Objective To understand the infection and molecular characteristics of Cryptosporidium spp. in pre-weaned dairy calves in Sichuan Province. Methods The fecal samples were collected from 278 pre-weaned dairy calves with age less than one month in 11 dairy farms(7 in-house farming and 4 free-range farming) located in 10 different regions of Sichuan Province from June 2016 to March 2017. The parasites in the collected fecal samples were concentrated using a modified floatation method with saturated sucrose solution and the fecal parasite DNA was extracted. The small subunit ribosomal RNA(SSU rRNA) of Cryptosporidium was amplified from the fecal DNAs by nested-PCR. The PCR positive fecal samples were used to calculate Cryptosporidium infection rate in these examined caves. Chi-square test was used to compare the difference of infection rates between in-house farming and free-range farming calves. The amplified SSU rRNA PCR products were sequenced and the obtained sequences were BLAST searched against Cryptosporidium sequences deposited in GenBank to identify the infected species. The phylogenetic tree was constructed based on the SSU rRNA sequences by MEGA 7 adjacency method. The M_r 60 000 glycoprotein(gp60) gene was further amplified from the samples identified as C. parvum by SSU rRNA sequencing to further determine their genotypes. Results A total of 278 calves were investigated and the fecal samples were collected from each of them. The SSU rRNA amplification results identified the total infection rate of Cryptosporidium was 14.4% (40/278) in the examined calves. Cryptosporidium infection was found in 10 of the 11 examined farms, among which the highest infection rate was 35.7% (10/28) in Mianyang followed by Aba(35%, 7/20) and Ziyang (30.8%, 8/26). No Cryptosporidium infection was found in Meishan. The infection rate was statistically different among the 10 positive farms(P < 0.05). The Cryptosporidium infection rate in calves in in-house farms was 17.4% (34/195) which is significantly higher than that in free-range farms (7.2%, 6/83) (P < 0.05). Among the 40 Cryptosporidium-positive samples, 28 were identified as C. bovis, 7 as C. parvum and 5 as C. ryanae. After removing the identical sequences, total 10 unique sequences were obtained (ABC881, AYC6953, AYC6969, CDC16111, CDC16117, DYC014, MYC116, MYC117, MYC126, ZYC6874, GenBank login number: MF671870-MF671879). Among them AYC6953, AYC6969, CDC16117 and ZYC6874 were 99% identical to C. bovis (JX515546), DYC014 is 99% identical to C. ryanae (HQ179574), MYC126 is 99% identical to C. bovis (JX416366). ABC881, CDC16111, MYC116 and MYC117 is 100% identical to C. parvum (AH006572), C. ryanae (HQ179574), C. bovis (HQ179573) and C. bovis (MH166335), respectively. The 7 C. parvum-positive samples were further determined as IIdA15G1 subtype which is 100% identical to the Ningxia isolate of KM067092, based on gp60 gene sequencing results. Conclusion The Cryptosporidium infection was prevalent in pre-weaned dairy calves in Sichuan Province and the major infected species were C. bovis, C. parvum and C. ryanae.