The Cryptosporidium Oocyst Wall Protein Is a Member of a Multigene Family and Has a Homolog in Toxoplasma

Abstract

Coccidian parasites are transmitted via a fecal oocyst stage that is exceptionally resistant to environmental stress and harsh chemical treatments, which allows parasites to stably persist outside a host. Because of its oocyst durability Cryptosporidium parvum is a significant water-and food-borne pathogen of humans, as well as animals of agricultural importance. To date, only one apicomplexan oocyst membrane protein has been identified, Cryptosporidium oocyst wall protein 1 (COWP1). COWP1 has a highly cysteine-rich periodicity due to arrays of two apicomplexan-specific motifs, designated the type I and type II domains. In this study, exhaustive BLAST screening of a complete C. parvum genome sequence database resulted in identification of eight additional genes encoding similar arrays of cysteine-rich type I and/or type II domains. Transcript expression analysis revealed that all COWP genes are abundantly expressed at a time when developing oocysts are observed, roughly 48 to 72 h after inoculation of in vitro cultures. A monoclonal antibody recognizing COWP8 specifically localized to the C. parvum oocyst wall, supporting the hypothesis that multiple COWPs play a role in the oocyst wall structure. BLAST screening of the Toxoplasma gondii genome sequence database resulted in identification of a gene encoding at least one COWP homolog (TgOWP1), and this multiexon sequence information was used to isolate a full-length cDNA. Exhaustive screening of Plasmodium sp. genome sequence databases by using COWP genes as BLAST queries failed to detect similar proteins in Plasmodium. We therefore propose that the COWP family of proteins have a structural role in apicomplexan species that produce durable shed cysts capable of surviving environmental stress.Cryptosporidium parvum is an obligate intracellular parasite that causes severe diarrhea via infection of the lower gastrointestinal tract epithelium of humans and other mammals. Cryptosporidium is one of several genera in the phylum Apicomplexa that have common life cycle and morphological stages, collectively referred to as coccidia (reviewed in reference 5). Coccidian oocyst stages are highly resistant to environmental stress and chemical disinfection (9), and this is attributed to a durable oocyst wall, a complex protective barrier consisting of a double layer of a protein-lipid-carbohydrate matrix (5). Ultrastructural studies have revealed that coccidian oocyst walls appear to form as a result of the interaction of wall-forming bodies and pellicular membranes (7,14). During oocyst maturation wall-forming bodies degranulate in the cytoplasm, and the vesicular material is delivered into the intermembrane space. Once the construction of the outer layer of the oocyst wall nears completion, a similar process begins for the inner layer. In the case of a related coccidian parasite, Eimeria nieschulzi, seven membranes and two types of wallforming bodies that appear to be involved in oocyst wall formation have been identified (14).Despite numerous studies of this unique structure...

Diversity of extracellular proteins during the transition from the ‘proto-apicomplexan’ alveolates to the apicomplexan obligate parasites

“…With the advent of whole genome sequence information it is now known that COWP genes are amplified in Cryptosporidium, which has 9 genes, and are also amplified in all cyst-forming coccidians and Gregarina ( Fig. 3A; Templeton et al 2004a;Templeton et al 2010). OWP modules are also found in genes amplified in the chromerids (e.g.…”

Factors mediating plastid dependency and the origins of parasitism in apicomplexans and their close relatives

“…Interestingly, chromerids, colpodellids, Perkinsus, and certain dinoflagellates (3,44) are known to multiply by a process similar to apicomplexan schizogony. The second group are oocyst wall proteins (OWPs), which are exclusively found in species that form reproductive/resting cysts: Cryptosporidium, coccidians, and all chrompodellids (3,12,44,45). Cell-wall maintenance has also been linked to the presence of multidomain polyketide synthases (PKSI/FASI) (26,46), which are found in all myzozoans (Fig.…”

Translational repression of the cpw-wpc gene family in the malaria parasite Plasmodium

“…Each domain typically contains 6 cysteines, likely paired in disulfide bridges, which is reminiscent of other cysteine-rich apicomplexan domains such as the COWP protein family of the oocyst wall in coccidians [26,27]; the 6-Cys family that includes the Plasmodium gamete proteins Pfs230, Pfs47 and Pfs48/45 [28][29][30][31]; and the Toxoplasma SAG proteins [28,30,31]. The cpw-wpc gene family is widely conserved among apicomplexans, including Table 3).…”