SINDERMANN: INTERNAL DEFENSES OF CRUSTACEA 



within 2 hr after injection was followed by a 

 l)lateau that remained constant for the duration 

 of the experiment; with labelled BSA, however, 

 the rate of clearance was concentration depen- 

 dent, but the foreign i)rotein declined to very low 

 levels by 3 days after injection. Fluorescent 

 material was apparently excreted in proportion 

 to its disaiipearance from lobster hemolymjih. 

 Pinocytosis by phag-ocytes was not demonstrated, 

 but tiny n-ranular fluorescent accretions were 

 observed in the hemolymph of lobsters injected 

 with labelled BSA, beginning- about 8 hr after in- 

 jection. In vitro studies, in which a standard 

 ring test with labelled or unlabelled BSA and 

 lobster serum was used, disclosed a clearly dis- 

 cernil)le ring after 16 hr at room temjierature, 

 and a precipitate at the bottom of the tulie after 

 36 hr. Precipitin titers of individual lobster 

 sera ranged from 2 to 16 and were independent 

 of the total protein concentration of the lobster 

 serum. Dialysis of lobster serum did not change 

 the titers, but preci]ntin activity was destroyed 

 by heating above 50° C. 



A number of interesting implications in the 

 study were pointed out by Stewart and Foley. 

 The clearance mechanism seemed able to dis- 

 tinguish between foreign and native in-oteins, 

 and the capacity for clearance seemed high, as 

 indicated by accelerated clearance of larger doses 

 of BSA. Attempts to increase the levels of pre- 

 cipitin by previous injection of lobsters with 

 BSA did not succeed, and in fact resulted in de- 

 creased jn-ecipitin levels in some individuals. 

 The authors suggested that the hemolymph fac- 

 tor responsible for clearance of foreign protein 

 may be maintained normally at low levels and 

 may be supplemented by further secretions when 

 requii'ed. They also suggested that the precipitin 

 principle in the hemolymph may be the first of 

 several steps or possibly the primary removal 

 factor and that digestion and excretion may take 

 place elsewhere — i)robal)ly in the hepatopan- 

 creas. 



The results of the studies of Stewart and Foley 

 are in agreement with those of Teague and Friou 

 (1964), who observed that injected foreign pro- 

 tein was rapidly removed fi-om the hemolymph 

 of the crayfish Camharus virilis. Previous in- 

 jection of the protein did not increase the clear- 



ance rate. Teague and Friou did not observe 

 precipitin activity against injected bovine and 

 human serum albumins but concluded that clear- 

 ance resulted from nonspecific degradation of the 

 foreign protein. 



Other evidence of clearance of injected pro- 

 teins but failure to induce heightened responsive- 

 ness in Crustacea was reported by Campbell and 

 (larvey ( 1961) . They mentioned that "It is also 

 of interest that we have made many attempts 

 to induce antibody formation in invertebrates, 

 e.g., lobsters. We have been unsuccessful so far, 

 but in every instance the antigens remained un- 

 digested and unchanged in the circulation and 

 tissues for many months." Although not men- 

 tioned specifically, the lobsters were probably 

 California spiny lobsters and the test antigens 

 probably included BSA, since this w^as the prin- 

 cipal antigen used in other studies reported in 

 the same paper. 



Phage Clearance 



Taylor, Taylor, and Collard (1964) and Nel- 

 strup, Taylor, and Collard (1968) presented 

 some evidence (from two crabs) of an increase 

 in the rate of secondary clearance of injected 

 Ti bacteriophage in the shore crab, Carcinus 

 maenus. Clearance was not complete until after 

 2 weeks at 16° to 18° C, and no neutralizing 

 antibody to Ti phage was detected in the hemo- 

 l.\Tn])h. Primary inoculation with Tu phage did 

 not increase clearance rates for Ti secondary 

 injections. The small number of animals used 

 in these experiments makes the conclusions 

 highly tentative. The authors suggested the ex- 

 istence of a "phylogenetically more primitive 

 type of immune response than the production of 

 humoral antibody," but did not state clearly what 

 the response was — except possibly that it was 

 "an apparently purely cellular secondary re- 

 sponse." 



Studies of phage clearance by Cushing and 

 McNeely (reported in Cushing, 1967) led to neg- 

 ative conclusions. Phage Tj persisted for up 

 to 168 days in the California spiny lobster and 

 disappeared at a steady rate, uninfluenced by 

 the size of the original inoculum. Two species 

 of crabs tested also failed to clear bacteriophage. 



469 



