Identificação molecular e caracterização tecnológica de bactérias ácido-lácticas isoladas de leite in natura e queijo colonial
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2016-07
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Souza, Claucia Fernanda Volken De
Banca
Souza, Claucia Fernanda Volken De
Pozzobon, Adriane
Granada, Camille Eichelberger
Maciel, Mônica Jachetti
Dullius, Carlos Henrique
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Culturas lácticas são consideradas um dos principais ingredientes da fabricação de derivados lácteos fermentados, sendo fundamental para a obtenção de um produto de boa qualidade e padronizado. No preparo destas culturas para a elaboração dos derivados, os microrganismos desejáveis são isolados, selecionados e preservados em culturas puras. O Vale do Taquari, RS, possui em sua constituição econômica e social a produção de leite e derivados como importante fator para o seu desenvolvimento. Desta forma, as indústrias de laticínios da região buscam a utilização de bactérias lácticas com características próprias capazes de conferir melhor aroma, sabor e textura aos seus produtos lácteos fermentados. O objetivo do presente trabalho foi identificar e caracterizar tecnologicamente isolados bacterianos de leite in natura e queijo colonial produzidos na região, visando o uso na indústria de laticínios. Foram coletadas 10 amostras de leite bovino in natura e 10 amostras de queijo colonial produzido sem o uso de cultura iniciadora. Obteve-se um total de 119 microrganismos, dentre eles, 109 bactérias. Os isolados bacterianos foram caracterizados quanto às propriedades tecnológicas: crescimento microbiano; produção de ácido láctico a partir do crescimento em leite desnatado reconstituído (LDR), soro de queijo e permeado de soro de queijo a 7 e 32 ºC; produção de diacetil; e atividade proteolítica e identificados a nível de gênero, espécie e subespécie por meio de sequenciamento de um fragmento do gene 16S rRNA. Identificou-se 81 isolados, sendo que a espécie e subespécie predominante foi Lactobacillus paracasei subsp. tolerans. As bactérias demonstraram uma boa produção de ácidos orgânicos no meio LDR a 32oC, sendo que se destacaram os isolados nº: 108 (Bacillus spp.), 112 (Lactobacillus pentosus), 117 (Lactobacillus plantarum), 118 (não identificado), 124 (Lactococcus spp.),129 (não identificado),131 (Lactobacillus spp.),136 (não identificado) e 141 (não identificado). A produção de proteases avaliada no estudo apresentou bom desempenho para a maioria dos microrganismos, sendo que se destacaram os isolados nº 4 (Lactobacillus parabuchneri), 5 (Bacillus methylotrophicus), 6 (não identificado), 12 (Lactobacillus spp.), 13 (Lactobacillus paracsei subsp. tolerans), 31 (Lactobacillus paracasei subsp. tolerans), 34 (Lactobacillus paracasei subsp. tolerans), 37 (Acetobacter sicerae), 58 (Lactobacillus parabuchneri), e 88a (Lactobacillus otakiensis). Os resultados verificados para a produção de diacetil (aroma) pelas BALs, mostraram que alguns microrganismos são capazes de promover o desenvolvimento de sabor, seis deles mostraram destaque: 21 (Lactobacillus paracasei subsp. tolerans), 28 (Lactobacillus zeae), 30 (Lactobacillus harbinensis), 55 (não identificado), 123 (Lactobacillus paracasei subsp. tolerans), 132 (Lactobacillus spp.). O teste de contagem de células viáveis mostrou-se similar em todos os isolados, exibindo uma contagem entre 9,0 e 11,7 log UFC/mL, atingindo a fase exponencial após 24 horas de incubação a 32 oC. De acordo com a caracterização tecnológica das bactérias isoladas na região do Vale do Taquari, Sul do Brasil, e sua identificação molecular através do sequenciamento do 16S rRNA, verifica-se que estas podem contribuir para o desenvolvimento regional através de sua aplicação na indústria de alimentos.
Lactic cultures are considered a key ingredient in the manufacturing of fermented dairy products, being instrumental for the obtainment of a product with good quality and standard. In the preparation of these cultures for the elaboration of these derivatives, the wanted microorganisms are isolated, selected and preserved in pure cultures. Vale do Taquari, RS, has in its economic and social setting up the production of dairy products as an important factor for its development. Thus, the dairy industries of the region seek the use of lactic bacteria with characteristics able to provide better aroma, flavor and texture to their fermented dairy products. The objective of this study was to identify and characterize bacterial technologically isolates of raw bovine milk and colonial cheese produced in the region, aiming for the use in dairy industry. 10 samples of raw bovine milk and 10 samples of colonial cheese produced without the use of starter culture were collected. We obtained a total of 119 microorganisms, among which 109 were bacteria. The bacterial isolates were characterized by technological properties: microbial growth; production of lactic acid from the growth of reconstituted skim milk (RSM), cheese whey and cheese whey permeate at 7 and 32°C; production of diacetyl; and proteolytic activity identified in genus, species and subspecies by sequencing a fragment of 16S rRNA. 81 isolates were identified, and the dominant species and subspecies were Lactobacillus paracasei subsp tolerans. The bacteria showed a good production of organic acids in the environment RSM at 32°C, and the isolates which stood out were: 108 (Bacillus spp.), 112 (Lactobacillus pentosus), 117 (Lactobacillus plantarum), 118 (unidentified), 124 (Lactococcus spp.), 129 (unidentified), 131 (Lactobacillus spp.), 136 (unidentified) and 141 (unidentified). The production of proteases evaluated in the study showed good performance for most microorganisms, and the isolates which stood out were: 4 (Lactobacillus parabuchneri), 5 (Bacillus methylotrophicus), 6 (unidentified), 12 (Lactobacillus spp.), 13 (Lactobacillus paracsei subsp. tolerans), 31 (Lactobacillus paracasei subsp. tolerans), 34 (Lactobacillus paracasei subsp. tolerans), 37 (Acetobacter sicerae), 58 (Lactobacillus parabuchneri) and 88a (Lactobacillus otakiensis). The results obtained for the production of diacetyl (aroma) by the lactic acid bacteria (LAB) showed that some microorganisms are capable of promoting the development of flavor, 6 of them showed prominence: 21 (Lactobacillus paracasei subsp. tolerans), 28 (Lactobacillus zeae), 30 (Lactobacillus harbinensis), 55 (unidentified), 123 (Lactobacillus paracasei subsp. tolerans), 132 (Lactobacillus spp.). The test of viable cell count was similar in all isolates, showing a count between 9.0 and 11.7 log CFU/ml, reaching the exponential phase after 24 hours of incubation at 32ºC. According to the technological characterization of the bacteria isolated in Vale do Taquari, southern of Brazil, and its molecular identification by sequencing of the 16S rRNA, we verify that they can contribute to the development of the region, through their application in the food industry.
Lactic cultures are considered a key ingredient in the manufacturing of fermented dairy products, being instrumental for the obtainment of a product with good quality and standard. In the preparation of these cultures for the elaboration of these derivatives, the wanted microorganisms are isolated, selected and preserved in pure cultures. Vale do Taquari, RS, has in its economic and social setting up the production of dairy products as an important factor for its development. Thus, the dairy industries of the region seek the use of lactic bacteria with characteristics able to provide better aroma, flavor and texture to their fermented dairy products. The objective of this study was to identify and characterize bacterial technologically isolates of raw bovine milk and colonial cheese produced in the region, aiming for the use in dairy industry. 10 samples of raw bovine milk and 10 samples of colonial cheese produced without the use of starter culture were collected. We obtained a total of 119 microorganisms, among which 109 were bacteria. The bacterial isolates were characterized by technological properties: microbial growth; production of lactic acid from the growth of reconstituted skim milk (RSM), cheese whey and cheese whey permeate at 7 and 32°C; production of diacetyl; and proteolytic activity identified in genus, species and subspecies by sequencing a fragment of 16S rRNA. 81 isolates were identified, and the dominant species and subspecies were Lactobacillus paracasei subsp tolerans. The bacteria showed a good production of organic acids in the environment RSM at 32°C, and the isolates which stood out were: 108 (Bacillus spp.), 112 (Lactobacillus pentosus), 117 (Lactobacillus plantarum), 118 (unidentified), 124 (Lactococcus spp.), 129 (unidentified), 131 (Lactobacillus spp.), 136 (unidentified) and 141 (unidentified). The production of proteases evaluated in the study showed good performance for most microorganisms, and the isolates which stood out were: 4 (Lactobacillus parabuchneri), 5 (Bacillus methylotrophicus), 6 (unidentified), 12 (Lactobacillus spp.), 13 (Lactobacillus paracsei subsp. tolerans), 31 (Lactobacillus paracasei subsp. tolerans), 34 (Lactobacillus paracasei subsp. tolerans), 37 (Acetobacter sicerae), 58 (Lactobacillus parabuchneri) and 88a (Lactobacillus otakiensis). The results obtained for the production of diacetyl (aroma) by the lactic acid bacteria (LAB) showed that some microorganisms are capable of promoting the development of flavor, 6 of them showed prominence: 21 (Lactobacillus paracasei subsp. tolerans), 28 (Lactobacillus zeae), 30 (Lactobacillus harbinensis), 55 (unidentified), 123 (Lactobacillus paracasei subsp. tolerans), 132 (Lactobacillus spp.). The test of viable cell count was similar in all isolates, showing a count between 9.0 and 11.7 log CFU/ml, reaching the exponential phase after 24 hours of incubation at 32ºC. According to the technological characterization of the bacteria isolated in Vale do Taquari, southern of Brazil, and its molecular identification by sequencing of the 16S rRNA, we verify that they can contribute to the development of the region, through their application in the food industry.
Descrição
Palavras-chave
Microrganismos; Fermentação; 16S rRNA
Citação
AGOSTINI, Camila. Identificação molecular e caracterização tecnológica de bactérias ácido-lácticas isoladas de leite in natura e queijo colonial. 2016. Dissertação (Mestrado) – Curso de Biotecnologia, Universidade do Vale do Taquari - Univates, Lajeado, 29 fev. 2016. Disponível em: http://hdl.handle.net/10737/1076.