Rodent-adapted Cryptosporidium infection in humans: Seven new cases and review of the literature.

Cases of cryptosporidiosis in humans have been reported with strong indication of transmission from rodents. Here, we report seven new human cases of cryptosporidiosis involving rodent-adapted species (Cryptosporidium ditrichi [n = 1], Cryptosporidium mortiferum [n = 4; previously known as Cryptosporidium chipmunk genotype I], Cryptosporidium tyzzeri [n = 1], and Cryptosporidium viatorum [n = 1]) and review cases of human infection caused by these four species published to date. The seven new cases were detected in Denmark within a period of twelve months from 2022 to 2023. Only the C. tyzzeri and C. viatorum cases were associated with travel outside Denmark. The total number of human cases of cryptosporidiosis due to C. ditrichi and C. tyzzeri documented to date globally are still limited (4 and 7, respectively), whereas cases involving C. viatorum and C. mortiferum have been detected to a larger extent (43 and 63 cases, respectively). The four new cases of C. mortiferum were all of the XIVaA20G2T1 subtype, which is the only subtype identified so far in Scandinavia, and which is a subtype not yet found outside of Scandinavia. The new C. viatorum case was identified as the XVaA3g subtype. The C. tyzzeri case was subtyped as IXbA6. No subtype data were produced for C. ditrichi due to lack of a subtype assay. Review of existing data suggests the presence of C. ditrichi and C. mortiferum primarily in northern countries and C. tyzzeri and C. viatorum primarily in warmer climates. While our data may further support the role of Cryptosporidium as a cause of zoonotic disease, case descriptions should be obtained where possible to determine if Cryptosporidium species primarily adapted to rodents are the likely cause of symptoms or just an incidental finding.

Complete sequencing of the Cryptosporidium suis gp60 gene reveals a novel type of tandem repeats-Implications for surveillance.

Cryptosporidiosis is an infectious enteric disease caused by species (some of them zoonotic) of the genus Cryptosporidium that in many countries are under surveillance. Typing assays critical to the surveillance of cryptosporidiosis typically involve characterization of Cryptosporidium glycoprotein 60 genes (gp60). Here, we characterized the gp60 of Cryptosporidium suis from two samples-a human and a porcine faecal sample-based on which a preliminary typing scheme was developed. A conspicuous feature of the C. suis gp60 was a novel type of tandem repeats located in the 5' end of the gene and that took up 777/1635 bp (48%) of the gene. The C. suis gp60 lacked the classical poly-serine repeats (TCA/TCG/TCT), which is usually subject to major genetic variation, and the length of the tandem repeat made a typing assay incorporating this region based on Sanger sequencing practically unfeasible. We therefore designed a typing assay based on the post-repeat region only and applied it to C. suis-positive samples from suid hosts from Norway, Denmark, and Spain. We were able to distinguish three different subtypes; XXVa-1, XXVa-2, and XXVa-3. Subtype XXVa-1 had a wider geographic distribution than the other subtypes and was also observed in the human sample. We think that the present data will inform future strategies to develop a C. suis typing assay that could be even more informative by including a greater part of the gene, including the tandem repeat region, e.g., by the use of long-read next-generation sequencing.

Cryptosporidium occultus in disguise.

As data accumulate in GenBank, the difficulties of delineating species of Cryptosporidium based on nuclear small subunit ribosomal RNA (ssu rRNA) gene information alone becomes increasingly evident. Here, we summarize currently available evidence suggesting that several ssu rDNA sequences primarily referred to as Cryptosporidium suis (some of them from non-suid hosts) should be considered Cryptosporidium occultus.

Cryptosporidium species and subtypes identified in human domestic cases through the national microbiological surveillance programme in Sweden from 2018 to 2022.

BACKGROUND: The intestinal protozoan parasite Cryptosporidium is an important cause of diarrheal disease worldwide. A national microbiological surveillance programme was implemented in Sweden in 2018 in order to increase knowledge of the molecular epidemiology of human cryptosporidiosis to better understand transmission patterns and potential zoonotic sources. This article summarises the results of the first five years of the surveillance programme. METHODS: Cryptosporidium-positive faecal and DNA samples from domestically acquired infections were collected from clinical microbiological laboratories in Sweden. Species and subtype determination was performed using 60 kDa glycoprotein and/or small subunit ribosomal RNA gene analysis. RESULTS: Between 2018 and 2022, 1654 samples were analysed and 11 different species were identified: C. parvum (n = 1412), C. mortiferum (n = 59), C. hominis (n = 56), C. erinacei (n = 11), C. cuniculus (n = 5), C. meleagridis (n = 3), C. equi (n = 2), C. ubiquitum (n = 2), and one each of C. canis, C. ditrichi and C. felis. Subtyping revealed seven subtype families of C. parvum (new subtype families IIy and IIz) and 69 different subtypes (11 new subtypes). The most common C. parvum subtypes were IIdA22G1c, IIdA24G1, IIdA15G2R1 and IIaA16G1R1b. For C. hominis, four different subtype families and nine different subtypes (two new subtypes) were identified. For additional species, two new subtype families (IIIk and VId) and nine new subtypes were identified. All successfully subtyped C. mortiferum cases were subtype XIVaA20G2T1, confirming previous findings in Sweden. Several outbreaks were identified of which the majority were foodborne and a few were due to direct contact with infected animals. CONCLUSION: Infection with C. parvum is the leading cause of human cryptosporidiosis acquired in Sweden, where more than 90% of domestic cases are caused by this zoonotic species and only a small proportion of cases are due to infection with other species. The rodent-associated C. mortiferum is considered an emerging zoonotic species in Sweden and the number of domestically acquired human cases has surpassed that of infection with C. hominis. A high diversity of species and subtypes, as well as diversity within the same subtype, was detected. Also, cryptosporidiosis appears to affect adults to a great extent in Sweden.

Cryptosporidium chipmunk genotype I - An emerging cause of human cryptosporidiosis in Sweden.

Most cases of cryptosporidiosis in humans are caused by Cryptosporidium parvum or Cryptosporidium hominis. However, more uncommon species are increasingly being recognised to cause infection in humans. Here we report that Cryptosporidium chipmunk genotype I, which has various rodents as its natural host, is the third most common source of human cryptosporidiosis in Sweden. We also describe the first small outbreak of cryptosporidiosis caused by Cryptosporidium chipmunk genotype I and report the first case of zoonotic transmission of Cryptosporidium chipmunk genotype I from a red squirrel to a human. Cryptosporidium chipmunk genotype I was identified in 20 human cases, including 16 sporadic cases, three outbreak-related cases, and one zoonotic case, as well as in two squirrel samples. Gp60 subtyping which was successful for 19 human cases and two squirrel samples showed that all samples harboured the same subtype, XIVaA20G2T1. The work presented here suggests that red squirrel is a natural host of Cryptosporidium chipmunk genotype I and that infection with Cryptosporidium chipmunk genotype I is an emerging cause of domestic cryptosporidiosis in Sweden and a potential source of outbreaks.

High Diversity of Cryptosporidium Species and Subtypes Identified in Cryptosporidiosis Acquired in Sweden and Abroad.

The intestinal protozoan parasite Cryptosporidium is an important cause of diarrheal disease worldwide. The aim of this study was to expand the knowledge on the molecular epidemiology of human cryptosporidiosis in Sweden to better understand transmission patterns and potential zoonotic sources. Cryptosporidium-positive fecal samples were collected between January 2013 and December 2014 from 12 regional clinical microbiology laboratories in Sweden. Species and subtype determination was achieved using small subunit ribosomal RNA and 60 kDa glycoprotein gene analysis. Samples were available for 398 patients, of whom 250 (63%) and 138 (35%) had acquired the infection in Sweden and abroad, respectively. Species identification was successful for 95% (379/398) of the samples, revealing 12 species/genotypes: Cryptosporidium parvum (n = 299), C. hominis (n = 49), C. meleagridis (n = 8), C. cuniculus (n = 5), Cryptosporidium chipmunk genotype I (n = 5), C. felis (n = 4), C. erinacei (n = 2), C. ubiquitum (n = 2), and one each of C. suis, C. viatorum, C. ditrichi, and Cryptosporidium horse genotype. One patient was co-infected with C. parvum and C. hominis. Subtyping was successful for all species/genotypes, except for C. ditrichi, and revealed large diversity, with 29 subtype families (including 4 novel ones: C. parvum IIr, IIs, IIt, and Cryptosporidium horse genotype Vic) and 81 different subtypes. The most common subtype families were IIa (n = 164) and IId (n = 118) for C. parvum and Ib (n = 26) and Ia (n = 12) for C. hominis. Infections caused by the zoonotic C. parvum subtype families IIa and IId dominated both in patients infected in Sweden and abroad, while most C. hominis cases were travel-related. Infections caused by non-hominis and non-parvum species were quite common (8%) and equally represented in cases infected in Sweden and abroad.

Subtype distribution of zoonotic pathogen Cryptosporidium felis in humans and animals in several countries.

Cryptosporidium felis is the major etiologic agent of cryptosporidiosis in felines and has been reported in numerous human cryptosporidiosis cases. Sequence analysis of the 60-kDa glycoprotein (gp60) gene has been developed for subtyping C. felis recently. In this study, 66 C. felis isolates from the United States, Jamaica, Peru, Portugal, Slovakia, Nigeria, Ethiopia, Kenya, China, India and Australia were subtyped using the newly established tool. Forty-four specimens yielded gp60 sequences, generating 23 subtypes clustered in 4 subtype families (XIXa, XIXc, XIXd and XIXe) with high bootstrap support in a phylogenetic analysis of sequence data. Among them, XIXa showed high genetic diversity at the nucleotide level, with the formation of 18 subtypes from both cats and humans with different geographic distribution. In contrast, all 11 XIXd isolates derived from humans from various countries had identical sequences. Results of this study improve our understanding of the genetic diversity, host specificity and transmission dynamics of C. felis.

Differentiation of Blastocystis and parasitic archamoebids encountered in untreated wastewater samples by amplicon-based next-generation sequencing.

BACKGROUND: Application of next-generation sequencing (NGS) to genomic DNA extracted from sewage offers a unique and cost-effective opportunity to study the genetic diversity of intestinal parasites. In this study, we used amplicon-based NGS to reveal and differentiate several common luminal intestinal parasitic protists, specifically Entamoeba, Endolimax, Iodamoeba, and Blastocystis, in sewage samples from Swedish treatment plants. MATERIALS AND METHODS: Influent sewage samples were subject to gradient centrifugation, DNA extraction and PCR-based amplification using three primer pairs designed for amplification of eukaryotic nuclear 18S ribosomal DNA. PCR products were sequenced using ILLUMINA® technology, and resulting sequences were annotated to species and subtype level using the in-house BION software, sequence clustering, and phylogenetic analysis. RESULTS: A total of 26 samples from eight treatment plants in central/southern Sweden were analysed. Blastocystis sp. and Entamoeba moshkovskii were detected in all samples, and most samples (n = 20) were positive for Entamoeba coli. Moreover, we detected Entamoeba histolytica, Entamoeba dispar, Entamoeba hartmanni, Endolimax nana, and Iodamoeba bütschlii in 1, 11, 4, 10, and 7 samples, respectively. The level of genetic divergence observed within E. nana and E. moshkovskii was 20.2% and 7.7%, respectively, across the ~400-bp region studied, and two clades of E. moshkovskii were found. As expected, Blastocystis sp. subtypes 1-4 were present in almost all samples; however, ST8 was present in 10 samples and was the only subtype not commonly found in humans that was present in multiple samples. CONCLUSIONS: Entamoeba and Blastocystis were identified as universal members of the "sewage microbiome". Blastocystis sp. ST8, which has been rarely reported in humans, was a very common finding, indicating that a hitherto unidentified but common host of ST8 contributed to the sewage influent. The study also provided substantial new insight into the intra-generic diversity of Entamoeba and Endolimax.

From mice to men: Three cases of human infection with Cryptosporidium ditrichi.

Most human cases of cryptosporidiosis are caused by Cryptosporidium parvum or Cryptosporidium hominis. However, the number of recognised Cryptosporidium species, some of which are capable of infecting humans, is continuously increasing. Here we present three human cases infected with Cryptosporidium ditrichi, a recently described species in Apodemus spp. (striped field mouse, yellow-necked mouse, and wood mouse) from various European countries. All three patients were infected in Sweden, but in different years and in different parts of the country. Two patients, from whom clinical data were available, showed symptoms consistent with cryptosporidiosis. For one patient, epidemiological data indicated a possible close contact with mice. The obtained sequences at the small subunit rRNA, actin, and Cryptosporidium oocyst wall protein loci showed 100% identity to C. ditrichi isolates from Apodemus spp., while no 70 kDa heat shock protein gene sequences from C. ditrichi were available for comparison. This study shows the importance of including molecular typing in Cryptosporidium surveillance programmes, and it adds one more species to the plethora of Cryptosporidium spp. hitherto diagnosed in Swedish patients.

Occurrence of Cryptosporidium spp. and Cystoisospora belli among adult patients with diarrhoea in Maputo, Mozambique.

Infections with Cryptosporidium spp. and Cystoisospora belli are important causes of diarrhoea in HIV patients. Nevertheless, information concerning these two parasites is scarce in many African countries, including Mozambique. In this study occurrence of Cryptosporidium spp. and C. belli was investigated by microscopy of stool specimens from 108 adult diarrhoeal patients, most with a confirmed HIV diagnosis. The Cryptosporidium isolates were further characterized by molecular methods. Cryptosporidium and C. belli oocysts were found in 8.3% (9/108), and 25.0% (27/108) of the study participants, respectively. Species identification was possible for all Cryptosporidium isolates with available DNA. The following Cryptosporidium species were detected (number of cases within parentheses): C. parvum (3), C. hominis (3), C. felis (1), and C. hominis/C. parvum (1). Subtyping targeting the gp60 gene revealed two C. hominis isolates with subtype IaA23R3, one C. parvum isolate with IIcA5G3d, and one with IIeA12G1. In summary the occurrence of C. hominis and anthroponotic subtypes of C. parvum indicates that the main route of Cryptosporidium transmission in the present study population was human to human (direct or via food and water). The high prevalence of C. belli highlights the need for early diagnosis of this parasite, for which a treatment exists.

Evaluation of a PCR Method for Detection of Entamoeba polecki, with an Overview of Its Molecular Epidemiology.

Entamoebapolecki is a parasite of human and nonhuman primates, other mammals, and birds. Due to overlapping morphological features, cysts of E. polecki may be confused with those of other Entamoeba species commonly found in human fecal samples, including immature cysts of Entamoeba histolytica Although the presence of E. polecki in human Entamoeba-positive stool samples may be rare, its prevalence is likely underestimated due to such confusion. Here, we give examples of diagnostic approaches applied so far and summarize data on the molecular epidemiology of E. polecki, including host specificity and phylogeography. Moreover, we evaluate a novel diagnostic conventional PCR developed for the screening of fecal samples for E. polecki The assay was highly sensitive and specific when used on genomic DNA extracted directly from stool and Swedish wastewater samples. The PCR enabled the identification of all four subtypes (ST1 to ST4) of E. polecki by PCR product sequencing. Most (23/28) subtyped E. polecki-positive samples detected in patients in Sweden between 2002 and 2015 reflected colonization by ST4 and were seen in travelers/foreigners. Two and three human cases of ST2 and ST3, respectively, were also detected. Subtypes 1, 2, and 3 were detected in 3/21 wastewater samples, suggesting local endemicity of these E. polecki subtypes; interestingly, ST4 was not detected in wastewater. In conclusion, the current PCR assay enables simple and cost-effective screening of fecal and wastewater samples for E. polecki Human cases of E. polecki appear to involve primarily ST4, while E. polecki detected in wastewater may be primarily of animal origin.

Molecular characterization and epidemiological investigation of Cryptosporidium hominis IkA18G1 and C. hominis monkey genotype IiA17, two unusual subtypes diagnosed in Swedish patients.

Cryptosporidium hominis is considered a strictly human-adapted species, and it is only occasionally diagnosed in animals. However, two variants, C. hominis monkey genotype and C. hominis Ik, were originally described in non-human hosts, monkeys and horses, respectively. During a Swedish national Cryptosporidium study, where all samples were analyzed at the small subunit rRNA and the 60 kDa (gp60) glycoprotein loci, we identified two patients infected with C. hominis monkey genotype (subtype IiA17) and two infected with C. hominis subtype IkA18G1. The isolates were further analyzed at the actin and the 70 kDa heat shock protein loci, and these analyses showed that these two subtype families are closely related to each other and to human-adapted C. hominis as well as to Cryptosporidium cuniculus. The two patients with C. hominis monkey genotype infection (a father and son) had visited a monkey farm in Thailand prior to infection, while the two cases with C. hominis Ik were unrelated, both probably infected in Sweden. This is the first time that a monkey genotype infection in humans has been related to contact with monkeys and where the gp60 subtype was identified. It is also the first time that human infection caused by C. hominis subtype Ik is described. Even though we were not able to detect any parasites in the animal samples, zoonotic transmission cannot be ruled out in any of these cases because both subtype families are regarded as animal adapted.

A novel high-resolution multilocus sequence typing of Giardia intestinalis Assemblage A isolates reveals zoonotic transmission, clonal outbreaks and recombination.

Molecular epidemiology and genotyping studies of the parasitic protozoan Giardia intestinalis have proven difficult due to multiple factors, such as low discriminatory power in the commonly used genotyping loci, which has hampered molecular analyses of outbreak sources, zoonotic transmission and virulence types. Here we have focused on assemblage A Giardia and developed a high-resolution assemblage-specific multilocus sequence typing (MLST) method. Analyses of sequenced G. intestinalis assemblage A genomes from different sub-assemblages identified a set of six genetic loci with high genetic variability. DNA samples from both humans (n = 44) and animals (n = 18) that harbored Giardia assemblage A infections, were PCR amplified (557-700 bp products) and sequenced at the six novel genetic loci. Bioinformatic analyses showed five to ten-fold higher levels of polymorphic sites than what was previously found among assemblage A samples using the classic genotyping loci. Phylogenetically, a division of two major clusters in assemblage A became apparent, separating samples of human and animal origin. A subset of human samples (n = 9) from a documented Giardia outbreak in a Swedish day-care center, showed full complementarity at nine genetic loci (the six new and the standard BG, TPI and GDH loci), strongly suggesting one source of infection. Furthermore, three samples of human origin displayed MLST profiles that were phylogenetically more closely related to MLST profiles from animal derived samples, suggesting zoonotic transmission. These new genotyping loci enabled us to detect events of recombination between different assemblage A isolates but also between assemblage A and E isolates. In summary, we present a novel and expanded MLST strategy with significantly improved sensitivity for molecular analyses of virulence types, zoonotic potential and source tracking for assemblage A Giardia.

Improving the genotyping resolution of Cryptosporidium hominis subtype IbA10G2 using one step PCR-based amplicon sequencing.

Cryptosporidium hominis gp60 subtype IbA10G2 is a common cause of cryptosporidiosis. This subtype is responsible for many waterborne outbreaks as well as sporadic cases and is considered virulent and highly important in the epidemiology of cryptosporidiosis. Due to low heterogeneity within the genome of C. hominis it has been difficult to identify epidemiological markers with higher resolution than gp60. However, new markers are required in order to improve outbreak investigations and studies of the transmission dynamics of this clinically important subtype. Based on the whole genome sequences of 17 C. hominis isolates, we have identified several differential loci and developed a new sequence based typing panel with higher resolution than gp60. An amplicon sequencing method was also developed which is based on a one-step PCR which can be sequenced using a Next Generation Sequencing (NGS) platform. Such a system provides a rapid and high-throughput workflow. A panel of nine loci with 10 single nucleotide variants (SNV) was selected and evaluated using clinical IbA10G2 isolates from sporadic, cluster and outbreak associated cases. The specimens were separated into 10 different genetic profiles named sequence types (STs). All isolates within an outbreak or cluster belonged to the same ST, including several samples from the two large waterborne outbreaks which occurred in Sweden between 2010 and 2011 indicating that these outbreaks might be linked. The results demonstrate the methods suitability for improved genotyping of C. hominis IbA10G2.

Early outbreak detection by linking health advice line calls to water distribution areas retrospectively demonstrated in a large waterborne outbreak of cryptosporidiosis in Sweden.

BACKGROUND: In the winter and spring of 2011 a large outbreak of cryptosporidiosis occurred in Skellefteå municipality, Sweden. This study summarizes the outbreak investigation in terms of outbreak size, duration, clinical characteristics, possible source(s) and the potential for earlier detection using calls to a health advice line. METHODS: The investigation included two epidemiological questionnaires and microbial analysis of samples from patients, water and other environmental sources. In addition, a retrospective study based on phone calls to a health advice line was performed by comparing patterns of phone calls between different water distribution areas. RESULTS: Our analyses showed that approximately 18,500 individuals were affected by a waterborne outbreak of cryptosporidiosis in Skellefteå in 2011. This makes it the second largest outbreak of cryptosporidiosis in Europe to date. Cryptosporidium hominis oocysts of subtype IbA10G2 were found in patient and sewage samples, but not in raw water or in drinking water, and the initial contamination source could not be determined. The outbreak went unnoticed to authorities for several months. The analysis of the calls to the health advice line provides strong indications early in the outbreak that it was linked to a particular water treatment plant. CONCLUSIONS: We conclude that an earlier detection of the outbreak by linking calls to a health advice line to water distribution areas could have limited the outbreak substantially.

Possible zoonotic transmission of Cryptosporidium felis in a household.

In humans, the risk of contracting cryptosporidiosis caused by Cryptosporidium felis is considered to be relatively low, and most of the confirmed cases have been observed in immunocompromised patients. Both anthroponotic and zoonotic transmission routes have been suggested. Here, we report a case of suspected zoonotic transmission of C. felis from a cat to a human. The cat developed diarrhea several months before such symptoms were displayed by its owner, a 37-year-old immunocompetent woman. The presence of identical C. felis SSU rRNA, HSP70, and COWP gene sequences was verified in both hosts. In conclusion, it is highly probable that the cat was the initial source of infection and not the opposite. Our results show that Cryptosporidium infection can be transmitted from pets to humans and that molecular analysis is needed to confirm the identity of the oocysts.

Comparative genomic analyses of freshly isolated Giardia intestinalis assemblage A isolates.

BACKGROUND: The diarrhea-causing protozoan Giardia intestinalis makes up a species complex of eight different assemblages (A-H), where assemblage A and B infect humans. Comparative whole-genome analyses of three of these assemblages have shown that there is significant divergence at the inter-assemblage level, however little is currently known regarding variation at the intra-assemblage level. We have performed whole genome sequencing of two sub-assemblage AII isolates, recently axenized from symptomatic human patients, to study the biological and genetic diversity within assemblage A isolates. RESULTS: Several biological differences between the new and earlier characterized assemblage A isolates were identified, including a difference in growth medium preference. The two AII isolates were of different sub-assemblage types (AII-1 [AS175] and AII-2 [AS98]) and showed size differences in the smallest chromosomes. The amount of genetic diversity was characterized in relation to the genome of the Giardia reference isolate WB, an assemblage AI isolate. Our analyses indicate that the divergence between AI and AII is approximately 1 %, represented by ~100,000 single nucleotide polymorphisms (SNP) distributed over the chromosomes with enrichment in variable genomic regions containing surface antigens. The level of allelic sequence heterozygosity (ASH) in the two AII isolates was found to be 0.25-0.35 %, which is 25-30 fold higher than in the WB isolate and 10 fold higher than the assemblage AII isolate DH (0.037 %). 35 protein-encoding genes, not found in the WB genome, were identified in the two AII genomes. The large gene families of variant-specific surface proteins (VSPs) and high cysteine membrane proteins (HCMPs) showed isolate-specific divergences of the gene repertoires. Certain genes, often in small gene families with 2 to 8 members, localize to the variable regions of the genomes and show high sequence diversity between the assemblage A isolates. One of the families, Bactericidal/Permeability Increasing-like protein (BPIL), with eight members was characterized further and the proteins were shown to localize to the ER in trophozoites. CONCLUSIONS: Giardia genomes are modular with highly conserved core regions mixed up by variable regions containing high levels of ASH, SNPs and variable surface antigens. There are significant genomic variations in assemblage A isolates, in terms of chromosome size, gene content, surface protein repertoire and gene polymorphisms and these differences mainly localize to the variable regions of the genomes. The large genetic differences within one assemblage of G. intestinalis strengthen the argument that the assemblages represent different Giardia species.

Subtyping novel zoonotic pathogen Cryptosporidium chipmunk genotype I.

Cryptosporidium chipmunk genotype I is an emerging zoonotic pathogen in humans. The lack of subtyping tools makes it impossible to determine the role of zoonotic transmission in epidemiology. To identify potential subtyping markers, we sequenced the genome of a human chipmunk genotype I isolate. Altogether, 9,509,783 bp of assembled sequences in 853 contigs were obtained, with an N50 of 117,886 bp and >200-fold coverage. Based on the whole-genome sequence data, two genetic markers encoding the 60-kDa glycoprotein (gp60) and a mucin protein (ortholog of cgd1_470) were selected for the development of a subtyping tool. The tool was used for characterizing chipmunk genotype I in 25 human specimens from four U.S. states and Sweden, one specimen each from an eastern gray squirrel, a chipmunk, and a deer mouse, and 4 water samples from New York. At the gp60 locus, although different subtypes were seen among the animals, water, and humans, the 15 subtypes identified differed mostly in the numbers of trinucleotide repeats (TCA, TCG, or TCT) in the serine repeat region, with only two single nucleotide polymorphisms in the nonrepeat region. Some geographic differences were found in the subtype distribution of chipmunk genotype I from humans. In contrast, only two subtypes were found at the mucin locus, which differed from each other in the numbers of a 30-bp minisatellite repeat. Thus, Cryptosporidium chipmunk genotype I isolates from humans and wildlife are genetically similar, and zoonotic transmission might play a potential role in human infections.

Low host-specific Enterocytozoon bieneusi genotype BEB6 is common in Swedish lambs.

Enterocytozoon bieneusi was found in 49/72 (68%) fecal samples from Swedish lambs while 37 samples from 24 adult sheep were negative. Molecular characterization of the internal transcribed spacer (ITS) revealed three genotypes: BEB6, OEB1, and OEB2, the latter two of which were novel and all three of which belonged to a group of genotypes (Group 2) of probably limited zoonotic importance. Our observations suggest that E. bieneusi is common in young Swedish sheep and add support to emerging evidence that the common BEB6 genotype, originally thought to be adapted to cattle, is capable of infecting a variety of hosts, suggesting low host specificity.