Effect of nitrate and incubation conditions on the production of catalase and nitrate reductase by staphylococci

doi:10.1016/S0168-1605(99)00127-0

International Journal of Food Microbiology

Volume 52, Issues 1–2, 1 November 1999, Pages 47-56

https://doi.org/10.1016/S0168-1605(99)00127-0 Get rights and content

Abstract

The objective of this work was to study the production of catalase and nitrate reductase by staphylococci in order to understand their role in lipid oxidation during sausage manufacturing. Catalase and nitrate reductase were measured in resting cells and supernatants of staphylococci grown in different conditions. All staphylococci (except S. warneri) synthetized nitrate reductase. In static condition, the synthesis was maximal during exponential growth phase, whereas in shaking condition, the synthesis was maximal at the beginning of stationary phase. The production of nitrate reductase was increased in presence of nitrate, this effect was particularly important for the two S. carnosus strains which exhibited the highest activity. For all staphylococci, the production of catalase was maximal at the end of the exponential growth phase. The lowest amount of catalase was produced by S. warneri and the highest by S. carnosus. Only S. xylosus 873 and S. saprophyticus 852 released high amounts of catalase in the supernatant growth. Staphylococci produced higher amounts of catalase in shaking conditions. Addition of nitrate in the growth media favoured the synthesis of catalase, with a pronounced effect for S. carnosus. Nitrate also favoured the release of catalase.

Introduction

Strains of Staphylococcus, either added as starter cultures or spoiling the raw material, contribute to the sensory properties of fermented sausages. They are well known for their participation in the reddening of sausage through their nitrate reductase activity (Weber, 1994). They also influence the level of many fragrant compounds from different metabolic Berdagué et al., 1993, Stahnke, 1994, Stahnke, 1995, Montel et al., 1996, Talon and Montel, 1997, Talon et al., 1998). Lipid oxidation generates numerous volatile compounds and staphylococci modify the proportion of the different volatiles. So, sausages inoculated with Staphylococcus carnosus or Staphylococcus xylosus were characterized by dry cured aroma in relation to a high content of ketones together with other volatiles originating from amino acids, and carbohydrates (Montel et al., 1996). Sausages manufactured with Staphylococcus warneri had rancid odours correlated with high content of aldehydes (Berdagué et al., 1993). So the balance between all these compounds has to be controlled.

To understand the role of staphylococci in lipid oxidation it is necessary to characterize their antioxidant properties. Catalase degrading hydrogen peroxide is an important antioxidant enzyme. Nitrate reductase produces nitrite that can limit lipid oxidation by three indirect mechanisms that may operate simultaneously (Igene et al., 1985). Nitrite may act by: binding heme and preventing release of catalytic iron, binding non heme iron thus inhibiting catalysis, and/or stabilizing olefinic lipids against oxidation. Few studies are available in the literature on the catalase and nitrate reductase of staphylococci. In fact, catalase has been used to establish immunological relationships between different species of staphylococci (Rupprecht and Schleifer, 1979, Schleifer et al., 1979). The catalase of Staphylococcus aureus has been studied to establish its role in virulence (Mandell, 1975), and in resistance to hydrogen peroxide stress (Martin and Chaven, 1987). Concerning nitrate reductase, most papers deal with that of S. aureus (Burke and Lascelles, 1975, Burke and Lascelles, 1979), although one is about S. carnosus nitrate reductase (Neubauer and Götz, 1996).

The aim of this work was to study the production of nitrate reductase and catalase by different Staphylococcus species in order to understand their role in lipid oxidation during sausage ripening.

Section snippets

Growth conditions of the strains

The following staphylococci were studied: Staphylococcus carnosus (833, 836), Staphylococcus xylosus (831, 873, 16), Staphylococcus warneri (863), Staphylococcus saprophyticus (852). All the strains were isolated from dry sausage except S. xylosus 16 isolated from cheese (Montel et al., 1996).

Strains were grown in MC media: meat extract 10 g/l (Difco, USA), yeast extract 5 g/l (Difco, USA), NaCl 5 g/l, Na₂HPO₄ 1 g/l at pH 7.0, or in MC supplemented with 0.03% (w/v) or 0.2% (w/v) of KNO₃ (E.

Effect of different factors on the growth of staphylococci

The growth rate (μ) of staphylococci was highly influenced by the conditional incubation (Table 1). It varied also with the strains and a significant interaction was noticed between these two factors. This interaction is shown in Table 2. It was obvious that growth rate was higher in shaking than in static incubations for all the strains (Table 2). Under shaking, the strains S. xylosus 873, 16 and S. saprophyticus 852 had the highest growth rates. The growth rate depended on the interaction

Discussion

As facultative anaerobes, growth of staphylococci is better under aerobic conditions. In anaerobic conditions, the presence of nitrate resulted in a higher growth. This result agreed with those of Neubauer and Götz (1996) on S. carnosus and those of Burke and Lascelles (1975) on S. aureus.

The synthesis of nitrate reductase by staphylococci was influenced by the incubation conditions. In static incubation, the synthesis of the nitrate reductase was maximal during exponential growth phase. In

Acknowledgements

This work had been supported by EU program (FAIR-CT97-32227) entitled ‘Control of bioflavour and safety in northern and mediterranean fermented meat products’.

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Nitrosyl complex targets iron-sulphur proteins, and thus inactivates aconitase and ferrochelatase of Clostridium botulinum (Cammack et al., 1999). Nitrate reductase activities differ considerably among CNS species, particularly in S. carnosus, S. xylosus, S. equorum, S. lentus, S. simulans, S. sciuri, and S. succinus (Pantel et al., 1998; Talon et al., 1999; Fedtke et al., 2002; Mauriello et al., 2004; Gøtterup et al., 2007; Casaburi et al., 2007; Mainar and Leroy, 2015). Six CNS strains including S. hominis, S. saprophyticus, S. xylosus, and S. equorum were isolated from Koozh (a finger millet-based fermented food of South India) and revealed considerable potentialities to reduce nitrate into nitrite (Khusro et al., 2018b).
Coagulase-negative staphylococci (CNS) are one of the most pervasive heterogeneous groups of bacteria which are used as starter/adjunct cultures to enhance the aroma and texture of fermented foods. The organoleptic characteristics of fermented foods rely on disparate metabolic attributes of CNS. Nitrate reductase production from CNS improves sensory characteristics of foods by converting nitrate into nitrite. These bacteria utilize arginine via arginine deiminase pathway in the cytosol, and thus, play effective role in the generation of colour of fermented foods. Coagulase-negative Staphylococcus spp. develop flavour in foods by fermenting carbohydrates, converting amino acids, inducing β-oxidation of lipids, and secreting esterases. Additionally, the characteristic flavour of foods depends on the proteolytic and lipolytic properties of CNS strains too. Coagulase-negative staphylococci strains have revealed exemplary functional or probiotic traits by showing tolerance to acidic pH and bile, depicting adhesion characteristics, producing exopolysaccharide, and secreting therapeutic bacteriocins. Unfortunately, some CNS strains have shown antibiotics resistance, enterotoxins secretions, biogenic amine productions, haemolytic activities, and biofilm formations, thereby indicated the utilization of CNS on strain-by-strain basis. This review sheds light not only on the metabolic heterogeneity and techno-functional traits but also the safety and pathogenic aspects of fermented foods-associated CNS strains.
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Citation Excerpt :
Lactobacillus fermentum has also been shown to possess nitrate reductase activity under anaerobic conditions (Xu & Verstraete, 2001). In the process of making fermented meat sausage, the presence of nitrate-reducing bacteria ensures that nitrate from the curing salt is reduced to nitrite (Talon et al., 1999). Nitrite limits lipid oxidation (Cammack et al., 1999) and chemical reduction of nitrite leads to nitric oxide (NO), which reacts with myoglobin to form nitrosomyoglobin (MbFeIINO), imparting the typical coloration (bright red) of these products (Gøtterup et al., 2007).
The development of standardized and safe food products with the typical characteristics of each region is highly desirable and can be obtained by using native starter cultures that influence the flavor, texture, and color of fermented foods. Therefore, scientists have been employing various techniques for screening and characterizing native bacteria (lactic acid bacteria and Gram-positive catalase-positive cocci) for application in fermented meat sausage. The present review outlines in vitro assays that evaluate the potential application and safety aspects of native isolates and introduces emerging omics technologies applied to the microbiology of fermented meat sausage. Results from current research are presented, and the strengths and limitations of each assay are provided, with references indicating where further details can be obtained. In choosing the most appropriate in vitro method, it is necessary to consider the available analytical infrastructure, the sensitivity and selectivity of the assay, the time it takes to get the results, the ease of the assay, and the costs involved.

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International Journal of Food Microbiology

Abstract

Introduction

Section snippets

Growth conditions of the strains

Effect of different factors on the growth of staphylococci

Discussion

Acknowledgements

References (24)

Catalase

Effects of starter cultures on the formation of flavour compounds in dry sausage

Meat Sci.

Mechanisms by which nitrite inhibits the development of warmed-over flavor in cured meat

Food Chem.

Probing the structure of catalase HPII of Escherichia coli, a review

Gene

Biochemical activities of Micrococcaceae and their effects on the aromatic profiles and odours of a dry sausage model

Food Microbiol.

Aroma components from dried sausages fermented with Staphylococcus xylosus

Meat Sci.

Hydrolysis of esters by staphylococci

Int. J. Food Microbiol.

Production of esters by staphylococci

Int. J. Food Microbiol.

The aerobic growth and product formation of Lactobacillus, Leuconostoc, Brochothrix and Carnobacterium in batch cultures

Appl. Microbiol. Biotechnol.

Nitrate reductase system in Staphylococcus aureus wild type and mutants

J. Bacteriol.

Partial purification and some properties of the Staphylococcusaureus cytoplasmic nitrate reductase

J. Bacteriol.

Chemical analysis

Cited by (105)

Genomic and metabolic properties of Staphylococcus gallinarum FCW1 MCC4687 isolated from naturally fermented coconut water towards GRAS assessment

Metabolic heterogeneity and techno-functional attributes of fermented foods-associated coagulase-negative staphylococci

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Nitrite reduction in fermented meat products and its impact on aroma

The effect of cranberry pomace ethanol extract on the growth of meat starter cultures, Escherichia coli O157:H7, Salmonella enterica serovar Enteritidis and Listeria monocytogenes

Selection of native bacterial starter culture in the production of fermented meat sausages: Application potential, safety aspects, and emerging technologies