基于宏基因组和宏转录组的发酵食品微生物研究进展

揭示传统发酵食品中微生物群落结构特征、演替变化规律和功能基因是多年来科学研究和工业生产共同关 注的焦点。近年来,高通量测序技术以其高效和相对廉价的优势,成为传统发酵食品微生物研究的重要工具。本文 从高通量测序技术的基因组和转录组层面的研究出发,综述了近8 年来宏基因组学与宏转录组学在研究发酵食品微 生物群落结构、相互作用及挖掘功能基因等方面的进展,并分析讨论其面临的主要问题和发展趋势,为未来发酵食 品的科学研究和工业生产提供一定参考。     

A review on selection of fermentative microorganisms for functional foods and beverages: the production and future perspectives

Fermentation has been employed as a traditional means of improving the shelf life and nutritional contents of foods, thus making fermented foods and beverages functional and therapeutic. Lactic acid bacteria (LAB) plays a major role in determining the health benefits of fermented milk and related products. This review takes into cognizance numerous investigations reporting certain microbial strains and mixes to contribute safety, quality, stability, health and organoleptic properties to fermented foods, due to increased consumers’ awareness of food products with health claims. A look was taken at selection criteria for the ideal fermentative microbial species and strains, their usefulness and prospective ‘OMICS‘-based approaches to elucidating fermentative complex communities in relation to their effects on fermented food products. Moreover, recommendations were given for improved fermentation of beverages and functional foods. It is projected that fermented foods and beverages will continually be a paramount in the global food and emerging functional food market.     

Non-gene-editing microbiome engineering of spontaneous food fermentation microbiota—Limitation control,design control,and integration

Non-gene-editing microbiome engineering (NgeME) is the rational design and control of natural microbial consortia to perform desired functions. Traditional NgeME approaches use selected environmental variables to force natural microbial consortia to perform the desired functions. Spontaneous food fermentation, the oldest kind of traditional NgeME, transforms foods into various fermented products using natural microbial networks. In traditional NgeME, spontaneous food fermentation microbiotas (SFFMs) are typically formed and controlled manually by the establishment of limiting factors in small batches with little mechanization. However, limitation control generally leads to trade-offs between efficiency and the quality of fermentation. Modern NgeME approaches based on synthetic microbial ecology have been developed using designed microbial communities to explore assembly mechanisms and target functional enhancement of SFFMs. This has greatly improved our understanding of microbiota control, but such approaches still have shortcomings compared to traditional NgeME. Here, we comprehensively describe research on mechanisms and control strategies for SFFMs based on traditional and modern NgeME. We discuss the ecological and engineering principles of the two approaches to enhance the understanding of how best to control SFFM. We also review recent applied and theoretical research on modern NgeME and propose an integrated in vitro synthetic microbiota model to bridge gaps between limitation control and design control for SFFM.     

Metaproteomics insights into traditional fermented foods and beverages

Traditional fermented foods and beverages (TFFB) are important dietary components. Multi‐omics techniques have been applied to all aspects of TFFB research to clarify the composition and nutritional value of TFFB, and to reveal the microbial community, microbial interactions, fermentative kinetics, and metabolic profiles during the fermentation process of TFFB. Because of the advantages of metaproteomics in providing functional information, this technology has increasingly been used in research to assess the functional diversity of microbial communities. Metaproteomics is gradually gaining attention in the field of TFFB research because it can reveal the nature of microorganism function at the protein level. This paper reviews the common methods of metaproteomics applied in TFFB research; systematically summarizes the results of metaproteomics research on TFFB, such as sauces, wines, fermented tea, cheese, and fermented fish; and compares the differences in conclusions reached through metaproteomics versus other omics methods. Metaproteomics has great advantages in revealing the microbial functions in TFFB and the interaction between the materials and microbial community. In the future, metaproteomics should be further applied to the study of functional protein markers and protein interaction in TFFB; multi‐omics technology requires further integration to reveal the molecular nature of TFFB fermentation.     

Cereal‐based fermented foods in developing countries: ancient foods for modern research

Cereal‐based fermented foods are major contributors to energy intake in developing countries (DC). Their microbiota is dominated by lactic acid bacteria and has been extensively investigated. Diversity studies have been facilitated by molecular methods enabling genotyping of isolates; the rapid development of ‘omics’ approaches should facilitate more comprehensive studies to describe the relation between microbial diversity, cell physiology and product characteristics. Also, the link between the food microbiota and health benefits, in particular in nutrition, should be investigated. There is a need to encourage researches in the field of DC cereal‐based foods in direction of more mechanistic approaches.     

传统甜醅细菌多样性及其与甜醅性质关系研究

以我国典型地区传统甜醅样品为对象，采用高通量扩增子测序技术开展甜醅样品细菌微生物区系研究，解析了甜醅细菌群落结构与甜醅性质关系。研究结果显示，甜醅样品中厚壁菌门（Firmicutes）含量最高。在属水平，以魏斯氏菌（Weissella）、乳杆菌（Lactobacillus）、葡萄球菌（Staphylococcus）等丰度最高。主坐标分析（PCo A）表明3种甜醅样品的细菌群落组成不同。利用Lefse分析发现了每种甜醅样品相关的生物标志分类单元，包括乳杆菌（Lactobacillus）和葡萄球菌（Staphylococcus）等。通过冗余分析（db-RDA）发现，甜醅细菌群落与甜醅性质（乳酸、黄酮、氨基酸、蛋白、淀粉、游离脂肪酸和还原糖含量）显著相关。基于16S r DNA预测分析，甜醅细菌功能主要涉及氨基酸转运和代谢及碳水化合物转运和代谢等。为了解传统全谷物发酵食品微生物资源多样性、利用微生物改善甜醅品质及今后工业化生产等奠定理论基础，也为其他发酵食品微生物多样性研究提供参考。     

Fermentation for tailoring the technological and health related functionality of food products

Abstract

Fermented foods are experiencing a resurgence due to the consumers’ growing interest in foods that are natural and health promoting. Microbial fermentation is a biotechnological process which transforms food raw materials into palatable, nutritious and healthy food products. Fermentation imparts unique aroma, flavor and texture to food, improves digestibility, degrades anti-nutritional factors, toxins and allergens, converts phytochemicals such as polyphenols into more bioactive and bioavailable forms, and enriches the nutritional quality of food. Fermentation also modifies the physical functional properties of food materials, rendering them differentiated ingredients for use in formulated foods. The science of fermentation and the technological and health functionality of fermented foods is reviewed considering the growing interest worldwide in fermented foods and beverages and the huge potential of the technology for reducing food loss and improving nutritional food security.     

豆豉和淡豆豉微生物组与功能成分研究进展

豆豉和淡豆豉作为中国传统大豆发酵食品,其中含有丰富的微生物资源,同时微生物组也可能对其中功能成分如异黄酮、γ-氨基丁酸等物质的含量产生影响。该文综述了组学技术在分析豆豉和淡豆豉发酵过程微生物变化研究中的应用,并对豆豉和淡豆豉中功能成分形成、转化和健康机制研究进行总结,同时根据临床药理学研究,两者中的部分功能成分对包括高血压、抑郁症等慢性病都显示出治疗潜力,通过收集有关其活性物质临床价值的信息,关注相关工艺改革进展,以期为进一步研究和开发豆豉和淡豆豉提供参考。     

Characterisation of lactic acid bacteria in spontaneously fermented camel milk and selection of strains for fermentation of camel milk

The microbial communities in spontaneously fermented camel milk from Ethiopia were characterised through metagenomic 16S rRNA sequencing and lactic acid bacteria were isolated with the goal of selecting strains suitable as starter cultures. The fermented camel milk microbiota was dominated either by Lactobacillales or by Enterobacteriaceae, depending on incubation temperature and the provider of the milk. Strains of species with a potential use as starter cultures i.e., Lactococcus lactis, Lactobacillus plantarum, and Pediococcus acidilactici, were isolated. Fast acidifiers of camel milk have been isolated from the species of Lc. lactis, P. acidilactici, and Streptococcus infantarius. Gram-negative and potentially pathogenic microorganisms were frequent in spontaneously fermented camel milk, indicating the need for improved hygiene in Ethiopian camel farms. The profiled microbiota of spontaneously fermented camel milk and the isolated LAB strains will significantly contribute towards improving food safety and food security in dry regions that depend on camel milk production.     

Identification of Microbial Profile of Koji Using Single Molecule,Real‐Time Sequencing Technology

Koji is a kind of Japanese traditional fermented starter that has been used for centuries. Many fermented foods are made from koji, such as sake, miso, and soy sauce. This study used the single molecule real‐time sequencing technology (SMRT) to investigate the bacterial and fungal microbiota of 3 Japanese koji samples. After SMRT analysis, a total of 39121 high‐quality sequences were generated, including 14354 bacterial and 24767 fungal sequence reads. The high‐quality gene sequences were assigned to 5 bacterial and 2 fungal plyla, dominated by Proteobacteria and Ascomycota, respectively. At the genus level, Ochrobactrum and Wickerhamomyces were the most abundant bacterial and fungal genera, respectively. The predominant bacterial and fungal species were Ochrobactrum lupini and Wickerhamomyces anomalus, respectively. Our study profiled the microbiota composition of 3 Japanese koji samples to the species level precision. The results may be useful for further development of traditional fermented products, especially optimization of koji preparation. Meanwhile, this study has demonstrated that SMRT is a robust tool for analyzing the microbial composition in food samples.     

ANTIOXIDANT PROPERTIES OF ETHANOLIC AND METHANOLIC EXTRACTS FROM MONASCUS-FERMENTED SOYBEANS

Soybeans are very common foods in oriental countries used as a meat substitute. Monascus species possess functional components and have been used on traditional fermented food. Solid-state fermentation of the soybean substrate by Monascus species is a new area of investigation. In the present study, methanolic extracts from three samples exhibited better antioxidant activity in the conjugated diene method, reducing power, scavenging ability on 1,1-diphenyl-2-picrylhydrazyl radicals and chelating ability on ferrous ions. Monascus species inoculated onto the soybean substrate contributed higher reducing power, scavenging and chelating abilities and higher amounts of phenol components than uninoculated soybean product. Overall, Monascus- fermented soybeans showed better antioxidant properties and can be developed as a new dietary supplement and functional food.

PRACTICAL APPLICATION

Soybeans and soy products are rich in isoflavones and are very common foods used as a meat substitute in oriental countries. Fungus Monascus has been used as a traditional fermented food and its metabolic products are also utilized as food pigments or biological agents in oriental countries for centuries. Many reporters showed that soy isoflavone aglycones and Monascus spp. metabolite exhibited physiological activities. In addition, previous studies showed a fermentation process using microorganisms to inoculate onto a nonsoluble material that can produce chemicals and enzymes. Therefore, the ethanolic and methanolic extracts from Monascus -fermented soybeans were evaluated for antioxidant properties, and the information obtained would be more valuable than that from soybeans to develop new functional foods.     

Prebiotics: Present and future in food science and technology

Because of its resident microbiota, the human colon is one of the body’s most metabolically active organs. The use of diet to fortify certain gut flora components is a popular current aspect of functional food sciences and prebiotics have a significant role. Prebiotics are selectively fermented ingredients that allow specific changes, both in the composition and/or activity in the gastrointestinal microbiota that confers benefits upon host well-being and health. Improved techniques for analysis of the gut microflora, new food manufacturing biotechnologies, and increased understanding of the metabolism of prebiotic inulin and oligosaccharides by probiotics are facilitating development. Such developments are leading us to the time when we will be able to rationally develop prebiotics for specific functional properties and health outcomes. Thus, this review will focus on the progress of prebiotics in food science and technology in understanding the important role of prebiotics in health, beginning at the rationale of gut microflora and interactions with prebiotics. Furthermore, the classification criteria, food applications and safety assessment of prebiotics as food ingredient is also discussed.     

多组学技术解析发酵水产食品风味形成机理研究进展

发酵水产食品风味形成机制复杂,制备和发酵过程中的原料、所用发酵剂以及设备和加工工艺中的多种微生物相互作用,导致形成的风味成分种类多样,从单一层面对不同发酵水产食品风味成分解析较为困难。近年来,通过利用对不同层面准确解析的组学技术,研究基因表达调控、蛋白质转录翻译及相互作用,并对代谢物进行定性及定量分析,可用于明确特征风味成分,揭示风味形成机制。因此,多组学技术可以用于动态检测发酵过程中水产品风味成分变化并解析风味形成机制、构建风味化合物代谢网络,探究风味相关微生物及酶作用关系。本文综述水产品发酵中风味形成的主要代谢途径、多组学技术应用于解析水产品发酵过程中风味形成的研究进展,以及多组学技术在发酵水产品风味研究中的重要作用。     

微生物发酵剂对四川风羊腿加工进程及产品特性的影响

传统发酵食品中微生物群落复杂,代谢途径多样,其中具有脱羧酶活性的微生物可代谢游离氨基酸形成潜在的危害因子—生物胺（biogenic amines,BAs）。BAs是一类具有生理活性的低分子碱性含氮化合物,主要由氨基酸脱羧酶脱羧产生。BAs根据其含氨量可分为单胺、二胺和多胺,根据其化学结构又可分为脂肪类胺、芳香类胺和杂环类胺,其主要形成途径包括微生物脱羧作用以及醛、酮氨基化和转胺作用。传统发酵食品中含有脱羧酶活性的乳酸菌、假单胞菌和肠杆菌等微生物是主要产胺菌。少量的BAs可以调节人体正常生理功能,但摄入过多则会导致中毒,甚至死亡。因此,传统发酵食品中的BAs问题一直备受关注。解析微生物多样性与BAs形成之间的关系,有利于探明发酵食品中BAs形成途径和机制,可以有效控制BAs的产生与积累,以期为提高传统发酵食品安全性及品质提供参考,保证食品安全。本文重点综述了发酵蔬菜、发酵豆制品、发酵乳制品、发酵肉制品以及发酵水产品等传统发酵食品中微生物多样性与BAs形成之间的相关性,明确了各种发酵食品中的主要产胺菌株,解析BAs形成机制,以期为提高传统发酵食品安全性及品质提供参考。

     

Tropical traditional fermented food,a field full of promise. Examples from the Tropical Bioresources and Biotechnology programme and other related French–Vietnamese programmes on fermented food

In tropical countries, traditional fermented foods are usually home‐made products obtained through spontaneous fermentation or backslopping. They are now facing an evolution aiming at responding to quality, safety and mass production issues. This requires acquisition of knowledge on raw materials, microbial ecosystems and fermentation processes. Vietnam is a laboratory for such studies as traditional fermented foods play an important role in the diet of Vietnamese and as these foods are very diverse. Among the most popular are nem chua (sausage reminding Thai Nham), dua muoi (cabbage reminding Kimchi), tom chua (shrimps) and the well‐known nuoc mam. The challenge for these products to enter the industrial era is to reach a level of quality and safety without losing their character. Beside the research for starters to elaborate these products, the world of fermented products is also a world of innovation and the microbial ecosystem of traditional products can also be used for the adaptation of fermented products from other regions, such as wines (from grape or other fruits), for cross‐cultural innovations such as soy‐yogurts, for technology transfer from one fermentation (soy sauce) to another one (fish sauce) and for evolution of traditional products towards higher nutritional qualities such as nem chua nam (replacing part of the sausage meat by mushrooms). Finally, these complex microbial ecosystems are a source of probiotic, antimicrobial compounds and biocatalysts, which can benefit health and improve food processes worldwide. After a presentation of Vietnamese traditional fermented foods, this article aims at illustrating the diverse applications of research on fermented products through examples obtained in past research and in the Tropical Bioresources and Biotechnology project.     

Using PacBio sequencing to investigate the bacterial microbiota of traditional Buryatian cottage cheese and comparison with Italian and Kazakhstan artisanal cheeses

Traditional fermented dairy foods including cottage cheese have been major components of the Buryatia diet for centuries. Buryatian cheeses have maintained not only their unique taste and flavor but also their rich natural lactic acid bacteria (LAB) content. However, relatively few studies have described their microbial communities or explored their potential to serve as LAB resources. In this study, the bacterial microbiota community of 7 traditional artisan cheeses produced by local Buryatian families was investigated using single-molecule, real-time sequencing. In addition, we compared the bacterial microbiota of the Buryatian cheese samples with data sets of cheeses from Kazakhstan and Italy. Furthermore, we isolated and preserved several LAB samples from Buryatian cheese. A total of 62 LAB strains (belonging to 6 genera and 14 species or subspecies) were isolated from 7 samples of Buryatian cheese. Full-length 16S rRNA sequencing of the microbiota revealed 145 species of 82 bacterial genera, belonging to 7 phyla. The most dominant species was Lactococcus lactis (43.89%). Data sets of cheeses from Italy and Kazakhstan were retrieved from public databases. Principal component analysis and multivariate ANOVA showed marked differences in the structure of the microbiota communities in the cheese data sets from the 3 regions. Linear discriminant analyses of the effect size identified 48 discriminant bacterial clades among the 3 groups, which might have contributed to the observed structural differences. Our results indicate that the bacterial communities of traditional artisan cheeses vary depending on geographic origin. In addition, we isolated novel and valuable LAB resources for the improvement of cottage cheese production.     

DNA-based, culture-independent strategies for evaluating microbial communities in food-associated ecosystems

Culture-independent molecular techniques are now available to study microbial ecosystems. They are opening interesting perspectives to problems related to composition and population dynamics of microbial communities in various environmental niches (e.g., soil, water) and foods. In fermented food products, estimates of true microbial diversity is often difficult chiefly on account of the inability to cultivate most of the viable bacteria. The increasing knowledge of gene sequences and the concomitant development of new culture-independent molecular techniques are providing new and effective tools to compare the diversity of microbial communities and to monitor population dynamics in minimally disturbed samples. In this review, recent advances in these techniques are reported. Possible applications to food-associated microbial ecosystems are emphasised.     

宏基因组测序技术在传统酿造食品微生物群落分析中应用研究进展

我国传统酿造食品发展历史悠久,种类丰富,以其独特的魅力深受消费者喜爱。传统酿造食品中微生物群落的多样性是影响产品风味、品质与安全的重要因素,因此微生物群落结构特征、演替变化和功能基因挖掘等成为近年来的研究热点。高通量测序技术以其通量高和结果准确等优势,成为传统酿造食品微生物群落研究中的重要工具。该文以基于高通量测序技术的宏基因组测序技术出发,综述了宏基因组测序技术在传统酿造食品微生物群落分析研究中的进展,并分析讨论其面临的主要问题和发展趋势,为酿造食品的科学研究和工业生产提供相关理论基础。     

发酵类食品中微生物鉴定方法及功能研究进展

发酵类食品是主要通过直接接种微生物而产生特殊风味或者将食物浸入含有微生物的臭卤水中,从而获得特殊食品风味的中国传统食品。其中,采用含有微生物的卤水进行发酵是常见的发酵方式。因此,微生物在该类食品中一直扮演着一种非常重要的角色,具有较高的研究价值。本文阐述了发酵类型的食品所需要的卤水中微生物的鉴定常用方法,以及卤水微生物与风味成分的产生之间的关系,同时论述了卤水中微生物在各种发酵食品制作过程中所起到的功能特性,包括提高豆制品的营养价值、延长保质期、去除有害成分等问题,并提出了较为可行的新型发酵类食品卤水改进方向,为进一步开发理想的食品微生物发酵剂,控制有害物质的残留、拓展相关有益微生物资源的研究提供较为明确的指导方向。