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During November-December 2017, a mass die-off of Guiana dolphins (Sotalia guianensis) began in Rio de Janeiro, Brazil. Molecular and pathologic investigations on 20 animals indicated that cetacean morbillivirus played a major role. Our findings increase the knowledge on health and disease aspects of this endangered species.

To the Editor: Four strains of cetacean morbillivirus (CeMV; family Paramyxoviridae, genus Morbillivirus) have been detected in the global cetacean population: porpoise morbillivirus (1), dolphin morbillivirus (2), pilot whale morbillivirus (PWMV) (3), and Longman’s beaked whale morbillivirus (4). In addition, 2 novel CeMV sequences or strains isolated from the Indo-Pacific bottlenose dolphin (Tursiops aduncus) and the Guiana dolphin (Sotalia guianensis) have been recently reported in the Southern Hemisphere (5,6). Pilot whales are known to be susceptible to 2 strains of CeMV, PWMV, and dolphin morbillivirus (3,7,8). Only 2 deaths of whales have been reported to be caused by PWMV: 1 long-finned pilot whale (Globicephala melas) (3) and 1 short-finned pilot whale (G. macrorhynchus) (8). We report deaths of 3 short-finned pilot whales caused by PWMV in the northeastern Atlantic Ocean along the coast of the Canary Islands, Spain. During mid-January–May 2015, a total of 3 whales (animals 1, 2, and 3) were found dead along the coasts of the Canary Islands (Table). Complete standardized necropsy was performed for all whales. Tissue samples from animals 1 and 2 were fixed in 10% neutral-buffered formalin for histologic and immunohistochemical analyses (Technical Appendix Figure). Immunohistochemical analysis was performed (brain, intestine, lymph nodes, lung, kidney, adrenal gland, uterus, ovary, testis, and spleen) by using a monoclonal antibody against the nucleoprotein of canine distemper virus (CDV-NP; VMRD, Inc., Pullman, WA, USA) (7). Samples of lung, pulmonary lymph nodes, larynx, laryngeal tonsil, intestine, spleen, and brain were frozen (−80°C) for virologic analysis. Table Characteristics for 3 short-finned pilot whales stranded along the Canary Islands, Spain, 2015* Grossly, the most remarkable findings in animal 1 were severe suppurative rhinitis, with clogged nasal passages by the accumulation of large quantity of purulent material, otitis media, sacculitis, and laryngitis. Severe diffuse epithelial hyperplasia and hyperkeratosis was observed along the upper respiratory tract and keratinized stomach. Animal 2 had severe proliferative dermatitis and cheilitis, and severe, suppurative, laryngeal tonsillitis. Animal 3 had advanced autolysis, which precluded pathologic analysis. Histologically, moderate, multifocal, bronchointerstitial pneumonia, severe suppurative tonsillitis and systemic lymphoid depletion were identified in animals 1 and 2. Severe nonsuppurative meningoencephalitis with neuronal and glial cell degeneration and necrosis, microgliosis and syncytial cells were observed in animal 2. Immunohistochemical analysis showed morbillivirus antigen in the bronchiolar epithelium, type 2 pneumocytes, and alveolar multinucleate cells. Syncytia from lymph nodes, laryngeal tonsil, spleen, and intestine also showed positive immunolabeling for morbillivirus. Epithelial tropism caused by the virus was suggested by identification of viral antigen in epithelia of the lung, larynx, keratinized stomach, intestine, kidney, urinary bladder, epididymis, and endometrial glands. In addition, intense immunolabeling was detected in neurons (soma, dendrites, axon hillock, and axons) and glial cells, primarily throughout the cerebral gray matter of animal 2. Molecular detection of CeMV was performed by a using a 1-step reverse transcription PCR for a 426-bp conserved region of the phosphoprotein gene (7). All tested samples from animals 1 and 2 and a laryngeal tonsil sample from animal 3 showed positive PCR results. Because co-infections with herpesvirus and morbillivirus were observed during morbillivirus epizootics in seals in 1988 and dolphins in 2006–2007, we also tested the same tissue for herpesvirus by conventional nested PCR (9). Herpesvirus DNA was detected in all samples from animal 1 except lung, although no specific lesions compatible with this infectious agent were observed. A pool containing all morbillivirus-positive PCR amplicons for animals 1 and 2 (GenBank accession nos. {"type":"entrez-nucleotide","attrs":{"text":"KT006289","term_id":"961377517"}}KT006289 and {"type":"entrez-nucleotide","attrs":{"text":"KT006290","term_id":"961377519"}}KT006290), a PCR amplicon for the brain sample from animal 2 (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KT006291","term_id":"961377521"}}KT006291), and a PCR amplicon for the larynx from animal 3 were sequenced. A BLAST search (http://www.ncbi.nlm.nih.gov/blast/Blast.cgi) showed that amplified samples were nearly identical to reference PWMV sequences (GenBank accession nos. {"type":"entrez-nucleotide","attrs":{"text":"AF200817","term_id":"7329127"}}AF200817 [3] and {"type":"entrez-nucleotide","attrs":{"text":"FJ842381","term_id":"239775410"}}FJ842381 [8]). The sequence obtained from animal 3 was too short and degenerated to be accurately classified as CeMV, although it showed high homology with PWMV and porpoise morbillivirus. It has been proposed that pilot whales might be enzootically infected with CeMV (10). These whales might be responsible for maintaining and transmitting CeMV over long distances or to other odontocetes. No die-offs have been observed in these species. However, an outbreak of a lethal morbillivirus infection in long-finned pilot whales caused by a dolphin morbillivirus strain occurred in the Mediterranean Sea during the end of October 2006–April 2007 (7). Results of this study support the previous hypothesis that pilot whales have a species-adapted morbillivirus but indicate that lethal infections are not as rare as previously believed (3). The tropism of the virus in these cases, the high number of multinucleated syncytial cells, and the severity of the lesions resemble the acute systemic symptoms observed in dolphins infected with morbillivirus (2). Thus, pilot whales in the northeastern Atlantic Ocean could be at risk for infection, especially in one of the main pilot whale–watching regions between La Gomera and Southern Tenerife Islands in the Canary Islands, which has >700,000 visitors each year. Technical Appendix. Microscopic images of tissue samples from 2 short-finned pilot whales (Globicephala macrorhynchus) from the eastern Atlantic Ocean stranded along the Canary Islands, Spain, 2015. Click here to view.(380K, pdf)

In January 2017, a yellow fever outbreak occurred in Espirito Santo, Brazil, where human immunization coverage is low. Histologic, immunohistologic, and PCR examinations were performed for 22 deceased nonhuman New World primates; typical yellow fever features were found in 21. Diagnosis in nonhuman primates prompted early public health response.

To the Editor: Since 1987, morbillivirus (family Paramyxoviridae, genus Morbillivirus) outbreaks among pinnipeds and cetaceans in the Northern Hemisphere have caused high rates of death (1,2). Two morbillivirus species are known to affect aquatic animals: Phocine distemper virus (PDV) and Cetacean morbillivirus (CeMV). PDV has been isolated from pinnipeds, and 3 strains of CeMV (porpoise morbillivirus [PMV], dolphin morbillivirus [DMV], and pilot whale morbillivirus [PWMV]) have been isolated from dolphins and whales (3,4). Serologic surveys indicate that morbilliviruses infect marine mammals worldwide (5); however, only 1 fatal case in a bottlenose dolphin (Tursiops truncatus) has been confirmed in the Southern Hemisphere (in the southwestern Pacific Ocean) (6). Positive DMV-specific antibody titers in 3 Fraser’s dolphins (Lagenodelphis hosei) stranded off Brazil and Argentina in 1999 indicate the exposure of South Atlantic cetaceans to morbillivirus (7). We report a case of lethal morbillivirus infection in a Guiana dolphin (Sotalia guianensis), a coastal marine and estuarine species that occurs off the Atlantic Coast of South and Central America. A female Guiana dolphin calf (108 cm in total body length) (8) was found stranded dead in Guriri (18°44’S; 39°44’W), Sao Mateus, Espirito Santo State, Brazil, on November 30, 2010; the dead calf was severely emaciated. Postmortem examination of the animal showed multifocal ulcers in the oral mucosa and genital slit, diffusely dark red and edematous lungs, and congested and edematous brain. Samples of selected tissues were collected, fixed in buffered formalin, and processed according to routine histopathologic methods. By microscopy, the most noteworthy lesions included marked lymphoplasmacytic and neutrophilic meningoencephalitis, optic nerve perineuritis, and hypophysitis. Lungs showed moderate acute diffuse lymphoplasmacytic and neutrophilic interstitial pneumonia; severe multicentric lymphoid depletion and multifocal necrotizing hepatitis were also observed. Immunohistochemistry was performed by using CDV-NP MAb (VMRD, Inc., Pullman, WA, USA), a monoclonal antibody against the nucleoprotein antigen of canine distemper virus that cross-reacts with cetacean morbilliviruses (9). Known positive and negative control tissues and test sections with omitted first-layer antibody were included. Viral antigen was detected in neurons in the brain, bronchiolar epithelium and macrophages in the lungs, bile duct epithelium in the liver, and macrophages and lymphocytes in lymph nodes. We extracted RNA from frozen lung samples by using TRIzol Reagent (Life Technologies Corporation, Carlsbad, CA, USA) according to the manufacturer’s instructions and amplified a 374-bp conserved fragment of the phosphoprotein (P) gene by reverse transcription PCR. The following Morbillivirus spp.–specific primers were used for PCR: 5′-ATGTTTATGATCACAGCGGT-3′ (forward) and 5′-ATTGGGTTGCACCACTTGTC-3′ (reverse) (3). MEGA5 (http://megasoftware.net/) was used to construct a neighbor-joining phylogenic tree based on the sequenced amplicon from this study (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KF711855","term_id":"566036178","term_text":"KF711855"}}KF711855) and 12 other GenBank sequences that represent the 6 morbillivirus species already described in the literature. The analysis placed the Guiana dolphin strain at the CeMV clade, but segregated it from the already described dolphin morbillivirus strains PMV, DMV, and PWMV (Figure). The sample shared 79.8% nt and 58.4% aa identity with PMV, 78.7% nt and 56.6% aa identity with DMV, and 78.7% nt and 57.1% aa identity with PWMV. Within the Morbillivirus spp., PDV shared the lowest sequence identity (51.1% nt and 26.8% aa). Figure Phylogenetic tree of a 374-bp conserved region from the phosphoprotein gene of a cetacean morbillivirus isolated from a Guiana dolphin (in boldface; GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KF711855","term_id":"566036178","term_text":"KF711855"}} ... In summary, sequence analysis of the morbillivirus from the dead Guiana dolphin suggests that the virus is a novel strain of the CeMV species; this conclusion is supported by phylogenic analysis and geographic distribution of the virus and by its distinct host. Emaciation, marked lymphoid depletion, interstitial pneumonia, and meningoencephalitis are common findings in morbillivirus-infected animals (1,2). Together with antigenic and genomic evidence, our findings indicate that morbillivirus infection is extant in Guiana dolphins in the waters off Brazil. Morbillivirus outbreaks have caused a high number of deaths among pinnipeds and cetaceans and are a major risk to previously unexposed nonimmune populations of aquatic mammals (1,2). A high number of morbillivirus-related deaths have not yet been reported among aquatic mammals in the waters off Brazil, but our findings shows that Guiana dolphin calves are susceptible to infection. Subclinical morbillivirus infection with immune suppression has been reported in bottlenose dolphins in Florida (10). It is unknown whether subclinical infection occurs in this host population or whether the virus has undergone species-adaptive changes, as proposed for PWMV (4). The sequence data from our study suggest that the virus from the Guiana dolphin calf is the fourth member of the CeMV group and is closer to the root of the CeMV clade than to that of DMV, PMV, or PWMV. Further studies are required to determine the epidemiology of morbillivirus infection in this and other cetacean species and to assess the risk for epizootic outbreaks among South Atlantic cetaceans.