Molecular Detection and Characterization of Mycoplasma spp. in Marine Mammals, Brazil.

Mycoplasma spp. are wall-less bacteria able to infect mammals and are classified as hemotropic (hemoplasma) and nonhemotropic. In aquatic mammals, hemoplasma have been reported in California sea lions (Zalophus californianus) and river dolphins (Inia spp.). We investigated Mycoplasma spp. in blood samples of West Indian manatees (Trichechus manatus), pinnipeds (5 species), and marine cetaceans (18 species) that stranded or were undergoing rehabilitation in Brazil during 2002-2022. We detected Mycoplasma in blood of 18/130 (14.8%) cetaceans and 3/18 (16.6%) pinnipeds. All tested manatees were PCR-negative for Mycoplasma. Our findings indicate that >2 different hemoplasma species are circulating in cetaceans. The sequences from pinnipeds were similar to previously described sequences. We also detected a nonhemotropic Mycoplasma in 2 Franciscana dolphins (Pontoporia blainvillei) that might be associated with microscopic lesions. Because certain hemoplasmas can cause disease and death in immunosuppressed mammals, the bacteria could have conservation implications for already endangered aquatic mammals.

The deep dive of organohalogen compounds: Bioaccumulation in the top predators of mesopelagic trophic webs, pygmy and dwarf sperm whales, from the Southwestern Atlantic ocean.

Kogia sima and Kogia breviceps are apex predators of mesopelagic trophic webs being far from most anthropogenic threats. However, chemical pollutants and naturally synthesized compounds may travel long distances. This study aimed to use kogiid whales as sentinels of mesopelagic trophic webs in the Southwestern Atlantic Ocean. Persistent organic pollutants (POPs), e.g., polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane (DDT) and metabolites, mirex, hexachlorobenzene (HCB), polybrominated diphenylethers (PBDEs), pentabromoethylbenzene (PBEB) and hexabromobenzene (HBB), and the naturally produced methoxylated BDE (MeO-BDEs) were determined in the blubber of 16 K. sima and 15 K. breviceps. Among the organochlorine compounds, DDTs were the main group found in K. sima and in K. breviceps (1636.6 and 3983.3 ng g-1 lw, respective medians), followed by PCBs (425.9 and 956.1 ng g-1 lw, respectively), mirex (184.1 and 375.6 ng g-1 lw, respectively), and HCB (132.4 and 340.3 ng g-1 lw, respectively). As for the organobromine, the natural MeO-BDEs were predominant (1676.7 and 501.6 ng g-1 lw, respectively), followed by PBDEs (13.6 and 10.3 ng g-1 lw, respectively) and PBEB (2.2 and 2.9 ng g-1 lw, respectively). In general, POPs concentration was higher in K. breviceps than in K. sima. Conversely, MeO-BDEs concentration was higher in K. sima than in K. breviceps. Differences in concentrations in these sympatric odontocetes were attributed to distinct species, sampling sites, and biological parameters and suggest some level of niche segregation. It is noteworthy the long-range reach and bioaccumulation of these synthetic compounds in an unexplored habitat, that present an increasing economic interest.

Franciscana dolphins, Pontoporia blainvillei, as environmental sentinels of the world's largest mining disaster: Temporal trends for organohalogen compounds and their consequences for an endangered population.

On November 5th, 2015, the Fundão dam collapsed in Minas Gerais, southeastern Brazil, releasing millions of cubic meters of mud containing mining residue into the Doce River. Two weeks later, the mud arrived to the marine environment, triggering changes in franciscana dolphin habitat, Pontoporia blainvillei, from Franciscana Management Area Ia. This is an isolated population of the most endangered cetacean species in the South Atlantic Ocean. Organohalogen compounds (OHCs) may pose a threat to this endangered population because of their endocrine disrupting properties. Hence, this study sought to determine if there were differences in the bioaccumulation profile of OHC (PCBs, DDTs, Mirex, HCB, HCHs, PBDEs, PBEB, HBBZ and MeO-BDEs) in franciscana dolphins before and after dam collapse and to build a temporal trend. Blubber of 33 stranded individuals was collected in Espírito Santo state for organohalogen assessment between 2003 and 2019. Differences were found between franciscana dolphins collected prior to and after the disaster. Additionally, significant temporal trends for organochlorine pesticides and natural and anthropogenic organobromine were detected. The increase in pesticide concentrations after 2015 is suggestive of their reavailability in the environment. The decline in organobromine over time could be due to their debromination in the marine environment and alterations in the composition of their natural producers. PCBs remained stable during the period of the study. Our findings show an increase in endocrine disruptor concentrations, which is of great concern for this endangered population.

Microsatellite markers for the endangered franciscana dolphin (Pontoporia blainvillei).

The franciscana (Pontoporia blainvillei) is the most threatened small cetacean in the South Atlantic. In this study we report the development of 13 microsatellite markers for franciscanas through next-generation sequencing, and the characterization of those loci in 38 samples from the species' northernmost population (Espírito Santo, Brazil). Besides providing diversity indices for the new, specific loci, we also report on the transferability of heterologous loci which had not been screened in franciscanas before, and review all loci used in previous studies. Expected heterozygosity in the new loci ranged between 0.107 and 0.595, and all but one were in Hardy Weinberg Equilibrium. These are the first microsatellite loci isolated from franciscanas, and they are an important addition to heterologous markers that were available previously.

Cetacean morbillivirus in Humpback whales' exhaled breath.

The humpback whale (HW; Megaptera novaeangliae) population that seasonally resides along the Brazilian coast concentrates in the Abrolhos Bank (Bahia and Espírito Santo states) for breeding during austral winter and spring. Cetacean morbillivirus (CeMV, Paramyxoviridae family) is currently one of the most significant biological threats to cetaceans worldwide with high infection and mortality rates. CeMV is pleiotropic yet it has special tropism for the respiratory, lymphoid and nervous system and is primarily transmitted by the aerogenous route. A new lineage of CeMV, the Guiana dolphin morbillivirus (GDMV), is known to affect cetaceans off Brazil. GDMV was first detected in a Guiana dolphin (Sotalia guianensis) stranded in the Abrolhos Bank region, in 2010. In addition to pathologic examinations on stranded HW, pathogen survey of free-ranging HW may provide valuable insight into the epidemiology of diseases. We hypothesized that HW in the Brazilian breeding ground could be exposed to CeMV. Thus, in the present study, we investigated the presence of CeMV in exhaled breath condensates (EBC) of HW in the Abrolhos Bank. Overall, 73 samples of EBC from 48 groups of HW were collected during the breeding seasons of 2011 (n = 16) and 2012 (n = 57). One sample failed to have the reference gene amplified and was excluded from the study. CeMV was detected by a RT-qPCR method in 2 EBC samples, representing 2 whale groups. Phylogenetic analysis of partial morbillivirus phosphoprotein gene showed 100% homology to GDMV. Our results show that HW in Brazil are infected by CeMV with a relative prevalence of 4.3% (2/47) and demonstrate the suitability of using EBC and RT-qPCR as a non-invasive tool for CeMV survey in free-ranging whales. This pioneer study provides scientific basis for non-invasive CeMV monitoring of HW, suggests HW may play a role in the dynamics of CeMV and raises concern for potential conservation implications for this species.

Toxoplasma gondii in cetaceans of Brazil: a histopathological and immunohistochemical survey.

Toxoplasmosis is a parasitic disease caused by the protozoan Toxoplasma gondii. In cetaceans, T. gondii infection is a significant cause of morbidity and mortality. Despite the worldwide range and broad cetacean host record of T. gondii infection, there is limited information on toxoplasmosis in cetaceans from the Southern hemisphere. We investigated the occurrence of T. gondii by histopathology and immunohistochemistry in tissue samples of 185 animals comprising 20 different cetacean species from Brazil. Three out of 185 (1.6%) animals presented T. gondii-associated lesions: a captive killer whale Orcinus orca, a free-ranging common bottlenose dolphin Tursiops truncatus and a free-ranging Guiana dolphin Sotalia guianensis. The main lesions observed in these animals were necrotizing hepatitis, adrenalitis and lymphadenitis associated with protozoal cysts or extracellular tachyzoites presenting immunolabeling with anti-T. gondii antibodies. This study widens the spectrum of species and the geographic range of this agent in Brazil, and provides the first reports of T. gondii infection in a captive killer whale and in a free-ranging common bottlenose dolphin in South America.

Toxoplasma gondii in cetaceans of Brazil: a histopathological and immunohistochemical survey / Toxoplasma gondii em cetáceos do Brasil: estudo histopatológico e imuno-histoquímico

Abstract Toxoplasmosis is a parasitic disease caused by the protozoan Toxoplasma gondii. In cetaceans, T. gondii infection is a significant cause of morbidity and mortality. Despite the worldwide range and broad cetacean host record of T. gondii infection, there is limited information on toxoplasmosis in cetaceans from the Southern hemisphere. We investigated the occurrence of T. gondii by histopathology and immunohistochemistry in tissue samples of 185 animals comprising 20 different cetacean species from Brazil. Three out of 185 (1.6%) animals presented T. gondii-associated lesions: a captive killer whale Orcinus orca, a free-ranging common bottlenose dolphin Tursiops truncatus and a free-ranging Guiana dolphin Sotalia guianensis. The main lesions observed in these animals were necrotizing hepatitis, adrenalitis and lymphadenitis associated with protozoal cysts or extracellular tachyzoites presenting immunolabeling with anti-T. gondii antibodies. This study widens the spectrum of species and the geographic range of this agent in Brazil, and provides the first reports of T. gondii infection in a captive killer whale and in a free-ranging common bottlenose dolphin in South America.

Resumo Toxoplasmose é uma doença parasitária causada pelo protozoário Toxoplasma gondii. A infecção por T. gondii é uma causa significativa de morbidade e mortalidade, nos cetáceos. Apesar da abrangência mundial e amplo registro de espécies de cetáceos infectadas por T. gondii, informações sobre toxoplasmose em cetáceos do hemisfério sul são limitadas. Neste estudo pesquisou-se por meio de histopatologia e imuno-histoquímica a ocorrência de T. gondii em amostras de tecido de 185 animais, compreendendo 20 diferentes espécies de cetáceos que ocorrem no Brasil. Três dos 185 (1,6%) animais apresentaram lesões associadas a T. gondii: uma orca Orcinus orca mantida em cativeiro, um golfinho-nariz-de-garrafa Tursiops truncatus e um boto-cinza Sotalia guianensis de vida livre. As principais lesões observadas nesses animais foram hepatite, adrenalite e linfadenite necrotizantes associadas a cistos protozoários ou taquizoítos extracelulares, marcados com anticorpos anti-T. gondii. O presente estudo amplia o espectro de espécies susceptíveis a esse agente e o seu alcance geográfico no Brasil, fornecendo o primeiro relato da infecção por T. gondii em uma orca mantida em cativeiro e em um golfinho-nariz-de-garrafa de vida livre na América do Sul.

Molecular, serological, pathological, immunohistochemical and microbiological investigation of Brucella spp. in marine mammals of Brazil reveals new cetacean hosts.

Brucella-exposure and infection is increasingly recognized in marine mammals worldwide. To better understand the epidemiology and health impacts of Brucella spp. in marine mammals of Brazil, molecular (conventional PCR and/or real-time PCR), serological (Rose Bengal Test [RBT], Competitive [c]ELISA, Serum Agglutination Test [SAT]), pathological, immunohistochemical (IHC) and/or microbiological investigations were conducted in samples of 129 stranded or by-caught marine mammals (orders Cetartiodactyla [n = 124], Carnivora [n = 4] and Sirenia [n = 1]). Previous serological tests performed on available sera of 27 of the 129 animals (26 cetaceans and one manatee), indicated 10 seropositive cetaceans. Conventional PCR and/or real-time PCR performed in cases with available organs (n = 119) and/or blood or swabs (n = 10) revealed 4/129 (3.1%) Brucella-infected cetaceans (one of them with positive serology; the remaining three with no available sera). Pathological, IHC and/or microbiological analyses conducted in PCR/real-time PCR and/or seropositive cases (n = 13) revealed Brucella-type lesions, including meningitis/meningoencephalitis, pneumonia, necrotizing hepatitis, pericarditis and osteoarthritis in some of those animals, and positive IHC was found in all of them (excepting two live-stranded animals without available organs). Brucella spp. culture attempts were unsuccessful. Our results demonstrated exposure, asymptomatic, acute and chronic Brucella sp. infection in several cetacean species in the Brazilian coast, highlighting the role of this pathogen in stranding and/or death, particularly in Clymene dolphin (Stenella clymene) and short-finned pilot whale (Globicephala macrorhynchus) off Ceará State. Novel hosts susceptible to Brucella included the franciscana (Pontoporia blainvillei), the Guiana dolphin (Sotalia guianensis) and the spinner dolphin (Stenella longirostris). Additionally, three coinfection cases involving Brucella spp. and cetacean morbillivirus, Edwarsiella tarda and Proteus mirabilis were detected. To the best of our knowledge, this is the first long-term and large-scale survey of Brucella spp. in marine mammals of South America, widening the spectrum of susceptible hosts and geographical distribution range of this agent with zoonotic potential.

Novel herpesviruses in riverine and marine cetaceans from South America.

Herpesvirus (HV) infections in cetaceans are frequently associated with skin and mucosal lesions. Although HV infections have been reported worldwide, their occurrence in southern Atlantic marine mammals is still poorly understood. We tested skin, oral and genital mucosal beta-actin PCR-positive samples from 109 free-ranging Brazilian cetaceans using a universal herpesvirus DNA polymerase PCR. Herpesvirus-positive skin samples from a Guiana dolphin (Sotalia guianensis), a dwarf sperm whale (Kogia sima), a Bolivian river dolphin (Inia boliviensis), and a lingual sample from an Atlantic spotted dolphin (Stenella frontalis) were histologically evaluated. Additional tissue samples from these animals were also PCR-positive for HV, including a novel sequence obtained from the dwarf sperm whale's stomach and mesenteric lymph node. Four novel HV species were detected in the Guiana dolphin (one), the dwarf sperm whale (two) and the Bolivian river dolphin (one). The cutaneous lesions (marked, focally extensive, chronic proliferative dermatitis) of the Guiana dolphin and the Bolivian river dolphin were similar to previous HV reports in cetaceans, despite the absence of intranuclear inclusion bodies. This is the largest HV survey in South American cetaceans and the first detection of HV infection in riverine dolphins worldwide.

Pathology and causes of death in stranded humpback whales (Megaptera novaeangliae) from Brazil.

This study describes the pathologic findings of 24 humpback whales (Megaptera novaeangliae) found stranded along the Brazilian coast from 2004 to 2016. Eighteen (75%) animals evaluated were found stranded alive. From these, 13 died naturally on shore and five were euthanized. Six died at sea and were washed ashore. Of the 24, 19 (79.2%) were calves, four (16.7%) were juveniles, and one (4.2%) was an adult. The most probable cause of stranding and/or death (CSD) was determined in 23/24 (95.8%) individuals. In calves, CSD included neonatal respiratory distress (13/19; 68.4%), infectious disease (septicemia, omphaloarteritis and urachocystitis; 3/19; 15.8%), trauma of unknown origin (2/19; 10.5%), and vehicular trauma (vessel strike; 1/19; 5.3%). In juveniles and adult individuals, CSD was: emaciation (2/5; 40%), sunlight-thermal burn shock (1/5; 20%); and discospondylitis (1/5; 20%). In one juvenile, the CSD was undetermined (1/5; 20%). This study integrates novel findings and published case reports to delineate the pathology of a South-western Atlantic population of humpback whales. This foundation will aid in the assessment of the population health and establish a baseline for development of conservation policies.

Occurrence of Prosthenorchis elegans in Free-living Primates from the Atlantic Forest of Southern Bahia, Brazil.

Parasite prevalence and abundance are important factors affecting species' conservation. During necropsies on a free-living golden-headed lion tamarin ( Leontopithecus chrysomelas ) and two Wied's marmosets ( Callithrix kuhlii ) in the Atlantic Forest of southern Bahia, Brazil, we collected a large number of adult intestinal parasites that we identified as Prosthenorchis elegans. This parasite is pathogenic for neotropical primates. Prosthenorchis spp. infestation is influenced by diet with increased risk of exposure from ingesting invertebrate intermediate hosts. The biological similarities and sympatric nature of these two nonhuman primates support that they may harbor similar infectious and parasitic agents.

Skeletal abnormalities in humpback whales Megaptera novaeangliae stranded in the Brazilian breeding ground.

Skeletal tissues of 49 humpback whales Megaptera novaeangliae that stranded between 2002 and 2011 along the Abrolhos Bank seashore and its adjacent waters in Brazil were studied. Twelve (24.5%) animals presented pathological changes in one or more bones. Degenerative changes and developmental malformations were most frequent (10.2% each), followed by inflammatory/infectious and traumatic lesions (8.2% each). Infectious diseases led to severe lesions of the caudal vertebrae of 2 whales. In one of these individuals, the lesions involved 6 caudal vertebrae, leading to ankylosis of 3 vertebrae. Degenerative changes were observed in the vertebral columns of 3 animals, involving the joints of 13 ribs of 1 individual, and in the humerus of 1 whale. Traumatic lesions, such as osseous callus in the ribs, were observed in 4 animals. In 1 whale, the rib showed severe osteomyelitis, possibly resulting from the infection of multiple fractures. Developmental abnormalities such as spina bifida on 3 cervical vertebrae of 1 whale, fusion of spinal processes on thoracic vertebrae of 1 individual and fusion of the first 2 ribs unilaterally or bilaterally in 4 animals were found. Chronic infectious conditions found in the axial skeleton may have restrained spinal mobility and had detrimental effects on the general health of the animals, contributing to stranding and death. To our knowledge, this is the first systematic study on skeletal lesions in stranded humpback whales.