[Advances in Food Research] Advances in Food Research Volume 15 Volume 15 || Ionizing Radiation for Control of Postharvest Diseases of Fruits and Vegetables

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<ul><li><p>IONIZING RADIATION FOR CONTROL OF POSTHARVEST DISEASES OF FRUITS AND VEGETABLES </p><p>I. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147 11. Fungicidal and Fungistatic Efiects of Radiation . . . . . . . . . . . . . . 149 </p><p>A. Radiobiology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 R. Factors Affecting Dose Requirer.ients . . . . . . . . . . . . . . . . . . . . . . . 151 C. Mutations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158 </p><p>159 A. I i i Vztro Studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160 B. 1~ Viwo Studies . . . . . . . . . . . . . . . . . . . . . . . . . . . 166 </p><p>IV. The Nature and Causes of ases . . . . . . . . . . . . . . . . 166 V. Disease-Control Investigations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170 </p><p>A. General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170 B. Irradiation-Induced Susceptibility to Infection . . . . . . . . . . . . . . . . 180 </p><p>VI. Protective Packa . . . . . . . . . . . . . . . . . . . . . . . . . 182 VII. Research Needs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 183 </p><p>References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 184 </p><p>111. Techniques for Postharvest Disease Radiation Studies . . . . . . . . . . . . </p><p>I. INTRODUCTION </p><p>The bactericidal and fungicidal properties of ionizing radiation have been studied from a time soon after Roentgens (1898) dis- covery of X-rays, in 1895, and Becquerels (1896) discovery of radioactivity. Notable early reports include those of Errera (1896) on the effects of X-rays on P h y c o m ~ c e s ; Dauphin (1904) on the influence of radium on the growth and development of certain lower fungi; and Minck (1896) and Rieder (1898) on the bactericidal properties of X-rays. Many of the earliest re- searchers reported no damage to the irradiated organism, but Rieder reported decided damage to bacteria. The early investiga- tions on the biological effects of ionizing radiation have been reviewed in detail by Duggar (1936) for bacteria and by Smith (1936) for fungi. </p><p>The possibility of utilizing ionizing radiation for disease con- trol was pointed up by the successful use of X-rays to cure </p><p>147 </p></li><li><p>148 N. F. SOMMER AND R. J. FORTLAGE </p><p>dermatophytoses (Jungling, 1919 ; Levy, 1913 ; Melchoir, 1916 ; Sardemann, 1914). Similarly, Heyderdahl (1926) cured actino- mycosis with gamma rays from radium. These demonstrations that parasites could be selectively treated in host tissue suggested the possible use of ionizing radiation to eliminate certain plant parasites in seed-borne diseases. Pichler and Wober (1922) re- ported that X-rays killed Ustilugo t ~ i t i c i (Pers.) Rostr. in wheat seeds, and C. mtda (Jens.) Rostr. in barley seeds. In addition, those researchers reported elimination of the wart disease fungus, Svnchyt?*ium endobioticum (Schilb.) Percival, from potato tubers. From later studies, however, Tascher (1933) concluded that radi- ation doses sufficient to kill fungi in seeds would injure em- bryos, resulting in abnormal germination and seedling growth. More recently, Lo (1964) reached similar conclusions. </p><p>Studies have been conducted on the effect of X-radiation on the crown gall disease and its causal organism, Ag~obacteiiuni tzime- facieizs (E. F. Sm. and Town.) Conn. (Levin and Levine, 1917; Rivera, 1929 ; Waggoner and Dimond, 1952a). </p><p>The possible application of ionizing radiation to disease control is evidently severely limited by the susceptibility of host plant meristems to damage by relatively low doses. Treatments sufficient to inactivate pathogens would generally be expected to result in abnormal or inhibited growth (Dimond, 1951 ; Hellmers, 1959 ; Schwinghamer, 1957 ; Waggoner, 1956 ; Waggoner and Dimond, 1952b). Thus, deeply penetrating ionizing radiation does not ap- pear promising for therapy of plants or plant parts destined for later growth. Consequently, such treatments are evidently ruled out for seed grains and propagules such its bulbs, tubers, and cuttings. Much more promising subjects for irradiation are har- vested plant parts destined for consumption. Although the com- modities a re alive, cell division has often virtually ceased. In other cases, cell division may occur after harvest but abnormal or reduced growth from irradiation may not be objectionable. Where the effects on growth can be ignored, relatively high doses can frequently be tolerated by the host. </p><p>The present capabilities for source construction and design suggest that radiation could economically be used to treat food if benefits were sufficient. An area of particular interest in this regard is the possible use of ionizing radiation as a fungicidal treatment for the control of postharvest diseases of fruits and vegetables (Droge, 1963). Here, radiation treatments appear to have important potential advantages. Not the least among them </p></li><li><p>RADIATION FOR DISEASE CONTROL I45 </p><p>is the complete absence of chemical residues. Furthermore, radia- tion provides a fungicidal treatment uniquely different from chemical applications. The extreme penetration, particularly of gamma rays, permits the host to be considered essentially trans- parent to the rays. Therefore, organisms deep within the host tissue niay be treated as readily as if they were on the surface. Because of this feature, pathogenic organisms may be treated within host tissue to provide a therapeutic effect in a manner ordinarily not possible with chemical fungicides. </p><p>With radiation as with chemicals, control of postharvest dis- eases requires a thorough understanding of the diseases to be controlled. Particularly important are the time, place, and manner of infection; the influence of temperature and other environmental conditions on disease development ; the propensity for spread by contact ; and the physiological length of the postharvest life of the fruit o r vegetable. Extended storage following radiation may affect host injury responses (Maxie and Abdel-Kader, 1965) as well as change the relative importance of various diseases. The effectiveness of radiation treatments may therefore be determined in large measure by factors relating to disease etiology. Equally important are those facets of radiation biology which affect the fungicidal and fungistatic consequences of radiation. Therefore, these topics are considered at this time to lay a foundation for discussions of recent investigations in the area of irradiation of fresh fruits and vegetables. </p><p>1 1 . FUNGICIDAL AND FUNGISTATIC EFFECTS OF RADIATION </p><p>A. RADIOBIOLOGY Ionizing radiation is a form of energy which, if absorbed, </p><p>acts upon living cells to produce injuries. The absorption of energy ionizes and electronically excites molecules in a way that pro- duces molecular changes. The action is termed direct if the damage occurs in the molecule in which the energy has been absorbed, and is called indirect if it results from highIy reactive free radicals formed in water and reacting with cell constituents. The injury may directly damage genetic material, producing mu- tations which may or may not prove lethal. Molecular change may cause so-called biochemical lesions which are developed or in- tensified by metabolic processes of the cell. Biochemical lesions may affect vital genetic syntheses to result in a mutation which, if sufficiently severe, may result in cell death. On the other hand, </p></li><li><p>150 N. F. SOMMER AND R. J. FORTLAGE </p><p>biochemical lesions may affect nongenetic processes and result in an altered physiology which may also prove lethal to the cell. The fundamentals of radiation biology are treated adequately in a number of excellent reports. Particular attention is called to those of Bacq and Alexander (1961a), Errera and Forssberg (19611, Harris (1961), Jenkinson (1963), Kimball (1957), Lea (1955), Romani (1965), Platzman (1952), Setlow e t al. (1961), and Spear (1953). Fungi were given special attention in the reports of Beck and Roschenthaler (1960), Pomper and Atwood (1955), and Tatum (1950). Certain aspects of radiation effects are of particular, and sometimes unique, importance to studies of the use of ionizing radiation in the pathology of harvested fruits and vegetables. These are the considerations which receive primary attention here. </p><p>Ionizing radiation may, in pathogens, cause morphological ab- normalities as well as genetic mutations and altered physiology (Atwood and Pittenger, 1955a ; Brace, 1950 ; Burns, 1955 ; John- son, 1932). The morphological effects on fungi are especially striking immediately after germination. When irradiated spores are plated on a medium conducive to prompt and rapid germina- tion, the germ tubes produced often have diameters much larger than normal (Berk, 1952a, 1953; Brace, 1950; Dimond and Dugger, 1940 ; Luyet, 1932). Frequently grotesque swellings occur at various places in the mycelium. Commonly, the germ tube may grow only a short distance and then round up at the end t o produce a monster germinant resembling a dumbbell. Occurring just as commonly, however, is extensive germ tube development, including branching. The amount of germ tube development is evidently inversely related to dose. Characteristically, cross walls are almost totally lacking in species that normally form regular and prominent walls in the germ tubes. Extensive growth may occur, but a point invariably seems to be reached at which all further development stops and death follows. The absorption of energy to cause lethal damage may be separated from death by many hours. If irradiated spores are stored in a n environment not conducive to germination, the time between energy absorption and death may be extended to a number of days. During that time respiration, the production of certain enzymes, and other metabolic functions are known to proceed at a normal or acceler- ated rate (Sommer e t aZ., 1963b, 1965a). The stimulation of me- tabolism or germination and growth (Buchwald and Wheldon, 1939 ; Vasudeva e t aZ., 1959) following lethal or sublethal irradia- </p></li><li><p>RADIATION FOR DISEASE CONTROL 151 </p><p>tion must presumably require an explanation based upon cell injury . </p><p>In considering control, the relation of radiation dose to the ability of fungus spores to germinate is of little importance (Uber and Goddard, 1934). Much greater significance must be attached t o loss of the ability t o form a colony capable of indeterminate growth. In fungi the capacity for indefinite growth is lost a t a much lower dose than the ability to germinate (Beraha et al., 1959a,b). In experiments with Rhixopzts stolonifer, a dose of 500 Krad reduced the surviving fraction (i.e., ability t o form a colony) to less than 1% but hardly affected the ability to germinate (though many germ tubes produced were abnormal). Reducing germination to nea.r 1% required a dose of 1500 Krad. An even more resistant process than germination is the ability of the spore to swell prior to emergence of the germ tube (Sommer ef al., 1963a). </p><p>When colonies are irradiated at a dose insufficient to inactivate permanently, growth is halted temporarily (Beraha et al., 1959a,b; Nelson et al., 1959), to be resumed after a delay which can amount to several days. The basis for the delay is not well understood. It is likely, however, that most of the mycelium has been irreversibly injured. Only certain portions of hypha may, with time, recover. Essentially normal growth then occurs from localized areas. </p><p>B. FACTORS AFFECTING DOSE REQUIREMENTS </p><p>1. Genetic </p><p>The use of radiation in postharvest pathology obviously de- pends on the radiation sensitivity of the fungus compared with the ability of the host t o withstand the treatment with little or no obvious injury or deleterious side effects. The apparent sen- sitivity of the fungus is determined by a number of factors. One factor is the inherent resistance of a fungus to inactivation by ionizing radiation. The resistance varies widely between species, is evidently of a complex nature, and is genetically controlled. The genetics of fungi are reviewed by Fincham and Day (1963), while radiation genetics have been discussed by Wolff (1961). </p><p>In fungi it has been demonstrated, particularly with yeast, that diploid cells are more radiation-resistant than haploid cells of the same species (Fig. 1 ) (Beam, 1955; Latarjet and Ephrussi, 1949 ; Mortimer, 1954 ; Mortimer, 1961 ; Zirkle and Tobias, 1953). </p></li><li><p>152 N. F. SOMMER A N D R. J. FORTLAGE </p><p>It appears tha t the greater radiation resistance of the diploid condition is a direct result of the presence of a second set of chromosomes. The nuclear redundancy of the diploid condition presumably provides protection against recessive lethal mutations. Data f rom higher plants suggest tha t the fur ther chromosome duplication of polyploids does not impart added radiation resist- ance (Sparrow and Miksche, 1961). Similarly, in fungi, Morti- mer (1961) found tha t hexaploid yeast cells were less resistant than triploids. </p><p>I 1 </p><p>\ </p><p>IC- L --L -1- 4 S 12 16 2 0 24%,000r </p><p>FIG. 1. Survival of haploid and diploid yeast cells a f t e r X-radiation. I . IIaploid. 11. Diploid. Redrawn from La ta r j e t and Ephrussi (1949). </p><p>The nuclei of fungus spores a re usually haploid, but, commonly, two or more nuclei may be present within a single-celled spore. Here, multinuclearity imparts a resistance analogous to the efiect of ploidy. In experiments with hrewospo?.a spores, Norman (1951 ) found uninucleate microconidia to be less radioresistant than conidia containing from one to several nuclei but averaging about two (Fig. 2 ) . Similarly, Atwood and Pittenger (195513) found multinucleate Sezwospo?*n ascopores more resistant than micro- conidia. The results suggest tha t the spore can survive if one ilucleus escapes inactivation. In the case of multicellular spores the presence of the several cells in a spore would presumably impart resistance: as long as lethal injury was escaped by any one constituent cell, the spore would retain the ability to form a colony. </p></li><li><p>RADIATION FOR DISEASE CONTROL 153 </p><p>Although the relation between nuclear conditions and radiation resistance is unmistakably important, damage to the cytoplasm is also important (Kimball, 1957). Furthermore, nuclear damage may not be all genetic damage. </p><p>I./ 0 10 2 0 3 0 40 </p><p>DOSE IN QUANTA CM* x FIG. 2 . Survival of uninucleate microconidia and multinucleate (av. 2.27) </p><p>conidia of .Vcztrospoi.a oassa. Redrawn from Norman, (1951). </p><p>2. Popu la t ion S i z e </p><p>The size or amount of fungus tissue present affects the ap- parent sensitivity of a species. With filamentous fungi, direct determination of population size of...</p></li></ul>