The spacelab experience: a synopsis.

The Spacelab 1 mission, a joint venture of the European Space Agency and the National Aeronautics and Space Administration took place during the period 28 November through 8 December 1983. An overview of the first flight of the orbiting laboratory is presented here. The payload crew members' view of Spacelab operations and results of the scientific investigations carried out on this mission are presented in the following reports.

Space plasma physics: space experiments with particle accelerators.

Electron and plasma beams and neutral gas plumes were injected into the space environment by instruments on Spacelab 1, and various diagnostic measurements including television camera observations were performed. The results yield information on vehicle charging and neutralization, beam-plasma interactions, and ionization enhancement by neutral beam injection.

Increased Extra-axial Cerebrospinal Fluid in High-Risk Infants Who Later Develop Autism.

BACKGROUND: We previously reported that infants who developed autism spectrum disorder (ASD) had increased cerebrospinal fluid (CSF) in the subarachnoid space (i.e., extra-axial CSF) from 6 to 24 months of age. We attempted to confirm and extend this finding in a larger independent sample. METHODS: A longitudinal magnetic resonance imaging study of infants at risk for ASD was carried out on 343 infants, who underwent neuroimaging at 6, 12, and 24 months. Of these infants, 221 were at high risk for ASD because of an older sibling with ASD, and 122 were at low risk with no family history of ASD. A total of 47 infants were diagnosed with ASD at 24 months and were compared with 174 high-risk and 122 low-risk infants without ASD. RESULTS: Infants who developed ASD had significantly greater extra-axial CSF volume at 6 months compared with both comparison groups without ASD (18% greater than high-risk infants without ASD; Cohen's d = 0.54). Extra-axial CSF volume remained elevated through 24 months (d = 0.46). Infants with more severe autism symptoms had an even greater volume of extra-axial CSF from 6 to 24 months (24% greater at 6 months, d = 0.70; 15% greater at 24 months, d = 0.70). Extra-axial CSF volume at 6 months predicted which high-risk infants would be diagnosed with ASD at 24 months with an overall accuracy of 69% and corresponding 66% sensitivity and 68% specificity, which was fully cross-validated in a separate sample. CONCLUSIONS: This study confirms and extends previous findings that increased extra-axial CSF is detectable at 6 months in high-risk infants who develop ASD. Future studies will address whether this anomaly is a contributing factor to the etiology of ASD or an early risk marker for ASD.

Glutathione activation of a cysteine proteinase from Schistosoma mansoni.

A cysteine proteinase isolated from Schistosoma mansoni adults requires reduction by thiols for activation. The proteinase is located in the parasite digestive tract where it degrades hemoglobin released from host red blood cells. Reduced glutathione (GSH) has been shown to be effective in activation. Total glutathione concentration and the GSH/oxidized glutathione (GSSG) ratio were measured in whole blood lysate (3.2 mM, 284), serum (24 microM, 9.8) and material collected from the parasite digestive tract (4.2 mM, 137). The ratio of GSH/GSSG at which the enzyme displays half-maximal activity (Kox) is 1.0. Proteinase activation as a function of glutathione concentration and time was determined. The first-order reaction yielded a half-time of activation of 13 min at 5 mM. The second-order rate constant was 12.7 M-1 X min-1. The function of the proteinase and its possible regulation by glutathione activation are discussed.

Generation of hydroxyeicosatetraenoic acids by human inflammatory cells: analysis by thermospray liquid chromatography-mass spectrometry.

Arachidonic acid was converted to a series of hydroxyeicosatetraenoic acids (HETEs) by mixed human inflammatory cells following stimulation with the calcium ionophore A23187. HETEs were purified by a simple one-step extraction procedure followed by HPLC. The HPLC was coupled to a Finnigan quadrupole mass spectrometer using the now commercially available thermospray liquid chromatography-mass spectrometry interface. The HPLC eluant was monitored 'on line' by the mass spectrometer. Soft ionisation occurs, generating intense molecular ion species in the negative ion mode (M - H-:m/z 319) for each of the isomeric HETEs. The (M + H+ - H2O) ion at m/z 303 is the major species in the positive ion spectra of HETEs. Mass spectra were obtained on-line post-HPLC for HETEs formed by the human cells, and the HPLC-MS profile compared with that obtained from standards; species corresponding to the 11-, 9- and 5-HETEs were observed.

Cryptosporidiosis in Houston, Texas. A report of 95 cases.

Cryptosporidiosis is an important cause of diarrhea. We identified 95 patients with cryptosporidiosis over a 6-year period in our county hospital system, including 9 children and 86 adults infected with the human immunodeficiency virus (HIV). Risk factors included male-to-male sexual practices and Hispanic race. Diarrhea, weight loss, and gastrointestinal complaints were the most common symptoms at presentation. Among the HIV-infected adults, 20 (23%) developed biliary tract disease. Biliary involvement was associated with low CD4 counts. Treatment with paromomycin and antimotility agents was effective in reducing diarrheal symptoms in 54 of 70 (77%) patients with the acquired immunodeficiency syndrome (AIDS), although there was a high rate of relapse. Paromomycin did not prevent the development of biliary disease. Biliary disease responded to cholecystectomy or sphincterotomy with stent placement. Though often a cause of morbidity, cryptosporidiosis was only rarely the cause of death, even among patients with HIV. Cryptosporidiosis continues to be an important medical problem even in developed-countries. Current methods of prevention and treatment are suboptimal.

Characterization of a cysteine proteinase from Taenia crassiceps cysts.

Human neurocysticercosis, due to infestation of the central nervous system with Taenia cysts, is a common cause of neurologic disease in endemic areas and is being increasingly recognized in the United States. Previous studies have suggested that Taenia cysts bind host IgG via Fc-like receptors and that bound IgG is degraded by the parasite, perhaps as a source of nutrients or a means of immune evasion. We now demonstrate that IgG degradation is thiol dependent and is inhibited by the cysteine proteinase inhibitor, E-64. The cysteine proteinase activity from Taenia crassiceps cysts was purified 682-fold by acid extraction, gel filtration chromatography, and anion-exchange FPLC. The cysteine proteinase appeared as a 43 kDa band on silver-stained gels. Its isoelectric point was 5.27. The purified enzyme was inhibited by cysteine proteinase inhibitors and also by chloromethyl ketone inhibitors, but not significantly by other inhibitors of serine, aspartic, or metallo-proteinases. Substrate studies showed pronounced cleavage of Z-Phe-Arg-7-amino-4-trifluoromethylcoumarin (Z-Phe-Arg-AFC), but not of substrates with neutral or positively charged amino acids in the P2 position. Km for Z-Phe-Arg-AFC was 1.0 x 10(-7) M, Kcat 58 s-1, and Kcat/Km 5.8 x 10(8) mol-1s-1. Amino acid sequencing of the amino terminus revealed a single cysteine residue with surrounding residues suggestive suggestive of a cysteine proteinase active site. The sequence, however, did not contain the conserved active site associated with enzymes of known cysteine proteinase families. This cysteine proteinase may play an important role in the interaction of Taenia cysts and host immunoglobulin and is a potential target for antiparasitic chemotherapy.

Properties of the acid thiol proteinase from Schistosoma mansoni adults.

Seven beta-naphthylamine-linked peptides were tested as substrates for a previously described thiol proteinase of adult Schistosoma mansoni. The enzyme was not active on carbobenoxy-arginyl-arginyl-4-methoxy-2-naphthylamide and carbobenzoxy-alanyl-arginyl-arginyl-4-methoxy-2-naphthylamide. Enzyme activity was maximal at acidic pH (4.9-5.5) with similar optima for both macromolecular and peptide substrates. Activity of partially purified enzyme preparations against carbobenzoxy-arginyl-arginyl-4-methoxy-2-naphthylamide was stimulated more than 10-fold by thiols. The properties of this proteinase differ from those of proteolytic enzymes from the cercariae ad eggs of S. mansoni.

Antibody response to a purified parasite proteinase (SMw32) in Schistosoma mansoni infected mice.

Indirect immunofluorescence of Schistosoma mansoni adult worm sections has revealed that the early immunoglobulin response is directed toward the parasite digestive tract. One of the components of the worm gut is a cysteine proteinase which degrades host hemoglobin ingested by the parasite. In this report the purified proteinase (SMw32) was used in ELISA and immunoblot analyses to study the specific antibody response during the course of an acute infection. We have found high titer IgG antibody in S. mansoni infected, but not uninfected, mice. The anti-proteinase response involves IgM, IgG1, IgG2a, and IgE isotypes. Total IgM and IgG levels increased by week 3 post-infection and remained elevated throughout the study (7 weeks). Increased titers (IgM, IgG) of specific anti-proteinase were also apparent by week 3 post-infection, long before fecal eggs were detectable. Mean anti-proteinase IgG stabilized at high titer by week 5 post-infection, while IgM titers decreased to near background levels. Anti-proteinase IgE was first detectable at week 4 and reached peak titers by weeks 6 and 7. The strong antibody response to the purified SMw32 proteinase is consistent with the early reactivity of S. mansoni infected mice and humans to a 31 kDa component of the worm gut described by others.

Antibody response to Schistosoma mansoni adult worm cysteine proteinases in infected individuals.

Antigens of the Schistosoma mansoni digestive tract are recognized early in the infective process. Two immunogenic components of the excretory/secretory products are proteolytic enzymes that degrade host hemoglobin in the lumen of the parasite gut. These enzymes, CP1 and CP2, belong to the class of cysteine proteinases. In this study, a preparation containing both proteinases has been used to detect proteinase antibodies in the sera of individuals living in Burundi. Of 133 individuals tested, 92% were excreting schistosome eggs. All patients with documented infections had positive anti-proteinase IgG titers (mean = 1:614), while 82% had positive IgM titers (mean = 1:267). Six weeks following praziquantel treatment, patients were assessed for egg excretion and antibody titer. Anti-proteinase IgG titers were significantly lower (mean = 1:259) than pre-treatment titers. Patients who were infected with S. japonicum or S. haematobium typically showed a cross-reactive IgG response. Patients from non-endemic regions yielded negative titers, and those with non-trematode parasites were negative (79%) or weakly positive. S. mansoni cysteine proteinases may be used for the detection of schistosome infections.

The hypersensitivity response to the adult worm proteinase, SMw32, in Schistosoma mansoni infected mice.

A cysteine proteinase (SMw32) from the digestive tract of an adult Schistosoma mansoni worm has previously been purified and characterized. During the course of infection with this parasite, a strong immune response to the enzyme occurs. We now have confirmed the presence of anti-proteinase IgE and IgG in S. mansoni infected mice and have investigated the in vivo cellular response to proteinase in infected and uninfected mice. Immediate and delayed type hypersensitivities were detected in uninfected mice sensitized by multiple injections of proteinase. In S. mansoni infected mice, immediate hypersensitivity reactions were seen at 6 and 8 weeks following infection, coincident with the increase in anti-proteinase IgE antibody. Histological sections of the injection site confirmed the presence of degranulating mast cells. In contrast, delayed type hypersensitivity could not be detected at any time during the course of the infection. In the murine model of acute infection, immediate hypersensitivity to the SMw32 proteinase was predictive of infection.

Risk factors for development of first symptomatic Giardia infection among infants of a birth cohort in rural Egypt.

Giardia infection is associated with diarrheal diseases among infants and young children in both industrialized and developing countries. A study was conducted to demonstrate the predisposing factors for occurrence of the first symptomatic Giardia infection among infants in rural Egypt. The study cohort was followed from birth through the first year of life. Univariate and multivariate analyses of data revealed that infants less than six months of age were at special risk for developing their first symptomatic infection compared with infants more than six months of age. Analysis of the data, furthermore, revealed an increased risk of infant Giardia infection associated with living in a household without a latrine (relative risk [RR] = 2.63, confidence interval [CI] = 1.4-4.9, P < 0.05), a mud floor in the sleeping rooms (RR = 1.79, CI = 1.O30-3.0, P < 0.05), and household exposure to more than 10 chickens (RR = 2.5, CI = 1.13-5.56, P < 0.05). In contrast, the mother's education beyond the primary level (RR = 0.28, CI = 0.09-0.85, P < 0.05), drinking water stored in metallic containers (RR = 0.33, CI = 0.11-0.98, P < 0.05), and male sex (RR = 0.52, CI = 0.3-0.89, P < 0.05) were associated with decreased risk of Giardia infection. These data suggest that in addition to age of infants, poverty, low education, gender discrimination, and certain environmental conditions potentiated the risk for developing the first symptomatic infection.

Coenuriasis in a spectacled langur (Presbytis obscura): praziquantel treatment and the antibody response to cyst antigens.

A diagnosis of coenuriasis was made in a spectacled langur raised in captivity. Multiple cysts removed from the subcutaneous tissues and later from the abdominal cavity were identified as coenuri, typical of the genus Taenia. Post-surgical treatment of the remaining cysts with praziquantel was assessed with whole body computerized tomography (CT). CT at 6 weeks post-treatment revealed a reduction in size and increased calcification of abdominal cysts as compared to pretreatment CT. Cyst fluid antigens in ELISA assays showed a high titer (1:5, 120) IgG response in the langur serum, while no IgM response could be detected. No decrease in IgG titer was seen 6 weeks after treatment. Immunoblot analyses identified several parasite-specific antigens with apparent molecular weights of greater than 92.5 (3 bands), 88, 41, 37, and 34 kDa.

Schistosoma mansoni: anti-SMw32 proteinase response in vaccinated and challenged baboons.

Antibodies to a cysteinyl proteinase of the trematode Schistosoma mansoni have been detected in serum from infected mice and humans. We have evaluated antiproteinase responses in infected baboons and in baboons vaccinated with irradiated, cryopreserved schistosomules prior to challenge. Prechallenge sera and normal, uninfected control sera were nonreactive by ELISA and immunoblots. Serum antibodies were first detectable by ELISA at two months post-challenge in both challenged (C) and vaccinated-challenged (V-C) baboons (serum dilution 1:200). By four months post-challenge, ELISA absorbance values for subgroup C baboons were significantly higher than for V-C counterparts. The immunoblot technique provided a more sensitive means of detecting antibody early in the infection. One month post-challenge, 7 of 12 C and V-C sera (diluted 1:100) contained measurable anti-proteinase antibody. By month two, 12 of 12 were immunoblot-positive. Baboons vaccinated but not challenged (subgroup V) remained negative. The presence of the anti-proteinase antibody appears to be a sensitive and early marker for infection by S. mansoni.

Infectivity of Cryptosporidium parvum in healthy adults with pre-existing anti-C. parvum serum immunoglobulin G.

A 50% infectious dose (ID50) of 132 Cryptosporidium parvum oocysts was previously determined in serologically negative individuals (ELISA). In this study, 17 healthy adults with pre-existing anti-C. parvum serum IgG were challenged with 500-50,000 oocysts. Infection and diarrhea were associated with the higher challenge doses. The ID50 was 1,880 oocysts, > 20-fold higher than in seronegative volunteers. Fecal oocysts were detected in only seven (53.8%) of 13 individuals with clinical cryptosporidiosis, indicating that the host response may effectively decrease the number of oocysts produced. Subjects with the highest absorbances prior to challenge had little to no increase in IgG following challenge, whereas volunteers with lower reactivities showed significant postchallenge increases. This suggests that an upper limit of serum IgG was present in some subjects, while others were further stimulated by an additional exposure. These data indicate that prior exposure to C. parvum provides protection from infection and illness at low oocyst doses.

Impact of breast-feeding on Giardia lamblia infections in Bilbeis, Egypt.

A total of 152 infants were followed from birth to 1 year of age in a rural community of Egypt to document Giardia lamblia infection and to determine the effect of breast-feeding on enteric infections by this protozoan. Asymptomatic Giardia infections persisted as long as 4 months, with a mean duration of excretion of 7.18 weeks. The incidence of asymptomatic infection was 4.5 episodes per child-year. Exclusively breast-fed infants had lower risk for asymptomatic (odds ratio [OR] = 0.66, 95% confidence interval [CI] = 0.45-0.96, P < 0.05) and symptomatic infections (relative risk [RR] = 0.50, 95% CI = 0.27-0.90, P < 0.05). Furthermore, breast-fed infants had fewer clinical manifestations, including mucus in stool (23.8% versus 76.2%, P = 0.08), loss of appetite (17.6% versus 82.3%, P < 0.05), and abdominal tenderness (17% versus 82.9%, P < 0.05) compared with infants who were not exclusively breast-fed. Breast-feeding should be considered as an effective means to prevent Giardia infections and should be encouraged in regions where G. lambia is highly endemic.

Risk assessment for Cryptosporidium: a hierarchical Bayesian analysis of human dose response data.

Three dose-response studies were conducted with healthy volunteers using different Cryptosporidium parvum isolates (IOWA, TAMU, and UCP). The study data were previously analyzed for median infectious dose (ID50) using a simple cumulative percent endpoint method (Reed and Muench, 1938). ID50s were derived using two definitions of infection: one as subjects having oocysts detected in stool by direct fluorescence assay, and the other by a clinical finding of diarrhea with or without detected oocysts (Chappell et al., 1998; Okhuysen et al., 1999). In the present study, the data were analyzed using the broader definition of infection (i.e., presence of oocysts in stool and/or diarrheal illness characteristic of cryptosporidiosis). Maximum likelihood dose-response parameter estimates for UCP, IOWA, and TAMU were 2980, 190, and 17.5, respectively. Based on these estimates, the ID50s of the three respective isolates were 2066, 132, and 12.1. The three oocyst isolates were considered representative of a larger population of human-infecting strains and analyzed as combined data using a hierarchical Bayesian model. Hyperparameters defined the distribution of dose-response parameters for the population of strains. Output from Markov Chain Monte Carlo analysis described posterior distributions for the hyperparameters and for the parameters of the IOWA, TAMU, and UCP strains. Point estimates of dose-response parameters produced by this analysis were similar to the maximum likelihood estimates. Finally, the utility of these results for probabilistic risk assessment was evaluated. The risk of infection from single oocyst doses was derived for a mixture of the three isolates (where IOWA, TAMU, or UCP are equally likely), and for an oocyst selected at random from the larger population of strains. These estimated risks of infection were 0.018 and 0.028, respectively.

Purification and partial characterization of cysteine proteinase from Spirometra mansoni plerocercoids.

Spirometra mansoni plerocercoids were dissected from the tissues of naturally infected snakes (Natrix trigrialateralia). Fresh plerocercoids were incubated in medium, and excretory-secretory products (E-S) were collected. In addition, soluble proteins from lyophilized plerocercoids (10 mg/ml) were extracted in 0.1 M sodium acetate. Proteinase activity was assayed with a synthetic fluorescent substrate, carbobenzoxy-phenylalanyl-arginyl-7-amino-4-trifluoromethylcoumarin. Proteinase was isolated from plerocercoid extract or E-S by diethylaminoethyl trisacryl M ion-exchange chromatography and thiolpropyl-Sepharose affinity chromatography. These separations resulted in a 12.2- (extract) and 15.6-fold (E-S) purification of proteinase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified materials revealed a 28-kDa band, consistent with the apparent native molecular weight (gel filtration chromatography) of approximately 35 kDa. Proteinases purified from whole extracts and E-S were compared for various biochemical characteristics; inhibitor profiles indicated that activities from both sources are cysteine proteinases, they exhibited identical pH curves with optima at pH 5.5 and a 50% activity range at pH 4.7-8.0, they cleaved collagen chains to 3 identical products, and they showed only minor activity toward hemoglobin. Further, the proteinase purified from plerocercoids was utilized in immunoblots with sera from sparganosis patients. Antibody (IgG) from the infected patients, but not uninfected controls, recognized the cysteine proteinase, suggesting that this antigen may be useful in the serodiagnosis of Spirometra mansoni infection.

Proteinase activity in miracidia, transformation excretory-secretory products, and primary sporocysts of Schistosoma mansoni.

Proteinase activity was detected in the culture medium of transforming miracidia and in detergent extracts of Schistosoma mansoni miracidia and primary sporocysts using a fluorescent substrate, carbobenzoxy-phenylalanyl-arginyl-7-amino-4- trifluoromethylcoumarin. Medium collected after the first 24 hr of miracidial cultivation (transformation medium; TM) contained most (80%) of the activity released during 5 days of in vitro culture. Based on proteinase activity contained in Triton X-100 extracts of whole larvae, miriacidia and primary sporocysts exhibited a similar amount of total activity per organism, whereas specific activity was about 2-fold greater in miracidia. Approximately 10% of total miracidial activity was released during the first 24 hr of transformation. This early release of proteinase is consistent with possible involvement of these enzymes in miracidial snail penetration. Proteinase activities from larval extracts and culture media were identical when characterized for thiol-dependence, inhibitor profile, and pH optimum and indicate that the proteinase(s) belongs to the cysteine class of acidic endopeptidases. Further studies with TM revealed a substrate preference for a hydrophobic amino acid in the P2 position. High performance liquid chromatography gel filtration showed 2 peaks of activity at 19,000 and 36,000 Da, whereas specific inhibitor labeling yielded heterogeneous banding in the molecular weight range of 33,000-44,000 Da. Lastly, sporocyst extracts incubated with snail plasma (cell-free hemolymph) revealed degradation of high molecular weight hemolymph proteins, including hemoglobin. The finding of significant cysteine proteinase activity in miracidia and primary sporocysts and the continued low level of secretion by sporocysts suggest a functional role of these proteinases in the establishment and/or maintenance of infections within the snail host.

Giardia antigen detection in patients with chronic gastrointestinal disturbances.

BACKGROUND: Giardia lamblia, a protozoan parasite, is transmitted by cysts in contaminated water or food or by person-to-person contact. The standard in diagnosis has been the microscopic demonstration of fecal cysts, which yields many false negatives due to high variability in cyst excretion. A new method that detects infection even when few parasites are present is now available. This immunodiagnostic test is rapid, sensitive, and specific, and typically requires only a single stool specimen. In this study patients with gastrointestinal (GI) complaints were screened for Giardia antigens, and the test results were compared to conventional microscopy. Costs incurred by patients with chronic GI problems were documented. METHODS: Twelve patients with GI complaints were tested for Giardia by microscopy and 13 patients by the immunodiagnostic test. Patient charts were evaluated for pertinent history and the diagnostic tests ordered before giardiasis was considered. RESULTS: For all patients, microscopy was uniformly negative, but 6 of 13 patients were antigen positive. Patients with chronic complaints, later found to test positive for Giardia, typically underwent five diagnostic tests at a cost of $338. CONCLUSIONS: Giardiasis, an increasing problem in family practice, should be considered early in patients with GI disturbances. New, sensitive immunodiagnostic tests that usually require a single specimen are more useful than microscopy. Prompt diagnosis of giardiasis not only relieves patients of unpleasant symptoms, but can avoid unnecessary and costly evaluations.