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Ticks are the second most important vector capable of transmitting diseases affecting the health of both humans and animals. Amblyomma testudinarium Koch 1844 (Acari: Ixodidae), is a hard tick species having a wide geographic distribution in Asia. In this study, we analyzed the composition of A. testudinarium whole mitogenomes from various geographical regions in Japan and investigated the population structure, demographic patterns, and phylogeographic relationship with other ixodid species. In addition, we characterized a potentially novel tick species closely related to A. testudinarium from Myanmar. Phylogeographic inference and evolutionary dynamics based on the 15 mitochondrial coding genes supported that A. testudinarium population in Japan is resolved into a star‐like haplogroup and suggested a distinct population structure of A. testudinarium from Amami island in Kyushu region. Correlation analysis using Mantel test statistics showed that no significant correlation was observed between the genetic and geographic distances calculated between the A. testudinarium population from different localities in Japan. Finally, demographic analyses, including mismatch analysis and Tajima's D test, suggested a possibility of recent population expansion occurred within Japanese haplogroup after a bottleneck event. Although A. testudinarium has been considered widespread and common in East and Southeast Asia, the current study suggested that potentially several cryptic Amblyomma spp. closely related to A. testudinarium are present in Asia. [ABSTRACT FROM AUTHOR]

Ticks are hematophagous ectoparasites that have a significant impact on their animal hosts. Along with mosquitoes, they are the main arthropod vectors of disease agents in domestic animals, wildlife and humans. To investigate the occurrence and prevalence of piroplasmids in ticks, DNA was extracted from 519 hard ticks collected from 116 hunted Hokkaido sika deer (Cervus nippon yesoensis). The success of the DNA extraction was confirmed by touchdown PCR targeting the mitochondrial 16S rDNA gene of ticks. Touchdown PCR and reverse line blot (RLB) hybridization targeting the 18S rRNA gene were used to detect 14 piroplasm species. All hard ticks parasitizing Hokkaido sika deer were identified as belonging to the genera Ixodes and Haemaphysalis. In total 163 samples (31.4%) were positive for Babesia and Theileria spp. among tick species according to RLB hybridization. Tick DNA hybridized to the oligonucleotide probes of Theileria sp. Thrivae (27.0% of ticks; 140/519), Theileria capreoli (10.6%; 55/519), Babesia divergens-like (1.7%; 9/519), Babesia sp. (Bab-SD) (0.6%; 3/519), Babesia microti U.S. (0.4%; 2/519), and B. microti Hobetsu (0.4%; 2/519). The partial sequencing and phylogenetic analyses of the 18S rRNA gene confirmed the RLB hybridization results. Further investigations are needed to reveal the epidemiology and respective vectors of these pathogens. [ABSTRACT FROM AUTHOR]

A previously undescribed Anaplasma species (herein referred to as AP-sd) has been detected in sika deer, cattle and ticks in Japan. Despite being highly similar to some strains of A. phagocytophilum , AP-sd has never been detected in humans. Its ambiguous epidemiology and the lack of tools for its specific detection make it difficult to understand and interpret the prevalence of this Anaplasma species. We developed a method for specific detection, and examined AP-sd prevalence in Hokkaido wildlife. Our study included 250 sika deer ( Cervus nippon yesoensis ), 13 brown bears ( Ursus arctos yesoensis ) and 252 rodents including 138 ( Apodemus speciosus ), 45 ( Apodemus argenteus ), 42 ( Myodes rufocanus ) and 27 ( Myodes rutilus ) were collected from Hokkaido island, northern Japan, collected during 2010 to 2015. A 770 bp and 382 bp segment of the 16S rRNA and gltA genes, respectively, were amplified by nested PCR. Results were confirmed by cloning and sequencing of the positive PCR products. A reverse line blot hybridization (RLB) based on the 16S rRNA gene was then developed for the specific detection of AP-sd. The prevalence of AP-sd by nested PCR in sika deer was 51% (128/250). We detected this Anaplasma sp. for the first time in wild brown bears and rodents with a prevalence of 15% (2/13) and 2.4% (6/252), respectively. The sequencing results of the 16S rRNA and gltA gene amplicons were divergent from the selected A. phagocytophilum sequences in GenBank. Using a newly designed AP-sd specific probe for RLB has enabled us to specifically detect this Anaplasma species. Besides sika deer and cattle, wild brown bears and rodents were identified as potential reservoir hosts for AP-sd. This study provided a high throughput molecular method that specifically detects AP-sd, and which can be used to investigate its ecology and its potential as a threat to humans in Japan. [ABSTRACT FROM AUTHOR]

Members of the genus Spiroplasma are Gram-positive bacteria without cell walls. Some Spiroplasma species can cause disease in arthropods such as bees, whereas others provide their host with resistance to pathogens. Ticks also harbour Spiroplasma, but their role has not been elucidated yet. Here, the infection status and genetic diversity of Spiroplasma in ticks were investigated using samples collected from different geographic regions in Japan. A total of 712 ticks were tested for Spiroplasma infection by PCR targeting 16S rDNA, and Spiroplasma species were genetically characterized based on 16S rDNA, ITS, dnaA, and rpoB gene sequences. A total of 109 samples originating from eight tick species were positive for Spiroplasma infection, with infection rates ranging from 0% to 84% depending on the species. A linear mixed model indicated that tick species was the primary factor associated with Spiroplasma infection. Moreover, certain Spiroplasma alleles that are highly adapted to specific tick species may explain the high infection rates in Ixodes ovatus and Haemaphysalis kitaokai. A comparison of the alleles obtained suggests that horizontal transmission between tick species may not be a frequent event. These findings provide clues to understand the transmission cycle of Spiroplasma species in wild tick populations and their roles in host ticks. [ABSTRACT FROM AUTHOR]

The tick Amblyomma testudinarium Koch, 1844 (Acari: Ixodidae) is known as a vector of several pathogens such as Rickettsia tamurae and severe fever with thrombocytopenia syndrome (SFTS) virus. This tick species is present in many Asian countries, including Japan, where its distribution is limited to the warm areas of Kanto region and the southwestern region. The present study reports the recovery of a partially engorged A. testudinarium from a wild brown bear captured in Shari town, Hokkaido. In addition to morphological identification, the specimen was genetically characterized by the complete mitochondrial genome sequencing. The results showed that the length of the obtained mitogenome is 14,835 bp that encodes 13 protein-coding, two ribosomal RNA (rRNA) (12S and 16S), and 22 transfer RNA genes with two non-coding control regions. The phylogenetic analysis indicated that our sample clustered with A. testudinarium from Nara, Japan, but separated from A. testudinarium from China. Although the introduction of the tick through livestock transportation cannot be ruled out, the detection of A. testudinarium in Hokkaido prefecture, which is separated from the main island where A. testudinarium is present in the south, may suggest the introduction by migratory birds. This study provides important insights on the distribution and host range of A. testudinarium. This will be useful for the future taxonomic analysis of ticks based on the complete mitogenome sequencing. To our knowledge, this is the northernmost detection point of the tropical tick A. testudinarium., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Many tick-borne pathogens (TBPs) are present in wildlife. The objective of this study is to reveal the role of wild bears in maintaining TBPs. A total of 49 brown bears (Ursus arctos yesoensis) from Hokkaido, and 18 Japanese black bears (Ursus thibetanus japonicus) from Tochigi, and 66 Japanese black bears from Nagano were examined by two molecular methods, reverse line blot (RLB) hybridization, and nested PCR. A total of 5 TBPs (Hepatozoon ursi, Babesia sp. UR2-like group, Cytauxzoon sp. UR1, Babesia sp. UR1, and Babesia microti) were detected from bear blood DNA samples. B. microti was detected from blood DNA samples of Japanese black bear for the first time, with the prevalence of 6.0% (5/84). Out of detected pathogens, H. ursi, Babesia sp. UR2-like pathogens, and Cytauxzoon sp. UR1 were considered as three of the most prevalent TBPs in bears. The prevalence of H. ursi were significantly higher in Japanese black bear (0% vs 96.4%) while that of Babesia sp. UR2-like group was higher in Hokkaido brown bears (89.8% vs 40.5%). The prevalence of Babesia sp. UR1 were significantly higher in Japanese black bears from Tochigi (44.4%), comparing with those from Nagano (18.2%). The prevalence of the detected TBPs were significantly higher in adult bears, comparing with those in younger bears. The present study suggests that Japanese bear species contribute in the transmission of several TBPs in Japan. The expanding distribution of bears might cause the accidental transmission of TBPs to humans and domestic animals.

Due to their toxicity and persistence, several families of brominated flame retardants (BFRs) have been listed as persistent organic pollutants (POPs) in the Stockholm Convention, a multilateral treaty overseen by the United Nations Environment Programme. This treaty mandates that parties who have signed must take administrative and legislative actions to prevent the environmental impacts that POPs pose, both within their jurisdictions and in the global environment. The specific BFRs listed in the Stockholm Convention are Polybrominated Diphenyl Ethers (PBDEs), Hexabromocyclododecane (HBCDD), and Hexabromobiphenyl (HBB), chemicals which must therefore be heavily restricted within the jurisdictions of the signatories. As an example, within the EU, hexabromobiphenyl (HBB), the PBDE commercial mixtures, and HBCDD are almost entirely prohibited in terms of both production and use in commercial goods. Waste articles containing excess concentrations of these BFRs are similarly restricted and must be disposed of in a manner that destroys or irreversible transforms the BFR in question. In some cases, specific exemptions for these limits are defined by the Convention for certain parties: for example, Penta- and Octa-BDE can be present in waste materials for recycling until 2030, while Deca-BDE can be applied to some aviation and automotive applications until 2036. However, in such cases, very specific criteria and guidelines apply for their use and/or production. Worldwide, China, Japan, India, and the United States of America have made significant advances in the regulation of POPs, in line with the provisions of the Stockholm Convention. China has established concentration limits for Penta- and Octa-BDEs in electronic goods. It is also currently availing of an exemption to allow for the use of HBCDD and has not yet ratified the Convention with regards to Deca-BDE. Japan meanwhile has classified HBB and Penta-/Octa-BDE compounds as Class I Specified Chemical Substances which virtually prohibits the manufacture, import, and use of these chemicals in all applications. India has banned the manufacture, trade, import, and use of HBB, HBCDD and some PBDEs, and has established concentration limits for all PBDEs in certain electrical goods. Finally, the United States has no federal mandate for the restriction of POPs and has not ratified the annexes to the Convention requiring them to do so. However, thirteen states have implemented their own state-wide concentration limits on a variety of flame retarding chemicals in various commercial applications. Though these limits worldwide are a very positive step for the removal of POP-BFRs from the environment, the increased use of replacement flame retardants renders such legislation only partially effective. The lack of effective screening mechanisms in waste management facilities means that BFR-treated plastics can be inadvertently recycled and remain in circulation. The rise in the use of novel BFRs (NBFRs) can furthermore hinder screening methods currently being developed and the additives themselves may pose similar issues to their predecessors owing to their similar chemical properties. Thus, restrictions on current BFRs will result in the use of new flame retardants, which may in turn be banned and replaced once again. Further research into and development of methods to screen for hazardous chemicals in end of life materials is therefore of the utmost importance. This must be coupled with pro-active legislation that eliminates the need for using such persistent and potentially harmful chemicals in the future., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)