Authors: Luyang Wang, Yuexin Wang, Zhaohui Cui...

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_5140 (Cpcgd7_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.

Topics: Animals; Cryptosporidium parvum; Membrane Proteins; Oocysts; Proteome; Proteomics; Protozoan Proteins

PubMed: 36151578
DOI: 10.1186/s13071-022-05448-8

Authors: Jahid Hasan Tipu, Audun Sivertsen, Jan-Egil Afset...

PCR-based diagnostics has revealed the previously largely unknown transmission and infections in high-income countries. This study aimed to determine domestic and imported subtypes of species in Norway, evaluate their demographic distribution, and identify potential small outbreaks. -positive human faecal samples were obtained from six medical microbiology laboratories between February 2022 and January 2024, together with 22 -positive animal samples. Species and subtypes were identified by sequencing PCR products from gp60 and SSU rRNA genes. Most cryptosporidiosis cases occurred during late summer/early autumn, primarily in children and young adults. Of 550 human samples, 359 were successfully characterized molecularly (65%), revealing infection with 10 different species. occurred in 245 (68%) human isolates with IIa and IId being major allele families, with distinct regional distribution patterns of common subtypes. A kindergarten outbreak with 5 cases was due to IIaA14G1R1. was identified in 33 (9.2%) human cases of which 24 were known to be of domestic origin, making it the second most common species in human autochthonous cases in Norway. All isolates were of the same genotype; XIVaA20G2T1, including 13 cases from a suspected small outbreak in Trøndelag. occurred in 68 typed cases (19%), but mostly in infections acquired abroad, with allele families Ib and If occurring most often. In conclusion, this study of recent spp. and subtypes in Norway, highlights the predominance of and the emergence of among autochthonous cases.

Topics: Cryptosporidiosis; Humans; Norway; Cryptosporidium; Child; Child, Preschool; Adult; Feces; Genotype; Animals; Female; Male; Adolescent; Young Adult; Infant; Middle Aged; Phylogeny; Disease Outbreaks; Aged; Cryptosporidium parvum; DNA, Protozoan

PubMed: 39361548
DOI: 10.1080/22221751.2024.2412624

Review

Authors: Lihua Xiao, Yaoyu Feng

The widespread usages of molecular epidemiological tools have improved the understanding of cryptosporidiosis transmission. Much attention on zoonotic cryptosporidiosis is centered on Cryptosporidium parvum. Results of genotype surveys indicate that calves are the only major reservoir for C. parvum infections in humans. The widespread presence of human-adapted C. parvum, especially in developing countries, is revealed by recent subtyping and multilocus typing studies, which have also demonstrated the anthroponotic transmission of C. parvum subtypes shared by humans and cattle. Developing and industrialized countries differ significantly in disease burdens caused by zoonotic species and in the source of these parasites, with the former having far fewer human infections caused by C. parvum and little zoonotic transmission of this species. Exclusive anthroponotic transmission of seemingly zoonotic C. parvum subtypes was seen in Mid-Eastern countries. Other zoonotic Cryptosporidium spp. are also responsible for substantial numbers of human infections in developing countries, many of which are probably transmitted by anthroponotic pathways. The lower pathogenicity of some zoonotic species in some populations supports the occurrence of different clinical spectra of Cryptosporidium spp. in humans. The use of a new generation of molecular diagnostic tools is likely to produce a more complete picture of zoonotic cryptosporidiosis.

Topics: Animals; Cattle; Cryptosporidiosis; Cryptosporidium parvum; Humans; Zoonoses

PubMed: 18205803
DOI: 10.1111/j.1574-695X.2008.00377.x

Authors: Katelyn A Walzer, Jayesh Tandel, Jessica H Byerly...

The parasite Cryptosporidium is a leading agent of diarrhoeal disease in young children, and a cause and consequence of chronic malnutrition. There are no vaccines and only limited treatment options. The parasite infects enterocytes, in which it engages in asexual and sexual replication, both of which are essential to continued infection and transmission. However, their molecular mechanisms remain largely unclear. Here we use single-cell RNA sequencing to reveal the gene expression programme of the entire Cryptosporidium parvum life cycle in culture and in infected animals. Diverging from the prevailing model, we find support for only three intracellular stages: asexual type-I meronts, male gamonts and female gametes. We reveal a highly organized program for the assembly of components at each stage. Dissecting the underlying regulatory network, we identify the transcription factor Myb-M as the earliest determinant of male fate, in an organism that lacks genetic sex determination. Conditional expression of this factor overrides the developmental program and induces widespread maleness, while conditional deletion ablates male development. Both have a profound impact on the infection. A large set of stage-specific genes now provides the opportunity to understand, engineer and disrupt parasite sex and life cycle progression to advance the development of vaccines and treatments.

Topics: Animals; Female; Humans; Male; Mice; Cryptosporidiosis; Cryptosporidium parvum; Gene Expression Regulation; Gene Regulatory Networks; Life Cycle Stages; Proto-Oncogene Proteins c-myb; Sex Determination Processes; Transcription, Genetic; Single-Cell Gene Expression Analysis

PubMed: 38811723
DOI: 10.1038/s41586-024-07466-1

Authors: Li Chen, Suhui Hu, Wen Jiang...

BACKGROUND

Non-human primates are often infected with human-pathogenic Cryptosporidium hominis subtypes, but rarely with Cryptosporidium parvum. In this study, 1452 fecal specimens were collected from farmed crab-eating macaques (Macaca fascicularis) in Hainan, China during the period April 2016 to January 2018. These specimens were analyzed for Cryptosporidium species and subtypes by using PCR and sequence analysis of the 18S rRNA and 60 kDa glycoprotein (gp60) genes, respectively.

RESULTS

Altogether, Cryptosporidium was detected using 18S rRNA-based PCR in 132 (9.1%) sampled animals, with significantly higher prevalence in females (12.5% or 75/599 versus 6.1% or 43/706), younger animals (10.7% or 118/1102 in monkeys 1-3-years-old versus 4.0% or 14/350 in those over 3-years-old) and animals with diarrhea (12.6% or 46/365 versus 7.9% or 86/1087). Four Cryptosporidium species were identified, namely C. hominis, C. parvum, Cryptosporidium muris and Cryptosporidium ubiquitum in 86, 30, 15 and 1 animal, respectively. The identified C. parvum, C. hominis and C. ubiquitum were further subtyped by using gp60 PCR. Among them, C. parvum belonged to subtypes in two known subtype families, namely IIoA14G1 (in 18 animals) and IIdA19G1 (in 2 animals). In contrast, C. hominis mostly belonged to two new subtype families Im and In, which are genetically related to Ia and Id, respectively. The C. hominis subtypes identified included ImA18 (in 38 animals), InA14 (in six animals), InA26 (in six animals), InA17 (in one animal) and IiA17 (in three animals). The C. ubiquitum isolates belonged to subtype family XIId. By subtype, ImA18 and IIoA14G1 were detected in animals with diarrhea whereas the remaining ones were mostly found in asymptomatic animals. Compared with C. parvum and C. muris, higher oocyst shedding intensity was observed in animals infected with C. hominis, especially those infected with the Im subtype family.

CONCLUSIONS

Data from the study suggest that crab-eating macaques are infected with diverse C. parvum and C. hominis subtypes. The C. parvum IIo subtype family previously seen in rodents in China has apparently expanded its host range.

Topics: Age Factors; Animals; China; Cryptosporidiosis; Cryptosporidium; Cryptosporidium parvum; DNA, Protozoan; Feces; Female; Genotype; Host Specificity; Macaca fascicularis; Monkey Diseases; Polymerase Chain Reaction; RNA, Ribosomal, 18S; Sequence Analysis, DNA

PubMed: 31307508
DOI: 10.1186/s13071-019-3604-7

Authors: Manasi Sawant, Sadia Benamrouz-Vanneste, Dionigia Meloni...

is a leading cause of diarrhoeal illness worldwide being a significant threat to young children and immunocompromised patients, but the pathogenesis caused by this parasite remains poorly understood. was recently linked with oncogenesis. Notably, the mechanisms of gene expression regulation are unexplored in and little is known about how the parasite impact host genome regulation. Here, we investigated potential histone lysine methylation, a dynamic epigenetic modification, during the life cycle of the parasite. We identified SET-domain containing proteins, putative lysine methyltransferases (KMTs), in the genome and classified them phylogenetically into distinct subfamilies (namely CpSET1, CpSET2, CpSET8, CpKMTox and CpAKMT). Our structural analysis further characterized CpSET1, CpSET2 and CpSET8 as histone lysine methyltransferases (HKMTs). The expression of the CpSET genes varies considerably during the parasite life cycle and specific methyl-lysine antibodies showed dynamic changes in parasite histone methylation during development (CpSET1:H3K4; CpSET2:H3K36; CpSET8:H4K20). We investigated the impact of infection on the host histone lysine methylation. Remarkably, parasite infection led to a considerable decrease in host H3K36me3 and H3K27me3 levels, highlighting the potential of the parasite to exploit the host epigenetic regulation to its advantage. This is the first study to describe epigenetic mechanisms occurring throughout the parasite life cycle and during the host-parasite interaction. A better understanding of histone methylation in both parasite and host genomes may highlight novel infection control strategies.

Topics: Child, Preschool; Cryptosporidiosis; Cryptosporidium; Cryptosporidium parvum; Epigenesis, Genetic; Histone-Lysine N-Methyltransferase; Histones; Humans; Lysine; Methylation

PubMed: 36097362
DOI: 10.1080/21505594.2022.2123363

Authors: Mattie C Pawlowic, Sumiti Vinayak, Adam Sateriale...

The apicomplexan parasite Cryptosporidium is a leading cause of diarrheal disease and an important contributor to overall global child mortality. We currently lack effective treatment and immune prophylaxis. Recent advances now permit genetic modification of this important pathogen. We expect this to produce rapid advances in fundamental as well as translational research on cryptosporidiosis. Here we outline genetic engineering for Cryptosporidium in sufficient detail to establish transfection in any laboratory that requires access to this key technology. This chapter details the conceptual design consideration, as well as the experimental steps required to transfect, select, and isolate transgenic parasites. We also provide detail on key in vitro and in vivo assays to detect, validate, and quantify genetically modified Cryptosporidium parasites. © 2017 by John Wiley & Sons, Inc.

Topics: Animals; Cell Culture Techniques; Cryptosporidiosis; Cryptosporidium parvum; Genetic Techniques; Humans; Microscopy, Fluorescence; Transfection

PubMed: 28800157
DOI: 10.1002/cpmc.33

Authors: Heng Jiang, Xu Zhang, Xin Li...

BACKGROUND

Cryptosporidium parvum is an important zoonotic parasite, which not only causes economic losses in animal husbandry but also harms human health. Due to the lack of effective measures for prevention and treatment, it is important to understand the pathogenesis and survival mechanism of C. parvum. Autophagy is an important mechanism of host cells against parasite infection through key regulatory factors such as microRNAs and MAPK pathways. However, the regulatory effect of C. parvum on autophagy has not been reported. Here, we demonstrated that C. parvum manipulated autophagy through host cellular miR-26a, miR-30a, ERK signaling and P38 signaling for parasite survival.

METHODS

The expression of Beclin1, p62, LC3, ERK and P38 was detected using western blotting in HCT-8 cells infected with C. parvum as well as treated with miR-26a-mimic, miR-30a-mimic, miR-26a-mimic or miR-30a-inhibitor post C. parvum infection. The qPCR was used to detect the expression of miR-26a and miR-30a and the number of C. parvum in HCT-8 cells. Besides, the accumulation of autophagosomes was examined using immunofluorescence.

RESULTS

The expression of Beclin1 and p62 was increased, whereas LC3 expression was increased initially at 0-8 h but decreased at 12 h and then increased again in C. parvum-infected cells. C. parvum inhibited miR-26a-mimic-induced miR-26a but promoted miR-30a-mimic-induced miR-30a expression. Suppressing miR-30a resulted in increased expression of LC3 and Beclin1. However, upregulation of miR-26a reduced ERK/P38 phosphorylation, and inhibiting ERK/P38 signaling promoted Beclin1 and LC3 while reducing p62 expression. Treatment with miR-26a-mimic, autophagy inducer or ERK/P38 signaling inhibitors reduced but treatment with autophagy inhibitor or miR-30a-mimic increased parasite number.

CONCLUSIONS

The study found that C. parvum could regulate autophagy by inhibiting miR-26a and promoting miR-30a expression to facilitate the proliferation of parasites. These results revealed a new mechanism for the interaction of C. parvum with host cells.

Topics: Animals; Humans; Beclin-1; Cryptosporidium parvum; Cryptosporidiosis; Cryptosporidium; Autophagy; MicroRNAs

PubMed: 36522638
DOI: 10.1186/s13071-022-05606-y

Authors: Semie Hong, Kyungjin Kim, Sejoung Yoon...

Cryptosporidium parvum is a zoonotic protozoan parasite that causes cryptosporidial enteritis. Numerous outbreaks of cryptosporidiosis have been reported worldwide. Cryptosporidium is transmitted to hosts via consumption of contaminated water and food but also by direct contact with contaminated soil or infected hosts. The present study investigated farm soil collected from 34 locations along the western Korean peninsula and 24 vegetables purchased from local grocery markets in Seoul. The soil and vegetable samples were examined by real-time polymerase chain reaction (qPCR) to estimate the risk of infection. Eleven of 34 locations (32.4%) and 3 of 24 vegetable samples (12.5%) were contaminated with Cryptosporidium parvum, as confirmed by TaqI enzyme digestion of qPCR products and DNA sequencing. It is suggested that Cryptosporidium infection can be mediated via farm soil and vegetables. Therefore, it is necessary to reduce contamination of this organism in view of public health.

Topics: Base Sequence; Cryptosporidiosis; Cryptosporidium parvum; DNA, Protozoan; Enteritis; Foodborne Diseases; Humans; Sequence Alignment; Sequence Analysis, DNA; Soil; Vegetables

PubMed: 25368489
DOI: 10.3346/jkms.2014.29.10.1367

Meta-Analysis

Authors: Philippa King, Kevin M Tyler, Paul R Hunter...

BACKGROUND

Globally cryptosporidiosis is one of the commonest causes of mortality in children under 24 months old and may be associated with important longterm health effects. Whilst most strains of Cryptosporidium parvum are zoonotic, C. parvum IIc is almost certainly anthroponotic. The global distribution of this potentially important emerging infection is not clear.

METHODS

We conducted a systematic review of papers identifying the subtype distribution of C. parvum infections globally. We searched PubMed and Scopus using the following key terms Cryptospor* AND parvum AND (genotyp* OR subtyp* OR gp60). Studies were eligible for inclusion if they had found C. parvum within their human study population and had subtyped some or all of these samples using standard gp60 subtyping. Pooled analyses of the proportion of strains being of the IIc subtype were determined using StatsDirect. Meta-regression analyses were run to determine any association between the relative prevalence of IIc and Gross Domestic Product, proportion of the population with access to improved drinking water and improved sanitation.

RESULTS

From an initial 843 studies, 85 were included in further analysis. Cryptosporidium parvum IIc was found in 43 of these 85 studies. Across all studies the pooled estimate of relative prevalence of IIc was 19.0% (95% CI: 12.9-25.9%), but there was substantial heterogeneity. In a meta-regression analysis, the relative proportion of all C. parvum infections being IIc decreased as the percentage of the population with access to improved sanitation increased and was some 3.4 times higher in those studies focussing on HIV-positive indivduals.

CONCLUSIONS

The anthroponotic C. parvum IIc predominates primarily in lower-income countries with poor sanitation and in HIV-positive individuals. Given the apparent enhanced post-infectious virulence of the other main anthroponotic species of Cryptosporidium (C. hominis), it is important to learn about the impact of this subtype on human health.

Topics: Cryptosporidiosis; Cryptosporidium parvum; Genetic Variation; Genotype; Humans; Poverty; Sanitation

PubMed: 30621759
DOI: 10.1186/s13071-018-3263-0