EFFECTS OF DETOMIDINE AND DEXMEDETOMIDINE ON PHYSIOLOGICAL PARAMETERS AND ANESTHETIC RECOVERY IN TAPIRUS TERRESTRIS ANESTHETIZED WITH KETAMINE, GLYCERYL GUAIACOL ETHER, AND MIDAZOLAM.

The aim of this study was to compare the effects of two anesthetic induction protocols for long procedures carried out in the field in Tapiridae. Sixteen tapirs were divided into two groups (n=8) receiving either detomidine (DET) or dexmedetomidine (DEX) for anesthetic induction. All animals were anesthetized by intramuscular administration of a combination of ketamine (1.5 mg/kg), midazolam (0.2 mg/kg), plus either DET (0.04 mg/kg) or DEX (0.007 mg/kg). Anesthetic maintenance was by continuous infusion of ketamine, midazolam, and glyceryl guaiacol ether at 2 mg/kg per hour, 0.1 mg/kg per hour, and 100 mg/kg per hour, respectively). The animals were kept anesthetized for a total of 50 min to allow physical examination and collection of biological material as part of a research program, and physiological variables (heart rate [HR], respiratory rate, oxyhemoglobin saturation [SpO2], rectal temperature [RT], mean arterial pressure [MAP], blood glucose [GLI], and cortisol) and electrocardiogram were recorded during anesthesia. Anesthetic recovery was monitored by two researchers who were not informed of the induction protocol group. The recorded results were statistically evaluated. In both groups there was an initial increase in MAP, which subsequently decreased; RT gradually decreased during anesthesia; HR and GLI increased throughout the procedure; SpO2 was below normal throughout the procedure. Cortisol levels were significantly higher in the DEX group than in the DET group. Also, the animals in the DEX group had a longer recovery time than those in the DET group. On the basis of the results, we conclude that the combination of alpha-2 agonists and midazolam, ketamine, and glyceryl guaicol ether is an appropriate protocol for the anesthesia of tapirs in the field. However, in moderately extended procedures oxygen supplementation is recommended. Additionally, DEX resulted in fewer cardiovascular effects and longer-lasting sedation than DET.

Relationship between SARS-CoV-2 vaccination and cases, program breakthrough cases, and deaths in Dois Vizinhos, Paraná, Brazil.

Several vaccines have been produced in 2 years of the COVID-19 pandemic to control the infection outbreak. This study demonstrated the success of vaccination in controlling COVID-19 cases and deaths in a small city (41 424 people) with a low population density in Brazil. This study was based on a 1-year dataset since the application of the first dose in January 2021. The results show a reduction in positive cases and deaths as the vaccination coverage increased in the city, mainly after 15 000 inhabitants were vaccinated (35.21% of the population) in July 2021. At the time, 49.06% of administered vaccines were ChAdOx1-S recombinant, 39.80% inactivated SARS-CoV-2 virus (CZ02 strain), 9.70% Tozinameran, and 1.44% Ad26.COV2-S recombinant. From August 2021, a marked reduction in daily positive cases and deaths was observed, and incidence (≤2.49 per 1000 inhabitants) and mortality (≤0.02 per 1000 inhabitants) rates remained stabilized until January 2022, when a new outbreak occurred upon the emergence of the Omicron variant. However, the mortality rate (0.07 per 1000 inhabitants) remained low regardless of the Omicron high incidence rate (68.41 per 1000 inhabitants). These data demonstrate the COVID-19 vaccination effectiveness with a threshold of 35.21% of the population vaccinated in this city model.

Placental Development and Physiological Changes in Pregnant Ewes in Silvopastoral and Open Pasture Systems during the Summer.

This study aimed to analyze the reproductive and physiological changes in ewes subjected to heat stress during pregnancy at UTFPR-Brazil. Twenty-four pregnant crossbred ewes were kept in a silvopastoral system (SP) or an open pasture system (OP) throughout the final trimester of pregnancy. Both systems were stressful, but the SP system had lower air temperature than the OP system (26.0 ± 0.38 and 26.9 ± 0.41 °C, respectively; p = 0.0288). Moreover, the radiant thermal load of the two groups presented a difference of 34 Wm-2 (p = 0.0288), and the grass temperature was also lower in the SP system compared to that in the OP system (23.4 ± 0.37 and 25.6 ± 0.44 °C, respectively; p = 0.0043). The respiratory and heart rates of animals from the OP group were higher than those from the SP group (p < 0.001), but no difference was observed in the mobilization of white blood cells (p = 0.4777), and the neutrophil count was only affected by time (p < 0.0001). As regards placental biometry, placentas in twin pregnancies had a greater membrane area (p = 0.0223), but no differences between the systems were observed in placental weight (p = 0.1522) and the number of cotyledons (p = 0.5457). We concluded that the type of rearing system used affects the thermal comfort of pregnant ewes, and that an SP system can offer more amenable microclimatic conditions, which result in greater comfort for the ewes.

Performance of RT-qPCR detection of SARS-CoV-2 in unextracted nasopharyngeal samples using the Seegene AllplexTM 2019-nCoV protocol.

The rapid spread of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has led the world to a pandemic. Therefore, rapid, sensitive, and reproducible diagnostic tests are essential to indicate which measures should be taken during pandemics. We retrospectively tested unextracted nasopharyngeal samples from consecutive patients with suspected SARS-CoV-2 infection (n = 334), and compared two different Ct cut-off values for interpretation of results using a modified Allplex protocol. Its performance was evaluated using the USA Centers for Disease Control and Prevention (CDC) as reference. The reduction on Ct cut-off to 35 increased the test NPA from 79.65 to 88.00 %, reducing the number of false positives, from 10.48 to 6.29 %, resulting in an almost perfect agreement between the Allplex and the CDC protocol (Cohen's Kappa coefficient = 0.830 ± 0.032). This study demonstrates that the Seegene Allplex™ 2019-nCoV protocol skipping the viral RNA extraction step using the Ct cut-off of 35 is a rapid and efficient method to detect SARS-CoV-2 in nasopharyngeal samples.

Trypanosoma cruzi infection in triatomines (Hemiptera: Reduviidae) from rural areas of the state of Pará, Brazil.

The present study aimed to identify the triatomine species and evaluate Trypanosoma cruzi infection in insects captured in endemic areas of Pará State, Brazil. Triatomines were captured in nine rural communities in the municipality of São Domingos do Capim in August, September and December 2014 using active searches and Malaise and Noireau traps. Additionally, from 2014 to 2018, residents and community health agents submitted captured triatomines to the study team. The analysis of T. cruzi infection in the insects was performed by direct parasitological examination and nested-PCR. A total of 225 triatomines were captured and identified: Rhodnius robustus (n = 111), Rhodnius pictipes (n = 54), Panstrongylus geniculatus (n = 44), Eratyrus mucronatus (n = 11), Panstrongylus lignarius (n = 4), and Panstrongylus rufotuberculatus (n = 1). Direct parasitological examination was performed in 27 living triatomines R. robustus (n = 14), P. geniculatus (n = 7) and R. pictipes (n = 6) and metacyclic trypomastigote forms similar to those of T. cruzi were observed in 66.6% (18/27) samples. Of 174 samples analysed by nested-PCR, 81.6% were positive for T. cruzi DNA: R. robustus (84.7%; 72/85), R. pictipes (84.1%; 37/44), P. geniculatus (69.4%; 25/36), P. lignarius (100%; 4/4), E. mucronatus (75%; 3/4) and P. rufotuberculatus (100%; 1/1). R. robustus, R. pictipes and P. geniculatus were the main vectors of T. cruzi in the studied areas; however, the detection of infections in P. lignarius, E. mucronatus and P. rufotuberculatus indicated that these species can also act as potential vectors of T. cruzi in the study areas.

Chagas disease in urban and peri­urban environment in the Amazon: Sentinel hosts, vectors, and the environment.

Chagas disease is an anthropozoonosis, caused by a flagellated protozoan, Trypanosoma cruzi, in which the enzootic cycle occurs between mammals and triatomines. Two dogs with a history of sudden death were necropsied at the Federal University of Pará (UFPA). One dog had a pale area in the myocardium, which on histopathological examination showed a T. cruzi amastigote nest; immunohistochemistry (IHC) analysis characterized it as acute Chagas disease (ACD). The second dog showed no macroscopic changes. Microscopically, a few cardiomyocytes were replaced by adipocytes, and IHC result was negative for T. cruzi. However, results of polymerase chain reaction (PCR) of the cardiac tissue of both dogs was positive for T. cruzi DNA. After that, an epidemiological study was conducted in the region. For this study, we selected four areas in Castanhal. One of the four areas (Area 1) is where one of the dogs lived. The other three areas were chosen because they were recently deforested for housing. Blood samples were collected from dogs, cats, wild small mammals (marsupials and rodents), and the digestive tract of triatomines. Nested PCR was performed on all the blood samples and the triatomine digestive tracts. In Area 1, T. cruzi DNA was detected in 50% (12/24) of the tested dogs, in the only tested cat (1/1), 50% (1/2) of the tested marsupials (Didelphis marsupials), and 100% of the captured triatomines (Rhodnius pictipes) (2/2). In Area 2, T. cruzi DNA was not detected in any of the 11 (0/11) dogs and two marsupials tested (0/2), and no triatomines were found in this area. In Area 3, T. cruzi DNA was detected in 42.25% (30/71) of the dogs, in 66,6% (2/3) of the cats, the only captured marsupial (D. marsupialis) (1/1), and all three triatomines (3/3) (R. pictipes) tested. In Area 4, the two dogs tested were negative (0/2), 25% (1/4) of the captured marsupials (D. marsupialis) was positive, and no triatomine was captured in this area. The data demonstrate the importance of detecting T. cruzi in dogs, cats, small rodents, and marsupials in the Amazon metropolitan areas, where ecotopes carry reservoirs and vectors capable of participating in the Chagas disease cycle. The proximity between humans and T. cruzi vectors in these places might contribute to increased disease transmission risk and maintenance of agents. It was noted that high-standard condominiums, previously thought to reduce the risk for this disease, presented a new epidemiological risk. The presence of T. cruzi DNA in a dog who, a year earlier had tested negative, when another dog in the same house died of ACD, shows that the transmission cycle is present and active, with a high possibility of disease transmission to animals and humans.

Applicability of Raman spectroscopy on porcine parvovirus and porcine circovirus type 2 detection.

Porcine parvovirus (PPV) is one of the major infectious causes of reproductive failure of swine. This disease is characterized by embryonic and fetal infection and death, responsible for important economic losses. PPV is also implicated as a trigger in the development of post-weaning multisystemic wasting syndrome (PMWS) caused by Porcine circovirus type 2 (PCV2). Their detection is PCR-based, which is quite sensitive and specific, but laborious, costly and time-demanding. Therefore, this study aimed to assess Raman spectroscopy (RS) as a diagnostic tool for PPV and PCV2 due to its label-free properties and unique ability to search and identify molecular fingerprints. Briefly, swine testis (ST) cells were inoculated with PPV or PCV2 and in vitro cultured (37 °C, 5% CO2) for four days. Fixed cells were then submitted to RS investigation using a 633 nm laser. A total of 225 spectra centered at 1300 cm-1 was obtained for each sample (5 spectra/cell; 15 cells/replicate; 3 replicates) of PPV-, PCV2-infected and uninfected (control) ST cells. Clear statistical discrimination between samples from both virus-infected cells was achieved with a Principal Component - Linear Discriminant Analysis (PCA-LDA) model, reaching sensitivity rates from 95.55% to 97.77%, respectively to PCV2- and PPV-infected cells. These results were then submitted to a Leave-One-Out (LOO) validation algorithm resulting in 99.97% of accuracy. Extensive band assignment was analyzed and compiled for better understanding of PPV and PCV2 virus-cell interaction, demonstrating that specific protein, lipids and DNA/RNA bands are the most important assignments related to discrimination of virus-infected from uninfected cells. In conclusion, these results represent promising bases for RS application on PCV2 and PPV detection for future diagnostic applications.

Molecular detection of Histoplasma capsulatum in bats of the Amazon biome in Pará state, Brazil.

Histoplasma capsulatum, the fungus causing histoplasmosis, has a strong impact on public health. Histoplasmosis is one of the most prevalent systemic mycoses in the Americas and occurs in several mammalian species. Bats are important in the epidemiological cycle of histoplasmosis because they disseminate the fungus throughout the environment. The aim of the present study was to investigate natural H. capsulatum infection in bats located in forested areas, which have undergone anthropogenic perturbations, as well as in the urban areas of the state of Pará. Twenty-two species of bats were captured in 18 municipalities of Pará; the samples obtained from these animals were subjected to nested PCR for amplification of H. capsulatum DNA. The HCI/HCII and HCIII/HCIV primers were used, and the final 210-pb fragment was amplified. Of the 100 bats analysed, two were confirmed to be positive for H. capsulatum. Samples amplified by nested PCR were sequenced and found to share identity and have 100% match with H. capsulatum DNA. H. capsulatum was detected in the area of study: the state of Pará has a wide diversity of bat species, and the region under investigation is situated in the north of the state, which suffers the most severe environmental and climatic changes. Therefore, it is essential to elucidate the distribution of H. capsulatum hosts in this region to facilitate the implementation of effective disease surveillance.

First report of natural infection by Trypanosoma cruzi in secretions of the scent glands and myocardium of Philander opossum (Marsupialia: Didelphidae): Parasitological and clinicopathological findings.

Trypanosoma cruzi is the etiologic agent of American trypanosomiasis and can infect humans and different species of domestic and wild animals. The marsupials are important wild reservoirs of T. cruzi, aiding in the maintenance of this agent in sylvatic and peri-domestic environments. The objective of this study was to report the parasitological and clinicopathological findings of a natural infection by T. cruzi in one specimen of Philander opossum that originated from the Brazilian Amazon. The animal was captured in a forest fragment near a rural community with reports of human Chagas disease. T. cruzi infection was diagnosed by blood smear examinations, blood culture, scent glands secretion culture, histopathological examination, and nested-PCR. Positive samples were subjected to PCR to characterize the discrete typing units (DTUs) of T. cruzi. Characteristic trypomastigotes of T. cruzi were observed in the blood smear, and spheromastigotes, epimastigotes, and trypomastigotes were visualized in the cultures. Non-suppurative myocarditis associated with amastigote clusters was the principal histopathological finding. DNA from T. cruzi was detected in samples of blood, blood cultures, scent glands secretion cultures, cardiac muscles, and the spleen. The TcI and the TcII/V/VI group DTUs were detected in blood culture and scent glands secretion cultures. Infection by T. cruzi can cause myocarditis in P. opossum and DTUs TcI and TcII/V/VI group mixed infection can be detected in the acute phase. P. opossum can be a source of infection for triatomine vectors and has the potential source for direct transmission of T. cruzi by secretions from the scent glands. These data are important to improve the understanding of the complex enzootic transmission cycle of T. cruzi in the Brazilian Amazon.

Natural Theileria equi infection in captive Tapirus terrestris (Perissodactyla: Tapiridae) in the Brazilian Amazon.

The objective of this study was to detect natural Theileria equi infection in captive tapirs (Tapirus terrestris) in the Brazilian Amazon. Samples from 19 captive tapirs were collected from zoological and botanical gardens and conservation parks in the Pará (n = 18) and Amazonas (n = 1) states. Whole-blood samples were collected for subsequent screening of T. equi DNA by PCR using the BEC-UF2 and EQUI-R primer set. Microscopic analyses of blood smears revealed T. equi trophozoites in 37% (7/19) of the animals examined, and T. equi DNA was detected in 58% (11/19) of the blood samples analyzed. Sequencing of amplified PCR products revealed an identity with T. equi isolates obtained from horses and waterbuck available in GenBank. In conclusion, T. equi infection occurs in captive tapirs in the Brazilian Amazon, and these mammals could potentially act as reservoirs.

Detection of canine parvovirus types 2b and 2c in canine faecal samples contaminating urban thoroughfares in Brazil.

Canine parvovirus type 2 (CPV-2) is a highly contagious virus that causes acute gastroenteritis in dogs all over the world. Because of its stability in the environment, CPV-2 can remain infective for a long time, especially if protected in organic matter. To demonstrate CPV-2's potential as an environmental hazard for nonimmunized susceptible hosts, we investigated 50 faecal samples collected from public areas in a municipality of Paraná state, Brazil. Seven samples tested positive for CPV by a PCR assay targeting the partial VP2 gene, with three strains being confirmed as CPV-2b variant and one as CPV-2c variant by sequence analysis. These findings were supported by phylogenetic analysis, and the species identity of faecal samples source was confirmed by canine mitochondrial DNA amplification and sequencing. Our results demonstrate the presence of CPV in canine faeces contaminating urban thoroughfares and reinforce the importance of environmental control to reduce the potential exposure risks to susceptible hosts.

Plasmodium vivax and Plasmodium falciparum infection in Neotropical primates in the western Amazon, Brazil.

The Brazilian Amazon is endemic for malaria and natural infections by Plasmodium spp. have been detected in Neotropical primates. Despite the diversity of primate species in the region, studies on infections by these agents are limited. The aim of the present study was to investigate the frequency of infection by Plasmodium vivax and P. falciparum in free-born primates that were kept in captivity, in the western Amazon, Brazil. Blood samples were collected from 98 Neotropical primates. Detection of P. vivax and P. falciparum DNA was performed using a semi-nested PCR, and the amplified products were sequenced. Plasmodium spp. DNA was detected in 6.12% (6/98) of the primates. P. vivax, and P. falciparum DNA was detected in 2.04% (2/98) and 4.08% (4/98) of these mammals, respectively. Sequencing and phylogenetic analysis confirmed the results obtained from the semi-nested PCR. The presence of infected non-human primates (NHP) can be auxiliary in the maintenance of P. falciparum and P. vivax and may have implications for the malaria surveillance and control in the Brazilian Amazon. It is necessary to structure an efficient surveillance system for the aetiological agents of malaria that infect NHP and humans to reduce the risk of Plasmodium spp. introduction into new areas, to protect all susceptible species.

Conhecimento sobre o manejo de estomias intestinais de eliminação / Knowledge about the management of intestinal stomas of elimination

Objetivo: verificar o conhecimento do profissional de Enfermagem sobre o cuidado com pacientes com estomias intestinais de eliminação. Método: trata-se de um estudo quantitativo, descritivo e exploratório desenvolvido em um hospital público de urgência. Registra-se que participaram do estudo 30 enfermeiros e 70 técnicos de Enfermagem da clínica cirúrgica que responderam a um questionário sociodemográfico e a um instrumento sobre o levantamento do conhecimento sobre os cuidados a pacientes com estomias intestinais de eliminação. Analisaram-se os dados estatisticamente por meio do programa SPSS® , versão 18.0, para Windows® . Apresentaram-se os resultados em forma de tabelas e figura. Resultados: verificou-se que o conhecimento da equipe de Enfermagem sobre estomias intestinais se encontra fragilizado, constatando uma frequência de acertos inferior a 50,0% nas questões relacionadas ao manejo das estomias intestinais de eliminação, tanto no período pré-operatório, como no pós-operatório. Conclusão: verifica-se que o nível de conhecimento dos profissionais mostrou-se relativamente incipiente, apontando-se a necessidade de promover a capacitação dos profissionais sobre o tema e a realização de novos estudos para avaliar o nível de conhecimento desta categoria.(AU)

Objective: to verify the knowledge of the nursing professional on the care of patients with intestinal stomas of elimination. Method: this is a quantitative study, descriptive and exploratory, developed in a public hospital of urgency. Register who participated in the study 30 nurses and 70 nursing technicians of the surgical clinic, who responded to a sociodemographic questionnaire and an instrument on the survey of knowledge about care of patients with intestinal stomas of elimination. The data were analyzed statistically using the SPSS® program, version 18.0 for Windows® . The results presented in the form of tables and figure. Results: it was found that the knowledge of the nursing team about intestinal stomas is weakened, noting a frequency of less than 50% correct answers on issues related to management of intestinal stomas disposal, both in the preoperative period, as in the postoperative. Conclusion: it appears that the level of knowledge of professionals showed relatively incipient, pointing to the need to promote training of professionals on the subject and completion of new studies to evaluate the level of knowledge of this category.(AU)

Objetivo: verificar los conocimientos de los profesionales de enfermería en la atención a pacientes con estomas intestinales de eliminación. Método: se trata de un estudio cuantitativo, descriptivo y exploratorio desarrollado en un hospital público de urgencia. Registrar que participaron en el estudio 30 enfermeras y 70 técnicos de enfermería de la clínica quirúrgica que respondieron a un cuestionario sociodemográfico y a una herramienta para la elevación de los conocimientos acerca del cuidado de los pacientes con estomas intestinales de eliminación. Los datos fueron analizados estadísticamente mediante el programa SPSS® versión 18.0 para Windows® . Los resultados se presentan en forma de tablas y figura. Resultados: se encontró que el conocimiento del equipo de enfermería acerca de estomas intestinales se debilita, observando una frecuencia de menos de 50% de respuestas correctas acerca de los temas relacionados con la gestión de la eliminación de estomas intestinales, tanto en el preoperatorio como en el postoperatorio. Conclusión: parece que el nivel de conocimiento de los profesionales mostró se relativamente incipiente, apuntando a la necesidad de promover la formación de profesionales en la materia y la realización de nuevos estudios para evaluar el nivel de conocimientos de esta categoría.(AU)

Detection of Toxoplasma gondii in Crassostrea spp. oysters cultured in an estuarine region in eastern Amazon.

The aim of the present study was to detect DNA of Toxoplasma gondii in Crassostrea spp. oysters cultured in the state of Pará, Brazil. A total of 400 oysters were directly collected from a fixed rack system. Gills, gastrointestinal tract (GIT) and intervalvular liquid were separated and grouped into pool samples of 10 animals, resulting in 40 samples each of gills, GIT and intervalvular liquid. DNA extraction was performed using a commercial kit, and T. gondii DNA was detected by nested PCR using the primers Toxo3 and Toxo4, which produced an amplification product of 155 bp of the T. gondii gene B1. Nucleotide sequencing was performed for positive samples, and the obtained sequences were identified by comparison with sequences in GenBank. The DNA of T. gondii was detected in 5.8% (7/120) of the pool samples, of which 7.5% (3/40) was in the GIT, 5% (2/40) in the gills, and 5% (2/40) was in the intervalvular liquid. The obtained sequences presented 100% identity and overlap with T. gondii DNA sequences. This is the report of detection of T. gondii DNA in oysters from genus Crassostrea spp. originating from the state of Pará, eastern Amazon.

Cellular and epigenetic changes induced by heat stress in bovine preimplantation embryos.

Exposure of the preimplantation embryo to heat stress triggers a series of cellular, molecular, and adaptive changes preventing a normal embryonic development. Heat stress disrupts the embryo cytoskeleton, intracellular calcium levels, mitochondrial function, and induces apoptosis. Moreover, heat stress can act indirectly through induction of reactive oxygen species (ROS), leading to a variety of cellular damage. Embryonic resistance to heat shock is determined by factors such as genotype, developmental stage, apoptosis, redox status, and regulatory molecules. The early embryo is very susceptible to heat stress; it acquires resistance to elevated temperature as development advances. One of the mechanisms involved in the developmental acquisition of thermotolerance is heat-induced apoptosis, which acts as a quality control mechanism to remove damaged blastomeres allowing the embryo to survive after stress. Although embryos at >8-cell stage can activate the apoptotic cascade as an adaptive response to stress, embryos at the two-cell stage are resistant to proapoptotic signals. This lack of apoptotic response has been associated to mitochondrial resistance to depolarization and epigenetic regulations, such as DNA methylation and histone deacetylation. Even though the cellular mechanisms triggered by heat stress have been studied, very little attention has been paid to the vulnerability of the epigenome to drastic temperature changes during the preimplantation period. Therefore, this review aims to characterize the effects of elevated temperature on the bovine embryo, especially addresissing developmental, cellular, and epigenetic alterations triggered in response to temperature.

Position- and Hippo signaling-dependent plasticity during lineage segregation in the early mouse embryo.

The segregation of the trophectoderm (TE) from the inner cell mass (ICM) in the mouse blastocyst is determined by position-dependent Hippo signaling. However, the window of responsiveness to Hippo signaling, the exact timing of lineage commitment and the overall relationship between cell commitment and global gene expression changes are still unclear. Single-cell RNA sequencing during lineage segregation revealed that the TE transcriptional profile stabilizes earlier than the ICM and prior to blastocyst formation. Using quantitative Cdx2-eGFP expression as a readout of Hippo signaling activity, we assessed the experimental potential of individual blastomeres based on their level of Cdx2-eGFP expression and correlated potential with gene expression dynamics. We find that TE specification and commitment coincide and occur at the time of transcriptional stabilization, whereas ICM cells still retain the ability to regenerate TE up to the early blastocyst stage. Plasticity of both lineages is coincident with their window of sensitivity to Hippo signaling.

Trypanosoma cruzi infection in captive Neotropical primates in the Brazilian Amazon.

The aim of this study was to detect the infection by Trypanosoma cruzi in captive Neotropical primates in the Brazilian Amazon. From February 2013 to July 2014, 112 blood samples were collected from Neotropical primates from the Amazonas, Amapá, and Pará States, north of Brazil. The subjects belonged to the families Cebidae (N = 59), Atelidae (N = 41), Callitrichidae (N = 5), Pitheciidae (N = 4), and Aotidae (N = 3). Blood smears also were examined for the presence of trypomastigotes by optical microscopy. For the detection of T. cruzi DNA, a Nested-PCR with primers TCZ1/TCZ2 and TCZ3/TCZ4 was performed. T. cruzi DNA was detected in 12.5% (14/112) of Neotropical primates examined. Positive samples were detected in 16%, 12.5%, and 11.11% of the different species of primates sampled from the Amapá, Pará, and Amazonas states, respectively. The analysis of the blood smears did not reveal trypomastigote forms of T. cruzi. In conclusion, Neotropical primates kept in captivity were infected by T. cruzi in the studied areas. We recommend that a health management protocol be put into place to prevent the transmission of infectious agents among captive populations, captive and wild populations, and between NHPs and the technicians who handle these animals.

In vitro culture of ovine mammary gland cells expressing beta-lactoglobulin and beta-casein / Células da glândula mamária ovina cultivadas in vitro expressam beta-lactoglobulina e beta-caseina

The expression of milk proteins in vitro is essential to exploit the mammary gland cells as a biological model. Enzymatic tissue disaggregation has been widely used to establish mammary cell culture, but its effect in long-term ovine mammary cell culture is not completely elucidated. This study aimed at comparing mechanical/enzymatic and mechanical dissociation methods to establish ovine mammary cell culture. We compared cellular differentiation induced by lactating ewe serum or fetal bovine serum based on the gene expression levels of milk proteins (beta-lactoglobulin, alpha s1-casein, and betacasein). Mechanically dissociated cells were positive immunostaining for cytokeratin 8.13, such as mammary epithelial cells. These cells are responsible for milk protein expression and they are low immunostaining for vimentin, mesenchymal marker. Mechanical/enzymatic dissociation cells were positive for vimentin. The fastest cell growth (cell/hour) was observed in the mechanical dissociation group cultured with 10% fetal bovine serum medium. Mechanically and mechanically/enzymatically derived cells were able to express beta-casein and beta-lactoglobulin, but not alpha s1-casein. The relative expression of beta-lactoglobulin was not affected by the tissue dissociation method or culture media, beta-casein relative expression was down regulated in mechanically dissociated cells cultured in the presence of lactating ewe serum, (P = 0.019). Beta-casein relative expression was also down regulated in mechanically/enzymatically dissociated cells cultured with fetal bovine serum (P = 0.021). In the present conditions, we conclude that mechanical dissociation followed by culture with 10% of fetal bovine serum was the most efficient method to induce milk proteins' mRNA expression by ovine mammary epithelial cells in vitro.(AU)

A expressão in vitro de proteínas do leite é essencial para explorar as células da glândula mamária como um modelo biológico. A desagregação tecidual via enzimática é amplamente utilizada para o estabelecimento cultivo de células mamárias. No entanto, seu efeito a longo prazo no cultivo de células da glândula mamária ovina ainda não é bem elucidado. Este estudo tem como objetivo comparar dois métodos de dissociação tecidual, mecânico/enzimático e mecânico, para estabelecer cultivo celular de glândula mamária ovina. A indução da diferenciação celular, por adição de soro de ovelha lactante ou soro fetal bovino, foi avaliada pelos níveis de expressão de proteínas do leite (beta-lactoglobulina, alpha s1-caseína e beta-caseína). Células mecanicamente dissociadas foram positivamente marcadas para a presença de citoqueratina 8.13, marcador para células epiteliais mamárias. Essas células são as responsáveis pela produção das proteínas do leite e são pouco marcadas para a presença de vimentina, marcador para células de origem mesenquimal. Já as células obtidas da dissociação mecânica/ enzimática foram positivamente marcadas para presença de vimentina. A maior velocidade de crescimento (células/hora) foi observado para o grupo com dissociação mecânica cultivado em meio com 10% de soro fetal bovino. As células obtidas tanto da dissociação mecânica quanto mecânica/enzimática foram capazes de expressar beta-lactoglobulina e beta-caseína, mas não alfa s1-caseína. A expressão relativa de beta-lactoglobulina não foi afetada pelo método de dissociação ou meio de cultivo. A expressão relativa da beta-caseína foi negativamente regulada para células mecanicamente dissociadas e cultivadas na presença de soro de ovelha lactante (P = 0,019). A expressão relativa da beta-caseína também foi negativamente regulada para células dissociadas de forma mecânica/enzimática e cultivadas com soro fetal bovino (P = 0,021). Nas condições do presente estudo, concluímos que o método de dissociação mecânica seguido pelo cultivo em meio com 10% de soro fetal bovino foi o método mais eficiente para induzir a expressão mRNA de proteínas do leite por células epiteliais mamárias ovinas in vitro.(AU)

Effect of age on expression of spermatogonial markers in bovine testis and isolated cells.

Spermatogonial stem cells (SSC) are the most undifferentiated germ cell present in adult male testes and, it is responsible to maintain the spermatogenesis. Age has a negative effect over stem cell, but the aging effect on SSC is not elucidated for bovine. The present study aim to evaluate the effect of age on the expression of undifferentiated spermatogonial markers in testis and in enriched testicular cells from prepubertal calves and adult bulls. In this matter, testicular parenchyma from calves (3-5 months) (n=5) and bulls with 3 years of age (n=5) were minced and, isolated cells were obtained after two enzymatic digestions. Differential platting was performed for two hours onto BSA coated dish. Cell viability was assessed by Trypan Blue solution exclusion method and testicular cells enriched for SSC was evaluated by expression of specific molecular markers by qRT-PCR (POU5F1, GDNF, CXCR4, UCHL1, ST3GAL, SELP, ICAM1 and ITGA6) and flow cytometry (GFRA1, CXCR4 and ITGA6). CXCR4 and UCHL1 expression was evaluated in fixated testes by immunohistochemistry. We observed that age just affected the expression of selective genes [SELP (Fold Change=5.61; p=0.0023) and UCHL1 (Fold Change=4.98; p=0.0127)]. By flow cytometry, age affected only the proportion of ITGA6+ cells (P<0.001), which was higher in prepubertal calves when compared to adult bulls. In situ, we observed an effect of age on the number of UCHL1+ (p=0.0006) and CXCR4+ (p=0.0139) cells per seminiferous tubule. At conclusion, age affects gene expression and the population of cells expressing specific spermatogonial markers in the bovine testis.