Boid Inclusion Body Disease Is Also a Disease of Wild Boa Constrictors.

Reptarenaviruses cause boid inclusion body disease (BIBD), a potentially fatal disease, occurring in captive constrictor snakes boas and pythons worldwide. Classical BIBD, characterized by the formation of pathognomonic cytoplasmic inclusion bodies (IBs), occurs mainly in boas, whereas in pythons, for example, reptarenavirus infection most often manifests as central nervous system signs with limited IB formation. The natural hosts of reptarenaviruses are unknown, although free-ranging/wild constrictor snakes are among the suspects. Here, we report BIBD with reptarenavirus infection in indigenous captive and wild boid snakes in Costa Rica using histology, immunohistology, transmission electron microscopy, and next-generation sequencing (NGS). The snakes studied represented diagnostic postmortem cases of captive and wild-caught snakes since 1989. The results from NGS on archival paraffin blocks confirm that reptarenaviruses were already present in wild boa constrictors in Costa Rica in the 1980s. Continuous sequences that were de novo assembled from the low-quality RNA obtained from paraffin-embedded tissue allowed the identification of a distinct pair of reptarenavirus S and L segments in all studied animals; in most cases, reference assembly could recover almost complete segments. Sampling of three prospective cases in 2018 allowed an examination of fresh blood or tissues and resulted in the identification of additional reptarenavirus segments and hartmanivirus coinfection. Our results show that BIBD is not only a disease of captive snakes but also occurs in indigenous wild constrictor snakes in Costa Rica, suggesting boa constrictors to play a role in natural reptarenavirus circulation. IMPORTANCE The literature describes cases of boid inclusion body disease (BIBD) in captive snakes since the 1970s, and in the 2010s, others and ourselves identified reptarenaviruses as the causative agent. BIBD affects captive snakes globally, but the origin and the natural host of reptarenaviruses remain unknown. In this report, we show BIBD and reptarenavirus infections in two native Costa Rican constrictor snake species, and by studying archival samples, we show that both the viruses and the disease have been present in free-ranging/wild snakes in Costa Rica at least since the 1980s. The diagnosis of BIBD in wild boa constrictors suggests that this species plays a role in the circulation of reptarenaviruses. Additional sample collection and analysis would help to clarify this role further and the possibility of, e.g., vector transmission from an arthropod host.

On the soil-bean-cup relationships in Coffea arabica L.

BACKGROUND: The relationships between soil and coffee beans variables were evaluated and then the influence of bean composition on cup quality attributes was computed by means of relation studies. A total of 139 coffee and soil samples were collected directly from the same number of coffee plantations in Chiapas, Mexico. RESULTS: In the elemental composition, only phosphorus, potassium, calcium, and copper in coffee beans had a significant (P < 0.05) relationship with the content of the same elements in soil. The level of macro- and microelements in the coffee bean affected some of the cup quality attributes, but variables such as texture, titratable acidity, and pH of water in soil had a major influence on those attributes. Caffeine, trigonelline, and 5-caffeoylquinic acid in green coffee beans also had a significant influence (P < 0.05) on the sensory attributes of the beverage. CONCLUSION: The elemental composition of soil and coffee beans was important in explaining the cup quality attributes, but the most important variables influencing the sensory quality of coffee were altitude of plantations and moisture of coffee beans. © 2020 Society of Chemical Industry.

A novel host-adapted strain of Salmonella Typhimurium causes renal disease in olive ridley turtles (Lepidochelys olivacea) in the Pacific.

Salmonella spp. are frequently shed by wildlife including turtles, but S. enterica subsp. enterica serovar Typhimurium or lesions associated with Salmonella are rare in turtles. Between 1996 and 2016, we necropsied 127 apparently healthy pelagic olive ridley turtles (Lepidochelys olivacea) that died from drowning bycatch in fisheries and 44 live or freshly dead stranded turtles from the west coast of North and Central America and Hawaii. Seven percent (9/127) of pelagic and 47% (21/44) of stranded turtles had renal granulomas associated with S. Typhimurium. Stranded animals were 12 times more likely than pelagic animals to have Salmonella-induced nephritis suggesting that Salmonella may have been a contributing cause of stranding. S. Typhimurium was the only Salmonella serovar detected in L. olivacea, and phylogenetic analysis from whole genome sequencing showed that the isolates from L. olivacea formed a single clade distinct from other S. Typhimurium. Molecular clock analysis revealed that this novel clade may have originated as recently as a few decades ago. The phylogenetic lineage leading to this group is enriched for non-synonymous changes within the genomic area of Salmonella pathogenicity island 1 suggesting that these genes are important for host adaptation.

Francisella marina sp. nov., Etiologic Agent of Systemic Disease in Cultured Spotted Rose Snapper (Lutjanus guttatus) in Central America.

Historically, piscine francisellosis in various warm-, temperate-, and cold-water fish hosts has been attributed to Francisella noatunensis From 2015 to 2016, an undescribed Francisella sp. was recovered during mortality events in cultured spotted rose snapper (Lutjanus guttatus) off the Pacific coast of Central America. Despite high mortality and emaciation, limited gross findings were observed in affected fish. Histological examination revealed multifocal granulomatous lesions, with the presence of numerous small, pleomorphic coccobacilli, predominantly in the peritoneum, spleen, kidneys, liver, pancreas, heart, and intestine. Sequencing of an ∼1,400-bp fragment of the 16S rRNA gene demonstrated these isolates to be most similar (99.9% identity) to Francisella sp. isolate TX077308 cultured from seawater in the Gulf of Mexico, while sharing <99% similarity to other Fransicella spp. Biochemical analysis, multilocus sequence comparisons of select housekeeping genes, repetitive extragenic palindromic PCR fingerprinting, matrix-assisted laser desorption ionization-time of flight mass spectrometry, and fatty acid methyl ester analysis revealed marked differences between these isolates and other described members of the genus. Koch's postulates were fulfilled by experimental intracoelomic injection and immersion trials using Nile (Oreochromis niloticus) and blue (Oreochromis aureus) tilapia. Based on observed phenotypic and genotypic differences from recognized Francisella spp., the name Francisellamarina sp. nov. (NRRL B-65518) is proposed to accommodate these novel strains.IMPORTANCE Finfish aquaculture is the fastest growing global food production sector. Infectious disease, particularly emergent pathogens, pose a significant threat to established and nascent aquaculture industries worldwide. Herein, we characterize a novel pathogen isolated from mortality events in cultured spotted rose snapper in Central America. The bacteria recovered from these outbreaks were genetically and phenotypically dissimilar from other known Francisella spp. from fish, representing a previously unrecognized member of the genus Francisella, for which the name Francisella marina sp. nov. is proposed.

Neotropical bats that co-habit with humans function as dead-end hosts for dengue virus.

Several studies have shown Dengue Virus (DENV) nucleic acids and/or antibodies present in Neotropical wildlife including bats, suggesting that some bat species may be susceptible to DENV infection. Here we aim to elucidate the role of house-roosting bats in the DENV transmission cycle. Bats were sampled in households located in high and low dengue incidence regions during rainy and dry seasons in Costa Rica. We captured 318 bats from 12 different species in 29 households. Necropsies were performed in 205 bats to analyze virus presence in heart, lung, spleen, liver, intestine, kidney, and brain tissue. Histopathology studies from all organs showed no significant findings of disease or infection. Sera were analyzed by PRNT90 for a seroprevalence of 21.2% (51/241), and by PCR for 8.8% (28/318) positive bats for DENV RNA. From these 28 bats, 11 intestine samples were analyzed by RT-PCR. Two intestines were DENV RNA positive for the same dengue serotype detected in blood. Viral isolation from all positive organs or blood was unsuccessful. Additionally, viral load analyses in positive blood samples by qRT-PCR showed virus concentrations under the minimal dose required for mosquito infection. Simultaneously, 651 mosquitoes were collected using EVS-CO2 traps and analyzed for DENV and feeding preferences (bat cytochrome b). Only three mosquitoes were found DENV positive and none was positive for bat cytochrome b. Our results suggest an accidental presence of DENV in bats probably caused from oral ingestion of infected mosquitoes. Phylogenetic analyses suggest also a spillover event from humans to bats. Therefore, we conclude that bats in these urban environments do not sustain DENV amplification, they do not have a role as reservoirs, but function as epidemiological dead end hosts for this virus.

Fatal Paratanaisia bragai (Digenea: Eucotylidae) infection in scarlet macaws (Ara macao) in Costa Rica.

We report on four fatal cases of renal infection due to Paratanaisia bragai in scarlet macaws (Ara macao) from two rescue centres in Costa Rica. At necropsy, birds had severe dehydration and cachexia. Two birds had hydropericardium, oedematous lungs, and liver lipidic degeneration. All birds had enlarged kidneys with brown pale discoloration and diffuse white spots on the cortical and sliced surfaces. Ureters were filled with many specimens of P. bragai. Histopathologically, the urinary system revealed multifocal interstitial lymphocytic-plasmacytic nephritis, multifocal mineralization of renal tubules, and interstitial fibrosis associated with flukes. Death of all scarlet macaws was related to severe nephritis leading to chronic renal failure due to P. bragai infection. It is plausible that P. bragai infection of scarlet macaws was accidental due to ingestion of the gastropod intermediate host inside the cages during the rainy season when humidity is higher and gastropods are more active. This represents the second report of parasitism by Eucotylidae digeneans in birds of Psittaciformes and the first in scarlet macaws.

Pathogenic potential of a Costa Rican strain of 'Candidatus Rickettsia amblyommii' in guinea pigs (Cavia porcellus) and protective immunity against Rickettsia rickettsii.

'Candidatus Rickettsia amblyommii' is a spotted fever group rickettsia that is not considered pathogenic, although there is serologic evidence of possible infection in animals and humans. The aim of this study was to evaluate the pathogenic potential of a Costa Rican strain of 'Candidatus R. amblyommii' in guinea pigs and determine its capacity to generate protective immunity against a subsequent infection with a local strain of Rickettsia rickettsii isolated from a human case. Six guinea pigs were inoculated with 'Candidatus R. amblyommii' strain 9-CC-3-1 and two controls with cell culture medium. Health status was evaluated, and necropsies were executed at days 2, 4, and 13. Blood and tissues were processed by PCR to detect the gltA gene, and end titers of anti-'Candidatus R. amblyommii' IgG were determined by indirect immunofluorescence. To evaluate protective immunity, another 5 guinea pigs were infected with 'Candidatus R. amblyommii' (IGPs). After 4 weeks, these 5 IGPs and 3 controls (CGPs) were inoculated with pathogenic R. rickettsii. Clinical signs and titers of anti-Rickettsia IgG were determined. IgG titers reached 1:512 at day 13 post-infection with 'Candidatus R. amblyommii'. On day 2 after inoculation, two guinea pigs had enlarged testicles and 'Candidatus R. amblyommii' DNA was detected in testicles. Histopathology confirmed piogranulomatous orchitis with perivascular inflammatory infiltrate in the epididymis. In the protective immunity assay, anti-Rickettsia IgG end titers after R. rickettsii infection were lower in IGPs than in CGPs. IGPs exhibited only transient fever, while CGP showed signs of severe disease and mortality. R. rickettsii was detected in testicles and blood of CGPs. Results show that the strain 9-CC-3-1 of 'Candidatus R. amblyommii' was able to generate pathology and an antibody response in guinea pigs. Moreover, its capacity to generate protective immunity against R. rickettsii may modulate the epidemiology and severity of Rocky Mountain spotted fever in areas where both species circulate.

Influencia del sexo en la susceptibilidad de ratones Swiss a Trypanosoma cruzi / Sex influence on the susceptibility of Swiss mice to Trypanosoma cruzi

Algunos estudios han reportado que los ratones hembras son más resistentes al trypanosoma cruzi que los machos. Para probar lo anterior, se realizo un estudio comparativo con cuatro cepas de T. cruzi de Costa Rica midiendo parámetros como niveles de parasitemia, porcentajes de mortalidad e histopatología, en un total de 240 ratones (160 para parasitemia y mortalidad y 80 para histopatología). Todas las cepas correspondieron al genotipo TcI. En todos los casos los ratones machos resultaron más susceptibles a la infección (P<0,001). Se observaron parasitemias hasta cinco veces más altas en los machos que, en general, mueren antes que las hembras. La cepa Oswaldo fue la que presentó una mayor mortalidad y niveles de parasitemia más altos. El tejido cardiaco fue el más afectado tanto en los ratones machos como en las hembras, con las cepas bolita y capitán el número de nidos en el miocardio fue significativamente mayor en los machos que en las hembras. Una vez concluido el análisis comparativo se eligió la cepa humana GA, para determinar su efecto en ratones machos y hembras castrados, al lado de ratones normales (150 ratones en total). Las hembras normales se mostraron como las más resistentes, con parasitemias menores que las castradas (P<0,001), y los machos normales como los más susceptibles, con parasitemias significativamente más altas (P<0,001) y supervivencias menores (P<0.01), que los ratones castrados. No se observó diferencia significativa en cuanto al número de nidos en los tejidos entre ratones normales y castrados. Se concluye que el sexo tiene influencia en la resistencia a la infección experimental por T. cruzi.

Some authors have reported that female mice are more resistant to Trypanosoma cruzi infections than male mice. In order to test this hypothesis we did a comparative study using four isolates of T. cruzi from Costa Rica in a total of 160 mice, and observed several parameters such as: parasitemia levels, percent mortality and histopathology. All isolates were identified as genotype TcI. Male mice were more susceptible to infection (P<0.001). Parasitemias reached levels up to five times higher in males and shorter survival periods were observed in males than in females. The Oswaldo isolate showed the highest parasitemias and mortality rates. The heart tissue was the most affected in both males and females. In two isolates the number of parasites in the heart was significantly higher in males than in females. In separate experiments, the human GA isolate was selected in order to observe its effect in castrated male and female mice. Non-castrated females were the most resistant, with lower parasitemias than castrated females, and non-castrated males were the most susceptible with higher parasitemias (P<0.001) and lower survival periods than castrated males (P<0.01). There was no significant difference in the number of parasites in tissues between normal and castrated mice. The final conclusion is that there is a marked sex influence in the susceptibility of mice to T. cruzi experimental infections.

Brucella ceti and brucellosis in cetaceans.

Since the first case of brucellosis detected in a dolphin aborted fetus, an increasing number of Brucella ceti isolates has been reported in members of the two suborders of cetaceans: Mysticeti and Odontoceti. Serological surveys have shown that cetacean brucellosis may be distributed worldwide in the oceans. Although all B. ceti isolates have been included within the same species, three different groups have been recognized according to their preferred host, bacteriological properties, and distinct genetic traits: B. ceti dolphin type, B. ceti porpoise type, and B. ceti human type. It seems that B. ceti porpoise type is more closely related to B. ceti human isolates and B. pinnipedialis group, while B. ceti dolphin type seems ancestral to them. Based on comparative phylogenetic analysis, it is feasible that the B. ceti ancestor radiated in a terrestrial artiodactyl host close to the Raoellidae family about 58 million years ago. The more likely mode of transmission of B. ceti seems to be through sexual intercourse, maternal feeding, aborted fetuses, placental tissues, vertical transmission from mother to the fetus or through fish or helminth reservoirs. The B. ceti dolphin and porpoise types seem to display variable virulence in land animal models and low infectivity for humans. However, brucellosis in some dolphins and porpoises has been demonstrated to be a severe chronic disease, displaying significant clinical and pathological signs related to abortions, male infertility, neurobrucellosis, cardiopathies, bone and skin lesions, strandings, and death.

Neurobrucellosis in stranded dolphins, Costa Rica.

Ten striped dolphins, Stenella coeruleoalba, stranded along the Costa Rican Pacific coast, had meningoencephalitis and antibodies against Brucella spp. Brucella ceti was isolated from cerebrospinal fluid of 6 dolphins and 1 fetus. S. coeruleoalba constitutes a highly susceptible host and a potential reservoir for B. ceti transmission.

Renal oxalosis in free-ranging green turtles Chelonia mydas.

Eighteen green turtles Chelonia mydas recovered from the Atlantic and Gulf coasts of Florida and Tortuguero National Park, Costa Rica, were diagnosed with renal oxalosis by histopathological examination. Affected sea turtles included 14 adults and 4 immature animals, which comprised 26% (18/69) of green turtle necropsy cases available for review. Calcium oxalate deposition ranged from small to moderate amounts and was associated with granuloma formation and destruction of renal tubules. All affected turtles died from traumatic events or health problems unrelated to renal oxalosis; however, 1 immature turtle had notable associated renal injury. Crystal composition was confirmed by infrared and scanning electron microscopy and energy dispersive X-ray analysis. The source of calcium oxalate is unknown and is presumed to be of dietary origin.

Digenetic trematode community in nesting green sea turtles (Chelonia mydas) from Tortuguero National Park, Costa Rica.

The digenetic community of 40 green sea turtles (Chelonia mydas) was investigated at Tortuguero National Park, Costa Rica. In total, 24,270 trematodes belonging to 29 species and 6 families including Clinostomidae 1, Microscaphidiidae 5, Paramphistomidae 2, Pronocephalidae 15, Rhytidodidae 2, and Spirorchiidae 4 were recorded from chelonians examined. Turtles harbored a mean of 7.63 +/- 3.5 SD species. Only 3 species, i.e., Learedius learedi, Microscaphidium reticulare, and Pyelosomum cochlear, infected more than 50% of the hosts sampled. Learedius learedi was the most prevalent (97.5%) and the second most abundant species with a mean of 25.6 +/- 21.6 SD. Only 1 core species, M. reticulare, was recovered from the gastrointestinal tract; it was the most abundant parasite, with a mean intensity of 477 +/- 1,180 SD, and the second most prevalent (77.5%). Diversity values ranged from 0.10 to 2.10, with a mean of 1.00 +/- 0.43 SD for the total component community and from 0.10 to 1.84, with a mean of 0.79 +/- 0.41 SD, for the gastrointestinal component community. Species richness was the highest recorded from a sea turtle species. All digenetic species were recorded from Costa Rica for the first time. This represents the first report on the helminth community of the green turtle.