<i>Cryptosporidium</i>infection of human small intestinal epithelial cells induces type III interferon and impairs infectivity of Rotavirus

Greigert, Valentin; Saraav, Iti; Son, Juhee; Dayao, Denise; Antia, Avan; Tzipori, Saul; Witola, William H.; Stappenbeck, Thaddeus S.; Ding, Siyuan; Sibley, L. David

doi:10.1101/2023.08.30.555581

Cited by 2 publications

(2 citation statements)

References 59 publications

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“…Infecting ALI with Cryptosporidium parental oocysts. The infection of ALI transwells has been previously described by our lab [10,45,46]. Before infection of ALI transwells, transgenic strains of parasites were treated with a 40% bleach solution for 10 min on ice, washed 3 times in DPBS w/ 1% BSA, and resuspended in DPBS w/ 1% BSA to be at 1 x 10 8 oocysts/mL for excystation.…”

Section: Cell Culturementioning

confidence: 99%

Mendelian segregation and high recombination rates facilitate genetic analyses inCryptosporidium parvum

Kimball,

Funkhouser-Jones,

Huang

et al. 2024

Preprint

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Despite the essentiality of sex in the life cycle, very little is known about the process of meiosis in Cryptosporidium. Most in vitro cell lines only support asexual growth of Cryptosporidium parvum (C. parvum), but stem cell derived intestinal epithelial cells grown under air-liquid interface (ALI) conditions support the sexual cycle. To examine chromosomal dynamics during meiosis in C. parvum, we generated two transgenic lines of parasites that were fluorescently tagged with mCherry or GFP on chromosomes 1 or 5, respectively. Infection of ALI cultures or Ifngr1-/- mice with mCherry and GFP parasites produced “yellow” oocysts generated by cross-fertilization. Outcrossed oocysts from the F1 generation were purified and used to infect HCT-8 cultures, and phenotypes of the progeny were observed by microscopy. All possible phenotypes predicted by independent segregation were represented equally (~25%) in the population, indicating that C. parvum chromosomes exhibit a Mendelian inheritance pattern. Unexpectedly, the most common pattern observed from the outgrowth of single oocysts included all possible parental and recombinant phenotypes derived from a single meiotic event, suggesting a high rate of crossover. To estimate the frequency of crossover, additional loci on chromosomes 1 and 5 were tagged and used to monitor intrachromosomal crosses in Ifngr1−/− mice. Both chromosomes showed a high frequency of crossover compared to other apicomplexans with map distances (i.e., 1% recombination) of 3-12 kb. Overall, a high recombination rate may explain many unique characteristics observed in Cryptosporidium spp. such as high rates of speciation, wide variation in host range, and rapid evolution of host-specific virulence factors.

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Section: Cell Culturementioning

confidence: 99%

Mendelian segregation and high recombination rates facilitate genetic analyses inCryptosporidium parvum

Kimball,

Funkhouser-Jones,

Huang

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Infecting ALI with Cryptosporidium parental oocysts. The infection of ALI transwells has been previously described by our lab [10,46,47]. Before infection of ALI transwells, transgenic strains of parasites were treated with a 40% bleach solution for 10 min on ice, washed 3 times in DPBS w/ 1% BSA, and resuspended in DPBS w/ 1% BSA to be at 1 x 10 8 oocysts/mL for excystation.…”

Section: Genetic Crosses In the Air Liquid Interface (Ali) Culture Sy...mentioning

confidence: 99%

Mendelian segregation and high recombination rates facilitate genetic analyses in Cryptosporidium parvum

Kimball,

Funkhouser-Jones,

Huang

et al. 2024

PLoS Genet

View full text Add to dashboard Cite

Very little is known about the process of meiosis in the apicomplexan parasite Cryptosporidium despite the essentiality of sex in its life cycle. Most cell lines only support asexual growth of Cryptosporidium parvum (C. parvum), but stem cell derived intestinal epithelial cells grown under air-liquid interface (ALI) conditions support the sexual cycle. To examine chromosomal dynamics during meiosis in C. parvum, we generated two transgenic lines of parasites that were fluorescently tagged with mCherry or GFP on chromosomes 1 or 5, respectively. Infection of ALI cultures or Ifngr1-/- mice with mCherry and GFP parasites resulted in cross-fertilization and the formation of “yellow” oocysts, which contain 4 haploid sporozoites that are the product of meiosis. Recombinant oocysts from the F1 generation were purified and used to infect HCT-8 cultures, and phenotypes of the progeny were observed by microscopy. All possible phenotypes predicted by independent segregation were represented equally (~25%) in the population, indicating that C. parvum chromosomes exhibit a Mendelian inheritance pattern. The most common pattern observed from the outgrowth of single oocysts included all possible parental and recombinant phenotypes derived from a single meiotic event, suggesting a high rate of crossover. To estimate the frequency of crossover, additional loci on chromosomes 1 and 5 were tagged and used to monitor intrachromosomal crosses in Ifngr1−/− mice. Both chromosomes showed a high frequency of crossover compared to other apicomplexans with map distances (i.e., 1% recombination) of 3–12 kb. Overall, a high recombination rate may explain many unique characteristics observed in Cryptosporidium spp. such as high rates of speciation, wide variation in host range, and rapid evolution of host-specific virulence factors.

show abstract

Cryptosporidiuminfection of human small intestinal epithelial cells induces type III interferon and impairs infectivity of Rotavirus

Cited by 2 publications

References 59 publications

Mendelian segregation and high recombination rates facilitate genetic analyses inCryptosporidium parvum

Mendelian segregation and high recombination rates facilitate genetic analyses inCryptosporidium parvum

Mendelian segregation and high recombination rates facilitate genetic analyses in Cryptosporidium parvum

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