Pascal Bonnarme

• Doctor of Philosophy
• Managing Director at French National Institute for Agriculture, Food, and Environment (INRAE)

Iron is involved in various microbial metabolisms and interactions and is an essential micronutrient for most microorganisms. This review focuses on the cheese ecosystem, in which iron is sparse (median concentration of 2.9 mg/kg based on a literature survey) and of limited bioavailability due to the presence of various metal-binding agents in the...

Iron is a vital micronutrient for nearly all microorganisms, serving as a co-factor in critical metabolic pathways. However, cheese is an iron-restricted environment. Furthermore, it has been demonstrated that iron represents a growth-limiting factor for many microorganisms involved in cheese ripening and that this element is central to many microb...

Investigating the diversity of a given species could give clues for the development of autochthonous starter cultures. However, few studies have focused on the intraspecies diversity of Lactobacillus delbrueckii strains, a technologically important lactic acid bacterium for the dairy industry. For this reason, Lactobacillus delbrueckii strains from...

Streptococcus thermophilus is of major importance for cheese manufacturing to ensure rapid acidification; however, studies indicate that intensive use of commercial strains leads to the loss of typical characteristics of the products. To strengthen the link between the product and its geographical area and improve the sensory qualities of cheeses,...

Meat represents an important protein source, even in developing countries, but its production is scarcely sustainable, and its excessive consumption poses health issues. An increasing number of Western consumers would replace, at least partially, meat with alternative protein sources. This review aims at: (i) depicting nutritional, functional, sens...

Moving to a more sustainable food system requires increasing the proportion of plant protein in our diet. Fermentation of plant product could thus be used to develop innovative and tasty food products. We investigated the impact of fermentation by synthetic microbial consortia (SMC) on the perception of pea protein-based gels, giving possible keys...

Aims The pea seeds (Pisum sativum) hosts a complex microbial community that is exposed to environmental constraints modulated by biotic, abiotic and anthropogenic factors. The aim of this study was to explore the effect of geographical origins on the composition of the microbial communities associated with pea seeds from storage locations in differ...

Studies of food microorganism domestication can provide important insight into adaptation mechanisms and lead to commercial applications. Penicillium roqueforti is a fungus with four genetically differentiated populations, two of which were independently domesticated for blue cheese-making, with the other two populations thriving in other environme...

Consumer demands for plant-based products have increased in recent years. However, their consumption is still limited due to the presence of off-flavor compounds, primarily beany and green notes, which are mainly associated with the presence of aldehydes, ketones, furans, and alcohols. To overcome this problem, fermentation is used as a lever to re...

The study of food microorganism domestication can bring important insights on adaptation mechanisms and have industrial applications. The Penicillium roqueforti mold is divided into four main populations, with two populations domesticated for blue-cheese making and two populations thriving in other environments. While most blue cheeses worldwide ar...

The use of pea protein is limited in food by the persistence of off-flavours. Fermentation by microbial consortia could be a way to circumvent this problem. The aim of our study was to investigate the adaptation potential and metabolic features of consortia as a function of the matrix composition (pea and/or milk proteins). Three designed consortia...

Ripening cultures containing fungi and bacteria are widely used in smear-ripened cheese production processes, but little is known about the biotic interactions of typical ripening microorganisms at the surface of cheese. We developed a lab-scale mini-cheese model to investigate the biotic interactions of a synthetic community that was composed of D...

Sodium reduction in the human diet is currently one of the main concerns for public health agencies and, consequently, has become a challenge for the food industries. In this study, the impact of reduced sodium chloride content (20%) or its partial substitution with potassium chloride in soft ("Camembert"-type) and semi-hard ("Reblochon"-type) chee...

Methionine deprivation induces growth arrest and death of cancer cells. To eliminate l-methionine we produced, purified, and characterized the recombinant pyridoxal 5'-phosphate (PLP)-dependent l-methionine γ-lyase (MGL)- BL929 from the cheese-ripening Brevibacterium aurantiacum Transformation of an Escherichia coli strain with the gene BL929 from...

Geotrichum candidum is a fungus-like yeast widely used as a starter culture for cheese ripening for its proteolytic and lipolytic activities and its contribution to the cheese flavours. The sequenced strain G. candidum CLIB 918 was isolated from cheese Pont-L'Evêque. This strain's ability to produce volatile compounds was compared to the ability of...

Smear-ripened cheeses develop a viscous, red-orange smear on their surfaces during ripening. For this reason, they are also called red-smear cheeses or bacterial surface-ripened cheeses. During the past decade, tremendous progress has been made in the understanding of the microbiology of the surface microbiota of these cheeses. This progress stems...

Yeasts play a crucial role in cheese ripening. They contribute to the curd deacidification, the establishment of acid-sensitive bacterial communities, and flavour compounds production via proteolysis and catabolism of amino acids (AA). Negative yeast-yeast interaction was observed between the yeast Yarrowia lipolytica 1E07 (YL1E07) and the yeast De...

The rheological and microstructural properties of pure pea, pure milk, and mixed pea/milk gels were investigated and compared. Three gelation techniques were used: a thermal treatment, an acid treatment employing glucono delta-lactone, and an enzyme treatment employing chymosin and transglutaminase. Considerable differences in the gels' rheological...

This study evaluated the ability of dairy matrices, different in composition (with and without fat) and structure (liquid and gel), to enhance microorganisms survival through digestion. The viability of three dairy microorganisms Streptococcus thermophilus, Brevibacterium aurantiacum and Hafnia alvei was measured during in vitro and in vivo digesti...

Surface-ripened cheeses host complex microbial communities responsible for the transformation of milk into cheese as well as the development of important properties in terms of texture, color and sensory perception. In this study, we used high-throughput amplicon sequencing to decipher the bacterial and fungal diversity of 60 cheeses belonging to 1...

Here, we report the draft genome sequence of Corynebacterium variabile Mu292, which was originally isolated from the surface of Munster, a French smear-ripened cheese. This genome investigation will improve our knowledge on the molecular determinants potentially involved in the adaptation of this strain during the Munster-type cheese manufacturing...

The microbial communities in cheeses are composed of varying bacteria, yeasts, and molds, which contribute to the development of their typical sensory properties. In situ studies are needed to better understand their growth and activity during cheese ripening. Our objective was to investigate the activity of the microorganisms used for manufacturin...

Many health authorities have targeted salt reduction in food products as a means to reduce dietary sodium intake due to the harmful effects associated with its excessive consumption. In the present work, we evaluated the effect of reducing sodium chloride (NaCl) content on the microbiological and biochemical characteristics of an experimental surfa...

Cheese ripening is a complex biochemical process driven by microbial communities composed of both eukaryotes and prokaryotes. Surface-ripened cheeses are widely consumed all over the world and are appreciated for their characteristic flavor. Microbial community composition has been studied for a long time on surface-ripened cheeses, but only limite...

Thirty-six microorganisms (twenty-one bacteria, twelve yeasts and three fungi) were isolated from surface-ripened cheeses and subjected to in vitro digestive stress. The approach mimicked gastric and/or duodenal digestion. Lactobacillus rhamnosus GG, Escherichia coli Nissle 1917 and Saccharomyces boulardii were used as reference strains. We studied...

Cheese ripening involves the activity of various bacteria, yeasts or molds, which contribute to the development of the typical color, flavor and texture of the final product. In situ measurements of gene expression are increasingly being used to improve our understanding of the microbial flora activity in cheeses. The objective of the present study...

Microbial communities living on cheese surfaces are composed of various bacteria, yeasts and molds that interact together, thus generating the typical sensory properties of a cheese. Physiological and genomic investigations have revealed important functions involved in the ability of microorganisms to establish themselves at the cheese surface. The...

Worldwide, the Food Industry uses a broad range of microorganism while making fermented products. Most of microorganisms are alive when the food is ingested; it therefore appears important to determine how such microorganisms react during the digestive transit as well as to characterize their potential impact on the large intestine. Our work aims a...

Due to their importance in flavouring capacities (e.g. production of volatile sulphur compounds (VSCs)), their role in redox regulation (e.g. production of thiols) and their possible role in microbial interactions, sulphur compounds play a key role in traditional fermented products. A better knowledge and control of their microbial production is th...

Yarrowia lipolytica, located at the frontier of hemiascomycetous yeasts and fungi, is an excellent candidate for studies of metabolism evolution. This yeast, widely recognized for its technological applications, in particular produces volatile sulfur compounds (VSCs) that fully contribute to the flavor of smear cheese. We report here a relevant glo...

The cheese microbiota contributes to a large extent to the development of the typical color, flavor, and texture of the final product. Its composition is not well defined in most cases and varies from one cheese to another. The aim of the present study was to establish procedures for gene transcript quantification in cheeses by reverse transcriptio...

Staphylococcus equorum subsp. equorum is a member of the coagulase-negative staphylococcus group and is frequently isolated from fermented food products and from food-processing environments. It contributes to the formation of aroma compounds during the ripening of fermented foods, especially cheeses and sausages. Here, we report the draft genome s...

Corynebacterium casei is one of the most prevalent species present on the surfaces of smear-ripened cheeses, where it contributes to the production of the desired organoleptic properties. Here, we report the draft genome sequence of Corynebacterium casei UCMA 3821 to provide insights into its physiology.

Hemiascomycetes are separated by considerable evolutionary distances and, as a consequence, the mechanisms involved in sulfur metabolism in the extensively studied yeast, Saccharomyces cerevisiae, could be different from those of other species of the phylum. This is the first time that a global view of sulfur metabolism is reported in the biotechno...

Among the numerous compounds involved in cheese aroma, sulfur compounds are of particular interest because of their very powerful odors and low perception thresholds. Nevertheless, little attention has been focused until now on the possible presence of polyfunctional thiols in cheese, even if these compounds have been found to be associated with th...

Despite their importance as potent odors that contribute to the aroma of numerous cheeses, S-methyl thioesters formation pathways have not been fully established yet. In a first part of our work, we demonstrated that Brevibacterium antiquum and Brevibacterium aurantiacum could produce S-methyl thioesters using short-chain fatty acids or branched-ch...

In this study, we combined metabolic reconstruction, growth assays, and metabolome and transcriptome analyses to obtain a global view of the sulfur metabolic network and of the response to sulfur availability in Brevibacterium aurantiacum. In agreement with the growth of B. aurantiacum in the presence of sulfate and cystine, the metabolic reconstru...

The yeast Yarrowia lipolytica has to develop dynamic metabolic adaptation mechanisms to survive within the cheese habitat. The availability of amino acids (AAs) is of major importance for microbial development and/or aroma production during cheese ripening. Using 2-D protein gel electrophoresis, we analyzed the adaptation mechanisms of Y. lipolytic...

To investigate the impact of Proteus vulgaris growth on a multispecies ecosystem and on volatile aroma compound production during cheese ripening. The microbial community dynamics and the production of volatile aroma compounds of a nine-species cheese ecosystem were compared with or without the presence of P. vulgaris in the initial inoculum. Prote...

The growth and aroma contribution of Microbacterium foliorum, Proteus vulgaris and Psychrobacter sp., some common but rarely mentioned cheese bacteria, were investigated in a cheese model deacidified by Debaryomyces hansenii during the ripening process. Our results show that these bacteria had distinct growth and cheese flavour production patterns...

The interactions that may occur between microorganisms in different ecosystems have not been adequately studied yet. We investigated yeast-bacterium interactions in a synthetic medium using different culture associations involving the yeast Yarrowia lipolytica 1E07 and two bacteria, Staphylococcus xylosus C2a and Lactococcus lactis LD61. The growth...

Multilocus sequence typing with nine selected genes is shown to be a promising new tool for accurate identifications of Brevibacteriaceae at the species level. A developed microarray also allows intraspecific diversity investigations of Brevibacterium aurantiacum showing that 13% to 15% of the genes of strain ATCC 9174 were absent or divergent in s...

The consumption of lactate and amino acids is very important for microbial development and/or aroma production during cheese ripening. A strain of Yarrowia lipolytica isolated from cheese was grown in a liquid medium containing lactate in the presence of a low (0.1×) or high (2×) concentration of amino acids. Our results show that there was a drama...

With the view to investigate the presence of thiols in cheese, the use of different methods of preparation and extraction with an organomercuric compound ( p-hydroxymercuribenzoate) enabled the isolation of a new compound. The analysis of cheese extracts by gas chromatography coupled with pulse flame photometry, mass spectrometry, and olfactometry...

The effect of cysteine on the ability of smear cheese-ripening bacteria (Brevibacterium linens and Arthrobacter spp) to produce volatile sulphur compounds (VSC) from methionine was studied. These bacteria were cultivated in a synthetic medium supplemented with various cysteine concentrations with or without methionine. Cultures with only cysteine s...

The formation of volatile sulfur compounds (VSC) in fermented food is a subject of interest. Such compounds are essential for the aroma of many food products like cheeses or fermented beverages, in which they can play an attractive or a repulsive role, depending on their identity and their concentration. VSC essentially arise from common sulfur-bea...

Yarrowia lipolytica is one of the yeasts most frequently isolated from the surface of ripened cheeses. In previous work, it has been shown that this yeast is able to convert l-methionine into various volatile sulfur compounds (VSCs) that may contribute to the typical flavors of several cheeses. In the present study, we show that Y. lipolytica does...

Twelve bacterial strains belonging to eight taxonomic groups: Brevibacterium linens, Microbacterium foliorum, Arthrobacter arilaitensis, Staphylococcus cohnii, Staphylococcus equorum, Brachybacterium sp., Proteus vulgaris and Psychrobacter sp., isolated from different surface-ripened French cheeses, were investigated for their abilities to generate...

Production of volatile sulphur compounds (VSC) was assessed in culture media supplemented with L-methionine or L-methionine/L-cysteine mixtures, using five cheese-ripening yeasts: Debaryomyces hansenii DH47(8), Kluyveromyces lactis KL640, Geotrichum candidum GC77, Yarrowia lipolytica YL200 and Saccharomyces cerevisiae SC45(3). All five yeasts produ...

A practical adaptation of the methylene blue reaction for hydrogen sulfide quantification was developed to perform microbial selection. Closed plate flasks containing a zinc-agar layer above the liquid microbial culture are proposed as a trap system where the H(2)S can be retained and then quantified by the methylene blue reaction. Using this quant...

The expression of genes possibly involved in L-methionine and lactate catabolic pathways were performed in Brevibacterium linens (ATCC9175) in the presence or absence of added L-methionine. The expression of 27 genes of 39 selected genes differed significantly in L-methionine-enriched cultures. The expression of the gene encoding L-methionine gamma...

DNA microarrays of 86 genes from the yeasts Debaryomyces hansenii, Kluyveromyces marxianus, and Yarrowia lipolytica were developed to determine which genes were expressed in a medium mimicking a cheese-ripening environment. These genes were selected for potential involvement in lactose/lactate catabolism and the biosynthesis of sulfur-flavored comp...

Cheese flavour is the result of complex biochemical transformations attributed to bacteria and yeasts grown in cheese curd. Volatile sulphur compounds (VSCs) are responsible for the characteristic aromatic notes of several cheeses, e.g., smear-ripened cheeses. The enzymatic degradation of L-methionine and subsequent formation of volatile sulfur com...

The enzymatic degradation of L-methionine and subsequent formation of volatile sulfur compounds (VSC) is believed to be essential for flavour development in cheese. L-methionine γ-lyase (MGL) can convert L-methionine to methanethiol (MTL) α-ketobutyrate and ammonia. The MGL gene encoding MGL was cloned from the type strain Brevibacterium linens ATC...

Volatile sulfur compounds (VSCs) are of major importance for flavour development in foodstuffs including cheeses. They primarily result from L-methionine degradation to methanethiol (MTL), a precursor for a variety of other VSCs. A plate assay based on double layer Petri dishes containing 5,5′-dithio-bis-2-nitrobenzoic acid (DTNB)—a chemical reacti...

Cheese flavour is the result of complex biochemical transformations attributed to bacteria and yeasts grown on the curd of smear-ripened cheeses. Volatile sulphur compounds (VSCs) are responsible for the characteristic aromatic notes of several cheeses. In the present study, we have assessed the ability of Kluyveromyces lactis, Kluyveromyces marxia...

Kluyveromyces lactis is one of the cheese-ripening yeasts and is believed to contribute to the formation of volatile sulfur compounds (VSCs) through degradation of L-methionine. L-methionine aminotransferase is potentially involved in the pathway that results in the production of methanethiol, a common precursor of VSCs. Even though this pathway ha...

Tracing experiments were carried out to identify volatile and nonvolatile l-methionine degradation intermediates and end products in the yeast Geotrichum candidum and in the bacterium Brevibacterium linens, both of which are present in the surface flora of certain soft cheeses and contribute to the ripening reactions. Since the acid-sensitive bacte...

Three cheese-ripening yeasts Kluyveromyces lactis, Geotrichum candidum and Debaryomyces hansenii were grown in a cheese slurry either individually or in association with the cheese-ripening bacterium Brevibacterium linens. Alcohols, aldehydes and esters were produced by the yeasts, and appeared in the early stages of ripening, whereas volatile sulp...

By its numerous properties and importance in cheese technology (production of colour, flavour, bacteriocins and resistance to salt) Brevibacterium linens is a major cheese ripening bacteria. However, the genetic approach of such biological functions has been hindered, up to now, by the lack of tools necessary to realise genetic modifications in thi...

Two mixtures of Propionibacterium freudenreichii commercial strains were tested as adjunct cultures in pasteurized milk Raclette cheese to investigate the ability of propionibacteria (PAB) to enhance flavor development. Cheese flavor was assessed by a trained sensory panel, and levels of free amino acids, free fatty acids, and volatile compounds we...

Volatile sulfur compounds (VSCs) are of major importance for flavor development in foodstuffs such as cheeses. Such compounds originate from the amino acid l-methionine, which can be degraded to methanethiol (MTL), a common precursor to a variety of other VSCs. A plate assay based on double-layer petri dishes containing 5,5'-dithio-bis-2-nitrobenzo...

The enzymatic degradation of l-methionine and the subsequent formation of volatile sulfur compounds (VSCs) are essential for the development of the typical flavor in cheese. In the yeast Yarrowia lipolytica, the degradation of l-methionine was accompanied by the formation of the transamination product 4-methylthio-2-oxobutyric acid. A branched-chai...

The enzymatic degradation of l-methionine and subsequent formation of volatile sulfur compounds (VSCs) is believed to be essential for flavor development in cheese. l-Methionine-γ-lyase (MGL) can convert l-methionine to methanethiol (MTL), α-ketobutyrate, and ammonia. The mgl gene encoding MGL was cloned from the type strain Brevibacterium linens A...

The aromatic potential of various cocultures of yeasts, Brevibacterium linens and lactic acid bacteria (LAB) was studied in cheese-based medium. Three yeasts (Debaryomyces hansenii, Geotrichum candidum and Kluyveromyces lochs) were cultivated in association with B. linens, in the presence or in the absence of LAB-added as the commercial lactic acid...

Volatile sulphur compounds (VSCs) production from L-methionine was studied in Lactococcus lactis. In vitro studies with radiolabelled L-methionine and resting cells of L. lactis revealed that L-methionine was initially converted to alpha-keto-gamma-methylthiobutyrate (KMBA) by a transamination reaction. A part of KMBA was subsequently chemically co...

Two Brevibacterium linens strains and the cheese-ripening yeast Geotrichum candidum were compared with regard to their ability to produce volatile sulfur compounds (VSCs) from three different precursors namely L-methionine, 4-methylthio-2-oxobutyric acid (KMBA) and 4-methylthio-2-hydroxybutyric acid (HMBA). All microorganisms were able to convert t...

The effect of the carbon source and l-methionine on the ability of Geotrichum candidum to produce volatile sulphur compounds (VSC) was studied. This yeast was cultivated in a synthetic medium supplemented with various carbon sources and l-methionine at different concentrations. Both glycerol and glucose significantly increased VSC production by G....

The biogeneration of sulfur compounds is an excellent model for illustrating the complexity of flavor production by microbial pathways. This paper will concentrate on the precursors of sulfur-containing flavor compounds. We will focus are discussion on sulfur compounds more commonly found in nature; such as, sulfates and sulfites, amino acids and p...

A study of the origins of volatile sulfur compounds isolated from soft cheeses during cheesemaking was presented. The degradation mechanisms were started by the deamination of methionine, and were necessary for the biosynthesis of flavors in cheeses. The results showed the effect of the pathways on the flavor quality changes in mixed cultures.

Five cheese-ripening yeasts (Geotrichum candidum, Saccharomyces cerevisiae, Kluyveromyces lactis, Yarrowia lipolytica and Debaryomyces hansenii) were compared with respect to their ability to generate volatile aroma compounds. K. lactis produced a variety of esters - ethylacetate (EA) being the major one - and relatively limited amounts of volatile...

L-methionine degradation products and catabolic enzymatic activities involved in methanethiol generation were investigated in Geotrichum candidum GcG. L-methionine was easily degraded by G. candidum and the transamination product, 4-methylthio-2-oxobutyric acid (KMBA), was found to transiently accumulate. In parallel, considerable L-methionine amin...

Volatile sulphur compounds are major flavouring compounds in many traditional fermented foods including cheeses. These compounds are products of the catabolism of L-methionine by cheese-ripening microorganisms. The diversity of L-methionine degradation by such microorganisms, however, remains to be characterized. The objective of this work was to c...

Several yeasts, Geotrichum candidum, Yarrowia lipolytica, Kluyveromyces lactis, Debaryomyces hansenii, and Saccharomyces cerevisiae, were studied for their ability to generate volatile sulfur compounds. These yeasts were cultivated on a synthetic culture medium supplemented with a biosynthetic precursor. With S-methylmethionine, dimethylsulfide (DM...

The synthesis of short chain S-methyl thioesters was investigated in Geotrichum candidum strain GcG. The results indicated the involvement of an enzymatic reaction in this microorganism that led to the synthesis of S-methyl thioacetate (MTA) when methanethiol and acetyl-CoA were used as substrates. MTA was generated from these substrates by enzymat...