目的 探索崇明地区蜱中常见顶复门原虫的感染情况和遗传进化特征，为崇明地区蜱传性疾病的防控工作提供参考。 方法 2023年7—9月，使用布旗法在上海崇明采集游离蜱，形态学鉴定后按蜱种和生长阶段分装（同种成蜱1只/份、若蜱5只/份、幼蜱10只/份），提取蜱基因组DNA，PCR扩增细胞色素c氧化酶亚基1（cox1）基因和16S rDNA基因鉴定蜱种。巢式PCR扩增巴贝虫/泰勒虫18S rRNA基因短片段（约400 bp），阳性样品进一步扩增18S rRNA基因长片段（约1 600 bp）。巢式PCR阳性产物测序后进行BLAST比对，使用MEGA 11软件计算不同样品的遗传距离并构建系统进化树。原虫检出率的比较使用卡方检验。 结果 共采集游离蜱622只，其中长角血蜱255只、褐黄血蜱367只。长角血蜱分装为83份样品（成蜱55份、若蜱16份、幼蜱12份），褐黄血蜱分装为43份样品（成蜱2份、若蜱9份、幼蜱32份）。18S rRNA基因短片段扩增获得17份阳性样品，其中长角血蜱的阳性样品16份（成蜱9份、若蜱7份），褐黄血蜱的阳性样品1份（成蜱）。17份阳性样品均扩增出18S rRNA基因长片段，测序序列的登录号分别为PX453257~PX453273。BLAST比对结果表明：1条序列与巴贝虫的18S rRNA基因序列（MK930513）一致性为99.52%，7条序列与新型原虫Colpodella的18S rRNA基因序列一致性为91.53%~99.94%，9条序列与肾形虫的18S rRNA基因序列一致性为95.66%~99.36%。遗传距离分析结果表明，7条Colpodella序列间的遗传距离为0.003~0.147，9条肾形虫序列间的遗传距离为0.009~0.060，种内序列保守性较高；Colpodella与肾形虫之间的遗传距离为0.124~0.221，不同属之间遗传距离较大。系统进化树结果显示，肾形虫和Colpodella聚为一大分支，巴贝虫与泰勒虫聚为另一大分支。长角血蜱的巴贝虫、Colpodella、肾形虫检出率和原虫总检出率分别为1.20%（1/83）、8.43%（7/83）、9.64%（8/83）和19.28%（16/83），褐黄血蜱的巴贝虫、Colpodella、肾形虫检出率和原虫总检出率分别为0（0/43）、0（0/43）、2.33%（1/43）和2.33%（1/43），长角血蜱的原虫总检出率高于褐黄血蜱（χ² = 6.29，P < 0.05）。 结论 崇明地区游离蜱中存在巴贝虫、肾形虫和新型原虫Colpodella感染，有引发蜱传性疾病的潜在风险。

Objective To investigate the prevalence and genetic characteristics of common Apicomplexan protozoan in ticks collected from Chongming, Shanghai, so as to provide insights into management of tick-borne diseases in Chongming area. Methods Free-living ticks were collected using the flagging method in Chongming, Shanghai, from July to September 2023. Following morphological identification, ticks were pooled according to species and developmental stage (one adult per pool, five nymphs per pool, or ten larvae per pool), and genomic DNA was extracted from each pool. Tick species were molecularly identified by PCR amplification of the cytochrome c oxidase subunit 1 (cox1) gene and 16S rDNA gene. The short fragment of the Babesia/Theileria 18S rRNA gene (approximately 400 bp in length) was amplified using nested PCR assay, and positive samples were further subjected to amplification of the long fragment of the 18S rRNA gene (approximately 1 600 bp in length), and positive nested PCR products were sequenced, followed by sequence alignment with BLAST. Genetic distances were calculated among different samples using the software MEGA 11 and phylogenetic trees were created. Differences in the detection of protozoan were tested for statistical significance with chi-square test. Results A total of 622 free-living ticks were collected, including 255 Haemaphysalis longicornis and 367 H. flava. H. longicornis ticks were divided into 83 pools (55 adult pools, 16 nymph pools and 12 larval pools), and H. flava were divided into 43 pools (2 adult pools, 9 nymph pools and 32 larval pools). Nested PCR amplification of the short fragment of the 18S rRNA gene yielded 17 positive pools, including 16 H. longicornis pools (9 adult pools and 7 nymph pools) and one H. flava pool (adult pool). and the long fragment of the 18S rRNA gene was successfully amplified from all 17 positive pools, with sequences deposited under accession numbers of PX453257 to PX453273. BLAST alignments revealed that one gene sequence showed 99.52% identity with the Babesia 18S rRNA gene sequence (MK930513), 7 gene sequences showed 91.53% to 99.94% identity with the 18S rRNA gene sequences of the novel protozoan Colpodella, and 9 gene sequences showed 95.66% to 99.36% identity with the Colpoda 18S rRNA gene sequences. Genetic distance analysis indicated high intraspecific sequence conservation, with genetic distances ranging from 0.003 to 0.147 among 7 Colpodella gene sequences and from 0.009 to 0.060 among 9 Colpoda gene sequences, and the genetic distance between Colpodella and Colpoda ranged from 0.124 to 0.221, indicating substantial interspecific divergence. Phylogenetic analysis revealed that Colpoda and Colpodella were clustered into a large clade, and Babesia and Theileria were clustered into another large clade. The detection rates of Babesia, Colpodella, Colpoda, and protozoan were 1.20% (1/83), 8.43% (7/83), 9.64% (8/83), and 19.28% (16/83) in H. longicornis, and 0 (0/43), 0 (0/43), 2.33% (1/43), and 2.33% (1/43) in H. flava, respectively. The overall detection of protozoan was significantly higher in H. longicornis than in H. flava (χ² = 6.29, P < 0.05). Conclusion There were Babesia, Colpoda, and novel protozoan Colpodella infections in free-living ticks collected from Chongming area, indicating a potential risk of tick-borne diseases.