Irradiation Processing of Fishand Shellfish Products
MARILYN B. KILGEN
Department of Biological Sciences, Nicholls State University, Thibodaux, Louisiana
7.1. INTRODUCTION
Humans have sought to preserve their food supply and control disease and insectinfestation from earliest times. However, the Food and Agricultural Organization ofthe United Nations (FAO) estimates that 25% of all food production and fisherycatch worldwide is lost postharvest due to insects, bacteria, and rodent infestationbecause of inadequate handling and processing facilities (ICGFI 1998, Venugopalet al. 1999). In the United States, the Council for Agricultural Science and Tech-nology (CAST) (Thayer et al. 1996) and a May 1997 presidential report, FoodSafety from Farm to Table, estimated that foodborne disease caused by bacterialpathogens and parasites causes 6-33 million cases of diarrheal disease and approxi-mately 9000 deaths annually, and is on the increase. In addition to this loss of lives,it is estimated that the annual economic losses associated with foodborne illness isas high as $5-$6 billion (Thayer et al. 1996, Henkel 1998). This does not includethe economic losses due to food spoilage postharvest.
Of all the postharvest elimination strategies for foodborne pathogens and spoil-age microorganisms, irradiation has been studied the most comprehensively, withmore than 40 years of worldwide investigation into the benefits and safety of thisprocessing technology for preservation and improvement of the microbiologicalquality and safety of foods (ICGFI 1998, Venugopal et al. 1999). The primary goalof irradiation processing is to greatly reduce or eliminate any spoilage or patho-genic microorganisms that may be present in foods without inducing sensorialchanges in the product.
Radiation processing has been approved for more than 40 different foods inabout 37 countries for human consumption (Venugopal et al. 1999). In the UnitedStates it has been approved by the Food and Drug Administration (FDA) for many
foods from wheat and flour in 1963 to fresh fruits, vegetables, dry spices, sea-sonings, enzymes, pork, poultry, and in 1997, meats (IAEA 1989, Henkel 1998,Venugopal et al. 1999). The U.S. National Fisheries Institute currently has a peti-tion before FDA to allow irradiation processing of live and processed molluscanshellfish products to eliminate potentially pathogenic naturallyoccurring Vibrio spp.bacteria and reduce other potential microbial pathogens like Salmonella spp.,Staphlococcus aureus, and Listeria monocytogenes (NFI 1999).
Many national and international agencies have actively investigated, supported,or approved the health and safety of irradiation technology to address the issues offoodborne disease throughout the world and in the United States. They include theWorld Health Organization (WHO), the Food and Agriculture Organization of theUnited Nations (FAO), the International Atomic Energy Agency (IAEA), theUSFDA, the U.S. Department of Agriculture (USDA), the American Medical As-sociation (AMA), the American Dietetic Association (ADA), the American Veter-inary Medical Association (AVMA), and the Codex Alimentarius (ICGFI 1998).Some U.S. industry trade groups supporting the technology include the NationalMeat Association (NMA), the Grocery Manufacturers of America (GMA), theNational Food Processors Association (NFPA) (Henkel 1998), and the NationalFisheries Institute (NFI) (NFI 1999). On the basis of a report of the Joint ExpertCommittee on Food Irradiation (JECFI) of the FAO/IAEA/WHO, the WHO con-cluded that a maximum dose of 1OkGy causes no toxicological hazard and intro-duces no nutritional or microbiological problems (WHO 1994, Venugopal et al.1999). On the basis of these findings, the Codex Alimentarius Commission devel-oped a General Standard for Irradiated Foods and a recommended InternationalCode of Practice for the Operation of Radiation Facilities Used for the Treatmentof foods (CAC 1984)
Aquatic or fishery products are an important and basic source of protein, buthave a relatively short shelf life unless frozen onboard or very shortly after harvest-ing. The shelf life of fresh, frozen, and processed marine and fresh fish and shellfishcan also be extended with irradiation processing using proper handling and packag-ing, and "vessel to table" Hazard Analysis Critical Control Point (HACCP) pro-grams with prerequisite Sanitary Standard Operating Procedures (SSOP's) andGood Manufacturing Practices (GMPs). Depending on the product and intendeduse, the effectiveness of irradiation processing can be enhanced with combinationtreatments such as heat, freezing, and "generally recognized as safe" (GRAS)preservatives. Irradiation processing of seafood products has also been shown tobe effective in eliminating potentially harmful microbiological pathogens fromeconomically important fresh and marine seafood species. (IAEA 1989, Grodnerand Andrews 1991, NAS 1991, Chen et al. 1996, ICGFI 1998, Kilgen et al. 1999,Venugopal et al. 1999).
The International Consultative Group on Food Irradiation (ICGFI) has compiledan extremely comprehensive monograph on irradiation of seafood products with192 worldwide references (ICGFI 1998). Additionally, Venugopal et al. (1999)compiled a critical review of 323 international references on radiation processingto improve the quality of fishery products.
The present chapter is intended to provide an update on irradiation of fish andother seafood. No attempt has been made to review the extensive literature pub-lished on this topic during the 1950s and 1960s. The interested reader is referred tothe excellent reviews by Josephson and Peterson (1983) and the IAEA (1989) for aninsight into the earlier literature, and to those by the ICGFI (1998) and Venugopalet al. (1999) for a more international perspective.
7.2. IRRADIATION FOR SHELF LIFE EXTENSIONOF SEAFOOD PRODUCTS
7.2.1. Finfish Products
The shelf life of fresh finfish is relatively short because these products are extre-mely susceptible to growth of spoilage organisms that are part of their naturalmicroflora. Microbial spoilage is the primary mechanism of spoilage of chilled fishand shellfish products (ICGFI 1998). The shelf life of fresh finfish is largely de-pendent on how it is handled immediately after catch and the time on board that itremains iced before it is processed. This is especially important in warm areaswhere greater amounts of ice must be stored on board. Highly perishable fresh fishwill spoil in 2-6 h if not iced (ICGFI 1998). Poor sanitary and quality practicesaboard fishing vessels is one of the main causes of spoilage and waste of fresh fishcatch (Venugopal et al. 1999). The FAO's Recommended International Code ofPractice for Fresh Fish (FAO 1973) indicates that fish should be immediatelycooled to melting ice temperature (—1 to -2 0 C). Since psychrophilic species ofbacteria are able to survive in iced fish, high-value fishery products can be preparedonly from fish iced for up to 5 days. By two weeks of ice storage, only low-qualityproducts can be prepared (Venugopal et al. 1999). Therefore, the application ofirradiation to prolong shelf life and quality of fresh fishery products should bewithin a few days of ice storage to produce the highest-quality product with anextended shelf life.
7.2.7.7. Normal Microflora of Finfish The initial microflora of finfish isdependent on the environment and can change seasonally. The natural flora aremainly in the outer slime layer of the skin, on the gills, and in the intestines. Themuscle flesh is initially sterile. The intestines of both marine and freshwater fishcontain the genera Achromobacter (Moraxelld), Pseudomonas, Flavobacterium,Vibrio, Bacillus, Clostridium, and Escherichia (ICGFI 1998). Fresh water fishadditionally harbor species of Aeromonas, Lactobacillus, Brevibacterium,Alcaligenes, and Streptococcus (ICGFI 1998, Venugopal et al. 1999).
Studies on the microflora of fresh marine fish caught in the north and mid-Atlantic have indicated that the predominant natural bacterial flora on gills, intes-tines, and slime of freshly caught fish and shellfish consists of Pseudomonas andAchromobacter (Moraxelld) sp. Species of Corynebacterium, Flavobacterium, andMicrococcus were a smaller percentage of the natural flora isolates (ICGFI 1998).
The psychrotrophs, Pseudomonas, Achromobacter (Moraxelld), and Flavobacter-ium, are typically associated with fish spoilage and impart slimy, smelly character-istics (Grodner and Andrews 1991).
In general, warm-water marine fish have more mesophilic Gram-positive micro-organisms, including Micrococcus, Bacillus, and Corynebacterium spp., and cold-water fish have a predominantly Gram-negative psychrophilic population, includingMoraxella, Acinetobacter, Pseudomonas, Flavobacterium, and Vibrio spp. (Mayerand Ward 1991, ICGFI 1998). The microflora of fish from temperate waters in-cludes Moraxella, Vibrionaceae, Pseudomonas, Alteromonas, Aeromonas, andFlavobacterium. Gram-positive microorganisms such as Bacillus, Micrococcus,Clostridium, Lactobacillus, and Corynebacterium are also found in fish from tem-perate waters (Venugopal et al. 1999).
7.2.1.2. Spoilage of Finfish Products Finfish spoilage is due to the presenceof indigenous microorganisms. A large number of these are psychrotophic and cangrow at O0C and higher. These organisms include Pseudomonas, Acromobacter(Moraxelld), and Flavobacterium spp. Shewanella putrefaciens and Pseudomonasspp. are the main spoilage bacteria of fresh fish iced 0-6 h after landing regardlessof the origin of the fish (Mayer and Ward 1991, ICGFI 1998, Venugopal et al.1999). Prolonged delays in icing result in a spoilage flora consisting of Bacillusspp., Aeromonas hydrophila, and Pseudomonas spp. (Mayer and Ward 1991).
7.2.1.3. Shelf Life Extension of Finfish Products by Irradiation Low-dose irradiation has been studied extensively as a method for extending the shelflife of fresh iced or refrigerated fish. Generally, Gram-negative bacteria are moresensitive to radiation than Gram-positive bacteria and tend to be mainly responsiblefor spoilage of fresh and marine finfish. This means that low dose levels of 1-3 kGy(cold pasteurization) can often reduce the initial load of potential spoilagemicroorganisms 1-3 Iog10 cycles to extend the shelf life of fresh fishsignificantly. In some investigations, doses up to 5 kGy were required. The doselevel for shelf life extension must be evaluated for each species to maintainacceptable flavor, texture, and wholesomeness (Grodner and Andrews 1991).This is crucial for more fatty fish that are more susceptible to rancid off-flavorsdue to oxidation by radiolytic hydroxyl ions (see Chapter 3). It is imperative thatonly high-quality products be irradiated for shelf life extension. Irradiation cannotimprove poor-quality or spoiled fish products. The longer the product is heldwithout proper icing or refrigeration, the less effective irradiation will be forshelf life extension. Selection of dose also requires evaluation of the productstorage temperature, origin, previous handling and processing, packagingmaterials, size, and density (ICGFI 1998, Venugopal et al. 1999).
The ICGFI (1998) monograph on irradiation of fish and shellfish provides anextensive literature review of each species of marine finfish for which data wereavailable concerning the minimum dose necessary to extend shelf life and themaximum acceptable dose that would not cause detrimental sensory effects. Fatcontent was a major consideration for maximum dose levels. Many high-fat fish
have to be vacuum-packaged or packaged in modified atmosphere to preventoxidative rancidity and other sensory changes such as bleached color. However,vacuum packaging often induces drip loss and provides conditions for potentialoutgrowth of Clostridium botulinum spores if the product is temperature abused.
Tables 7.1 and 7.2 present selected data on the relationship between dose,storage temperature, and shelf life extension achieved with various species offreshwater and marine finfish, respectively, adapted from the excellent reviews ofNickerson et al. (1983), ICGFI (1998) and Venugopal et al. (1999). [The Interna-tional Atomic Energy Agency (IAEA 1989) also published a technical report onradiation preservation of fish and fishery products.] This table contains data offresh iced fish products held near O0C. However, Bari et al. (2000) discoveredthat a combination treatment of irradiation at 5kGy and ascorbic acid wouldextend the shelf life of processed fish cutlets for up to 5 weeks at ambienttemperature.
7.2.2. Shellfish and Crustaceans
7.2.2.7. Normal Microflora of Molluskan Bivalve Shellfish: Oysters,Clams, Mussels, and Scallops The normal microflora of molluskanbivalve shellfish is greatly dependent on their aquatic environment because theyare filterfeeders that concentrate potential nutrients and particles from the largequantities of water that they filter through their gills. Environmental parametersaffecting such microflora include temperature, salinity, nutrients, and fecal orsewage pollution of surrounding waters. In a review, Cook (1991) notes thatvarious studies have shown that the normal microflora that could be related tospoilage in bivalves consist primarily of Gram-negative rods like Pseudomonasand Vibrio species. Flavobacterium, Acinetobacter, coryneforms, Achromobacter(Moraxella), Alcaligenes, Micrococcus, and Bacillus are also often present inshellfish such as American Pacific and Eastern oysters. MoraxellalAcinetobacterhas been mistaken for Achromobacter in some earlier studies (Cockey andChai 1991). Microflora of oysters from the Gulf of Mexico are mainlyVibrio, Aeromonas, Moraxella, and Pseudomonas species (Cook 1991). In chilledoysters, Pseudomonas and Achromobacter (Moraxella) are often mainly
TABLE 7.1. Shelf Life Extension of Selected Freshwater Finfish Irradiated at Low toMedium Doses
Source: Adapted from Nickerson et al. (1983), ICGFI (1998), and Venugopal et al. (1990).
Sources: Adapted from Nickerson et al. (1983), ICGFI (1998), and Venugopal et al. (1999).
responsible for spoilage, in addition to Flavobacterium and Micrococcus.Presence of glycogen and sugars in oyster meat will result in spoilage withsouring due to acid formation by enteric and lactic acid producers likeStreptococcus (ICGFI 1998).
7.2.2.2. Normal Mlcroflora of Crustaceans: Shrimp and Prawns, Crabs,Lobster, and Crawfish Like other seafood species, the microflora of shrimp,crabs, lobsters, and crawfish reflect their geographic environment and are thussubject to change or variation due to seasonal, temperature, and waterquality factors (Cockey and Chai 1991, ICGFI, 1998, Venugopal et al. 1999). Ingeneral, most researchers have reported that seafood from tropical or temperatewarmer waters initially have a microflora that is composed predominately ofmesophilic Gram-positive bacteria such as Micrococci, and Gram-negativeVibrionaceae. They have fewer numbers of the potential psychrotrophic Gram-negative microorganisms such as Pseudomonas and Moraxella species (Miget1991). The different physiological characteristics of the initial microflora ofwarm- and cold-water crustaceans have led some researchers to conclude thatfresh crustaceans held on ice will spoil more quickly if they are from cold-waterenvironments than will those from warmer-water environments, because theirpredominant microflora is potentially psychrotrophic, Gram-negative spoilageorganisms (Miget 1991).
Miget (1991) published a review of the main bacterial genera found in freshshrimp from the Gulf of Mexico and from the Pacific coast. The genera mostpredominant in Gulf of Mexico (warm water) fresh shrimp (Penaeus sp.) includedVibrio, Pseudomonas, Moraxella, and Acinetobacter. Penaeus shrimp in aquacul-ture ponds also contained Aeromonas. Species of Pandalus shrimp from thePacific (cold water) contained Arthrobacter, Micrococcus, Bacillus, Pseudomonas,
TABLE 7.2. Shelf Life Extension of Selected Marine Finfish Irradiated at Low toMedium Doses
of frozen shrimp from Japan, the predominant bacteria in the samples werethe psychrotrophs Moraxella-Acenitobacter, Flavobacterium, Arthrobacter, andMicrococcus, with fewer numbers of Pseudomonas, Corynebacterium, andStreptococcus sp. (Venugopal et al. 1999). Pseudomonas spp. were the predominantspoilage organisms in the freshwater prawn Macrobrachium rosenbergii stored atrefrigerator temperatures (Angel et al. 1986). Decapitation of shrimp on board canreduce 75% of the bacterial load, but can also contaminate the vessel holdingfacilities (Miget 1991, ICGFI 1998).
Vibrio parahaemolyticus has been found in blue crab hemolymph. Other speciesof Vibrio, Pseudomonas, and Acinetobacter are the dominant microorganismsin blue crabs. Aeromonas, Bacillus, and Flavobacterium have also been associatedwith blue crabs. Clostridium botulinum has occasionally been found in the gillsand viscera. Cockey and Chai (1991) compiled an extensive list of bacteria foundin four different types of commercial raw crab species, including blue crab,Dungeness, rock, and Tanner (Bering Sea). Organisms listed were from the naturalenvironment, potential sewage pollution, and from processing and handling.Chilled crab meats are spoiled mainly by Pseudomas, which may have beenmistaken for Proteus and Alcaligenes in some early studies, and Moraxella-Acinetobacter, which has been mistaken for Achromobacter (Cockey and Chai1991).
The microflora of fresh crawfish have been found to consist of species ofMicrococcus, Staphylococcus, and Alcaligenes. It was noted that with storage, thismicroflora changed as the spoilage psychrotrophs became more predominant. AtO0C, Pseudomonas became more dominant than the Achromobacter sp. (Moraxelld).At 50C, the shift in the spoilage population was more toward Achromobacter(Moraxelld) than Psuedomonas sp. (Miget 1991). In fresh chilled crab meat, asimilar spoilage was noted with Pseudomonas sp. at O0C and Moraxella sp. at50C, but additionally, at chilled storage temperatures greater than 1O0C, Proteussp. were the main spoilage organisms (ICGFI 1998).
7.2.2.3. Shelf Life Extension of Fresh Shellfish Using Low- andMedium-Dose Irradiation Many commercially important shellfish worldwidehave been evaluated for shelf life extension by low-dose irradiation. Tables 7.3 and7.4 present compilations of selected data adapted from several extensive reviews.
Kilgen et al. (1988), using an experimental irradiator at Louisiana StateUniversity, determined the LD50 of live shellstock Eastern oysters (Cmssostreavirginicd) to be 2.25 kGy. This is significant because later work by Kilgen et al.(1995, 1999) and Kilgen and Hemard (1996) in a commercial irradiation facilityin Florida, using commercial, 60-lb boxes of select shellstock (about 200 oystersper box), showed that a sublethal dose of 1.5 kGy was optimum for elimination ofVibrio vulnificus, an indigenous potential human pathogen in estuarine waters. Thisdose also extended the shelf life of commercially shucked and packaged oyster meatfrom 14 to 24 days at a storage temperature of 40C in a commercial cooler.
Sources: Adapted from Nickerson et al. (1983), ICGFI (1998), Kilgen et al. (1999), and Venugopal et al.(1999).
7.3. POTENTIAL HUMAN PATHOGENS OF PUBLICHEALTH CONCERN IN SEAFOOD PRODUCTS
Pathogens that represent a potential public health risk to consumers of fresh andfrozen finfish and shellfish products are from three main potential sources (NAS1991)
1. Indigenous species from the natural aquatic environment.2. Nonindigenous species introduced into the aquatic environment from human
and/or animal sewage or fecal wastes.3. Microorganisms associated with the processing, distribution, and storage envir-
onment:
a. from human and/or animal cross-contaminationb. from cross-contamination of raw products with processed or cooked pro-
ducts;c. from time or temperature abuse of fresh products that allows outgrowth of
both indigenous and nonindigenous mesophilic species.
7.3.1. Indigenous Potential Pathogens Associatedwith the Natural Aquatic Environment
A number of potential human pathogens may be indigenous to fresh, estuarine, andmarine waters used for harvest, mariculture, and aquaculture of commercially im-portant seafood species (NAS 1991, Rodrick, 1991, Garrett et al. 1997). Many ofthe species that are associated with freshwater environments are also found in
TABLE 7.3. Shelf Life Extension of Selected Fresh Molluskan Bivalve ShellfishUsing Low-Dose Ionizing Irradiation
Lobser — American (Homarus gammarus)Lobster — Norwegian (Nephrops norvegicus)
TABLE 7.4. Shelf Life Extension of Selected Fresh Crustacean Shellfish Using Low-Dose Irradiation
Sources: Adapted from Angel et al. (1986), Chen et al. (1996), ICGFI (1998), and Venugopal et al. (1999).
estuarine and marine environments. Those generally associated mostly with freshwater include Listeria mono cy to genes, Aeromonas hydrophila, Yersinia enterocoli-tica, Plesiomonas shigelloides, and Clostridium botulinum type E. Additionally, theprotozoan parasite, Giardia lamblia, has been linked to fishborne illness (CDC1989, NAS 1991). These same highly ubiquitous microorganisms can also be in-digenous to estuarine and marine vegetation and waters.
The Vibrionacea are the most significant potential indigenous pathogens in mar-ine and estuarine waters. They include species of Vibrio, Aeromonas, and Pleisio-monas. About 11 of the 66 species of vibrios in marine environments can causeillness in humans under certain circumstances. Some of the species of vibrios asso-ciated with seafoodborne illness include V. parahaemolyticus, V. cholerae Ol, V.choleraenon-Ol, V.vulnificus, V.mimicus, V hollisae, V.fluvialis, V. alginolyticus, V.furnissii, and V. damsiella. V. cholerae, V. mimicus, Aeromonas sp., and Pleisomonassp. may also be found in fresh waters. Of these, V. cholerae and V parahaemolyticusare the only two species that have caused reported outbreaks involving more thantwo individuals (NAS 1991, Rodrick 1991). An epidemic of V cholerae ongoing inSouth and Central America since the mid-1990s has resulted in more than 200,000cases and thousands of deaths (Albert et al. 1997). In Asian countries, a majority ofseafood-associated illnesses are attributed to V. parahaemolyticus. The other speciesof vibrios cause isolated and sporadic incidents of disease in single individuals, andoverwhelmingly in high-risk individuals who have predisposing underlying diseaseof the liver, blood, stomach, or immune system, which may include one or a combi-nation of the following underlying illnesses: liver disease, alcoholism, diabetes,peptic ulcer, renal disease, gastric surgery, heart disease, haematologic disease,immunodeficiency (including AIDS), and cancer, or who are undergoing chemother-apy (CDC 1989, Rodrick 1991, NAS 1991).
A study by the U.S. National Academy of Science's Committee on Evaluation ofthe Safety of Fishery Products and data from the Centers for Disease Control(CDC) and the U.S. FDA for the period 1978-1987, revealed few cases of fin-fish-associated illnesses from indigenous pathogens. There were only two casesfrom Vibrio cholerae Ol and 29 cases from Giardia lamblia (CDC 1989, NAS1991). Listeria monocytogenes has been recovered from finfish, smoked fish, andsurimi-based products. However, there was only one outbreak involving 29 cases inNew Zealand where the cause was not discovered, but raw fish and shellfish werepossibly implicated (Kvenberg 1991, Venugopal et al. 1999).
Indigenous microorganisms that have been implicated in outbreaks and cases offoodborne illness have been mainly associated with .molluskan shellfish that wereconsumed raw or partially cooked (Cook 1991, Kilgen 1991, NAS 1991, Garrettet al. 1997). The indigenous Vibrionaceae have been the main concern since 1980in cases of foodborne illness associated mainly with raw molluskan bivalve shell-fish. In the reporting period 1978-1987, the CDC and FDA reported 52 cases ofshellfish-associated illnesses from V parahaemolyticus, 27 cases from V choleraeOl, 131 cases of V cholerae non-Ol, 100 cases of V vulnificus, 5 cases of Vmimicus, 5 cases of V fluvialis, 5 cases of V hollisae, 18 cases of Plesiomonasshigelloides, and 7 cases of Aeromonas hyprophila. In more recent years,
V. vulnificus has been considered the most significant indigenous potential pathogenof public health concern in molluskan shellfish products because it is the maincause of mortality associated with consumption of seafoods in the United States(CDC 1996). As of August 2000, there were 11 cases of V vulnificus with sixassociated deaths in the United States (EPA 2000).
7.3.2. Potential Pathogenic MicroorganismsAssociated with Human and/or Animal Fecal Pollution
Potential pathogens associated with human and/or animal fecal pollution are theenteric bacteria and viruses. Protozoan parasites that are sewage related includeCryptosporidia and Cyclospora. However, the literature shows that the majority ofseafood-associated illnesses from raw and partiallycooked shellfish is due to theNorwalk and Norwalk-like human enteric viruses (Kilgen and Cole 1991, NAS1991).
7.3.2.1. Enteric Bacteria In the United States, only a few cases of illnessfrom enteric bacteria were reported to be associated with seafood consumption in1989 and the 1990s, and most of these cases were from recontamination orcross-contamination by food handlers and not from fecal pollution of thegrowing and harvest waters (CDC 1989, NAS 1991, Rippey 1994). The mostimportant potential enteric bacterial pathogens from human and/or animal sewagepollution of estuarine waters include Salmonella spp., Shigella spp, Campylobacterjejuni, Yersinia enterocolitica, and enterotoxigenic strains of Escherichia coli(Kvenberg, 1991, NAS 1991). Salmonella is an important pathogen that can betransmitted by both humans and animals. Kvenberg (1991) noted that the earlyliterature indicates that Salmonella sp. could be isolated from fish handled in highlypolluted fresh and marine waters, and that the fish often remained positive forSalmonella as long as 30 days after the initial contamination. Salmonella fromcontaminating runoff from farms, livestock, or feeds is also a potential problemin aquaculture ponds and facilities. Farm raised catfish in the southeastern UnitedStates had a 5% incidence of contamination with Salmonella in one study.Salmonella has also been found in frog legs and smoked fish. Shigella species,mainly S. flexneri and S. sonnet, are generally waterborne pathogens. Infectedhumans handle seafood or dump infected sewage into harvest waters (Kvenberg,1991). Salmonella and Shigella have been a significant problem in frozen seafoodfrom some countries (Kvenberg 1991, Venugopal et al. 1999); these seafood-associated pathogens may originate from sewage pollution of the aquaticenvironment or cross contamination from improper processing, handling, andpreparation (NAS 1991).
7.3.2.2. Enteric Human Viruses The human enteric viruses, Norwalk andNorwalk-like viruses, hepatitis A virus (HAV), and hepatitis E virus (HEV)(Kilgen 1991, NAS 1991) have the potential of contaminating seafood harvestwaters from partially treated or untreated human sewage. These viruses tend to
be more of a problem in raw shellfish. In all seafood products they may also beassociated with improper processing, handling, and preparation (NAS 1991). In theNational Academy of Sciences "Seafood Safety" study for the period 1978-1987,the CDC reported three outbreaks of Hepatitis A with 33 associated cases. Therewere only 42 documented cases of Norwalk and related viruses. However, therewere about 3500 cases of seafood-associated illnesses listed as "unknown"etiology. Although these were not all microbiological pathogens, the majorityexhibited the typical pathology of Norwalk and Norwalk-like Caliciviruses.
7.3.3. Potential Pathogenic MicroorganismsAssociated with Processing and Preparation
7.3.3.1. Bacterial Pathogens Pathogenic bacteria often associated withcontamination, recontamination, or cross-contamination of seafood productsinclude Vibrio parahaemolyticus, Clostridium perfringens, Clostridiumbotulinum, Salmonella (nontyphoidal), Shigella, Staphylococcus aureus andBacillus cereus. Listeria monocy to genes is an ubiquitous bacteria commonly as-sociated with the seafood processing environment. (NAS 1991). In the reportingperiod 1978-1987, the CDC reported 15 outbreaks of V parahaemolyticus with 176cases, 3 outbreaks of C. perfringens with 74 cases, 3 outbreaks of nontyphoidalSalmonella with 67 cases, 26 outbreaks of C. botulinum with 38 cases (all due tohome preparation of ethnic finfish products). Additionally, there were 3 outbreakswith 60 cases of Shigella, 2 outbreaks of Staphylococcus aureus with 12 cases, and3 outbreaks of B. cereus with 10 cases. All were associated with improperprocessing and preparation (CDC 1989, NAS 1991).
The possibility of botulinal toxin being formed in temperature-abused irradiatedseafood products by C. botulinum type E, which is able to grow at refrigeratedtemperatures above 3.30C and is the predominant type in seafood, has hamperedthe clearance of irradiation of seafood to this date. Of particular concern hasbeen the potential for botulism from products irradiated and then packaged invacuum or modified atmospheres. However, it has generally been shown that iffish products are kept below 30C, botulinal toxin production would not take place(ICGFI 1998).
7.3.3.2. Enteric Human Viruses The human enteric viruses, Norwalk andNorwalk-like viruses, hepatitis A virus (HAV), and hepatitis E virus (HEV), havethe potential to be more of a problem in raw shellfish and other seafood products,and may also be associated with improper processing, handling, and preparation. Inthe United States from 1978-1987, the CDC reported 2 outbreaks with 92 cases ofHAV due to contamination of fin-fish products from infected food handlers, and 33cases from shellfish washed with water contaminated with human sewage (Kilgen1991, NAS 1991). The vast majority (>3000 cases) of seafood-associated illnessesin the NAS "Seafood Safety" report were of an "unknown etiology." Although notall were of microbiological origin, the pathology of many cases was consistent with
that of the Calicivirus Norwalk virus or Norwalk-like viruses (Kilgen 1991, NAS1991).
7.4. LOW- AND MEDIUM-DOSE IRRADIATIONFOR PATHOGEN CONTROL IN SEAFOOD PRODUCTS
It is extremely important from both public health and economic standpoints tocontrol potential human pathogens in seafood products. European countries havecommon marketing standards for certain fishery products. Both the FDA and theEuropean common marketing standards require zero tolerance for Salmonella infishery products. The FDA also requires zero tolerance for Listeria monocytogenesin ready-to-eat seafood products. The standards on Staphylococcus aureus andEscherichia coli are designed to serve as indices of poor hygiene, as they pointto contamination from human and/or animal sources.
There is also a standard on the total mesophilic aerobic plate count related toshelf life and quality (Venugopal et al. 1999). The FDA, in collaboration with theInterstate Shellfish Sanitation Conference (ISSC) and the National Shellfish Sanita-tion Program (NSSP), is currently considering a possible guideline for reduction ofVibrio vulnificus and toxigenic strains of V parahaemolyticus that would potentiallyrequire some type of postharvest treatment of raw oysters.
There is a very large body of scientific information concerning the effectivenessof low, medium, and even high levels of ionizing radiation on fresh, frozen, andprocessed seafood products (Giddings 1984, Kilgen et al. 1988, 1999; Grodner andAndrews 1991; Kilgen and Hemard 1996; ICGFI 1998; Cisneros et al. 1999; Gelliet al. 1999; Lopez 1999; Torres et al. 1999; Venugopal et al. 1999). In general,Gram-negative bacteria are more sensitive to ionizing radiation than Gram-positivemicroorganisms. Spores are extremely resistant, and the Gram-positive bacteriaMicrococcus (Deionococcus) radiodurans and Micrococcus radiophilus are extre-mely resistant due to their highly efficient DNA repair enzymes (ICGFI 1998;Venugopal et al. 1999). A dose of 4kGy has been found to be sufficient to eliminatenon-spore-forming pathogens in many different kinds of foods, including frozenseafoods (Giddings 1984; IAEA 1989; Grodner and Andrews 1991; ICGFI 1991,1998; Torres et al. 1999; Venugopal et al. 1999). The naturally occurring Vibriospecies are relatively sensitive to low-dose irradiation, and can generally be easilyeliminated, as has been determined in vitro in saline or seafood homogenates, andin vivo by inoculation, or seeding, or by natural contamination of the product.Kilgen et al. (1995, 1999) evaluated Gulf of Mexico live shellstock oysterscontaminated with naturally high levels of Vibrio vulnificus [4.6 x 105 MPN(most probable number)per gram]. Commercially harvested, processed, and pack-aged shellstock oysters were irradiated at doses of 0.5-2.OkGy. Reduction to 0.9MPN/g occurred with the 1.0-kGy dose, which and had no significant effect onmortality of the live shellfish after 14 days of storage at 4O0C.
Tables 7.5 and 7.6 are compilations of data from various reviews and researchpapers. However, while the experimental work represented in Table 7.5 was done in
Aeromonas (sp. not identified)Aeromonas hydrophila
Clostridium perfringensClostridium botulinum type EEscherichia coliSalmonella (sp. not identified)Salmonella typhimuriumSalmonella enteritidisStaphylococcus aureus
Vibrio cholerae (unspecified)
V cholerae Ol El Tor
Vibrio parahaemolyticusVibrio vulnificus
aCFU/g = colony-forming units per gram.^Radiation dose necessary to reduce bacterial numbers to a nondetectable level.Sources: Adapted from Giddings (1984), IAEA (1989), Grodner and Andrews (1991), ICGFI (1991), ICGFI (1998), Torres et al. (1999), and Venugopal et al. (1999).
TABLE 7.5. In vitro Reduction or Elimination of Some Potential Pathogens in Fresh and Frozen Finfish and Frog Legs through Low-andMedium-Dose Irradiation
Extinction Dose^(kGy)
.2
.5
.2
.2
.2
.0
.0
.00.31.5
>2.02.5
Initial Contamination
(CFu/gr107 (inoculated)
5 XlO 5
106-108
106-108
106-108
104
107
104
107
104
104
1010
Product(s)
Oysters, live (inoculated) (Crassostrea virginica)Oysters, live (natural contamination) (Crassostrea virginica)
flCFU/g = colony-forming units per gram.^Radiation dose necessary to reduce bacterial numbers to a non-detectable level.Sources: Adapted from Giddings (1984), Kilgen and Bernard (1988,1995,1999), Grodner and Andrews (1991), ICGFI (1998), Cisneros et al. (1999), Gelli et al. (1999),Lopez (1999), Torres et al. (1999), and Venugopal et al. (1999).
TABLE 7.6. Reduction or Elimination of Some Potential Pathogens in Fresh and Frozen Shellfish Products Using Low- and Medium-DoseIrradiation
vitro in fish or shellfish medium artificially inoculated with the pathogens, Table 7.6presents results of experimental work in vivo using inoculated commercial speciesof fish and other seafood.
7.5. RESEARCH NEEDS IN SEAFOOD IRRADIATION
Studies on commercial-scale harvesting, processing, packaging, transportation, anddistribution of seafood products treated with ionizing irradiation in bulk at com-mercial irradiation facilities is greatly needed to update the vast body of literatureon irradiation processing technology that is available but may not relate to currentcommercial practices. Much of the work that has been done in the past was con-ducted under experimental conditions, and often with pathogens seeded or inocu-lated artificially into products. The incidence of and doses required for reduction ofVibrio spp. numbers below detectable levels in many products are fairly wellcharacterized. However, the effectiveness of irradiation on other microbial patho-gens in vivo for elimination in processed seafood on the commercial scale is notwell characterized. Comprehensive commercial evaluation of dose levels andpackaging for optimal doses for both spoilage microorganisms and potentiallypathogenic microorganisms in commercial species is very important.
7.6. THE FUTURE OF SEAFOOD IRRADIATION
A database of clearances of all seafood products approved for irradiation from allcountries worldwide through 1998 is maintained by the International ConsultativeGroup on Food Irradiation (IGCFI 1998). It includes product, country, type ofclearance (e.g. disinfestation, microbial control, shelf life extension, and parasitecontrol), date of clearance, and maximum doses (kGy). Approximately 20 countriesworldwide have approved various applications and doses for different seafoodproducts, including fresh and frozen fish, dried fish, fish powder, fish products,fresh or frozen frog legs, fresh and frozen "seafood," shellfish, shellfish powder,and fresh and frozen shrimp. To date, the United States has approved irradiation ofpork, poultry, and red meat, but is still reviewing applications for similar clearancesfor fish and other seafood. The National Fisheries Institute currently has a petitionbefore the FDA to approve irradiation of live, shucked, fresh and frozen molluskanshellfish at levels from 0.5 to 7.5 kGy, depending on the species and product form ofthe shellfish (NFI 1999).
The future of seafood irradiation is largely an economic issue of supply anddemand. It depends on the willingness of industry to invest in irradiation of pro-cessed and packaged seafood products to offer the consumer a value-added productwith a higher level of quality and safety, and on whether consumers will demandthese safer, higher-quality products.
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