**1. Introduction**

Cryptosporidiosis is a global parasitic disease, which usually presents with selflimiting diarrhea. However, the disease can be severe, especially in immunocompromised and malnourished individuals [1]. The causative agent is the intestinal protozoan parasite *Cryptosporidium*, which infects a wide range of animals, including humans [2]. There are more than 40 recognized species described, but 2 in particular (*Cryptosporidium hominis* and *Cryptosporidium parvum*) account for most cases of human cryptosporidiosis [3]. In addition, around 20 other species/genotypes have also been observed in humans, including *Cryptosporidium meleagridis*, *Cryptosporidium cuniculus*, *Cryptosporidium ubiquitum*, *Cryptosporidium canis*, *Cryptosporidium felis*, *Cryptosporidium viatorum*, and *Cryptosporidium* chipmunk genotype I [3,4].

Cryptosporidiosis has been a notifiable disease in Sweden since 2004, with an increasing incidence from 69 (0.8 cases/100,000 inhabitants) cases annually in 2005 to 1088

Winiecka-Krusnell, J.; Stensvold, C.R.; Beser, J. High Diversity of *Cryptosporidium* Species and Subtypes Identified in Cryptosporidiosis Acquired in Sweden and Abroad. *Pathogens* **2021**, *10*, 523. https:// doi.org/10.3390/pathogens10050523

Academic Editors: David Carmena, David González-Barrio and Pamela Carolina Köster

Received: 9 April 2021 Accepted: 23 April 2021 Published: 26 April 2021

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(10.3/100,000 inhabitants) cases in 2019 [5]. There are several reasons for this increase, including better general knowledge of cryptosporidiosis, and the introduction of more sensitive diagnostic methods and, perhaps more importantly, multiple-agent diagnostic approaches, whereby clinicians do not need to specifically request testing for *Cryptosporidium.* Where this approach is used, the number of reported cases tends to increase [6,7]. Nevertheless, there is substantial local variation in the number of reported cases, which suggests that cryptosporidiosis is still underdiagnosed in Sweden [5].

Knowledge about the distribution of *Cryptosporidium* species and subtypes within a country is crucial for the management of cryptosporidiosis and to identify and understand transmission patterns and potential zoonotic sources [8]. The first larger molecular study of sporadic cryptosporidiosis cases in Sweden was conducted between April 2006 and November 2008. Patients from the Stockholm metropolitan area diagnosed with cryptosporidiosis were investigated, and 194 samples were successfully genotyped. A high occurrence of *C. parvum* was found; 111 cases vs. 65 of *C. hominis*. Only 17 (26%) of the *C. hominis* infections were acquired in Sweden, compared with 57 (51%) of the *C. parvum* infections. In addition, less common species such as *C. meleagridis* (*n* = 11), *C. felis* (*n* = 2), *C. viatorum* (*n* = 2), and *Cryptosporidium* chipmunk genotype I (*n* = 2) were found [9].

The largest known *Cryptosporidium* outbreaks in Sweden (and Europe) occurred in Östersund (Jämtland County) and Skellefteå (Västerbotten County) in 2010 and 2011. Both these outbreaks were caused by contamination of the municipal drinking water with *C. hominis* subtype IbA10G2, together affecting an estimated 45,500 persons [10,11]. *Cryptosporidium parvum* has been documented in water- and foodborne outbreaks as well as in outbreaks caused by animal contacts [12–14].

The primary aim of the present study was to identify if observations from the Stockholm metropolitan area (i.e., predominance of *C. parvum* but with relatively frequent detection of unusual species) reflected the situation in the rest of Sweden, and thereby to geographically expand the molecular investigation of cryptosporidiosis cases in Sweden. The second aim was to investigate if the *C. hominis* subtype IbA10G2 was established in the population following the large waterborne outbreaks in 2010 and 2011.

#### **2. Materials and Methods**
