Dithiocarbamate toxicity toward thymocytes involves their copper-catalyzed conversion to thiuram disulfides, which oxidize glutathione in a redox cycle without the release of reactive oxygen species.

We have reported previously that diethyldithio-carbamate (DDC) and pyrrolidine dithiocarbamate (PDTC) induce apoptosis in rat thymocytes. Apoptosis was shown to be dependent upon the transport of external Cu ions into the cells and was accompanied by the oxidation of intracellular glutathione, indicating the inducement of pro-oxidative conditions (C. S. I. Nobel, M. Kimland, B. Lind, S. Orrenius, and A. F. G. Slater, J. Biol. Chem. 270, 26202-26208, 1995). In the present investigation we have examined the chemical reactions underlying these effects. Evidence is presented to suggest that dithiocarbamates undergo oxidation by CuII ions, resulting in formation of the corresponding thiuram disulfides, which are then reduced by glutathione, thereby generating the parent dithiocarbamate and oxidized glutathione (glutathione disulfide). Although DDC and PDTC were found to partially stabilize CuI ions, limited redox cycling of the metal ion was evident. Redox cycling did not, however, result in the release of reactive oxygen species, which are believed to be scavenged in situ by the dithiocarbamate. DDC and PDTC were, in fact, shown to prevent copper-dependent hydroxyl radical formation and DNA fragmentation in model reaction systems. The thiuram disulfide disulfiram (DSF) was found to induce glutathione oxidation, DNA fragmentation, and cell killing more potently than its parent dithiocarbamate, DDC. Of particular importance was the finding that, compared with DDC, the actions of DSF were less prone to inhibition by the removal of external copper ions with a chelating agent. This observation is consistent with our proposed mechanism of dithiocarbamate toxicity, which involves their copper-catalyzed conversion to cytotoxic thiuram disulfides.

Susceptibility of four species of copepods, from areas of endemic Dracunculus medinensis, to the North American D. insignis.

Studies of the relative importance of different copepod species in areas of endemic dracunculiasis have been restricted, in part, by limited access to Dracunculus medinensis. However, development of the D. insignis-ferret model provides a source of larvae of a species closely related to D. medinensis. Using larvae derived from this model, the susceptibility of four species of copepods from D. medinensis-endemic areas was compared to that of Canadian Acanthocyclops vernalis, which is fully susceptible to D. insignis. Cryptocyclops linjanticus, from Cameroon, was susceptible but highly sensitive to infection, resulting in high mortality. Mesocyclops aequatorialis similis, also from Cameroon, was fully susceptible, but M. leuckarti leuckarti from Pakistan was partially refractory to D. insignis, as was Thermocyclops emini from Cameroon. Comparisons of infection rates and mean and maximum larval recoveries at three to four and 16-20 days after infection indicated that exposure to 15 first-stage larvae exceeded the threshold of tolerance of A. vernalis, C. linjanticus and M. a. similis to infection with D. insignis. This may account for a decreased survival rate in these copepods which is not evident in the partially refractory T. emini and M. l. leuckarti. The patterns of susceptibility of these copepods to D. insignis are the same as those of copepods exposed to D. medinensis, and indicate that D. insignis can be used to examine factors influencing the susceptibility of copepods to dracunculid infection.

Threshold of detection of Cryptosporidium oocysts in human stool specimens: evidence for low sensitivity of current diagnostic methods.

To determine the minimum number of Cryptosporidium oocysts that can be detected in stool specimens by diagnostic procedures, stool samples seeded with known numbers of Cryptosporidium parvum oocysts were processed by the modified Formalin-ethyl acetate (FEA) stool concentration method. FEA concentrates were subsequently examined by both the modified cold Kinyoun acid-fast (AF) staining and fluorescein-tagged monoclonal antibody (immunofluorescence [IF]) techniques. Oocysts were more easily detected in watery diarrheal stool specimens than they were in formed stool specimens. For watery stool specimens, a 100% detection rate was accomplished at a concentration of 10,000 oocysts per g of stool by both the AF staining and IF techniques. In formed stool specimens, 100% of specimens seeded with 50,000 oocysts per gram of stool were detected by the IF technique, whereas 500,000 oocysts per g of stool were needed for a 100% detection rate by AF staining. Counting of all oocysts on IF slides indicated a mean oocyst loss ranging from 51.2 to 99.6%, depending on the stool consistency as determined by the FEA concentration procedure. Our findings suggest that the most commonly used coprodiagnostic techniques may fail to detect cryptosporidiosis in many immunocompromised and immunocompetent individuals.

Marking snails with numbered, colored discs: a technique for identifying individual specimens.

Numbered, colored discs, measuring 2.5 mm in diameter, were evaluated under laboratory conditions as tags for marking and identifying individual snails. Discs were affixed to the shells of specimens of Helisoma duryi (Pulmonata: Planorbidae) with an adhesive supplied with the discs and with waterproof epoxy. All snails survived the initial tagging procedure and showed no observable behavioral responses to the tagging. The supplied adhesive, intended for use with terrestrial insects, was inadequate for securing the tags for prolonged periods under aquatic conditions. Use of waterproof epoxy, however, resulted in 100% tag retention for over 5 months without significant mortality. The permanence of the marks, their coding of individuals, the rapidity and ease with which the tags can be applied without causing behavioral changes or snail mortality suggest that this technique will be useful in studies requiring identification of individual aquatic snails.