Handbook of Fruits and Fruit Processing Editor Y. H. Hui Associate Editors J´ zsef Barta, M. Pilar Cano, Todd W. Gusek, o Jiwan S. Sidhu, and Nirmal K. Sinha
C 2006 Blackwell Publishing Danvers, MA 01923. For those organizations that haveAll rights reserved been granted a photocopy license by CCC, a sepa- rate system of payments has been arranged. The feeBlackwell Publishing Professional codes for users of the Transactional Reporting Service2121 State Avenue, Ames, Iowa 50014, USA are ISBN-13: 978-0-8138-1981-5; ISBN-10: 0-8138- 1981-4/2006 $.10.Orders: 1-800-862-6657Ofﬁce: 1-515-292-0140 First edition, 2006Fax: 1-515-292-3348Web site: www.blackwellprofessional.com Library of Congress Cataloging-in-Publication DataBlackwell Publishing Ltd Handbook of fruits and fruit processing / editor, Y.H.9600 Garsington Road, Oxford OX4 2DQ, UK Hui; associate editors, J´ zsef Barta . . . oTel.: +44 (0)1865 776868 [et al.].— 1st ed. p. cm.Blackwell Publishing Asia Includes index.550 Swanston Street, Carlton, Victoria 3053, Australia ISBN-13: 978-0-8138-1981-5 (alk. paper)Tel.: +61 (0)3 8359 1011 ISBN-10: 0-8138-1981-4 (alk. paper) 1. Food industry and trade. 2. Fruit—Processing.Authorization to photocopy items for internal or per- I. Hui, Y. H. (Yiu H.) II. Barta, J´ zsef. osonal use, or the internal or personal use of speciﬁcclients, is granted by Blackwell Publishing, provided TP370.H264 2006that the base fee of $.10 per copy is paid directly to 664 .8—dc22the Copyright Clearance Center, 222 Rosewood Drive, 2005013055 The last digit is the print number: 9 8 7 6 5 4 3 2 1
Contents Contributors, vii Preface, xiPart I Processing Technology 1. Fruit Microbiology, 3 A. Kalia and R. P. Gupta 2. Nutritional Values of Fruits, 29 C. S´ nchez-Moreno, S. De Pascual-Teresa, B. De Ancos, and M. P. Cano a 3. Fruit Processing: Principles of Heat Treatment, 45 I. K¨ rmendy o 4. Fruit Freezing Principles, 59 B. De Ancos, C. S´ nchez-Moreno, S. De Pascual-Teresa, and M. P. Cano a 5. Fruit Drying Principles, 81 J. Barta 6. Non-Thermal Pasteurization of Fruit Juice Using High Voltage Pulsed Electric Fields, 95 Zs. Cserhalmi 7. Minimally Processed Fruits and Fruit Products and Their Microbiological Safety, 115 Cs. Balla and J. Farkas 8. Fresh-Cut Fruits, 129 O. Mart´n-Belloso, R. Soliva-Fortuny, and G. Oms-Oliu ı 9. Food Additives in Fruit Processing, 145 P. S. Raju and A. S. Bawa10. Fruit Processing Waste Management, 171 J. Monspart-S´ nyi ePart II Products Manufacturing11. Manufacturing Jams and Jellies, 189 H. S. Vibhakara and A. S. Bawa12. Manufacturing Fruit Beverages, 205 E. Horv´ th-Kerkai a13. Fruit as an Ingredient in a Fruit Product, 217 Gy. P´ tkai a14. Fruit Processing Plant, 231 J. Barta15. Fruits: Sanitation and Safety, 245 S. Al-Zenki and H. Al-Omariah v
vi ContentsPart III Commodity Processing16. Apples, 265 N. K. Sinha17. Apricots, 279 M. Siddiq18. Horticultural and Quality Aspects of Citrus Fruits, 293 M. J. Rodrigo and L. Zacar´as ı19. Oranges and Citrus Juices, 309 K. S. Sandhu and K. S. Minhas20. Sweet Cherries, 359 J. Alonso and R. Alique21. Cranberry, Blueberry, Currant, and Gooseberry, 369 K. K. Girard and N. K. Sinha22. Date Fruits Production and Processing, 391 J. S. Sidhu23. Grape Juice, 421 O. Mart´n-Belloso and A. R. Marsell´ s-Fontanet ı e24. Grapes and Raisins, 439 N. R. Bhat, B. B. Desai, and M. K. Suleiman25. Grape and Wine Biotechnology: Setting New Goals for the Design of Improved Grapevines, Wine Yeast, and Malolactic Bacteria, 453 I. S. Pretorius26. Olive Processing, 491 B. Gandul-Rojas and M. I. M´nguez-Mosquera ı27. Peach and Nectarine, 519 M. Siddiq28. Pear Drying, 533 R. de Pinho Ferreira Guin´ e29. Plums and Prunes, 553 M. Siddiq30. Processing of Red Pepper Fruits (Capsicum annuum L.) for Production of Paprika and Paprika Oleoresin, 565 A. P´ rez-G´ lvez, M. Jar´ n-Gal´ n, and M. I. M´nguez-Mosquera e a e a ı31. Strawberries and Raspberries, 581 N. K. Sinha32. Tropical Fruits: Guava, Lychee, Papaya, 597 J. S. Sidhu33. Banana, Mango, and Passion Fruit, 635 L. G. Occe˜ a-Po n34. Nutritional and Medicinal Uses of Prickly Pear Cladodes and Fruits: Processing Technology Experiences and Constraints, 651 M. Hamdi35. Speciality Fruits Unique to Hungary, 665 M. St´ ger-M´ t´ e ae Index, 679
ContributorsRafael Alique (Chapter 20) Csaba Balla (Chapter 7)Instituto del Frío (CSIC) Corvinus University of Budapest, Faculty ofC/José Antonio Novais n◦ 10 Food Science, Department of Refrigeration28040 Madrid, Spain and Livestock Products TechnologyPhone: +34915492300 Hungary 1118, Budapest, Ménesi út 45 Phone: 36-1-482-6064Jesús Alonso (Chapter 20) Fax: 36-1-482-6321Instituto del Frío (CSIC) E-mail: firstname.lastname@example.orgC/José Antonio Novais n◦ 1028040 Madrid, Spain J´ zsef Barta, Ph.D. (Chapters 5, 14) oPhone: +34915492300 Head of the DepartmentE-mail: email@example.com Corvinus University of Budapest Faculty of Food ScienceHusam Al-Omariah (Chapter 15) Department of Food PreservationBiotechnology Department Budapest, Ménesi út 45Kuwait Institute for Scientiﬁc Research Hungary 1118P.O. Box 24885, 13109-Safat, Kuwait Phone: 36-1-482-6212 Fax: 36-1-482-6327Sameer Al-Zenki (Chapter 15) E-mail: firstname.lastname@example.orgBiotechnology DepartmentKuwait Institute for Scientiﬁc Research A.S. Bawa (Chapters 9, 11)P.O. Box 24885, 13109-Safat, Kuwait Fruits and Vegetables TechnologyPhone: (965)-483-6100 Defence Food Research LaboratoryFax: (965)-483-4670 Siddarthanagar, Mysore-570 011, IndiaE-mail: email@example.com Phone: 0821-247-3783 Fax: 0821-247-3468Begoña De Ancos (Chapters 2, 4) E-mail: firstname.lastname@example.orgDepartment of Plant Foods Scienceand Technology, Instituto del Frío N. R. Bhat (Chapter 24)Consejo Superior de Investigaciones Arid Land Agriculture DepartmentCientíﬁcas (CSIC) Ciudad Universitaria Kuwait Institute for Scientiﬁc ResearchE-28040 Madrid, Spain P.O. Box 24885, 13109-Safat, KuwaitE-mail: email@example.com E-mail: firstname.lastname@example.org vii
viii ContributorsM. Pilar Cano, Ph.D. (Chapters 2, 4) Beatriz Gandul-Rojas (Chapter 26)Director Group of Chemistry and BiochemistryInstituto del Frío-CSIC of Pigments. Food Biotechnology DepartmentC/Jose Antonio Novais, 10 Instituto de la Grasa (CSIC).Ciudad Universitaria Av. Padre García Tejero 4, 4101228040-Madrid, Spain Sevilla, SpainPhone: 34-91-5492300Fax: 34-91-5493627 Kristen K. Girard (Chapter 21)E-mail: email@example.com Principal Scientist Ocean Spray Cranberries, Inc.Zsuzsanna Cserhalmi (Chapter 6) IngredientsCentral Food Research Institute 1 Ocean Spray Dr.Hungary 1022 Budapest, Hermann O. u. 15 Middleboro MA 02349, USAPhone: 36-1-214-1248 E-mail: firstname.lastname@example.orgFax: 36-1-355-8928E-mail: email@example.com Rajinder P. Gupta (Chapter 1) Department of Microbiology,Sonia De Pascual-Teresa (Chapters 2, 4) College of Basic Sciences and HumanitiesDepartment of Plant Foods Science Punjab Agricultural Universityand Technology, Instituto del Frío Ludhiana-141004, IndiaConsejo Superior de Investigaciones firstname.lastname@example.orgCientíﬁcas (CSIC) Ciudad UniversitariaE-28040 Madrid, Spain Todd W. Gusek, Ph.D.E-mail: email@example.com Principal Scientist, Central Research Cargill, Inc.B. B. Desai (Chapter 24) PO Box 5699Arid Land Agriculture Department Minneapolis, MN 55440, USAKuwait Institute for Scientiﬁc Research Phone: (952)742-6523P.O. Box 24885, 13109-Safat, Kuwait Fax: (952)742-4925 E-mail: todd firstname.lastname@example.orgJ´ zsef Farkas (Chapter 7) oCorvinus University of Budapest M. Hamdi (Chapter 34)Faculty of Food Science, Department Director, Department of Biochemical and Chemicalof Refrigeration and Livestock Products Engineering Microbial and Food ProcessesTechnology and Central Food Research Institute Higher School of Food IndustriesHungary 1118, Budapest, Ménesi út 45 National Institute of Applied Sciencesand, 1022, Budapest, Hermann O. u. 15 and Technology. BP: 676. 1080 TunisiaPhone: 36-1-482-6303 Phone: 216-98-326675Fax: 36-1-482-6321 Fax: 216-71-704-329E-mail: email@example.com E-mail: firstname.lastname@example.orgRaquel de Pinho Ferreira Guin´ (Chapter 28) e Emoke Horváth-Kerkai (Chapter 12)Associate Professor Corvinus University of Budapest, FacultyDepartment of Food Engineering of Food Science, Department ofESAV, Polytechnic Institute of Viseu Food Preservation Hungary 1118Campus Politécnico, Repeses Budapest, Ménesi út 45.3504-510 Viseu, Portugal Phone: 36-1-482-6035E-mail: email@example.com Fax: 36-1-482-6327 E-mail: firstname.lastname@example.org
Contributors ixY. H. Hui, Ph.D. Av. Padre García Tejero 4, 41012Senior Scientist Sevilla, SpainScience Technology System Phone: +34954691054P.O. Box 1374 Fax: +34954691262West Sacramento, CA 95691, USA E-mail: email@example.com.Phone: 916-372-2655Fax: 916-372-2690 Kuldip Singh Minhas (Chapter 19)E-mail: firstname.lastname@example.org Professor Food Science and TechnologyManuel Jarén-Galán (Chapter 30) Punjab Agricultural UniversityGroup of Chemistry and Biochemistry Ludhiana, Punjab, Indiaof Pigments. Food Biotechnology Department Phone: 0161-2401960 Extn. 305Instituto de la Grasa (CSIC)Av. Padre García Tejero 4, 41012 Judit Monspart-Sényi (Chapter 10)Sevilla, Spain Corvinus University of Budapest, Faculty of Food Science, Department of Food PreservationAnu Kalia (Chapter 1) Hungary 1118, Budapest, Ménesi út 45Department of Microbiology, Phone: 36-1-482-6037College of Basic Sciences and Humanities Fax: 36-1-482-6327Punjab Agricultural University E-mail: email@example.comLudhiana-141004, Indiakaliaanu@rediffmail.com Lillian G. Occeña-Po (Chapter 33) Department of Food Science and Human NutritionImre Körmendy (Chapter 3) Michigan State UniversityCorvinus University of Budapest, East Lansing, MI 48824, USAFaculty of Food Science, Department Phone: 517-432-7022of Food Preservation Hungary 1118 Fax: 517-353-8963Budapest, Ménesi út 45 E-mail: firstname.lastname@example.orgPhone: 36-1-482-6212Fax: 36-1-482-6327 Gemma Oms-Oliu (Chapter 8)E-mail: email@example.com Department of Food Technology, University of Lleida Av. Alcalde Rovira Roure, 191. 25198Olga Martín-Belloso (Chapters 8, 23) Lleida, SpainDepartment of Food Technology, University Phone: +34-973-702-593of Lleida Av. Alcalde Rovira Roure, 191. 25198 Fax: +34-973-702-596Lleida, Spain E-mail: firstname.lastname@example.orgPhone: +34-973-702-593Fax: +34-973-702-596 Györgyi Pátkai (Chapter 13)E-mail: email@example.com Corvinus University of Budapest, Faculty of Food Science, Department of Food PreservationAngel Robert Marsellés-Fontanet (Chapter 23) Hungary 1118, Budapest, Ménesi út 45Department of Food Technology, University Phone: 36-1-482-6212of Lleida Av. Alcalde Rovira Roure, 191. 25198 Fax: 36-1-482-6327Lleida, Spain E-mail: firstname.lastname@example.orgPhone: +34 973 702 593Fax: +34 973 702 596 Antonio Pérez-Gálvez (Chapter 30)E-mail: email@example.com Group of Chemistry and Biochemistry of Pigments, Food Biotechnology DepartmentM. Isabel Mínguez-Mosquera (Chapters 26, 30) Instituto de la Grasa (CSIC).Group of Chemistry and Biochemistry Av. Padre García Tejero 4, 41012,of Pigments. Food Biotechnology Department Sevilla, SpainInstituto de la Grasa (CSIC)
x ContributorsIsak S. Pretorius (Chapter 25) East Lansing, MI 48824, USAThe Australian Wine Research Institute Phone: 517-355-8474PO Box 197, Glen Osmond Fax: 517-353-8963Adelaide, SA 5064 E-mail: firstname.lastname@example.orgAustraliaPhone: +61-8-83036835 Nirmal K. Sinha, Ph.D. (Chapters 16, 21, 31)Fax: +61-8-83036601 VP, Research and DevelopmentE-mail: Sakkie.Pretorius@awri.com.au Graceland Fruit, Inc. 1123 Main StreetP.S. Raju (Chapter 9) Frankfort, MI 49635, USAFruits and Vegetables Technology Phone: 231-352-7181Defence Food Research Laboratory Fax: 231-352-4711Siddarthanagar, Mysore-570 011, India E-mail: email@example.comPhone: 0821-247-3783Fax: 0821-247-3468 Robert Soliva-Fortuny (Chapter 8)E-mail: firstname.lastname@example.org Department of Food Technology, University of Lleida Av. Alcalde Rovira Roure, 191. 25198María Jesús Rodrigo (Chapter 18) Lleida, SpainInstituto de Agroquímica y Tecnología Phone: +34-973-702-593de Alimentos (CSIC). Apartado Postal 73 Fax: +34-973-702-59646100 Burjasot, Valencia, Spain E-mail: email@example.comConcepción Sánchez-Moreno (Chapters 2, 4) Mónika Stéger-Máté (Chapter 35)Department of Plant Foods Science and Corvinus University of Budapest, FacultyTechnology, Instituto del Frío, Consejo Superior of Food Science, Department of Food Preservationde Investigaciones Cientíﬁcas (CSIC) Hungary 1118, Budapest, Ménesi út 45Ciudad Universitaria, E-28040 Madrid, Spain Phone: 36-1-482-6034E-mail: firstname.lastname@example.org Fax: 36-1-482-6327 E-mail: email@example.comKulwant S. Sandhu (Chapter 19)Sr. Veg. Technologist (KSS) M. K. Suleiman (Chapter 24)Department of Food Science and Technology Arid Land Agriculture DepartmentPunjab Agricultural University Kuwait Institute for Scientiﬁc ResearchLudhiana - 141 004, Punjab, India P.O. Box 24885, 13109-Safat, KuwaitPhone: 0161-2405257, 2401960 extn. 8478(KSS) H.S. Vibhakara (Chapter 11)E-mail: firstname.lastname@example.org Fruits and Vegetables Technology Defence Food Research LaboratoryJiwan S. Sidhu, Ph.D. (Chapters 22, 32) Siddarthanagar, Mysore-570 011, IndiaProfessor, Department of Family Science Phone: 0821-247-3949College for Women, Kuwait University Fax: 0821-247-3468P.O. Box 5969, Safat-13060, KuwaitPhone: (965)-254-0100 extn. 3307 Lorenzo Zacarías (Chapter 18)Fax: (965)-251-3929 Instituto de Agroquímica y TecnologíaE-mails: email@example.com; de Alimentos (CSIC). Apartado Postal firstname.lastname@example.org 46100 Burjasot, Valencia, Spain Phone: 34 963900022Muhammad Siddiq (Chapters 17, 27, 29) Fax: 34 963636301Food Processing Specialist E-mail: email@example.com orDepartment of Food Science & Human Nutrition firstname.lastname@example.orgMichigan State University
PrefaceIn the past 30 years, several professional reference Part III is from the commodity processing perspec-books on fruits and fruit processing have been pub- tive, covering important groups of fruits such as:lished. The senior editor of this volume was part of r Applesan editorial team that published a two-volume text on r Apricotsthe subject in the mid-nineties. r Citrus fruits and juices It may not be appropriate for us to state the ad- r Sweet cherriesvantages of our book over the others available in the r Cranberries, blueberries, currants, andmarket, especially in contents; however, each profes-sional reference treatise has its strengths. The deci- gooseberries r Date fruitssion is left to the readers to determine which title best r Grapes and raisins, including juices and winesuits their requirement. r Olives This book presents the processing of fruits from r Peaches and nectarinesfour perspectives: scientiﬁc basis; manufacturing and r Pearsengineering principles; production techniques; and r Plums and Prunesprocessing of individual fruits. r Red pepper fruits Part I presents up-to-date information on the funda- r Strawberries and raspberriesmental aspects and processing technology for fruits r Tropical fruits (guava, lychee, papaya, banana,and fruit products, covering: mango, and passion fruit)r Microbiologyr Nutrition Although many topical subjects are included in ourr Heat treatment text, we do not claim that the coverage is comprehen-r Freezing sive. This work is the result of the combined effortsr Drying of nearly ﬁfty professionals from industry, govern-r New technology: pulsed electric ﬁelds ment, and academia. They represent eight countriesr Minimal processing with diverse expertise and backgrounds in the disci-r Fresh-cut fruits pline of fruit science and technology. An internationalr Additives editorial team of six members from four countriesr Waste management led these experts. Each contributor or editor was re- sponsible for researching and reviewing subjects ofPart II covers the manufacturing aspects of processed immense depth, breadth, and complexity. Care andfruit products: attention were paramount to ensure technical accu-r Jams and jellies racy for each topic. In sum, this volume is unique inr Fruit beverages many respects. It is our sincere hope and belief that itr Fruit as an ingredient will serve as an essential reference on fruits and fruitr A fruit processing plant processing for professionals in government, industry,r Sanitation and safety in a fruit processing plant and academia. xi
xii Preface We wish to thank all the contributors for sharing TechBooks, Inc. for their time, effort, advice, andtheir expertise throughout our journey. We also thank expertise. You are the best judges of the quality ofthe reviewers for giving their valuable comments on this work.improving the contents of each chapter. All these pro-fessionals are the ones who made this book possible. Y. H. HuiWe trust that you will beneﬁt from the fruits of their J. Bartalabor. M. P. Cano We know ﬁrsthand the challenges in developing T. W. Guseka book of this scope. What follows are the difﬁcul- J. S. Sidhuties in producing the book. We thank the editorial N. Sinhaand production teams at Blackwell Publishing and
4 Part I: Processing Technologyunidentiﬁed etiological agents. These new outbreaks NORMAL MICROFLORA OFof fresh-produce-related food poisoning include ma- PROCESSED FRUIT PRODUCTSjor outbreaks by tiny culprits as Escherichia coli0157:H7, Salmonella, Shigella, Cyclospora, Hepati- Postharvest processing methods include diversetis A virus, Norwalk disease virus, on a variety of range of physical and chemical treatments to enhancefruits as cantaloupes, apples, raspberries, and other the shelf life of fresh produce. The minimally pro-fruits. Erickson and Kornacki (2003) have even ad- cessed fresh-cut fruits remain in a raw fresh statevocated the appearance of Bacillus anthracis as a without freezing or thermal processing, or additionpotential food contaminant. Factors include global- of preservatives or food additives, and may be eatenization of the food supply, inadvertent introduction of raw or conveniently cooked and consumed. Thesepathogens into new geographical areas (Frost et al., minimally processed fruits are washed, diced, peeled,1995; Kapperud et al., 1995), the development of trimmed, and packed, which lead to the removal ofnew virulence factors by microorganisms, decreases fruit’s natural cuticle, letting easy access by outerin immunity among certain segments of the popula- true or opportunistic normal microﬂora to the internaltion, and changes in eating habits. disrupted tissues abrassed during processing. Gorny and Kader (1996) observed that pear slices cut with a freshly sharpened knife retained visual quality longer than the fruits cut with a dull hand-slicer.NORMAL MICROFLORA Rinsing of fresh produce with contaminated wa-OF FRESH FRUITS ter or reusing processed water adds E. coli 0157:H7,Fresh fruits have an external toughness, may be water Enterobacter, Shigella, Salmonella sp., Vibrio chlo-proof, wax-coated protective covering, or skin that reae, Cryptosporidium parvum, Giardia lamblia, Cy-functions as a barrier for entry of most plant clospora caytanensis, Toxiplasma gondii, and otherpathogenic microbes. The skin, however, harbors a causative agents of foodborne illnesses in humans,variety of microbes and so the normal microﬂora of thus increasing the microbial load of the fresh pro-fruits is varied and includes both bacteria and fungi duce that undergo further processing including addi-(Hanklin and Lacy, 1992). These microbes get tion of undesirable pathogens from the crop.associated with fruits, since a variety of sources such Fruits processed as fruit concentrates, jellies, jams,as the blowing air, composted soil, insects as preserves, and syrups have reduced water activ-Drosophila melanogaster or the fruit ﬂy inoculate ity (aw ) achieved by sufﬁcient sugar addition andthe skin/outer surface with a variety of Gram- heating at 60–82◦ C, that kills most of xerotolerantnegative bacteria (predominantly Pseudomonas, fungi as well as restrains the growth of bacteria.Erwinia, Lactobacillus). Likewise, hand-picking Thus, the normal microﬂora of such diligently pro-the fresh produce inoculates the fruit surfaces cessed fruit products may include highly osmophilicwith Staphylococcus. Contact with soil, especially yeasts and certain endospore-forming Clostridium,partially processed compost or manure, adds di- Bacillus sp. that withstand canning procedures. Sim-verse human pathogenic microbes generally of the ilar ﬂora may appear for processed and pasteurizedfecal-oral type including the Enterobacter, Shigella, fruit juices and nectars that loose most vegetative bac-Salmonella, E. coli 0157:H7, Bacillus cereus, as teria, yeasts, and molds while retaining heat-resistantwell as certain viruses such as Hepatitis A Virus, ascospores or sclerotia producing Paecilomyces sp.,Rotavirus, and Norwalk disease viruses that are Aspergillus sp., and Penicillum sp. (Splittstoesser,transmitted by consumption of raw fruits. Normal 1991). Recently, Walls and Chuyate (2000) reportedfungal microﬂora of fruits includes molds such the occurrence of Alicyclobacillus acidoterrestris,as Rhizopus, Aspergillus, Penicillum, Eurotium, an endospore-forming bacteria in pasteurized orangeWallemia, while the yeasts such as Saccharomyces, and apple juices.Zygosaccharomyces, Hanseniaspora, Candida,Debaryomyces, and Pichia sp. are predominant.These microbes are restrained to remain outside on FACTORS AFFECTINGfruit surfaces as long as the skins are healthy and MICROBIAL GROWTHintact. Any cuts or bruises that appear during the Fruits are composed of polysaccharides, sugars, or-postharvest processing operations allow their entry ganic acids, vitamins, minerals which function as em-to the less protected internal soft tissue. inent food reservoirs or substrates dictating the kind
1 Fruit Microbiology 5of microorganisms that will ﬂourish and perpetuate as ATP and DNA require neutrality (Brown, 1964).in the presence of speciﬁc microﬂora and speciﬁc The pH changes also affect the morphology of someenvironmental prevailing conditions. Hence, one can microbes as Penicillum chrysogenum that show de-predict the development of microﬂora on the basis creased length of hyphae at pH above 6.0. Corlettof substrate characteristics. Fresh fruits exhibit the and Brown (1980) observed varying ability of or-presence of mixed populations, and growth rate of ganic acids as microbial growth inhibitors in relationeach microbial type depends upon an array of factors to pH changes.that govern/inﬂuence the appearance of dominatingpopulation, which include the following. Water Activity (Moisture Requirement)Intrinsic Factors Water is a universal constituent required by all the liv- ing cells, and microbes are no exceptions but the exactThese imply the parameters that are an inherent part amount of water required for growth of microorgan-of the plant tissues (Mossel and Ingram, 1955) and isms varies. Hence, several preservation methods in-thus are characteristics of the growth substrates that volve drying or desiccation of the produce (Worboinclude the following. and Padilla-Zakour, 1999). The water requirement of microbes is deﬁned as water activity (aw ) or ratio ofHydrogen Ion Concentration (pH) water vapor pressure of food substrate to that of vapor pressure of pure water at same temperatureMicrobial cells lack the ability to adjust their inter-nal pH, hence are affected by change in pH, so could p aw = ,grow best at pH values around neutral. Bacteria ex- pohibit a narrow pH range with pathogenic bacteria be- where p is the vapor pressure of the solution and poing the most fastidious; however, yeasts and molds is the vapor pressure of the solvent.are more acid-tolerant than bacteria. Fruits possess Christian (1963) related water activity to relativemore acidic pH (<4.4) favoring growth of yeasts and humidity as (Table 1.2)molds. Microbes, in general, experience increasedlag and generation times at either extremes of the RH = 100 × aw .optimum pH range, which is usually quite narrow.The small ﬂuctuations in pH have elaborate impact Thus, the relative humidity of a food correspondingon microbial growth rates, and the pH changes be- to a lower aw tends to dry the surface and vice versa.come more profound if the substrate has low buffer- In general, most fresh produce has aw value aboveing capabilities leading to rapid changes in response 0.99 which is sufﬁcient for the growth of both bacte-to metabolites produced by microorganisms during ria and molds; however, bacteria, particularly Gram-fermentation (Table 1.1). negative, are more stringent regarding aw changes, Adverse pH affects the functioning of respiring while molds could grow at aw as low as 0.80. Themicrobial enzymes and the transport of nutrients into lowest range of permeable aw values for halophilicthe cell. The intracellular pH of microbial cytoplasm bacteria, xerophilic fungi, and osmophilic yeasts isremains reasonably constant due to relative imper- 0.75–0.61. Morris (1962) elaborated the interactionmeability of cell membrane to hydrogen (H+ ) and of aw values with temperature and nutrition and ob-hydroxyl (OH− ) ions as key cellular compounds such served that at optimum temperature, range of aw val- ues remain wide, while lowering/narrowing aw values reduces growth and multiplication of microbes, andTable 1.1. Approximate pH Values of Some nutritive properties of substrate increase the range ofFresh Fruits aw over which microorganisms can survive (Fig. 1.1). Hence, each microbe has its own characteristic awFruits pH Values Fruits pH Values range and optimum for growth and multiplicationApples 2.9–3.3 Limes 1.8–2.0 which are affected by temperature, pH, oxygen avail-Bananas 4.5–4.7 Melons 6.3–6.7 ability, and nutritive properties of substrate as wellGrapefruit 3.4–4.5 Figs 4.6 as the presence of organic acids or other secondaryWatermelons 5.2–5.6 Plums 2.8–4.6 metabolites performing inhibitory action, thus nar-Oranges 3.6–4.3 rowing the aw range that culminates in decreasedSource: Adapted from Jay (1992). yield of cells and increased lag phase for growth,
6 Part I: Processing Technology Table 1.2. Lower Limit aw Values of Certain Microorganisms Bacteria Minimum aw Values Fungi Minimum aw Values Pseudomonas 0.97 Mucor 0.62 (0.94) E. coli 0.96 Rhizopus 0.62 Staphylococcus aureus 0.86 Botyritis 0.62 Bacillus subtilis 0.95 Aspergillus 0.85 Clostridium botulinum 0.93 Penicillum 0.95 Enterobacter aerogenes 0.94 Source: Adapted from Jay (1992). Pseudomonas 1 S.aureus 0.9 E.coli 0.8 B.subtilis 0.7 C.botulinum 0.6 0.5 E.aerogenes 0.4 Mucor 0.3 Rhizopus 0.2 Botyritis 0.1 Aspergillus 0 Penicillum Pseudomonas S.aureus E.coli B.subtilis Mucor C.botulinum E.aerogenes Rhizopus Botyritis Aspergillus PenicillumFigure 1.1. Graphical representation ofaW values of various microbes.and results in decreased growth rate and size of ﬁnal accumulate polyhydric alcohols (Troller, 1986). Mi-population (Wodzinsky and Frazier, 1961). Lowering crobes thus attempt to compensate for increasedof water activity builds up stress and exerts adverse stress by accumulating compatible solutes.inﬂuence on all vital metabolic activities that requireaqueous environment. Charlang and Horowitz (1974) Redox Potential/Redox Poising Capacityobserved the appearance of non-lethal alterations incell membrane permeability of Neurospora crassa The type of microbial growth depends upon oxidationcells resulting in loss of various essential molecules, and reduction power of the substrate. The oxidation–as the dynamic cell membrane should remain in ﬂuid reduction potential of a substrate may be deﬁned asstate. the ease with which the substrate loses or gains elec- The exception to normal aw requirements are ba- trons and, in turn, gets oxidized or reduced, respec-sically the halophilic bacteria that grow under low tively. Aerobic microbes require oxidized (positiveaw values by accumulating potassium ions in the Eh values) substrates for growth and it is reverse forcell (Csonka, 1989), while osmophilic yeasts concen- the anaerobes (Walden and Hentges, 1975). The fruitstrate polyols as osmoregulators and enzyme protec- contain sugars and ascorbic acid for maintaining thetors (Sperber, 1983). Brown (1976) reported proline reduced conditions, though plant foods tend to haveaccumulation in response to low aw in halotolerant positive values (300–400 mV). Hence, aerobic bac-Staphylococcus aureus strains. Xerotolerant fungi teria and molds most commonly spoil fruits and fruit
1 Fruit Microbiology 7products. The O/R potential of food can be deter- be furnished by substrate since microorganisms aremined by unable to synthesize essential vitamins. In general,r Characteristic pH of food Gram-positive bacteria are least synthetic and requirer Poising capacity supply of certain vitamins before growth, whiler Oxygen tension of the atmosphere Gram-negative bacteria and molds are relatively in-r Atmospheric access of food dependent and could synthesize most of the vitamins. Thus, these two groups of microbes grow profusely Poising capacity could be deﬁned as the extent to on foods relatively low in B-complex vitamins suchwhich a food resists externally effected changes in as fruits under the inﬂuence of usual low pH and pos-pH that depend on the concentration of oxidizing or itive Eh values.reducing compounds in the substrate. The capacity Each microbe has a deﬁnite range of food require-alters the ability of the living tissues to metabolize ments, with some species having wide range and abil-oxygen at speciﬁcally low Eh values that exist in ity to grow on a variety of substrates, while othersthe vacuum-packed foods. Aerobic microbes include having narrow range and fastidious requirement al-bacilli, micrococci, pseudomonas, and actinobacters, lowing growth on limited substrates.and require positive Eh values, while anaerobes suchas clostridia and bacteriodes require negative Eh val- Antimicrobial Factorsues. However, most yeast and molds are aerobic andfew tend to be facultative anaerobes. In the presence Certain naturally occurring substances in substrateof limited amounts of oxygen, aerobic or faculta- (food) work against the microbes, thus maintain-tive microbes may produce incompletely oxidized or- ing stability of food; however, these are directed to-ganic acids. Processing procedures such as heating or ward a speciﬁc group of microorganism and havepasteurization, particularly of fruit juices, make mi- weak activity. Song et al. (1996) reported that thecrobes devoid of reducing substances, but favorable presence of aroma precursor Hexal readily gets con-for the growth of yeasts. verted to aroma volatiles in vivo by fresh-cut apple slices. Hexal acts as antibrowning agent as well as inhibits growth of molds, yeasts, mesophilic and psy-Available Nutrients chrotropic bacteria (Lanciotti et al., 1999). HexanalFruits as substrate act as a reservoir of sugars (source and (E)-Hexenal in modiﬁed atmosphere packagingof energy), water, minerals, vitamins, and other (MAP) of sliced apples reduce spoilage microbe pop-growth-promoting factors, while the protein content ulations (Corbo et al., 2000).or nitrogen source appears to be little less in fruits. Spices contain essential oils such as eugenolCarbohydrates include sugars or other carbon sources (clove), allicin (garlic), cinnamic aldehyde andthat act as sources of energy because breakage of eugenol (cinnamon), allyl isothiocynate (mustard),bonds or oxidation of these compounds helps in the eugenol and thymol (sage), thymol and isothymolformation of energy currency of cell or ATP. (oregano) that have antimicrobial activity (Shelef, Microorganisms have varied nutrient require- 1983). Buta and Molin (1998) observed reductionments, which are inﬂuenced by other conditions such in mold growth on fresh-cut peppers by exogenousas temperature, pH and Eh values. The microbes be- application of methyl jasmonate.come more demanding at decreased temperatures, The antimicrobial compounds may originally bewhile under optimum temperature conditions, nutri- present in food, added purposely or developed byents control the microbial growth only when present associated microbial growth, or by processing meth-in limiting quantities. Thus, microorganisms that ods. Certain antifungal compounds applied to fruitsgrow on a product become the best-suited by ex- include benomyl, biphenyl, and other phenylic com-ploiting the product, as pectinolytic bacteria such as pounds that exist in small quantities as by-product ofErwinia cartovora, Pseudomonas sp., or pectinolytic phenol synthesis pathways. Beuchat (1976) observedmolds grow best on fruits and vegetables. that essential oils of oregano, thyme, and sassafras Nitrogen requirement is usually fulﬁlled by pro- have bacteriocidal activity, at 100 ppm, to V. para-teolysis of protein present in substrate and the use haemolyticus in broth, while cinnamon and clove oilsof amino acids, nucleotides, certain polysaccharides, at 200–300 ppm inhibit growth and aﬂatoxin pro-and fats under usual microbe-speciﬁc conditions. duction by Aspergillus parasiticus (Bullerman et al.,The accessory food substances or vitamins are to 1977). The hydroxy-cinnamic acid derivatives as
8 Part I: Processing Technologyp-coumaric, ferulic, caffeic, and chlorogenic acids Bacillus cereus, Staphylococcus aureus, andand benzoic acid in cranberries have antibacterial Clostridium perfringens. There exists a relationand antifungal activities and are present in most plant of temperature to growth rate of microorganismsproducts including fruits. between minimum and maximum temperature range by (Ratowsky et al., 1982) √Extrinsic Factor r = b(T − T0 ),Extrinsic factors include parameters imposed from where r is the growth rate, b is the slope of regres-the external environment encountered during storage sion line, and T0 is the conceptual temperature of nothat affect food, and the microbes that tend to develop metabolic signiﬁcance.on it. These factors include the following. Relative Humidity of EnvironmentTemperature of Storage Success of a storage temperature depends on the rel-Microbes grow over a wide range of temperature, and ative humidity of the environment surrounding thechange in temperature at both extremes lengthens the food. Thus, relative humidity affects aw within a pro-generation time and lag periods. The range is quite cessed food and microbial growth at surfaces. A lowwide from −34◦ C to 90◦ C, and according to range aw food kept at high R.H. value tends to pick up mois-microbes could be grouped as follows. ture until the establishment of equilibrium, and foods with high aw lose moisture in a low-humidity envi-Psychrotrophs. These microorganisms grow well ronment. Fruits and vegetables undergo a variety ofat 7◦ C or below 7◦ C with the optima ranging surface growth by yeasts and molds as well as bac-from 20◦ C to 30◦ C. For example, Lactobacillus, teria, and thus are liable to spoilage during storageMicrococcus, Pseudomonas, Enterococcus, Psy- at low R.H. conditions. However, this practice maychrobacter, Rhodotorula, Candida and Saccha- cause certain undesirable attributes such as ﬁrmnessromyces (yeasts), Mucor, Penicillum, Rhizopus and texture loss of the climacteric (perishable) fruits(molds) and Clostridium botulinum, Listeria mono- calling for the need of altered gas compositions to re-cytogenes, Yersinia enterocolitica, Bacillus cereus tard surface spoilage without lowering R.H. values.(pathogenic psychrotrophs). The group of microbesthat grow from −10◦ C to 20◦ C with the optima of10–20◦ C are included as Psychrophiles and include Modiﬁed Atmosphere Storagecertain overlapping genera mentioned above. Altering the gaseous composition of the environ-Mesophiles. These include microbes growing best ment that retards the surface spoilage without re-between 20◦ C and 45◦ C with optimum range of ducing humidity includes the general practice of in-30–40◦ C. For example, Enterococcus faecalis, Strep- creasing CO2 (to 10%) and is referred as “controlledtococcus, Staphylococcus, and Leuconostoc. or modiﬁed atmosphere” (MA). MA retards senes- cence, lowers respiration rates, and slows the rate ofThermophiles. Microbes that grow well above tissue softening or texture loss (Rattanapanone and45◦ C with the optima ranging between 55◦ C and Watada, 2000; Wright and Kader 1997a; Qi et al.,65◦ C and with maximum of above 60–85◦ C are 1999). MA storage has been employed for fruitsknown as thermotolerant thermophiles. For exam- (apples and pears) with CO2 applied mechanicallyple, Thermus sp. (extreme thermophile), Bacillus or as dry ice, and this retards fungal rotting of fruitssternothermophilus, Bacillus coagulans, Clostrid- probably by acting as competitive inhibitor of ethy-ium thermosaccharolyticum are endospore-forming lene action (Gil et al., 1998; Wright and Kaderthermotolerants and grow between 40◦ C and 60◦ C 1997b).and create major problems in the canning industry. The inhibitory effect increases with decrease in temperature due to increase in solubility of CO2 atThermotrophs. This group includes microbes lower temperatures (Bett et al., 2001). Elevated CO2similar to mesophiles but grows at slightly higher levels are generally more microbiostatic than micro-temperature optima and includes pathogenic bac- biocidal probably due to the phenomena of catabo-teria in foods. For example, Salmonella, Shigella, lite repression. However, an alternative to CO2 ap-enterovirulent E. coli, Campylobacter, toxigenic plication includes the use of ozone gas at a few ppm
1 Fruit Microbiology 9concentration that acts as ethylene antagonist as well targeted toward inhibition of a narrow spectrumas a strong oxidizer that retards microbial growth. of microbes. Other bacteriocins produced by lac-Sarig et al. (1996) and Palou et al. (2002) reported tic acid bacteria include lactococcins, lacticins,control of postharvest decay of table grapes caused by lactacins, diplococcin, sakacins, acidophilocins, pe-Rhizopus stolonifera. A similar report on effect of diocins, and leuconosins. As an inhibitor of spore-ozone and storage temperature on postharvest dis- forming Clostridium spp., which cause cheese blow-eases of carrots was observed by Liew and Prange ing due to undesirable gas production, nisin was the(1994). In general, gaseous ozone introduction to ﬁrst bacteriocin produced by lactic acid bacteria topostharvest storage facilities or refrigerated shipping be isolated and approved for use in cheese spreads.and temporary storage containers is reported to be op- Although mostly active against Gram-positive bacte-timal at cooler temperatures and high relative humid- ria, bacteriocins can be microbiocidal under certainity (85–95%) (Graham, 1997). The most reproducible conditions, even toward Gram-negative bacteria andbeneﬁts of such storage are substantial reduction of yeasts, provided that their cell walls have been sen-spore production on the surface of infected produce sitized to their action. The antimicrobial action ofand the exclusion of secondary spread from infected nisin and of similar bacteriocins is believed to in-to adjacent produce (Kim et al., 1999; Khadre and volve cell membrane depolarization leading to leak-Yousef, 2001). age of cellular components and to loss of electrical Ozone treatment has been reported to induce pro- potential across the membrane. Propioniobacteriumduction of natural plant defense response compounds produces propionic acid that has inhibitory effectinvolved in postharvest decay resistance. Ozone de- on other bacteria. Certain microorganisms may pro-struction of ethylene in air ﬁltration systems has been duce wide spectrum antimicrobial substances or sec-linked to extended storage life of diverse ethylene- ondary metabolites capable of killing or inhibitingsensitive commodities. wide range of microbes called “antibiotics.” How- ever, growth of one kind of microbe could lead to lowering of pH of substrate, making the environ-Implicit Factors ment unsuitable for other microbes to grow, whileImplicit factors include the parameters depending organic acid production or hydrogen peroxide for-on developing microﬂora. The microorganisms while mation could also interfere with the growth of back-growing in food may produce one or more inhibitory ground microbial population (Jay, 1992).substances such as acids, alcohols, peroxides, andantibiotics that check the growth of other microor- Bioﬁlm Formationganisms. Most of the Gram-negative bacteria exhibit quorum sensing or the cell-to-cell communication phenom-General Interference ena that leads to the formation of a multicellularThis phenomena works when competition occurs be- structure in the life of a unicellular prokaryote thattween one population of microbes and another re- provides protection to bacterial species from the dele-garding the supply of the same nutrients. Normal terious environment by precipitation. Adoption ofmicroﬂora of fresh produce helps prevent the col- bioﬁlm formation involves release of autoinducers,onization of pathogens and succeeds in overcoming particularly called the N-acyl homoserine lactonesthe contaminant number by overgrowth and efﬁcient that either activate or repress the target genes in-utilization of available resources. volved in bioﬁlm formation (Surette et al., 1999). Quorum sensing has a profound role in food safety in association with behavior of bacteria in food matrixProduction of Inhibitory Substances and regulates prime events such as spore germina-Some microbes can produce inhibitory substances tion, bioﬁlm formation on surfaces (Frank, 2000b),and appear as better competitors for nutrient sup- and virulence factor production. Cells in bioﬁlm areply. The inhibitory substances may include “bac- more resistant to heat, chemicals, and sanitizers dueteriocins,” the commonest being “nisin” produced to diffusional barrier created by biomatrix as well asby certain strains of Lactobacillus lactis, which is very slow growth rates of cells in bioﬁlms (Costerton,heat stable, attached by digestive enzymes, labile 1995). Morris et al. (1997) have reported certainand non-toxic for human consumption, and is quite methods for observing microbial bioﬁlms directly
10 Part I: Processing Technologyon leaf surfaces and also to recover the constituent the causative agent to other fruits. The postharvestmicrobes for isolation of cultivable microorganisms. rots are most prevalent in fruits, particularly the dam-Thus, bioﬁlm formation has been emerging as a chal- aged or bruised ones (Sanderson and Spotts, 1995;lenge for the decontamination techniques routinely Bachmann and Earles, 2000). The processing meth-used in the food and beverage industries, and requires ods involve the use of temperature, moisture content,the advent of new revolutionary methods for decon- and ethylene control, thus include the extrinsic pa-tamination or the modiﬁcation of the older techniques rameters discussed earlier.in vision of the current scenario (Frank, 2000a). FRUIT SPOILAGEFACTORS AFFECTING The fruit spoilage is manifested as any kind of phys-MICROBIAL QUALITY ical change in color or ﬂavor/aroma of the productAND FRUIT SPOILAGE that is deteriorated by microﬂora that affects the cel-From quality standpoint, the fresh fruits and the pro- lulose or pectin content of cell walls which, in turn,cessed fruit products should possess certain charac- is the fundamental material to maintain the structuralteristics such as fresh-like appearance, taste, aroma, integrity of any horticultural product. Fresh fruitsand ﬂavor that should be preserved during stor- possess more effective defense tactics including theage. Thus, if the primary quality attributes of pro- thicker epidermal tissue and relatively higher con-duce remain unoffended, the shelf-life characteristics centration of antimicrobial organic acids. The higherlengthen. As discussed before, fruits possess normal water activity, higher sugar content, and more acidicmicroﬂora as well as the microﬂora that is added dur- pH (<4.4) of fresh fruits favor the growth of xero-ing the handling and postharvest processing of fruits, tolerant fungi or osmophilic yeasts. Lamikarna et al.though harsh treatments during processing can kill or (2000) have reported bacterial spoilage in neutral pHinhibit certain or most of the microﬂora while letting fruits.speciﬁc types to become predominant and prevail in Normal microﬂora of fruits is diverse and includesthe ﬁnished product. A variety of factors that affect bacteria such as Pseudomonas, Erwinia, Enterobac-the microbial quality of fruits include the following. ter, and Lactobacillus sp. (Pao and Petracek, 1997), and a variety of yeasts and molds. These microbes remain adhered to outer skin of fruits and come fromPreharvest Factors several sources such as air, soil, compost, and insectThese factors basically involve production practices infestation. Brackett (1987) reported inoculation ofthat have tremendous explicit effect on the micro- Rhizopus sp. spores by egg laying in ruptured epi-bial quality of fruits. Management practices can af- dermal ﬁssures of fruits by Drosophila melanogasterfect product quality since stressed produce or me- or the common fruit ﬂy. The microbial load of thechanical injuries permit microbial contamination. fresh produce could be reduced by rinsing with waterMold growth and decay on winter squash caused by (Splittstoesser, 1987). However, the source and qual-Rhizoctoina result from fruits lying on the ground. ity of water dictate the potential for human pathogenFood safety begins in ﬁeld as a number of food- contamination upon contact with the harvested pro-borne disease outbreaks have potential sources in duce.ﬁeld that contaminate the fresh produce such as Lund and Snowdon (2000) reported certain com-the use of partially treated manure, irrigation with mon molds to be involved in fruit spoilage suchlivestock-used farm pond water, or storage near as Penicillum sp., Aspergillus sp., Eurotium sp.,roosting birds (Trevor, 1997). Wallace et al. (1997) Alternaria sp., Cladosporium sp., and Botrytisreported the presence of verocytotoxin producing cinerea of fresh and dried fruits (Fig. 1.2), whileE. coli O157:H7 from wild birds. certain molds producing heat-resistant ascospores or sclerotia such as Paecilomyces fulvus, P. niveus, Aspergillus ﬁscheri, Penicillum vermiculatum, andPostharvest Handling P. dangeardii were observed to cause spoilage ofand Processing thermally processed fruits or the fruit productsImproper or harsh handling of produce causes skin exhibiting characteristic production of off-ﬂavors,breaks, bruises, or lesions leading to increased visible mold growth, starch and pectin solubilization,chances of microbial damage. Handlers picking fresh and fruit texture breakdown (Beuchat and Pitt, 1992;produce with skin lesions could potentially transfer Splittstoesser, 1991).
1 Fruit Microbiology 11 tive external protective system, thus causing active invasion and active spoilage in fruits. The degrada- tive enzyme brigade includes the following. Pectinases These enzymes depolymerize the pectin, which is a polymer of ␣-1, 4-linked d-galactopyranosyluronic acid units interspersed with 1, 2-linked rhamnopy- ranose units. On the basis of site and type of reac- tion on the pectin polymer, pectinases are of three main types, i.e., pectin methyl esterases produced by Botrytis cinerea, Monilinia fructicola, Penicillum citrinum, and Erwinia cartovora (Cheeson, 1980), polygalacturonase, and pectin lyase.Figure 1.2. Degradation of fruit texture due to growthof cellulase/pectinase-producing bacteria followed byfungal growth. Cellulases Several types of cellulase enzymes attack the na- tive cellulose and cleave the cross-linkage between ␤-d-glucose into shorter chains. Cellulases con- Fruit safety risks could be increased by certain tribute toward tissue softening and maceration as wellspoilage types that create microenvironments suit- as yield glucose, making it available to opportunisticable for the growth of human pathogens as the pri- microﬂora.mary spoilage by one group of phytopathogens pro-duces substances required for nurturing growth and Proteasesdevelopment of human pathogens. Wade and Beuchat(2003) have well documented the crucial role of pro- These enzymes degrade the protein content of freshteolytic fungi and the associated implications on the produce giving simpler units of polypeptides, i.e.,changes in pH of the pericarp of the decayed and amino acids. The action of proteases is limiting indamaged raw fruits in survival and growth of various fruit spoilage as fruits are not rich in proteins.foodborne pathogens. Botrytis or Rhizopus spoilageof fruits could help create environment for the prolif- Phosphatidaseseration of Salmonella enterica serovar typhimurium These enzymes cleave the phosphorylated com-(Wells and Butterﬁeld, 1997), while Dingman (2000) pounds present in cell cytoplasm and the energy re-observed the growth of E. coli 0157:H7 in bruised leased is utilized to cope with the increased respira-apple tissues. Similar reports of Riordan et al. (2000) tion rates.and Conway et al. (2000) depicted the impact of priormold contamination of wounded apples by Penicil-lum expansum and Glomerella cingulata on survival Dehydrogenasesof E. coli 0157:H7 and Listeria monocytogenes. These enzymes dehydrogenate the compounds, thus Technically, the fresh produce deteriorating mi- increasing the amount of reduced products that maycroﬂora is diverse and remains on surface skin of lead to increased fermentation reaction under mi-fruits, and the basis of invasion process could be of croaerobic/anaerobic conditions.two types. Opportunistic PathogensTrue Pathogens These microorganisms lack the degradative enzymeThese microbes possess ability to actively infect plant brigade and thus gain access only when the normaltissues as they produce one or several kinds of cellu- plant product defense system weakens, which is thelytic or pectinolytic and other degradative enzymes situation of mechanical injury or cuticular damageto overcome tough and impervious outer covering of caused by the insect infestation or by natural openingsfruits which acts as the ﬁrst and the foremost effec- present on the surface of the fresh produce. Thus,
12 Part I: Processing Technology MODES OF FRUIT SPOILAGE Fruit spoilage occurs as a result of relatively strong interdependent abiotic and biotic stresses posed par- ticularly during the postharvest handling of produce (Fig. 1.5). Harvested fruits continue to respire by uti- lizing the stored available sugars and adjunct organic acids culminating to signiﬁcant increase in stress- related/stress-induced carbon dioxide and ethylene production that leads to rapid senescence (Brecht, 1995). Moreover, postharvest processing that in- volves washing, rinsing, peeling, and other treat- ments result in major protective epidermal tissue damage and disruption which in turn leads to un-Figure 1.3. Growth of Aspergillus on surface of apple sheathing of the vacuole-sequestered enzymes andfruits visible due to formation of spores. related substrates and their amalgamation with the cy- toplasmic contents. Cutting/dicing increases the aw and surface area as well as stress-induced ethylenean opportunistic pathogen slips in through the dam- production which accelerates the water loss, while theage caused by biotic and abiotic stresses on the pro- sugar availability promptly invites enhanced micro-duce and generally involves movement via natural bial invasion and rapid growth (Wiley, 1994; Watadagateways as the lenticels, stomata, hydathodes, or and Qi, 1999). The physiological state of fruit alsothe other pores/lesions caused by insect infestation determines the pattern of spoilage to be followed asor invasion by true pathogens. Damage of the prod- with increase in age/maturity, the normal defenseuct during harvesting or by postharvest processing tactics of the plant produce deteriorates. Harvestedtechniques and equipments enables opportunistic mi- produce loses water by transpiration, thus gets de-croﬂora to invade the internal unarmed tissue and hydrated, followed by climacteric ripening, enzy-causes spoilage (Fig. 1.3). matic discoloration of cut surfaces to senescence, Hence, spoilage connotes any physical change in thus increasing possibilities of damage by microﬂoracolor, taste, ﬂavor, texture, or aroma caused by micro- (Fig. 1.6). Harsh handling and ill-maintained equip-bial growth in fruit/fruit product, thereby resulting in ment during processing lead to increased damage orproduct that becomes unacceptable for human con-sumption (Fig. 1.4).Figure 1.4. Fungal hyphae and spores of Aspergillus Figure 1.5. White hyphal mass of Aspergillusniger on guava fruits. fumigatus on surface of orange fruit.
1 Fruit Microbiology 13 Abiotic forces Biotic forces Damage by Damage by external Preharvest Postharvest State of sources insect damage by produce infestation microbes damage pH wind blown sand lesions invasion water activity rubbing egg-laying fermentation transpiration harvesting degradative ethylene processing enzymes production procedures damage of and outer senescence equipments layer INTERNAL TISSUE INVASION PHYSICAL CHANGES IN PRODUCE RAPID SOFTENING OF PRODUCE SHRINKAGE OF PRODUCE DECAY DECREASED SHELF LIFE OF PRODUCEFigure 1.6. Modes of fruit spoilage and factors responsible for spoilage.removal of the outer cuticle leading to tissue disrup- grow faster than the molds and this usually includestion that provokes stress-induced increased respira- the genera such as Cryptococcus, Rhodotorula, andtion and microbial decay (Gorny and Kader, 1996). Saccharomyces sp. in fresh fruits, and Zygosaccha-Spanier et al. (1998) reported the development of romyces rouxii, Hanseniaspora, Candida, Debary-off-ﬂavors in fresh-cut pineapple that appeared un- omyces, and Pichia sp. in dried fruits.damaged physically, in lower portion of container Thus, senescence and spoilage depend on prod-kept at 4◦ C for 7–10 days. Walls and Chuyate (2000) uct type, abiotic factors, and microbes involved inreported survival of acid- and heat-tolerant Alicy- deterioration process, and it is convenient to de-clobacillus acidoterrestris that produces 2-methoxy scribe spoilage on the basis of visible symptoms.phenol or guaiacol imparting phenolic off-ﬂavor in Thus, a customary approach is to name the spoilagepasteurized orange and apple juices. Jay (1992) re- type by symptomatological appearance such as softported osmophilic yeasts to be associated primarily rot or black rot. However, this deﬁnitely results inwith the spoilage of cut fruits due to their ability to discrepancy in ascertaining the causal pathogen of
14 Part I: Processing Technologyspoilage and this ambiguity could be overruled by present in a given sample. This method ushers littleclassifying on the basis of causal microbe such as value for the determination of microbiological statusRhizopus rot, Cladosporium rot, etc. of a food sample as usually total cell counts exceed 105 cfu per g or ml of the sample. New variationsMETHODS TO EVALUATE of microscopes render researchers the capability toMICROBIAL QUALITY predict the presence of pathogens on the surfaces of fruits clinging or attached to internal surfaces. Con-Food quality and safety are ensured by analysis of focal scanning laser microscopy has been reported tofood for the presence of microbes, and such mi- show the presence of E. coli 0157:H7 on surfacescrobial analyses are routinely performed as quaran- and internal structures of apple (Burnett et al.,tine/regulatory procedures. The methods employed 2000).for adjudging the quality of food include an array of Drawbacks: This technique suffers from a ma-microbiological to biochemical assays to ascertain jor drawback of not providing the types of bacteriathe acceptability or unacceptability of a food prod- present in the sample as well as it does not differenti-uct for human consumption or a processing/handling ate between the normal microﬂora and the pathogen-practice that needs to be followed. Thus, enumerating causing spoilage.the microbial load of the produce could help in de-termining the quality as well as the related safety as-pects of product and effectiveness of the processingtechnique employed to kill spoilage microbes. Aerobic Plate Counts (APC) or Total Plate Microbiological methods for pathogen identiﬁca- Counts (TPC)tion primarily involve conventional cultural tech- It is the most practical approach to determine theniques of growing microbes on culture media and ob- presence of cultivatable microbes in a sampled foodserving the ability to form viable countable colonies product having ability to spoil food. This technique,showing characteristic growth on such media as well thus, reveals the total number of microbes in a foodas the direct microscopic methods for various groups product under a particular set of incubation temper-of microbes. ature, time, or culture media and can be used to pref- Hence, microbiological criteria are speciﬁcally erentially screen out a speciﬁc group of microbes,employed to assess: thereby, helping in determining the utility of foodr Safety of food or food ingredient added for speciﬁc purpose. How-r Shelf life of perishable products ever, the APC of the refrigerated fruits/fruit productsr Suitability of food or ingredient for speciﬁc indicate utensil or equipment conditions prevailing purpose during storage and distribution of the product.r Adherence to general manufacturing practices Drawbacks: Though APC bacterial count is the most practical and easy technique, it suffers from The routine culturing techniques require longer certain inherent drawbacks as listed below:time to obtain results. To overcome this hurdle, r It provides the viable cell count that does notnowadays, use of indicator organisms that providerapid, simple, and reliable information without the reﬂect the quality of raw material used forrequirement of isolation and identiﬁcation of speciﬁc processing. r It is unable to record the extent of quality loss atpathogens is performed. However, such tests could beused as the presumptive ones with the conﬁrmation low count levels. r It provides negligible information regardingprovided by a battery of biochemical tests, and mayinclude specialized serological typing also (Swami- organoleptic quality that is degraded at lownathan and Feng, 1994). The microbiological tech- counts. r It requires scrupulous researcher to interpret APCniques could be summarized as follows. results.Conventional Techniques Certain variations to APC method are now available to classify according to the types of microbes asDirect Microscopic Count molds, yeasts, or thermophilic spore counts. TheseThis method involves the microscopic examination counts are basically used for microbiological analy-for evaluating the viable or non-viable microbes sis of the canned fruits/fruit products.
1 Fruit Microbiology 151. Howard Mold Count. This technique involves the formats and diverse technologies that are quite spe- enumeration of molds in products such as the ciﬁc and more sensitive (Mermelstein et al., 2002). canned fruits and provides the inclusion of the Some of the assays involved in the rapid enumeration moldy material. of pathogens in food samples are as follows.2. Yeasts and Mold Counts. The high sugar prod- ucts such as fruit drinks or fruit beverages are Modiﬁcation of Conventional Techniques prone to contamination and overgrowth by yeasts and molds more than the bacterial counterparts r Miniaturized Biochemical Assays: The use of and thus enumeration of these microbes gives the certain biochemical test kits for identiﬁcation of presumptive glimpse of the microbiological status pure cultures of bacterial isolates delivers results of the product. A similar kind of count involves in less than 1 day with high accuracy of 90–99% the heat-resistant mold count providing the pres- comparable to conventional techniques making ence of molds such as Aspergillus ﬁscheri and the procedure simpler, cost- and Byssochlamys fulva in heat-processed fruit prod- performance-effective (Hartman et al., 1992). ucts such as the fruit concentrates. r Modiﬁed Process/Specialized Media: Use of3. Thermophilic Spore Count. The technique again petriﬁlms (Curiale et al., 1991) and hydrophobic advocates the presence of spore-forming bacteria grid membrane ﬁlters eliminates the need for as the major contaminants of canned fruits, fruit media preparation, thus economizes storage and beverages, and fruit juices that are being thermally incubation space as well as simpliﬁes disposal processed by pasteurization and thus speciﬁcally after analysis while the use of chromogenic enriches the spore-forming genera. (ONPG/X-gal) or ﬂuorogenic (MUG/GUD) substances provides quick measure of speciﬁc enzyme activities to quickly ascertain theNew Methods for Rapid Analysis presence of a speciﬁc microbe, and theThe physical characteristics of food result in non- bioluminescence assays provide quick assessmentuniform distribution of microbes and thus such a non- of direct live cell counts with sensitivity to provideuniform homogenate results in inconsistent presence results with low counts within few minutes.of speciﬁc pathogen providing non-reproducible re-sults following the analysis of the same sample. Thus, DNA-Based Assaysthe drawbacks of the conventional microbiologicalanalysis criteria are: Use of DNA probes technically ﬁshes out the tar-r Requirement of the selective or enrichment media get gene sequence speciﬁc to a particular pathogenic microbe in the concoction of sample DNA obtained for isolation of foodborne pathogen suffers from from the food sample with unique sensitivity and involvement of several days to provide results.r Normal microﬂora interferes with the isolation reproducibility, and has been developed for detec- tion of most of the foodborne pathogens (Guo et al., and identiﬁcation protocols of low infectious dose 2000; Feng et al., 1996; Lampel et al., 1992; Saiki and low number pathogens that may be et al., 1988; Schaad et al., 1995). However, if the sub-lethally injured during the accomplishment of target DNA contains several targets, then PCR as- a variety of processing procedures employed. says can be used in a multiplex format that ensures These microorganisms that exist in state of shock the elimination of culturing steps prior to produc- after vigorous heat/chemical/radiation treatments ing the results (Chen and Grifﬁth, 2000; Hill, 1996; need speciﬁc enriched culture media to overcome Jones and Bej, 1994). PCR protocols can detect very the shock (Jiang and Doyle, 2003). Thus, unless small number/few cells of particular pathogens and the injured cells could resuscitate, they could be have been successfully developed for various fas- easily outgrown by other bacteria in the sample. tidious/uncultivatable pathogens (Guo et al., 2000, Zhao and Doyle (2001) have reported the use of a 2002). DNA ﬁngerprinting methods are the most re- universal pre-enrichment broth for growth of cent ones for the detection of pathogens in fresh pro- heat-injured pathogens in food. duce and a semi-automated ﬂuorescent AFLP tech-Hence, these rapid methods shorten the assay time nique for genomic typing of E. coli 0157:H7 hasby a simple modiﬁcation of conventional methods been developed (Zhao et al., 2000). Another report ofor may also involve an array of molecular assay occurrence of Acidovorax avenae subsp. citrulli in