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Context The overproduction of reactive oxygen species (ROS) during in vitro culture of ovarian tissues impairs follicular development and survival. Aims To evaluate the effects of punicalagin on the development and survival of primordial follicles, stromal cell and collagen fibres, as well as on the levels of mRNA for nuclear factor erythroid 2-related factor 2 (NRF2 ), superoxide dismutase 1 (SOD1 ), catalase (CAT ), glutathione peroxidase 1 (GPX1 ) and perirredoxin 6 (PRDX6 ), and activity of antioxidant enzymes in cultured bovine ovarian tissues. Methods Bovine ovarian cortical tissues were cultured for 6days in α-MEM+ alone or with 1.0, 10.0, or 100.0µM punicalagin at 38.5°C with 5% CO2 . Follicle morphology and growth, stromal cell density, and collagen fibres were evaluated by classical histology, while the expression of mRNA was evaluated by real-time PCR. The activity of enzymes was analysed by the Bradford method. Key results Punicalagin improved follicle survival and development, reduced mRNA expression for SOD1 and CAT , but did not influence stromal cells or collagen fibres. Punicalagin (10.0µM) increased the levels of thiol and activity of SOD1, CAT , and GPX1 enzymes. Conclusions Punicalagin (10.0µM) promotes follicle survival and development and activates SOD1, CAT , and GPX1 enzymes in bovine ovarian tissues. Implications Punicalagin improves follicle development and survival in cultured ovarian tissues.
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Catalase , Glutationa Peroxidase GPX1 , Glutationa Peroxidase , Taninos Hidrolisáveis , Folículo Ovariano , Animais , Feminino , Bovinos , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Folículo Ovariano/enzimologia , Taninos Hidrolisáveis/farmacologia , Glutationa Peroxidase/metabolismo , Glutationa Peroxidase/genética , Catalase/metabolismo , Catalase/genética , Ovário/efeitos dos fármacos , Ovário/enzimologia , Ovário/metabolismo , Superóxido Dismutase-1/metabolismo , Superóxido Dismutase-1/genética , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Técnicas de Cultura de Tecidos , Superóxido Dismutase/metabolismoRESUMO
Oxidative stress during in vitro of ovarian tissues has adverse effects on follicle survival. α-pinene is a monoterpenoid molecule with antioxidant activity that has great potential to maintain cell survival in vitro. This study investigated the effect of α-pinene (1.25, 2.5, 5.0, 10.0, or 20.0 µg/mL) on primordial follicle growth and morphology, as well as on stromal cells and collagen fibers in bovine ovarian slices cultured for six days. The effect of α-pinene on transcripts of catalase (CAT), superoxide dismutase (SOD), peroxiredoxin 6 (PRDX6), glutathione peroxidase (GPX1), and nuclear factor erythroid 2-related factor 2 (NRF2) was investigated by real-time PCR. The tissues were processed for histological analysis to evaluate follicular growth, morphology, stromal cell density, and collagen fibers. The results showed that 2.5, 5.0, or 10.0 µg/mL α-pinene increased the percentages of normal follicles but did not influence follicular growth. The α-pinene (10.0 µg/mL) kept the stromal cell density and collagen levels in cultured bovine ovarian tissue like uncultured tissues. Ovarian tissues cultured in control medium had reduced expression of mRNA for NRF2, SOD, CAT, GPX1, and PRDX6, but α-pinene (10.0 µg/mL) increased mRNA levels for NRF2 and PRDX6. In conclusion, 10.0 µg/mL α-pinene improves the follicular survival, preserves stromal cell density and collagen levels, and increases transcripts of NRF2 and PRDX6 after in vitro culture of bovine ovarian tissue.
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Light-Emitting Diodes (LEDs) have been effective light sources in attracting Anopheles mosquitoes, but the broad-spectrum white light, even with a wide-ranging application in lighting, have not been evaluated yet. In this study, the white light was field evaluated against the green one in the light trapping of anopheline mosquitoes by using two non-suction Silva traps and two CDC-type suction light traps. Anopheline mosquitoes were captured for two 21-night periods of collecting (2022 and 2023). In the first period, two LEDs were used per Silva trap, but three were used in the second one to increase the luminance/illuminance at traps. A CDC-type suction light trap equipped with an incandescent lamp was used in 2022 and a CDC-type suction light trap equipped with a 6 V-white light (higher luminance/illuminance) in 2023. A total of eight species and 3,289 specimens were captured in both periods. The most frequent species were Anopheles triannulatus s.l., An. goeldii, An. evansae and An. argyritarsis. In 2022, white LEDs were less attractive to anopheline mosquitoes than the other light sources, but without statistical difference among treatments (F = 2.703; P = 0.0752; df = 2). In 2023, even with an increased luminance/illuminance at traps, no statistical difference was found between the two LED-baited Silva traps (F = 6.690; P = 0.0024; df = 2), but rather between the 6 V-white-baited CDC-type suction light trap and green-baited Silva traps. Due to some drawbacks and the lower abundance of individuals caught by using white LEDs, the narrow-banded green LEDs is preferable to white ones for attracting anophelines.
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Anopheles , Luz , Controle de Mosquitos , Animais , Anopheles/fisiologia , Anopheles/efeitos da radiação , Controle de Mosquitos/métodos , Controle de Mosquitos/instrumentaçãoRESUMO
This study aims to investigate the effects of melatonin on follicular growth, viability and ultrastructure, as well as on the levels of mRNA for antioxidant enzymes, reactive oxygen species (ROS) and meiotic progression in oocytes from in vitro cultured bovine early antral follicles. To this end, isolated early antral follicles (500-600 µm) were cultured in TCM-199+ alone or supplemented with 10-6 , 10-7 or 10-8 M melatonin at 38.5°C with 5% CO2 for 8 days. Follicle diameters were evaluated at days 0, 4 and 8 of culture. At the end of culture, ultrastructure, chromatin configuration, viability (calcein-AM and ethidium homodimer-1 staining), and the levels of ROS and mRNA for catalase (CAT), superoxide dismutase (SOD) and peroxiredoxin 6 (PRDX6) and glutathione peroxidase (GPx) were investigated in oocyte-granulosa cell complexes (OGCs). The results showed that early antral follicles cultured with 10-6 and 10-8 M melatonin had a progressive and significant increase in their diameters throughout the culture period (p < .05). Additionally, oocytes from follicles cultured with 10-7 or 10-8 M melatonin had increased fluorescence for calcein-AM, while those cultured with 10-6 or 10-7 M had reduced fluorescence for ethidium homodimer-1. Different from follicles cultured in other treatments, those cultured with 10-8 M melatonin had well-preserved ultrastructure of oocyte and granulosa cells. Melatonin, however, did not influence the levels of ROS, the mitochondrial activity, oocyte meiotic resumption and expression mRNA for SOD, CAT, GPX1 and PRDX6. In conclusion, the presence of 10-8 M melatonin in culture medium improves viability and preserves the ultrastructure of oocyte and granulosa cells of early antral follicles cultured in vitro.
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Fluoresceínas , Melatonina , Feminino , Animais , Bovinos , Melatonina/farmacologia , Melatonina/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Oócitos , Superóxido Dismutase , RNA Mensageiro/metabolismoRESUMO
The current work assessed the infection with Ehrlichia and Anaplasma species, and exposure to Rickettsia spp. in free-ranging capybaras in the Iberá wetlands ecoregion in Argentina. By indirect immunofluorescence assay, 37 out of 51 (73%) capybara sera were seropositive to Rickettsia spp., with 23.5% and 4% samples considered homologous to Rickettsia parkeri and Rickettsia bellii, respectively (or very closely related serotypes). Anaplasmataceae DNA was found to be highly prevalent in capybaras, with 33 out of 62 samples positive for Anaplasma sp. with Ct values of 28.64 ± 0.35 (average ± standard error), and 12 samples positive for Ehrlichia sp. with Ct values of 31.74 ± 0.87. Anaplasma sp. from capybaras was closely related to Anaplasma sp. reported to infect Amblyomma dubitatum in Iberá wetlands and to Anaplasma odocoilei, while the detected Ehrlichia sp. was closely related to "Candidatus Ehrlichia hydrochoerus" previously reported to infect capybaras in Brazil and A. dubitatum in Iberá wetlands. Structures compatible with Anaplasma morulae were observed in the cytoplasm of platelets from Anaplasma-positive capybaras. Our findings show that capybaras from the Iberá wetlands were exposed to Rickettsia species related to R. bellii and to the pathogen R. parkeri, and were infected with "Ca. Ehrlichia hydrochoerus" and a novel Anaplasma species, herein named "Candidatus Anaplasma capybara".
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Rickettsiaceae , Roedores , Animais , Áreas Alagadas , Argentina , BrasilRESUMO
Phlebotomine sand flies are crepuscular and nocturnal small dipteran insects in the family Psychodidae. Several disease agents, including Leishmania parasites, are transmitted to humans and other vertebrate hosts by the bite of an infected female sand fly. As part of leishmaniasis surveillance programs, light traps have been routinely used in sand fly collections. In this context, new trapping devices are always being required to improve vector monitoring. Here, the efficiency of a new suction light trap, named Silva suction trap or SS trap, was field evaluated in collecting sand flies. Two SS traps, one with green (520 nm, 15,000 mcd) and the other with white (wide spectrum, 18,000 mcd) LEDs, and one CDC-type trap were deployed in a rural forested environment. A total of 4686 phlebotomine sand flies were captured. The most frequent species were females of the Ps. Chagasi series (77.8%) followed by males of Ps. wellcomei (11.6%), Nyssomyia whitmani (3.3%), and Bichromomyia flaviscutellata (2.4%). The CDC-type light trap collected 101.9 ± 20.89 sand flies and 14 species, followed by the white-baited SS trap (87.78 ± 16.36, 14), and the green-baited SS trap (70.61 ± 14.75, 15), but there were no statistically significant differences among traps. A discussion on the considerable advantages of the use of SS traps over CDC traps is included. In this study, the Silva suction trap proved to be efficient and can be an alternative to CDC traps for monitoring adult phlebotomine sand fly populations.
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Leishmania , Leishmaniose , Phlebotomus , Psychodidae , Humanos , Masculino , Animais , Feminino , SucçãoRESUMO
This study investigated the effects of cyclic adenosine monophosphate modulating during cumulus-oocyte complexes (COCs) pre-maturation and the role of melatonin on in vitro maturation (IVM) of bovine COCs. In experiment one, COCs were pre-matured for 8 h in control medium or with 3-isobutyl-1-methylxanthine (IBMX) and forskolin, IBMX and C-type natriuretic peptide, c-type natriuretic peptide and forskolin or IBMX, forskolin and c-type natriuretic peptide. Then, meiotic progression was evaluated. In experiment two, COCs were pre-matured, followed by IVM in control medium alone or with 10-6, 10-7 or 10-8 M melatonin. After IVM, chromatin configuration, transzonal projections (TZPs), reactive oxygen species, mitochondrial distribution, ultrastructure and mRNA expression for antioxidant enzymes were evaluated. In experiment 1, COCs pre-matured with both C-type natriuretic peptide and forskolin or C-type natriuretic peptide, forskolin and IBMX had lower meiotic resumption rate when compared to control. Considering that IBMX had not an additional effect to potentiate inhibition of meiotic resumption, a combination of C-type natriuretic peptide and forskolin was chosen. In experiment 2, COCs matured with 10-8 M melatonin had greater rates of meiotic resumption when compared to the other treatments (P < 0.05). The COCs matured with 10-7 or 10-8 M melatonin had greater mitochondrial activity (P < 0.05), while those matured with 10-6 or 10-8 M of melatonin had greater levels of TZPs. Ultrastructure of oocyte and cumulus cells after IVM with melatonin was relatively well preserved. COCs matured with 10-8 M melatonin increased mRNA expression for superoxide dismutase (SOD) and catalase (CAT) (P < 0.05), when compared to non-cultured and pre-matured COCs, respectively. In conclusion, bovine COC pre-maturation with C-type natriuretic peptide and forskolin, followed by IVM with 10-8 M melatonin improves meiotic resumption rates, TZPs, mitochondrial distribution and mRNA expression for SOD and CAT.
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Melatonina , Animais , Bovinos , Feminino , Melatonina/farmacologia , Melatonina/metabolismo , 1-Metil-3-Isobutilxantina/farmacologia , Técnicas de Maturação in Vitro de Oócitos/veterinária , Peptídeo Natriurético Tipo C/farmacologia , Colforsina/farmacologia , Colforsina/metabolismo , Oócitos/fisiologia , AMP Cíclico/metabolismo , RNA Mensageiro/metabolismo , Superóxido Dismutase/metabolismo , Monofosfato de Adenosina/metabolismo , Monofosfato de Adenosina/farmacologia , Células do CúmuloRESUMO
Opossums are synanthropic marsupials able to interchange among wild, periurban and urban environments, playing an epidemiologically important role as hosts for emerging pathogens and ectoparasites of relevance in public health. The present study aimed to detect and molecularly characterize vector-borne agents in a population of common opossums (Didelphis marsupialis) from the Island of São Luís do Maranhão, northeastern Brazil. Of the 45 animals analyzed, one (2.22%) was positive in the nested PCR assay based on the 18S rRNA gene of piroplasmids. The obtained sequence was phylogenetically positioned in a clade containing sequences of Babesia sp. previously detected in Didelphis aurita, Didelphis albiventris and associated ticks from Brazil. Eight (17.77%) samples were positive in PCR for Ehrlichia spp. based on the dsb gene; four samples were sequenced and positioned into a new clade, sister to E. minasensis and Ehrlichia sp. clade detected in Superorder Xenarthra mammals. No samples tested positive in the screening PCR assays based on the 16S rRNA gene of Anaplasma spp. Two samples were positive in the qPCR for Bartonella spp. based on the nuoG gene. Seven animals (15.56%) were positive in the nPCR based on the 16S rRNA gene of hemoplasmas. Of these, three were positive in a PCR based on the 23S rRNA gene. The phylogenies based on both 16S rRNA and 23S rRNA genes corroborated to each other and positioned the sequences in the same clade of hemoplasmas previously detected in D. aurita and D. albiventris sampled in Brazil. Finally, three (6.66%) animals were positive in the PCR for Hepatozoon spp.; the obtained 18S rRNA sequence was positioned into the H. felis clade.The present study showed, for the first time, the circulation of piroplasmids, Hepatozoon spp., Ehrlichia spp., hemoplasmas and Bartonella spp. in D. marsupialis sampled in northeastern Brazil, with description of putative novel genotypes of Ehrlichia and Hepatozoon and copositivity by different vector-borne agents. The present work consolidates the "South American Marsupialia" piroplasmid clade, adding one more genotype of Babesia sp. to this clade.
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Babesia , Bartonella , Didelphis , Carrapatos , Animais , Brasil/epidemiologia , RNA Ribossômico 16S/genética , Carrapatos/parasitologia , Anaplasma/genética , Ehrlichia/genética , Babesia/genética , Bartonella/genética , MamíferosRESUMO
Ticks are vectors for several pathogens, including bacteria belonging to the Rickettsia genus, such as Rickettsia rickettsii and Rickettsia parkeri, the causative agents of spotted fever. The aim of the present study was to investigate the tick species richness and rickettsial agents associated with wild birds captured in the Humaita Forest Reserve, Acre, in the Western Amazon region. Wild birds were captured with ornithological nets for visual inspection with the purpose of collecting ticks, which were identified through morphological analyses and molecular tests for several genes (12S rDNA, 16S rDNA, gltA, ompA, and sca4). A total of 607 wild birds were captured, 12% of which were parasitized by 268 ticks of the Amblyomma genus, with new host-parasite associations reported for Amblyomma calcaratum, Amblyomma geayi, Amblyomma longirostre, Amblyomma naponense, Amblyomma nodosum, and Amblyomma varium. Of the total ticks collected, 113 were tested for the presence of rickettsial DNA fragments, with 19 testing positive for R. parkeri in A. geayi, Rickettsia tamurae-like in Amblyomma sp., and Rickettsia amblyommatis in A. geayi, A. longirostre, and Amblyomma sp. We detected R. tamurae-like in Amblyomma larvae for the first time in the Western Brazilian Amazon biome, and registered spotted fever group rickettsiae, although the relevance of the detected species in a public health context should be further explored in South America, as well as new host-parasite interactions in this underexplored region.
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Ixodidae , Rickettsia , Rickettsiose do Grupo da Febre Maculosa , Infestações por Carrapato , Carrapatos , Animais , Carrapatos/microbiologia , Ixodidae/microbiologia , Brasil/epidemiologia , Infestações por Carrapato/epidemiologia , Infestações por Carrapato/veterinária , Infestações por Carrapato/parasitologia , Rickettsia/genética , Animais Selvagens , Rickettsiose do Grupo da Febre Maculosa/veterinária , Amblyomma/genética , Aves , DNA RibossômicoRESUMO
Visceral Leishmaniasis is a serious public health problem and dogs are considered to be the main source of infection in urban areas. In Brazil, this disease is present in all regions, but with high concentration of cases in the Northeast, and the state of Maranhão is considered to be an endemic region. The aim of this study was to conduct an epidemiological, spatial, molecular and serological survey on Leishmania infantum among domestic dogs in the municipality of Belágua, Maranhão. Blood samples were collected from dogs and questionnaires were applied to their owners to obtain epidemiological data and risk factors relating to this zoonosis in the region. The coordinates of the dogs' homes were obtained to produce a disease risk map. Serological diagnoses were made using the indirect immunofluorescence reaction (IFAT) and the dual-path platform chromatographic immunoassay test (DPP®) (Bio-Manguinhos/FIOCRUZ, Brazil). A molecular investigation was undertaken using the polymerase chain reaction (PCR). Georeferencing was performed using the global positioning system (GPS) and cases of canine visceral leishmaniasis in the municipality were spatially represented and analyzed using QGIS version 3.16.6 (QGIS Development Team, 2021). A total of 205 blood samples were collected, of which 122 (59.51%) were seroreactive for L. infantum through IFAT, while the DPP test showed 84 reactive samples (40.97%). IFAT and DPP detected 16 positive animals simultaneously. One sample that was seroreactive through IFAT was also positive through PCR. In the clinical evaluation, it was observed that among the seropositive dogs, 112 (91.80%) were symptomatic and 10 (8.20%) were asymptomatic. In the spatial analysis, the Kernel density estimator enabled determination of the place at greatest risk of occurrence of the disease. The areas with the highest concentrations of cases were in districts with large quantities of precarious housing and lack of basic sanitation. This was the first report on the occurrence of L. infantum among dogs in the municipality of Belágua. The results show that canine visceral leishmaniasis is well dispersed in this municipality, thus putting at risk the human population.