RESUMO: O objetivo do presente trabalho foi avaliar e diagnosticar alterações do sistema reprodutor de ovelhas abatidas em abatedouros frigoríficos no estado da Bahia. A pesquisa foi realizada em abatedouros frigoríficos do estado da Bahia com Serviços de Inspeção Federal e Estadual no abate de ovinos. No período de julho de 2018 a fevereiro de 2019 foram realizadas visitas e acompanhamento do abate de 1.072 ovelhas. Os animais eram provenientes de 22 municípios do estado da Bahia, com idade entre 6 a 18 meses. Durante o abate, na evisceração foram seccionados os sistemas reprodutores das ovelhas para avaliação e coleta das lesões. Adicionalmente foram obtidos inquéritos epidemiológicos relacionados à procedência, idade e raça. Para o exame bacteriológico, as coletas foram realizadas com lâminas de bisturi estéreis e swabs em tubos estéreis com meio Stuart e refrigeradas em caixa térmica. Para análise histopatológica, os fragmentos foram fixados em formol a 10% e processados de forma rotineira para histologia e corados pela hematoxilina e eosina (HE). Entre 1.072 sistemas genitais avaliados, identificou-se 211 alterações. Os órgãos do sistema reprodutor mais acometidos foram os ovários (65,3%), útero (29,4%) e tubas uterinas (5,3%). Nos ovários, a lesão mais frequente foi o cisto folicular (34,1%); no útero a endometrite (9%) e nas tubas uterinas, os cistos representaram (3,3%). Outras lesões identificadas nos ovários foram: cisto luteinizado (2,3%); tumor de células da granulosa (0,5%); adenoma (0,5%); agenesia unilateral (0,5%), além de outras alterações de pouco significado clínico, como cistos paraovarianos (15,6%) e corpo lúteo cístico (11,8%). Nas tubas uterinas observou-se, além dos cistos tubo-ovarianos (3,3%), hidrossalpinge (1%), adenoma (0,5%) e agenesia (0,5%). As lesões uterinas foram endometrite (9%), hidrometra (2,3%); hiperplasia endometrial cística, abscesso e piometra (1,4% cada); adenomiose, petéquias na serosa do útero e aplasia segmentar (1% cada); dois úteros gravídicos, em início de gestação, apresentaram piometra (0,9%), porém os fetos não apresentaram alterações; e pólipo uterino (0,5%). Observou-se uma gestação ectópica com maceração fetal (0,5%); a vaginite ocorreu em 0,5%, e melanose endometrial em 8,5%. Destaca-se a elevada incidência de cistos foliculares e endometrite que são doenças que reduzem a eficiência reprodutiva dos rebanhos, provocando infertilidade e perdas econômicas na produção.

ABSTRACT: The present study aimed to evaluate and diagnose pathologies of the reproductive system of slaughtered sheep in slaughterhouses in the state of Bahia since there are few data on the subject as well as it is important to note the diagnosis of these diseases to apply appropriate control and prophylaxis measures. The research was carried out in slaughterhouses in the state of Bahia according to the “Serviço de Inspeção Federal” and “Serviço de Inspeção Estadual” for sheep slaughter. From July 2018 to February 2019, visits and monitoring of 1,072 slaughtered sheep were carried out. The animals came from 22 municipalities in the state of Bahia, aged from six to 18 months. During slaughter, the sheep reproductive systems were sectioned for evisceration and lesion collection. Additionally, epidemiological surveys related to origin, age, and race were obtained. For bacteriological examination, collections were performed with sterile scalpel slides and swabs in Stuart medium sterile tubes and refrigerated in a thermal box. For histopathological analysis, fragments were fixed in 10% formaldehyde and routinely processed for histology, stained with Hematoxylin and Eosin (HE). A study of the sheep’s main pathological changes in slaughterhouses in the state of Bahia was carried out, in which 1.072 genital systems were evaluated, and 211 alterations were identified. The most affected reproductive organs were the ovaries (65.3%), followed by the uterus (29.4%) and uterine tubes (5.3%). In the ovaries, the most frequent lesion was a follicular cyst (34.1%); in the fallopian tubes, the cysts represented 3.3% of the lesions, and in the womb, endometritis was observed in 9% of the animals. Other pathologies identified in the ovaries were: luteinized cyst (2.3%); cystic granulosa cell tumor (0.5%); benign lesion (0.5%); agenesis unilateral (0.5%), in addition to other changes of little clinical significance, such as corpus luteum cysts (11.8%) and paraovarian cysts (15.6%). In the fallopian tubes, hydrosalpinx was observed (1%), as well as adenoma (0.5%), agenesis (0.5%), and cysts (3.3%). Uterine lesions included hydrometra (2.3%); Cystic Endometrial Hyperplasia (CEH), abscess, and pyometra (1.4% each); adenomyosis, womb sera petechiae, and total segmental aplasia (1% each); two pregnant uterus, in early pregnancy, presented pyometra (0.9%), however, no change was observed in the fetuses; and uterine polyp (0.5%). Ectopic pregnancy with fetal maceration (0.5%) was observed; vaginitis occurred in 0.5% of the animals, and endometrial melanosis in 8.5%. The high incidence of follicular cysts and endometritis are characterized as diseases that reduce the reproductive efficiency in herds, causing infertility and economic losses in production.

RESUMO Com o objetivo de estudar o efeito da condroitinase associada às células-tronco mesenquimais na lesão aguda da medula espinhal, utilizaram-se 50 ratos Lewis, distribuídos igualmente nos grupos: controle negativo (CN), tratamento com placebo (PLA), condroitinase (CDN), células-tronco mesenquimais (CTM) e condroitinase mais células-tronco mesenquimais (CDN+CTM). Todos os animais tiveram a medula espinhal exposta por laminectomia, e os grupos PLA, CDT, CTM e CDT+CTM sofreram também trauma medular compressivo. Após sete dias, procedeu-se à reexposição da medula espinhal, quando os grupos PLA e CTM receberam 4µL de líquido cefalorraquidiano artificial via intralesional, e os grupos CDT e CDT+CTM receberam o mesmo líquido contendo 2,2U de condroitinase. Após 14 dias da cirurgia inicial, todos os animais receberam 0,2mL de PBS via endovenosa, contudo, nos grupos CTM e CDT+CTM, esse líquido continha 1x106 CTM. Avaliou-se a capacidade motora até o 28o dia pós-trauma e, posteriormente, as medulas espinhais foram analisadas por RT-PCR, para quantificação da expressão gênica para BDNF, NT-3, VEGF, KDR e PECAM-1, e por imunoistoquímica, para detecção das células-tronco GFP injetadas (anti-GFP), quantificação dos neurônios (anti-NeuN) e da GFAP e vimentina, para avaliação da cicatriz glial. As análises estatísticas foram realizadas com o auxílio do Prism 5 for Windows, com o nível de significância de 5%. Não houve diferença entre os grupos quanto à capacidade motora. O grupo CDT+CTM apresentou maior imunoexpressão de neurônios viáveis do que o placebo. No CTM, houve maior expressão dos fatores neurotróficos BDNF e VEGF. E no CDT, houve menor imunoexpressão de vimentina. Concluiu-se que a associação CDT+CTM favorece a viabilidade neuronal após o trauma, que o tratamento com CTM promove aumento na expressão dos fatores tróficos BDNF e VEGF e que o tratamento com condroitinase é efetivo na redução da cicatriz glial.

ABSTRACT The aim of this work was to study the effect of chondroitinase associated with mesenchymal stem cells in acute spinal cord injury. Therefore, 50 Lewis rats were distributed in the following groups: negative control (NC), treatment with placebo (PLA), chondroitinase (CDT), mesenchymal stem cells (MSC), and chondroitinase associated with mesenchymal stem cells (CDT + MSC). All animals had their spinal cord exposed by laminectomy, and the groups named PLA, CDT, MSC and CDT + MSC also suffered compressive spinal cord trauma. After seven days, the spinal cord was re-exposed, when the PLA and MSCs groups received 4uL of artificial cerebrospinal fluid through the lesion, and the CDT group and CDT + MSC received the same fluid containing 2,2U of chondroitinase. 14 days after the first surgery, all animals received 0.2ml of PBS intravenously; however, the MSC and CDT + MSC groups received the same liquid also containing 1x106 MSCs. The motor skills were evaluated up to 28 days post-injury and, subsequently, the spinal cords were analyzed by RT-PCR for BDNF, NT-3, VEGF, PECAM-1 and KDR gene expression quantification, immunohistochemistry to detect injected stem cells GFP (anti-GFP), to quantify neurons (anti-NeuN), GFAP and detect vimentin in order to evaluate the glial scar. Statistical analyzes were performed by Prism 5 for Windows using a 5% level of significance. There was no difference between groups with regarding motor capacity. The CDT + MSC group showed increased immunoreactivity of viable neurons than placebo. In MSC, there was a greater expression of neurotrophic factors BDNF and VEGF. Also, there was less vimentin immunostaining in group CDT. It was concluded that CDT + MSC association promotes neuronal viability after trauma, in which treatment with MSC promotes increased expression of BDNF and VEGF trophic factors, and also that treatment with chondroitinase is effective in reducing the glial scar.

RESUMO OBJETIVOS: Verificar a prevalência de níveis séricos elevados de fibrinogênio e sua associação com os fatores de risco cardiometabólicos em crianças e adolescentes com sobrepeso ou obesidade. MÉTODOS: Estudo transversal com 138 crianças e adolescentes (obesos ou com sobrepeso) acompanhados em um ambulatório de referência da rede pública. A concentração do fibrinogênio foi distribuída em quartis e considerada elevada quando os valores eram iguais ou superiores ao terceiro quartil. A associação entre o valor elevado do fibrinogênio com os fatores de risco cardiometabólicos foi verificada com o teste qui-quadrado de Pearson ou o teste exato de Fisher, quando necessário. A regressão logística foi usada para ajuste das variáveis preditoras do nível do fibrinogênio. As análises foram feitas no SPSS 22.0 e SAS e considerou-se o intervalo de confiança de 95%. RESULTADOS: Os níveis séricos de fibrinogênio estiveram elevados em 28,3% dos indivíduos e apresentaram associação com a PCR elevada (RP: 2,41; IC95%: 1,30-4,46, p = 0,003) e com a presença de quatro ou mais fatores de risco (RP: 1,78; IC95%: 1,00-3,17; p = 0,042). Após a regressão logística, apenas o PCR elevado continuou associado ao fibrinogênio alterado (RP: 1,32; IC95% 1,09-5,25; p = 0,024). CONCLUSÕES: O aumento do fibrinogênio foi prevalente na população estudada, esteve associado à proteína C reativa ultrassensível e ao número igual ou superior a quatro de fatores de risco cardiovasculares e deve ser incluído na avaliação de indivíduos sob risco.

ABSTRACT OBJECTIVES: To determine the prevalence of increased serum fibrinogen levels and its association with cardiometabolic risk factors in overweight or obese children and adolescents. METHODS: Cross-sectional study with 138 children and adolescents (overweight or obese) followed at a reference outpatient clinic of the public health care network. Fibrinogen concentration was divided into quartiles, and values above or equal to the third quartile were considered high. The association between high fibrinogen values and cardiometabolic risk factors was assessed using Pearson's chi-squared test or Fisher's exact test, as necessary. Logistic regression was used to adjust variables predictive of fibrinogen levels. Analyses were performed using SPSS version 22.0 and SAS software, considering a confidence interval of 95%. RESULTS: Serum fibrinogen levels were elevated in 28.3% of individuals, showing association with the presence of high CRP (p = 0.003, PR: 2.41, 95% CI: 1.30-4.46) and the presence of four or more risk factors (p = 0.042; PR: 1.78, 95% CI: 1.00-3.17). After a logistic regression, only elevated CRP remained associated with altered fibrinogen levels (p = 0.024; PR: 1.32; 95% CI: 1.09-5.25). CONCLUSIONS: Increased fibrinogen was prevalent in the study population and was associated with ultrasensitive C-reactive protein and the presence of four or more cardiovascular risk factors; it should be included in the assessment of individuals at risk.

Recognition of pathogens is performed by specific receptors in cells of the innate immune system, which may undergo modulation during the continuum of clinical manifestations of sepsis. Monocytes and neutrophils play a key role in host defense by sensing and destroying microorganisms. This study aimed to evaluate the expression of CD14 receptors on monocytes; CD66b and CXCR2 receptors on neutrophils; and TLR2, TLR4, TLR5, TLR9, and CD11b receptors on both cell types of septic patients. Seventy-seven septic patients (SP) and 40 healthy volunteers (HV) were included in the study, and blood samples were collected on day zero (D0) and after 7 days of therapy (D7). Evaluation of the cellular receptors was carried out by flow cytometry. Expression of CD14 on monocytes and of CD11b and CXCR2 on neutrophils from SP was lower than that from HV. Conversely, expression of TLR5 on monocytes and neutrophils was higher in SP compared with HV. Expression of TLR2 on the surface of neutrophils and that of TLR5 on monocytes and neutrophils of SP was lower at D7 than at D0. In addition, SP who survived showed reduced expression of TLR2 and TLR4 on the surface of neutrophils at D7 compared to D0. Expression of CXCR2 for surviving patients was higher at follow-up compared to baseline. We conclude that expression of recognition and cell signaling receptors is differentially regulated between SP and HV depending on the receptor being evaluated.

Antibodies play an important role in both medicine and analytical biotechnology. Recently, the demand for monoclonal antibodies (mAbs) has been increasing exponentially, since their potential for therapeutic use was disclosed and improved techniques to produce them at higher titers have become available. Although upstream processing of mAbs has suffered a tremendous improvement, in the last decades, me downstream processing lias been considered the bottleneck in providing antibodies at reliable costs, being, in fact, the major cost factor with 50 to 80% of total production costs. Therefore, alternative methods for mAbs's purification may be required. Aqueous two-phase systems (ATPS) have been described, for the last decade, as a viable alternative to the current establish platform, since clarification, concentration and purification can be achieved in just one step using a biocompatible environment. Therefore, the use of ATPS for integrative purification of mAbs it will be reviewed.

Los anticuerpos tienen un papel importante tanto en la biotecnología médica como en la analítica. Desde que su potencial para uso terapéutico fue descubierto y las técnicas para producirlos en concentraciones más altas fueron mejoradas y se hicieron disponibles, la demanda de anticuerpos monoclonales (mAbs) ha estado incrementado exponencialmente. A pesar de que en las últimas décadas los procesos de fermentación de mAbs han mejorado notablemente, los procesos de recuperación y purificación han sido considerados el cuello de botella para la producción de anticuerpos a precios confiables. De hecho, es esta última etapa la que involucra un mayor costo abarcando del 50 al 80% de los costos totales de producción. Debido a esto, métodos alternativos de producción de anticuerpos son requeridos. Los sistemas de dos fases acuosas (SDFA) han sido descritos en la ultima década, como una alternativa viable a la plataforma actual establecida, dado que la clarificación, concentración y purificación pueden ser llevadas a cabo en un solo paso dentro de un ambiente biocompatible. Es por tanto que el uso de SDFA para una purificación integral será revisado.

Avaliaram-se o consumo, a digestibilidade aparente total e parcial dos nutrientes, o pH e a concentração de amônia ruminal em bovinos alimentados com silagem de capim-mombaça e concentrado nas seguintes proporções: 80:20, 65:35, 50:50 e 35:65, com base na matéria seca. Foram utilizados quatro animais Holandês x Zebu, com peso corporal médio inicial de 229kg, canulados no rúmen e abomaso, e distribuídos em quadrado latino 4x4. Os consumos de matéria seca (MS), matéria orgânica (MO), proteína bruta (PB), extrato etéreo (EE) e carboidratos totais (CHOT), expressos em kg/dia, e a digestibilidade parcial dos carboidratos não fibrosos (CNF) apresentaram comportamento linear crescente, com resposta platô nos níveis de concentrado de 54,1; 54,8; 52,9; 62,2; 55,2 e 52,7%. O consumo dos demais nutrientes, exceto da fibra em detergente neutro (FDN), e as digestibilidades aparente total de MS, MO e CNF e a parcial de MO aumentaram linearmente com o incremento do concentrado nas dietas. Não foram encontradas diferenças no consumo e nas digestibilidades aparente total e parcial da FDN. Para concentração de amônia e pH ruminal, observou-se efeito quadrático de tempo de amostragem, com valores máximos de 24,76mg/dL e 6,53 em 2,8 e 3,5 horas após a alimentação, respectivamente.

Intake; total and partial digestibilities of nutrients; and ruminal pH and ammonia concentration were evaluated in beef cattle fed Mombaça-grass silage and concentrate at the following proportions: 80:20, 65:35, 50:50, and 35:65, in the dry matter basis. Four Holstein x Zebu steers, with average live weight of 229kg, fitted with ruminal and abomasal canullae, were used. The animals were allotted in a 4x4 latin square design. The intakes of dry matter (DM), organic matter (OM), crude protein (CP), ether extract (EE), and total carbohydrates (kg/day), as well the partials digestibilities of non fibrous carbohydrates (NFC) increased linearly with plateau at the concentrate levels of 54.1, 54.8, 52.9, 62.2, 55.2, and 52.7%. The intakes of others nutrients, except neutral detergent fiber (NDF), and the total apparent digestibilities of DM, OM, and NFC and the partial digestibility of OM increased linearly as the concentrate levels in the diets increased. No differences were found on the intake and the total and partial apparent digestibilities of NDF. The ruminal pH and ammonia concentration presented a quadratic affect in relation to the time of sampling, with maximum register at 2.8 and 3.5 hours after feeding, respectively, corresponding to pH 6.53 and 24.76mg/dL of ammonia.

Lactococcus lactis, the model lactic acid bacterium, is a good candidate for heterologous protein production in both foodstuffs and the digestive tract. We attempted to produce Streptomyces tendae antifungal protein 1 (Afp1) in L. lactis with the objective of constructing a strain able to limit fungal growth. Since Afp1 activity requires disulfide bond (DSB) formation and since intracellular redox conditions are reportedly unfavorable for DSB formation in prokaryotes, Afp1 was produced as a secreted form. An inducible expression-secretion system was used to drive Afp1 secretion by L. lactis; Afp1 was fused or not with LEISSTCDA, a synthetic propeptide (LEISS) that has been described to be a secretion enhancer. Production of Afp1 alone was not achieved, but production of LEISS-Afp1 was confirmed by Western blot and immunodetection with anti-Afp1 antibodies. This protein (molecular mass: 9.8 kDa) is the smallest non-bacteriocin heterologous protein ever reported to be secreted in L. lactis via the Sec-dependent pathway. However, no anti-fungal activity was detected, even in concentrated samples of induced supernatant. This could be due to a too low secretion yield of Afp1 in L. lactis, to the absence of DSB formation, or to an improper DSB formation involving the additional cysteine residue included in LEISS propeptide. This raises questions about size limits, conformation problems, and protein secretion yields in L. lactis.

Gene vaccines represent a new and promising approach to control infectious diseases, inducing a protective immune response in the appropriate host. Several routes and methods of genetic immunization have been shown to induce antibody production as well as T helper (Th) cell and cytotoxic T lymphocyte activation. However, few studies have compared the nature of the immune responses generated by different gene vaccination delivery systems. In the present study we reviewed some aspects of immunity induced by gene immunization and compared the immune responses produced by intramuscular (im) DNA injection to gene gun-mediated DNA transfer into the skin of BALB/c mice. Using a reporter gene coding for ß-galactosidase, we have demonstrated that im injection raised a predominantly Th1 response with mostly IgG2a anti-ßgal produced, while gene gun immunization induced a mixed Th1/Th2 profile with a balanced production of IgG2a and IgG1 subclasses. Distinct types of immune responses were generated by different methods of gene delivery. These findings have important implications for genetic vaccine design. Firstly, a combination between these two systems may create optimal conditions for the induction of a broad-based immune response. Alternatively, a particular gene vaccine delivery method might be used according to the immune response required for host protection. Here, we describe the characteristics of the immune response induced by gene vaccination and the properties of DNA involved in this process.

DNA-based immunization has initiated a new era of vaccine research. One of the main goals of gene vaccine development is the control of the levels of expression in vivo for efficient immunization. Modifying the vector to modulate expression or immunogenicity is of critical importance for the improvement of DNA vaccines. The most frequently used vectors for genetic immunization are plasmids. In this article, we review some of the main elements relevant to their design such as strong promoter/enhancer region, introns, genes encoding antigens of interest from the pathogen (how to choose and modify them), polyadenylation termination sequence, origin of replication for plasmid production in Escherichia coli, antibiotic resistance gene as selectable marker, convenient cloning sites, and the presence of immunostimulatory sequences (ISS) that can be added to the plasmid to enhance adjuvanticity and to activate the immune system. In this review, the specific modifications that can increase overall expression as well as the potential of DNA-based vaccination are also discussed.