c871df80febb5773d619ac524cda641c vetsci-12-01101-v2.pdf 3f4daeaa8970092292a690d5e10618a2d8a2390c vetsci-12-01101-v2.pdf bf7778bd9bbdcea0d257f3c56c444aaf107ba854a1b64d5aed141c244999d2cc vetsci-12-01101-v2.pdf Title: Exposure of Wild Ruminants to Toxoplasma gondii in Alpine Ecosystems, NE Spain Subject: Toxoplasma gondii is a protozoan parasite with a complex life cycle that involves warm-blooded animals as intermediate hosts and felids as definitive hosts. Its epidemiology in alpine ecosystems remains poorly understood. This study presents the first long-term investigation of T. gondii exposure in wild ruminants in the Pyrenees (NE Spain), where definitive hosts are scarce. Pyrenean chamois (Rupicapra pyrenaica; n = 1045) and mouflon (Ovis aries musimon; n = 115) sera collected between 2001 and 2024 were tested (Modified Agglutination Test, ELISA-IDvet, ELISA-IDEXX) for the presence of T. gondii antibodies. Sera from 53 chamois and 27 mouflon foetuses and hearts and brains from 38 chamois and 35 mouflon foetuses were analysed for the presence of antibodies and parasite’s DNA, respectively. Moreover, heart and brain (n = 3) and faeces (n = 91) from Pyrenean wildcats (Felis silvestris) were analysed for parasite’s DNA. Seroprevalence was overall low (chamois: 5.24%; mouflon: 1.74%). In multivariate analyses performed in chamois, seroprevalence variation was mainly associated with geographic origin and the diagnostic method used, with little influence of the individual traits of sex or age. No antibodies or T. gondii DNA were detected in chamois or mouflon foetuses. However, chamois did not show a clear age-related increase in exposure to T. gondii, as would be expected if indirect horizontal transmission were the predominant route. In contrast, T. gondii DNA was detected in brain and faecal samples from wildcats, confirming their role as definitive hosts in this ecosystem. Overall, our results reinforce the hypothesis that alpine ecosystems are environments with low T. gondii oocyst contamination and that the parasite most probably relies on sylvatic cycles. These results suggest that wild ruminants, such as chamois and mouflon, could serve as effective sentinels of changes in the extent of this parasite under ongoing environmental and ecological changes. Keywords: Felis silvestris; mouflon; Ovis aries musimon; parasite; pyrenees; pyrenean chamois; Rupicapra pyrenaica; Toxoplasma gondii; wildcat; wildlife population health Author: Alejandra Escudero, Maria Puig Ribas, Sonia Almería, Hojjat Gholipour, Lola Pailler-García, Natalia Sastre, Jordi Ruiz-Olmo, Santiago Palazón, Ferran Sayol, Johan Espunyes, Xavier Fernández Aguilar and Oscar Cabezón Creator: LaTeX with hyperref Producer: pdfTeX-1.40.25; modified using OpenPDF 1.4.2 CreationDate: Wed Nov 19 04:47:21 2025 CET ModDate: Wed Nov 19 05:47:26 2025 CET Custom Metadata: yes Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 15 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1068154 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- VPGCOS+VnURWPalladioL Type 1 Custom yes yes yes 86 0 SKOGGW+URWPalladioL-Bold Type 1 Custom yes yes yes 87 0 BOPBFP+URWPalladioL-Roma Type 1 Custom yes yes yes 88 0 SVNGBP+URWPalladioL-BoldItal Type 1 Custom yes yes yes 89 0 HBITDL+URWPalladioL-Ital Type 1 Custom yes yes yes 90 0 AWDAAW+VnURWPalladioL-Italic Type 1 Custom yes yes yes 166 0 YZIJXT+CMSY10 Type 1 Builtin yes yes yes 167 0 HGJNID+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 140 0 HGJNIB+PalatinoLinotype-Bold CID TrueType Identity-H yes yes yes 141 0 HGJNHA+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 142 0 RFECCF+EURM10 Type 1 Builtin yes yes yes 210 0 MPUDFL+PazoMath Type 1 Builtin yes yes yes 234 0 HGJNID+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 226 0 HGJONC+Arial-ItalicMT TrueType WinAnsi yes yes no 227 0 HGJNIB+PalatinoLinotype-Bold CID TrueType Identity-H yes yes yes 228 0 HGJNHA+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 229 0 HGJONA+ArialMT TrueType WinAnsi yes yes no 230 0 HGJOND+SymbolMT CID TrueType Identity-H yes yes yes 231 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2025-12-18 03:14:05 CET RepresentationInformation: vetsci-12-01101-v2.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2025-12-17 10:57:46 CET Size: 1068154 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 496 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: ViewerPreferences: HideToolbar: false HideMenubar: false HideWindowUI: false FitWindow: true CenterWindow: false DisplayDocTitle: false NonFullScreenPageMode: UseNone Direction: L2R ViewArea: CropBox ViewClip: CropBox PrintArea: CropBox PageClip: CropBox PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Item: Title: Materials and Methods Children: Item: Title: Study Area Item: Title: Animal Samples Item: Title: Serological Test Item: Title: Molecular Analysis Item: Title: Statistical Analyses Item: Title: Results Item: Title: Discussion Item: Title: References Info: Title: Exposure of Wild Ruminants to Toxoplasma gondii in Alpine Ecosystems, NE Spain Author: Alejandra Escudero, Maria Puig Ribas, Sonia Almería, Hojjat Gholipour, Lola Pailler-García, Natalia Sastre, Jordi Ruiz-Olmo, Santiago Palazón, Ferran Sayol, Johan Espunyes, Xavier Fernández Aguilar and Oscar Cabezón Subject: Toxoplasma gondii is a protozoan parasite with a complex life cycle that involves warm-blooded animals as intermediate hosts and felids as definitive hosts. Its epidemiology in alpine ecosystems remains poorly understood. This study presents the first long-term investigation of T. gondii exposure in wild ruminants in the Pyrenees (NE Spain), where definitive hosts are scarce. Pyrenean chamois (Rupicapra pyrenaica; n = 1045) and mouflon (Ovis aries musimon; n = 115) sera collected between 2001 and 2024 were tested (Modified Agglutination Test, ELISA-IDvet, ELISA-IDEXX) for the presence of T. gondii antibodies. Sera from 53 chamois and 27 mouflon foetuses and hearts and brains from 38 chamois and 35 mouflon foetuses were analysed for the presence of antibodies and parasite’s DNA, respectively. Moreover, heart and brain (n = 3) and faeces (n = 91) from Pyrenean wildcats (Felis silvestris) were analysed for parasite’s DNA. Seroprevalence was overall low (chamois: 5.24%; mouflon: 1.74%). In multivariate analyses performed in chamois, seroprevalence variation was mainly associated with geographic origin and the diagnostic method used, with little influence of the individual traits of sex or age. No antibodies or T. gondii DNA were detected in chamois or mouflon foetuses. However, chamois did not show a clear age-related increase in exposure to T. gondii, as would be expected if indirect horizontal transmission were the predominant route. In contrast, T. gondii DNA was detected in brain and faecal samples from wildcats, confirming their role as definitive hosts in this ecosystem. Overall, our results reinforce the hypothesis that alpine ecosystems are environments with low T. gondii oocyst contamination and that the parasite most probably relies on sylvatic cycles. These results suggest that wild ruminants, such as chamois and mouflon, could serve as effective sentinels of changes in the extent of this parasite under ongoing environmental and ecological changes. 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