% (2/310) infection rate in girls. Three Cryptosporidium species were identified including C. felis (37.5%; 3/8), C. hominis (37.5%; 3/8), and C. parvum (25.0%; 2/8). Three C. hominis subtypes (IbA9G3, IdA14, and IfA12G1) and two C. parvum subtypes (IIdA14G1 and IIdA15G1) were also found. This study is the first to identify the presence of Cryptosporidium in kindergarten children in Southern Xinjiang, China. The presence of zoonotic C. parvum subtypes IIdA14G1 and IIdA15G1 indicates the possible cross-species transmission of Cryptosporidium between children and animals., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)