iSeq 100 for metagenomic pathogen screening in ticks

Abstract

Background Ticks are blood-sucking ectoparasites that play a pivotal role in the transmission of various pathogens to humans and animals. In Korea, Haemaphysalis longicornis is the predominant tick species and is recognized as the vector of pathogens causing various diseases such as babesiosis, borreliosis, rickettsiosis, and severe fever with thrombocytopenia syndrome. Methods In this study, the targeted high-throughput sequencing of the 16S rRNA V4 region was performed using the state-of-the-art sequencing instrument, iSeq 100, to screen bacterial pathogens in H. longicornis, and the findings were compared with those using conventional PCR with specific primers. Microbiome analyses were performed with EzBioCloud, a commercially available ChunLab bioinformatics cloud platform. ANOVA-Like Differential Expression tool (ALDEx2) was used for differential abundance analysis. Results Rickettsia spp. were detected in 16 out of 37 samples using iSeq 100, and this was confirmed using a PCR assay. In the phylogenetic analysis using gltA and ompA sequences of the detected Rickettsia, the highest sequence similarity was found with ‘Candidatus Rickettsia jingxinensis’ isolate Xian-Hl-79, ‘Ca. R. jingxinensis’ isolate F18, and ‘Ca. R. longicornii‘ isolate ROK-HL727. In the microbiome study, Coxiella AB001519, a known tick symbiont, was detected in all 37 tick samples. Actinomycetospora chiangmaiensis was more abundant in Rickettsia-positive samples than in Rickettsia-negative samples. Conclusions In this study, iSeq 100 was used to investigate the microbiome of H. longicornis, and the potentially pathogenic Rickettsia strain was detected in 16 out of 37 ticks. We believe that this approach will aid in large-scale pathogen screening of arthropods to be used in vector-borne disease control programs. Graphical Abstract

Comparative microbiome analysis of Haemaphysalis longicornis ticks at the Korea Combat Training Center in 2022

“…This study also found that all samples from military training centers in Gangwon Province contained Coxiella (Fig. 2), identified as a Coxiella-like endosymbiont (AB001519), consistent with previous studies in the same region (Lee et al 2004;Kim et al 2021a;Sang et al 2022).…”

Parasites and blood-meal hosts of the tsetse fly in Tanzania: a metagenomics study

“…Our method can theoretically detect all potentially pathogenic taxa in samples and simultaneously analyze 96 samples at once. The approximate cost of one run of the iSeq 100 machine is US$ 2000, which takes 18 h to complete [ 19 ].…”

“…The recent development of high-throughput sequencing allows the use of a metagenomic approach to detect all prokaryotic and eukaryotic species in a sample in a single sequencing run at a low cost [ 17 – 19 ]. In this study, we used amplicon deep sequencing of the 12S rRNA and 18S rRNA genes to identify the mammalian hosts of the tsetse fly and associated eukaryotic pathogens (including Trypanosoma spp.…”

First detection of Rickettsia aeschlimannii in Hyalomma marginatum in Tibet, China

“…marginatum feeding on yaks in Tibet, China. mNGS has been successfully used to identify known and/ or unknown microorganisms in arthropods [38,39], thus enabling analysis of microorganism diversity. By mNGS, the most abundant bacteria identified in ticks include Rickettsia spp., Coxiella spp., Francisella spp., and "Candidatus Midichloria mitochondrii" [40].…”