*2.1. Occurrence of Cryptosporidium*

All fecal samples were screened for *Cryptosporidium* by nested PCR targeting of the SSU rRNA gene. In total, 36 samples were *Cryptosporidium*-positive, resulting in an overall infection rate of 7.6% (36/476). In total, 11 of 16 Bactrian camel herds tested contained individuals positive for *Cryptosporidium* spp., and the infection rate at the different collection sites ranged from 0–33.3%; the highest infection rate was observed in Qapqal Xibe County (Table 1).


**Table 1.** Occurrence of *Cryptosporidium* species/subtypes in Bactrian camels in Xinjiang, China.

N = Number of positives for *Cryptosporidium*; T = Total analyzed samples.

#### *2.2. Cryptosporidium Species and Subtypes*

Six species were detected from the 36 *Cryptosporidium*-positive samples. *C. andersoni* (*n* = 24) was the predominant species, followed by *C. parvum* (*n* = 6), *C. ubiquitum* (*n* = 2), *C. occultus* (*n* = 2), *C. hominis* (*n* = 1), and *C. bovis* (*n* = 1) (Table 1). The six *C. parvum*-positive samples were identified once again by restriction fragment length polymorphism (RFLP) analysis, and no mixed infections were found. Phylogenetic analysis revealed that all *C. andersoni* sequences were identical to the GenBank sequence KX710084, derived from Bactrian camels in China. Two types of sequences were identified from the six *C. parvum* isolates: *C. parvum* type 1 (*n* = 5) and *C. parvum* type 2 (*n* = 1) were identical to Genbank sequences KX259139 and KX259140, respectively, derived from deer in China. The two sequences of *C. occultus* were identical to sequence MK982467, derived from calves in Bangladesh. Moreover, the sequence of *C. hominis* was identical to sequence KU200955, derived from horses, while the sequence of *C. bovis* was identical to sequence MF074602, derived from dairy cattle in China. Two sequence types were identified in the two *C. ubiquitum* isolates: *C. ubiquitum* type 1 was identical to sequence KT235697, derived from goats in China, while *C. ubiquitum* type 2 represented a new sequence, bearing two single-nucleotide polymorphism (SNP) deletions at positions 485 and 486 and one SNP substitution at position 298 (A to G), compared with KT235697.

Sequence and phylogenetic analysis of the *gp60* gene revealed two subtypes present in the five *C. parvum* isolates: If-like-A15G2 (*n* = 5) and IIdA15G1 (*n* = 1). The sequence of If-like-A15G2 was similar to an isolate derived from a Swedish patient infected in South Africa (JN867334), except for the copy number differences in the trinucleotide repeat (A15G2 versus A12G2). The sequence of IIdA15G1 was identical to sequence KT964798, derived from dairy cattle in China. The single *C. hominis* isolate was subtyped as IkA19G1 and was similar to sequence KU727290, derived from an infected human patient in Sweden (A19G1 versus A18G2). The two new sequences of *C. ubiquitum* identified were identical to one another and subtyped to family XIIa. All of the subtype sequences, If-like, IId, Ik, and XIIa, clustered with published sequences, If-like, IId, Ik, and XIIa, respectively (Figure 2).

**Figure 2.** Phylogenetic relationships between *Cryptosporidium* spp. partial *gp60* sequences obtained in this study and sequences retrieved from the GenBank database. Phylogenetic trees were constructed using neighbor-joining methods based on genetic distance, calculated using the Kimura two-parameter model implemented in MEGA 7.0. Bootstrap values >50% from 1000 replicates are indicated at each node. Isolates from this study are shown in bold.
