Dataset Information

Utility of a Sequence-Independent, Single-Primer-Amplification (SISPA) and Nanopore Sequencing Approach for Detection and Characterization of Tick-Borne Viral Pathogens.

ABSTRACT: Currently, next generation sequencing (NGS) is the mainly used approach for identification and monitorization of viruses with a potential public health threat in clinical and environmental samples. To facilitate detection in NGS, the sequence-independent, single-primer-amplification (SISPA) is an effective tool for enriching virus sequences. We performed a preliminary assessment of SISPA-nanopore sequencing as a potential approach for screening tick-borne viruses in six specimens with detectable Crimean-Congo hemorrhagic fever virus (CCHFV) and Jingmen tick virus (JMTV) sequences. A comparison of unbiased NGS and SISPA followed by nanopore sequencing was carried out in 4 specimens with single and pooled ticks. The approach was further used for genome sequencing in culture-grown viruses. Overall, total/virus-specific read counts were significantly elevated in cell culture supernatants in comparison to single or pooled ticks. Virus genomes could be successfully characterized by SISPA with identities over 99%. Genome coverage varied according to the segment and total read count. Base calling errors were mainly observed in tick specimens and more frequent in lower viral loads. Culture-grown viruses were phylogenetically-related to previously-reported local viruses. In conclusion, the SISPA + nanopore sequencing was successful in generating data comparable to NGS and will provide an effective tool for broad-range virus detection in ticks.

SUBMITTER: Brinkmann A

PROVIDER: S-EPMC7911436 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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Utility of a Sequence-Independent, Single-Primer-Amplification (SISPA) and Nanopore Sequencing Approach for Detection and Characterization of Tick-Borne Viral Pathogens.

Brinkmann Annika A Uddin Steven S Krause Eva E Surtees Rebecca R Dinçer Ender E Kar Sırrı S Hacıoğlu Sabri S Özkul Aykut A Ergünay Koray K Nitsche Andreas A

Viruses 20210129 2

Currently, next generation sequencing (NGS) is the mainly used approach for identification and monitorization of viruses with a potential public health threat in clinical and environmental samples. To facilitate detection in NGS, the sequence-independent, single-primer-amplification (SISPA) is an effective tool for enriching virus sequences. We performed a preliminary assessment of SISPA-nanopore sequencing as a potential approach for screening tick-borne viruses in six specimens with detectable ...[more]

PMID: 33572847

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Project description:Ticks are blood-feeding arthropods and transmit a variety of medically important viral, bacterial, protozoan pathogens to animals and humans. Ticks also harbor a diverse community of microbes linked to their biological processes, such as hematophagy, and hence affect vector competence. The interactions between bacterial and/or protozoan pathogens and the tick microbiome is a black-box, and therefore we tested the hypothesis that the presence of a protozoan or bacterial pathogen will alter the microbial composition within a tick. Hence, this study was designed to define the microbial composition of two tick species, Hyalomma (H.) anatolicum and Rhipicephalus (R.) microplus. We used a combination of PCR based pathogen (Anaplasma marginale and Theileria species) and symbiont (Wolbachia species) identification followed by metagenomic sequencing and comparison of the microbial communities in PCR positive and negative ticks. A total of 1786 operational taxonomic units was identified representing 25 phyla, 50 classes, and 342 genera. The phylum Proteobacteria, Firmicutes, Actinobacteriota, and Bacteroidota were the most represented bacteria group. Alpha and beta diversity were not significantly affected in the presence or absence of Theileria sp. and A. marginale as see with H. anatolicum ticks. Interestingly, bacterial communities were significantly reduced in Theileria sp. infected R. microplus ticks, while also exhibiting a significant reduction in microbial richness and evenness. Putting these observations together, we referred to the effect the presence of Theileria sp. has on R. microplus a "pathogen-induced dysbiosis". We also identify the presence of Plasmodium falciparum, the causative agent of human malaria from the microbiome of both H. anatolicum and R. microplus ticks. These findings support the presence of a "pathogen-induced dysbiosis" within the tick and further validation experiments are required to investigate how they are important in the vector competence of ticks. Understanding the mechanism of "pathogen-induced dysbiosis" on tick microbial composition may aid the discovery of intervention strategies for the control of emerging tick-borne infections.

Project description:BackgroundIxodes ricinus and Ixodes persulcatus are the main vectors of Lyme borreliosis spirochetes and several other zoonotic bacteria in northern Europe and Russia. However, few studies screening bacterial pathogens in Finnish ticks have been conducted. Therefore, reports on the occurrence and prevalence of several bacterial pathogens detected from ticks elsewhere in Europe and Russia are altogether missing from Finland. The main aim of the current study was to produce novel data on the occurrence and prevalence of several tick-borne bacterial pathogens in ticks collected from southwestern Finland.MethodsTicks were collected in 2013-2014 by blanket dragging from 25 localities around southwestern Finland, and additionally from a dog in Lempäälä. Collected ticks were molecularly identified and screened for Borrelia burgdorferi s.l., Borrelia miyamotoi, Rickettsia, Bartonella and Candidatus Neoehrlichia mikurensis using quantitative PCR. Furthermore, detected Rickettsia spp. were sequenced using conventional PCR to determine species.ResultsA total of 3169 ticks in 1174 DNA samples were screened for the listed pathogens. The most common bacteria detected was B. burgdorferi (s.l.) (18.5 % nymphal and 23.5 % adult ticks), followed by Rickettsia spp. (1.1 %; 5.1 %) and B. miyamotoi (0.51 %; 1.02 %). B. miyamotoi and Rickettsia spp. were also detected in larval samples (minimum infection rates 0.31 % and 0.21 %, respectively). Detected Rickettsia spp. were identified by sequencing as R. helvetica and R. monacensis. All screened samples were negative for Bartonella spp. and Ca. N. mikurensis.ConclusionsIn the current study we report for the first time the presence of Rickettsia in Finnish ticks. Furthermore, Rickettsia spp. and B. miyamotoi were found from larval tick samples, emphasizing the importance they may have as vectors of these pathogens. Comparisons of tick density estimates and B. burgdorferi (s.l.) prevalence made between the current study and a previous study conducted in 2000 in ten out of the 25 study localities suggest that an increase in tick abundance and B. burgdorferi (s.l.) prevalence has occurred in at least some of the study localities.

Dataset Information

Utility of a Sequence-Independent, Single-Primer-Amplification (SISPA) and Nanopore Sequencing Approach for Detection and Characterization of Tick-Borne Viral Pathogens.

Publications

Utility of a Sequence-Independent, Single-Primer-Amplification (SISPA) and Nanopore Sequencing Approach for Detection and Characterization of Tick-Borne Viral Pathogens.

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