Figure 1

Figure 1. Stranding locations and animal species in study of Neospora caninum infection in marine mammals stranding in the northeastern Pacific Ocean region. NES, northern elephant seal (Mirounga angustirostris...

Throughout the northeastern Pacific region, local and regional marine mammal stranding networks respond to live stranded and dead marine mammals. In this case series, wildlife officials delivered a California sea lion (Zalophus californianus) for rehabilitation. Despite supportive care, the animal declined and was euthanized. We identified a solitary N. caninum infection using PCR, observing no discernible parasites by histopathology. As a result of that finding, we conducted a retrospective database analysis on 410 stranded marine mammals previously screened by PCR-DNA sequencing at the internal transcribed spacer (ITS) 1 marker. We identified 7 additional cases of N. caninum infection in another 5 species of marine mammals. The geographic range of the N. caninum–infected animals (Figure 1) suggested multiple points, rather than a single point source, of parasite exposure.

We documented the stranding location, recorded signalment, compiled morphometrics, and performed a necropsy for all 8 cases (Appendix Table). We also harvested representative tissues for histopathology and immunohistochemistry and froze samples for ancillary diagnostic studies. We conducted PCR-DNA sequencing using pan-Apicomplexan primers that amplify across the ITS1 region for speciation. We carried out immunohistochemistry for N. caninum, T. gondii, and S. neurona on available tissues from 4 PCR-positive animals, following previously reported protocols (1,12).

Figure 2

Figure 2. Phylogenetic relationships among Neospora caninum–like species that circulate in a marine cycle compared with terrestrial-sourced species from study of N. caninuminfection in marine mammals stranding in...

Phylogenetic analysis at the ITS1 locus established unequivocally that all 8 animals were infected with an identical sequence type that resolved as N. caninum (GenBank accession nos. PX529932–8) (Figure 2). For comparison, we included in the tree sequences recovered from other pinnipeds infected by N. caninum–like parasites that commonly circulate within a marine transmission cycle (5). We conducted PCR testing, determining 7 of the 8 N. caninum–infected animals had polyparasite infections with 1 or 2 other terrestrially sourced coccidian agents (T. gondii, S. neurona) in their tissues. Infected animals were 2 sea otters (Enhydra lutris), 2 harbor seals (Phoca vitulina), 1 northern elephant seal (Mirounga angustirostris), 1 Guadelupe fur seal (Arctocephalus townsendi), and 1 Steller sea lion (Eumetopias jubatus) (Appendix Table).

Figure 3

Figure 3. Immunohistochemistry of a harbor seal pup with Neospora caninum antigen localized within a neuron from study of N. caninuminfection in marine mammals stranding in the northeastern...

We diagnosed nonsuppurative and necrotizing encephalitis (n = 2), meningoencephalomyelitis (n = 1), and meningoencephalitis (n = 1) by histology of the brain for 4 of 6 cases, including the pup and subadult harbor seals, adult Steller sea lion, and yearling California sea lion. We conducted immunohistochemistry in 4 encephalitic cases, observing N. caninum antigen in brain samples from the harbor seal pup and T. gondii and S. neurona (one or both) antigens in the other 3 animals (Figure 3). We found no pathognomonic lesions in this case series directly attributed to N. caninum infection. The cause of death for all 8 animals was independent of N. caninum infection (Appendix Table).

We identified 8 animals from 6 marine mammal species unequivocally infected with N. caninum. Infection by this parasite alone was confirmed in 1 animal, a yearling California sea lion, and the infection represents another example of land-to-sea flow of a pollutagen of parasite origin. The contribution of this pathogen to marine mammal health is unknown. All infected animals we studied were from species with varying degrees of philopatry, with either short natal dispersal distances or extended oceanic to pelagic migrations. Seasonal haul out associated with pupping and breeding, coupled with accounts of wild canids scavenging at rookeries (13), provide insights into potential parasite introduction, persistence, and dissemination. For sea otters, bioaccumulation of parasites by invertebrate prey species may be inferred from observations of transmission dynamics with T. gondii (4). On the basis of gross and microscopic findings, we determined N. caninum infection to be unrelated to the proximate cause of death in the examined animals. Nevertheless, detection of N. caninum in a pregnant Steller sea lion, as well as in a harbor seal pup and northern elephant seal pup (weanling), is concerning. Recrudescence of latent infection during pregnancy contributes to parasite reactivation that results in vertical transmission in cattle and dogs (6), and we propose this phenomenon might be occurring sporadically in marine mammals.

Documented reports have confirmed land-to-sea transmission of protozoan parasites, and serosurveys for N. caninum have detected titers in marine mammal species. A recent review documents the global extent of land-to-sea pathogen flow (14). Seals and sea lions are monophyletic, members of the order Carnivora, and share a common ancestry with terrestrial canids. This evolutionary relationship may predispose these marine mammals to pollutagens defecated by canids. Cross reactivity with previously described nonpathogenic N. caninum–like parasites that circulate among pinnipeds in a marine transmission cycle, including coccidia types A and B (with California sea lions as definitive hosts) (5), may have contributed to false-positive results in prior serosurveys. The contribution of these N. caninum–like parasites to immune protection of hosts against N. caninum is unknown. In this limited series, we confirmed N. caninum infection in 2 sea otters, 2 harbor seals, 1 California sea lion, 1 northern elephant seal, 1 Steller sea lion, and 1 Guadelupe fur seal, none of which had been infected with previously described N. caninum–like parasites. Those findings extend the host range and ecologic niche for N. caninum. Infections were predominantly mixed with T. gondii, S. neurona, or both. Climate change, ecologic marine regime shifts, rural and urban development, weather events, and other environmental perturbations may lead to incursion of previously recognized land-based pathogens into the marine environment.

In conclusion, our results indicate that further investigations to characterize the life history of N. caninum in the marine environment, the role of polyparasitism in disease manifestation, and the potential pathogenicity in susceptible host species are warranted. Whereas we cannot definitively attribute N. caninum as the cause of illness or death for the animals described here, future investigations of similar unexpected deaths, particularly those involving abortion storms among coastal breeding marine mammals in the northeastern Pacific, should consider molecular screening for this pathogen.