Archives of Virology https://doi.org/10.1007/s00705-021-05296-4 ORIGINAL ARTICLE Screening for tick‑borne and tick‑associated viruses in ticks collected in Ghana Michael Amoa‑Bosompem1,2,3,8 · Daisuke Kobayashi1 · Astri Nur Faizah1,4 · Shohei Kimura5 · Ama Antwi3 · Esinam Agbosu6 · Deborah Pratt6 · Mitsuko Ohashi3,5 · Joseph H. Kofi Bonney6 · Samuel Dadzie3 · Hiroko Ejiri1 · Nobuo Ohta7 · Kyoko Sawabe1 · Shiroh Iwanaga5,9 · Haruhiko Isawa1 Received: 12 February 2021 / Accepted: 1 October 2021 © The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature 2021 Abstract Ticks are blood-sucking arthropods that transmit many pathogens, including arboviruses. Arboviruses transmitted by ticks are generally referred to as tick-borne viruses (TBVs). TBVs are known to cause diseases in humans, pets, and livestock. There is, however, very limited information on the occurrence and distribution of TBVs in sub-Saharan Africa. This study was designed to determine the presence and distribution of ticks infesting dogs and cattle in Ghana, as well as to identify the tick-borne or tick-associated viruses they harbour. A more diverse population of ticks was found to infest cattle (three genera) relative to those infesting dogs (one genus). Six phleboviruses and an orthonairovirus were detected in tick pools screened by RT-PCR. Subsequent sequence analysis revealed two distinct phleboviruses and the previously reported Odaw virus in ticks collected from dogs and a virus (16GH-T27) most closely related to four unclassified phleboviruses in ticks collected from cattle. The virus 16GH-T27 was considered a strain of Balambala tick virus (BTV) and named BTV strain 16GH-T27. Next-generation sequencing analysis of the BTV-positive tick pool detected only the L and S segments. Phylogenetic analysis revealed that BTV clustered with viruses previously defined as M-segment-deficient phleboviruses. The orthonairovirus detected in ticks collected from cattle was confirmed to be the medically important Dugbe virus. Furthermore, we discuss the importance of understanding the presence and distribution of ticks and TBVs in disease prevention and mitigation and the implications for public health. Our findings contribute to the knowledge pool on TBVs and tick-associated viruses. Introduction Ticks are among the most important vectors of arboviruses, transmitting many infectious pathogens of medical and veterinary importance [1, 2]. Arbovirus-transmitting tick Handling Editor: Patricia Aguilar. * Haruhiko Isawa 5 Department of Environmental Parasitology, Tokyo Medical hisawa@nih.go.jp and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan 1 Department of Medical Entomology, National Institute 6 of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Department of Virology, Noguchi Memorial Institute Tokyo 162-8640, Japan for Medical Research, University of Ghana, College of Health Sciences, P.O. Box LG581, Legon, Accra, Ghana 2 Laboratory of Sanitary Entomology, Faculty 7 of Agriculture, Kyushu University, 744 Motooka, Nishi-ku, Faculty of Health Science, Suzuka University of Medical Fukuoka 819-0395, Japan Science, 1001-1 Kishioka-cyo, Suzuka-shi, Mie 510-0293, Japan 3 Department of Parasitology, Noguchi Memorial Institute 8 for Medical Research, University of Ghana, College Present Address: Department of Biomedical and Diagnostic of Health Sciences, P.O. Box LG581, Legon, Accra, Ghana Sciences, School of Veterinary Medicine, University of Tennessee, 2407 River Drive, Knoxville, TN 37996, USA 4 Graduate School of Agricultural and Life Science, 9 The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Present Address: Research Institute for Microbial Diseases, Tokyo 113-8657, Japan 3-1 Yamadaoka, Osaka 565-0871 Suita, Japan Vol.:(012 3456789) M. Amoa-Bosompem et al. species can be broadly categorized as soft ticks belonging (DUGV), have been detected in ticks infesting livestock to the genera Ornithodoros, Carios and Argas and hard ticks in Ghana [12, 13]; however, information on the prevalence of the genera Ixodes, Haemaphysalis, Hyalomma, Ambly- and distribution of TBVs in Ghana is limited. Therefore, omma, Dermacentor, Rhipicephalus, and Boophilus [3–5]. this study was designed to investigate the presence and The ability of ticks to infest multiple host species throughout distribution of TBVs in ticks collected from dogs and cat- their different life cycle stages compounds the prominence tle across Ghana and assess the risk of TBV infection in and risk of ticks as vectors of arboviruses as host migration humans and livestock. facilitates the spread of tick-borne viruses into new areas [6]. The survival of ticks in any environment, however, depends on climatic conditions such as temperature, humidity, and precipitation [7, 8]. For example, the activity of Hyalomma Materials and methods marginatum ticks, the primary vector of Crimean-Congo hemorrhagic fever virus (CCHFV), peaks in Turkey during Study site and sample collection the summer when the temperatures range from 30°C to 40°C and maximum relative humidity ranges from 20% to 50% Ticks were collected from three different vegetations [6]. Thus, the distribution and density of ticks are depend- spanning four regions in Ghana in August and Septem- ent on and limited by the presence of hosts as well as the ber 2016. Accra (5o38’28.0”N 0o10’32.7”W), a coastal prevailing climatic conditions. savannah in the Greater Accra region, is an urban area; Tick-borne viruses (TBV) are vertebrate-infecting viruses Hohoe (7o09’32.8”N 0o28’39.6”E), a deciduous forest in transmitted by ticks. There are approximately 160 known the Volta region, is semi-urban; Larabanga (9o13’01.2”N TBVs, transmitted by about 10% of known tick species [4]. 1o51’25.4”W) in the savannah region and Jirapa TBVs are classified into eight RNA viral families (Flavi- (10o31’53.2”N 2 o41’59.8”W) in the Upper West region viridae, Nairoviridae, Orthomyxoviridae, Reoviridae, Rhab- are rural areas with a Guinea savannah vegetation (Fig. 1) doviridae, Phenuiviridae, Peribunyaviridae, and Nyamiviri- [14]. dae) and one DNA viral family (Asfarviridae) [4]. These Tick samples were collected from dogs in households TBVs cause at least 25 diseases in humans, pets, and live- in Larabanga and dogs that had been brought to veterinary stock [9]. hospitals in Accra. Tick samples from cattle were collected At least five genera of ticks have been identified in in Hohoe, Larabanga, and Jirapa (Supplementary Table S1). Ghana, a country located along the coast of the Gulf of Questing ticks were collected from cattle-grazing fields in Guinea in West Africa, where it is warm all year round. Larabanga by dragging a flannel sheet (70 × 100 cm) as The five genera, all of which are medically important for described by Kobayashi et al. [15]. Sampled ticks were TBV transmission, include Ixodes, Amblyomma, Haema- classified to the species or genus level. In addition, the sex, physalis, Rhipicephalus, and Hyalomma [3, 10, 11]. Two developmental stage, and feeding status were recorded (Sup- medically important TBVs, CCHFV and Dugbe virus plementary Table S1) [13, 16]. Fig. 1 Map showing sample collection sites. Map of Africa Jirapa (inset) showing the location of Ghana (shaded). Map of Ghana, showing the sample collection sites (yourfreetemplate.com) Larabanga Hohoe Accra 1 3 Tick-borne and tick-associated viruses in Ghana Virus isolation and detection BLASTx. The virus-positive pools were subjected to NGS analysis to determine unknown sequences [15, 18]. Virus isolation was performed using BHK-21 cells (derived from Syrian hamster kidney, Japan Health Sci- Phylogenetic analysis ence Research Resources Bank, Osaka, Japan) as described previously [17]. Briefly, homogenized tick pools were The online version of MAFFT 7 was used to align amino inoculated on a monolayer of BHK-21 cells and incubated acid sequences [19]. MEGA ver. 7 was used for determi- at 37 °C and 5% CO2 for 7 days. After two blind passages, nation of a suitable amino acid substitution model and for cell culture supernatants were stored at −80 °C until used. dendrogram construction [20]. The culture supernatant from the virus isolation process was subjected to next-generation sequencing (NGS) analy- Complete coding sequence (CDS) determination sis using an Illumina MiniSeq System as described pre- of DUGV strain 15AC‑T25 viously [14]. CLC Genomics Workbench software (CLC Bio, Aarhus, Denmark) was used to assemble reads into NGS analysis was performed on the cell culture supernatant contigs, which were used to search the National Centre for after six passages of DUGV strain 15AC-T25, isolated from Biotechnology Information database using the Basic Local Amblyomma variegatum ticks collected from cattle in Accra Alignment Search Tool (BLAST)n and BLASTx. in 2015 [13]. The complete CDS of each genome segment of In addition to virus isolation and NGS analysis, virus the strain was determined, and similarities to known strains detection was performed by conventional RT-PCR. Total of DUGV were assessed. RNA was extracted from tick homogenates using ISO- GEN II (Nippon Gene, Tokyo, Japan). Extracted RNA was screened for selected medically important tick-borne Results or tick-associated viruses of the genera Alphavirus, Fla- vivirus, Orthonairovirus, Phlebovirus, and Thogotovirus, Tick sampling and classification using a PrimeScript One Step RT-PCR Kit Ver. 2 (Takara Bio, Shiga, Japan) and universal primers (Table 1). The Ticks were collected from dogs, cattle, and cattle-graz- reaction conditions for RT-PCR were as follows: 50°C for ing fields in Ghana and classified morphologically at the 30 min, 94°C for 2 min, and 35 cycles of 94°C for 30 genus or species level. Ticks of at least three genera were s, 53°C for 30 s, and 72°C for 30 s. Amplified products detected in this study (Supplementary Table S1). All ticks were purified from an agarose gel after electrophoresis, collected from dogs (in both Accra and Larabanga) were sequenced directly, and identified using BLASTn and of the genus Rhipicephalus, while a more diverse collec- tion of ticks was obtained from cattle across the collection Table 1 Primers used for RT-PCR-based virus screening Target viruses Primer name Primer sequence (5' - 3') Reference Genus Orthonairovirus Nairo Forward TCTC AA AGA AAC ACGT GC CGC Lambert and Lanciotti, 2009 Nairo Reverse GTC CTT CCTC CA CTT GWG RGC AGC CTG CTGG TA Tick-borne phleboviruses TBPVL2759F CAG CAT GGIGGICTIAGA GAG AT Matsuno et al., 2015 TBPVL3267R TGIAGIATSCCYT GC ATCAT Genus Flavivirus* FU1 TAC AAC ATG ATG GGA AAGA GAG AGA A Kuno et al., 1998 cFD2 GTG TCC CAG CCG GCG GTG TCAT CA GC FU2 GCT GATG AC ACC GCC GGC TGG GAC AC cFD3 AGCA TGT CT TCCG TG GTC ATCCA Genus Alphavirus AJUN CTSTAC GGY KRWC CTA AAT Miller et al., 2000 CCAP RTA YTGS ACWGCKCCR TGRT GCCA Genus Orthobunyavirus BCS82C ATG ACT GAG TTGG AG TTT CAT GATG TCGC Kuno et al., 1996 (Bunyamwera and California BCS332V TGT TCC TGT TGCC AG GAAA AT serogroups) Genus Thogotovirus THOV-uni-1-170F AAR AGRT ACA CTA CR AGC AAGAA Current study THOV-uni-1-600R GCTG WA TTG GGG RCA GAA SACTTG *Two different primer sets (FU1 and cFD2, FU2 and cFD3) were used in detecting flaviviruses in this study. 1 3 M. Amoa-Bosompem et al. sites (Supplementary Table S1). The ticks collected from and amino acid sequence identity with both the L and S cattle were Amblyomma spp. (including A. variegatum), Rhi- segments of 16GH-T27. 16GH-T27 can therefore be consid- picephalus spp., Hyalomma spp., and Ixodidae spp. (Sup- ered a strain of BTV (Table 2; Fig. 2). The amino acid and plementary Table S1). In total, 354 ticks divided into 93 nucleotide sequence identity values for the L and S segments pools were subjected to virus isolation, RT-PCR, and NGS of the four related unclassified phleboviruses are shown in analysis (Supplementary Table S1). Table 3 and Supplementary Table S2, respectively. The RT-PCR-amplified product of the orthonairovirus- Virus detection and sequence analysis positive pool, 16GH-T89, was subjected to Sanger sequenc- ing analysis. The resulting sequence shared 97% identity In screening for tick-borne or tick-associated viruses, the with the S segment of DUGV strain IbAr 1792 detected in second supernatant from virus isolation and the homogen- A. variegatum ticks picked off cattle in Nigeria [22]. Sub- ate from pooled tick samples were subjected to NGS and sequent determination of the complete genome sequence of RT-PCR analysis, respectively. No viruses were detected or DUGV strain 15AC-T25, consisting of a 12,201-nt L seg- isolated by the virus isolation method. RT-PCR analysis of ment (GenBank accession number LC579816), a 4,849- tick homogenates revealed that, out of 93 pools, seven were nt M segment (GenBank accession number LC579817), positive for RNA viruses. We detected phleboviruses in six and a 1,691-nt S segment (GenBank accession number pools and orthonairoviruses in one pool (Table 2). The phle- LC579818), showed the S segment of 15AC-T25 and boviruses detected in pools 16GH-T12, 16GH-T13, 16GH- 16GH-T89 to be nearly identical. 16GH-T89 was therefore T14, and 16GH-T15 were confirmed to be strains of Odaw confirmed to be DUGV (Table 2). In addition, pairwise virus (ODWV), which was detected previously in Rhipiceph- alignment analysis of 15AC-T25 by BLASTn also showed alus ticks from Accra [13] (Table 2). The remaining two 97%–98% sequence identity to the L, M, and S segments phlebovirus-positive pools, 16GH-T24 and 16GH-T27, had of DUGV strain IbAr 1792 (GenBank accession numbers 95% nucleotide sequence identity to Balambala tick virus KU925455, KU925456, and KU925457, respectively) [22], (BTV) detected in H. rufipes in Kenya and 92% nucleotide confirming that 15AC-T25 has the typical genome organisa- sequence identity to a tick phlebovirus detected in H. mar- tion of DUGV. ginatum sampled from sheep in Turkey [21]. The viruses in 16GH-T24 and 16GH-T27 were considered to be the same, as their nucleotide sequences were 100% identical within Discussion the limits of the RT-PCR-amplified region. NGS analysis of 16GH-T27 and subsequent de novo assembly coupled with Ghana is a West African country slightly north of the equa- Sanger sequencing resulted in the detection of 5,691-nt and tor with an average temperature range of 24°–30 °C [23] and 1,547-nt partial-yet-continuous regions of the viral L and 50–80% humidity. It is home to an abundance of domesti- S segment, respectively. Phylogenetic analysis using amino cated animals, including pets and livestock [24]. Dogs and acid sequences showed 16GH-T27 to form a clade with four cats are the most common companion animals in Ghana; unclassified phleboviruses, the most closely related being dogs also serve as a form of security in some households. BTV (Fig. 2). Similarly, BTV shared the highest nucleotide The most common livestock in Ghana are cattle, goats, Table 2 Viruses detected in this study Source Virus name Strain Location Tick species Number, stage, and sex of pooled Host animals GenBank accession number ticks Odaw virus 16GH-T12 Accra Rhipicephalus sp. 13 adult males* Dogs LC579647 16GH-T13 Accra Rhipicephalus sp. 14 adult males* Dogs LC589706 16GH-T14 Accra Rhipicephalus sp. 10 adult females* Dogs LC589707 16GH-T15 Accra Rhipicephalus sp. 8 adult females* Dogs LC589708 Balambala tick virus 16GH-T24 Jirapa Amblyomma sp. 3 adult females*, 1 adult female** Cattle LC579820 16GH-T27 Jirapa Hyalomma sp. 9 adult males* Cattle LC579819 (L segment) LC589978 (S segment) Dugbe virus 16GH-T89 Hohoe Rhipicephalus sp. 5 adult females** Cattle LC579815 * Non-engorged ** Engorged (partially or fully engorged) 1 3 Tick-borne and tick-associated viruses in Ghana 87 Kaisodi virus MG581739.1 Kaisodi group Khasan virus KF892046.1 Odaw virus strain 16GH-T12 Odaw virus LC193452.1 62 92 Phlebovirus sp. SL3-L ASD49914.1 Odaw virus strain 16GH-T14 85 Odaw virus strain 16GH-T13 Odaw virus strain 16GH-T15 M segment-deficient phlebovirus Phlebovirus sp LC146411.1 99 Tick phlebovirus KY965999.1 73 Bole Tick Virus 1 AJG39234.1 74 Balambala tick virus strain 16GH-T27 52 Balambala tick virus MW561966.1 Xinjiang tick phlebovirus QBQ64947.1 Uukuniemi virus NC 005214.1 Kabuto mountain virus LC483650.1 Tick phlebovirus KY965998.1 99 Lesvos virus KX452150.1 M segment-deficient phlebovirus Dabieshan Tick Virus KM817666.1 Yongjia Tick Virus 1 KM817704.1 Lone Star virus KC589005.1 100 Bhanja virus JX961619.1 Bhanja group 88 Palma virus JX961628.1 0.20 Fig. 2 Phylogenetic analysis of Balambala tick virus (BTV) strain 16GH-T14, and 16GH-T15, respectively). The Bhanja virus group 16GH-T27. An outgroup-rooted maximum-likelihood tree was con- was used as the outgroup. Bootstrap support values from 1000 boot- structed, using the LG + G substitution model, based on complete strap replicates are indicated on the branches. Viruses detected/iso- RdRp amino acid sequences with GenBank accession numbers lated in this study are preceded by a bullet (•) and shown in boldface LC579819 (BTV strain 16GH-T27) and LC579820, LC589706, type. The scale bar represents the number of substitutions per site. LC589707, and LC589708 (ODWV strains 16GH-T12, 16GH-T13, Table 3 L segment nucleotide and amino acid sequence identity values for the four related unclassified phleboviruses Virus name Accession number Reference virus Accession number Region analyzed Nucleotide Amino acid sequence iden- sequence identity tity (%) (%) Balambala tick virus LC579819 Balambala tick virus MW561966.1 Complete CDS 93.13 97.65 strain 16GH-T27* Bole tick virus 1 AJG39234.1 Complete CDS 71.20 77.95 Tick phlebovirus KY965999.1 Partial CDS 78.00 81.18 Xinjiang tick phlebo- QBQ64947.1 Complete CDS 72.90 81.45 virus Balambala tick virus MW561966.1 Bole tick virus 1 AJG39234.1 Complete CDS 73.98 79.38 Tick phlebovirus KY965999.1 Partial CDS 78.52 78.74 Xinjiang tick phlebo- QBQ64947.1 Complete CDS 74.42 82.11 virus Bole tick virus 1 AJG39234.1 Tick phlebovirus KY965999.1 Partial CDS 74.51 90.00 Xinjiang tick phlebo- QBQ64947.1 Complete CDS 74.30 81.15 virus Tick phlebovirus KY965999.1 Xinjiang tick phlebo- QBQ64947.1 Complete CDS 75.88 90.59 virus *Virus detected in this study sheep, pigs, and poultry [25]. Livestock animals in the cities suggests, the intensive model restricts livestock movement, are reared in intensive or semi-intensive systems, while rural whereas semi-intensive and extensive models, which are less areas employ extensive farming models [24]. As the name capital-intensive, allow animals varying degrees of freedom 1 3 M. Amoa-Bosompem et al. to move and interact with the community and ecosystem such as monocytes and macrophages, from tick feeding sites [24]. This interaction between humans and animals in a cli- facilitates the transmission of viruses between infected and mate conducive to tick survival and breeding increases the uninfected ticks. The potential for horizontal transmission likelihood of transmission of tick-borne infections of public through co-feeding raises concerns about the evolutionary health and veterinary importance. This study was therefore and public health implications of coinfection of an MdPV designed to identify and characterize tick-borne or tick- and related phleboviruses (with an M segment). It has been associated viruses harboured by ticks infesting domesticated postulated that phlebovirus L and N proteins recognize the animals in different regions of Ghana. A total of 354 nymph untranslated region of the M segment of a related virus as or adult ticks collected from four different geographic loca- a functional promoter for transcription and replication, pro- tions in Ghana were subjected to virus isolation, RT-PCR, ducing viable reassortant progeny [32]. and NGS analysis. DUGV is an orthonairovirus of public health impor- Ticks of at least three genera were collected across Ghana; tance that has been reported to cause thrombocytopenia however, members of only one genus (Rhipicephalus) were and febrile illnesses in humans [33]. Although there are no found on dogs, consistent with a previous report by Kob- reports of pathology in livestock animals or pets, the pub- ayashi et al. [13]. Furthermore, tick samples were collected lic health implications of the widespread distribution of from dogs in Accra and Jirapa, but only those from Accra DUGV in Africa, and possibly Ghana, must be evaluated were positive for an RNA virus, ODWV. The detection of [33, 34]. DUGV was originally isolated from A. variegatum, ODWV-positive ticks in Accra is consistent with the findings the main vector. Other tick species, Hyalomma truncatum, of Kobayashi et al. [13]. However, in this study, ODWV was Boophilus decoloratus, and Rhipicephalus appendiculatus, detected in pools of non-engorged ticks, whereas, previously, are also regarded as major vectors of DUGV [35]. In this it was detected only in pools containing engorged ticks [13]. study, DUGV was detected in Rhipicephalus ticks collected These results confirm that ODWV is a tick-associated virus in Hohoe. Kobayashi et al. [13] isolated DUGV from A. that is widely distributed in Rhipicephalus ticks infesting variegatum collected in Accra, demonstrating that DUGV is dogs in Accra; however, whether ODWV is infectious to distributed throughout southern Ghana and may be transmit- dogs and/or humans is yet to be determined. ted by multiple vectors. Among the cattle-infesting ticks, three pools, each repre- The results of this study suggest that the close interaction senting a genus, were positive for an RNA virus. Two pools between people and livestock/pets in Ghana may increase the were positive for BTV, and the third was positive for DUGV. transmission potential of TBVs in the country. Thus, screen- BTV was detected in Amblyomma and Hyalomma ticks from ing for TBVs (such as DUGV and CCHFV) when diagnos- Jirapa, while DUGV was detected in Rhipicephalus ticks ing febrile illnesses is important, particularly in malaria- from Hohoe. negative patients. A public health adjustment of this nature BTV, like ODWV, is a phlebovirus belonging to the may help reduce the number of (and issues surrounding) group defined as the M-segment-deficient phleboviruses undiagnosed febrile cases in Ghana [36]. Furthermore, the (MdPVs) [15]. To our knowledge, this is the first report on results of this study highlight the need to comprehensively the evolutionary history of any BTV strain that confirms examine the actual disease burden of TBVs in Ghana, which its classification as a member of the MdPV group. These will facilitate the development of health policies to include MdPVs can further be classified into three distinct groups TBV-screening in the diagnoses of febrile illnesses. based primarily on their phylogeny [15]. The host ticks of the MdPVs also appear to be distinct, with only group II, Supplementary Information The online version contains supplemen- which includes BTV and ODWV, detected in ticks of mul- tary material available at https://d oi.o rg/1 0.1 007/s 00705-0 21-0 5296-4. tiple genera: Dermacentor, Hyalomma, and Rhipicephalus Acknowledgements The authors are grateful to all the staff of Nogu- [15, 26–30]. Owing to the apparent relationship between chi Memorial Institute for Medical Research for supporting this study. MdPVs and their host ticks, they are postulated to have coevolved with their hosts through vertical transmission. Author contributions Conceptualization: MA-B, DK, HI. Methodol- Nevertheless, the possibility of horizontal transmission has ogy: MA-B, DK, ANF, SK, AA, EA, DP, MO, HE. Formal analysis not been ruled out. In this study, BTV was detected in Hya- and investigation: MA-B, DK, HI. Writing—original draft preparation: MA-B. Writing—review and editing: MA-B, DK, HI. Funding acqui- lomma and Amblyomma ticks, representing the first report sition: HI. Resources: HI. Supervision: JHKB, SD, NO, KS, SI, HI. of a group II MdPV in ticks of the genus Amblyomma. Fur- thermore, the detection of BTV in both Amblyomma and Funding This study was supported by the Japan Initiative for Hyalomma ticks infesting cattle in Jirapa may be an indica- Global Research Network on Infectious Diseases (Grant numbers tion of possible horizontal transmission between ticks of JP19fm0108010 and JP20wm0225007) and the Research Program on Emerging and Re-emerging Infectious Diseases (Grant num- different genera infesting the same host, possibly through co- ber JP20fk0108067) from the Japan Agency for Medical Research feeding. Labuda et al. [31] reported that migration of cells, and Development. This study was also supported in part by JSPS 1 3 Tick-borne and tick-associated viruses in Ghana KAKENHI (Grant numbers JP16J09470, JP18K19220, and at an abattoir in Ghana. BMC Infect Dis 16:324. https:// doi. org/ JP18H02856). 10. 1186/s 12879- 016- 1660-6 1 3. Kobayashi D, Ohashi M, Osei JHN, Agbosu E, Opoku M, Availability of data and materials The datasets used and/or analysed Agbekudzi A, Joannides J, Fujita R, Sasaki T, Bonney JHK, during the current study are available from the corresponding author Dadzie S, Isawa H, Sawabe K, Ohta N (2017) Detection of a novel on reasonable request. putative phlebovirus and first isolation of Dugbe virus from ticks in Accra, Ghana. Ticks Tick Borne Dis 8(4):640–645. https://d oi. org/1 0. 1016/j.t tbdis. 2017. 04. 010 Declarations 14. Amoa-Bosompem M, Kobayashi D, Murota K, Faizah AN, Itokawa K, Fujita R, Osei JHN, Agbosu E, Pratt D, Kimura S, Conflict of interest The authors declare that they have no competing Kwofie KD, Ohashi M, Bonney JHK, Dadzie S, Sasaki T, Ohta N, interests. Isawa H, Sawabe K, Iwanaga S (2020) Entomological assessment of the status and risk of mosquito-borne arboviral transmission in Ghana. Viruses 12(2):147. https:// doi. org/1 0. 3390/ v1202 0147 15. 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