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Antimicrobial Resistance (AMR) poses a global threat to both human health and environmental well-being. Our study delved into Costa Rican wildlife reserves, uncovering a substantial human impact on these ecosystems and underscoring the imperative to pinpoint AMR hotspots. Embracing a One Health perspective, we advocated for a comprehensive landscape analysis that intricately intertwined geographic, climatic, forest, and human factors. This study illuminated the link between laboratory results and observed patterns of antimicrobial use, thereby paving the way for sustainable solutions. Our innovative methodology involved deploying open-ended questions to explore antimicrobial usage across livestock activities, contributing to establishing a comprehensive methodology. Non-invasive sampling in wildlife emerged as a critical aspect, shedding light on areas contaminated by AMR. Feline species, positioned at the apex of the food chain, acted as sentinels for environmental health due to heightened exposure to improperly disposed waste. Regarding laboratory findings, each sample revealed the presence of at least one antimicrobial resistance gene (ARG). Notably, genes encoding resistance to tetracyclines dominated (94.9%), followed by beta-lactams (75.6%), sulfonamides (53.8%), aminoglycosides (51.3%), quinolones (44.9%), phenicols (25.6%), and macrolides (20.5%). Genes encoding polymyxins were not detected. Moreover, 66% of samples carried a multi-resistant microbiome, with 15% exhibiting resistance to three antimicrobial families and 51% to four. The absence of a correlation between forest coverage and ARG presence underscored the profound human impact on wildlife reserves, surpassing previous estimations. This environmental pressure could potentially modify microbiomes and resistomes in unknown ways. As not all antimicrobial families encoding ARGs were utilized by farmers, our next step involved evaluating other human activities to identify the primary sources of contamination. This comprehensive study contributed crucial insights into the intricate dynamics of AMR in natural ecosystems, paving the way for targeted interventions and sustainable coexistence.

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Abstract Introduction: Wildlife hematological patterns are fundamental for health monitoring, and allows elucidating variations both within and between populations. Among these, hematological parameters are particularly valuable to evaluate the health status of neotropical primate species in the wild. Objective: To define hematological reference values for two species of monkeys in Costa Rica. Methods: During 2014, we collected blood samples from free-ranging mantled howler monkeys, Alouatta palliata (17 females, 18 males) and white-faced capuchin monkeys, Cebus imitator (5 females, 7 males) in seven localities of the Costa Rican Pacific coast. Results: For both species, the hematological values were higher in males, and howler monkey populations differed significantly except for platelets. Conclusions: These hematological values, which differ by sex and locality, will help evaluate the health status of these neotropical primate populations.

Resumen Introducción: Los patrones hematológicos de la vida silvestre son fundamentales para el monitoreo de la salud y permiten dilucidar las variaciones tanto dentro como entre poblaciones. Entre estos, los parámetros hematológicos son particularmente valiosos para evaluar el estado de salud de las especies de primates neotropicales en la naturaleza. Objetivo: Definir valores de referencia hematológicos para dos especies de monos en Costa Rica. Métodos: Durante el 2014 recolectamos muestras de sangre de monos aulladores de manto, Alouatta palliata (17 hembras, 18 machos) y monos capuchinos cariblancos, Cebus imitador (5 hembras, 7 machos) en siete localidades de la costa Pacífica de Costa Rica. Resultados: Para ambas especies, los valores hematológicos fueron mayores en los machos, y las poblaciones de monos aulladores difirieron significativamente con excepción de las plaquetas. Conclusiones: Estos valores hematológicos, que difieren según el sexo y la localidad, ayudarán a evaluar el estado de salud de estas poblaciones de primates neotropicales.

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BACKGROUND: In South and Central America, Plasmodium malariae/Plasmodium brasilianum, Plasmodium vivax, Plasmodium simium, and Plasmodium falciparum has been reported in New World primates (NWP). Specifically in Costa Rica, the presence of monkeys positive to P. malariae/P brasilianum has been identified in both captivity and in the wild. The aim of the present study was to determine the presence of P. brasilianum, P. falciparum, and P. vivax, and the potential distribution of these parasites-infecting NWP from Costa Rica. METHODS: The locations with PCR (Polymerase Chain Reaction) positive results and bioclimatic predictors were used to construct ecological niche models based on a modelling environment that uses the Maxent algorithm, named kuenm, capable to manage diverse settings to better estimate the potential distributions and uncertainty indices of the potential distribution. RESULTS: PCR analysis for the Plasmodium presence was conducted in 384 samples of four primates (Howler monkey [n = 130], White-face monkey [n = 132], Squirrel monkey [n = 50], and red spider monkey [n = 72]), from across Costa Rica. Three Plasmodium species were detected in all primate species (P. falciparum, P. malariae/P. brasilianum, and P. vivax). Overall, the infection prevalence was 8.9%, but each Plasmodium species ranged 2.1-3.4%. The niche model approach showed that the Pacific and the Atlantic coastal regions of Costa Rica presented suitable climatic conditions for parasite infections. However, the central pacific coast has a more trustable prediction for malaria in primates. CONCLUSIONS: The results indicate that the regions with higher suitability for Plasmodium transmission in NWP coincide with regions where most human cases have been reported. These regions were also previously identified as areas with high suitability for vector species, suggesting that enzootic and epizootic cycles occur.

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The major histocompatibility complex (MHC) is an important gene complex contributing to adaptive immunity. Studies of platyrrhine MHC have focused on identifying experimental models of immune system function in the equivalent Human Leukocyte Antigen (HLA). These genes have thus been explored primarily in captive platyrrhine individuals from research colonies. However, investigations of standing MHC variation and evolution in wild populations are essential to understanding its role in immunity, sociality and ecology. Capuchins are a promising model group exhibiting the greatest habitat diversity, widest diet breadth and arguably the most social complexity among platyrrhines, together likely resulting in varied immunological challenges. We use high-throughput sequencing to characterize polymorphism in four Class II DR and DQ exons for the first time in seven capuchin species. We find evidence for at least three copies for DQ genes and at least five for DRB, with possible additional unrecovered diversity. Our data also reveal common genotypes that are inherited across our most widely sampled population, Cebus imitator in Sector Santa Rosa, Costa Rica. Notably, phylogenetic analyses reveal that platyrrhine DQA sequences form a monophyletic group to the exclusion of all Catarrhini sequences examined. This result is inconsistent with the trans-species hypothesis for MHC evolution across infraorders in Primates and provides further evidence for the independent origin of current MHC genetic diversity in Platyrrhini. Identical allele sharing across cebid species, and more rarely genera, however, does underscore the complexity of MHC gene evolution and the need for more comprehensive assessments of allelic diversity and genome structure.

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Arthropod-borne viruses belonging to the flavivirus genus possess an enormous relevance in public health. Neotropical non-human primates (NPs) have been proposed to be susceptible to flavivirus infections due to their arboreal and diurnal habits, their genetic similarity to humans, and their relative closeness to humans. However, the only known flavivirus in the American continent maintained by sylvatic cycles involving NPs is yellow fever virus (YFV), and NPs' role as potential hosts of other flaviviruses is still unknown. Here, we examined flavivirus exposure in 86 serum samples including 83.7% samples from free-range and 16.3% from captive NPs living in flavivirus-endemic regions of Costa Rica. Serum samples were opportunistically collected throughout Costa Rica in 2000-2015. We used a highly specific micro-plaque reduction neutralization test (micro-PRNT) to determine the presence of antibodies against YFV, dengue virus 1-4 (DENV), Zika virus, West Nile virus (WNV), and Saint Louis encephalitis virus (SLEV). We found evidence of seropositive NPs with homotypic reactivity to SLEV 11.6% (10/86), DENV 10.5% (9/86), and WNV 2.3% (2/86). Heterotypic reactivity was determined in 3.5% (3/86) of individuals against DENV, 1.2% (1/86) against SLEV, and 1.2% (1/86) against WNV. We found that 13.9% (12/86) of NPs were positive for an undetermined flavivirus species. No antibodies against DENV-3, DENV-4, YFV, or ZIKV were found. This work provides compelling serological evidence of flavivirus exposure in Costa Rican NPs, in particular to DENV, SLEV, and WNV. The range of years of sampling and the region from where positives were detected coincide with those in which peaks of DENV in human populations were registered, suggesting bidirectional exposure due to human-wildlife contact or bridging vectors. Our work suggests the continuous exposure of wildlife populations to various flaviviruses of public health importance and underscores the necessity of further surveillance of flaviviruses at the human-wildlife interface in Central America.

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The apicomplexan parasite Toxoplasma gondii (T. gondii) has been found in more than 350 species of homoeothermic vertebrates in diverse climates and geographic areas. In most animals, T. gondii produces mild or asymptomatic infection. However, acute and hyperacute toxoplasmosis is associated with high mortality rates observed in Neotropical primates (NP) in captivity. These primates are distributed in 20 countries across the Americas, and although infection has been reported in certain countries and species, toxoplasmosis in the wild and its impact on NP population survival is unknown. Differences among species in exposure rates and disease susceptibility may be due in part to differences in host behavior and ecology. Four species of NP are found in Costa Rica, i.e., howler (Alouatta palliata), spider (Ateles geoffroyi), capuchin (Cebus imitator), and squirrel monkeys (Saimiri oerstedii). This study reports NP exposure to T. gondii using the modified agglutination test in 245 serum samples of NP (198 wild and 47 from captivity) from Costa Rica. Associations of serostatus with environmental (forest cover, annual mean temperature), anthropogenic (human population density), and biological (sex) variables in howler and capuchin monkeys were evaluated. The seroprevalence among wild NP was 11.6% (95% CI = 7.7-17.34), compared with 60% in captive monkeys (95% CI = 44.27-73.63), with significant differences between species (X 2 = 20.072; df = 3, p = 0.000164), suggesting an effect of behavior and ecology. In general, antibody titers were low for wild NP (<1:128) and high for captive NP (>1:8192), suggesting higher exposure due to management factors and increased life span in captivity. Seropositivity in howler monkeys was positively related to forest cover and inversely related to annual rainfall. For capuchins, annual rainfall was inversely related to seropositivity. Surveillance of T. gondii exposure in NP in captivity and in the wild is required to understand drivers of the infection and develop novel strategies to protect them.

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Parvoviruses in the genera Bocaparvovirus (HBoV), Erythroparvovirus (B19) and Tetraparvovirus (PARV4) are the only autonomous parvoviruses known to be associated with human and non-human primates based on studies and clinical cases in humans worldwide and non-human primates in Asia and Africa. Here, the presence of these agents with pathogenic potential was assessed by PCR in blood and faeces from 55 howler monkeys, 112 white-face monkeys, 3 squirrel monkeys and 127 spider monkeys in Costa Rica and El Salvador. Overall, 3.7% (11/297) of the monkeys had HboV DNA, 0.67% (2/297) had B19 DNA, and 14.1% (42/297) had PARV4 DNA, representing the first detection of these viruses in New World Primates (NWP). Sex was significantly associated with the presence of HBoV, males having greater risk up to nine times compared with females. Captivity was associated with increased prevalence for PARV4 and when all viruses were analysed together. This study provides compelling molecular evidence of parvoviruses in NWPs and underscores the importance of future research aimed at understanding how these viruses behave in natural environments of the Neotropics and what variables may favour their presence and transmission.

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Fur rubbing or anointing is a well known behavior in capuchin monkeys (Cebus and Sapajus), and may have medicinal and/or social functions. Observations of anointing in capuchins have recorded the application of substances derived from both plants (orange, onion, garlic, citronella, and lemongrass) and animals (ants and millipedes). The present study reports on anointing behavior in free-ranging white-headed capuchins, Cebus capucinus, which involved a commercial insect repellent. After looting a bottle of repellent from the bag of a visitor to the Manuel Antonio National Park in Costa Rica, an adult male bit open the bottle and rubbed the leaking liquid over its entire body, focusing mainly on its belly. Other members of the group rubbed themselves against the male's body and were eventually able to retrieve the bottle of repellent and anoint themselves. The repellent is composed mainly of extracts of eucalyptus and citronella. The capuchins may have been attracted by the strong citric scent of the citronella, which is known to stimulate fur-rubbing behavior in these monkeys. This is reinforced by the fact that the sequence of events was quite distinct from that associated with an earlier event, in which a juvenile male looted, tasted, and then discarded a stick of lip gloss and a tube of sunblock. Overall, the observations indicate that the citric scent of the repellent was attractive to the capuchins, especially in comparison with other man-made substances. As the animals partially ingested all the substances, there is clearly a need for more effective regulation of the contact between animals and visitors in the park.

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Two-hundred-nine free ranging non-human primates from 31 locations throughout Costa Rica were captured and released between 1993 and 2012, and blood samples, sera or plasma were collected, to detect antigens and antibodies, and so assess the distribution of active and passive flavivirus infections over time. A competitive enzyme-linked immunoassay for the detection of antibodies was used to determine the distribution of past flavivirus infections over time, while Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) was used to detect active West Nile Virus (WNV) and Dengue virus (DENV) infections. The first serological evidence of flavivirus in these animals was determined in 1993, at the same time when DENV re-emerged in humans from Costa Rica. An increase in the number of seropositive wild monkeys to flavivirus was determined over time in the country (11.3% seropositivity in 1993-1996, 20.7% in 2001-2008, and finally 52.9% in 2010-2012). Furthermore, the presence of DENV2 was detected in samples from four howler monkeys collected in 2001-2002, whereas DENV2, DENV3, and DENV4 were found in samples from four white-faced monkeys, and WNV in three howler monkeys living in the Pacific coast of Costa Rica during 2010-2012. The habitat where the positive PCR individuals lived were characterized as fragmented forests, having temperatures ranging from 26°C to 28°C, altitudes below 250 meters above sea level, high precipitation during 7 to 9 months (1500-4000 mm), and a marked dry season of 3 to 5 months. All these animals were living near mangroves; however, they did not show clinical signs of illness at the time of sampling. Results obtained show that the number of seropositive wild non-human primates to flavivirus were increasing during time in the country, longitudinal studies are needed to investigate their role as sentinels of these viruses and to determine if flavivirus infections can affect these species.

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La angiostrongilosis abdominal es una parasitosis esporádica, que ocurre en países del neotrópico, especialmente en Costa Rica. Es una enfermedad cuyo diagnóstico es infrecuente y su tratamiento controversial. Se presenta un caso de un adulto que muestra un malestar generalizado, dolor abdominal y una intensa eosinofilia que lleva a la detección de una masa en la fosa iliaca inferior derecha. El diagnóstico definitivo se hace mediante la prueba de Morera y el tratamiento incluye el uso de azitromicina y metronidazol, no obstante no se usan antihelmínticos ni se realiza cirugía. El caso descrito pone de manifiesto que tanto el diagnóstico como el tratamiento de esta enfermedad parasitaria continúan siendo inciertos. No se descarta que exista un subregistro importante de esta parasitosis o que los síntomas sean confundidos con otras patologías.

Abdominal angiostrogyliasis is a sporadic parasitosis that occurs in Neotropical countries, mainly in Costa Rica. It is a disease with an infrequent diagnosis and a controversial treatment. We discuss a case of an adult that presents a severe generalized discomfort, abdominal pain and an intense eosinophilia that leads to the detection of a mass at the right lower quadrant. Definitive diagnosis is achieved by the Morera Test and the treatment includes the use of azythromicin and metronidazole: no antihelmintic drugs or surgery were used. The case shows that both the diagnosis and treatment of this parasitosis are uncertain. An important underreporting of the disease may occur, as well as confusion of the symptoms with other pathologies.