tested, while all chiselmouths tested were 

 negative for the three extracts. Particularly 

 significant is the Cytisus sroparius seed extract 

 reaction due to the apparently high species 

 specificity of this reagent for squawfish cells. 

 On the other hand, the negative reaction of the 

 shiner-dace hybrid with lima bean extract 

 compared to the positive reaction of all shiners 

 tested suggests that the gene controlling the 

 inheritance of this reacting site may be 

 segregating in the shiner population or that 

 dosage may be involved. The soy bean extract 

 reacts for the first time in this group demon- 

 strating qualitative individual variations in 

 dace. This indicates a possible absence of 

 reacting sites specific for this extract in more 

 primitive species and is consistent with the 

 recent work of Sprague and Nakashima (1962) 

 which shows that certain individual tuna react 

 with soy bean extract. 



The reactions of two species of the cod 

 family are shown in table 6. Since eight of the 

 extracts give evidence of individual variation 

 in one or both species, it appears likely that 

 further research can produce a number of 

 phytoagglutinin reagents useful for blood typing 

 studies in this group. 



Table 7 includes species of the order 

 P ere i formes tested. The only reactions to the 

 peanut extract, including evidence of individual 

 variation in Sebastodes caurinus, are found in 

 this group. 



Reactions of three species of the family 

 Pleuroneciidae are given in table 8. The mid- 

 shipman is included in this table although 

 taxonomically distinct from the Pleuronectidae. 

 Species separation of the three pleuronectid 

 species appears possible on the basis of these 

 limited samples. The starry flounder is the 

 only flatfish reacting with the soy bean extract. 

 The sand sole is the only flatfish reacting 

 with the lima bean extract. The English sole 

 can be tentatively separated from the other 

 two species by its lack of reaction with either 

 extract. 



Results of this study indicate a relationship 

 between seed extracts and bark extracts that 



varies according to species. The Cytisus 

 scoparius seed extract was specific for the 

 squawfish and its hybrids throughout the range 

 of fish species tested, whereas the C. scoparius 

 bark extract agglutinated red blood cells of a 

 broad range of fish species but failed to 

 agglutinate squawfish erythrocytes. Other 

 direct comparisons that can be made are the 

 reactions of seed and bark extracts of Caragana 

 macrophvlum. Laburnum vulf^are. and Robinia 

 pseudoacacia. In none of these is there strong 

 evidence of a qualitative difference between 

 the seed and bark agglutinins as is found 

 in C. sfuparius. 



Variations in the strengths of reactions 

 when the cells age is a problem that has 

 complicated the interpretation of many of the 

 apparently promising reactions of the salmonid 

 group. Though the degree of variation changed 

 from one fish species-plant extract system to 

 another, cells kept over 1 week have shown 

 detectable alteration in specificity in every 

 variable system studied. The alteration of 

 specificity proceeded at approximately the 

 same rate in untreated samples as in those 

 in which an antibiotic was used to inhibit 

 bacterial growth (chlortetracycline at 80 

 parts/million). Perhaps the most striking 

 example of this alteration is seen in the 

 reaction of chinook salmon cells against lima 

 bean extract: no fresh cells reacted, but week- 

 old cells reacted strongly. A reverse situation 

 was found in the reaction of chinook cells with 

 C\tisus scoparius bark extract, for fresh cells 

 reacted strongly and 10-day old cells reacted 

 considerably less. Table 9 and figure 1 illus- 

 trate the aging phenomenon when the same 

 cells were tested at various time intervals 

 with a given reagent. The immune antiserum 

 reaction with chinook cells is included to 

 demonstrate the relative reactive stability of 

 fish erythrocytes with immune reagents when 

 compared with phytoagglutinins. The score 

 used is the sum of the agglutination strengths 

 of three consecutive serial dilutions. Each 

 individual test is scored ranging from 4 to 0. 

 A maximum score for the three dilutions is 

 therefore 12. 



11 



