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Updated: Jul 6, 2025
Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
Published on: August 29, 2014
Detection of zoonotic Cryptosporidium spp. in small wild rodents using amplicon-based next-generation sequencing
Rosalina Rotovnik1, Tatiana Siegler Lathrop2, Jakob Skov3
1Laboratory of Parasitology, Department of Bacteria, Parasites & Fungi, Statens Serum Institut, Artillerivej 5, DK-2300 Copenhagen S, Denmark.
Abstract:
Rodents may serve as reservoirs of zoonotic species of Cryptosporidium; however, data from molecular surveys in support of this hypothesis are still scarce. In this study, we screened faeces and rectal content from murid and cricetid rodents (N = 58) caught around three farms in Zealand, Denmark, for Cryptosporidium spp. by amplicon-based next-generation sequencing (NGS) of ribosomal genes. Selected samples were further examined using nested conventional PCR targeting SSU rRNA, gp60, and actin genes. Cryptosporidium-specific DNA was identified in 40/58 (69%) samples, and in 12 (30%) of the 40 positive animals, mixed cryptosporidial infections were observed. Cryptosporidium ditrichi was the species most commonly identified, found in 28 (48%) of the animals. Cryptosporidium parvum was identified in 4 (7%) of the animals, all of which were co-infected with C. ditrichi. The present study is the first to utilize NGS-based screening for Cryptosporidium species in wild rodents. Moreover, it is the first study to provide molecular data on Cryptosporidium in rodents sampled in Denmark and to detect DNA of C. ditrichi in Mus musculus, Myodes glareolus, and Microtus agrestis. The NGS approach was successfully applied to yield new knowledge, and the results showed that zoonotic species of Cryptosporidium are common in murid and cricetid rodents in Zealand, Denmark.
Insights
Rodents in Denmark carry common zoonotic Cryptosporidium species, including Cryptosporidium parvum. This study used next-generation sequencing (NGS) to identify Cryptosporidium in wild rodents, revealing high infection rates and mixed infections.
Area of Science:
- Veterinary Parasitology
- Molecular Epidemiology
- Zoonotic Diseases
Background:
- Rodents are potential reservoirs for zoonotic Cryptosporidium species, but molecular data are limited.
- Understanding Cryptosporidium prevalence in rodent populations is crucial for assessing zoonotic risk.
Purpose of the Study:
- To investigate the presence and diversity of Cryptosporidium species in wild rodents in Denmark.
- To utilize next-generation sequencing (NGS) for molecular screening of Cryptosporidium in rodents.
- To determine the prevalence of zoonotic Cryptosporidium species in murid and cricetid rodents.
Main Methods:
- Faecal and rectal samples from 58 rodents (murid and cricetid) in Zealand, Denmark, were analyzed.
- Amplicon-based next-generation sequencing (NGS) of ribosomal genes was employed for Cryptosporidium screening.
- Conventional PCR targeting SSU rRNA, gp60, and actin genes was used for confirmation and genotyping.
Main Results:
- Cryptosporidium DNA was detected in 69% (40/58) of the rodents.
- Mixed Cryptosporidium infections were found in 30% (12/40) of positive animals.
- Cryptosporidium ditrichi was the most prevalent species (48%), followed by Cryptosporidium parvum (7%), often in co-infections.
Conclusions:
- This study is the first to use NGS for Cryptosporidium screening in wild rodents and provides molecular data from Denmark.
- Zoonotic Cryptosporidium species, including C. parvum, are common in wild rodents in Zealand, Denmark.
- The findings highlight the importance of rodents as potential reservoirs for zoonotic Cryptosporidium.
