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Cryptosporidium parvum

Table of Contents
  1. General Information
    1. NCBI Taxonomy ID
    2. Disease
    3. Introduction
    4. Microbial Pathogenesis
    5. Host Ranges and Animal Models
    6. Host Protective Immunity
  2. Vaccine Related Pathogen Genes
    1. CP15/60 (Protective antigen)
    2. CP23 (Protective antigen)
  3. Vaccine Related Host Genes
    1. Ighg1
    2. Ighv1-9
  4. Vaccine Information
    1. C. parvum DNA vaccine encoding CP15/60
    2. C. parvum DNA vaccine encoding CP23
  5. References
I. General Information
1. NCBI Taxonomy ID:
5807
2. Disease:
Cryptosporidiosis
3. Introduction
Cryptosporidium parvum is one of several species that cause cryptosporidiosis, a parasitic disease of the mammalian intestinal tract. Primary symptoms of C. parvum infection are acute, watery, and non-bloody diarrhoea. C. parvum infection is of particular concern in immunocompromised patients, where diarrhea can reach 10–15L per day. Other symptoms may include anorexia, nausea/vomiting and abdominal pain.

Infection is caused by ingestion of sporulated oocysts transmitted by the fecal-oral route. In healthy human hosts, the median infective dose is 132 oocysts. The general C. parvum life cycle is shared by other members of the genus. Invasion of the apical tip of ileal enterocytes by sporozoites and merozoites causes pathology seen in the disease. Infection is generally self-limiting in immunocompetent people. In immunocompromised patients, such as those with AIDS or those undergoing immunosuppressive therapy, infection may not be self-limiting, leading to dehydration and, in severe cases, death.

The diagnosis of C. parvum consists of serological tests and microscopic evaluation of oocysts in stools using Kinyoun acid-fast staining. C. parvum is considered to be the most important waterborne pathogen in developed countries. It is resistant to all practical levels of chlorination, surviving for 24hrs at 1000 mg/L free chlorine (Wiki: Cryptosporidium parvum).
4. Microbial Pathogenesis
Invasion of the apical tip of ileal enterocytes by sporozoites and merozoites causes pathology seen in the disease. Infection is generally self-limiting in immunocompetent people. In immunocompromised patients, such as those with AIDS or those undergoing immunosuppressive therapy, infection may not be self-limiting, leading to dehydration and, in severe cases, death (Wiki: Cryptosporidium parvum).
5. Host Ranges and Animal Models
Cryptosporidiosis is an enteric diarrheal disease caused mainly by Cryptosporidium parvum, an obligate intracellular protozoan parasite of the intestinal epithelium that infects a variety of mammalian species, including rodents, livestock, and humans (Ehigiator et al., 2007).
6. Host Protective Immunity
Cryptosporidium sp. infection induces both cellular and humoral immune responses in infected hosts, but protection is mediated primarily through cellular immunity (Ehigiator et al., 2007).
1. CP15/60
  • Gene Name : CP15/60
  • Sequence Strain (Species/Organism) : Cryptosporidium parvum
  • VO ID : VO_0011309
  • NCBI Nucleotide GI : 170676501
  • NCBI Protein GI : 170676502
  • Protein Accession : ACB30404.1
  • Taxonomy ID : 5807
  • Gene Strand (Orientation) : ?
  • Protein Name : sporozoite surface antigen CP15/60
  • Protein Length : 148
  • DNA Sequence : Show Sequence
    >gi|170676501|gb|EU532137.1| Cryptosporidium parvum sporozoite surface antigen CP15/60 mRNA, complete cds
    ATGGGTAACTTGAAATCCTGTTGTTCTTTTGCCGATGAACACTCCCTAACCCCTACTCAACTAGTAGTTG
    GAAATGGTTCAGGAGCTTCAGAAACTGCTTCCAACCACCCCCAAGAAGAAGTTAATGATATTAATACTTT
    TAATGTAAAGTTAACAATGCAAGATAGAAGTAAGCTTGACTGTGAGGTAGTATTTGATAGCACAAGTATT
    TCGCTTTCTGGAGATGGAAAATGCAGAAATATTGCTTTGGATGAAATCCACCAATTATTATATTCAAAGG
    AAGAGCTCTCTAGAGTTGAAAGTAGTGCTGGAATCAGTGATTCCGACAATTGTGTTGCAATTCATCTCAA
    AGAATCAGGAAACTGTATTCCCCTTTTCTTTAATAATTCGCAAGACAAAGAAAGATTTGTTGCAACAGCA
    AACAAATTCAAACCAAACTTTAACTAAAACAGGAATTTCATTAATTCTGGATTTGTTTTTTCCAAACCCT
    ATAAATAAAATCCAAATAGCTTTTCTTCAATAACAGGGAG
  • Protein Sequence : Show Sequence
    >gi|170676502|gb|ACB30404.1| sporozoite surface antigen CP15/60 [Cryptosporidium parvum]
    MGNLKSCCSFADEHSLTPTQLVVGNGSGASETASNHPQEEVNDINTFNVKLTMQDRSKLDCEVVFDSTSI
    SLSGDGKCRNIALDEIHQLLYSKEELSRVESSAGISDSDNCVAIHLKESGNCIPLFFNNSQDKERFVATA
    NKFKPNFN
  • Molecule Role : Protective antigen
  • Molecule Role Annotation : Preparturient cows were immunized three times over a six-week period with recombinant plasmid DNA encoding the Cryptosporidium parvum CP15/60 antigen by injecting the DNA in the mammary gland. Colostrum from these cows contained Ab specific for C. parvum sporozoites and oocysts as indicated by immunofluorescence Ab (IFA) staining. Mice receiving immune colostrum showed partial protection (about 50% reduction) against intestinal C. parvum development compared to mice receiving control colostrum. This protection was evident at a challenge dose of 103 C. parvum oocysts per mouse (Jenkins et al., 1999).
  • Related Vaccine(s): C. parvum DNA vaccine encoding CP15/60
2. CP23
  • Gene Name : CP23
  • Sequence Strain (Species/Organism) : Cryptosporidium parvum
  • VO ID : VO_0011308
  • NCBI Protein GI : 88659549
  • Taxonomy ID : 5807
  • Gene Strand (Orientation) : ?
  • Protein Name : Cp23 antigen
  • Protein Length : 124
  • Protein Note : genotype: cervine
  • Protein Sequence : Show Sequence
    >gi|88659549|gb|ABD47677.1| Cp23 antigen [Cryptosporidium parvum]
    MGCSSSKPETKVADNKSAADANKQRELAEKKAQLAKAVKNPAPISNQPQAKPEEPKKAEPAPVPAPAPVP
    APAPAPVPTPAPVPEAPAAEAPSNQAPAPVPAEAPAAEAPVDQNGVPQEKPADA
  • Molecule Role : Protective antigen
  • Molecule Role Annotation : In this study, Cp23 antigen was investigated as a vaccine candidate using the DNA vaccine model in adult interleukin-12 (IL-12) knockout (KO) mice, which are susceptible to C. parvum infection. Cp23-DNA vaccination induced a 50-60% reduction in oocysts shedding, indicating a partial protection against C. parvum infection in IL-12 KO mice (Ehigiator et al., 2007).
  • Related Vaccine(s): C. parvum DNA vaccine encoding CP23
1. Ighg1
2. Ighv1-9
  • Gene Name : Ighv1-9
  • Sequence Strain (Species/Organism) : Mus musculus
  • NCBI Gene ID : 668478
  • Genbank Accession : AC073561
  • Taxonomy ID : 10090
  • Chromosome No : 12
  • Gene Starting Position : 114583568
  • Gene Ending Position : 114583861
  • Protein Name : immunoglobulin heavy variable V1-9
  • Protein Note : Also known as Igg2a; Gm16697
  • DNA Sequence : Show Sequence
    >gi|372099098:114583568-114583861 Mus musculus strain C57BL/6J chromosome 12, GRCm38 C57BL/6J
    GTCTTGCACAGTAATAGATGGCAGAGTCCTCAGTTGTCAGGCTGCTGAGTTGCATGTAGGCTGTGTTGGA
    GGATGTATCTGCAGTGAATGTGGCCTTGCCCTTGAACTTCTCATTGTAGTTAGTACTACCACTTCCAGGT
    AAAATCTCTCCAATCCACTCAAGGCCATGTCCAGGCCTCTGCTTTACCCACTCTATCCAGTAGCCAGTGA
    ATGTGTAGCCAGTAGCCTTGCAGGAAAGCTTCACTGAGGCCCCAGGCTTCATCAGCTCAGCTCCAGACTG
    CTGCAGCTGAACCT
  • Molecule Role : Vaximmutor
  • Related Vaccine(s): C. parvum DNA vaccine encoding CP23
IV. Vaccine Information
1. C. parvum DNA vaccine encoding CP15/60
a. Vaccine Ontology ID:
VO_0011502
b. Type:
DNA vaccine
c. Status:
Research
d. Antigen
C. parvum sporozoite surface antigen CP15/60
e. Gene Engineering of CP15/60
  • Type: DNA vaccine construction
  • Description: Recombinant pCP15/60 plasmid DNA was prepared using standard procedures and resuspended to 1 mg/ml in 0.15 M NaCl. Sera was collected from multiparous Holstein–Fresian cows at 8–12 weeks prior to parturition and tested by IFA for presence of Ab to C. parvum oocysts (Jenkins et al., 1999).
  • Detailed Gene Information: Click here.
f. Vector:
pCP15/60 (Jenkins et al., 1999)
g. Immunization Route
Parenchymal injection
h. Mouse Response
  • Host Strain: C57BL/6NCr
  • Vaccination Protocol: Cows (n=2) with negligible titers (<1:50) to C. parvum oocysts were immunized at 6, 4 and 2 weeks prior to parturition with 1 mg of plasmid DNA by needle-syringe injection into the parenchymal tissue of the mammary gland. Colostrum from the cows was also tested for conferring passive immunity against C. parvum infection by oral administration to immunosuppressed adult inbred mice. Immune colostrum and control colostrum were administered to separate groups of dexamethasone (DEX)-treated adult C57BL/6NCr mice beginning 12 h before and at 12 h intervals for 3 days after oral C. parvum oocyst infection (Jenkins et al., 1999).
  • Challenge Protocol: In three separate experiments, DEX-treated mice (n=4 per group, six groups per treatment) received 250 μl of HBC or NBC by gastric intubation using a 26 ga. gavage needle 12 h prior to C. parvum infection. At 0 h, the mice were infected with either 103 or 104 C. parvum oocysts and given 250 μl of either HBC or NBC. Control mice were given H2O and either infected or not infected with C. parvum oocysts (Jenkins et al., 1999).
  • Efficacy: Mice receiving immune colostrum showed partial protection (about 50% reduction) against intestinal C. parvum development compared to mice receiving control colostrum. This protection was evident at a challenge dose of 103 C. parvum oocysts per mouse (Jenkins et al., 1999).
2. C. parvum DNA vaccine encoding CP23
a. Vaccine Ontology ID:
VO_0011362
b. Type:
DNA vaccine
c. Status:
Research
d. Antigen
Cp23 antigen
e. Gene Engineering of CP23
  • Type: DNA vaccine construction
  • Description: The C. parvum 27-kDa antigen coding sequence (GenBank accession number U34390) was amplified from a Cp23- pGEX 4T-2 clone. The fragment was cloned into the EcoRI and NotI restriction enzyme sites of the pUMVC4b expression vector (Adevron, Fargo, ND, USA). This vector has a cytomegalovirus promoter and immunoadjuvant site that enhances immune responses. Plasmids containing inserts (pUMVC4b-Cp23) and plasmids without inserts (pUMVC4b) were transformed into UltraMAX™ DH5a-FT™ Competent Cells (Invitrogen, Carlsbad, CA, USA), purified, and dissolved in sterile Tris–borate–ethylenediamine tetraacetic acid (TBE). After cloning, the sequence of the resulting clone was confirmed by automated DNA sequencing (Ehigiator et al., 2007).
  • Detailed Gene Information: Click here.
f. Vector:
pUMVC4b expression vector
g. Immunization Route
Subcutaneous injection
h. Mouse Response
  • Host Strain: C57BL/6 KO
  • Vaccination Protocol: Mice (ten per group) were injected subcutaneously in the ear with 100-μg plasmid pUMVC4b-Cp23 in TBE on days 0, 14, and 29. Mice in the control group were injected with either the vector pUMVC4b without the insert or with PBS (Ehigiator et al., 2007).
  • Challenge Protocol: The different treatment groups of IL-12p40KO mice were challenged with a dose of 1,000 C. parvum oocysts by oral gavage 2 weeks after the last immunization (Ehigiator et al., 2007).
  • Efficacy: Cp23-DNA vaccination induced a 50-60% reduction in oocysts shedding, indicating a partial protection against C. parvum infection in IL-12 KO mice (Ehigiator et al., 2007).
  • Host Gene Response of Ighg1
    • Gene Response: Six weeks post immunization, mice were bled and antibody responses were measured by ELISA. IgG1 responses were significantly higher in Cp23 vaccinated mice than vector (control) immunized mice (Ehigiator et al., 2007).
    • Detailed Gene Information: Click here.
  • Host Gene Response of Ighv1-9
    • Gene Response: Six weeks post immunization, mice were bled and antibody responses were measured by ELISA. IgG2a responses were significantly higher in Cp23 vaccinated mice than vector (control) immunized mice (Ehigiator et al., 2007).
    • Detailed Gene Information: Click here.
V. References
1. Ehigiator et al., 2007: Ehigiator HN, Romagnoli P, Priest JW, Secor WE, Mead JR. Induction of murine immune responses by DNA encoding a 23-kDa antigen of Cryptosporidium parvum. Parasitology research. 2007; 101(4); 943-950. [PubMed: 17487508].
2. Jenkins et al., 1999: Jenkins MC, O'Brien C, Trout J, Guidry A, Fayer R. Hyperimmune bovine colostrum specific for recombinant Cryptosporidium parvum antigen confers partial protection against cryptosporidiosis in immunosuppressed adult mice. Vaccine. 1999; 17(19); 2453-2460. [PubMed: 10392628].
3. Wiki: Cryptosporidium parvum: Cryptosporidium parvum [http://en.wikipedia.org/wiki/Cryptosporidium_parvum]