- Email: [email protected]
Foodomics for investigations of food toxins
Accepted Manuscript Title: Foodomics for investigations of food toxins Author: Dina Reˇsetar Sandra Kraljevi´c Paveli´c Djuro Josi´c PII: DOI: Reference:
S2214-7993(15)00073-9 http://dx.doi.org/doi:10.1016/j.cofs.2015.05.004 COFS 54
To appear in:
Please cite this article as: Reˇsetar, D., Paveli´c, S.K., Josi´c, D.,Foodomics for investigations of food toxins, COFS (2015), http://dx.doi.org/10.1016/j.cofs.2015.05.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Highlights: Food contamination coming from bacterial toxins, mycotoxins and toxins from eucariotic algae
ip t
Poisoning by toxins coming from contaminated food
cr
Use of foodomics methods for identification and quantification of food borne pathogens and
Ac ce p
te
d
M
an
us
food toxins
1 Page 1 of 17
Foodomics for investigations of food toxins
ip t
Dina Rešetar1, Sandra Kraljević Pavelić1, Djuro Josić1,2*
University of Rijeka, Department of biotechnology, Radmile Matejčić 2, 51000 Rijeka, Croatia
2
Warren Alpert Medical School, Brown University, Rhode Island, Providence RI, USA
us
cr
1
an
* Corresponding author: Djuro Josić
M
University of Rijeka
Ac ce p
51000 Rijeka
te
Radmile Matejčić 2
d
Department of biotechnology
Croatia
e-mail: [email protected] , [email protected] tel: +385 51 584 560
fax: +385 51 584 599
2 Page 2 of 17
Abstract: New nutritional trends and globalization of food market are substantially increasing food-borne outbreaks that remain a world-wide problem. Indeed, changes in consumers’ behavior, available
ip t
production methods, food processing approaches, climate changes and microbial resistance bring some weak points within the food production and distribution lines where foodomics methods and
cr
novel protocols have been increasingly used to ensure food safety within the whole food production
us
and distribution line. In this paper, foodomics technologies are described for precise identification of food pathogens and their toxins even on the strain/subtype level. In the near future further
an
improvements in the methodologies and protocols should be mainly directed to generation of new databases, better throughput and standardization while increased food monitoring within the food
M
production and distribution lines will be growingly demanded.
Keywords: Food toxins, bacterial endo- and exotoxins, mycotoxins, toxins from algae, foodomics,
Ac ce p
te
d
toxin identification, toxin quantitation
3 Page 3 of 17
Highlights: Food contamination coming from bacterial toxins, mycotoxins and toxins from eucariotic algae
ip t
Poisoning by toxins coming from contaminated food
cr
Use of foodomics methods for identification and quantification of food borne pathogens and
Ac ce p
te
d
M
an
us
food toxins
4 Page 4 of 17
1. Introduction Recent topics in food research deal with arising food-related problems and topics such as obesity and unhealthy diets, food allergies, but also with health benefits of food ingredients or growing problems
2, 3].
[DR1]New
ip t
in industrial food production and distribution related to the microbiological aspect of food safety [1, nutritional trends suggest consume of the fresh, pre-packaged, raw food, the food
cr
with low concentration of salt, dry products and exotic ingredients and foods from all over the world.
us
These trends and globalization of food market are substantially increasing food-borne outbreaks and food safety remains a world-wide problem [4]. Food safety as a matter of global importance
an
prompted the World Health Organization (WHO) to launch an initiative Global Burden of Foodborne Diseases aimed to globally map and estimate disease burden associated with unsafe food [5].
M
Several issues might be nowadays identified in the field of food microbial safety that include changes in consumers’ behavior, changes in available production methods (free, outdoor and organic
d
production or increasing herd size in industrial production), food processing approaches (mildly
te
preserved foods, lack of understanding of the preservation systems used in traditional ethnic foods), climate changes and microbial resistance [6]. These are weak points within the food production and
Ac ce p
distribution lines where food pathogens remain only partially destroyed, re-enter and adapt to food production and processing conditions, and therefore grow and produce toxic compounds. Food safety has to be ensured and remains a responsibility of all participants involved in the production chain [7].
Bacteria, fungi and microalgae may produce toxins in foods while leaving the food appearance, odor or flavor unaltered. Intoxication occurs after ingestion of such food products [7]. Bacteria can in some instances, produce toxins in the intestine of the host, causing toxico-infections. Since toxin producing bacteria and molds are basically ubiquitous, strict preventive measures during food preparation, storage and transportation up to the final product are the best practice to follow as prevention of toxico-infections [8]. Crucial food safety parameters therefore, remain complex 5 Page 5 of 17
problems inlcuding spore forming bacteria, thermostable bacterial toxins and mycotoxins, biofilm forming microorganisms, and marine biotoxins [6, 7]. This article brings an overview of the pathogenic bacteria, eukaryotic food pathogens and their food toxins with a particular focus on
ip t
recent advances in foodomics methods for investigations of food toxins. 2. Food toxins in foodborne illnesses
cr
2.1. Pathogenic bacteria and their food toxins
us
Lipopolysaccharide bacterial endotoxins are structural components of outer bacterial membrane, moderately toxic, heat stable, and fatal to animals only in large doses. In vivo, endotoxins are rarely
an
released during bacterial growth but always after autolysis or external lysis and phagocytic digestion of bacterial cells. Every bacterial endotoxin structure consists of three regions: highly conserved Lipid
M
A, R polysaccharide (core polysaccharide common to all members of a bacterial genus) and O polysaccharide (attached to the core polysaccharide repeating oligosaccharide subunits). Lipid A is
d
toxic while polysaccharide side chains are nontoxic even though immunogenic. Most of the work on
te
the chemical structure of endotoxin has been done with species of Salmonella and E. coli [9].
Ac ce p
On the other side Gram-positive and Gram-negative bacteria secrete the most powerful known human poisons as exoproteins (enterotoxins, neurotoxins, leukocidins and hemolysins) that act at the site of bacterial growth or spread through host’s organism towards target organs or cells [8]. A less common, but equally important from a food safety point of view, is food poisoning as a result of ingestion of pre-formed bacterial toxins or bacteria producing toxins that continue to grow in the host gut, i.e. food-borne outbreaks caused by Bacillus cereus, Clostridium botulinum, Clostridium perfringens and Staphylococcus aureus exotoxins (figure 1.) [10]. Heat stable exoproteins cannot be inactivated by thermal processing of food while heat labile botulinum toxin can be easily denatured by heat and acid or proteolyzed by proteolytic enzymes. Bacterial toxins may have a range of complex activities in the human body, which might be further
6 Page 6 of 17
complicated by individual susceptibility to toxins where the amounts of ingested toxin influence the onset and severity of the symptoms [9, 11]. Furthermore, almost nothing is known about effects resulting from ingestion of low or moderate levels of exotoxins [8].
ip t
Staphylococcal enterotoxins (SE) are a group of potent gastrointestinal protease-tolerant single chain exoproteins, that are pyrogenic toxins resistant to heat [12, 13]. As shown recently, S. aureus can
cr
produce 23 diverse enterotoxins but most frequently staphylococcal illnesses are generated by the
us
best characterized enterotoxins SEA (80%) and SEB (10%), and their most common way for food contamination is due to poor hygiene during the production process [13]. Enterotoxigenic strains of
an
E. coli produce both heat-labile and/or heat-stable enterotoxins, the most common cause of acute watery diarrhea in developing countries where an increasing degree of complexity and new virulence
M
factors was assessed in recent years [14]. B. cereus, another food pathogen, produces heat-labile diarrheagenic nonhemolytic enterotoxin Nhe [15] and/or hemolytic enterotoxin HBL [16] in the small
d
intestine of the host causing food poisoning symptoms [17]. The same species of B. cereus can
te
produce emetic heat-stable exotoxin cereulide during growth on the food [18]. Preformed cereulide is resistant to high and low pH, proteolysis, and high temperatures, generally it survives cooking and
Ac ce p
causes outbreaks of food intoxication with similar symptoms to food poisoning with staphylococcal and C. perfringens enterotoxins [19]. In food-borne botulism, the botulinum heat-labile pro-exotoxin is produced by proteolytic and non-proteolytic C. botulinum, C. baratii and C. butyricum. Extracellular proteases or proteolytic enzymes in the hosts gut cleave the single pro-exotoxin chain and convert it to a more toxic di-chain form [10]. This form of exotoxin is the well-known deadly botulinum toxin. It is extremely potent neurotoxin that is stable for days [20]. 2.2. Eukaryotic food pathogens and their food toxins The mycotoxin producing pathogenic fungi and toxin producing microalgae are eukaryotic food pathogens with the highest toxigenic potential. Large part of secondary metabolites produced by pathogenic funghi Aspergillus, Fusarium, Penicilium genera under favorable environmental 7 Page 7 of 17
conditions include mycotoxins: aflatoxins, fumonisins, trichothecenes, ochratoxins, patulin and zearalenone. Until now, it has been proved that mycotoxins incite autoimmune illnesses, allergic reactions, gastrointestinal and kidney disorders, some of them bear teratogenic, carcinogenic and
ip t
mutagenic properties. Frequently, co-occurrence of several mycotoxins in cereal has been reported with potential synergistic toxic effects. These substances enter the food chain since majority of
cr
mycotoxins are relatively heat-stable. Even though 400 of these substances and their metabolites have been identified so far, only few of them are studied in details. At last, marine toxins as
us
structurally diverse group of food toxins include eukaryotic algae toxins known to accumulate in fish
an
and shellfish [7, 21]. 3. Identification of food toxins
M
In the last decade rapid and sensitive screening methods for monitoring of food toxins were introduced: bioassays, molecular biology and/or immunological techniques, gel-clot technique,
d
turbidimetric technique, biosensors and chromatographic as well as mass spectrometry-based
te
methods [7, 22-24]. Bioassays are used when fast analysis of potentially contaminated food is required [25] and/or for assessment of cytotoxic action of food toxin in cell cultures. These methods
Ac ce p
provide a fast toxicity assessment however they are not suitable for characterization of toxins because of their relatively low specificity. More specific methods are required for determination of exact toxin origin and identification, i.e. highly sensitive and high-throughput molecular methods such as real time polymerase chain reaction (RT-PCR) and RT quantitative PCR (RT-qPCR). These are golden standards for detection of exotoxin coding genes in strains isolated from contaminated foods, i.e. staphylococcal enterotoxin coding se genes or cereulide synthetase coding structural genes cesA and cesB where exact quantification of relative transcript levels are possible [26]. Limitations of these methods include the need for previous isolation of food-poisoning strains from food where only presence or absence of toxin encoding genes are possible and no information on these genes’ expression or their respective levels in food are available. Further on, immunological testing by use of
8 Page 8 of 17
commercially developed kits is a method of choice only for detection of bacterial toxin effects (antibody production) in the host organism. For example, ELISA double antibody sandwich and automated enzyme-linked fluorescent immunoassay (ELFA) are used for detection and semi-
ip t
quantification of Staphylococus, Clostridium exotoxins and cyanotoxin microcystin [24, 27]. Although, immunological assays are officially approved they lack specificity and sensitivity and produce false
cr
positives and false negatives signal due to cross-reactions. Again, the method is reliable for assessment of toxin presence while almost nothing can be deduced about its active/inactive form.
us
The same problem may appear when polyacrylamide gel electrophoresis (PAGE) methods combined with Western blotting detection of certain toxins are used [28]. However, 2D-electrophoretic
an
fractionation, proved to be highly useful in secretomics studies of food pathogenic fungi, i.e.
M
extracellular proteome analysis of Asspergillus terreus [29] and Fusarium graminearum [30]. 3.1. Foodomics methods
d
Food safety nowadays, remains focused on prevention and monitoring of potentially harmful effects
te
growingly correlated with a number of different microbial subtypes and sub strains that may produce toxic products not easily detectable by previously described, standard methods [31]. [DR2]Therefore,
Ac ce p
development of specific, rapid and sensitive methods for analytical determination of food pathogens and identification and quantification of preformed toxins in food has attracted major interest of scientists, in particular those developing high-throughput foodomics methodologies (genomics, proteomics and metabolomics).
For example, next generation sequencing and whole-genome sequencing (WGS) technologies, capillary gel electrophoresis (CGE) and combination of molecular techniques with other analytical approaches (i.e. multiplex PCR-based procedure followed by capillary gel electrophoresis with laserinduced fluorescence detection, multiplex-PCR-CGE-LIF) have been used for detection of new foodborne pathogens with the power to differentiate closely related serotypes [32-37]. [DR3]Furthermore,
highly sensitive and accurate mass spectrometry soft ionization methods, namely 9 Page 9 of 17
matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and electrospray ionization mass spectrometry (ESI-MS) are also growingly used in the field of food safety assessment [38]. These methods are frequently combined with chromatography, in order to improve
ip t
their sensitivity [39]. MALDI TOF MS is already accepted as a promising analytical method for bacterial food pathogen identification and characterization. MALDI spectra, known as MALDI TOF MS
cr
fingerprints, provide characteristic intact or trypsin digested ribosomal or intracellular protein and peptide profiles of whole bacterial cells, whole cell suspensions or cell extracts that might be easily
us
correlated with databases [40, 41]. Furthermore, interpretation of MALDI TOF MS fingerprints is used for differentiation of bacterial strains without the necessity for proteins identification: representative
an
peaks patterns suffice for conclusion about species, genus or even strain level [42]. Still, MALDI– based identification remains highly dependent to the number of well-characterized food pathogen
M
protein biomarker sequences available in proteome databases. So far, a number of reference databases [43-45] have been developed that paved the way for introduction of the MALDI-based MS
d
identification of microbes in food as a routine. The major bottleneck for its further implementation in
te
food monitoring remains the problem of incorrect interpretation due to presence or absence of
Ac ce p
unique peak patterns [46]. The only possible solution to this problem is further development of MALDI TOF fingerprint reference databases that will host a growing number of correct MALDI TOF fingerprint profiles (for example reference library SpectraBank) coupled to bioinformatics pipelines and user friendly software [47]. This might provide an accurate tool for fast and precise identification since no statistical analysis is needed for data interpretation, and the costs of materials and staff in the investigation of food poisoning outbreaks without a known causative agent may be substantially lower in a long-term. Another interesting approach for fast discrimination of bacteria, without need for sample preparation, is paper spray mass spectrometry ambient ionization combined with principal component statistical analysis of obtained phospholipid spectra [48]. Both MALDI and ESI-MS are appropriate for the study of intact proteins and non-covalent protein complexes such as complexes of botulinum neurotoxin and neurotoxin associated proteins, which 10 Page 10 of 17
contain hemoglutinins and non-toxin non-hemogglutinins [49]. Recently, a new ultraviolet photodissociation (UVPD) online liquid chromatographic tandem mass spectrometry (LC MS/MS) strategy was implemented into analysis of structural variants of E. coli lipid A on the Orbitrap mass
ip t
spectrometer [50]. Since food is a complex matrix, sample preparation protocols have been developed to facilitate
cr
isolation and identification of food toxins: immunocapture, precipitation, filtration, solid phase
us
extraction (SPE), solid-liquid extraction (SLE), SDS-PAGE and different chromatographic methods. All these separation steps remain a challenge, i.e. cereulide extraction from foods with high lipid
an
concentration. This is why multidimensional liquid chromatography tandem mass spectrometry (MDLC MS/MS) has been developed as a faster alternative to laborious sample preparation of highly
M
complex samples [32]. In recent years, much effort was put on optimization of extraction techniques QuEChERS or dispersive liquid-liquid microextraction (DLLME) for multi-mycotoxin separation and
d
isolation although SPE, SLE [38]. QuEChERS combined with multi-mycotoxin liquid chromatography
te
tandem mass spectrometry (LC-MS/MS) screening method enabled simultaneous identification of 36 mycotoxins in wines [51] and up to 56 mycotoxins in animal feed [52]. Graphitised carbon solid phase
Ac ce p
desalting clean-up step has been used more recently for sample preparation prior to hydrophilic interaction liquid chromatography (HILIC) UPLC–MS/MS detection of paralytic shellfish toxins. This approach represents a major technical breakthrough and holds potential as new routine monitoring method for hydrophilic marine toxins [53]. Moreover, nanoscale LC-MS/MS may also provide identification of proteolytically digested exoproteins isolated from low-protein samples [54]. Recently, combined protocols for in vivo protein targeting, including stable isotope in culture (SILAC) with extraction and proteolytical strategies and mass spectrometry were introduced [55]. Such an approach was used for example, during MALDI-TOF MS or LC-MS/MS multiple reaction monitoring (MRM) for identification and quantification of cerulide toxin with a synthetic cereulide employed as internal standards. Such approach is superior to previousl methods that relayed on surrogate standards, i.e. antibiotic valinomycin [56]. At last, isobaric tandem mass tags coupled to ion-mobility 11 Page 11 of 17
spectrometry (IMS) protocols are emerging alternatives to label-free approaches and mass difference tags for quantitative proteomics and separation of isobaric exopeptides [57]. Another versatile, robust and cost-effective foodomic method is capillary electromigration (CE),
ip t
sometimes combined to mass spectrometry (CE-MS). It provides fast, efficient and automated separation with small sample volumes and low consumption of solvents and reagents [58]. Micellar
cr
electrokinetic chromatography (MEKC) is a modified CE, where an analyte is separated by differential
us
partitioning between micellar pseudo-stationary phase and mobile aqueous phase, providing separation of neutral analytes [59]. MEKC proved to be more precise technique compared to HPLC in
an
identification and quantification of patulin mycotoxin [60] and detection of emetic toxin cereulide from contaminated rice samples [61].
M
In conclusion, foodomics technologies have matured in recent years and may robustly provide precise identification of food pathogens and their toxins even on the strain/subtype level from highly
d
complex food samples. In the near future further improvements in the methodologies and protocols
te
should be mainly directed to generation of new databases, better throughput and standardization while increased food monitoring within the food production and distribution lines will be growingly
Ac ce p
demanded.
4. References
(*)[DR5][1] Cifuentes A: Food analysis and foodomics, J. Chromatogr. A 2009, 1216, 7109. [2] Simó C, Domínguez-Vega E, Marina ML, García MC, Dinelli G, Cifuentes A: CE-TOF MS analysis of complex protein hydrolyzates from genetically modified soybeans. A tool for foodomics. Electrophoresis 2010, 31, 1175-1183. [3] Ibáñez C, Valdés A, García-Cañas V, Simó C, Celebier M, Rocamora L, Gómez A, Herrero M, Castro M, Segura-Carretero A et all.: Global foodomics strategy to investigate the health benefits of dietary constituents. J. Chromatogr A 2012, 1248, 139-153.[DR6] [4] Akhtar S., Sarker MR, Hossain A: Microbiological food safety: a dilemma of developing societies. Crit Rev Microbiol 2012, 40, 348-359. [5]WHO, 2006: http://www.who.int/foodsafety/foodborne_disease/ferg/en/ 12 Page 12 of 17
[6]Havelaar AH, Brul S, Jong A, Jonge R, Zwietering MH, Kuile BH: Future challenges to microbial food safety. Int J Food Microbiol 2010, 139, 79-94. [7] Giacometti J., Buretić Tomljanović A, Josic Dj: Application of proteomics and metabolomics for investigation of food toxins. Food Res Int 2013, 54, 1042–1051.
ip t
(*)[DR7][8] Rajković A: Microbial toxins and low level of foodborne exposure. Trends Food Sci Tech 2014, 38, 149-157. [9] Henkel JS, Baldwin MR, Barbieri JT: Toxins from bacteria. EXS. 2010, 100, 1–29.
cr
[10] EFSA: The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks in 2013. EFSA Journal 2015, 13, 1-162.
us
[11] Peck MW, Stringer SC, Carter AT: Clostridium botulinum in the post-genomic era. Food Microbiol 2011, 28, 183-191.
an
[12] Grumanna D, Nübelb U, Bröker BM: Staphylococcus aureus toxins – their functions and genetics. Infection, Genetics and Evolution 2014, 21, 583–592.
M
[13] Gustafson JE, Muthaiyan A, Dupre JM, Ricke SC: Staphylococcus aureus and understanding the factors that impact enterotoxin production in foods: A review. Food Control 2014, DOI: 10.1016/j.foodcont.2014.10.016.
d
[14] Fleckenstein JM, Hardwidge PR, Munson GP, Rasko DA, Sommerfelt H, Steinsland H: Molecular mechanisms of enterotoxigenic Escherichia coli infection. Microbes Infec 2010, 12, 89-98.
te
[15] Ceuppens S, Rajkovic A, Hamelink S, van De Wiele T, Boon N, Uyttendaele M: Enterotoxin production by Bacillus cereus under gastrointestinal conditions and their immunological detection by commercially available kits. Foodborne Pathog 2012, 9, 1130–1136.
Ac ce p
[16] Senesi S, Ghelardi E: Production, secretion and biological activity of Bacillus cereus enterotoxins. Toxins 2010, 2, 1690–1703. [17] Ceuppens S, Uyttendaele M, Drieskens K, Rajkovic A, Boon N, van DeWiele T: Survival of Bacillus cereus vegetative cells and spores during in vitro simulation of gastric passage. J Food Prot 2012, 75, 690–694. [18] Marxen S, Stark TD, Frenzel E, Rütschle A, Lücking G, Pürstinger G, Pohl EE, Scherer S, EhlingSchulz M, Hofmann T: Chemodiversity of cereulide, the emetic toxin of Bacillus cereus. Anal Bioanal Chem 2015, 407, 2439-2453. [19] Stenfors Arnesen LP, Fagerlund A, Granum PE: From soil to gut: Bacillus cereus and its food poisoning toxins. FEMS Microbiol Rev 2008, 32, 579-606. [20] Hou HJM: Analysis of botulinum neurotoxin detection by mass spectrometry in forensic samples. J Forensic Res 2013, 4, 2-6. (**)[DR8][21] Giacometti J, Josic Dj: Foodomics in microbial safety. Trend Anal Chem 2013, 52, 16-22. [22] Adley, CC: Past, present and future of sensors in food production. Foods 2014, 3, 491-510. 13 Page 13 of 17
(*)[DR9][23] Yeni F, Acar S, Polat ÖG, Soyer Y, Alpas H: Rapid and standardized methods for detection of foodborne pathogens and mycotoxins on fresh produce. Food Control 2014, 40, 359-367. [24] Zhang C, Zhang J: Current techniques for detecting and monitoring algal toxins and causative harmful algal blooms. J Environ Anal Chem 2015, DOI: 10.4172/JREAC.1000123.
ip t
[25] Hu DL, Omoe K, Shimoda Y, Nakane A, Shinagawa K: Induction of emetic response to staphylococcal enterotoxins in the House Musk Shrew (Suncus murinus). Infect Immun 2003, 71, 567–570.
cr
[26] Ikeda T, Tamate N, Yamaguchi K, Makino S: Mass outbreak of food poisoning disease caused by small amounts of staphylococcal enterotoxins A and H. Appl Environ Microbiol 2005, 71, 2793–2795.
us
(*)[DR10][27] Zhao X, Lin CW, Wang J, Oh DH: Advances in rapid detection methods for foodborne pathogens. J Microbiol Biotechnol 2014, 24, 297–312.
an
[28] Hennekinne JA, Ostyn A, Guillier F, Herbin S, Prufer AL, Dragacci S: How should staphylococcal food poisoning outbreaks be characterized? Toxins 2010, 2, 2106–2116. [29] Kim Y, Nanadakumar MP, Marten MR: The state of proteome profiling in the fungal genus Aspergillus. Brief Funct Genomic Proteomic 2008, 7, 87-94.
d
M
[30] Yang FEN, Jensen JD, Svensson B, Jørgensen HJL, Collinge DB, Finnie C: Secretomics identifies Fusarium graminearum proteins involved in the interaction with barley and wheat. Molecular Plant Pathology 2012, 13, 445-453.
te
[31] García-Cañas V, Simó C, Herrero M, Ibáñez E, Cifuentes A: Present and future challenges in food analysis. Foodomics. Anal. Chem. 2012, 84, 10150-10159.[DR11][32] Cunsolo V, Muccilli V, Saletti R, Foti S: Mass spectrometry in food proteomics: a tutorial. J Mass Spectrom 2014, 49, 768-784.
Ac ce p
(*)[DR12][33] Bakker HC, Allard MW, Bopp D, Brown EW, Fontana J, Iqbal Z, Kinney A, Limberger R, Musser KA, Shudt M, et al: Rapid whole-genome sequencing for surveillance of Salmonella enterica serovar enteritidis. Emerg Infect Dis 2014, 20, 1306–1314. [34] Bergholz TM, Moreno Switt AI, Wiedmann M: Omics approaches in food safety: fulfiling the promise? Trends Microbiol 2014, 22, 275-281. [35] García-Cañas V, González R, Cifuentes A: Combined use of molecular techniques and capillary electrophoresis in food analysis. TRAC-Trend Anal.Chem. 2004, 23, 637-643. [36] Alarcón B, García-Cañas V, Cifuentes A, González R, Aznar R: Simultaneous and sensitive detection of three food-borne pathogens by multiplex PCR, capillary gel electrophoresis and laser induced fluorescence. J. Agric. Food Chem. 2004, 52, 7180-7186. [37] García-Cañas V, Cifuentes A: Detection of microbial food contaminants and their products by capillary electromigration techniques. Electrophoresis 2007, 28, 4013-4030.[DR13] (**)[DR14][38] Berthiller F, Brera C, Crews C, Iha MH, Krska R, Lattanzio VMT, MacDonald S, Malone RJ, Maragos C, Solfrizzo M, Stroka J, Whitaker TB: Developments in mycotoxin analysis: an update for 2013-2014. World Mycotoxin J 2015, 8, 5-36. 14 Page 14 of 17
(**)[DR15][39] Hird SJ, Lau BPY, Schuhmacher R, Krska R: Liquid chromatography-mass spectrometry for the determination of chemical contaminants in food. Trends Anal Chem 2014, 59, 59–72. [40] Sauer S, Kliem M: Mass spectrometry tools for the classification and identification of bacteria. Nat Rev Microbiol 2010, 8, 74–82.
ip t
(*)[DR16][41] Schumann P, Maier T: MALDI-TOF mass spectrometry applied to classification and identification of bacteria. Method Microbiol 2014, 41, 275–306.
cr
(**)[DR17][42] Novais A, Sousa C, de Dios Caballero J, Fernandez-Olmos A, Lopes J, Ramos H, Coque TM, Cantón R, Peixe L: MALDI-TOF mass spectrometry as a tool for the discrimination of high-risk Escherichia coli clones from phylogenetic groups B2 (ST131) and D (ST69, ST405, ST393). Eur J Clin Microbol 2014, 33, 1391-1399.
an
us
[43] Böhme K, Fernández-No IC, Barros-Velázquez J, Gallardo JM, Canas B, Calo-Mata P: Comparative analysis of protein extraction methods for the identification of seafood-borne pathogenic and spoilage bacteria by MALDI-TOF mass spectrometry. Anal Methods 2010, 2, 1941– 1947.
M
[44] Böhme K, Fernández-No IC, Barros-Velázquez J, Gallardo JM, Calo-Mata P, Canas B: Species differentiation of seafood spoilage and pathogenic gram-negative bacteria by MALDI-TOF mass fingerprinting. J Proteome Res 2010, 9, 3169–3183.
d
[45] AlMasoud N, Nicolaou N, Goodacre R: Optimization of matrix assisted desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) for the characterization of Bacillus and Brevibacillus species. Anal Chimica Acta 2014, 840, 49–57.
Ac ce p
te
[46] Fernandez-No IC, Bohme K, Gallardo JM, Barros-Velazquez J, Canas B, Calo-Mata P: Differential characterization of biogenic amine-producing bacteria involved in food poisoning using MALDI-TOF mass fingerprinting. Electrophoresis 2010, 31, 1116-1127. [47] Bohme K, Fernandez-No IC, Barros-Velazquez J, Gallardo JM, Canas B, Calo-Mata P: SpectraBank: an open access tool for rapid microbial identification by MALDI-TOF MS fingerptinting. Electrophoresis 2012, 33, 2138-2142. [48] Hamid AM, Jarmusch AK, Pirro V, Pincus DH, Clay BG, Gervasi G, Cooks RG: Rapid discrimination of bacteria by paper spray mass spectrometry. Anal Chem 2014, 86, 7500–7507. [49] McFarland MA, Andrzejewski D, Musser SM, Callahan JH: Platform for identification of Salmonella serovar differentiating bacterial proteins by top-down mass spectrometry: S. Typhimurium vs S. Heidelberg. Anal Chem 2014, 86, 6879–6886. [50] O’Brien JP, Needham BD, Henderson JC, Nowicki EM, Trent MS, Brodbelt, JS: 193 nm ultraviolet photodissociation mass spectrometry for the structural elucidation of lipid A compounds in complex mixtures. Anal Chem 2014, 86, 2138–2145. (*)[51] Pizzutti IR, De Kok A, Scholten J, Righi LW, Cardoso CD, NecchiRohers G, Da Silva RC: Development, optimization and validation of a multimethod for the determination of 36
15 Page 15 of 17
mycotoxins in wines by liquid chromatography-tandem mass spectrometry. Talanta 2014, 129, 352363. [52] Dzuman Z, Zachariasova M, Lacina O, Veprikova Z, Slavikova P, Hajslova J: A rugged highthroughput analytical approach for the determination and quantification of multiple mycotoxins in complex feed matrices. Talanta 2014, 121, 263-272.
cr
ip t
(**)[DR18][53] Boundy MJ, Selwood AI, Harwood DT, McNabb PS, Turner AD: Development of a sensitive and selective liquid chromatography–mass spectrometry method for high throughput analysis of paralytic shellfish toxins using graphitised carbon solidphase extraction. Journal of Chromatogr A 2015, 1387, 1–12.
us
[54] Janga KS, Sweredoskib MJ, Grahamb RLJ, Hessb S, Clemons Jr WM: Comprehensive proteomic profiling of outer membrane vesicles from Campylobacter jejuni. J Proteomics 2014, 98, 90–98.
an
[55] Bauer T, Stark T, Hofmann T, Ehling-Schulz M: Development of a stable isotope dilution analysis for the quantification of the Bacillus cereus toxin cereulide in foods. J Agric Food Chem 2010, 58, 1420–1428.
M
(**)[56] Muratovic AZ, Tröger R, Granelli K, Hellenäs KE: Quantitative analysis of cereulide toxin from Bacillus cereus in rice and pasta using synthetic cereulide standard and 13C6-cereulide standard—A short validation study. Toxins 2014, 6, 3326-3335.
d
[57] Lanucara F, Holman SW, Gray CJ, Eyers CE: The power of ion mobility-mass spectrometry for structural characterization and the study of conformational dynamics. Nature Chemistry 2014, 6, 281–294.
Ac ce p
te
[58] Castro-Puyana M, García-Canas V, Simó C, Cifuentes A: Recent advances in the application of capillary electromigration methods for food analysis and foodomics. Electrophoresis 2012, 33, 147– 167. [59] Hancu G, Simon B, Rusu A, Mircia E, Gyéresi Á: Principles of micellar electrokinetic capillary chromatography applied in pharmaceutical analysis. Adv Pharm Bull 2013, 3, 1–8. [60] Murillo-Arbizu M, González-Penas E, Amézqueta, S: Patulin in apple juice for infants. Food Chem Toxicol 2010, 48, 2429–2434. [61] Oh MH, Cox JM: Development and application of a centrifugation-plating method to study the biodiversity of Bacillus species in rice products. Food Control 2010, 21, 652–665.
16 Page 16 of 17
ip t cr us an
Ac ce p
te
d
M
Figure 1. Distribution of food-borne outbreaks per causative agent in the EU during 2013 [10].
17 Page 17 of 17
S2214-7993(15)00073-9 http://dx.doi.org/doi:10.1016/j.cofs.2015.05.004 COFS 54
To appear in:
Please cite this article as: Reˇsetar, D., Paveli´c, S.K., Josi´c, D.,Foodomics for investigations of food toxins, COFS (2015), http://dx.doi.org/10.1016/j.cofs.2015.05.004 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Highlights: Food contamination coming from bacterial toxins, mycotoxins and toxins from eucariotic algae
ip t
Poisoning by toxins coming from contaminated food
cr
Use of foodomics methods for identification and quantification of food borne pathogens and
Ac ce p
te
d
M
an
us
food toxins
1 Page 1 of 17
Foodomics for investigations of food toxins
ip t
Dina Rešetar1, Sandra Kraljević Pavelić1, Djuro Josić1,2*
University of Rijeka, Department of biotechnology, Radmile Matejčić 2, 51000 Rijeka, Croatia
2
Warren Alpert Medical School, Brown University, Rhode Island, Providence RI, USA
us
cr
1
an
* Corresponding author: Djuro Josić
M
University of Rijeka
Ac ce p
51000 Rijeka
te
Radmile Matejčić 2
d
Department of biotechnology
Croatia
e-mail: [email protected] , [email protected] tel: +385 51 584 560
fax: +385 51 584 599
2 Page 2 of 17
Abstract: New nutritional trends and globalization of food market are substantially increasing food-borne outbreaks that remain a world-wide problem. Indeed, changes in consumers’ behavior, available
ip t
production methods, food processing approaches, climate changes and microbial resistance bring some weak points within the food production and distribution lines where foodomics methods and
cr
novel protocols have been increasingly used to ensure food safety within the whole food production
us
and distribution line. In this paper, foodomics technologies are described for precise identification of food pathogens and their toxins even on the strain/subtype level. In the near future further
an
improvements in the methodologies and protocols should be mainly directed to generation of new databases, better throughput and standardization while increased food monitoring within the food
M
production and distribution lines will be growingly demanded.
Keywords: Food toxins, bacterial endo- and exotoxins, mycotoxins, toxins from algae, foodomics,
Ac ce p
te
d
toxin identification, toxin quantitation
3 Page 3 of 17
Highlights: Food contamination coming from bacterial toxins, mycotoxins and toxins from eucariotic algae
ip t
Poisoning by toxins coming from contaminated food
cr
Use of foodomics methods for identification and quantification of food borne pathogens and
Ac ce p
te
d
M
an
us
food toxins
4 Page 4 of 17
1. Introduction Recent topics in food research deal with arising food-related problems and topics such as obesity and unhealthy diets, food allergies, but also with health benefits of food ingredients or growing problems
2, 3].
[DR1]New
ip t
in industrial food production and distribution related to the microbiological aspect of food safety [1, nutritional trends suggest consume of the fresh, pre-packaged, raw food, the food
cr
with low concentration of salt, dry products and exotic ingredients and foods from all over the world.
us
These trends and globalization of food market are substantially increasing food-borne outbreaks and food safety remains a world-wide problem [4]. Food safety as a matter of global importance
an
prompted the World Health Organization (WHO) to launch an initiative Global Burden of Foodborne Diseases aimed to globally map and estimate disease burden associated with unsafe food [5].
M
Several issues might be nowadays identified in the field of food microbial safety that include changes in consumers’ behavior, changes in available production methods (free, outdoor and organic
d
production or increasing herd size in industrial production), food processing approaches (mildly
te
preserved foods, lack of understanding of the preservation systems used in traditional ethnic foods), climate changes and microbial resistance [6]. These are weak points within the food production and
Ac ce p
distribution lines where food pathogens remain only partially destroyed, re-enter and adapt to food production and processing conditions, and therefore grow and produce toxic compounds. Food safety has to be ensured and remains a responsibility of all participants involved in the production chain [7].
Bacteria, fungi and microalgae may produce toxins in foods while leaving the food appearance, odor or flavor unaltered. Intoxication occurs after ingestion of such food products [7]. Bacteria can in some instances, produce toxins in the intestine of the host, causing toxico-infections. Since toxin producing bacteria and molds are basically ubiquitous, strict preventive measures during food preparation, storage and transportation up to the final product are the best practice to follow as prevention of toxico-infections [8]. Crucial food safety parameters therefore, remain complex 5 Page 5 of 17
problems inlcuding spore forming bacteria, thermostable bacterial toxins and mycotoxins, biofilm forming microorganisms, and marine biotoxins [6, 7]. This article brings an overview of the pathogenic bacteria, eukaryotic food pathogens and their food toxins with a particular focus on
ip t
recent advances in foodomics methods for investigations of food toxins. 2. Food toxins in foodborne illnesses
cr
2.1. Pathogenic bacteria and their food toxins
us
Lipopolysaccharide bacterial endotoxins are structural components of outer bacterial membrane, moderately toxic, heat stable, and fatal to animals only in large doses. In vivo, endotoxins are rarely
an
released during bacterial growth but always after autolysis or external lysis and phagocytic digestion of bacterial cells. Every bacterial endotoxin structure consists of three regions: highly conserved Lipid
M
A, R polysaccharide (core polysaccharide common to all members of a bacterial genus) and O polysaccharide (attached to the core polysaccharide repeating oligosaccharide subunits). Lipid A is
d
toxic while polysaccharide side chains are nontoxic even though immunogenic. Most of the work on
te
the chemical structure of endotoxin has been done with species of Salmonella and E. coli [9].
Ac ce p
On the other side Gram-positive and Gram-negative bacteria secrete the most powerful known human poisons as exoproteins (enterotoxins, neurotoxins, leukocidins and hemolysins) that act at the site of bacterial growth or spread through host’s organism towards target organs or cells [8]. A less common, but equally important from a food safety point of view, is food poisoning as a result of ingestion of pre-formed bacterial toxins or bacteria producing toxins that continue to grow in the host gut, i.e. food-borne outbreaks caused by Bacillus cereus, Clostridium botulinum, Clostridium perfringens and Staphylococcus aureus exotoxins (figure 1.) [10]. Heat stable exoproteins cannot be inactivated by thermal processing of food while heat labile botulinum toxin can be easily denatured by heat and acid or proteolyzed by proteolytic enzymes. Bacterial toxins may have a range of complex activities in the human body, which might be further
6 Page 6 of 17
complicated by individual susceptibility to toxins where the amounts of ingested toxin influence the onset and severity of the symptoms [9, 11]. Furthermore, almost nothing is known about effects resulting from ingestion of low or moderate levels of exotoxins [8].
ip t
Staphylococcal enterotoxins (SE) are a group of potent gastrointestinal protease-tolerant single chain exoproteins, that are pyrogenic toxins resistant to heat [12, 13]. As shown recently, S. aureus can
cr
produce 23 diverse enterotoxins but most frequently staphylococcal illnesses are generated by the
us
best characterized enterotoxins SEA (80%) and SEB (10%), and their most common way for food contamination is due to poor hygiene during the production process [13]. Enterotoxigenic strains of
an
E. coli produce both heat-labile and/or heat-stable enterotoxins, the most common cause of acute watery diarrhea in developing countries where an increasing degree of complexity and new virulence
M
factors was assessed in recent years [14]. B. cereus, another food pathogen, produces heat-labile diarrheagenic nonhemolytic enterotoxin Nhe [15] and/or hemolytic enterotoxin HBL [16] in the small
d
intestine of the host causing food poisoning symptoms [17]. The same species of B. cereus can
te
produce emetic heat-stable exotoxin cereulide during growth on the food [18]. Preformed cereulide is resistant to high and low pH, proteolysis, and high temperatures, generally it survives cooking and
Ac ce p
causes outbreaks of food intoxication with similar symptoms to food poisoning with staphylococcal and C. perfringens enterotoxins [19]. In food-borne botulism, the botulinum heat-labile pro-exotoxin is produced by proteolytic and non-proteolytic C. botulinum, C. baratii and C. butyricum. Extracellular proteases or proteolytic enzymes in the hosts gut cleave the single pro-exotoxin chain and convert it to a more toxic di-chain form [10]. This form of exotoxin is the well-known deadly botulinum toxin. It is extremely potent neurotoxin that is stable for days [20]. 2.2. Eukaryotic food pathogens and their food toxins The mycotoxin producing pathogenic fungi and toxin producing microalgae are eukaryotic food pathogens with the highest toxigenic potential. Large part of secondary metabolites produced by pathogenic funghi Aspergillus, Fusarium, Penicilium genera under favorable environmental 7 Page 7 of 17
conditions include mycotoxins: aflatoxins, fumonisins, trichothecenes, ochratoxins, patulin and zearalenone. Until now, it has been proved that mycotoxins incite autoimmune illnesses, allergic reactions, gastrointestinal and kidney disorders, some of them bear teratogenic, carcinogenic and
ip t
mutagenic properties. Frequently, co-occurrence of several mycotoxins in cereal has been reported with potential synergistic toxic effects. These substances enter the food chain since majority of
cr
mycotoxins are relatively heat-stable. Even though 400 of these substances and their metabolites have been identified so far, only few of them are studied in details. At last, marine toxins as
us
structurally diverse group of food toxins include eukaryotic algae toxins known to accumulate in fish
an
and shellfish [7, 21]. 3. Identification of food toxins
M
In the last decade rapid and sensitive screening methods for monitoring of food toxins were introduced: bioassays, molecular biology and/or immunological techniques, gel-clot technique,
d
turbidimetric technique, biosensors and chromatographic as well as mass spectrometry-based
te
methods [7, 22-24]. Bioassays are used when fast analysis of potentially contaminated food is required [25] and/or for assessment of cytotoxic action of food toxin in cell cultures. These methods
Ac ce p
provide a fast toxicity assessment however they are not suitable for characterization of toxins because of their relatively low specificity. More specific methods are required for determination of exact toxin origin and identification, i.e. highly sensitive and high-throughput molecular methods such as real time polymerase chain reaction (RT-PCR) and RT quantitative PCR (RT-qPCR). These are golden standards for detection of exotoxin coding genes in strains isolated from contaminated foods, i.e. staphylococcal enterotoxin coding se genes or cereulide synthetase coding structural genes cesA and cesB where exact quantification of relative transcript levels are possible [26]. Limitations of these methods include the need for previous isolation of food-poisoning strains from food where only presence or absence of toxin encoding genes are possible and no information on these genes’ expression or their respective levels in food are available. Further on, immunological testing by use of
8 Page 8 of 17
commercially developed kits is a method of choice only for detection of bacterial toxin effects (antibody production) in the host organism. For example, ELISA double antibody sandwich and automated enzyme-linked fluorescent immunoassay (ELFA) are used for detection and semi-
ip t
quantification of Staphylococus, Clostridium exotoxins and cyanotoxin microcystin [24, 27]. Although, immunological assays are officially approved they lack specificity and sensitivity and produce false
cr
positives and false negatives signal due to cross-reactions. Again, the method is reliable for assessment of toxin presence while almost nothing can be deduced about its active/inactive form.
us
The same problem may appear when polyacrylamide gel electrophoresis (PAGE) methods combined with Western blotting detection of certain toxins are used [28]. However, 2D-electrophoretic
an
fractionation, proved to be highly useful in secretomics studies of food pathogenic fungi, i.e.
M
extracellular proteome analysis of Asspergillus terreus [29] and Fusarium graminearum [30]. 3.1. Foodomics methods
d
Food safety nowadays, remains focused on prevention and monitoring of potentially harmful effects
te
growingly correlated with a number of different microbial subtypes and sub strains that may produce toxic products not easily detectable by previously described, standard methods [31]. [DR2]Therefore,
Ac ce p
development of specific, rapid and sensitive methods for analytical determination of food pathogens and identification and quantification of preformed toxins in food has attracted major interest of scientists, in particular those developing high-throughput foodomics methodologies (genomics, proteomics and metabolomics).
For example, next generation sequencing and whole-genome sequencing (WGS) technologies, capillary gel electrophoresis (CGE) and combination of molecular techniques with other analytical approaches (i.e. multiplex PCR-based procedure followed by capillary gel electrophoresis with laserinduced fluorescence detection, multiplex-PCR-CGE-LIF) have been used for detection of new foodborne pathogens with the power to differentiate closely related serotypes [32-37]. [DR3]Furthermore,
highly sensitive and accurate mass spectrometry soft ionization methods, namely 9 Page 9 of 17
matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and electrospray ionization mass spectrometry (ESI-MS) are also growingly used in the field of food safety assessment [38]. These methods are frequently combined with chromatography, in order to improve
ip t
their sensitivity [39]. MALDI TOF MS is already accepted as a promising analytical method for bacterial food pathogen identification and characterization. MALDI spectra, known as MALDI TOF MS
cr
fingerprints, provide characteristic intact or trypsin digested ribosomal or intracellular protein and peptide profiles of whole bacterial cells, whole cell suspensions or cell extracts that might be easily
us
correlated with databases [40, 41]. Furthermore, interpretation of MALDI TOF MS fingerprints is used for differentiation of bacterial strains without the necessity for proteins identification: representative
an
peaks patterns suffice for conclusion about species, genus or even strain level [42]. Still, MALDI– based identification remains highly dependent to the number of well-characterized food pathogen
M
protein biomarker sequences available in proteome databases. So far, a number of reference databases [43-45] have been developed that paved the way for introduction of the MALDI-based MS
d
identification of microbes in food as a routine. The major bottleneck for its further implementation in
te
food monitoring remains the problem of incorrect interpretation due to presence or absence of
Ac ce p
unique peak patterns [46]. The only possible solution to this problem is further development of MALDI TOF fingerprint reference databases that will host a growing number of correct MALDI TOF fingerprint profiles (for example reference library SpectraBank) coupled to bioinformatics pipelines and user friendly software [47]. This might provide an accurate tool for fast and precise identification since no statistical analysis is needed for data interpretation, and the costs of materials and staff in the investigation of food poisoning outbreaks without a known causative agent may be substantially lower in a long-term. Another interesting approach for fast discrimination of bacteria, without need for sample preparation, is paper spray mass spectrometry ambient ionization combined with principal component statistical analysis of obtained phospholipid spectra [48]. Both MALDI and ESI-MS are appropriate for the study of intact proteins and non-covalent protein complexes such as complexes of botulinum neurotoxin and neurotoxin associated proteins, which 10 Page 10 of 17
contain hemoglutinins and non-toxin non-hemogglutinins [49]. Recently, a new ultraviolet photodissociation (UVPD) online liquid chromatographic tandem mass spectrometry (LC MS/MS) strategy was implemented into analysis of structural variants of E. coli lipid A on the Orbitrap mass
ip t
spectrometer [50]. Since food is a complex matrix, sample preparation protocols have been developed to facilitate
cr
isolation and identification of food toxins: immunocapture, precipitation, filtration, solid phase
us
extraction (SPE), solid-liquid extraction (SLE), SDS-PAGE and different chromatographic methods. All these separation steps remain a challenge, i.e. cereulide extraction from foods with high lipid
an
concentration. This is why multidimensional liquid chromatography tandem mass spectrometry (MDLC MS/MS) has been developed as a faster alternative to laborious sample preparation of highly
M
complex samples [32]. In recent years, much effort was put on optimization of extraction techniques QuEChERS or dispersive liquid-liquid microextraction (DLLME) for multi-mycotoxin separation and
d
isolation although SPE, SLE [38]. QuEChERS combined with multi-mycotoxin liquid chromatography
te
tandem mass spectrometry (LC-MS/MS) screening method enabled simultaneous identification of 36 mycotoxins in wines [51] and up to 56 mycotoxins in animal feed [52]. Graphitised carbon solid phase
Ac ce p
desalting clean-up step has been used more recently for sample preparation prior to hydrophilic interaction liquid chromatography (HILIC) UPLC–MS/MS detection of paralytic shellfish toxins. This approach represents a major technical breakthrough and holds potential as new routine monitoring method for hydrophilic marine toxins [53]. Moreover, nanoscale LC-MS/MS may also provide identification of proteolytically digested exoproteins isolated from low-protein samples [54]. Recently, combined protocols for in vivo protein targeting, including stable isotope in culture (SILAC) with extraction and proteolytical strategies and mass spectrometry were introduced [55]. Such an approach was used for example, during MALDI-TOF MS or LC-MS/MS multiple reaction monitoring (MRM) for identification and quantification of cerulide toxin with a synthetic cereulide employed as internal standards. Such approach is superior to previousl methods that relayed on surrogate standards, i.e. antibiotic valinomycin [56]. At last, isobaric tandem mass tags coupled to ion-mobility 11 Page 11 of 17
spectrometry (IMS) protocols are emerging alternatives to label-free approaches and mass difference tags for quantitative proteomics and separation of isobaric exopeptides [57]. Another versatile, robust and cost-effective foodomic method is capillary electromigration (CE),
ip t
sometimes combined to mass spectrometry (CE-MS). It provides fast, efficient and automated separation with small sample volumes and low consumption of solvents and reagents [58]. Micellar
cr
electrokinetic chromatography (MEKC) is a modified CE, where an analyte is separated by differential
us
partitioning between micellar pseudo-stationary phase and mobile aqueous phase, providing separation of neutral analytes [59]. MEKC proved to be more precise technique compared to HPLC in
an
identification and quantification of patulin mycotoxin [60] and detection of emetic toxin cereulide from contaminated rice samples [61].
M
In conclusion, foodomics technologies have matured in recent years and may robustly provide precise identification of food pathogens and their toxins even on the strain/subtype level from highly
d
complex food samples. In the near future further improvements in the methodologies and protocols
te
should be mainly directed to generation of new databases, better throughput and standardization while increased food monitoring within the food production and distribution lines will be growingly
Ac ce p
demanded.
4. References
(*)[DR5][1] Cifuentes A: Food analysis and foodomics, J. Chromatogr. A 2009, 1216, 7109. [2] Simó C, Domínguez-Vega E, Marina ML, García MC, Dinelli G, Cifuentes A: CE-TOF MS analysis of complex protein hydrolyzates from genetically modified soybeans. A tool for foodomics. Electrophoresis 2010, 31, 1175-1183. [3] Ibáñez C, Valdés A, García-Cañas V, Simó C, Celebier M, Rocamora L, Gómez A, Herrero M, Castro M, Segura-Carretero A et all.: Global foodomics strategy to investigate the health benefits of dietary constituents. J. Chromatogr A 2012, 1248, 139-153.[DR6] [4] Akhtar S., Sarker MR, Hossain A: Microbiological food safety: a dilemma of developing societies. Crit Rev Microbiol 2012, 40, 348-359. [5]WHO, 2006: http://www.who.int/foodsafety/foodborne_disease/ferg/en/ 12 Page 12 of 17
[6]Havelaar AH, Brul S, Jong A, Jonge R, Zwietering MH, Kuile BH: Future challenges to microbial food safety. Int J Food Microbiol 2010, 139, 79-94. [7] Giacometti J., Buretić Tomljanović A, Josic Dj: Application of proteomics and metabolomics for investigation of food toxins. Food Res Int 2013, 54, 1042–1051.
ip t
(*)[DR7][8] Rajković A: Microbial toxins and low level of foodborne exposure. Trends Food Sci Tech 2014, 38, 149-157. [9] Henkel JS, Baldwin MR, Barbieri JT: Toxins from bacteria. EXS. 2010, 100, 1–29.
cr
[10] EFSA: The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks in 2013. EFSA Journal 2015, 13, 1-162.
us
[11] Peck MW, Stringer SC, Carter AT: Clostridium botulinum in the post-genomic era. Food Microbiol 2011, 28, 183-191.
an
[12] Grumanna D, Nübelb U, Bröker BM: Staphylococcus aureus toxins – their functions and genetics. Infection, Genetics and Evolution 2014, 21, 583–592.
M
[13] Gustafson JE, Muthaiyan A, Dupre JM, Ricke SC: Staphylococcus aureus and understanding the factors that impact enterotoxin production in foods: A review. Food Control 2014, DOI: 10.1016/j.foodcont.2014.10.016.
d
[14] Fleckenstein JM, Hardwidge PR, Munson GP, Rasko DA, Sommerfelt H, Steinsland H: Molecular mechanisms of enterotoxigenic Escherichia coli infection. Microbes Infec 2010, 12, 89-98.
te
[15] Ceuppens S, Rajkovic A, Hamelink S, van De Wiele T, Boon N, Uyttendaele M: Enterotoxin production by Bacillus cereus under gastrointestinal conditions and their immunological detection by commercially available kits. Foodborne Pathog 2012, 9, 1130–1136.
Ac ce p
[16] Senesi S, Ghelardi E: Production, secretion and biological activity of Bacillus cereus enterotoxins. Toxins 2010, 2, 1690–1703. [17] Ceuppens S, Uyttendaele M, Drieskens K, Rajkovic A, Boon N, van DeWiele T: Survival of Bacillus cereus vegetative cells and spores during in vitro simulation of gastric passage. J Food Prot 2012, 75, 690–694. [18] Marxen S, Stark TD, Frenzel E, Rütschle A, Lücking G, Pürstinger G, Pohl EE, Scherer S, EhlingSchulz M, Hofmann T: Chemodiversity of cereulide, the emetic toxin of Bacillus cereus. Anal Bioanal Chem 2015, 407, 2439-2453. [19] Stenfors Arnesen LP, Fagerlund A, Granum PE: From soil to gut: Bacillus cereus and its food poisoning toxins. FEMS Microbiol Rev 2008, 32, 579-606. [20] Hou HJM: Analysis of botulinum neurotoxin detection by mass spectrometry in forensic samples. J Forensic Res 2013, 4, 2-6. (**)[DR8][21] Giacometti J, Josic Dj: Foodomics in microbial safety. Trend Anal Chem 2013, 52, 16-22. [22] Adley, CC: Past, present and future of sensors in food production. Foods 2014, 3, 491-510. 13 Page 13 of 17
(*)[DR9][23] Yeni F, Acar S, Polat ÖG, Soyer Y, Alpas H: Rapid and standardized methods for detection of foodborne pathogens and mycotoxins on fresh produce. Food Control 2014, 40, 359-367. [24] Zhang C, Zhang J: Current techniques for detecting and monitoring algal toxins and causative harmful algal blooms. J Environ Anal Chem 2015, DOI: 10.4172/JREAC.1000123.
ip t
[25] Hu DL, Omoe K, Shimoda Y, Nakane A, Shinagawa K: Induction of emetic response to staphylococcal enterotoxins in the House Musk Shrew (Suncus murinus). Infect Immun 2003, 71, 567–570.
cr
[26] Ikeda T, Tamate N, Yamaguchi K, Makino S: Mass outbreak of food poisoning disease caused by small amounts of staphylococcal enterotoxins A and H. Appl Environ Microbiol 2005, 71, 2793–2795.
us
(*)[DR10][27] Zhao X, Lin CW, Wang J, Oh DH: Advances in rapid detection methods for foodborne pathogens. J Microbiol Biotechnol 2014, 24, 297–312.
an
[28] Hennekinne JA, Ostyn A, Guillier F, Herbin S, Prufer AL, Dragacci S: How should staphylococcal food poisoning outbreaks be characterized? Toxins 2010, 2, 2106–2116. [29] Kim Y, Nanadakumar MP, Marten MR: The state of proteome profiling in the fungal genus Aspergillus. Brief Funct Genomic Proteomic 2008, 7, 87-94.
d
M
[30] Yang FEN, Jensen JD, Svensson B, Jørgensen HJL, Collinge DB, Finnie C: Secretomics identifies Fusarium graminearum proteins involved in the interaction with barley and wheat. Molecular Plant Pathology 2012, 13, 445-453.
te
[31] García-Cañas V, Simó C, Herrero M, Ibáñez E, Cifuentes A: Present and future challenges in food analysis. Foodomics. Anal. Chem. 2012, 84, 10150-10159.[DR11][32] Cunsolo V, Muccilli V, Saletti R, Foti S: Mass spectrometry in food proteomics: a tutorial. J Mass Spectrom 2014, 49, 768-784.
Ac ce p
(*)[DR12][33] Bakker HC, Allard MW, Bopp D, Brown EW, Fontana J, Iqbal Z, Kinney A, Limberger R, Musser KA, Shudt M, et al: Rapid whole-genome sequencing for surveillance of Salmonella enterica serovar enteritidis. Emerg Infect Dis 2014, 20, 1306–1314. [34] Bergholz TM, Moreno Switt AI, Wiedmann M: Omics approaches in food safety: fulfiling the promise? Trends Microbiol 2014, 22, 275-281. [35] García-Cañas V, González R, Cifuentes A: Combined use of molecular techniques and capillary electrophoresis in food analysis. TRAC-Trend Anal.Chem. 2004, 23, 637-643. [36] Alarcón B, García-Cañas V, Cifuentes A, González R, Aznar R: Simultaneous and sensitive detection of three food-borne pathogens by multiplex PCR, capillary gel electrophoresis and laser induced fluorescence. J. Agric. Food Chem. 2004, 52, 7180-7186. [37] García-Cañas V, Cifuentes A: Detection of microbial food contaminants and their products by capillary electromigration techniques. Electrophoresis 2007, 28, 4013-4030.[DR13] (**)[DR14][38] Berthiller F, Brera C, Crews C, Iha MH, Krska R, Lattanzio VMT, MacDonald S, Malone RJ, Maragos C, Solfrizzo M, Stroka J, Whitaker TB: Developments in mycotoxin analysis: an update for 2013-2014. World Mycotoxin J 2015, 8, 5-36. 14 Page 14 of 17
(**)[DR15][39] Hird SJ, Lau BPY, Schuhmacher R, Krska R: Liquid chromatography-mass spectrometry for the determination of chemical contaminants in food. Trends Anal Chem 2014, 59, 59–72. [40] Sauer S, Kliem M: Mass spectrometry tools for the classification and identification of bacteria. Nat Rev Microbiol 2010, 8, 74–82.
ip t
(*)[DR16][41] Schumann P, Maier T: MALDI-TOF mass spectrometry applied to classification and identification of bacteria. Method Microbiol 2014, 41, 275–306.
cr
(**)[DR17][42] Novais A, Sousa C, de Dios Caballero J, Fernandez-Olmos A, Lopes J, Ramos H, Coque TM, Cantón R, Peixe L: MALDI-TOF mass spectrometry as a tool for the discrimination of high-risk Escherichia coli clones from phylogenetic groups B2 (ST131) and D (ST69, ST405, ST393). Eur J Clin Microbol 2014, 33, 1391-1399.
an
us
[43] Böhme K, Fernández-No IC, Barros-Velázquez J, Gallardo JM, Canas B, Calo-Mata P: Comparative analysis of protein extraction methods for the identification of seafood-borne pathogenic and spoilage bacteria by MALDI-TOF mass spectrometry. Anal Methods 2010, 2, 1941– 1947.
M
[44] Böhme K, Fernández-No IC, Barros-Velázquez J, Gallardo JM, Calo-Mata P, Canas B: Species differentiation of seafood spoilage and pathogenic gram-negative bacteria by MALDI-TOF mass fingerprinting. J Proteome Res 2010, 9, 3169–3183.
d
[45] AlMasoud N, Nicolaou N, Goodacre R: Optimization of matrix assisted desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) for the characterization of Bacillus and Brevibacillus species. Anal Chimica Acta 2014, 840, 49–57.
Ac ce p
te
[46] Fernandez-No IC, Bohme K, Gallardo JM, Barros-Velazquez J, Canas B, Calo-Mata P: Differential characterization of biogenic amine-producing bacteria involved in food poisoning using MALDI-TOF mass fingerprinting. Electrophoresis 2010, 31, 1116-1127. [47] Bohme K, Fernandez-No IC, Barros-Velazquez J, Gallardo JM, Canas B, Calo-Mata P: SpectraBank: an open access tool for rapid microbial identification by MALDI-TOF MS fingerptinting. Electrophoresis 2012, 33, 2138-2142. [48] Hamid AM, Jarmusch AK, Pirro V, Pincus DH, Clay BG, Gervasi G, Cooks RG: Rapid discrimination of bacteria by paper spray mass spectrometry. Anal Chem 2014, 86, 7500–7507. [49] McFarland MA, Andrzejewski D, Musser SM, Callahan JH: Platform for identification of Salmonella serovar differentiating bacterial proteins by top-down mass spectrometry: S. Typhimurium vs S. Heidelberg. Anal Chem 2014, 86, 6879–6886. [50] O’Brien JP, Needham BD, Henderson JC, Nowicki EM, Trent MS, Brodbelt, JS: 193 nm ultraviolet photodissociation mass spectrometry for the structural elucidation of lipid A compounds in complex mixtures. Anal Chem 2014, 86, 2138–2145. (*)[51] Pizzutti IR, De Kok A, Scholten J, Righi LW, Cardoso CD, NecchiRohers G, Da Silva RC: Development, optimization and validation of a multimethod for the determination of 36
15 Page 15 of 17
mycotoxins in wines by liquid chromatography-tandem mass spectrometry. Talanta 2014, 129, 352363. [52] Dzuman Z, Zachariasova M, Lacina O, Veprikova Z, Slavikova P, Hajslova J: A rugged highthroughput analytical approach for the determination and quantification of multiple mycotoxins in complex feed matrices. Talanta 2014, 121, 263-272.
cr
ip t
(**)[DR18][53] Boundy MJ, Selwood AI, Harwood DT, McNabb PS, Turner AD: Development of a sensitive and selective liquid chromatography–mass spectrometry method for high throughput analysis of paralytic shellfish toxins using graphitised carbon solidphase extraction. Journal of Chromatogr A 2015, 1387, 1–12.
us
[54] Janga KS, Sweredoskib MJ, Grahamb RLJ, Hessb S, Clemons Jr WM: Comprehensive proteomic profiling of outer membrane vesicles from Campylobacter jejuni. J Proteomics 2014, 98, 90–98.
an
[55] Bauer T, Stark T, Hofmann T, Ehling-Schulz M: Development of a stable isotope dilution analysis for the quantification of the Bacillus cereus toxin cereulide in foods. J Agric Food Chem 2010, 58, 1420–1428.
M
(**)[56] Muratovic AZ, Tröger R, Granelli K, Hellenäs KE: Quantitative analysis of cereulide toxin from Bacillus cereus in rice and pasta using synthetic cereulide standard and 13C6-cereulide standard—A short validation study. Toxins 2014, 6, 3326-3335.
d
[57] Lanucara F, Holman SW, Gray CJ, Eyers CE: The power of ion mobility-mass spectrometry for structural characterization and the study of conformational dynamics. Nature Chemistry 2014, 6, 281–294.
Ac ce p
te
[58] Castro-Puyana M, García-Canas V, Simó C, Cifuentes A: Recent advances in the application of capillary electromigration methods for food analysis and foodomics. Electrophoresis 2012, 33, 147– 167. [59] Hancu G, Simon B, Rusu A, Mircia E, Gyéresi Á: Principles of micellar electrokinetic capillary chromatography applied in pharmaceutical analysis. Adv Pharm Bull 2013, 3, 1–8. [60] Murillo-Arbizu M, González-Penas E, Amézqueta, S: Patulin in apple juice for infants. Food Chem Toxicol 2010, 48, 2429–2434. [61] Oh MH, Cox JM: Development and application of a centrifugation-plating method to study the biodiversity of Bacillus species in rice products. Food Control 2010, 21, 652–665.
16 Page 16 of 17
ip t cr us an
Ac ce p
te
d
M
Figure 1. Distribution of food-borne outbreaks per causative agent in the EU during 2013 [10].
17 Page 17 of 17