Individual Mobilization Augmentation: recipe for an effective total force partnership.

The ability to carry out rapid mobilization and full integration of well-trained reserve/guard personnel with active component personnel proved to be a decisive factor during the Gulf War. Such an approach can also be effective in fulfilling peacetime military missions, especially in an environment of increasing requirements, nontraditional missions, and pervasive force reductions. This paper describes the role played by Army Reserve Individual Mobilization Augmentation (IMA) assets in support of two important Army Medical Department missions involving new initiatives in scientific peer review and extramural grant administration: military nursing research and breast cancer. Successful planning, development, and implementation of these programs is attributed in part to the fostering of an effective partnership between reserve and active components on both individual and organizational levels. Critical factors influencing this partnership are analyzed and the relevance of such peacetime IMA taskings to wartime missions is discussed.

Production of anti-idiotypic antibodies as potential immunoreagents for the serological diagnosis of bovine cysticercosis.

Although serological assays have been developed for cysticercosis, difficulties remain in isolating antigens from cross-reacting components and in obtaining reliable sources of parasite material for purification. The object of this study was to demonstrate the feasibility of using anti-idiotypic antibodies as alternative immunoreagents for the serological diagnosis of bovine cysticercosis. A fraction of Taenia hydatigena cyst fluid (ThFAS) had previously been used as a diagnostic antigen for Taenia saginata cysticercosis. Rabbits and mice were inoculated with ThFAS to produce either polyclonal or monoclonal antibodies (idiotypes), respectively. Idiotypes were purified by preparative protein A column, then inoculated into other rabbits to produce anti-idiotypes. The presence of anti-idiotypes in hyperimmune serum was demonstrated by competitive inhibition ELISA. Then anti-idiotypes were purified and used as coating antigen in a plate ELISA; results were compared to those using ThFAS as coating antigen. Anti-idiotypes yielded results comparable to those obtained with the native antigen and thus can be used as synthetic antigens for diagnostic purposes.

Morphological adaptations of intestinal helminths.

Nematodes, trematodes, cestodes, and acanthocephalans each have become adapted in different ways to the microenvironment of the vertebrate intestine. Life in this specialized habitat affords parasites a reliable source of nutrients, a relatively homeostatic environment, and protection from predators but, in exchange for these advantages, presents the special challenges of exposure to digestive enzymes, normal peristalsis, and host immune response to infection. Logically, the surface of the parasite should be the first part of the organism to encounter such challenges, and, for this reason, any response or reaction by the parasite is expected to be manifested at the parasite-host interface. Morphological adaptations of intestinal helminths to their microenvironment include modification of the tegumental surface that affords protection and increases absorptive surface area, development of specialized attachment organs, and, in some cases, complete loss of their own internal digestive system. Representative examples of such adaptations by helminths are described and discussed in terms of the parasite's nutritional requirements, site selection, and host specificity, and the possibility is suggested that some helminths may have adapted in ways that exploit host defensive mechanisms for their own benefit.

Isolation and purification of a diagnostic antigen for bovine cysticercosis by hydrophobic chromatography.

An ammonium sulfate fraction of Taenia hydatigena cyst fluid (ThFAS) was further fractionated by hydrophobic interaction chromatography, using alkylagarose and omega-amino alkylagarose columns, in an effort to isolate and purify a specific diagnostic antigen in the ThFAS preparation. The less than 12 kDa antigen was found to have an affinity for immobilized alkanes with chain length of six carbons or greater. The antigen was recovered in an ethylene glycol eluate from a hexylagarose column then analyzed by Western blot; it reacted with bovine and human cysticercosis infection sera and with specific monoclonal antibodies but not with control sera or fascioliasis infection sera. When the eluate was used as coating antigen in a plate ELISA assay no false positive reactions were seen in sera from cattle infected with Fasciola hepatica; false positive reactions were observed for the unfractionated ThFAS antigen preparation.

Development of a serologic assay for cysticercosis, using an antigen isolated from Taenia spp cyst fluid.

An ammonium sulfate-soluble fraction of Taenia hydatigena cyst fluid (ThFAS) was further evaluated for use in the immunodiagnosis of cysticercosis. Analysis of ThFAS by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and protein immunoblot analysis confirmed earlier reports of a highly specific, low molecular weight antigen in this preparation; in contrast, other components of ThFAS were shown to react nonspecifically. Antibodies against the less than 12-kD diagnostic antigen were detected in sera from 10 cattle and 4 swine inoculated with metacestodes of T saginata and T solium, respectively, but not in animals inoculated with Fasciola hepatica, Trichinella spiralis, Brucella abortus, or Toxoplasma gondii, or in noninoculated controls. Isolation and immobilization of the less than 12-kD antigen on a hydrophobic transfer membrane resulted in development of an unambiguous dipstick assay capable of correctly identifying fully developed (10-week) experimentally induced infections in cattle and swine. In addition, the dipstick assay was highly specific for diagnosis of the disease in human beings, and offers the potential of distinguishing between human clinical cases of cysticercosis and taeniasis. A similar reactive antigen of diagnostic potential was also identified and isolated from T crassiceps and T taeniaeformis cyst fluids.

Evaluation of a 'dipstick' immunoassay to detect cysticercosis in experimentally infected cattle.

A 'dipstick' immunoassay for bovine cysticercosis, using an antigen isolated from Taenia hydatigena cyst fluid, was evaluated in cattle experimentally infected with Taenia saginata. The assay correctly identified six out of seven infected cattle, including an animal in which only 12 living cysticerci were found. Cattle became seropositive as early as 3 weeks post-infection. A false-negative reaction was found for one very lightly infected animal, from which only four living cysticerci were recovered at necropsy. The assay was also used to detect circulating antibodies in experimentally infected cattle before and after therapeutic treatment with anthelminthics. The results suggest that praziquantel-treated animals gradually revert to being seronegative after the cysticerci are killed.

Evidence for selective incorporation of host immunoglobulin by strobilocerci of Taenia taeniaeformis.

Strobilocerci of Taenia taeniaeformis were obtained from laboratory rats 90 days after experimental infection. Cyst fluid, whole parasite homogenate, and rat serum each were fractionated by SDS-PAGE, immobilized on nitrocellulose by western blot, and probed with conjugated goat anti-rat IgG. Reactive bands with relative mobilities corresponding to rat IgG were found in all 3 samples. Additional bands in cyst fluid and parasite homogenate may represent enzymatic degradation of IgG. The pattern of reactive bands in the homogenate discounts the nonspecific adsorption of host molecules onto the tegument and suggests selective incorporation of serum proteins. The presence of an IgG-like molecule of atypical molecular weight is consistent with either molecular mimicry or enzymatic cleavage of IgG bound to the tegument. The relevance of serum protein utilization by the parasite to evasion of the host immune response is discussed.

Early diagnosis of Schistosoma mansoni in mice using assays directed against cercarial antigens isolated by hydrophobic chromatography.

Two diagnostic assays are described for the early diagnosis of acute schistosomiasis, using a defined cercarial antigen preparation obtained by hydrophobic chromatography. Circulating IgM antibodies against this antigen fraction could be detected by ELISA as early as 1 wk after exposure in experimentally infected mice; IgM levels against other antigens and IgG levels against all the preparations examined were not significantly elevated until approximately 4-5 wk postinfection. Circulating antigen was detected as early as 3 days after exposure by a competitive inhibition ELISA using rabbit serum prepared against the cercarial antigen; antigen levels in the serum of mice with a 100-worm infection were found to exceed 100 ng/ml. Studies using sera from infected humans indicate that the assay can also recognize chronic S. mansoni, S. haematobium or S. japonicum infections. In a very limited field study, the specificity of the circulating antigen assay with regard to other helminthic infections was found to be 85%; sensitivity 100%. Preliminary characterization of the relevant antigen indicates that it is a relatively hydrophobic polypeptide with a molecular weight of approximately 41,000 daltons. The implications of these findings with regard to the treatment of travelers or the conduct of seroepidemiological studies in endemic areas are discussed.

Expression of lectin-binding surface glycoproteins during the development of Schistosoma mansoni schistosomula.

Tegumental glycoproteins of Schistosoma mansoni cercariae, mechanically produced 24-hr and 48-hr schistosomula, and adult worms were radioiodinated with the Bolton-Hunter reagent, then isolated by lectin affinity chromatography. SDS-PAGE revealed Con A binding glycoproteins with apparent molecular weights of 180,000, 150,000, 43,000, and 30,000 in detergent extracts of the tegument of cercariae. These glycoproteins are retained by 24-hr mechanically produced, cultured schistosomula and are accompanied by the appearance of 2 additional labeled glycoproteins, mol. wt. 66,000 and 57,000. In 48-hr schistosomula, there is a marked increase in the relative size of the 66,000 mol. wt. peak. In contrast, the 57,000 mol. wt. glycoprotein is the major radiolabeled Con A binding component of the adult tegument; the other peaks are either reduced or absent in adults. Similar findings were obtained following affinity chromatography using immobilized Lens culinaris lectin or Ricinus communis agglutinin, and following metabolic labeling of glycoproteins with tritiated galactose.

Development of circulating antigen assay for rapid detection of acute schistosomiasis.

A circulating antigen assay to detect acute schistosomiasis mansoni in experimentally infected mice has been developed. The competitive-inhibition enzyme-linked immunosorbent assay uses rabbit serum prepared against purified cercarial antigen. The assay detects a 100-worm infection as early as 1 week after exposure; circulating antigen levels in mouse serum exceeded 0.1 microgram/ml. Preliminary characterisation of the cercarial antigen shows that it is a hydrophobic polypeptide of approximate molecular weight 41,000. A rapid and accurate diagnosis of acute schistosomiasis in travellers would facilitate appropriate chemotherapy before the onset of pathology. A circulating antigen assay would also be valuable for seroepidemiological studies in areas where the disease is endemic.

Characterization of surface glycoproteins on Schistosoma mansoni adult worms by lectin affinity chromatography.

Adult Schistosoma mansoni were radiolabeled by direct radioiodination using the Bolton-Hunter reagent or by metabolic labeling using radioactive hexose precursors. Tegumental material was extracted by freeze-thaw or by incubation in the non-ionic detergent Nonidet P-40, then applied to chromatography columns containing the following immobilized lectins: Con A, lentil lectin, wheat germ agglutinin, soybean agglutinin and the agglutinins from Ricinus communis and Helix pomatia. SDS-PAGE analysis of the sugar eluates from these columns revealed the presence of 15 glycoproteins with apparent molecular weights greater than or equal to 300,000, 215,000, 168,000, 152,000, 134,000, 122,000, 108,000, 83,000, 58,000, 53,000, 46,000, 41,000, 34,000, 30,000 and 23,500. Many of the glycoproteins reacted with more than one lectin. Information about carbohydrate content and lectin binding provides a preliminary characterization of the tegumental glycoprotein antigens of adult worms.

Strategies to determine the molecular basis of chemical communication by trematodes.

The identification of pheromones and chemicals that may inhibit or stimulate growth and reproduction necessarily leads to the consideration of biochemical methods to isolate and characterize these molecules. Analysis ofSchistosoma mansoni surface antigens by direct radioiodination, metabolic labeling with tritiated and(14)C-tagged hexose precursors, affinity chromatography, isoelectric focusing, hydrophobic chromatography, and competitive inhibition is presented to illustrate methods of immunochemical analysis and antigen purification. Technical problems that may arise when investigating parasite molecular biology are described. Evidence of diminished fecundity of female worms in acutely infected mice supports the theory of a "crowding effect" in murine schistosomiasis and suggests the possibility that worm secretions or metabolites may function as chemical messages to inhibit oviposition. There is also evidence that the immune response of mice to an isolated surface antigen from adult worms results in the attenuation of hepatic granulomata of challenge infections. Several hypotheses are proposed to elucidate the molecular basis for chemical communication between trematodes and analytical approaches to test these hypotheses are outlined.

Leishmanial differentiation in vitro: induction of heat shock proteins.

During temperature-induced in vitro differentiation from the promastigote to the amastigote form of the parasitic protozoan Leishmania mexicana, seven actively synthesized proteins were identified. These proteins corresponded precisely in molecular weight to the well-known heat shock proteins seen in species such as Drosophila. The minimal DNA synthesis observed in the differentiating parasites indicated that little cell division occurs during this process. The absence of RNA synthesis during early temperature-induced differentiation suggests that the leishmanial heat shock proteins are post-transcriptionally regulated. Heat shock proteins may play an adaptive role in the transition of Leishmania from arthropod to mammalian host.

Some problems associated with radiolabeling surface antigens on helminth parasites: a brief review.

Recent developments in technology have facilitated substantial advances in the characterization of surface antigens from a wide variety of both normal and neoplastic cells. However, the immunochemistry of parasites has lagged behind. Efforts to apply conventional radiolabeling methods to helminths have not always been successful. Experimental work with Schistosoma mansoni is reviewed to illustrate common problems encountered in surface labeling studies. These findings should provide insight for the future investigation of other helminth species.