Shotgun Metagenomics and Volatilome Profile of the Microbiota of Fermented Sausages

ABSTRACT Changes in the microbial gene content and abundance can be analyzed to detect shifts in the microbiota composition due to the use of a starter culture in the food fermentation process, with the consequent shift of key metabolic pathways directly connected with product acceptance. Meat fermentation is a complex process involving microbes that metabolize the main components in meat. The breakdown of carbohydrates, proteins, and lipids can lead to the formation of volatile organic compounds (VOCs) that can drastically affect the organoleptic characteristics of the final products. The present meta-analysis, performed with the shotgun DNA metagenomic approach, focuses on studying the microbiota and its gene content in an Italian fermented sausage produced by using a commercial starter culture (a mix of Lactobacillus sakei and Staphylococcus xylosus ), with the aim to discover the connections between the microbiota, microbiome, and the release of volatile metabolites during ripening. The inoculated fermentation with the starter culture limited the development of Enterobacteriaceae and reduced the microbial diversity compared to that from spontaneous fermentation. KEGG database genes associated with the reduction of acetaldehyde to ethanol (EC 1.1.1.1), acetyl phosphate to acetate (EC 2.7.2.1), and 2,3-butanediol to acetoin (EC 1.1.1.4) were most abundant in inoculated samples (I) compared to those in spontaneous fermentation samples (S). The volatilome profiles were highly consistent with the abundance of the genes; elevated acetic acid (1,173.85 μg/kg), ethyl acetate (251.58 μg/kg), and acetoin (1,100.19 μg/kg) were observed in the presence of the starters at the end of fermentation. Significant differences were found in the liking of samples based on flavor and odor, suggesting a higher preference by consumers for the spontaneous fermentation samples. Inoculated samples exhibited the lowest scores for the liking data, which were clearly associated with the highest concentration of acetic acid. IMPORTANCE We present an advance in the understanding of meat fermentation by coupling DNA sequencing metagenomics and metabolomics approaches to describe the microbial function during this process. Very few studies using this global approach have been dedicated to food, and none have examined sausage fermentation, underlying the originality of the study. The starter culture drastically affected the organoleptic properties of the products. This finding underlines the importance of starter culture selection that takes into consideration the functional characteristics of the microorganism to optimize production efficiency and product quality.

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Microbiological and Physicochemical Characterization of Small-Scale Cocoa Fermentations and Screening of Yeast and Bacterial Strains To Develop a Defined Starter Culture

Applied and Environmental Microbiology ◽

10.1128/aem.01144-12 ◽

2012 ◽

Vol 78 (15) ◽

pp. 5395-5405 ◽

Cited By ~ 86

Author(s):

Gilberto Vinícius de Melo Pereira ◽

Maria Gabriela da Cruz Pedrozo Miguel ◽

Cíntia Lacerda Ramos ◽

Rosane Freitas Schwan

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Citric Acid ◽

Dominant Species ◽

Starter Culture ◽

Physicochemical Characterization ◽

Cocoa Bean ◽

Small Scale ◽

Content Type ◽

Cocoa Bean Fermentation

ABSTRACTSpontaneous cocoa bean fermentations performed under bench- and pilot-scale conditions were studied using an integrated microbiological approach with culture-dependent and culture-independent techniques, as well as analyses of target metabolites from both cocoa pulp and cotyledons. Both fermentation ecosystems reached equilibrium through a two-phase process, starting with the simultaneous growth of the yeasts (withSaccharomyces cerevisiaeas the dominant species) and lactic acid bacteria (LAB) (Lactobacillus fermentumandLactobacillus plantarumwere the dominant species), which were gradually replaced by the acetic acid bacteria (AAB) (Acetobacter tropicaliswas the dominant species). In both processes, a sequence of substrate consumption (sucrose, glucose, fructose, and citric acid) and metabolite production kinetics (ethanol, lactic acid, and acetic acid) similar to that of previous, larger-scale fermentation experiments was observed. The technological potential of yeast, LAB, and AAB isolates was evaluated using a polyphasic study that included the measurement of stress-tolerant growth and fermentation kinetic parameters in cocoa pulp media. Overall, strainsL. fermentumUFLA CHBE8.12 (citric acid fermenting, lactic acid producing, and tolerant to heat, acid, lactic acid, and ethanol),S. cerevisiaeUFLA CHYC7.04 (ethanol producing and tolerant to acid, heat, and ethanol), andAcetobacter tropicalisUFLA CHBE16.01 (ethanol and lactic acid oxidizing, acetic acid producing, and tolerant to acid, heat, acetic acid, and ethanol) were selected to form a cocktail starter culture that should lead to better-controlled and more-reliable cocoa bean fermentation processes.

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Starter Culture Selection for Making Chinese Sesame-Flavored Liquor Based on Microbial Metabolic Activity in Mixed-Culture Fermentation

Applied and Environmental Microbiology ◽

10.1128/aem.00905-14 ◽

2014 ◽

Vol 80 (14) ◽

pp. 4450-4459 ◽

Cited By ~ 35

Author(s):

Qun Wu ◽

Jie Ling ◽

Yan Xu

Keyword(s):

Metabolic Activity ◽

Dominant Species ◽

Starter Culture ◽

Isobutyric Acid ◽

Spontaneous Fermentation ◽

Mixed Culture Fermentation ◽

Chinese Liquor ◽

Content Type ◽

Issatchenkia Orientalis ◽

Pichia Membranaefaciens

ABSTRACTSelection of a starter culture with excellent viability and metabolic activity is important for inoculated fermentation of traditional food. To obtain a suitable starter culture for making Chinese sesame-flavored liquor, the yeast and bacterium community structures were investigated during spontaneous and solid-state fermentations of this type of liquor. Five dominant species in spontaneous fermentation were identified:Saccharomyces cerevisiae,Pichia membranaefaciens,Issatchenkia orientalis,Bacillus licheniformis, andBacillus amyloliquefaciens. The metabolic activity of each species in mixed and inoculated fermentations of liquor was investigated in 14 different cocultures that used different combinations of these species. The relationships between the microbial species and volatile metabolites were analyzed by partial least-squares (PLS) regression analysis. We found thatS. cerevisiaewas positively correlated to nonanal, andB. licheniformiswas positively associated with 2,3-butanediol, isobutyric acid, guaiacol, and 4-vinyl guaiacol, whileI. orientaliswas positively correlated to butyric acid, isovaleric acid, hexanoic acid, and 2,3-butanediol. These three species are excellent flavor producers for Chinese liquor. AlthoughP. membranaefaciensandB. amyloliquefacienswere not efficient flavor producers, the addition of them alleviated competition among the other three species and altered their growth rates and flavor production. As a result, the coculture of all five dominant species produced the largest amount of flavor compounds. The result indicates that flavor producers and microbial interaction regulators are important for inoculated fermentation of Chinese sesame-flavored liquor.

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Interesting Starter Culture Strains for Controlled Cocoa Bean Fermentation Revealed by Simulated Cocoa Pulp Fermentations of Cocoa-Specific Lactic Acid Bacteria

Applied and Environmental Microbiology ◽

10.1128/aem.00594-11 ◽

2011 ◽

Vol 77 (18) ◽

pp. 6694-6698 ◽

Cited By ~ 33

Author(s):

Timothy Lefeber ◽

Maarten Janssens ◽

Frédéric Moens ◽

William Gobert ◽

Luc De Vuyst

Keyword(s):

Acetic Acid ◽

Lactic Acid ◽

Citric Acid ◽

Lactic Acid Bacteria ◽

Starter Culture ◽

Lactobacillus Fermentum ◽

Cocoa Bean ◽

Bacterial Strains ◽

Content Type ◽

Cocoa Bean Fermentation

ABSTRACTAmong various lactic acid bacterial strains tested, cocoa-specific strains ofLactobacillus fermentumwere best adapted to the cocoa pulp ecosystem. They fermented glucose to lactic acid and acetic acid, reduced fructose to mannitol, and converted citric acid into lactic acid and 2,3-butanediol.

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Benefits of the Use of Lactic Acid Bacteria Starter in Green Cracked Cypriot Table Olives Fermentation

Foods ◽

10.3390/foods9010017 ◽

2019 ◽

Vol 9 (1) ◽

pp. 17 ◽

Cited By ~ 2

Author(s):

Dimitrios A. Anagnostopoulos ◽

Vlasios Goulas ◽

Eleni Xenofontos ◽

Christos Vouras ◽

Nikolaos Nikoloudakis ◽

...

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Salt Concentration ◽

Starter Culture ◽

Total Phenolic ◽

Table Olives ◽

Acid Fermentation ◽

Spontaneous Fermentation ◽

Organoleptic Characteristics ◽

Nacl Concentration

Table olives are one of the most established Mediterranean vegetables, having an exponential increase consumption year by year. In the natural-style processing, olives are produced by spontaneous fermentation, without any chemical debittering. This natural fermentation process remains empirical and variable since it is strongly influenced by physicochemical parameters and microorganism presence in olive drupes. In the present work, Cypriot green cracked table olives were processed directly in brine (natural olives), using three distinct methods: spontaneous fermentation, inoculation with lactic acid bacteria at a 7% or a 10% NaCl concentration. Sensory, physicochemical, and microbiological alterations were monitored at intervals, and major differences were detected across treatments. Results indicated that the predominant microorganisms in the inoculated treatments were lactic acid bacteria, while yeasts predominated in control. As a consequence, starter culture contributed to a crucial effect on olives fermentation, leading to faster acidification and lower pH. This was attributed to a successful lactic acid fermentation, contrasting the acetic and alcoholic fermentation observed in control. Furthermore, it was established that inhibition of enterobacteria growth was achieved in a shorter period and at a significantly lower salt concentration, compared to the spontaneous fermentation. Even though no significant variances were detected in terms of the total phenolic content and antioxidant capacity, the degradation of oleuropein was achieved faster in inoculated treatments, thus, producing higher levels of hydroxytyrosol. Notably, the reduction of salt concentration, in combination with the use of starter, accented novel organoleptic characteristics in the final product, as confirmed from a sensory panel; hence, it becomes obvious that the production of Cypriot table olives at reduced NaCl levels is feasible.

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EFFECT OF MICROBIAL STARTER CULTURE ON SOME MICROBIAL PROPERTIES OF FERMENTED SAUSAGE

10.36295/asro.2020.231817 ◽

2020 ◽

Vol 23 (18) ◽

Author(s):

RanjbarShwan Abdulrahman ◽

Abduljabar Omer Qoja

Keyword(s):

Starter Culture ◽

Fermented Sausage ◽

Microbial Properties

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Native Yeasts and Lactic Acid Bacteria Isolated from Spontaneous Fermentation of Seven Grape Cultivars from the Maule Region (Chile)

Foods ◽

10.3390/foods10081737 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1737

Author(s):

Wendy Franco ◽

Sergio Benavides ◽

Pedro Valencia ◽

Cristian Ramírez ◽

Alejandra Urtubia

Keyword(s):

Lactic Acid ◽

Lactic Acid Bacteria ◽

Regional Growth ◽

Starter Culture ◽

Growth Conditions ◽

Microbial Evolution ◽

Spontaneous Fermentation ◽

Wine Production ◽

Grape Juices ◽

Grape Cultivars

Grapes are a source of native yeasts and lactic acid bacteria (LAB); however, the microbial make up is dependent on the grape cultivar and the regional growth conditions. Therefore, the aim of this study was to characterize the yeast and LAB in seven grape cultivars cultivated in Chile. Grape juices were fermented at 25 °C for 7 days. Samples were collected to analyze sugar, organic acids, and ethanol. Microbial evolution was measured with culture-dependent and molecular approaches. Then, a native isolated Candida oleophila was selected for further sequential fermentations with Saccharomyces cerevisiae. The grape cultivars in the Maule showed a diversity of non-Saccharomyces yeasts, with a greater diversity observed at the beginning of the fermentation. However, species from the Hansenasporia, Metschnikowia, Torulaspora, Lachancea, and Candida genera were detected after 7 days, suggesting tolerance to environments rich in ethanol, capability may be associated to the terroir studied, which is characterized by torrid weather and antique and traditional vineyards. The alcoholic fermentation negatively impacted the LAB population, and after 7 days only Leuconostoc mesenteroides was isolated. In the sequential fermentations, C. oleophila was able to produce fermented grape juices with <1.5 g/L glucose, 12.5% (v/v) alcohol, and low concentrations of malic (<1.00 g/L) and succinic (2.05 g/L) acids, while acetic acid reached values >0.3 (g/L). To our knowledge this is the first time C. oleophila has been reported as a potential starter culture for wine production. However, more studies are necessary to fully characterize the potential of C. oleophila on wine attributes.

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Applicability of Bacteriocinogenic Lactobacillus pentosus 31-1 as a Novel Functional Starter Culture or Coculture for Fermented Sausage Manufacture

Journal of Food Protection ◽

10.4315/0362-028x-73.2.292 ◽

2010 ◽

Vol 73 (2) ◽

pp. 292-298 ◽

Cited By ~ 12

Author(s):

GUORONG LIU ◽

MANSEL W. GRIFFITHS ◽

NAN SHANG ◽

SHANGWU CHEN ◽

PINGLAN LI

Keyword(s):

Sensory Quality ◽

Lipolytic Activity ◽

Starter Culture ◽

Meat Product ◽

Fatty Acid Analysis ◽

Fermented Sausage ◽

Lactobacillus Pentosus ◽

Fermented Sausages ◽

Challenge Tests ◽

Sensory Profiles

The technological feasibility of producing fermented sausages using the bacteriocin-producing Lactobacillus pentosus 31-1, isolated from a traditional Chinese fermented meat product (Xuanwei ham), was evaluated. Strain 31-1 was used both as a single starter and in coculture for manufacture of fermented sausages. The microbiological and physicochemical properties (color, texture, and sensory quality) and the production of bacteriocin during ripening of these products were compared with those characteristics of sausages produced with a commercial meat starter. Challenge tests were performed using Listeria innocua or Staphylococcus aureus as target strains. The addition of L. pentosus 31-1 can significantly reduce L. innocua and S. aureus populations during all ripening phases. Free amino acid and free fatty acid analysis suggested that strain 31-1 might have proteolytic and lipolytic activity. The use of this strain resulted in a final product with a brighter surface and better texture and sensory profiles. A maximum bacteriocin (pentocin 31-1) concentration of 640 AU/g was detected in homogenized sausages with added L. pentosus 31-1. The bacteriocin-producing strain L. pentosus 31-1 could be used as a novel functional starter culture or coculture for sausage fermentation.

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Biochemical and Genetic Bases of Indole-3-Acetic Acid (Auxin Phytohormone) Degradation by the Plant-Growth-Promoting Rhizobacterium Paraburkholderia phytofirmans PsJN

Applied and Environmental Microbiology ◽

10.1128/aem.01991-16 ◽

2016 ◽

Vol 83 (1) ◽

Cited By ~ 19

Author(s):

Raúl Donoso ◽

Pablo Leiva-Novoa ◽

Ana Zúñiga ◽

Tania Timmermann ◽

Gonzalo Recabarren-Gajardo ◽

...

Keyword(s):

Energy Source ◽

Gene Regulation ◽

Acetic Acid ◽

Plant Growth ◽

Plant Hormone ◽

Content Type ◽

Gene Functions ◽

Plant Growth Promoting ◽

Growth Promoting ◽

Indole 3 Acetic Acid

ABSTRACT Several bacteria use the plant hormone indole-3-acetic acid (IAA) as a sole carbon and energy source. A cluster of genes (named iac) encoding IAA degradation has been reported in Pseudomonas putida 1290, but the functions of these genes are not completely understood. The plant-growth-promoting rhizobacterium Paraburkholderia phytofirmans PsJN harbors iac gene homologues in its genome, but with a different gene organization and context than those of P. putida 1290. The iac gene functions enable P. phytofirmans to use IAA as a sole carbon and energy source. Employing a heterologous expression system approach, P. phytofirmans iac genes with previously undescribed functions were associated with specific biochemical steps. In addition, two uncharacterized genes, previously unreported in P. putida and found to be related to major facilitator and tautomerase superfamilies, are involved in removal of an IAA metabolite called dioxindole-3-acetate. Similar to the case in strain 1290, IAA degradation proceeds through catechol as intermediate, which is subsequently degraded by ortho-ring cleavage. A putative two-component regulatory system and a LysR-type regulator, which apparently respond to IAA and dioxindole-3-acetate, respectively, are involved in iac gene regulation in P. phytofirmans. These results provide new insights about unknown gene functions and complex regulatory mechanisms in IAA bacterial catabolism. IMPORTANCE This study describes indole-3-acetic acid (auxin phytohormone) degradation in the well-known betaproteobacterium P. phytofirmans PsJN and comprises a complete description of genes, some of them with previously unreported functions, and the general basis of their gene regulation. This work contributes to the understanding of how beneficial bacteria interact with plants, helping them to grow and/or to resist environmental stresses, through a complex set of molecular signals, in this case through degradation of a highly relevant plant hormone.

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Could new information influence attitudes to foods supplemented with edible insects?

British Food Journal ◽

10.1108/bfj-11-2016-0541 ◽

2017 ◽

Vol 119 (9) ◽

pp. 2027-2039 ◽

Cited By ~ 28

Author(s):

Fanny Barsics ◽

Rudy Caparros Megido ◽

Yves Brostaux ◽

Catherine Barsics ◽

Christophe Blecker ◽

...

Keyword(s):

A Priori ◽

Human Consumption ◽

Edible Insects ◽

Content Type ◽

Edible Insect ◽

Information Session ◽

Organoleptic Characteristics ◽

New Information ◽

Gender Information ◽

The Impact

Purpose Broader acceptance of entomophagy (i.e. human consumption of insects) will depend on factors that impact consumers’ perceptions of edible insects. The purpose of this paper is to examine how a broad-based information session would affect consumers’ perceptions and attitudes about an edible insect product. Design/methodology/approach During a taste testing session, preceded or followed by an information session about entomophagy, participants rated the organoleptic characteristics of two bread samples on nine-point hedonic scales. The two bread samples were identical, though one was faux-labelled as containing an insect product. Findings Generalised linear model (GLM) analysis showed effects of gender, information session exposure, entomophagy familiarity, and entomophagy experience on participants’ ratings of the samples. Wilcoxon-Mann-Whitney ranked sum tests showed that appearance, flavour, and overall liking were significantly better rated for the bread sample labelled as insect free by participants who attended the presentation a priori. Potential ways to improve information content and delivery in favour of encouraging dietary shifts are discussed. Practical implications This study shows that information about insect-based products could change consumers’ perceptions of such products. The results provide clues regarding how the food industry can adapt communication for target audiences. Originality/value Actual edible insect products were not used in this study. Paradoxically, it is the first to show the impact of an information session on the acceptability of edible insect products, by revealing participants’ perceptual expectations.

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The impact of sequencing depth on the inferred taxonomic composition and AMR gene content of metagenomic samples

10.1101/593301 ◽

2019 ◽

Author(s):

H. Soon Gweon ◽

Liam P. Shaw ◽

Jeremy Swann ◽

Nicola De Maio ◽

Manal AbuOun ◽

...

Keyword(s):

Data Processing ◽

Open Source ◽

Open Source Software ◽

River Sediment ◽

Taxonomic Composition ◽

Sequencing Depth ◽

Gene Content ◽

Metagenomic Data ◽

Software Pipeline ◽

Shotgun Metagenomics

ABSTRACTBackgroundShotgun metagenomics is increasingly used to characterise microbial communities, particularly for the investigation of antimicrobial resistance (AMR) in different animal and environmental contexts. There are many different approaches for inferring the taxonomic composition and AMR gene content of complex community samples from shotgun metagenomic data, but there has been little work establishing the optimum sequencing depth, data processing and analysis methods for these samples. In this study we used shotgun metagenomics and sequencing of cultured isolates from the same samples to address these issues. We sampled three potential environmental AMR gene reservoirs (pig caeca, river sediment, effluent) and sequenced samples with shotgun metagenomics at high depth (∼200 million reads per sample). Alongside this, we cultured single-colony isolates ofEnterobacteriaceaefrom the same samples and used hybrid sequencing (short- and long-reads) to create high-quality assemblies for comparison to the metagenomic data. To automate data processing, we developed an open-source software pipeline, ‘ResPipe’.ResultsTaxonomic profiling was much more stable to sequencing depth than AMR gene content. 1 million reads per sample was sufficient to achieve <1% dissimilarity to the full taxonomic composition. However, at least 80 million reads per sample were required to recover the full richness of different AMR gene families present in the sample, and additional allelic diversity of AMR genes was still being discovered in effluent at 200 million reads per sample. Normalising the number of reads mapping to AMR genes using gene length and an exogenous spike ofThermus thermophilusDNA substantially changed the estimated gene abundance distributions. While the majority of genomic content from cultured isolates from effluent was recoverable using shotgun metagenomics, this was not the case for pig caeca or river sediment.ConclusionsSequencing depth and profiling method can critically affect the profiling of polymicrobial animal and environmental samples with shotgun metagenomics. Both sequencing of cultured isolates and shotgun metagenomics can recover substantial diversity that is not identified using the other methods. Particular consideration is required when inferring AMR gene content or presence by mapping metagenomic reads to a database. ResPipe, the open-source software pipeline we have developed, is freely available (https://gitlab.com/hsgweon/ResPipe).

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