RESUMO
Lentinus crinitus (Basidiomycota: Polyporales) is a saprophytic fungus with biotechnological importance described more than 20 years ago. However, there are few studies on the long-term preservation of this basidiomycete. Cryopreservation is a long-term storage technique that reduces the metabolic activity of microorganisms, but its success depends on the adjustment of the freezing process, the cryoprotectants, and the protective substrates for each species. This study aimed to assess the mycelial viability and genetic stability of L. crinitus strains cryopreserved at -86 °C for two years by the wheat grain technique using different cryoprotectants and freezing methods. Three strains of L. crinitus (U9-1, U13-5, and U15-12) were subjected to different concentrations and types of cryoprotectants (dimethyl sulfoxide, glycerol, glucose, and sucrose), freezing methods such as immediate freezing from 25 to -86 °C and progressing freezing from 25 to -86 °C in a freezing container with isopropyl alcohol to control the rate of cell freezing at -1 °C min-1, protective substrate (wheat grain and 2% malt extract agar), and cryopreservation period (1, 6, 12, and 24 months). After thawing, samples were evaluated for mycelial viability, time to mycelial recovery, mycelial stability, and genetic stability of the fungus. All techniques achieved effective cryopreservation at -86 °C, mainly with the wheat grain technique. All cryoprotectants (3.5% glycerol, 1.5% dimethyl sulfoxide, 25% sucrose, and 5% glucose), freezing methods (immediate and gradual), and protective substrate (wheat grain and malt extract agar) were effective for cryopreservation of the three L.crinitus strains in an ultra-low temperature freezer for two years. Mycelial viability, mycelial stability, and genetic stability of the fungus were not affected after two-year cryopreservation, evidencing the robustness of the long-term cryopreservation technique and the fungus.
Assuntos
Basidiomycota , Dimetil Sulfóxido , Ágar , Basidiomycota/metabolismo , Criopreservação/métodos , Crioprotetores/farmacologia , Congelamento , Glucose , Glicerol , Lentinula , Extratos Vegetais , Sacarose , TriticumRESUMO
Prochilodus brevis is a migratory fish, an important component of its river ecosystem and an appreciated animal in northeastern cuisine. However, human activities have threatened its survival. Thus, researchers have become interested in developing genetic material storage protocols, such as seminal cryopreservation. Therefore, determination of the appropriate freezing media and thawing rate is a fundamental step toward the use of this biotechnology in the production of common curimatã and for reducing risks to the species survival. As such, the aim of the current study was to evaluate the effect of different freezing media and thawing rates on the quality of cryopreserved semen from P. brevis. For this study, males received a single dose of pituitary extract of carp 18 hours before semen collection. The semen samples were diluted in 5% glucose + 10% methyl glycol (MG), 5% glucose + 10% dimethyl sulfoxide (DMSO), 0.9% NaCl + 10% methyl glycol, and 0.9% NaCl + 10% dimethyl sulfoxide, loaded into 0.25-mL straws and frozen in liquid nitrogen vapor. The semen from each treatment was thawed at three different thawing rates: 25 C for 30 sec, 30 C for 16 sec and 40 C for 12 sec. Motility, vitality and morphology analyses were performed by computer-assisted sperm analysis (CASA). The characteristics of the fresh sperm mostly resembled those found in the literature. For the parameter...(AU)
O Prochilodus brevis é um peixe reofílico, importante componente do ecossistema fluvial e apreciado na culinária nordestina. No entanto, ações antrópicas têm ameaçado sua sobrevivência. Desta forma, surge, nos pesquisadores, o interesse no desenvolvimento de protocolos de conservação do material genético, como a criopreservação seminal. Logo, a determinação do meio de congelação e da taxa de descongelação adequados, são passos fundamentais que possibilitarão a utilização dessa biotecnologia na produção de curimatã comum, reduzindo os riscos à sua sobrevivência. Portanto, o objetivo desse trabalho foi avaliar o efeito de diferentes meios de congelação e taxas de descongelação sobre a qualidade do sêmen criopreservado de P. brevis. Para isso, 18 horas antes da coleta de sêmen, os machos receberam dose única de extrato hipofisário de carpa. Cada animal foi sedado com solução à base de eugenol e o sêmen foi coletado. As amostras foram diluídas em quatro meios de congelação (5% Glicose + Metilglicol 10%; 5% Glicose + DMSO 10%; 0,9% NaCl + Metilglicol 10%; 0,9% NaCl + DMSO 10%) envasadas em palhetas de 0,25 mL e congeladas em vapor de nitrogênio líquido. O sêmen foi descongelado após sete dias em três taxas de descongelação: 25 C 30 s-1; 30 C 16 s-1; 40 C 12 s-1. Foram feitas as análises de motilidade, vitalidade e morfologia com auxílio de sistema automatizado de análise seminal...(AU)
Assuntos
Animais , Caraciformes/embriologia , Caraciformes/fisiologia , Criopreservação , Recuperação Espermática , CrioprotetoresRESUMO
Prochilodus brevis is a migratory fish, an important component of its river ecosystem and an appreciated animal in northeastern cuisine. However, human activities have threatened its survival. Thus, researchers have become interested in developing genetic material storage protocols, such as seminal cryopreservation. Therefore, determination of the appropriate freezing media and thawing rate is a fundamental step toward the use of this biotechnology in the production of common curimatã and for reducing risks to the species survival. As such, the aim of the current study was to evaluate the effect of different freezing media and thawing rates on the quality of cryopreserved semen from P. brevis. For this study, males received a single dose of pituitary extract of carp 18 hours before semen collection. The semen samples were diluted in 5% glucose + 10% methyl glycol (MG), 5% glucose + 10% dimethyl sulfoxide (DMSO), 0.9% NaCl + 10% methyl glycol, and 0.9% NaCl + 10% dimethyl sulfoxide, loaded into 0.25-mL straws and frozen in liquid nitrogen vapor. The semen from each treatment was thawed at three different thawing rates: 25 C for 30 sec, 30 C for 16 sec and 40 C for 12 sec. Motility, vitality and morphology analyses were performed by computer-assisted sperm analysis (CASA). The characteristics of the fresh sperm mostly resembled those found in the literature. For the parameter...
O Prochilodus brevis é um peixe reofílico, importante componente do ecossistema fluvial e apreciado na culinária nordestina. No entanto, ações antrópicas têm ameaçado sua sobrevivência. Desta forma, surge, nos pesquisadores, o interesse no desenvolvimento de protocolos de conservação do material genético, como a criopreservação seminal. Logo, a determinação do meio de congelação e da taxa de descongelação adequados, são passos fundamentais que possibilitarão a utilização dessa biotecnologia na produção de curimatã comum, reduzindo os riscos à sua sobrevivência. Portanto, o objetivo desse trabalho foi avaliar o efeito de diferentes meios de congelação e taxas de descongelação sobre a qualidade do sêmen criopreservado de P. brevis. Para isso, 18 horas antes da coleta de sêmen, os machos receberam dose única de extrato hipofisário de carpa. Cada animal foi sedado com solução à base de eugenol e o sêmen foi coletado. As amostras foram diluídas em quatro meios de congelação (5% Glicose + Metilglicol 10%; 5% Glicose + DMSO 10%; 0,9% NaCl + Metilglicol 10%; 0,9% NaCl + DMSO 10%) envasadas em palhetas de 0,25 mL e congeladas em vapor de nitrogênio líquido. O sêmen foi descongelado após sete dias em três taxas de descongelação: 25 C 30 s-1; 30 C 16 s-1; 40 C 12 s-1. Foram feitas as análises de motilidade, vitalidade e morfologia com auxílio de sistema automatizado de análise seminal...
Assuntos
Animais , Caraciformes/embriologia , Caraciformes/fisiologia , Criopreservação , Crioprotetores , Recuperação EspermáticaRESUMO
O reconhecimento da grande relevância da biodiversidade microbiana para o desenvolvimento humano tem conduzido ao aprimoramento de técnicas destinadas à conservação dos mais diversos espécimes microbiológicos. Contudo, ainda não existe uma fórmula singular, ideal ou universal que possibilite a eficiência da estocagem e preservação microbiana a longo prazo. Consequentemente, diversos micro-organismos para os quais há uma crescente demanda, ainda esperam pelo desenvolvimento de metodologias adequadas à sua conservação, fazendo da criobiologia um campo de vasto potencial para o desenvolvimento de pesquisas. Nesse contexto, a proposta dessa revisão é discutir os princípios básicos da criomicrobiologia, com enfoque na preservação dos variados tipos de micro-organismos e nos principais agentes crioprotetores, tomando como base estudos disponíveis na literatura científica.
The recognizing the great importance of microbial biodiversity for human development has led to the improvement of techniques for the conservation of diverse microbiological specimens. However, there isn't a singular, ideal or universal method to provice the efficiency of the long-term microorganisms stock and preservation. Many microorganisms for which there is a growing demand still wait for the development of satisfactory methodologies to their conservation, making the cryobiology a field of vast potential for the development of researches. Then, the proposal of this review is to discuss the basic principles of cryomicrobiology, focusing the preservation of kinds microorganisms and principals agents cryoprotectants, using the available studies in the comtemporary scientific literature as base.
Assuntos
Criopreservação , Crioprotetores , Biodiversidade , MicrobiologiaRESUMO
The present investigation represents the effect of freeze drying on some properties as acid and bile tolerance of Streptococcus thermophilus MTCC 1938 culture isolated from dairy products. The cell paste obtained from milk based medium was freeze dried with a pressure of 50-100 mtorr for 24h at -40ºC. Acid and bile tolerance test exhibited 3.8-4.9 and 3.2-3.8 log counts reduction after freeze drying respectively.
Assuntos
Ácido Láctico/análise , Crioprotetores/análise , Fermento Seco/análise , Laticínios/análise , Leite , Streptococcus thermophilus/isolamento & purificação , Microbiologia de Alimentos , Amostras de Alimentos , Liofilização , Métodos , MétodosRESUMO
The present investigation represents the effect of freeze drying on some properties as acid and bile tolerance of Streptococcus thermophilus MTCC 1938 culture isolated from dairy products. The cell paste obtained from milk based medium was freeze dried with a pressure of 50-100 mtorr for 24h at -40°C. Acid and bile tolerance test exhibited 3.8-4.9 and 3.2-3.8 log counts reduction after freeze drying respectively.
RESUMO
The present investigation represents the effect of freeze drying on some properties as acid and bile tolerance of Streptococcus thermophilus MTCC 1938 culture isolated from dairy products. The cell paste obtained from milk based medium was freeze dried with a pressure of 50-100 mtorr for 24h at -40ºC. Acid and bile tolerance test exhibited 3.8-4.9 and 3.2-3.8 log counts reduction after freeze drying respectively.