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In mice infected with virulent blood (trypomastigote) forms of Trypanosoma cruzi, complement depletion with cobra venom factor caused a marked exacerbation of the disease evidenced by significantly increased parasitemia levels and early mortality as compared with those of untreated infected animals. The effect was greater in mice receiving cobra venom factor on day 7 postinfection, i.e., at the time when the parasites had had time to localize and multiply in the tissues and appeared in the circulation in appreciable numbers. The possibility that complement participates in host defense against T. cruzi infection through a mechanism involving immune lysis was explored in vitro. T. cruzi trypomastigotes were found to undergo immune lysis in sera of patients with chronic Chagas' disease, in sera of immunized mice, and in solutions containing both immune mouse gamma globulin and a source of active complement. This phenomenon failed to take place either in the absence of complement or after complement inactivation by heat or utilizing complement inactivators. The lytic capacity of heated sera was restored by the addition of active complement to the system. During the immune lysis of T. cruzi blood forms, complement was activated in human sera via both the classical and the alternate pathways. In mouse sera, activation followed at least the alternate pathway.

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Complement (C) activity present in normal human serum has been reported to lyse circulating forms of Trypanosoma cruzi following activation by specific host antibodies bound to the surface of the parasites. In view of this observation, we examined the possibility that a similar phenomenon may cause lysis of T. cruzi by avian complement, a mechanism postulated to be responsible for the natural resistance of birds to T. cruzi infection and previously described as being antibody independent. Trypomastigote forms of T. cruzi grown either in lethally irradiated mice or in cell cultures were lysed readily by the sera of agammaglobulinemic chickens. Lytic activity and titers of normal and agammaglobulinemic sera were comparable. The lytic reaction was inhibited by heat inactivation of the sera, or by addition of EDTA or cobra venom factor to the assay mixtures. Lysis of T. cruzi was observed when calcium, but no magnesium ions, were chelated with EGTA. Furthermore, a significant loss of lytic activity was observed when sera from C-depleted chickens were tested. Normal and agammaglobulinemic chickens cleared intravenously injected parasites (from either lethally irradiated mice or cell cultures) from their circulation in 7 min or less whereas C-depleted animals required 1,740 min or longer. Routine examination of the parasites from these two sources by immunofluorescence confirmed the absence of immunoglobulins on their surface. These results emphasize the lack of requirement of antibodies for, and the important role of complement in both the natural resistance that birds exhibit against T. cruzi infection and the lytic activity displayed by avain serum on virulent forms of T. cruzi.

An enzyme-linked immunosorbent assay (ELISA) for the detection of herpes simplex virus is described that can be performed in approximately four hours. The test, which does not require specialized equipment and uses relatively inexpensive, commercially available reagents, detected herpes simplex virus in 51% of specimens found to be positive by a time-consuming cell culture technic. The ELISA test compared favorably with a direct immunofluorescence method that detected HSV in only 1% of the cell culture-positive specimens. The ELISA test was readily carried out even with specimens unsuitable for cell culture and did not require cellular material as is the case with immunofluorescence technics. An advantage of the ELISA test for herpes simplex virus over the cell culture method was the detection of nonviable virus.

Treatment of highly purified preparations of the third component of complement (C3) with 0.5M hydroxylamine at 20 degrees C for 15 to 30 minutes, followed by acidification, resulted in dissociation of a peptide from the C3 molecule. The isolated fragment (molecular weight, 7600) resembled enzymatically liberated anaphylatoxin (C3a) with respect to size, charge, amino acid composition, and biological activity. Its capacity to contract smooth muscle was inhibitable by antihistamines; it also produced tachyphylaxis and desensitization of the guinea pig ileum to C3a. Thus native C3 probably contains an esterlike bond and hydroxylamine-liberated anaphylatoxin may represent one of the polypeptide chains of the C3 molecule.

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Protection against infection with virulent blood (trypomastigote) forms of Trypanosoma cruzi was accomplished in mice by immunization with culture (mainly epimastigote) forms killed by treatment with sodium perchlorate. Sodium chloride, used instead of sodium perchlorate, with all other conditions kept the same, failed to kill all the organisms, indicating that the effects of the perchlorate anion were not simply ionic or osmotic, suggesting that they might be chaotropic. A single dose of the immunogen, without adjuvants, was sufficient to significantly protect against the infection. Protection was achieved by either intraperitoneal, intramuscular, or subcutaneous immunization, though the first two routes appeared to be more effective. After challenge, parasitemias were negative in 25, 29, and 17% of the animals immunized intraperitoneally, intramuscularly, and subcutaneously, respectively.