Serological Diagnosis of Brucella Infections in Odontocetes▿

Brucella ceti causes disease in Odontoceti. The absence of control serum collections and the diversity of cetaceans have hampered the standardization of serological tests for the diagnosis of cetacean brucellosis. Without a “gold” standard for sensitivity and specificity determination, an alternative approach was followed. We designed an indirect enzyme-linked immunosorbent assay (iELISA) that recognizes immunoglobulins G (IgGs) from 17 odontocete species as a single group. For the standardization, we used Brucella melitensis and Brucella abortus lipopolysaccharides, serum samples from seven resident odontocetes with no history of infectious disease displaying negative rose bengal test (RBT) reactions, and serum samples from seven dolphins infected with B. ceti. We compared the performance of the iELISA with those of the protein G ELISA (gELISA), the competitive ELISA (cELISA), and the immunofluorescence (IF) and dot blot (DB) tests, using 179 odontocete serum samples and RBT as the reference. The diagnostic potential based on sensitivity and specificity of the iELISA was superior to that of gELISA and cELISA. The correlation and agreement between the iELISA and the gELISA were relatively good (Ri/g2 = 0.65 and κi/g = 0.66, respectively), while the correlation and agreement of these two ELISAs with cELISA were low (Ri/c2 = 0.46, Rg/c2 = 0.37 and κi/c = 0.62, κg/c = 0.42). In spite of using the same anti-odontocete IgG antibody, the iELISA was more specific than were the IF and DB tests. An association between high antibody titers and the presence of neurological symptoms in dolphins was observed. The prediction is that iELISA based on broadly cross-reacting anti-dolphin IgG antibody would be a reliable test for the diagnosis of brucellosis in odontocetes, including families not covered in this study

Demographic clusters identified within the northern Gulf of Mexico common bottlenose dolphin (Tursiops truncates) unusual mortality event: January 2010-June 2013.

A multi-year unusual mortality event (UME) involving primarily common bottlenose dolphins (Tursiops truncates) was declared in the northern Gulf of Mexico (GoM) with an initial start date of February 2010 and remains ongoing as of August 2014. To examine potential changing characteristics of the UME over time, we compared the number and demographics of dolphin strandings from January 2010 through June 2013 across the entire GoM as well as against baseline (1990-2009) GoM stranding patterns. Years 2010 and 2011 had the highest annual number of stranded dolphins since Louisiana's record began, and 2011 was one of the years with the highest strandings for both Mississippi and Alabama. Statewide, annual numbers of stranded dolphins were not elevated for GoM coasts of Florida or Texas during the UME period. Demographic, spatial, and temporal clusters identified within this UME included increased strandings in northern coastal Louisiana and Mississippi (March-May 2010); Barataria Bay, Louisiana (August 2010-December 2011); Mississippi and Alabama (2011, including a high prevalence and number of stranded perinates); and multiple GoM states during early 2013. While the causes of the GoM UME have not been determined, the location and magnitude of dolphin strandings during and the year following the 2010 Deepwater Horizon oil spill, including the Barataria Bay cluster from August 2010 to December 2011, overlap in time and space with locations that received heavy and prolonged oiling. There are, however, multiple known causes of previous GoM dolphin UMEs, including brevetoxicosis and dolphin morbillivirus. Additionally, increased dolphin strandings occurred in northern Louisiana and Mississippi before the Deepwater Horizon oil spill. Identification of spatial, temporal, and demographic clusters within the UME suggest that this mortality event may involve different contributing factors varying by location, time, and bottlenose dolphin populations that will be better discerned by incorporating diagnostic information, including histopathology

Caryospora-like Coccidia infecting green turtles (Chelonia mydas): an emerging disease with evidence of interoceanic dissemination

Protozoa morphologically consistent with Caryospora sp. are one of the few pathogens associated with episodic mass mortality events involving free-ranging sea turtles. Parasitism of green turtles (Chelonia mydas) by these coccidia and associated mortality was first reported in maricultured turtles in the Caribbean during the 1970s. Years later, epizootics affecting wild green turtles in Australia occurred in 1991 and 2014. The first clinical cases of Caryospora-like infections reported elsewhere in free-ranging turtles were from the southeastern US in 2012. Following these initial individual cases in this region, we documented an epizootic and mass mortality of green turtles along the Atlantic coast of southern Florida from November 2014 through April 2015 and continued to detect additional, sporadic cases in the southeastern US in subsequent years. No cases of coccidial disease were recorded in the southeastern US prior to 2012 despite clinical evaluation and necropsy of stranded sea turtles in this region since the 1980s, suggesting that the frequency of clinical coccidiosis has increased here. Moreover, we also recorded the first stranding associated with infection by a Caryospora-like organism in Hawai'i in 2018. To further characterize the coccidia, we sequenced part of the 18S ribosomal and mitochondrial cytochrome oxidase I genes of coccidia collected from 62 green turtles found in the southeastern US and from one green turtle found in Hawai'i. We also sequenced the ribosomal internal transcribed spacer regions from selected cases and compared all results with those obtained from Caryospora-like coccidia collected from green turtles found in Australia. Eight distinct genotypes were represented in green turtles from the southeastern US. One genotype predominated and was identical to that of coccidia collected from the green turtle found in Hawai'i. We also found a coccidian genotype in green turtles from Florida and Australia with identical 18S and mitochondrial sequences, and only slight inter-regional differences in the internal transcribed spacer 2. We found no evidence of geographical structuring based on phylogenetic analysis. Low genetic variability among the coccidia found in green turtle populations with minimal natural connectivity suggests recent interoceanic dissemination of these parasites, which could pose a risk to sea turtle populations

Map of the U.S. Gulf Coast showing county groups used in analysis of regional monthly baseline common bottlenose dolphin (<i>Tursiops truncatus</i>) stranding numbers.

<p>Baselines for Texas and Western Florida were determined by combining multiple regions outlined below. Gulf Florida includes the Florida Panhandle and the Western Florida combined region.</p

Magnitude of numbers of stranded common bottlenose dolphins (<i>Tursiops truncatus</i>) above the upper baseline limit (1990–2009) in the northern Gulf of Mexico during the ongoing unusual mortality event (January 2010-June 2013).

<p>From west to east; A) Texas, B) Western Louisiana C) Barataria Bay, Louisiana, D) Northern Louisiana, E) Mississippi, and F) Alabama, G) Florida Panhandle, and H) Western Florida. The solid box marked ‘Pre-spill’ represents time before the <i>Deepwater Horizon</i> oil spill in the northern Gulf of Mexico.</p

Comparison of demographics for common bottlenose dolphins (<i>Tursiops truncatus</i>) stranded in Mississippi and Alabama during shared large-scale mortality years (LSMY) 1990 and 2011.

<p>Louisiana 2011 was compared to Mississippi and Alabama 2011 due to Louisiana’s relative longevity of high stranding rates. The ongoing dolphin Gulf of Mexico (GoM) unusual mortality event (UME) includes years 2010 and 2011.</p><p>Comparison of demographics for common bottlenose dolphins (<i>Tursiops truncatus</i>) stranded in Mississippi and Alabama during shared large-scale mortality years (LSMY) 1990 and 2011.</p

Baseline historical stranding numbers of common bottlenose dolphins (<i>Tursiops truncatus</i>) in the northern Gulf of Mexico, fitted values, and the upper 95% confidence limit from negative-binomial regression model for regions with statewide annual increases during the ongoing northern Gulf of Mexico unusual mortality event; A) Texas, B) Western Louisiana, C) Barataria Bay, Louisiana D) Northern Louisiana, E) Mississippi, F) Alabama, G) Florida Panhandle, H) Western Florida.

<p>Baseline historical stranding numbers of common bottlenose dolphins (<i>Tursiops truncatus</i>) in the northern Gulf of Mexico, fitted values, and the upper 95% confidence limit from negative-binomial regression model for regions with statewide annual increases during the ongoing northern Gulf of Mexico unusual mortality event; A) Texas, B) Western Louisiana, C) Barataria Bay, Louisiana D) Northern Louisiana, E) Mississippi, F) Alabama, G) Florida Panhandle, H) Western Florida.</p