three Atlantic fishing areas. J. Fish. Res. Board Can. 

 11:171-197. 



Simpson, a. C. 



1951. The fecundity of the plaice. Fish. Invest. Minist. 

 Agric. Fish. Food (G.B.), Ser. II, 17(5), 27 p. 



Snedecor, G. w., and w. G. Cochran. 



1967. Statistical methods. 6th ed. Iowa State Univ. 

 Press, Ames, 593 p. 

 TYLER, A. V., AND R. S. DUNN. 



1976. Ration, growth, and measures of somatic and organ 

 condition in relation to meal frequency in winter flounder, 

 Pseudopleuronectes americanus, with hypotheses regard- 

 ing population homeostasis. J. Fish. Res. Board Can. 

 33:63-75. 



Department of Zoology 

 University of Rhode Island 

 Kingston, RI 02881 



Division of Biological Sciences 

 University of Michigan 

 Ann Arbor, MI 48109 



W. HUNTTING HOWELL 



DAVID H. KESLER 



structurally with cheesecloth was devised. The 

 mock fish allowed us to control: total number and 

 composition of the microbial flora; location of mi- 

 crobial contamination, e.g., surface or evenly dis- 

 persed throughout the sample; uniformity of dis- 

 tribution of microbes from sample to sample; size 

 and thickness of the samples; and the handling 

 history and physiological state of the samples. 

 This system permits the quantitative recovery of 

 the inoculated microbes by simply melting the 

 gelatin at 31°-32°C. 



This note describes the application of mock fish 

 in studying the effects of disodium ethylenedi- 

 amine tetraacetate (EDTA, Fisher Scientific Co. 1 ) 

 with or without an iodophor (Wyandotte Co.) con- 

 tained in ice for controlling microbial outgrowth of 

 a mixture of four Pseudomonas species. This pro- 

 cedure is not recommended as a means of predict- 

 ing the effectiveness of an inhibitor on a specific 

 species of fish. Its role is to screen inhibiting 

 agents for general effectiveness and to permit a 

 comparison among them. 



"MOCK FISH" METHOD FOR STUDYING 

 MICROBIAL INHIBITING AGENTS 



Materials and Methods 



Mixture of Pseudomonas Species 



In experiments intended to study the effects of 

 various agents or conditions on the microbial out- 

 growth in food products, it is desirable to approach 

 efficacy similar to those conditions of actual han- 

 dling and marketing. However, in experiments on 

 fishery products, when one wishes to find effects of 

 an agent or condition, the use of whole fish or fish 

 fillets adds variables to any experimental design 

 These undesired variables are: variations in the 

 total microbial population and in the composition 

 of the microbial flora from fish to fish; different 

 time intervals and other storage variations in the 

 handling history offish even from the same catch; 

 different fillet or sample thicknesses which will 

 affect the counts per gram ratio from sample to 

 sample; different physiological conditions, age, 

 wounds, etc., of the fish which might affect ex- 

 perimental comparisons; and possible presence of 

 inherent antibiotics in the substrate. The latter 

 variable does not permit a separation of the an- 

 tibiotic effects of the additives from the antibiotic 

 effects of the substrate. 



In order to study what effects agents might ac- 

 tually have on specific microbial outgrowth in an 

 efficacious situation, a "mock fish," composed of 

 gelatin (containing nutrients) and supported 



Four Pseudomonas species, previously isolated 

 from iced fish in our laboratory, were used in these 

 experiments. Each species of Pseudomonas was 

 grown in separate Eugon Broth (BBL) test tube 

 culture for 18 h at 20°C. Then 2 ml from each 

 culture were pooled and well mixed in a sterile test 

 tube to prepare an inoculum mixture. From this 

 mixture 1 ml was inoculated into 1 liter of melted 

 gelatin medium described below to give an esti- 

 mated 10 4 to 10 5 bacteria/ml of the final prepara- 

 tion. 



Mock Fish Preparation 



1 ) Cheesecloth discs were cut to size to fit inside 

 glass Petri dishes, and then they were cut in half. 

 The Petri dishes were then sterilized at 121°C for 

 15 min. 



2) Ten milliliters of melted, inoculated 10% 

 gelatin and 1% Eugon Broth medium were pi- 

 petted into each sterile Petri dish. A sterile needle 

 was used to make sure that the cheesecloth disc 

 halves did not overlap during gelatin solidifica- 



1 Reference to trade names does not imply endorsement by the 

 National Marine Fisheries Service, NOAA. 



880 



