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Analysis of Relationship Between Air Cavities and Weissella Viridescens

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Analysis of Relationship Between Air Cavities and Weissella Viridescens Analysis of the relationship between air cavities and Weissella viridescens in meat products and possibilities for air cavity elimination Josef Kameník, Marta Dušková University of Veterinary and Pharmaceutical Sciences Brno, Czech Republic Abstract The cavities in cooked hams are not necessarily associated with bacterial contamination and the growth of bacteria. If we disregard the mechanical way of cavity formation, in which air gets into the meat and penetrates between the muscle fibres in pork, the bacterial origin of cavities is associated with shortcomings in the production process. A large number of genera and species of lactic acid bacteria (LAB) have been isolated from spoiled meat and meat products. Heat treatment plays an important role in the selection of bacteria that may be brought into the product with the used raw material (meat) or additives, or from the production environment. However, cooking is not always effective when it comes to thermo- tolerant vegetative bacteria. The very important species in meat processing is Weissella viridescens which may contribute to the spoilage of meat products. As weissellas are, to a certain extent, thermo-resistant, they could survive cooking process. In the scientific literature, isolated cases have been described in which weissellas contributed to the formation of cavities in cooked hams. However, LAB of the genus Leuconostoc were isolated from these products far more often than weissellas. W. viridescens has also been described as spoilers for other groups of meat products, such as hot smoked dry sausages. Key Words: cooked meat products; cross contamination; lactic acid bacteria; Leuconostoc spp.; Weissella spp. 1 Contents 1. Lactic acid bacteria: characteristics, occurrence and importance in foods………....…….. 3 2. Lactic acid bacteria: their occurrence and importance in meat…………………………… 7 2.1. Lactic acid bacteria and packaged meat……………………………………………… 8 2.2. The importance of selected genera of LAB in meat spoilage………………………. 11 3. Lactic acid bacteria: their occurrence and importance in cooked meat products……...... 13 3.1. Sources of lactic acid bacteria in products………………………………………….. 13 3.2. Lactic acid bacteria and comminuted meat products……………………………….. 14 3.3. Lactic acid bacteria and cooked hams……………………………………………… 16 4. Weissella viridescens in meat products……………………...……………. …………..... 19 4.1. Methods of identifying Weissella spp…………………………….………………… 21 4.2. The occurrence of Weissella spp…………………………………………………… 22 4.3. Weissella viridescens in meat products…………………………………………….. 22 5. Cavities inside whole-muscle meat products: their causes and significance……...…….. 30 5.1. Bacteria as a possible cause of cavities in whole-muscle meat products…………... 33 6. Conclusions………………………………………………………….…………………... 36 References…………………………………………………………………….……………... 37 2 1. Lactic acid bacteria: characteristics, occurrence and importance in foods Lactic acid bacteria (LAB) are among the most widely studied bacterial groups at the present time. We can find them as agents of the spoilage of many foods and drinks, such as cooked meat products, mayonnaises, cheeses, salad dressings, beer, etc. Certain LAB may have virulence factors and belong among pathogens studied in human and veterinary medicine (e.g. certain representatives of the genus Streptococcus). The participation of LAB in the fermentation of foods of animal and plant origin is, however, far more widely known. They are one of most important groups of bacteria industrially and their use in fermentation processes has an extremely long history (Giraffa, 2014). The first starter culture was probably used for industrial purposes in Denmark, Germany and the USA in 1890 for the production of cheese and fermented milk (Holzapfel and Wood, 2014). LAB are a group of functionally related bacteria that are capable of producing lactic acid by homofermentative and/or heterofermentative metabolism (Ruiz et al., 2014). LAB are gram- positive bacteria of the phylum Firmicutes with a “low” G+C ratio (≤ 55 mol %) in the DNA (Holzapfel and Wood, 2014). The phylum Firmicutes takes in a number of classes. LAB are assigned to the class Bacilli. Two orders – Bacillales and Lactobacillales – belong to the class Bacilli. The order Lactobacillales currently has 6 families and 40 genera. This order is comprised of LAB characterised by enormous diversity. The six families are: • Aerococcaceae (7 genera, e.g. Aerococcus) • Carnobacteriaceae (16 genera, e.g. Carnobacterium) • Enterococcaceae (7 genera, e.g. Enterococcus, Tetragenococcus, Vagococcus) • Lactobacillaceae (3 genera, Lactobacillus, Paralactobacillus, Pediococcus) • Leuconostocaceae (4 genera, e.g. Leuconostoc, Oenococcus, Weissella) • Streptococcaceae (3 genera, Lactococcus, Lactovum, Streptococcus) Bacterial groups that have a similar metabolism, but which are not phylogenetically related, are sometimes classified among LAB. A typical example is the genus Bifidobacterium, though it belongs to a different phylum – Actinobacteria. This phylum includes microorganisms with a G+C ratio in the DNA of ≥ 55 mol % (Stolaki et al., 2012). LAB are demanding microorganisms that require rich nutrients for growth, such as saccharides, amino acids, vitamins and minerals. Certain LAB require special growth factors, such as tomato juice, whey, etc. LAB decompose a large number of saccharides and related compounds by means of various metabolic pathways. ATP is generated by substrate-level phosphorylation and serves for the transport of dissolved substances across the cell membrane and for the purposes of biosynthesis. Environmental conditions influence the metabolic pathways used (Endo and Dicks, 2014). The phenotypic characteristics of LAB include a positive reaction to Gram staining, the absence of endospores, a negative oxidase and catalase 3 reaction (Leroi, 2010), a saccharide fermentation model, production of D(-) and L(+) lactic acid, aesculin and arginine hydrolysis, nitrate reduction, gelatin liquefaction, growth under various conditions of temperature, pH and NaCl content, and tolerance to oxygen. As LAB do not have a functional respiratory system, they have to obtain energy by the phosphorylation of suitable substrates (Wright and Axelsson, 2012). Two metabolic pathways exist in LAB depending on the fermentation of glucose – homofermentative and heterofermentative (Endo and Dicks, 2014). Homofermentative LAB use glycolysis – the Embden-Meyerhof-Parnas pathway (EMP, Figure 1). Species of LAB that exhibit a homofermentative metabolism produce more than 85 % of their lactic acid from hexoses (Salvetti et al., 2013). The EMP is characterised by the formation of fructose-1,6-diphosphate (FDP), which is split by the enzyme FDP aldolase into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). GAP (and also DHAP via GAP) is subsequently converted into pyruvic acid (Pot and Tsakalidou, 2009). Heterofermentative LAB use the phosphoketolase pathway (the pentose phosphate pathway, PPP, Figure 2). In addition to lactic acid, the heterofermentative pathway also produces a significant quantity of carbon dioxide and ethanol or acetic acid (Wright and Axelsson, 2012). The choice of metabolic pathway is determined at the family level (Endo and Dicks, 2014). The glycolytic pathway is used by representatives of the families Enterococcaceae, Lactobacillaceae and Streptococcaceae, with the exception of one group in the genus Lactobacillus. This pathway converts glucose into lactic acid (2 molecules of lactic acid to 1 molecule of glucose). Two molecules of ATP are generated from one molecule of glucose. Representatives of the family Leuconostocaceae and a number of species in the genus Lactobacillus metabolise glucose by the phosphoketolase pathway, during which one molecule of lactic acid, one molecule of CO2 and one molecule of ethanol are released from one molecule of glucose. The formation of ethanol from acetyl phosphate, during which NADH is oxidised into NAD+, is important in this pathway. This reaction is associated with the speed with which glucose is fermented, and thereby with the intensity of growth of heterofermentative LAB. LAB generally produce D(-) and L(+) lactic acid. The ratio at which these isomers are released is almost identical among strains of the same species and is considered one of the key characteristics for the classification of LAB into subgroups. Nevertheless, not all species respect this characteristic. LAB from the family Carnobacteriaceae produce lactic acid, formic acid, acetic acid and ethanol from glucose. CO2 is not produced, with the exception of Carnobacterium spp. The ratios of metabolites differ depending on the bacteria, though the main end products are lactic acid and formic acid. 4 Figure 1: Glycolysis (Embden-Meyerhof-Parnas pathway). The diagram indicates the involvement of certain enzymes – a: glucokinase, b: fructose-1,6-diphosphate aldolase, c: lactate dehydrogenase (source: Endo and Dicks, 2014) The metabolism of disaccharides includes transport across the cell membrane either as free molecules or following phosphorylation. This is followed by cleavage into two monosaccharide molecules or a molecule of monosaccharide and a molecule of monosaccharide phosphate. Certain LAB prefer disaccharides to monosaccharides as growth substrates. Examples include the fermentation of lactose by dairy LAB or fermentation of maltose by dough LAB. LAB are divided into three groups according to the metabolic pathway used for the fermentation of glucose and the metabolism of pentoses
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