Characterization of Clostridium tyrobutyricum Strains Using Three Different Typing Techniques

Johanna Burtscher,Emmanuelle Arias-Roth,Konrad J. Domig,Franziska Küller,David Drissner,Matthias Dreier

doi:10.3390/microorganisms8071057

Abstract

Clostridium tyrobutyricum is well known as one of the main causative agents of severe cheese spoilage. The metabolism of this anaerobic bacterium during ripening leads to textural and sensory defects in cheese and consequential loss of product value. The potential to induce cheese spoilage, however, may vary among different strains of the same species. Therefore, a better understanding of the intra-species diversity of C. tyrobutyricum may be of practical relevance for the dairy industry. In the present study, we compared the ability of three typing techniques to differentiate 95 C. tyrobutyricum strains on the subspecies level: (1) repetitive element palindromic PCR (rep-PCR) fingerprinting combined with conventional agarose gel electrophoresis, (2) hexaplex-PCR followed by an automated capillary electrophoresis and (3) matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) typing. MALDI-TOF MS fingerprinting provided only moderate reproducibility and low discriminatory power. Both PCR-based methods were highly reproducible and discriminative, with hexaplex-PCR fingerprinting being slightly more discriminative than rep-PCR typing. Overall, a high intra-species diversity was observed among the tested strains, indicating that further investigations on the strain level may be of interest.

Highlights

Clostridium tyrobutyricum is recognized as one of the main causative agents of severe cheese spoilage
One of the main conclusions of the listed studies was that the species C. tyrobutyricum is relevant for cheese spoilage
Some of the techniques used in the studies listed above would be suitable for bacterial fingerprinting (e.g., randomly amplified polymorphic DNA (RAPD)-PCR, automated ribosomal intergenic spacer analysis (ARISA) or amplified 16S ribosomal DNA restriction analysis (16S ARDRA)), only a few studies have devoted special attention to the differentiation of butyric acid-producing clostridia on the subspecies or strain level [1]

Summary

Introduction

Clostridium tyrobutyricum is recognized as one of the main causative agents of severe cheese spoilage. The principal products of clostridial metabolic activity in cheese are gases (H2 and CO2) and organic acids (mainly butyric acid) [1] The presence of these metabolites leads to rancid off-flavors and pronounced textural defects in cheese, such as slits, cracks and undesired eyes [2,3]. Many studies have investigated the clostridial diversity in the dairy environment to gain a better understanding of how clostridia may affect cheese quality In this context, naturally occurring clostridial populations along the dairy supply chain have been characterized in depth on the species level [4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19]. Some of the techniques used in the studies listed above would be suitable for bacterial fingerprinting (e.g., randomly amplified polymorphic DNA (RAPD)-PCR, automated ribosomal intergenic spacer analysis (ARISA) or amplified 16S ribosomal DNA restriction analysis (16S ARDRA)), only a few studies have devoted special attention to the differentiation of butyric acid-producing clostridia on the subspecies or strain level [1]

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