Multiple Lineages of Hantaviruses Harbored by the Iberian Mole (Talpa occidentalis) in Spain

Abstract

The recent detection of both Nova virus (NVAV) and Bruges virus (BRGV) in European moles (Talpa europaea) in Belgium and Germany prompted a search for related hantaviruses in the Iberian mole (Talpa occidentalis). RNAlater^®-preserved lung tissue from 106 Iberian moles, collected during January 2011 to June 2014 in Asturias, Spain, were analyzed for hantavirus RNA by nested/hemi-nested RT-PCR. Pairwise alignment and comparison of partial L-segment sequences, detected in 11 Iberian moles from four parishes, indicated the circulation of genetically distinct hantaviruses. Phylogenetic analyses, using maximum-likelihood and Bayesian methods, demonstrated three distinct hantaviruses in Iberian moles: NVAV, BRGV, and a new hantavirus, designated Asturias virus (ASTV). Of the cDNA from seven infected moles processed for next generation sequencing using Illumina HiSeq1500, one produced viable contigs, spanning the S, M and L segments of ASTV. The original view that each hantavirus species is harbored by a single small-mammal host species is now known to be invalid. Host-switching or cross-species transmission events, as well as reassortment, have shaped the complex evolutionary history and phylogeography of hantaviruses such that some hantavirus species are hosted by multiple reservoir species, and conversely, some host species harbor more than one hantavirus species.

Keywords: Hantaviridae; Spain; evolution; hantavirus; talpid.

Figure 1

(A) Adult Iberian mole (Talpa occidentalis) and European mole (Talpa europaea), showing the comparatively smaller size of the former species. (B) Map showing sites in Asturias in northwestern Spain, where hantavirus-infected Iberian moles and crowned shrews were collected. The number of hantavirus RNA-positive animals and the number tested are shown for each site.

Figure 2

Phylogenetic trees, based on sequences of the S-, M-, and L-genomic segments, respectively, generated by the Bayesian Markov chain Monte Carlo estimation method, under the GTR + I + Γ model of evolution. Hantavirus 3879, 3884, 3890, 3914, 3930 and 3947 (shown in green) from Iberian moles clustered with Bruges virus (BRGV), while 3873, 3931, 3943 and 3945 (shown in blue) clustered with Nova virus (NVAV). Accordingly, these have been named BRGV and NVAV, respectively. By contrast, 3877 (shown in red) was distinct from all other hantaviruses described to date and was named Asturias virus (ASTV). Sequence lengths of the S, M and L segments for ASTV and the L segment of BRGV and NVAV from Iberian moles are shown in Table 2. Also shown are BRGV Vieux-Genappe/TE/2013/1 (S: KX551960; M: KX551961; L: KX551962) and BRGV Wandlitz/TE/2013/1 (S: MF683844; M: MF683845; L: MF683846) from European moles in Belgium and Germany, respectively; ACDV Academ-Ta450 (S: MK340905; M: OL871119; L: MH784614) from a Siberian mole in Russia; and LDRV MNHN-ZM-2017-2257 (S: ON944104) from an Aquitanian mole in France. Other mole-borne hantaviruses include Asama virus (ASAV) N10 (S: EU929072; M: EU929075; L: EU929078) from Urotrichus talpoides; Oxbow virus (OXBV) Ng1453 (S: FJ5339166; M: FJ539167; L: FJ593497) from Neurotrichus gibbsii; Rockport virus (RKPV) MSB57412 (S: HM015223; M: HM015222; L: HM015221) from Scalopus aquaticus; Dahonggou Creek virus (DHCV) DGR36708 (L: HQ616595) from Scaptonyx fusicaudus; and Nova virus (NVAV) Te34 (S: KR072621; M: KR072622; L: KR072623) and MSB95703 (S: FJ539168; M: HQ840957; L: FJ593498) from Talpa europaea. Bat-borne hantaviruses include Láibīn virus (LAIV) BT20 (S: KM102247; M: KM102248; L: KM102249); Xuân Sơn virus (XSV) F42682 (S: KF704709; M: KJ000538; L: KF704714); Magboi virus (MGBV) 1209 (L: JN037851); Mouyassué virus (MOYV) KB576 (L: JQ287716); Huángpí virus (HUPV) Pa-1 (S: JX473273; L: JX465369); Lóngquán virus (LQUV) Ra10 (S: JX465413; M: JX465396; L: JX465379). Shrew-borne hantaviruses include Cao Bằng virus (CBNV) CBN-3 (S: EF543524; M: EF543526; L: EF543525) from Anourosorex squamipes; Jemez Springs virus (JMSV) MSB144475 (S: FJ593499; M: FJ593500; L: FJ593501) from Sorex monticolus; Seewis virus (SWSV) mp70 (S: EF636024; M: EF636025; L: EF636026) from Sorex araneus; Kenkeme virus (KKMV) MSB148794 (S: GQ306148; M: GQ306149; L: GQ306150) from Sorex roboratus; and Qian Hu Shan virus (QHSV) YN05-284 (S: GU566023; M: GU566022; L: GU566021) from Sorex cylindricauda; as well as Thottapalayam virus (TPMV) VRC66412 (S: AY526097; M: NC_010708; L: EU001330) from Suncus murinus; Imjin virus (MJNV) Cl05-11 (S: EF641804; M: EF641798; L: EF641806) from Crocidura lasiura; Azagny virus (AZGV) KBM15 (S: JF276226; M: JF276227; L: JF276228) from Crocidura obscurior; Tanganya virus (TGNV) Tan826 (S: EF050455; L: EF050454) from Crocidura theresea; Bowé virus (BOWV) VN1512 (S: KC631782; M: KC631783; L: KC631784) from Crocidura douceti; Jeju virus (JJUV) SH42 (S: HQ663933; M: HQ663934; L: HQ663935) from Crocidura shantungensis; Uluguru virus (ULUV) FMNH158302 (S: JX193695; M: JX193696; L: JX193697) from Myosorex geata; and Kilimanjaro virus (KMJV) FMNH174124 (S: JX193698; M: JX193699; L: JX193700) from Myosorex zinki. Rodent-borne orthohantaviruses include Sin Nombre virus (SNV) NMH10 (S: NC_005216; M: NC_005215; L: NC_005217); Andes virus (ANDV) Chile9717869 (S: AF291702; M: AF291703; L: AF291704); Prospect Hill virus (PHV) PH-1 (S: Z49098; M: X55129; L: EF646763); Tula virus (TULV) M5302v (S: NC_005227; M: NC_005228; L: NC_005226); Puumala virus (PUUV) Sotkamo (S: NC_005224; M: NC_005223; L: NC_005225); Dobrava virus (DOBV) Greece (S: NC_005233; M: NC_005234; L: NC_005235); Hantaan virus (HTNV) 76-118 (S: NC_005218; M: NC_005219; L: NC_005222); Soochong virus (SOOV) SOO-1 (S: AY675349; M: AY675353; L: DQ056292); and Seoul virus (SEOV) 80-39 (S: NC_005236; M: NC_005237; L: NC_005238). The numbers at selected nodes are Bayesian posterior probabilities (>0.70) based on 150,000 trees. Two replicate Markov chain Monte Carlo runs, consisting of six chains of 10 million generations, each sampled every 100 generations with a burn-in of 25,000 (25%). The scale bars indicate nucleotide substitutions per site.

Figure 3

Phylogenetic tree based on full-length sequences of the cytochrome b mtDNA using the maximum-likelihood method. The percentage of trees in which the associated taxa clustered together is shown at the respective node. Initial trees for the heuristic search were obtained automatically by applying neighbor-joining and BioNJ algorithms to a matrix of pairwise distances estimated using the maximum composite likelihood (MCL) approach and then selecting the topology with superior log likelihood value. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. Talpa occidentalis from this study are shown in red lettering.