Good practices in the rearing and maintenance of zebrafish (Danio rerio) in Brazilian laboratories / Boas práticas na criação e manutenção de zebrafish (Danio rerio) em laboratório no Brasil

Good Laboratory Practice (GLP) is a management quality control system that encompasses the organizational process and conditions under which non-clinical health and environmental studies are carried out. According to the World Health Organization, GLP must contain five topics: resources, characterization, rules, results, and quality control. The aim of this work is to address a review according to WHO standard to the implementation of Good Laboratory Practices in zebrafish (Danio rerio) facility. Considering that the promotion of one health (animal, human, and environmental) associated with an education plan, protocols, and records are fundamental to guarantee the safety and integrity of employees, animals and the environment as well as reliability in the results generated. In a way, Brazil still needs improvements related to the wellfare of aquatic organisms (national laws, international agreements, corporate programs, and others); especially in relation to its use in research and technological development. In this way, the implementation of GLPs provide valuable guidance for improving animal welfare and worker safety, facilitating the standardization of research.

As Boas Práticas de Laboratório (BPL) são um sistema de controle de qualidade gerencial que abrange o processo organizacional e as condições sob as quais os estudos não clínicos de saúde e meio ambiente são desenvolvidos. Conforme a Organização Mundial da Saúde (OMS) as BPL devem conter cinco tópicos: recursos, caracterização, regras, resultados e controle de qualidade. O objetivo deste trabalho foi apresentar uma revisão conforme o padrão da OMS para a implementação das BPL em biotério de zebrafish. Considerando que a promoção da saúde única (animal, humana e ambiental) associada a um plano de educação, protocolos e registros são fundamentais para garantir a segurança e a integridade dos trabalhadores/pesquisadores, animais e meio ambiente assim como confiabilidade nos resultados gerados. De certa forma o Brasil ainda necessita de melhorias relacionadas ao bem-estar de organismos aquáticos (leis nacionais, acordos internacionais, programas corporativos e outros); especialmente em relação à utilização destes na pesquisa e desenvolvimento tecnológico. Desta forma, a implementação de BPL fornece uma orientação valiosa para a melhoria do bem-estar animal, e segurança do trabalhador vindo a facilitar a padronização da pesquisa.

The effect of adjusting settings within a Computer-Assisted Sperm Analysis (CASA) system on bovine sperm motility and morphology results

Abstract Semen motility is the most widely recognized semen quality parameter used by Artificial Insemination (AI) centers. With the increasing worldwide export of semen between AI centers there is an increasing need for standardized motility assessment methods. Computer-Assisted Sperm Analysis (CASA) technology is thought to provide an objective motility evaluation; however, results can still vary between laboratories. The aim of present study was to verify the impact of different setting values of the CASA IVOS II on motility, concentration, and morphology of bovine semen samples frozen in an extender with or without egg yolk and then decide on optimal settings for a further validation step across AI centers. Semen straws from 30 different bulls were analyzed using IVOS II with twelve modified settings. No significant changes were observed in semen concentration, percentage of motile sperm or kinetic results for either extender type. However, increasing settings for both STR and VAP progressive (%) from Low, Medium, and High cut-off values significantly (p<0.05) reduced the percentage of detected progressive spermatozoa, in egg yolk extender from 49.5±15.2, 37.2±11.9 to 11.9±5.3%, and in clear extender from 51.9±9.1, 35.8±7.3 to 10.0±2.4%, respectively. In clear extender only, the modification of droplet proximal head length significantly affected the detection of normal sperm percentages (88.0± 4.7 to 95.0±0.6 and 96.0±0.6%) and of the percentage of detected proximal droplets (12.2±4.7, 2.5±2.7 to 0.6±0.2%) for Low, Medium and High values respectively (p<0.05). The identification of sensitivity within the CASA system to changes in set parameters then led to the determination of an optimal IVOS II setting. The existing variability among centers for these phenotypes was reduced when the standardized settings were applied across different CASA units. The results clearly show the importance of applied settings for the final CASA results and emphasize the need for standardized settings to obtain comparable data.

The effect of adjusting settings within a ComputerAssisted Sperm Analysis (CASA) system on bovine sperm motility and morphology results

Semen motility is the most widely recognized semen quality parameter used by Artificial Insemination (AI) centers. With the increasing worldwide export of semen between AI centers there is an increasing need for standardized motility assessment methods. Computer-Assisted Sperm Analysis (CASA) technology is thought to provide an objective motility evaluation; however, results can still vary between laboratories. The aim of present study was to verify the impact of different setting values of the CASA IVOS II on motility, concentration, and morphology of bovine semen samples frozen in an extender with or without egg yolk and then decide on optimal settings for a further validation step across AI centers. Semen straws from 30 different bulls were analyzed using IVOS II with twelve modified settings. No significant changes were observed in semen concentration, percentage of motile sperm or kinetic results for either extender type. However, increasing settings for both STR and VAP progressive (%) from Low, Medium, and High cut-off values significantly (p<0.05) reduced the percentage of detected progressive spermatozoa, in egg yolk extender from 49.5±15.2, 37.2±11.9 to 11.9±5.3%, and in clear extender from 51.9±9.1, 35.8±7.3 to 10.0±2.4%, respectively. In clear extender only, the modification of droplet proximal head length significantly affected the detection of normal sperm percentages (88.0± 4.7 to 95.0±0.6 and 96.0±0.6%) and of the percentage of detected proximal droplets (12.2±4.7, 2.5±2.7 to 0.6±0.2%) for Low, Medium and High values respectively (p<0.05). The identification of sensitivity within the CASA system to changes in set parameters then led to the determination of an optimal IVOS II setting. The existing variability among centers for these phenotypes was reduced when the standardized settings were applied across different CASA units. The results clearly show the importance of applied settings for the final CASA results and emphasize the need for standardized settings to obtain comparable data.(AU)

Protocol for the in vitro rearing of Frieseomelitta varia workers (Hymenoptera: Apidae: Meliponini)

In vitro rearing protocols already established for honeybees are currently being adapted to assess the risk of pesticides, and to conduct comparative developmental biology studies on stingless bees. However, differences in critical life-history traits (development time and the type of larval nutrition leading to caste differentiation process) among social bees require the development of an in vitro rearing protocol for each species and caste. We generated a protocol to produce workers of Frieseomelitta varia (Lepeletier, 1836), a non-endangered and highly eusocial pollinator species with wide geographical distribution. We tested the viability of using either the eggs or the first instar larvae as the starting point for in-vitro transfer. In vitro rearing was performed in acrylic plates at 30 °C and 99% relative humidity during the larval feeding phase. The humidity was subsequently reduced to 75% during the following days of development. The experimental larvae were offered either 25 µL or 27 µL of larval food. The development time, emergence and mortality rates, and morphological parameters of the emerged workers were assessed. In the process of validating the protocol, the adults that emerged after in vitro rearing were compared with colony-reared adults. In our results, 27 µL of larval food allowed 90% of workers to emerge. No significant differences were found between the emerging workers reared in vitro and those reared in the colony. The described protocol is a useful method for rearing F. varia workers in vitro, which can be used for diverse types of experimental approaches.

Standardization of resazurin use in susceptibility testing of natural products against yeasts in planktonic cells and in biofilms formation

Natural products, such as the ethanolic propolis extract (PE), have been shown to be a safe and effective therapeutic alternative for the treatment of fungal skin and nail diseases. However, the presence of the resin and the physicochemical characteristics of the extract sometimes difficult the reading and determination of breakpoints of the in vitrotests, evidencing the need for alternatives that facilitate the reading. The present study aimed to standardize the use of resazurin in tests of susceptibility of PE with planktonic yeast cells and biofilm forms. The antifungal activity of PE was determined by minimum inhibitory concentration (MIC) and we observed that, for all Candida spp. tested, the most reproducible MIC results were obtained when resazurin was placed after 24 hours of incubation and remained more 24 hourswith yeasts plus PE. For encapsulated yeasts, there was no dye reduction and color transition. Resazurin was also used for the evaluation of minimal biofilm inhibitory concentration and minimal biofilm eradication concentration and it was metabolized and reproducedthe action of PE on Candida biofilms. In addition, microdilution checkerboard plates were made with the dye, which assisted reading the result of the interaction between PE and nystatin. We observed that the resin, the color and the turbidity of the PE slightly changed the color of the resazurin in high concentrations of the extract and did not impair the reading. Therefore, the resazurin standardization tests were proven to be efficient and grounds that it should be used as an auxiliary methodology for reading and interpretation of the susceptibility tests for non-encapsulated yeasts with natural products, which form turbidity or precipitation, such as propolis.

Standardization of resazurin use in susceptibility testing of natural products against yeasts in planktonic cells and in biofilms formation

Natural products, such as the ethanolic propolis extract (PE), have been shown to be a safe and effective therapeutic alternative for the treatment of fungal skin and nail diseases. However, the presence of the resin and the physicochemical characteristics of the extract sometimes difficult the reading and determination of breakpoints of the in vitrotests, evidencing the need for alternatives that facilitate the reading. The present study aimed to standardize the use of resazurin in tests of susceptibility of PE with planktonic yeast cells and biofilm forms. The antifungal activity of PE was determined by minimum inhibitory concentration (MIC) and we observed that, for all Candida spp. tested, the most reproducible MIC results were obtained when resazurin was placed after 24 hours of incubation and remained more 24 hourswith yeasts plus PE. For encapsulated yeasts, there was no dye reduction and color transition. Resazurin was also used for the evaluation of minimal biofilm inhibitory concentration and minimal biofilm eradication concentration and it was metabolized and reproducedthe action of PE on Candida biofilms. In addition, microdilution checkerboard plates were made with the dye, which assisted reading the result of the interaction between PE and nystatin. We observed that the resin, the color and the turbidity of the PE slightly changed the color of the resazurin in high concentrations of the extract and did not impair the reading. Therefore, the resazurin standardization tests were proven to be efficient and grounds that it should be used as an auxiliary methodology for reading and interpretation of the susceptibility tests for non-encapsulated yeasts with natural products, which form turbidity or precipitation, such as propolis.(AU)

Influencing factors for preparation of platelet-rich plasma in horses / Fatores que influenciam o preparo do plasma rico em plaquetas na espécie equina

There is no consensus in the literature as to the best platelet-rich plasma (PRP) acquisition method or the ideal platelet concentration to stimulate tissue repair. Besides that, most studies do not provide a clear and replicable description of the methods used, which makes standardization and result comparison difficult. Thus, this study aimed to accurately describe the method and evaluate factors influencing PRP preparation in equines. In this regard, two protocols were used, P1 and P2, based on two centrifugation methods, which differed in blood volume, speed and time, and platelet pipetting and resuspension. In conclusion, factors such as centrifugation strength and time are essential to obtain PRP with adequate platelet count. However, other factors such as tube type and plasma pipetting and resuspension methods directly interfere with the replicability of the technique, and hence influencing PRP applicability.(AU)

Não há consenso na literatura quanto ao melhor método de obtenção do plasma rico em plaquetas (PRP) ou à concentração plaquetária ideal com o objetivo de estimular o reparo tecidual. Além disso, grande parte dos trabalhos não faz descrição clara e reprodutível dos métodos utilizados, o que dificulta a padronização e a comparação dos resultados. Dessa forma, este trabalho teve por objetivo descrever detalhadamente a metodologia e avaliar fatores que influenciam o preparo do PRP em equinos. Para isso, foram utilizados dois protocolos, P1 e P2, baseados em duas centrifugações, que diferiram quanto ao volume de sangue colhido, velocidade e tempo das centrifugações e método de pipetagem e ressuspensão das plaquetas. Conclui-se que fatores como força e tempo de centrifugação são essenciais à obtenção de um PRP com concentração plaquetária adequada. Contudo, fatores adicionais como tipo de tubo e métodos de pipetagem e ressuspensão do plasma interferem diretamente na repetibilidade da técnica, influenciando a aplicabilidade prática do PRP.(AU)

Standardization of blood meal production process at a rendering plant using design of experiments / Padronização do processo de produção de farinha de sangue em uma usina de processamento por meio do planejamento de experimentos

This research focuses on the standardization of the blood meal production process at a Colombian rendering plant through a design of experiments. Initially, 108 samples of blood meal were taken where only 23% achieved the moisture target (7.5% to 8.5%). Therefore, an analysis of the measurement system was performed using a repeatability and reproducibility (R&R) study. Results showed that 39.96% of the observed variability was caused by the measurement system that was out of control. So, it was necessary to improve the method of sampling reducing the participation of the measurement system in the variability of the process to only 3.79%. Later, several experiments were accomplished with a 2k factorial design. Each experiment consisted of a response variable (blood meal moisture), two controllable factors (drying chamber temperature and percentage of rotation of the metering screw), and an uncontrollable factor (initial blood meal moisture). Finally, experiments were carried out and validated observing that, with a drying chamber temperature of 160 °C and a percentage of screw rotation of 29%, more than 97% of the blood meal was according to the moisture target. In conclusion, is confirmed that the design of experiments is a tool that allows a clear path towards optimization and standardization of processes.(AU)

Esta pesquisa se concentra na padronização do processo de produção de farinha de sangue de uma usina de beneficiamento colombiana por meio de um planejamento de experimentos. Em primeiro lugar, foi realizado um estudo de 108 amostras de farinha de sangue onde apenas 23% atingiram a meta de umidade (7.5% a 8.5%). Portanto, uma análise do sistema de medição foi realizada usando um estudo de repetibilidade e reprodutibilidade (R&R). Os resultados mostraram que 39.96% da variabilidade observada foi causada pelo sistema. Assim, foi necessário aprimorar o método de amostragem, reduzindo a participação do sistema na variabilidade para apenas 3.79%. Posteriormente, vários experimentos foram realizados com um planejamento fatorial 2k. Cada experimento consistiu em uma variável de resposta (umidade da farinha de sangue), dois fatores controláveis (temperatura da câmara de secagem e porcentagem de rotação da rosca dosadora) e um fator incontrolável (umidade inicial da farinha de sangue). Finalmente, experimentos foram realizados e validados observando que, com uma temperatura da câmara de secagem de 160 ° C e um percentual de rotação da rosca de 29%, mais de 97% da farinha de sangue estava de acordo com a umidade desejada. Em conclusão, confirma-se que o planejamento de experimentos é uma ferramenta que permite um caminho claro para a otimização e padronização de processos.(AU)

Elaboração do padrão de qualidade para cortes de frango

Foi elaborada um padrão de qualidade de cortes de frango (peito com osso, asas, coxa e sobrecoxa e filé de coxa e sobrecoxa) em um abatedouro no Sudeste Goiano. A análise foi efetuada por meio da obtenção de amostras, análise visual e captura de imagem, com 9 amostras de peito com osso, 13 de asa, 20 de coxa e sobrecoxa e 9 de filé de coxa e sobrecoxa. O padrão de qualidade não aceitável (presença de fraturas, presença de pés, hematomas, ausência de pele, entre outros) apresentou maior número de amostras. Foi possível concluir que a padronização é uma forma de melhorar continuamente o processo, agregar valor ao produto e evitar insatisfação do cliente.(AU)

A quality standard of chicken cuts (bone-in breast, wings, thigh and drumstick and thigh and drumstick fillet) was prepared in a slaughterhouse in Southeast Goiás. The analysis was performed by obtaining samples, visual analysis and image capture, with 9 samples of breast with bone, 13 of wing, 20 of thigh and drumstick and 9 of thigh and drumstick fillet. The unacceptable quality standard (presence of fractures, presence of feet, bruises, absence of skin, among others) showed a greater number of samples. It was concluded that standardization is a way to continuously improve the process, add value to the product and avoid customer dissatisfaction.(AU)

Elaboração do padrão de qualidade para cortes de frango

Foi elaborada um padrão de qualidade de cortes de frango (peito com osso, asas, coxa e sobrecoxa e filé de coxa e sobrecoxa) em um abatedouro no Sudeste Goiano. A análise foi efetuada por meio da obtenção de amostras, análise visual e captura de imagem, com 9 amostras de peito com osso, 13 de asa, 20 de coxa e sobrecoxa e 9 de filé de coxa e sobrecoxa. O padrão de qualidade não aceitável (presença de fraturas, presença de pés, hematomas, ausência de pele, entre outros) apresentou maior número de amostras. Foi possível concluir que a padronização é uma forma de melhorar continuamente o processo, agregar valor ao produto e evitar insatisfação do cliente.

A quality standard of chicken cuts (bone-in breast, wings, thigh and drumstick and thigh and drumstick fillet) was prepared in a slaughterhouse in Southeast Goiás. The analysis was performed by obtaining samples, visual analysis and image capture, with 9 samples of breast with bone, 13 of wing, 20 of thigh and drumstick and 9 of thigh and drumstick fillet. The unacceptable quality standard (presence of fractures, presence of feet, bruises, absence of skin, among others) showed a greater number of samples. It was concluded that standardization is a way to continuously improve the process, add value to the product and avoid customer dissatisfaction.

Musculoskeletal ultrasonography of the elbow joint in dogs: applicability and evaluation protocol / Ultrassonografia musculoesquelética da articulação do cotovelo em cães: aplicabilidade e protocolo de avaliação

The elbow is a complex joint and has great clinical relevance in small animal medicine. Previous research in this area has been performed using radiographic and tomographic methods; however, there are limited studies on ultrasonography. The aims of this study was suggesting an evaluation protocol for elbow scan and describe the ultrasonographic anatomy of the elbow joint in dogs. Ten cross-breed dogs weighing 5-15kg underwent radiography and were selected for this ultrasonographic study. The protocol was established for the ultrasonographic description dividing the articular areas in the proximal, middle, and distal, lateral, cranial, medial, and caudal faces. The approach was performed in the longitudinal, transverse and oblique planes and the musculoskeletal structures were described according to the architecture, echogenicity and echotexture. Computed tomography and magnetic resonance imaging scans were obtained for one animal for comparison. Ultrasonography was effective in visualizing and analyzing muscles, tendons and ligaments. Bone contours and regions that have clinical significance such as the medial coronoid process and anconeus process were identified, but with limited access. Prior knowledge of the normal sonographic anatomy of the elbow joint, as well as its technical advantages and limitations will allow further studies related to the identification of musculoskeletal disorders.(AU)

O cotovelo é uma articulação complexa e tem grande relevância clínica na medicina veterinária de pequenos animais. Pesquisas prévias nesta área foram realizadas utilizando radiografias e tomografia computadorizada, entretanto há limitados estudos com ultrassonografia. O objetivo desse estudo é sugerir um protocolo de avaliação da articulação do cotovelo e descrever sua anatomia ultrassonográfica. Dez cães sem raça definida, pesando 5-15kg foram submetidos à radiografias e foram selecionados para o estudo ultrassonográfico. O protocolo foi estabelecido para a descrição anatômica ultrassonográfica dividindo as articulações em proximal, média e distal, faces lateral, cranial, medial e caudal. A abordagem foi realizada nos planos longitudinal, transverso e oblíquo e as estruturas foram descritas de acordo com a arquitetura, ecogenicidade e ecotextura. Tomografia computadorizada e ressonância magnética foram realizadas em um animal para comparação. A ultrassonografia foi efetiva na visualização e análise de músculos, tendões e ligamentos. Os contornos ósseos e regiões com significado clínico como o processo coronóide medial e o processo ancôneo foram identificados, mas com acesso limitado. Conhecimento prévio da anatomia ultrassonográfica normal da arquitetura do cotovelo, bem como suas vantagens e limitações, irão permitir estudos adicionais relacionados à identificação de desordens musculoesqueléticas.(AU)

Musculoskeletal ultrasonography of the elbow joint in dogs: applicability and evaluation protocol / Ultrassonografia musculoesquelética da articulação do cotovelo em cães: aplicabilidade e protocolo de avaliação

The elbow is a complex joint and has great clinical relevance in small animal medicine. Previous research in this area has been performed using radiographic and tomographic methods; however, there are limited studies on ultrasonography. The aims of this study was suggesting an evaluation protocol for elbow scan and describe the ultrasonographic anatomy of the elbow joint in dogs. Ten cross-breed dogs weighing 5-15kg underwent radiography and were selected for this ultrasonographic study. The protocol was established for the ultrasonographic description dividing the articular areas in the proximal, middle, and distal, lateral, cranial, medial, and caudal faces. The approach was performed in the longitudinal, transverse and oblique planes and the musculoskeletal structures were described according to the architecture, echogenicity and echotexture. Computed tomography and magnetic resonance imaging scans were obtained for one animal for comparison. Ultrasonography was effective in visualizing and analyzing muscles, tendons and ligaments. Bone contours and regions that have clinical significance such as the medial coronoid process and anconeus process were identified, but with limited access. Prior knowledge of the normal sonographic anatomy of the elbow joint, as well as its technical advantages and limitations will allow further studies related to the identification of musculoskeletal disorders.(AU)

O cotovelo é uma articulação complexa e tem grande relevância clínica na medicina veterinária de pequenos animais. Pesquisas prévias nesta área foram realizadas utilizando radiografias e tomografia computadorizada, entretanto há limitados estudos com ultrassonografia. O objetivo desse estudo é sugerir um protocolo de avaliação da articulação do cotovelo e descrever sua anatomia ultrassonográfica. Dez cães sem raça definida, pesando 5-15kg foram submetidos à radiografias e foram selecionados para o estudo ultrassonográfico. O protocolo foi estabelecido para a descrição anatômica ultrassonográfica dividindo as articulações em proximal, média e distal, faces lateral, cranial, medial e caudal. A abordagem foi realizada nos planos longitudinal, transverso e oblíquo e as estruturas foram descritas de acordo com a arquitetura, ecogenicidade e ecotextura. Tomografia computadorizada e ressonância magnética foram realizadas em um animal para comparação. A ultrassonografia foi efetiva na visualização e análise de músculos, tendões e ligamentos. Os contornos ósseos e regiões com significado clínico como o processo coronóide medial e o processo ancôneo foram identificados, mas com acesso limitado. Conhecimento prévio da anatomia ultrassonográfica normal da arquitetura do cotovelo, bem como suas vantagens e limitações, irão permitir estudos adicionais relacionados à identificação de desordens musculoesqueléticas.(AU)

Duplex nested-PCR for detection of small ruminant lentiviruses

Abstract Small ruminant lentiviruses (SRLV) have high genetic variability which results in different viral strains around the world. This create a challenge to design sensible primers for molecular diagnosis in different regions. This work proposes a protocol of duplex nested-PCR for the precise diagnosis of SRLV. The technique was designed and tested with the control strains CAEV Co and MVV 1514. Then, field strains were submitted to the same protocol of duplex nested-PCR. Blood samples of sheep and goats were tested with AGID and nested PCR with specific primers for pol, gag and LTR. The AGID results showed low detection capacity of positive animals, while the nested PCR demonstrated a greater capacity of virus detection. Results demonstrated that LTR-PCR was more efficient in detecting positive sheep samples, whereas gag-PCR allowed a good detection of samples of positive goats and positive sheep. In addition, pol-PCR was more efficient with goat samples than for sheep. Duplex nested PCR performed with standard virus samples and field strains demonstrated that the technique is more efficient for the detection of multiple pro-viral DNA sequences. This study demonstrated a successful duplex nested PCR assay allowing a more accurate diagnosis of SRLV.

Duplex nested-PCR for detection of small ruminant lentiviruses

Small ruminant lentiviruses (SRLV) have high genetic variability which results in different viral strains around the world. This create a challenge to design sensible primers for molecular diagnosis in different regions. This work proposes a protocol of duplex nested-PCR for the precise diagnosis of SRLV. The technique was designed and tested with the control strains CAEV Co and MVV 1514. Then, field strains were submitted to the same protocol of duplex nested-PCR. Blood samples of sheep and goats were tested with AGID and nested PCR with specific primers for pol, gag and LTR. The AGID results showed low detection capacity of positive animals, while the nested PCR demonstrated a greater capacity of virus detection. Results demonstrated that LTR-PCR was more efficient in detecting positive sheep samples, whereas gag-PCR allowed a good detection of samples of positive goats and positive sheep. In addition, pol-PCR was more efficient with goat samples than for sheep. Duplex nested PCR performed with standard virus samples and field strains demonstrated that the technique is more efficient for the detection of multiple pro-viral DNA sequences. This study demonstrated a successful duplex nested PCR assay allowing a more accurate diagnosis of SRLV.(AU)

EXPRESSÃO GÊNICA DA CALPASTATINA E CARACTERÍSTICAS QUALITATIVAS EM CARNES DE BOVINOS NELORE SUBMETIDAS À ESTIMULAÇÃO ELÉTRICA E MATURAÇÃO

A existência de variabilidade na concentração de calpastatina entre indivíduos do mesmo grupo genético (Bos taurus indicus) contribui para a falta de padronização da maciez dos cortes cárneos oriundos desses animais e resulta em menor valor agregado ao produto, o que afeta negativamente a competitividade do setor brasileiro no mercado mundial. A estimulação elétrica das carcaças e a maturação tem sido importantes ferramentas tecnológicas utilizadas com o intuito de melhorar a maciez da carne bovina. O objetivo do presente estudo foi associar a expressão do gene da calpastatina (CAST) e suas isoformas (CAST 1 e CAST 2) com a dureza da carne de bovinos Nelore submetidos à estimulação elétrica (EE) e à maturação. Foram selecionadas 10 carcaças inteiras, onde as meias carcaças esquerdas foram estimuladas eletricamente, enquanto as suas respectivas direitas, não. Dois grupos experimentais foram formados com base nos valores de força de cisalhamento (WBSF, Warner-Bratzler shear force) das amostras de carnes com 48 horas post-mortem (meias carcaças direitas), sendo: Carne moderadamente dura(CMD, 5,0 ± 0,17 kg) e Carne muito dura (CD, 7,28 kg ± 0,43). Amostras do M. longissimus thoracis et lumborum de ambas meias carcaças foram submetidas à maturação (7, 14, 21 e 28 dias) e análise da qualidade da carne (48 horas, 7, 14, 21 e 28 dias). Amostras de M. longissimus thoracis das meias carcaças direitas foram utilizadas para avaliação da expressão gênica de CAST, CAST 1 e CAST 2. As expressões gênicas de CAST 1 e CAST 2 foram maiores no grupo de CD comparada ao CMD (P < 0,05) e se mostraram superiores à CAST (P < 0,05), independente do grupo experimental. A EE acelerou o declínio do pH nas primeiras 9 horas postmortem, influenciou (P < 0,05) os parâmetros L*, a*, b*, saturação e tonalidade da carne de ambos os grupos experimentais e eliminou as diferenças de WBSF entre os grupos CD e CMD já com 48 horas post-mortem. Os presentes resultados sugerem que a EE e maturação contribuem para a redução da dureza da carne de bovinos Nelore mesmo daquelas que apresentam maior expressão de CAST 1 e CAST 2.

The existence of variability in the concentration of calpastatin among individuals of the same genetic group (Bos taurus indicus) contributes to the lack of standardization of the tenderness of meat cuts from these animals and results in lower added value to the product, which negatively affects the competitiveness of the sector Brazilian market in the world. The electrical stimulation of carcasses and maturation have been important technological tools used in order to improve the tenderness of beef. The aim of the present study was to associate the expression of the calpastatin gene (CAST) and its isoforms (CAST 1 and CAST 2) with the toughness of meat from Nellore cattle submitted to electrical stimulation (EE) and maturation. Ten whole carcasses were selected, where the left half carcasses were electrically stimulated, while their respective right half carcasses were not. Two experimental groups were formed based on the values of shear force (WBSF, Warner-Bratzler shear force) of meat samples with 48 hours post-mortem (right half carcasses), as follows: Moderately tough meat (CMD, 5.0 ± 0.17 kg) and Very tough meat (CD, 7.28 kg ± 0.43). Samples of M. longissimus thoracis et lumborum from both half carcasses were submitted to maturation (7, 14, 21 and 28 days) and meat quality analysis (48 hours, 7, 14, 21 and 28 days). Samples of M. longissimus thoracis from the right half carcasses were used to evaluate the gene expression of CAST, CAST 1 and CAST 2. Gene expressions of CAST 1 and CAST 2 were higher in the CD group compared to CMD (P < 0.05 ) and were superior to CAST (P < 0.05), regardless of the experimental group. The EE accelerated the pH decline in the first 9 hours post-mortem, influenced (P < 0.05) the parameters L*, a*, b*, saturation and meat hue of both experimental groups and eliminated the WBSF differences between CD and CMD groups with 48 hours post-mortem. The present results suggest that EE and maturation contribute to the reduction of meat toughness in Nellore cattle, even those with higher expression of CAST 1 and CAST 2.

O princípio da padronização como uma boa prática de Gestão em Biotérios: Licitação de Rações Comerciais para Animais de Laboratório

As licitações são práticas utilizadas pela Administração Pública, com a finalidade de obter produtos e serviços selecionando a proposta mais vantajosa ao alcance do interesse público. Entretanto, o critério de vantajosidade nem sempre contempla o melhor produto e serviço a ser adquirido. Se a vantajosidade não estiver devidamente fundamentada através de criteriosos estudos técnicos e mercadológicos, as aquisições demonstram fragilidades. Considerando estas fragilidades encontradas, o objetivo deste trabalho é propor melhorias aos ritos procedimentais das licitações, adotando o processo de padronização as aquisições realizadas pelo Instituto de Ciência em Tecnologia em Biomodelos (ICTB), da Fundação Oswaldo Cruz (FIOCRUZ), com as compras de rações para roedores de laboratório. Para essa investigação foram feitas as análises dos atuais procedimentos licitatórios, investigando tempo excedente (demora) para a homologação do certame que impactam em insegurança e ineficiência para os gestores envolvidos nesta aquisição de rações processadas, que são vitais ao atendimento nutricional destes animais que subsidiam as pesquisas científicas desenvolvidas pela FIOCRUZ. Foi definido como fator crítico de sucesso ao aprimoramento processual a utilização de algumas estratégias/instrumentos de gestão. Desta forma, através da modelagem do processo para o novo rito procedimental de compras, foi elaborada a cesta de indicadores de desempenho a fim de monitorar os procedimentos de avaliação da ração e da licitação objetivando a uniformização das práticas, e com isso minimizar a variação nos resultados experimentais, propiciando aos pesquisadores o uso de animais que atendam às suas necessidades experimentais a bem da saúde pública. Espera-se assim, a geração de um banco de fornecedores com as marcas validadas, evitando desperdício de tempo/retrabalho, o que pode colocar em risco o bem-estar animal. Portanto, pretende-se com a adoção da padronização benefícios não somente aos animais de laboratório, bem como à Administração Pública, considerando os princípios da eficiência e celeridade, propiciando maior segurança jurídica e técnica aos gestores.

Tenders are practices used by the Public Administration, with the purpose of obtaining products and services by selecting the most advantageous proposal within the reach of the public interest. However, the criterion of advantage does not always include the best product and service to be purchased. If the advantage is not properly substantiated through careful technical and market studies, acquisitions show weaknesses. Considering these weaknesses found, the objective of this work is to propose improvements to the procedural rites of bidding, adopting the process of standardizing the acquisitions carried out by the Institute of Science in Technology in Biomodels (ICTB), of the Oswaldo Cruz Foundation (FIOCRUZ), with the purchases of rations for laboratory rodents. For this investigation, the analyzes of the current bidding procedures were carried out, investigating excess time (delay) for the approval of the event that impacts insecurity and inefficiency for the managers involved in this acquisition of processed feed, which are vital to the nutritional care of these animals that subsidize the scientific research developed by FIOCRUZ. The use of some management strategies/instruments was defined as a critical success factor for procedural improvement. In this way, through the modeling of the process for the new procedural rite of purchases, the basket of performance indicators was prepared in order to monitor the procedures for evaluating the feed and the bidding, aiming to standardize practices, and thereby minimize the variation in the experimental results, providing researchers with the use of animals that meet their experimental needs for the sake of public health. Thus, it is expected the generation of a supplier bank with validated brands, avoiding waste of time/rework, which can put animal welfare at risk. Therefore, it is intended with the adoption of standardization benefits not only to laboratory animals, as well as to the Public Administration, considering the principles of efficiency and speed, providing greater legal and technical security to managers.

Epidemiologia da toxoplasmose ocular em Bancos de Olhos do Estado de São Paulo, Brasil

Toxoplasma gondii é um parasita intracelular obrigatório amplamente disseminado pelo mundo. Estima-se que um terço da população mundial esteja cronicamente infectada por esse coccídio. No Brasil, essa prevalência chega a ser estimada em 70% em algumas regiões do país. Apesar de apresentar uma alta prevalência, a primo-infecção por T. gondii acaba passando despercebida em pacientes imunocompetentes, com quadros subclínicos ou com sintomas semelhantes aos de uma gripe. Em pacientes imunocomprometidos e em mulheres grávidas, infecções recentes merecem grande atenção, pois a evolução clínica da doença é mais severa, podendo levar ao desenvolvimento de sequelas irreversíveis ou até à morte. Uma das manifestações mais comuns da infecção sistêmica por T. gondii é a forma ocular, acometendo principalmente a retina e a coroide. O isolamento de T. gondii a partir de amostras de seres humanos é imprescindível para verificar se existe ou não uma correlação entre a expressão fenotípica da doença e a linhagem do parasita. No Brasil, ainda há poucos relatos de isolamentos obtidos a partir de amostras humanas. O objetivo do presente estudo foi determinar a epidemiologia da toxoplasmose ocular em Bancos de Olhos das cidades de Ribeirão Preto (BORP) e São José do Rio Preto (BOSJRP), no interior do Estado de São Paulo. Foram recebidos um total de 395 globos oculares correspondentes a 201 doadores. Destes globos oculares, 289 foram captados pelo BORP, enquanto 106 foram captados pelo BOSJRP. Dentre os doadores do BORP, 42/148 (28,3%) possuíam ambos os tecidos (olhos e sangue) disponíveis para pesquisa e 5,4% (8/148) dos doadores possuíam lesões sugestivas para toxoplasmose ocular (TO). Três deles apresentavam lesões macroscópicas bilaterais, isto é, em ambos os olhos, e cinco possuíam lesões unilaterais. Dentre os 53 doadores do BOSJRP, 86,8% (46/53) possuíam ambos os tecidos disponíveis para pesquisa e nenhuma amostra de tecido ocular apresentava lesões sugestivas para TO. Do total de 88 soros analisados, 70,5% (62/88) apresentavam apenas anticorpos IgG anti-T. gondii, 6,8% (6/88) foram positivos para ambos os anticorpos (IgG e IgM) anti-T. gondii e 22,7% (20/88) foram considerados soronegativos. Todos os globos oculares com lesão potencialmente causada por T. gondii foram submetidos ao bioensaio em camundongos para tentativa de isolamento do parasita. Ao término do período de bioensaio, nenhum camundongo apresentou anticorpos anti-T. gondii no Teste de Aglutinação Modificado (MAT) ou cistos nos tecidos analisados durante a necropsia. Na cPCR, a amplificação dos fragmentos de interesse do gene B1 e da região repetitiva do genoma REP-529 foi observada em uma amostra que não apresentava lesão macroscópica aparente; esta amostra foi designada como PS-TgOTBrSP1. Além dessa amostra, outras oito apresentaram amplificação para os marcadores selecionados na PCR em tempo real, portanto, 2,3% (9/395) dos globos oculares foram diagnosticados como positivos para T. gondii. As demais amostras, incluindo todas as onze amostras com lesão macroscópica sugestiva para TO, foram diagnosticadas como negativas. A amostra PS-TgOTBrSP1 foi identificada como não-arquétipa por meio da análise de 15 marcadores microssatélites e um genótipo incompleto, não-arquétipo, foi obtido por meio de PCR-RFLP. O genótipo MS de T. gondii da amostra PS-TgOTBrSP1 é um genótipo único, o que contribui para corroborar a baixa circulação de linhagens clássicas clonais e a ampla diversidade genética de T. gondii no Brasil. E, embora só tenha sido possível examinar um número reduzido de amostras, o presente estudo também contribui para validar dados encontrados em outros estudos que apontam que 50-80% da população adulta tem anticorpos para T. gondii no Brasil.

Toxoplasma gondii is an obligate intracellular parasite widely spread around the world. It is estimated that one third of the world population is chronically infected by this coccidia. In Brazil, this prevalence is estimated at 70% in some regions of the country. Despite having a high prevalence, the primary infection by T. gondii ends up going unnoticed in immunocompetent patients, with subclinical conditions or with symptoms similar to those in a flu. In immunocompromised patients and pregnant women, recent infections deserve great attention, as the clinical course of the disease is more severe and can lead to the development of irreversible sequelae or even death. One of the most common manifestations of systemic infection by T. gondii is the ocular form, affecting mainly the retina and choroid. The isolation of T. gondii from human samples is essential to verify whether or not there is a correlation between the phenotypic expression of the disease and the parasite lineage. In Brazil, there are few reports of isolations obtained from human samples. The aim of this study was to determine the epidemiology of ocular toxoplasmosis in eye banks in the cities of Ribeirão Preto (BORP) and São José do Rio Preto (BOSJRP), from São Paulo State. A total of 395 eyes from 201 donors were received. Of these samples, 289 were captured by BORP, while 106 were captured by BOSJRP. Among the BORP donors, 42/148 (28.3%) had both tissues (eyes and blood) available for research and 5.4% (8/148) of the donors had lesions compatible with ocular toxoplasmosis (OT). Three of them had bilateral lesions and five had unilateral lesions. Among the 53 BOSJRP donors, 86.8% (46/53) had both tissues available for research and no ocular tissue sample had lesions compatible with OT. Of the total 88 sera analyzed, 70.5% (62/88) had only IgG antibodies against T. gondii, 6.8% (6/88) were positive for both antibodies (IgG and IgM) against T. gondii and 22.7% (20/88) were considered seronegative. All eyes with lesions potentially caused by T. gondii were submitted to a bioassay in mice to isolate the parasite. At the end of the bioassay period, no mouse showed antibodies against T. gondii in the Modified Agglutination Test (MAT) and no cysts were found in tissues analyzed during necropsy. In cPCR, the amplification of the fragments of B1 gene and the repetitive region of the REP-529 genome was observed in a sample, designated as PS-TgOTBrSP1 which did not show macroscopic lesion. In addition to this positive sample, eight others showed amplification for selected markers in real-time PCR, therefore, 2.3% (9/395) of the eyes were diagnosed as positive for T. gondii. The remaining samples, including all eleven samples with macroscopic lesions, were diagnosed as negative. The PS-TgOTBrSP1 sample was identified as non-archetype through the analysis of 15 microsatellite markers and an incomplete, non-archetype genotype was obtained through PCR-RFLP. The MS genotype of T. gondii from the PSTgOTBrSP1 sample is a unique genotype, which contributes to corroborate that T. gondii has a low circulation of classical clonal strains and has a wide genetic variability in Brazil. Although it was only possible to examine a small number of samples in this study, the data herein obtained also contributes to validate findings from other groups showing that 50-80% of the adult population has antibodies against T. gondii in Brazil.

PARÂMETROS OFTÁLMICOS EM CÃES ADULTOS DA RAÇA SPITZ ALEMÃO ANÃO

A raça Spitz Alemão Anão ganhou recente aumento de popularidade e algumasdas particularidades oftalmológicas desta ainda são desconhecidas. Com o presente estudo objetivou-se conhecer parâmetros oftálmicos na raça. Estudaram-se 48 olhos de 24 cães, machos e fêmeas, com idades entre 1 e 7,5 anos, oriundos de canil de criação. Sob contenção física delicada, foram empregados teste lacrimal de Schirmer, tempo de ruptura do filme lacrimal, teste de Jones, estesiometria de Cochet-Bonnet, biomicroscopia com lâmpada em fenda, pressão intraocular, microscopia especular de não contato, flaremetria a laser, oftalmoscopia indireta e ultrassonografia. Aplicou-se estatística descritiva para análise dos dados. As médias e erros padrões encontrados foram, quanto ao teste lacrimal de Schirmer24,310±0,448mm/min e ao tempo de ruptura do filme lacrimal 14,570±0,406s. Ao teste de Jones 8,420±1,571min sendo 62,5% dos olhos positivos. À estesiometria, a medida foi 3,667±0,157cm. À biomicroscopia, 43,75% dos olhos apresentaram pelos na carúncula, 25% cromodacriorreia, 25% discreto entrópio de canto medial, 10,42% distiquíase, 6,25% mácula corneal, 4,16% nódulo palpebral e 2,08% triquíase de canto medial. À tonometria de rebote, os valores obtidos média e erro padrão foram 17,960±0,697mmHg. À microscopia especular foi encontrada densidade de células endoteliais 1958,285±36,760células/mm². À hexagonalidade, 62,229±2,183% e para área das células endoteliais, 519,231±9,853mm². A espessura corneal central foi de 0,567±0,006mm. À flaremetria10,440±2,514PC/ms. À oftalmoscopia, foi observado padrão descrito em outras raças. Padrão amelanótico foi reportado em 8,33% dos olhos. À ultrassonografia obtiveram-se médias e erros padrões de 16,859±0,148mm para diâmetro axial horizontal. Espessura da lente 6,651±0,081mm, profundidade da câmara anterior 2,673±0,091mm e,profundidade de câmara vítrea 7,485±0,101mm. Os parâmetros e dados encontrados corresponderam à literatura em sua maioria. Alterações discretas nos parâmetros de avaliação lacrimal foram identificadas. Constatou-se boa sensibilidade corneal. À ultrassonografia, os diâmetros correspondentes foram inferiores quando comparado à cães braquicefálicos.

The Spitz German Klein breed has shown a substantial increase in popularity and ophthalmic particularities of this breed are still unknown. The present study aimed to establish ophthalmic parameters in German Spitz Klein. The research for this study was made through 48 eyes of 24 male and female dogs, aged between 1 and 7.5 years old, from a breeding kennel. Thru physical restraint it was used Schirmer tear test, break up time test, Jones test, Cochet-Bonnet esthesiometry, slit lamp biomicroscopy, rebound tonometry, specular microscopy of non-contact, laser flaremetry, ophthalmoscopy and ultrasonography. With the purpose of analyzing data this study was applied descriptive statistics. The means and standard errors found were, for the Schirmer tear test 24.310±0.448mm/min and the break up time test 14.570±0.406s. For the Jones test 8.420±1.571min being 62.5% of the positive eyes. For the esthesiometry was found 3.667±0.157cm. The biomicroscopy, it was found that 43.75% of the eyes presented hair on the caruncle, 25% chromodacryreia, 25% discrete medial entropion, 10.42% of distichiasis, 6.25% of corneal macula, 4.16% of palpebral nodule and 2.08% of medial trichiasis. For the rebound tonometry, the values obtained and standard error were 17.960±0.679mmHg. Furthermore, under the specular microscopy was found density of endothelial cells 1958.285±36.760cells/mm². The hexagonality, 62.229±2.183% and for endothelial cell area, 519.231±9.853mm². The central corneal thickness was 0.567±0.006mm. The flaremetry 10,440±2,514PC/ms.The ophthalmoscopy showed a fundus described in other races. Amelanotic fundus was reported in 8.33% of eyes. The ultrasonography obtained an average and standard error of 16.859±0.148mm for horizontal axial diameter. Lens thickness 6.651±0.081mm, depth of the anterior chamber 2.673±0.091mm and, depth of the vitreous chamber 7.485±0.101mm.Parameters and data found corresponded to the literature in the majority. Discrete changes in lacrimal evaluation parameters were identified. Good corneal sensitivity was found. At ultrasonography, the corresponding diameters were lower compared to the brachycephalic dogs.

Qualidade de ovos de codorna em diferentes tempos de armazenamento / Quality of quail eggs at different times of storage

A trial was conducted to evaluate quality of quail eggs from two companies (A and B) at two storage times (5 and 15 days after packing). A total of 480 eggs were kept at room temperature and distributed in a completely randomized design in 2x2factorial arrangement  (companies and storage time). The results indicated that trays from company B presented higher weight, as well as mean egg weight. However, the eggs in trays from company A showed greater uniformity. Evaluating the external quality, we found that above 90% of eggshell were considered with total integrity, and no difference for this characteristic between companies was found. The eggs from company A presented lower average weight in comparison to those from company B, regarding storage time. No differences were observed between eggs from companies A and B for Haugh unit value. Storage time of 15 days after packing promoted reduction in Haugh unit value in comparison to those stored for 5 days. Quail eggs from company A presented lower variation in weight, although heavier eggs are produced by company B. Internal quality of quail eggs from both companies was similar. Storage time affected eggs quality.

O estudo foi conduzido para avaliar a qualidade de ovos de codorna de duas empresas (A e B) em dois tempos de armazenamento (5 e 15 dias após data de embalagem). Um total de 480 ovos foram mantidos em temperatura ambiente e distribuídos em delineamento inteiramente casualizado em esquema fatorial 2x2 (empresas e tempo de armazenamento). Os resultados indicam que as bandejas de ovos da companhia B apresentaram maior peso, como também no peso médio dos ovos. No entanto, os ovos em bandejas da empresa A apresentou maior uniformidade. Avaliando-se a qualidade externa, foi observada que acima de 90% dos ovos apresentaram cascas com total integridade, e não houve diferença para esta característica entre as empresas. Os ovos da empresa A apresentaram peso médio menor em comparação com os da empresa B, indenpendente do tempo de armazenamento. Não foram observadas diferenças entre os ovos de empresas A e B para valores de unidade Haugh. O tempo de armazenamento de 15 dias após a embalagem promoveu a redução nos valores de unidade Haugh em relação aos armazenados por 5 dias. Ovos de codorna da empresa A apresentaram menor variação de peso, embora os ovos da empresa B tenham apresentado maior peso. Os ovos de ambas as empresas apresentaram mais de 90% de ovos com integridade total da casca. qualidade interna de ovos de codorna de ambas asempresas foram semelhantes, apesar do tempo de armazenamento ter afetado as características qualitativas dos mesmos.

Quality of quail eggs at different times of storage

A trial was conducted to evaluate quality of quail eggs from two companies (A and B) at two storage times (5 and 15 days after packing). A total of 480 eggs were kept at room temperature and distributed in a completely randomized design in 2x2factorial arrangement (companies and storage time). The results indicated that trays from company B presented higher weight, as well as mean egg weight. However, the eggs in trays from company A showed greater uniformity. Evaluating the external quality, we found that above 90% of eggshell were considered with total integrity, and no difference for this characteristic between companies was found. The eggs from company A presented lower average weight in comparison to those from company B, regarding storage time. No differences were observed between eggs from companies A and B for Haugh unit value. Storage time of 15 days after packing promoted reduction in Haugh unit value in comparison to those stored for 5 days. Quail eggs from company A presented lower variation in weight, although heavier eggs are produced by company B. Internal quality of quail eggs from both companies was similar. Storage time affected eggs quality.

O estudo foi conduzido para avaliar a qualidade de ovos de codorna de duas empresas (A e B) em dois tempos de armazenamento (5 e 15 dias após data de embalagem). Um total de 480 ovos foram mantidos em temperatura ambiente e distribuídos em delineamento inteiramente casualizado em esquema fatorial 2x2 (empresas e tempo de armazenamento). Os resultados indicam que as bandejas de ovos da companhia B apresentaram maior peso, como também no peso médio dos ovos. No entanto, os ovos em bandejas da empresa A apresentou maior uniformidade. Avaliando-se a qualidade externa, foi observada que acima de 90% dos ovos apresentaram cascas com total integridade, e não houve diferença para esta característica entre as empresas. Os ovos da empresa A apresentaram peso médio menor em comparação com os da empresa B, indenpendente do tempo de armazenamento. Não foram observadas diferenças entre os ovos de empresas A e B para valores de unidade Haugh. O tempo de armazenamento de 15 dias após a embalagem promoveu a redução nos valores de unidade Haugh em relação aos armazenados por 5 dias. Ovos de codorna da empresa A apresentaram menor variação de peso, embora os ovos da empresa B tenham apresentado maior peso. Os ovos de ambas as empresas apresentaram mais de 90% de ovos com integridade total da casca. A qualidade interna de ovos de codorna de ambas as empresas foram semelhantes, apesar do tempo de armazenamento ter afetado as características qualitativas dos mesmos.