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. 2026 Jul 7;125(1):117. doi: 10.1007/s00436-026-08725-y

Validating a point-of-care test for Toxoplasma gondii infection in southern sea otters (Enhydra lutris nereis)

Ellis Song 1, Devinn Sinnott 1, Cara L Field 2, Michael Murray 3, Pádraig Duignan 2, Melissa Miller 4, Karen Shapiro 1,✉
PMCID: PMC13627161  PMID: 42412205

Abstract

Toxoplasma gondii is a zoonotic protozoan parasite that infects a high proportion of threatened southern sea otters (Enhydra lutris nereis) and is an important cause of mortality in this host species. Recently, a point-of-care rapid antibody test (POCT) for T. gondii infection was developed for detection of IgG and IgM antibodies in human sera. We aimed to validate the POCT using southern sea otter sera against a gold standard state of infection, based on histopathology, immunohistochemistry, parasite isolation, and PCR. In this study, we hypothesized that the POCT rapid screening tool would offer both high sensitivity and specificity (> 90%) for screening T. gondii infection in archived serum samples from southern sea otters with known T. gondii infection status. We applied the POCT assay to sera from 109 sea otters (49 negative, 60 positive), and this assay demonstrated an overall sensitivity of 93.3% and specificity of 93.9%. These results indicate that the POCT may be a useful screening tool for T. gondii exposure in sea otters. Utilization of this test in wildlife rehabilitation centers would allow for rapid, on-site, and cost-efficient screening of T. gondii exposure that could aid in the diagnosis and clinical management of sea otters with suspected toxoplasmosis. Future efforts could target POCT validation in species such as Hawaiian monk seals and Hector’s dolphins, for which T. gondii is listed as a threat to species survival.

Supplementary Information

The online version contains supplementary material available at https://doi.org/10.1007/s00436-026-08725-y.

Keywords: Antibody, Detection, Immunochromatographic assay, Point-of-care test, Protozoa, Serology

Introduction

Toxoplasma gondii is a zoonotic, protozoan parasite that can infect all warm-blooded animals, including birds, humans, and other mammals. In the environment, this parasite is transmitted via environmentally resistant oocysts shed in the feces of definitive hosts (wild and domestic felids) which can contaminate food and water ingested by humans and other susceptible intermediate hosts (Dubey 2016; Shapiro et al. 2019a). Transmission of T. gondii in threatened southern sea otters (Enhydra lutris nereis) has been associated with the consumption of benthic invertebrates, such as snails (Johnson et al. 2009), which can concentrate oocysts in their tissues from contaminated seawater (Krusor et al. 2015). Once infective oocysts are ingested by an intermediate host such as sea otters, rapidly multiplying T. gondii tachyzoites spread systemically in tissues. Clinical toxoplasmosis in sea otters is typically characterized by meningoencephalitis with severe neurologic signs including seizures, tremors, ataxia, paralysis, and death (Miller et al. 2018). In response to host immune defenses, T. gondii tachyzoites transform into slowly replicating bradyzoites that form tissue cysts in the central nervous system, heart, and skeletal muscles and can cause recurrent, chronic clinical disease (Miller et al. 2018).

With an estimated seroprevalence of 57%, infection with T. gondii is widespread in southern sea otters (SSO) (Miller et al. 2020) and mortalities can occur following both acute and chronic infections (Kreuder et al. 2003; Miller et al. 2023). The highly virulent COUG strain of T. gondii was implicated in SSO mortalities in 2020–2024, causing death in otherwise healthy animals due to severe protozoa-associated steatitis and systemic toxoplasmosis or more classic meningoencephalitis (Miller et al. 2023; Sinnott et al. 2025). This T. gondii strain and its associated steatitis syndrome was not observed in this host prior to 2020, and the cause of its emergence in this species remains unknown. The discovery of this highly virulent parasite strain and the importance of toxoplasmosis as a cause of death for SSOs underscores the need for a rapid, cost-efficient test that can be performed on-site at rehabilitation centers to expedite diagnosis and treatment. Currently, the only validated serological test for T. gondii exposure in SSOs is the indirect fluorescent antibody test (IFAT) (Miller et al. 2002). The need to ship sera to a specialty lab where the IFAT is performed can create a delay of several days or longer between the time blood is drawn and receiving results, potentially delaying clinicians from initiating timely appropriate antiprotozoal treatment.

In Europe, an immunochromatographic (ICT) antibody point-of-care test designed to detect both IgG and IgM anti-Toxoplasma antibodies has been recently approved for rapid exposure detection in humans (Toxoplasma ICT IgG-IgM®, LDBIO Diagnostics, Lyon, France, hereafter referred to as point-of-care-Toxoplasma, or POCT) (Khan and Noordin 2020). Previous reports of application of the POCT in veterinary species include studies conducted on black bears, ring-billed gulls, and black vultures, where the utility of this assay for screening wildlife for T. gondii exposure was compared with another serologic test, the modified agglutination test (MAT). However, animals with molecularly and/or histopathologically confirmed positive or negative T. gondii infection status were not available to definitively validate the POCT in these hosts (Strules et al. 2023; Rosypal von Dohlen et al. 2024). This lack of ancillary test confirmation of infection status can make it difficult to evaluate and interpret results of real-life applications of the POCT, highlighting the need for validation against a gold standard state of infection.

While the POCT is not able to discriminate between IgM and IgG antibodies, it can yield positive results for acute infections (in which IgM antibodies would be present before IgG antibodies are produced). For these acute cases, conventional testing methods that typically rely only on measuring IgG antibody levels (IFAT and Enzyme-Linked Immunosorbent Assay, or ELISA) may yield false negative results (Khan and Noordin 2020). The POCT is also cost and time efficient compared to other serological tests, which can be limiting factors to optimal diagnostics and care in a wildlife rehabilitation setting. Considering the potential benefits of the POCT, we aimed to evaluate the performance of this test using serum from SSOs that are commonly infected with T. gondii and may require veterinary care in rehabilitation centers. We hypothesized that the POCT will offer both high sensitivity (> 90%) and specificity (> 90%) for diagnosing T. gondii infection when used to evaluate serum from SSOs with confirmed T. gondii infection status.

Methods

In collaboration with The Marine Mammal Center (TMMC), Monterey Bay Aquarium (MBA), and the California Department of Fish and Wildlife (CDFW), we selected T. gondii-infected and uninfected SSOs that stranded along the California coast between 1999 and 2025 for which cryoarchived premortem or perimortem sera was available. Individuals were selected based on records of previous diagnostic testing to identify confirmed T. gondii infected and uninfected SSOs. Positive cases were defined as animals with histopathological identification of protozoal organisms in tissues and confirmation of parasite identity as T. gondii by immunohistochemistry, isolation of T. gondii using cell culture, and/or polymerase chain reaction (PCR) followed by sequencing (Shapiro et al. 2019b; Miller et al. 2020) in at least one tissue (brain +/- heart or skeletal muscle). Negative animals (uninfected controls) were defined as SSOs with no histopathologic evidence of T. gondii infection and no amplification of T. gondii DNA in three independent biological replicates of at least one tissue (brain +/- heart or skeletal muscle).

Following manufacturer’s instructions, we performed the POCT on SSO sera from the selected cases and controls. Briefly, 15 ul of serum was applied to a well in the lateral flow cassette followed by 4 drops of eluent, per manufacturer instructions. The results were recorded after 30 min as positive (black line at the ‘T’ (test) mark and blue line at the ‘C’ (control) mark) or negative (no line at the ‘T’ mark, blue line at the ‘C’ mark (Fig. 1). Samples for which a control line was not visible were repeated. To evaluate the performance of the test, we calculated the sensitivity, specificity, positive predictive value, and negative predictive value (Trevethan 2017).

Fig. 1.

Fig. 1

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Examples of a negative (left) and positive (right) test results using the ICT IgG-IgM® point-of-care-Toxoplasma (POCT) test. The blue line at the “C” mark indicates the control and the presence or absence of the black line at the “T” mark indicates a positive or negative result for the detection of anti-T. gondii antibodies

In the test manual, the manufacturer advises against the use of hemolyzed sera as hemolysis can make interpretation of test results challenging; however, the manual also states that there is no evidence of hemolysis influencing the performance of the test itself. Obtaining serum from wildlife species, including SSOs, can often be difficult and can lead to hemolyzed samples. To further evaluate the effect of serum hemolysis on test accuracy, we recorded the degree of hemolysis for both the serum itself and how the assay cassette appeared once each sample was applied to the test strip (Fig. 2). The degrees of hemolysis for the tubes and test strips were scored separately on a scale from 0 (no hemolysis) to + 3 (severely hemolyzed). The test manufacturer also recommends using either fresh serum or samples that were frozen for two years or less for this test. However, cryoarchived samples exceeding this recommended two-year limit were needed in order to achieve an adequate sample size to validate the POCT for use in SSOs. The minimum sample size required to calculate our predicted sensitivity (90%) and specificity (90%) with 95% confidence intervals and 80% power was N = 70 (35 positive and 35 negative SSOs) (Hajian-Tilaki 2014).

Fig. 2.

Fig. 2

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Examples of varying levels of hemolysis (scored as 0, +1, +2, +3) of the serum (top) and the assay cassette window (bottom). Despite the reddish background for hemolyzed samples on the test cassette, a positive line could still be discerned for SSO infected with Toxoplasma gondii

Performance of the POCT was assessed by calculating sensitivity, specificity, and positive and negative predictive values:

graphic file with name d33e469.gif
graphic file with name d33e472.gif
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Where “a” is true positives, “b” is false positives, “c” is false negatives, and “d” is true negatives.

To evaluate the effects of hemolysis and sample age of serum on test accuracy, we applied the Fisher’s exact test using Stata (StataCorp, 2025). Significance was determined at P ≤ 0.05.

Results and discussion

Based on our selection criteria and the previously performed gold-standard diagnostic results available, we identified 60 SSO with histopathologic, culture, and/or PCR-confirmed T. gondii infection and 49 negative controls for a total sample size of 109. A positive POCT result was obtained for 56 of the 60 SSO with confirmed T. gondii infection, yielding a test sensitivity of 93.3%. Of the 49 negative controls, a positive POCT result was recorded for three animals (false positives) yielding a specificity of 93.9%. The positive and negative predictive values were calculated as 94.9 and 92.0%, respectively (Table 1).

Table 1.

Sensitivity and specificity of Toxoplasma gondii point-of-care test (POCT) compared with a gold standard of confirmation T. gondii infection or non-infection based on histopathology, immunohistochemistry, culture, and/or PCR in SSOs (Enhydra lutris nereis) from California.

POCT test result T. gondii positive cases T. gondii negative cases Totals
Positive 56 3 59 PPV = 94.9%
Negative 4 46 50 NPV = 92.0%
Totals 60 49 109
Se = 93.3% Sp = 93.9%
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Se sensitivity, Sp specificity, PPV positive predictive value, NPV negative predictive value

We evaluated the association between test performance and the level of hemolysis, both for the serum directly and its appearance on the test strip (Fig. 2), using a Fisher’s exact test. The level of hemolysis was analyzed as both a categorical variable ranging from no, mild, moderate to marked hemolysis (denoted as 0, + 1, +2, and + 3, respectively) and as a binomial variable (unhemolyzed vs. hemolyzed). No significant association was observed between test performance (correct classification of infection status) and level of serum hemolysis in categorical analyses (p = 0.117). Overall, no negative effects of hemolysis were observed on test accuracy, when analyzing both the hemolysis score of the sera directly or discoloration of the test cassette, using either binary or categorical classification of hemolysis.

We also evaluated whether serum age affected test performance, defining accuracy as the ability of the assay to correctly determine infection status compared with the gold standard. We categorized the age of serum as a binomial variable (older than two years vs. younger than two years) based on the year of collection and the year the POCT was performed. No significant association was observed between serum age and test accuracy (Fisher’s exact test, p = 0.211).

Our study validates the POCT for use in SSOs using molecular and histopathologic confirmation as the gold standard state of infection. Previous studies in other veterinary species have evaluated the POCT in comparison to another serological test (Strules et al. 2023; Rosypal von Dohlen et al. 2024). Use of a molecular and histopathology-based gold standard more closely reflects the true state of infection and has enabled us to more precisely evaluate the sensitivity and specificity of the POCT in SSOs and assess its potential utility as an antemortem and postmortem diagnostic test for T. gondii infection in this species. Our results revealed that the POCT has a high sensitivity (93.3%) and specificity (93.9%) for detecting anti-T. gondii antibodies in SSO serum, making it an effective tool for rehabilitators to rapidly screen for T. gondii infection. This is a more conservative estimate than its performance in humans (100% sensitivity and 97.7% specificity per manufacturer’s records).

By evaluating the POCT against molecular and histopathological evidence of infection rather than an alternative serological test, we intended to evaluate POCT performance relative to evidence of infection rather than relative agreement with an existing serologic platform. This method limits bias estimates of diagnostic sensitivity and specificity when measuring biologic processes using serologic assays (Gardner et al. 2000) and is consistent with recommendations outlined by the World Organization for Animal Health (WOAH, 2023; formerly OIE). However, when interpreting sensitivity and specificity in light of this reference standard, we must also acknowledge that PCR and histopathology have their limitations: both PCR and histopathology assess small sections of tissue that can potentially miss parasites that are randomly or sparsely distributed within tissue, even if multiple replicates are evaluated. This may result in apparent POCT false positive cases, as some chronically infected animals with low parasite burdens may have been misclassified as uninfected by PCR and histopathology.

To date, the only validated serological test for assessing T. gondii infection in SSO is the IFAT (Miller et al. 2002). The sensitivity and specificity of the IFAT for detecting anti-T. gondii antibodies in SSO serum were previously evaluated to be 96.4% and 67.3%, respectively (Miller et al. 2002). The case definitions, gold standard criteria, and samples used for validation of the IFAT assay differed in criteria used in this study; therefore, a true comparison of test performance between the IFAT and POCT cannot be drawn. While the POCT provides a time- and cost-efficient screening test for assessing T. gondii exposure, the IFAT can provide additional clinically valuable information such as a semi-quantitative titer for T. gondii antibodies, and can be run in parallel to detect Sarcocystis neurona antibodies, another closely related protozoan parasite that commonly infects SSOs and can cause a similar clinical presentation. The POCT may serve as a rapid benchtop screening tool for veterinary staff in marine mammal rehabilitation facilities to help triage animals, expedite screening for T. gondii infection, and guide treatment. Because the effect of serum hemolysis was found to be negligible, the POCT might also be a helpful benchtop screening tool for necropsied SSOs as well.

Our study was limited by the serum samples available from SSOs with corresponding histopathological data and tissues for molecular diagnostics. The samples used in this study were cryoarchived between < 1 and 25 years since the initial date of collection, and many samples had been previously frozen and thawed before use. It is possible that these factors may have caused some degree of antibody degradation in some serum samples which could have influenced the results of the POCT. Our analysis revealed that there was no significant difference in test accuracy when using serum greater than or less than the two-year cut-off that was recommended in the manufacturer’s instructions. However, continued evaluation of the POCT using freshly obtained serum may yield higher sensitivity and specificity estimates. The samples used in this study also had varying degrees of hemolysis. While non-hemolyzed sera is preferred by the manufacturer for use in humans, providing veterinary care for wildlife in rehabilitation centers presents a unique set of challenges that may unavoidably result in the collection of hemolyzed serum. While our results suggest that hemolysis does not adversely affect the test accuracy, it is important for veterinary staff to exercise caution when using this test with hemolyzed sera.

Several considerations should be taken into account when interpreting our study’s findings. First, given the high prevalence of infection of both T. gondii and S. neurona in SSOs, potential cross-reactivity between these protozoal parasites is an important consideration. Due to logistical and cost constraints, we were unable to identify and test sera from a substantial number of SSOs with confirmed S. neurona infections and without evidence of T. gondii infection. In a pilot sample of eight SSOs with confirmed S. neurona infection and without molecular evidence of concurrent T. gondii infection, the POCT results were negative for 7 of these 8 animals (Table S2). Interpretation of POCT results in the context of clinical toxoplasmosis versus chronic subclinical infection is also relevant for rehabilitation and future surveillance applications. The methods used to classify true infection status (parasite isolation, histology, and PCR) can identify both clinical toxoplasmosis and latent (subclinical) infections (Miller et al. 2020; Shapiro et al. 2019b) We were able to identify the cause of death (COD) for 45 T. gondii-infected SSOs in our study. Of these 45 animals, T. gondii was determined to be a primary or contributing COD for 19 SSOs, while 26 had incidental infections unrelated to the identified COD (Table S1). This suggests that the POCT can detect anti-T. gondii antibodies in both clinical and incidental cases of T. gondii infection. Finally, because T. gondii is fairly prevalent in SSOs, the predictive values reported here may differ in populations with different infection prevalences.

Considering the many differential diagnoses for SSOs that strand with neurological clinical signs, including protozoal infections (T. gondii and S. neurona) (Miller et al. 2018), domoic acid toxicosis (Miller et al. 2021), morbilliviruses (Thomas et al. 2020), and trauma, establishing a rapid and accurate diagnosis for these similarly presenting conditions is often challenging. However, wildlife veterinarians and rehabilitation professionals must respond quickly to implement appropriate treatment that can reduce morbidity and mortality. Our study demonstrates that the POCT is a highly sensitive and specific test for screening SSO sera for T. gondii infection and can provide rapid, onsite results to rule out T. gondii infection or determine the need for additional diagnostics when managing SSOs in rehabilitation facilities. Future application of the POCT in other T. gondii susceptible wildlife would provide a valuable clinical screening tool for these vulnerable marine mammal species, such as Hawaiian monk seals (Monachus schauinslandi) and Hector’s dolphins (Cephalorhynchus hectori) for which T. gondii is listed as a threat to species survival (Roe et al. 2013; Robinson et al. 2023).

Supplementary information

Below is the link to the electronic supplementary material.

Supplementary Material 1 (87.8KB, xlsx)
Supplementary Material 2 (77.5KB, xlsx)

Acknowledgements

Our study was supported by the NSF/NIH Ecology and Evolution of Infectious Diseases program and funded by National Institute of General Medical Sciences of the National Institutes of Health under award number R01GM085232. This study was also supported by NIH grant 5T35OD010956-25 through the University of California, Davis (UC Davis) - Students Training in Advanced Research (STAR) Program. We are thankful to Andrea Packham and Magdalena Plancarte at the One Health Institute at UC Davis and Marissa Young at the Monterey Bay Aquarium for facilitating access to archived serum samples. We are grateful for the many volunteers and staff working at CDFW, TMMC and MBA that respond to SSO strandings and have made this work possible.

Author contributions

Conceptualization: Karen Shapiro; Methodology: Ellis Song, Devinn Sinnott; Formal analysis and investigation: Ellis Song, Karen Shapiro; Writing - original draft preparation: Ellis Song; Writing - review and editing: Devinn Sinnott, Cara L. Field, Michael Murray, Pádraig Duignan, Melissa Miller, Karen Shapiro; Resources: Karen Shapiro; Supervision: Karen Shapiro.

Funding

Our study was supported by the NSF/NIH Ecology and Evolution of Infectious Diseases program and funded by National Institute of General Medical Sciences of the National Institutes of Health under award number R01GM085232.

Data availability

All data supporting the findings of this study are available within the paper and its Supplementary Information.

Declarations

Ethics approval

Serum samples from southern sea otters included in this study were obtained from animals recovered through authorized stranding response by the California Department of Fish and Wildlife (CDFW), and/or rehabilitation of animals that initially stranded alive by The Marine Mammal Center (TMMC), and Monterey Bay Aquarium (MBA). All handling protocols were approved through applicable federal permits and regulatory authorizations as detailed below. Animals included in this study consisted of stranded free-ranging southern sea otters that were found dead, died during care, or were humanely euthanized when necessary for animal welfare reasons; no animals were euthanized specifically for the purposes of this research.

CDFW handled stranded sea otters under Sect.  109(h) of the U.S. Marine Mammal Protection Act (MMPA), in accordance with U.S. Fish and Wildlife Service (USFWS) regulations implementing the MMPA (50 CFR 18.22[a]) and the U.S. Endangered Species Act (50 CFR 17.21[c][3]). TMMC and MBA conducted animal recovery, rehabilitation, humane euthanasia, and sample collection under USFWS permits MA101713-1 and PER4324646 (TMMC), and MA032027 and MA186914 (MBA), respectively. These statutes and permits authorize wildlife agencies and rehabilitation centers to intervene on behalf of free-ranging marine mammals for their protection and welfare, including rehabilitation and humane euthanasia when indicated. All animal handling and euthanasia procedures were conducted under standardized institutional animal care and veterinary oversight protocols, and gross necropsies were performed on all sea otters by veterinary pathologists and trained staff at CDFW and TMMC. Serum and other biological samples were collected from fresh or previously frozen/thawed carcasses during postmortem examination.

Consent to participate

Not applicable.

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

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References

  1. Dubey JP (2016) Toxoplasmosis of Animals and Humans, 2nd edn. CRC, Boca Raton, Florida, p 336. 10.1201/9781420092370 [DOI] [Google Scholar]
  2. Gardner IA, Stryhn H, Lind P, Collins MT (2000) Conditional dependence between tests affects the diagnosis and surveillance of animal diseases. Prev Vet Med 45(1–2):107–122. 10.1016/S0167-5877(00)00119-7 [DOI] [PubMed] [Google Scholar]
  3. Hajian-Tilaki K (2014) Sample size estimation in diagnostic test studies of biomedical informatics. J Biomed Inform 48:193–204. 10.1016/j.jbi.2014.02.013 [DOI] [PubMed] [Google Scholar]
  4. Johnson CK, Tinker MT, Estes JA, Conrad PA, Staedler M, Miller MA, Jessup DA, Mazet JAK (2009) Prey choice and habitat use drive sea otter pathogen exposure in a resource-limited coastal system. Proc Natl Acad Sci 106:2242–2247. 10.1073/pnas.0806449106 [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Khan AH, Noordin R (2020) Serological and molecular rapid diagnostic tests for Toxoplasma infection in humans and animals. Eur J Clin Microbiol Infect Dis 39:19–30. 10.1007/s10096-019-03680-2 [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kreuder C, Miller MA, Jessup DA, Lowenstine LJ, Harris MD, Ames JA, Carpenter TE, Conrad PA, Mazet JAK (2003) Patterns of mortality in southern sea otters (Enhydra lutris nereis) from 1998–2001. J Wildl Dis 39:495–509. 10.7589/0090-3558-39.3.495 [DOI] [PubMed] [Google Scholar]
  7. Krusor C, Smith WA, Tinker MT, Silver M, Conrad PA, Shapiro K (2015) Concentration and retention of Toxoplasma gondii oocysts by marine snails demonstrate a novel mechanism for transmission of terrestrial zoonotic pathogens in coastal ecosystems. Environ Microbiol 17:4527–4537. 10.1111/1462-2920.12927 [DOI] [PubMed] [Google Scholar]
  8. Miller MA, Gardner IA, Packham A, Mazet JK, Hanni KD, Jessup D, Estes J, Jameson R, Dodd E, Barr BC, Lowenstine LJ, Gulland FM, Conrad PA (2002) Evaluation of an indirect fluorescent antibody test (IFAT) for demonstration of antibodies to Toxoplasma gondii in the sea otter (Enhydra lutris). J Parasitol 88:594–599. 10.1645/0022-3395(2002)088[0594:EOAIFA]2.0.CO;2 [DOI] [PubMed] [Google Scholar]
  9. Miller M, Shapiro K, Murray MJ, Haulena M, Raverty S (2018) Protozoan parasites of marine mammals. In: Gulland FMD, Dierauf LA, Whitman KL (eds) CRC Handbook of Marine Mammal Medicine, 3rd edn. CRC, Boca Raton, Florida., pp 425–470. doi: 10.1201/9781315144931 [DOI] [Google Scholar]
  10. Miller MA, Moriarty ME, Henkel L, Tinker MT, Burgess TL, Batac FI, Dodd E, Young C, Harris MD, Jessup DA, Ames J, Conrad PA, Packham AE, Johnson CK (2020) Predators, disease, and environmental change in the nearshore ecosystem: mortality in southern sea otters (Enhydra lutris nereis) from 1998–2012. Front Mar Sci 7:582. 10.3389/fmars.2020.00582 [DOI] [Google Scholar]
  11. Miller MA, Moriarty ME, Duignan PJ, Zabka TS, Dodd E, Batac FI, Young C, Reed A, Harris MD, Greenwald K, Kudela RM, Murray MJ, Gulland FMD, Miller PE, Hayashi K, Gunther-Harrington CT, Tinker MT, Toy-Choutka S (2021) Clinical signs and pathology associated with domoic acid toxicosis in southern sea otters (Enhydra lutris nereis). Front Mar Sci 8:585501. 10.3389/fmars.2021.585501 [DOI] [Google Scholar]
  12. Miller MA, Newberry CA, Sinnott DM, Batac FI, Greenwald K, Reed A, Young C, Harris MD, Packham AE, Shapiro K (2023) Newly detected, virulent Toxoplasma gondii COUG strain causing fatal steatitis and toxoplasmosis in southern sea otters (Enhydra lutris nereis). Front Mar Sci 10:1116899. 10.3389/fmars.2023.1116899 [DOI] [Google Scholar]
  13. Robinson SJ, Amlin A, Barbieri MM (2023) Terrestrial pathogen pollutant, Toxoplasma gondii, threatens Hawaiian monk seals (Neomonachus schauinslandi) following heavy runoff events. J Wildl Dis 59(1):1–11. 10.7589/JWD-D-21-00179 [DOI] [PubMed] [Google Scholar]
  14. Roe WD, Howe L, Baker EJ, Burrows L, Hunter SA (2013) An atypical genotype of Toxoplasma gondii as a cause of mortality in Hector’s dolphins (Cephalorhynchus hectori). Vet Parasitol 192(1–3):67–74. 10.1016/j.vetpar.2012.11.001 [DOI] [PubMed] [Google Scholar]
  15. Shapiro K, Bahia-Oliveira L, Dixon B, Dumètre A, de Wit LA, VanWormer E, Villena I (2019a) Environmental transmission of Toxoplasma gondii: Oocysts in water, soil and food. Food Waterborne Parasitol 15:e00049. 10.1016/j.fawpar.2019.e00049 [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Shapiro K, VanWormer E, Packham A, Dodd E, Conrad PA, Miller M (2019b) Type X strains of Toxoplasma gondii are virulent for southern sea otters (Enhydra lutris nereis) and present in felids from nearby watersheds. Proc Royal Soc B 286(1909):20191334. 10.1098/rspb.2019.1334 [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sinnott DM, Miller M, Arranz-Solís D, Rodriguez F, Saeij JPJ, Batac F, Greenwald K, Young C, Harris MD, Gomes M, Shapiro K (2025) Fatal Toxoplasma gondii COUG strain infections in southern sea otters (Enhydra lutris nereis): New insight on contributing factors and parasite serotyping. PLoS ONE 20:e0332223. 10.1371/journal.pone.0332223 [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. StataCorp (2025) Stata Statistical Software: Release 19. College Station. StataCorp LLC, TX [Google Scholar]
  19. Strules J, Dawant T, Riese K, Gerhold R, Brown J, Olfenbuttel C, Deperno CS, Hunt BJ, von Dohlen AR (2023) Use of a point of care test to determine the prevalence of antibodies to Toxoplasma gondii in black bears from North Carolina and Pennsylvania. J Parasitol 109:221–224. 10.1645/22-72 [DOI] [PubMed] [Google Scholar]
  20. Thomas N, White CL, Saliki J, Schuler K, Lynch D, Nielsen O, Dubey JP, Knowles S (2020) Canine distemper virus in the sea otter (Enhydra lutris) population in Washington State, USA. J Wildl Dis 56(4):873–883. 10.7589/JWD-D-19-00008 [DOI] [PubMed] [Google Scholar]
  21. Trevethan R (2017) Sensitivity, specificity, and predictive values: foundations, pliabilities, and pitfalls in research and practice. Front Public Health 5:307. 10.3389/fpubh.2017.00307 [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. von Rosypal A, Randall S, Grays J, Dugo MA, Hunt BJ, Brown J, Van Why K, Ammar S, Gerhold R (2024) Rapid point-of-care testing for detection of antibodies to Toxoplasma gondii in black vultures and ring-billed gulls from Pennsylvania. J Parasitol 110:206–209. 10.1645/24-24 [DOI] [PubMed] [Google Scholar]
  23. World organisation for animal health (WOAH) (2023) Principles and methods of validation of diagnostic assays for infectious diseases. In: manual of diagnostic tests and vaccines for terrestrial animals. WOAH, Paris. Available at: https://www.woah.org/fileadmin/Home/fr/Health_standards/tahm/1.01.06_VALIDATION.pdf. Accessed 13 May 2026

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Supplementary Material 1 (87.8KB, xlsx)
Supplementary Material 2 (77.5KB, xlsx)

Data Availability Statement

All data supporting the findings of this study are available within the paper and its Supplementary Information.

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