Thirteen reference genes were investigated to determine their stability to be used as a housekeeping in gene expression studies in skeletal muscle of chickens. Five different algorithms were used for ranking of reference genes and results suggested that individual rankings of the genes differed among them. The stability of the expression of reference genes were validated using samples obtained from the Pectoralis major muscle in chicken. Samples were obtained from chickens in different development periods post hatch and under different nutritional diets. For gene expression calculation the ΔΔCt approach was applied to compare relative expression of pairs of genes within each of 52 samples when normalized to mitochondrially encoded cytochrome c oxidase II (MT-CO2) target gene. Our findings showed that hydroxymethylbilane synthase (HMBS) and hypoxanthine phosphoribosyl transferase 1 (HPRT1) are the most stable reference genes while transferrin receptor (TFRC) and beta-2-microglobulin (B2M) ranked as the least stable genes in the Pectoralis major muscle of chickens. Moreover, our results revealed that HMBS and HPRT1 gene expression did not change due to dietary variations and thus it is recommended for accurate normalization of RT-qPCR data in chicken Pectoralis major muscle.

Quantitative RT-PCR is an important technique for assessing gene expression. However, a proper normalization of reference genes prior to expression analyses of target genes is necessary. The best normalizer is that gene which remains stable in all samples from different treatments. The aim of this study was to identify stable reference genes for normalization of target genes in muscle tissue from three genetically divergent chickens groups (Peloco, Cobb 500® and Caneluda) under environmental (heat stress and comfort) and sex influence. Expressions of ten reference genes were tested for stability in breast muscular tissue (Pectoralis major muscle). Samples were obtained from 36 males and females of two backyard breeds (Caneluda and Peloco) and one commercial line (Cobb 500®) under two environments. The heat stress and comfort temperature were 39 and 23°C, respectively. Animals were housed in the Animal Science Department at Universidade Estadual do Sudoeste da Bahia. We analyzed the expression data by four statistical tools (SLqPCR, NormFinder, Bestkeeper and Comparative CT). According to these tools, genes stability varied according to sex, genetic group and environment, however, some genes remained stable in all analyzes. There was no difference between the most stable genes for sex effect, being MRPS27 more stable for both males and females. In general, MRPS27 was the most stable gene. Within the three genetic groups, the most stable genes were RPL5, HMBS and EEF1 to Cobb 500®, Peloco and Caneluda, respectively. Within the environment, the most stable gene under comfort and heat stress conditions was HMBS and MRPS27, respectively. BestKeeper and Comparative Ct were less correlated (28%) and SLqPCR and NormFinder were the most correlated (98%). MRPS27, RPL5 and MRPS30 genes were considered stable according the overall ranking and can be used as normalizer of relative expression of target genes in muscle tissue of chickens under heat stress.

ABSTRACT. Leishmaniasis is an endemic disease present in 98 countries. In Brazil, the northeast region accounts for approximately half of the cases in humans, and has experienced an increased number of positive cases in dogs. In this study, we investigated the epidemiology of canine leishmaniasis in the city of Ilhéus, Bahia, using serological and molecular techniques and evaluated the possible environmental risk factors and associated clinical signs. Blood samples were collected from 560 dogs in urban and peri-urban areas in Ilhéus, northeastern Brazil. Genomic DNA was extracted from the selected animals and 12063 Leishmania (Viannia) braziliensis in dogs ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (4): 12062-12073 (2015) subjected to molecular analysis using Leishmania species-specific primers and diagnosis of Trypanosoma cruzi. A total of 54.72% of dogs were positive for Leishmania braziliensis, and animals positive for both Leishmania infantum and T. cruzi were not identified. Hematologic variables were not statistically associated with cases of L. braziliensis. However, the positive animal group showed lower red blood cell and platelet counts and higher levels of urea and serum creatinine. Few dogs presented clinical signs compatible with the presence of Leishmania. Age of more than 2 years and specific hair colors were associated with positive results for L. braziliensis. The geoclimatic characteristics of the region may improve parasite survival, reproduction, and vectors. This may explain the higher rate of dogs identified as positive in this study.

BACKGROUND Sepsis is a set of serious organic manifestations caused by an infection, whose progression culminates in exacerbated inflammation and oxidative stress, poor prognosis, and high hospital costs. Antioxidants used against sepsis have been evaluated, including essential oils such as β‐caryophyllene (BCP), and polyunsaturated fatty acids, such as docosahexaenoic acid (DHA). The aim of this study was to evaluate the anti‐inflammatory activity of the association of these two compounds. RESULTS Treatment with BCP‐DHA, at a dose of 200 μL/animal, significantly inhibited the migration of neutrophils in a Cg‐induced peritonitis model. After Staphylococcus aureus infection, in the groups treated with BCP‐DHA there was a significant decrease in the total and differential count of leukocytes, increased expression of cytokines TNF‐α and IFN‐γ in treated groups, an increase of IL‐4 and IL‐5 in B/D and B/D + SA groups, and an augmentation of IL‐6 and IL‐12 groups in B/D + SA groups. Histological and bacterial analysis revealed lower neutrophil migration and lower bacterial load in the infected and treated groups. CONCLUSION In general, the BCP‐DHA association presented anti‐inflammatory activity against two different models of acute inflammation and infection, showing promising potential as a therapeutic adjuvant in sepsis. © 2019 Society of Chemical Industry

Abstract Spontaneous abortion is considered a public health problem having several causes, including infections. Among the infectious agents, bacteria of the vaginal microbiota and Ureaplasma parvum have been associated with abortion, but their participation needs to be further elucidated. This study aims to evaluate the influence of Mollicutes on the development of spontaneous abortion. Women who underwent spontaneous abortion and those with normal birth (control) were studied. Samples of cervical mucus (CM) and placental tissue were collected to identify Mollicutes using the quantitative polymerase chain reaction methodology. Eighty-nine women who had a miscarriage and 20 women with normal pregnancies were studied. The presence of Mollicutes in placental tissue increased the chance of developing miscarriage sevenfold. The prevalence of U. parvum in women who experienced spontaneous abortion was 66.3% in placental tissue. A positive association was observed between the detection of U. parvum in samples of placental tissue and abortion. There was a significant increase in microbial load in placental tissue for M. hominis, U. urealyticum and U. parvum compared to the control group. Detection of U. parvum in CM in pregnant women can ascend to the region of the placental tissue and trigger a spontaneous abortion.

This experiment was designed to evaluate the effects of the inclusion of detoxified castor bean meal (CM) content on the chemical-bromatological composition, as well as the fermentation characteristics and dry matter losses of sugarcane silage. The treatments consisted of four levels (0, 7, 14 and 21% natural matter) of addition of castor bean meal. The design was completely randomized, with five replicates. The material was ensiled in PVC silos of 50-cm in height and 10-cm in diameter, which were opened after 60 days. The density of fodder maintained in the silos was equal to 750kg of natural matter/m 3 . In order to quantify the gas and effluent losses, the silos were weighed at the beginning and at the end of the experiment. The addition of castor bean meal has increased the levels of DM and crude protein and reduced the levels of neutral detergent fiber, N-NH 3 , ethanol, and gas and effluent losses from silages, but did not affect pH values. During ensiling, alcoholic fermentation was controlled with the inclusion of the additive. RESUMOO experimento foi desenvolvido para avaliar os efeitos da inclusão de níveis de farelo de mamona destoxificado sobre a composição químico-bromatológica, as características fermentativas e as perdas de matéria seca da silagem de cana-de-açúcar. Os tratamentos consistiam em quatro níveis (0, 7, 14 e 21% da matéria natural) de inclusão do farelo de mamona. O delineamento foi inteiramente ao acaso, com cinco repetições. Foram usados silos de PVC, com 50cm de altura e 10cm de diâmetro, para a produção das silagens, que foram abertos após 60 dias. A densidade de forragem nos silos foi correspondente a 750kg de matéria natural/m 3 . Os silos foram pesados no início e ao final do período experimental para quantificar as perdas por gases e efluentes. A inclusão de farelo de mamona elevou os teores de MS e proteína bruta e reduziu os teores de fibra em detergente neutro, N-NH 3 , etanol e as perdas por gases e efluentes das silagens, não ocasionando efeito nos valores de pH. A fermentação alcoólica durante a ensilagem foi controlada com a inclusão do aditivo.

This study reports the pathogen Perkinsus beihaiensis in oysters of the genus Crassostrea on the coast of the State of Bahia (Brazil), its prevalence, infection intensity and correlation with salinity. Oysters (n = 240) were collected between October and December 2014 at eight sampling stations between latitudes 13°55'S and 15°42'S. The laboratory procedures included macroscopic analysis, histology, culture in Ray's fluid thioglycollate medium (RFTM), Polymerase Chain Reaction (PCR) and DNA sequencing. PCR and sequencing have been used for the genetic identification of oysters as well. Two species of oysters have been identified: Crassostrea rhizophorae and C. brasiliana. In both oyster species P. beihaiensis was the only Perkinsus species detected. In C. rhizophorae, the average prevalence was 82.8% by histology and 65.2% by RFTM. In C. brasiliana, the prevalences were 70.5% and 35.7%, respectively. The higher prevalence of P. beihaiensis in C. rhizophorae was probably influenced by salinity, with which was positively correlated (r> 0.8). In both oysters, P. beihaiensis was located mainly in the gastric epithelium. The infection was generally mild or moderate, without apparent harm to the hosts, but in cases of severe infection, there was hemocytical reaction and tissue disorganization. The generally high prevalence in the region suggests that oysters should be monitored with respect to this pathogen, especially in growing areas.Keywords: bivalves, diseases, pathology, salinity. Perkinsus beihaiensis (Perkinsozoa) em ostras do Estado da Bahia ResumoEste estudo relata o patógeno Perkinsus beihaiensis em ostras do gênero Crassostrea no litoral do Estado da Bahia (Brasil), sua prevalência, intensidade de infecção e correlação com a salinidade. As ostras (n = 240) foram coletadas entre outubro e dezembro de 2014 em oito estações amostrais entre as latitudes 13°55'S e 15°42'S. Os procedimentos laboratoriais incluíram análise macroscópica, histologia, cultivo em meio de tioglicolato de Ray (RFTM), reação em cadeia da polimerase (PCR) e sequenciamento de DNA. PCR e sequenciamento foram também utilizados para a identificação genética das ostras. Foram identificadas duas espécies de ostras: Crassostrea rhizophorae e C. brasiliana. Em ambas as espécies, P. beihaiensis foi a única espécie de Perkinsus detectada. Em C. rhizophorae, a prevalência média foi de 82,8% por histologia e de 65,2% por RFTM. Em C. brasiliana, as prevalências foram de 70,5% e 35,7%, respectivamente. A maior prevalência de P. beihaiensis em C. rhizophorae foi provavelmente influenciada pela salinidade, com a qual este apresentou correlação positiva (r>0,8). Em ambas as espécies, P. beihaiensis esteve localizada principalmente no epitélio gástrico. A infecção foi geralmente leve ou moderada, sem danos aparentes aos hospedeiros, mas em casos de infecção severa, houve reação hemocitária e desorganização de tecidos. As prevalências geralmente altas na região sugerem que as ostras devam ser monitoradas com relação a este patógeno, principalmente e...

SUMMARYThe experiment was developed to evaluate the effects of wilting and inclusion of cassava meal level on nutritive value, fermentative characteristics and losses of elephant grass silage. The 4 x 2 factorial scheme was adopted, with four repetitions, in a completely randomized design. The factors evaluated were wilting (without and with) and cassava meal level (0; 7.5; 15 and 22.5% of natural matter, DN). PVC silos with 50 cm height and 10 cm diameter were used for silages production, which were opened after 60 days. The wilting and cassava meal addition had increasing linear effect on DM content. Silages with cassava meal showed reduction of total nitrogen, ether extract, acid detergent insoluble nitrogen, neutral detergent fiber ash and protein free, acid detergent fiber, celluloses, hemicelluloses and lignin contents and increase of organic matter content, DM in vitro digestibility and total digestible nutrients. Relative to variables inherent to silages' fermentative characteristics, pH presented quadratic behavior. Concerning to ammonia nitrogen, it was observed quadratic and decreasing linear behavior for silages without and with wilting, respectively. Wilting and cassava meal inclusion provided smaller effluent losses.