**3. Life cycle and classification**

The parasite has a complex monoxenous life cycle with both asexual (merogony) and sexual (gametogony) stages. Ingestion of an infective oocyst (containing four sporozoites) by a susceptible host initiates the excystation process in the gastrointestinal tract. The sporulated oocyst ruptures, releasing sporozoites that invade the enterocytes, inducing the cell membrane to enclose the parasite in the parasitophorous vacuole, which then differentiates into a trophozoite. Trophozoites undergo merogony and form either a further type I meront or a type II meront, which contains four merozoites that are destined for gametogony. Merozoites can differentiate into sexually distinct stages called macro- and microgametocytes in a process called gametogony. New oocysts are formed in the epithelial cells from the fusion of a macro- and a microgametocyte to form a diploid zygote. The new fused cell evolves and sporulates in situ in a process called sporogony, becoming oocysts containing four sporozoites. Type II meronts attach to the epithelial cell and differentiate into either macrogamonts or microgamonts. The microgametes from the microgamont are released, and each can fertilised a macrogamont to form a diploid zygote. This cell undergoes a process like meiosis (sporogony) to produce an oocyst, either thinor thick-walled, containing four sporozoites (sporulated oocysts). The thin-walled oocysts are involved in autoinfection, and thick-walled oocysts are released within the faeces to infect new hosts [104–107] (**Figure 1**).

Until relatively recently, *Cryptosporidium* was classified as a coccidian parasite. However, the taxonomic placement of *Cryptosporidium* was altered after revisions to higher-order classifications due to recent particularities observed in *Cryptosporidium*. The parasite can develop in a cell-free culture, while extracellular stages have been observed in both cell-free and cell cultures, in biofilms, and in vivo [108–111]. It presents the ability to grow and amplify without host cell attachment and encapsulation, as well as the insensitivity of all anticoccidial agents [26]. Moreover, the parasite lacks a micropyle, sporocyst, and polar granular [111–113]. Although initially considered

**337**

**Figure 1.**

*State of the Art and Future Directions of* Cryptosporidium *spp.*

to be a coccidian, *Cryptosporidium* spp. share features of both the coccidia and gregarines, confirmed by morphological and molecular data. Major similarities between *Cryptosporidium* and gregarine parasites are as follows: (1) the ability to complete its life cycle in the absence of host cells, (2) extracellular gamont-like stages, (3) the process in which two mature trophozoites pair up before the formation of gametocyst (szygy), and (4) changing cell architecture to adapt to diverse environments (biofilms, coelom, intestines, soil, and water) [107, 108, 111, 114]. The most recent classification considers *Cryptosporidium* as a separate group within the Apicomplexa. Analyses of comparative genomics and of phylogenetic inference and the ability of *Cryptosporidium* to complete its life cycle extracellularly confirm its close relationship with gregarines and corroborate the transference of *Cryptosporidium* to the Gregarinomorphea class as a new subclass of Cryptogregaria [111, 115]. Early taxonomy at species level was based originally on morphology and host specificity. Nowadays, the description of species

*well as (l) involved in autoinfection process (adapted from Ref. [104]).*

*A schematic diagram of* Cryptosporidium *life cycle. After ingestion of contaminated water and/or food, the oocyst wall opens (excystation) triggered by temperature, stomach acid, and bile salts. Then, sporulated oocyst ruptures releasing (a) sporozoites that (b) invade the host cell (c) inducing the cell membrane to enclose the parasite in the parasitophorous vacuole, (d) which then differentiates into a trophozoite that undergoes an asexual reproduction, (e) forming a type I meront that contains 6–8 merozoites. These merozoites can reinfect the epithelial cell, where they undergo merogony and form type I meront or (f) type II meront. (g) Merozoites can differentiate into sexually distinct stages called (g) micro- and (h) macrogametocytes. (i) Zygote is formed after the fertilisation of macrogametocyte by the microgametocyte, (j) and this cell undergoes sporogony and produces a thin-walled oocyst. (k) These thin-walled oocysts are released within faeces to infect new hosts, as* 

*DOI: http://dx.doi.org/10.5772/intechopen.88387*

*State of the Art and Future Directions of* Cryptosporidium *spp. DOI: http://dx.doi.org/10.5772/intechopen.88387*

#### **Figure 1.**

*Parasitology and Microbiology Research*

necessary for accurate identification.

tracking for further in vivo testing in animals and humans.

the faeces to infect new hosts [104–107] (**Figure 1**).

**3. Life cycle and classification**

cryptosporidiosis, which could also be used for therapeutic or vaccine trials, are

Current treatment options for cryptosporidiosis are limited. So far, there is no vaccine against *Cryptosporidium* [97]*,* and nitazoxanide (NTZ) is the only drug approved by the FDA for treatment of cryptosporidiosis in children and immunocompetent adults [98]. However, it is not effective without an appropriate immune status and, consequently, is ineffectual for the treatment of immune-compromised patients, particularly those with AIDS [25, 99]. NTZ is a nitrothiazole benzamide compound with a broad spectrum of activity against a wide range of parasites, bacteria, and viruses. In protozoa, NTZ inhibits the enzyme pyruvate ferredoxin oxidoreductase, which is essential to anaerobic energy metabolism [100]. Due to the prevalence of cryptosporidiosis, the development of novel therapeutic targets and vaccines against *Cryptosporidium* spp. is a public health priority. The ongoing need to develop new anti-cryptosporidial drugs has spurred the process of finding new uses for existing drugs. Repurposing drug provides an attractive alternative to drug development [101]. Two compounds, 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor, pitavastatin and auranofin (approved for the treatment of rheumatoid arthritis), have been shown to be effective against *Cryptosporidium* in vitro [102]. Auranofin has been shown to be 10 times more potent than metronidazole against *Entamoeba histolytica*, the protozoan agent of human amoebiasis [103]. HMG-CoA and auranofin have particular promise in fast-

The parasite has a complex monoxenous life cycle with both asexual (merogony) and sexual (gametogony) stages. Ingestion of an infective oocyst (containing four sporozoites) by a susceptible host initiates the excystation process in the gastrointestinal tract. The sporulated oocyst ruptures, releasing sporozoites that invade the enterocytes, inducing the cell membrane to enclose the parasite in the parasitophorous vacuole, which then differentiates into a trophozoite. Trophozoites undergo merogony and form either a further type I meront or a type II meront, which contains four merozoites that are destined for gametogony. Merozoites can differentiate into sexually distinct stages called macro- and microgametocytes in a process called gametogony. New oocysts are formed in the epithelial cells from the fusion of a macro- and a microgametocyte to form a diploid zygote. The new fused cell evolves and sporulates in situ in a process called sporogony, becoming oocysts containing four sporozoites. Type II meronts attach to the epithelial cell and differentiate into either macrogamonts or microgamonts. The microgametes from the microgamont are released, and each can fertilised a macrogamont to form a diploid zygote. This cell undergoes a process like meiosis (sporogony) to produce an oocyst, either thinor thick-walled, containing four sporozoites (sporulated oocysts). The thin-walled oocysts are involved in autoinfection, and thick-walled oocysts are released within

Until relatively recently, *Cryptosporidium* was classified as a coccidian parasite. However, the taxonomic placement of *Cryptosporidium* was altered after revisions to higher-order classifications due to recent particularities observed in *Cryptosporidium*. The parasite can develop in a cell-free culture, while extracellular stages have been observed in both cell-free and cell cultures, in biofilms, and in vivo [108–111]. It presents the ability to grow and amplify without host cell attachment and encapsulation, as well as the insensitivity of all anticoccidial agents [26]. Moreover, the parasite lacks a micropyle, sporocyst, and polar granular [111–113]. Although initially considered

**336**

*A schematic diagram of* Cryptosporidium *life cycle. After ingestion of contaminated water and/or food, the oocyst wall opens (excystation) triggered by temperature, stomach acid, and bile salts. Then, sporulated oocyst ruptures releasing (a) sporozoites that (b) invade the host cell (c) inducing the cell membrane to enclose the parasite in the parasitophorous vacuole, (d) which then differentiates into a trophozoite that undergoes an asexual reproduction, (e) forming a type I meront that contains 6–8 merozoites. These merozoites can reinfect the epithelial cell, where they undergo merogony and form type I meront or (f) type II meront. (g) Merozoites can differentiate into sexually distinct stages called (g) micro- and (h) macrogametocytes. (i) Zygote is formed after the fertilisation of macrogametocyte by the microgametocyte, (j) and this cell undergoes sporogony and produces a thin-walled oocyst. (k) These thin-walled oocysts are released within faeces to infect new hosts, as well as (l) involved in autoinfection process (adapted from Ref. [104]).*

to be a coccidian, *Cryptosporidium* spp. share features of both the coccidia and gregarines, confirmed by morphological and molecular data. Major similarities between *Cryptosporidium* and gregarine parasites are as follows: (1) the ability to complete its life cycle in the absence of host cells, (2) extracellular gamont-like stages, (3) the process in which two mature trophozoites pair up before the formation of gametocyst (szygy), and (4) changing cell architecture to adapt to diverse environments (biofilms, coelom, intestines, soil, and water) [107, 108, 111, 114]. The most recent classification considers *Cryptosporidium* as a separate group within the Apicomplexa. Analyses of comparative genomics and of phylogenetic inference and the ability of *Cryptosporidium* to complete its life cycle extracellularly confirm its close relationship with gregarines and corroborate the transference of *Cryptosporidium* to the Gregarinomorphea class as a new subclass of Cryptogregaria [111, 115]. Early taxonomy at species level was based originally on morphology and host specificity. Nowadays, the description of species

takes molecular analyses, mainly DNA sequencing and PCR-related methods, into account for the detection and differentiation of *Cryptosporidium* spp.
