Elsevier

Food Microbiology

Volume 89, August 2020, 103434
Food Microbiology

Raw meat quality and salt levels affect the bacterial species diversity and community dynamics during the fermentation of pork mince

https://doi.org/10.1016/j.fm.2020.103434Get rights and content

Highlights

  • Normal meat fermentation is governed by L. sakei, L. curvatus prevails in DFD meat.

  • High salt favours staphylococci, without much effect on their species diversity.

  • Salt hurdle does not prevent Enterobacterales at a favourable pH for growth.

Abstract

Acidification level and temperature modulate the beneficial consortia of lactic acid bacteria (LAB) and coagulase-negative staphylococci (CNS) during meat fermentation. Less is known about the impact of other factors, such as raw meat quality and salting. These could for instance affect the growth of the pathogen Staphylococcus aureus or of Enterobacterales species, potentially indicative of poor fermentation practice. Therefore, pork batters from either normal or borderline quality (dark-firm-dry, DFD) were compared at various salt concentrations (0–4%) in meat fermentation models. Microbial ecology of the samples was investigated with culture-dependent techniques and (GTG)5-PCR fingerprinting of genomic DNA. Whilst Lactobacillus sakei governed the fermentation of normal meat, Lactobacillus curvatus was more prominent in the fermentation of the DFD meat variant. CNS were favoured during fermentation at rising salt concentrations without much effects on species diversity, consisting mostly of Staphylococcus equorum, Staphylococcus saprophyticus, and Staphylococcus xylosus. During fermentation of DFD meat, S. saprophyticus was less manifest than during that of normal meat. Enterobacterales mainly emerged in DFD meat during fermentation at low salt concentrations. The salt hurdle was insufficient to prevent Enterobacterales when acidification and initial pH were favourable for their growth.

Introduction

Fermented meats are produced based on the various metabolic actions of lactic acid bacteria (LAB; Ravyts et al., 2012) and catalase-positive cocci, in particular coagulase-negative staphylococci (CNS; Sánchez Mainar et al., 2017; Stavropoulou et al., 2018a). These valuable food products are part of a culinary legacy, offering nutritional value, gastronomic pleasure, and convenience (Leroy et al., 2015a; Leroy and Degreef, 2015). Within this framework of traditionality, there is a growing interest in some of the more artisan-type production features, such as a return to spontaneous fermentation set-ups rather than the use of starter cultures (Janssens et al., 2012; Leroy et al., 2013). As spontaneously fermented and often home-made fermented foods are becoming increasingly popular, the need for sufficient empirical know-how should not be underestimated (Pot and Leroy, 2017). Deviations from validated expertise and practice may indeed pose serious quality and safety concerns (Centers for Disease Control and Prevention, 2012).

As a first constraint, a satisfactory level of initial meat quality needs to be respected. Pork, for instance, comes in various gradings, ranging from pale-soft-exudative (PSE) to dark-firm-dry (DFD) quality. Long-term ante mortem stressors are culpable for the DFD defect (Faustman and Cassens, 1990; Adzitey and Huda, 2011), which is characterized by a high final pH (≥6.0) and, thus, an elevated risk of microbial spoilage (Newton and Gill, 1981; Lister, 1989; Adzitey and Huda, 2011; Prieto et al., 2014; Ponnampalam et al., 2017). Although the use of DFD meat for sausage fermentation is acceptable, the higher risk of spoilage may require the use of starter cultures for enhanced process control (Ruiz and Pérez Palacios, 2014).

Substandard processing with non-conform raw materials and procedures is particularly risky when coupled to misconceptions about the relevance of technologically important factors (Leroy et al., 2018). Contemporary meat processing is often seen as harmful to health and has generated a demand for milder production technologies (Leroy et al., 2015b). However, added sugar is mostly converted into lactic acid and helps to kickstart the fermentation by the LAB, whilst the presence of sufficient salt is required for proper gel formation and microbial stabilisation (Työppönen et al., 2003; Desmond, 2006; Hutkins, 2006; Ockerman and Basu, 2007). Too zealous reduction of added sugar and salt may thus affect both the microbial communities as well as the formation of texture and flavour in the end-products (Corral et al., 2013; Desmond, 2006).

Because the technological conditions, such as the pH and salt level, play a primordial role in the shaping of microbial consortia during meat fermentation (Janssens et al., 2013; Leroy et al., 2014, 2015b; Yıldırım Yalçın and Seker, 2015), any alteration in processing conditions needs to be carefully addressed. This concern is not only related to the potential growth of undesirable microorganisms, including pathogens and spoilage-inducing Enterobacterales, but also with respect to the technological microbiota (i.e., the LAB and CNS). On the level of the CNS communities for instance, Staphylococcus equorum and Staphylococcus xylosus overtake Staphylococcus saprophyticus, and especially Staphylococcus carnosus, in less acidic environments (Janssens et al., 2013; Stavropoulou et al., 2018b). Also, suboptimal meat fermentation may give rise to the pathogenic species Staphylococcus aureus (Messier et al., 1989; Stavropoulou et al., 2018a). Therefore, the purpose of the present study was to delineate between a safe operating space and malpractice when using different raw meat qualities (normal versus DFD) and salt levels (0–4%) in fermented meat models that were spontaneously fermented without added carbohydrates. With respect to the microbiota, focus was on LAB, catalase-positive cocci (in particular CNS), and Enterobacterales.

Section snippets

Meat model preparation and experimental set-up

Fermented meat models were used, as described previously (Stavropoulou et al., 2018a). An experimental set-up with two different raw meat qualities was performed (normal meat with a pH of 5.74 ± 0.01 and DFD meat with a pH of 6.02 ± 0.02). DFD meat was purchased from a local butcher based on colour and exudation level. Each sausage meat batter consisted of fresh pork mince [2 kg, 14% fat fraction; obtained from a local butcher (Brussels, Belgium) based on colour and exudation level], sodium

The effect of different salt levels on the bacterial community dynamics during the fermentation of pork of normal raw meat quality

In a first experiment, raw meat batters consisting of pork mince of a normal raw meat quality level were used. The initial aw of the unsalted batter was 0.979 ± 0.01 but decreased with increasing salt concentrations, and also as fermentation proceeded over time (Table S1). After 14 days of fermentation, the aw at 0%, 1%, 2%, 3%, and 4% of salt was 0.969 ± 0.01, 0.964 ± 0.01, 0.955 ± 0.01, 0.950 ± 0.01, and 0.948 ± 0.01, respectively (Table S1). The initial pH of the sausage meat batter was

Discussion

Meat fermentation is a well understood biotechnological process, carried out either spontaneously (artisan-scale) or starter culture-initiated with LAB and/or CNS (Hutkins, 2006). Yet, the species diversity of bacterial consortia within fermented meat matrices is known to vary according to the raw material used and the processing methods applied (Talon et al., 2007; Janssens et al., 2012; Ravyts et al., 2012; Stavropoulou et al., 2018a, 2018b). Consequently, more information on the ecological

Conclusions

Both the initial raw meat quality and the degree of salting can interfere with the growth of the microbiota during meat fermentation processes. This may not only affect the technologically important bacteria, in particular the LAB and CNS, but also cause outgrowth of potentially harmful ones, in particular members of the Enterobacterales group. The use of DFD meat specifically warrants for appropriate salting and sufficient control over the acidification. The latter may be especially

Declarations of competing interest

None.

Acknowledgements

Funding: This work was supported by the Research Council of the Vrije Universiteit Brussel (SRP7 and IOF342 projects), and in particular the Interdisciplinary Research Program IRP11 ‘Tradition and naturalness of animal products within a societal context of change’ and the Hercules Foundation (project UABR 09/004).

References (98)

  • G. Comi et al.

    Characterisation of naturally fermented sausages produced in the North-East of Italy

    Meat Sci.

    (2005)
  • S. Coppola et al.

    Microbial succession during ripening of Naples type salami, a Southern Italian fermented sausage

    Meat Sci.

    (2000)
  • S. Corral et al.

    Salt reduction in slow fermented sausages affects the generation of aroma active compounds

    Meat Sci.

    (2013)
  • E. Desmond

    Reducing salt: a challenge for the meat industry

    Meat Sci.

    (2006)
  • A.I. Doulgeraki et al.

    Characterization of the Enterobacteriaceae community that developed during storage of minced beef under aerobic or modified atmosphere packaging conditions

    Int. J. Food Microbiol.

    (2011)
  • A.I. Doulgeraki et al.

    Spoilage microbiota associated to the storage of raw meat in different conditions

    Int. J. Food Microbiol.

    (2012)
  • E.H. Drosinos et al.

    Characterization of the microbial flora from a traditional Greek fermented sausage

    Meat Sci.

    (2005)
  • E.H. Drosinos et al.

    Phenotypic and technological diversity of lactic acid bacteria and staphylococci isolated from traditionally fermented sausages in southern greece

    Food Microbiol.

    (2007)
  • S. Fonseca et al.

    Monitoring the bacterial population dynamics during the ripening of Galician chorizo, a traditional dry fermented Spanish sausage

    Food Microbiol.

    (2013)
  • M.C. García Fontán et al.

    Microbiological characteristics of “androlla”, a Spanish traditional pork sausage

    Food Microbiol.

    (2007)
  • M. García-Varona et al.

    Characterisation of Micrococcaceae isolated from different varieties of “chorizo”

    Int. J. Food Microbiol.

    (2000)
  • W. Geeraerts et al.

    Diversity of the dominant bacterial species on sliced cooked pork products at expiration date in the Belgian retail

    Food Microbiol.

    (2017)
  • A. Greppi et al.

    Monitoring of the microbiota of fermented sausages by culture independent rRNA based approaches

    Int. J. Food Microbiol.

    (2015)
  • L. Höll et al.

    Identification and growth dynamics of meat spoilage microorganisms in modified atmosphere packaged poultry meat by MALDI-TOF MS

    Food Microbiol.

    (2016)
  • L. Iacumin et al.

    Ecology and dynamics of coagulase-negative cocci isolated from naturally fermented Italian sausages

    Syst. Appl. Microbiol.

    (2006)
  • L. Iacumin et al.

    Catalase-positive cocci in fermented sausage: variability due to different pork breeds, breeding systems and sausage production technology

    Food Microbiol.

    (2012)
  • M. Janssens et al.

    Species diversity and metabolic impact of the microbiota are low in spontaneously acidified Belgian sausages with an added starter culture of Staphylococcus carnosus

    Food Microbiol.

    (2012)
  • M. Janssens et al.

    Community dynamics of coagulase-negative staphylococci during spontaneous artisan-type meat fermentations differ between smoking and moulding treatments

    Int. J. Food Microbiol.

    (2013)
  • M. Janssens et al.

    The use of nucleosides and arginine as alternative energy sources by coagulase-negative staphylococci in view of meat fermentation

    Food Microbiol.

    (2014)
  • J.M. Jay et al.

    Profile and activity of the bacterial biota of ground beef held from freshness to spoilage at 5–7 oC

    Int. J. Food Microbiol.

    (2003)
  • D.-W. Jeong et al.

    Safety and technological characterization of Staphylococcus equorum isolates from jeotgal, a Korean high-salt-fermented seafood, for starter development

    Int. J. Food Microbiol.

    (2014)
  • G. Landeta et al.

    Characterization of coagulase-negative staphylococci isolated from Spanish dry cured meat products

    Meat Sci.

    (2013)
  • M. Laranjo et al.

    Impact of salt reduction on biogenic amines, fatty acids, microbiota, texture and sensory profile in traditional blood dry-cured sausages

    Food Chem.

    (2017)
  • I. Lebert et al.

    Diversity of micro-organisms in environments and dry fermented sausages of French traditional small units

    Meat Sci.

    (2007)
  • F. Leroy et al.

    Convenient meat and meat products. Societal and technological issues

    Appetite

    (2015)
  • F. Leroy et al.

    Meat fermentation at the crossroads of innovation and tradition: a historical outlook

    Trends Food Sci. Technol.

    (2013)
  • F. Leroy et al.

    Elements of innovation and tradition in meat fermentation: conflicts and synergies

    Int. J. Food Microbiol.

    (2015)
  • F. Leroy et al.

    Fermented meats (and the symptomatic case of the Flemish food pyramid): are we heading towards the vilification of a valuable food group?

    Int. J. Food Microbiol.

    (2018)
  • A.M. Lindberg et al.

    Enterobacteriaceae found in high numbers in fish, minced meat and pasteurised milk or cream and the presence of toxin encoding genes

    Int. J. Food Microbiol.

    (1998)
  • G. Lizaso et al.

    Microbiological and biochemical changes during ripening of salchichon, a Spanish dry cured sausage

    Food Microbiol.

    (1999)
  • M.L. Martins et al.

    Genetic diversity of Gram-negative, proteolytic, psychrotrophic bacteria isolated from refrigerated raw milk

    Int. J. Food Microbiol.

    (2006)
  • E. Marty et al.

    Identification of staphylococci and dominant lactic acid bacteria in spontaneously fermented Swiss meat products using PCR-RFLP

    Food Microbiol.

    (2012)
  • G. Mauriello et al.

    Isolation and technological properties of coagulase negative staphylococci from fermented sausages of Southern Italy

    Meat Sci.

    (2004)
  • C. Molinero et al.

    The effects of extended curing on the microbiological, physicochemical and sensorial characteristics of “Cecina de León”

    Meat Sci.

    (2008)
  • K.G. Newton et al.

    The microbiology of DFD fresh meats: a review

    Meat Sci.

    (1981)
  • P.T. Olesen et al.

    Generation of flavor compounds in fermented sausages – the influence of curing ingredients, Staphylococcus starter culture and ripening time

    Meat Sci.

    (2004)
  • E. Papamanoli et al.

    Characterization of Micrococcaceae isolated from dry fermented sausage

    Food Microbiol.

    (2002)
  • E. Papamanoli et al.

    Characterization of lactic acid bacteria isolated from a Greek dry-fermented sausage in respect of their technological and probiotic properties

    Meat Sci.

    (2003)
  • V. Pisacane et al.

    Microbial analyses of traditional Italian salami reveal microorganisms transfer from the natural casing to the meat matrix

    Int. J. Food Microbiol.

    (2015)
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