Multiple Reactivity Research Articles

Determination of Bovine Rotavirus G and P Serotypes in Italy by PCR

Determination of the G and P serotypes of group A bovine rotaviruses from 149 samples of feces or intestinal contents collected from calves showing clinical signs of neonatal diarrhea was performed by a nested reverse transcription-PCR typing assay. The G6 serotype was the most prevalent, accounting for viruses in 55.7% of the samples; viruses of the G10 and G8 serotypes were found in 34.9 and 4.7% of the samples, respectively. The virus in one sample (0.7%) was not classified due to concomitant infection with G6 and G8 strains, whereas viruses in six samples (4.0%) could not be characterized with any of the three G serotype-specific primers selected for the present study. When examined for their P-serotype specificities, viruses in 55 and 42.3% of the samples were characterized as P[11] and P[5], respectively, no P[1] serotype was identified, and viruses in 2.7% of the samples could not be classified due to multiple reactivity with both P[5]- and P[11]-specific primers. Various combinations of G and P serotypes were observed, the most frequent being G6,P[5] (38.3%), G10,P[11] (31.5%), and G6,P[11] (15.4%). The results of the present study, while contributing to a better understanding of the epidemiology of bovine rotaviruses in Italy, address the relevance of serotype specificity with regard to the constancy of the quality of bovine rotavirus vaccines under different field conditions.

The mycotoxin patulin induces intra- and intermolecular protein crosslinks in vitro involving cysteine, lysine, and histidine side chains, and α-amino groups

As previous studies have indicated a multiple electrophilic reactivity of patulin (PAT) towards simple thiol nucleophiles, we have methodically investigated the ability of PAT to covalently crosslink proteins in vitro. By means of sodium dodecylsulphate polyacrylamide gel electrophoresis, the formation of PAT-induced intermolecular protein-protein crosslinks was clearly demonstrated for bovine serum albumin containing one thiol group per molecule, but also for the thiol-free hen egg lysozyme. Characterization of the crosslink sites was carried out by (1) modulation of the thiol groups with N-ethylimaleimide and 2-iminothiolane; (2) comparison with various known crosslinking agents, i.e. phenylenedimaleimide, glutardialdehyde, and dimethylsuberimidate, and (3) fluorescence incorporation studies using dansyl-labeled amino acids and a fluorescent glutathione derivative. The thiol group of cysteine was preferred for PAT-mediated crosslink reactions, but the side chains of lysine and histidine, and α-amino groups also exhibited reactivity. PAT can act both as a homobifunctional as well as a heterobifunctional crosslinking agent. The initial formation of a monoadduct with a thiol group appears to activate PAT for the subsequent reaction with an amino group, but also leads to rapid loss of further electrophilic properties when no second nucleophile for crosslink completion is available. Studies using microtubule proteins as a protein with experimentally controllable quarternary structure and a proposed cellular target for PAT toxicity emphasized the influence of specific sterical conditions on crosslink formation at low protein concentrations. Non-polymerized microtubule proteins, i.e. tubulin α,β-dimers, formed a defined product with PAT consisting of an intramolecularly crosslinked β-tubulin, whereas guanosine triphosphate- or paclitaxel-induced polymerization to microtubule-like quarternary structures prior to treatment with PAT gave rise to intermolecular crosslink formation between α- and β-tubulin. In contrast, denaturated tubulin yielded none of those two new protein species, but only unspecific intramolecular crosslinks and highly crosslinked aggregates. Thus, in addition to the amino acid composition, the tertiary and quarternary superstructures of proteins appear to markedly influence their reactivity towards PAT. Under appropriate conditions, the generation of protein crosslinks could easily be observed at concentrations of PAT equal to or even below the concentration of the protein. The relevance of these novel reaction pathways of PAT demonstrated in vitro for its in vivo mechanisms of toxicity remains to be investigated.

Black Fire Ant ( Solenopsis Richteri ) Sting Producing Anaphylaxis: A Report Of 10 Cases from Najran

Black Fire Ant ( <i>Solenopsis Richteri</i> ) Sting Producing Anaphylaxis: A Report Of 10 Cases from Najran

Specific immunoglobulin E to peanut, hazelnut and brazil nut in 731 patients: similar patterns found at all ages.

Previous studies have reported reactions to an increasing range of nuts as patients with nut allergy grow older. Most patients with symptoms suggesting nut allergy have specific IgE to more than one nut. Furthermore, fatal reactions have followed eating nuts different from any causing the deceased's previous reactions. To explore the pattern of specific IgE to three distantly related nuts in patients of all ages with nut allergy. This study includes all patients referred to our laboratory for nut allergy testing from January 1994 to August 1998 who were tested for peanut, hazelnut and brazil nut, and had specific IgE to at least one of these nuts. All tests were performed using the Pharmacia Unicap system. Seven hundred and thirty-one patients (age 7 months to 65 years, median 6.6 years) had specific IgE >0.35 kU(A)/L to at least one of these three nuts: 282 had IgE to one nut, 130 to two nuts, and 319 to all three nuts. When analysed by gender and age quartile, very similar patterns were found in all subgroups though significant age trends and age interactions were found for IgE to individual nuts and combinations of nuts. The probability of a patient with nut allergy having specific IgE to a particular combination of peanut, hazelnut and brazil nut is similar, whatever their age or sex. The apparent increase in multiple nut reactivity with increasing age may therefore be due to exposure of previously unchallenged sensitivity. The frequency of multiple-nut specificity is sufficiently high that patients should always be tested for allergy to a range on nuts if they have a history of reacting to any nut.

Catalytic Upgrading of Fermentation-Derived Organic Acids.

The production of organic acids in low-cost, high-efficiency fermentation processes makes available a new route to chemical production from biomass. Because of their multiple functional groups and high reactivity, organic acids can undergo a variety of reactions that are effectively catalyzed by inorganic heterogeneous or homogeneous catalysts. Lactic acid and succinic acid, in particular, are approaching large-scale production via fermentation and show excellent promise as feedstocks for catalytic conversion routes such as hydrogenation, dehydration, or condensation. A number of catalytic conversion pathways of organic acids are potentially competitive with petroleum-based routes in the current economic environment, particularly when integrated into existing biomass/crop processing schemes. This article reviews some of the key reaction pathways available using fermentation-derived organic acids as feedstocks and presents recent results from the authors' lab on succinate hydrogenation to 1,4-butanediol and tetrahydrofuran. By a judicious choice of support properties and reaction conditions, it is possible to achieve yields of either of these two products in excess of 80%.

Sperm, nuclear, phospholipid, and red blood cell antibodies and isotype RF in infertile couples and patients with autoimmune rheumatic diseases.

To determine if measuring of nonorgan-specific autoantibodies is useful for better understanding and management of unexplained infertility. Sera were obtained from 70 infertile couples, 57 rheumatic patients, and 76 fertile donors. Sperm antibodies (SA) were detected by the tests of Kibrick and Friberg, anti-histones, anti-cardiolipin antibodies, and RF isotypes by ELISA, antinuclear antibodies by indirect immunofluorescence, and anti-red blood cell antibodies by Capture-R. Multiple autoimmune reactivity (both partners positive and/or more than one type of autoantibody involved), higher than naturally occurring in fertile individuals, was found in 55% of the idiopathically infertile couples. IgA-RF was the dominant autoimmune marker. SA revealed similar rates in patients with rheumatic diseases and in infertiles with or without other autoantibodies. Although no single autoimmunity marker could predict occurrence of SA, the coincidence of enhanced polyclonal autoimmunity in both partners of infertile couples might potentiate their negative effect on reproduction.

Antiplatelet glycoprotein autoantibodies in patients with autoimmune diseases with and without thrombocytopenia.

The presence and specificity of antiplatelet autoantibodies in 32 patients with primary and 18 patients with secondary autoimmune thrombocytopenic purpura (AITP), as well as 11 non-thrombocytopenic patients with systemic autoimmune diseases, were studied. By means of the direct and indirect monoclonal antibody immobilization of platelet antigen (MAIPA) assay, antiplatelet autoantibodies were detected using monoclonal antibodies specific for platelet glycoproteins (GPs) Ib, IIb/IIIa, Ia/IIa, and IV. Serum antiplatelet autoantibodies were found in 18 of 32 primary AITP patients (56%), 6 of 18 secondary AITP patients (33%), and 5 of 11 nonthrombocytopenic patients (45%). Platelet-associated autoantibodies were detected in five of eight patients with primary (62%) and in four of eight patients with secondary AITP (50%) and in two of four patients without thrombocytopenia (50%). Multiple antibody reactivity, mainly against GPs IIb/IIIa and Ib and, in a few patients, against Ia/IIa, was found. Using MAIPA, platelet xylene eluates from 20 patients were also studied. Antiplatelet elutable autoantibodies were related to thrombocytopenia; autoantibodies against membrane GPs Ib and IIb/IIIa were demonstrable in 84 and 63% of eluates from patients with primary and secondary AITP, respectively, but not in eluates from nonthrombocytopenic patients. The presence of antiplatelet antibodies thus appears to be a common feature of many autoimmune diseases apart from the thrombocytopenia, but the (primary or secondary) etiology of the immune thrombocytopenia cannot be differentiated on the grounds of their specificity.

Multiple false reactions in viral antibody screening assays after influenza vaccination.

In December 1991, US blood centers reported an unusual increase in donations that tested falsely reactive for antibodies to two or more (multiple false positive) of the following viruses: human immunodeficiency virus type 1 (HIV-1), human T-cell lymphotrophic virus type I (HTLV-I), and hepatitis C virus. Many of these donations were from people who had recently received the 1991-1992 influenza vaccine, raising the possibility that this vaccine had somehow specifically caused the problem of multiple false reactivity. A case-control study of 101 affected donors and 191 matched controls found that recent receipt of any brand of influenza vaccine was significantly associated with testing multiple false positive (p < 0.05), as was a history of recent acute illness (p < 0.05) and of allergies (p < 0.05). Surveillance for monthly rates of multiple reactive donations from May 1990 through December 1992 linked the seasonal cluster of multiple false-positive donations to the use of viral screening test kits thought to react nonspecifically to donor immunoglobulin M. There was no similar increase in multiple false-positive donations during the 1992-1993 influenza vaccination season after the HIV-1 and hepatitis C virus tests were replaced; however, the number of donations that were falsely reactive for only HTLV-I almost doubled, indicating that false reactivity was not specifically associated with the 1991-1992 influenza vaccine. Retesting of affected donors found that the duration of HTLV-I and hepatitis C virus false reactivity was 3-6 months. The cluster of multiple false-positive donations in 1991 was most likely caused by the test kits used, rather than by the influenza vaccine.

Donor follow-up of influenza vaccine-related multiple viral enzyme immunoassay reactivity.

False-positive enzyme immunoassay (EIA) tests in blood donors receiving influenza vaccine were first reported in 1991. We conducted follow-up testing for 6 months of those donors with multiply reactive, but unconfirmed EIA (at least 2 positives in anti-HCV-1.0, anti-HIV-1, and anti-HTLV-I assays) with a history of recent flu vaccine to determine the duration of false positivity. Of 133,000 donors tested, 16 met study criteria; all 16 were reactive for anti-HCV, 10 were reactive for anti-HIV-1, and 12 were reactive for anti-HTLV-I. Fifteen donors were available for follow-up testing (using the original screening and supplemental tests): 10 (67%) reverted to negative for the 3 tests and 5 remained false positive for various markers at last sampling (3-6 months after vaccination). The mean duration of false positivity for those reverting to negative EIA test status, was 4.2 months (range 2-7 months) indicating a transient phenomenon and supporting studies which suggest a role for IgM in the mechanism.

Donor Follow-Up of Influenza Vaccine-Related Multiple Viral Enzyme Immunoassay Reactivity

False-positive enzyme immunoassay (EIA) tests in blood donors receiving influenza vaccine were first reported in 1991. We conducted follow-up testing for 6 months of those donors with multiply reactive, but unconfirmed EIA (at least 2 positives in anti-FICV-1.0, anti-HIV-1, and anti-HTLV-I assays) with a history of recent flu vaccine to determine the duration of false positivity. Of 133,000 donors tested, 16 met study criteria; all 16 were reactive for anti-HCV, 10 were reactive for anti-HIV-1, and 12 were reactive for anti-HTLV-I. Fifteen donors were available for follow-up testing (using the original screening and supplemental tests): 10 (67%) reverted to negative for the 3 tests and 5 remained false positive for various markers at last sampling (3-6 months after vaccination). The mean duration of false positivity for those reverting to negative EIA test status, was 4.2 months (range 2-7 months) indicating a transient phenomenon and supporting studies which suggest a role for IgM in the mechanism.

Multiple reactivity of freund's “methyl isothiocyanate sulfide” in cycloaddition-elimination reactions

4-Methyl-5-methylimino-1,2,4-dithiazolidine-3-thione (l)and 2,4-dimethyl-1,2,4-thiadiazolidine-3,5-dithione (2) are interconvertible in acetone at 50°. They react with ketenes and electrophilic acetylenes to give the thiazole derivatives 6 and 7 after elimination of methyl isothiocyanate or carbon disulfide. Electrophilic isothiocyanates and phenylsulfonyl isocyanate transform the C = S function of 1 in an imine function (8), and a similar transformation of the thiourea C = S function of 2 is observed on reaction with phenylsulfonyl isocyanate, giving 9.

Characterization of nematode glycoproteins: the major O-glycans of Toxocara excretory-secretory antigens are O-methylated trisaccharides

Toxocara excretory-secretory antigens (TES) were isolated from the culture media of T.canis and T.cati larvae and their O-glycan content was investigated using fast atom bombardment-mass spectrometry (FAB-MS), gas chromatography and electron impact mass spectrometry. The major oligosaccharides released by reductive elimination of T.canis TES glycoproteins were shown to be two, approximately equi-abundant, trisaccharides: 2-O-Me-Fucp(alpha 1----2)-4-O-Me-Galp(beta 1----3)GalNAcitol and 2-O-Me-Fucp(alpha 1----2)-Galp(beta 1----3)GalNAcitol. In contrast T.cati TES O-glycans are predominantly one component, shown by FAB-MS to be a di-O-methylated trisaccharide, which is probably identical to the di-O-methylated trisaccharide from T.canis. The O-methylated trisaccharides are strong candidates for the carbohydrate epitopes recognized by a panel of monoclonal antibodies which exhibit multiple reactivity against TES antigens. This study constitutes the first rigorous characterization of glycans from a parasitic nematode.

IgG Antibodies to Gram-negative endotoxin in human sera. I. Lipopolysaccharide (LPS) cross-reactivity due to antibodies to LPS core

A series of 700 blood donor sera were screened for IgG antibodies to the core of Gram-negative bacterial endotoxin with a quantitative enzyme-linked immunosorbent-assay (ELISA), based on a cocktail of incomplete-core R-LPS from four different Gram-negative bacterial species, and further serum samples were obtained from donors exhibiting a range of different reactivity for isolation of serum IgG. Analysis of the different IgG samples by ELISA employing a panel of individual LPS from 31 different Gram-negative bacteria covering a range of species, serotypes and R-LPS chemotyes showed that high-titer sera from the screening ELISA expressed IgG with multiple reactivity to LPS in the complex ELISA. We investigated this multiple reactivity in three serum IgGs by inhibition and absorption of isolated serum IgG ELISA reactivity to R-LPS, employing purified LPS and whole bacteria respectively. In two cases the ELISA reactivity appeared to be predominantly attributable to a single antibody component directed to the inner LPS core structure in the lipid A to KDO region. For the third serum IgG, the results suggested that the cross-reactivity may be attributable to more than one specificity-group of cross-reactive antibodies, although still restricted to the LPS inner core structures.

Multiple Lineage Reactivity in Childhood Leukemia

The leukemia cells from 63 children with acute leukemia were evaluated by flow cytometry with a panel of monoclonal antibodies that included lymphoid and myeloid lineage-specific antigens. Surface markers from patients with acute lyphocytic leukemia (ALL) did not correlate with their FAB classification except for L3 leukemia. Myeloid leukemias of FAB class M1-M4 were positive for CD13 and CD33, whereas CD14 and MY8 were only detected in FAB class M4 leukemia. Mixed leukemias were subclassified as intralineage (B+T+) or interlineage (B+ or T+/M+). Interlineage leukemias represented 5.6% of ALLs, and all patients are alive after treatment with ALL protocols. Interlineage mixed leukemias represent 7.9% of all leukemias occurring in 3.7% of ALLs and 33% of acute myeloid leukemias (AMLs). All children with mixed interlineage leukemias are alive after treatment with the protocol for the dominant leukemia; however, follow-up periods are too short to predict final outcome. The high proportion of mixed interlineage leukemias in AMLs supports Greaves' theory of lineage promiscuity, that is, there is a normal period of hematopoietic development when individual cells co-express multiple lineage antigens on the cell surface.

Immunophenotyping in the classification of acute leukemia in adults. Interpretation of multiple lineage reactivity

Fifty-nine adult patients with acute leukemia were classified using a combination of the French-American-British (FAB) criteria and characterization by immunophenotyping using flow cytometric study. The authors identified 51 patients with acute myeloblastic leukemia and eight with acute lymphoblastic leukemia. This procedure permitted lineage assignment in leukemias that otherwise might have been unclassifiable. In addition, the authors demonstrated that the leukemic blasts of 29% of patients with myeloblastic disease exhibited one or more T-cell antigens on their surface. The use of immunophenotyping has greatly enhanced the authors' ability to correctly identify the lineage of acute leukemias. The data, however, must be interpreted with caution with respect to diagnosing acute mixed lineage leukemias and must be integrated with the morphologic and cytochemical evaluation of traditional classification schemes. The possible significance of T-cell markers in myeloblastic leukemia is discussed.

Allergens in Hymenoptera venom : XXI. Cross-reactivity and multiple reactivity between fire ant venom and bee and wasp venoms

The relationships between fire ant venom and bee and wasp venoms were explored by studying sera from five groups of subjects. Group 1 included adults not allergic to any venoms and who were not exposed to fire ants. Group 2 included adults with fire ant exposure who were not allergic to venoms. Group 3 included patients with recent systemic reactions to fire ant venom. Group 4 included patients allergic to bee and vespid venoms with no fire ant exposure. Last, group 5 included patients allergic to bee and vespid venoms with fire ant exposure. None of the serum samples from group 1 was RAST reactive to fire ant venom, but 24% of those from group 2 were fire ant positive, as were 100% of those from group 3, 51% of those from group 4, and 87% of those from group 5. The RAST-positive patients in groups 2 and 5 were also skin test positive. RAST inhibition studies demonstrated cross-reactivity in some cases and multiple reactivity in others. The serum samples were further investigated via nondenaturing electrophoretic immunoblot studies and RAST with highly purified allergens. Serum samples from group 4 reacted to a single band on immunoblots and with only one of the four purified allergens from fire ant venom ( Solenopsis invicta I, or Sol i I). Serum samples from groups 2, 3, and 5 showed various patterns of allergen reactivity. All serum samples from patients allergic to fire ant venom who also reacted to bee and/or vespid venoms by RAST contained IgE antibodies binding to Sol i I. A few serum samples that did not bind to bee or vespids did not have IgE antibodies against Sol i I. Sol i I, the 37,000 dalton molecular weight allergen, is the major cross-reactive allergen of imported fire ant venom.

144 Cross reactivity and multiple reactivity between fire ant and bee and vespid venoms

144 Cross reactivity and multiple reactivity between fire ant and bee and vespid venoms

Characterization of a monoclonal antibody that induces the acrosome reaction of sea urchin sperm.

A monoclonal antibody, J18/29, induces the acrosome reaction (AR) in spermatozoa of the sea urchin Strongylocentrotus purpuratus. J18/29 induces increases in both intracellular Ca2+ and intracellular pH similar to those occurring upon induction of the AR by the natural inducer, the fucose sulfate-rich glycoconjugate of egg jelly. Lowering the Ca2+ concentration or the pH of the seawater inhibits the J18/29-induced AR, as does treatment with Co2+, an inhibitor of Ca2+ channels. The J18/29-induced AR is also inhibited by verapamil, tetraethylammonium chloride, and elevated K+. All these treatments cause similar inhibition of the egg jelly-induced AR. J18/29 reacts with a group of membrane proteins ranging in molecular mass from 340 to 25 kD, as shown by immunoprecipitation of lysates of 125I-labeled sperm and Western blots. The most prominent reacting proteins are of molecular masses of 320, 240, 170, and 58 kD. The basis of the multiple reactivity appears to reside in the polypeptide chains of these proteins, as J18/29 binding is sensitive to protease digestion but resistant to periodate oxidation. There are approximately 570,000 sites per cell for J18/29 binding. J18/29 is the only reagent of known binding specificity that induces the AR; it identifies a subset of sperm membrane proteins whose individual characterization may lead to the isolation of the receptors involved in the triggering of the AR at fertilization.

Density Polarization and Chemical Reactivity

A qualitative analysis of the main polarization effects in molecular systems is presented. Both the single and multiple site reactivity cases are discussed and illustrative applications are given for selected molecular systems: acrylonitrile, propene, formamide, and the trans-nitrous acid. The polarization effects are expected to determine the directional specificity of an early approach (before the substantial charge transfer and the intramolecular rearrangements) of both reactants in a chemical reaction. The preferred direction of such an approach is therefore postulated as that generating the minimum energy change of reactants in the Field of their respective reaction partners, including the stabilization due to the relaxation of their densities in a changed environment. The example of the electrophilic substitution in the five membered heterocycles is examined. It demonstrates the importance of the density polarization effects for explaining the observed α-preference in these reactions for furan, pyrrole and N-methyl pyrrole. Next we discuss the relation between the density relaxation and the internal stability of molecular systems. Finally, we briefly comment on the Atoms-in-a-Molecule description.

Standardized extracts. Stinging and biting insects.

The state of the art in insect allergy before 1973 consisted of obtaining a history, sometimes performing skin tests with one or more insect whole body extracts (WBEs), and, if convinced of the veracity of the history, treating the patient with WBE. Although some allergists, especially Dr. Mary Loveless, 1 challenged this practice and suggested that venom and not insect bodies contained the allergens, they were generally ignored for lack of convincing scientific evidence. A 4-year-old boy was seen at the allergy clinic of Johns Hopkins University in 1973 who had experienced 2 near fatal reactions to honeybee stings, the second following a course of WBE immunotherapy. His sister had previously died from bee sting anaphylaxis, and his father was a commercial beekeeper. Lichtenstein and colleagues 2 had the father collect bee venom and attempted rush desensitization. At the equivalent of 1/3 sting (15 ~g), systemic symptoms began and they became severe at a dose of 50 ~,g. The patient was placed on a slower injection protocol and eventually could tolerate 100 p,g. At this time serologic studies showed a substantial rise in IgG antibodies against venom and the patient tolerated an intentional sting challenge. This single case was the beginning of an extensive research program at Johns Hopkins University and other institutions, which led to controlled scientific studies of many of the variables in insect venom allergy and, ultimately, to the licensing of bee venom and vespid venom protein extracts for human use. Present standards for dose, regimen, and some of the indications for immunotherapy are based on controlled studies; but many questions are still unanswered, such as the ability to predict severe reactions in untreated individuals, cross-reactivity and multiple reactivity among the venoms, and the duration of immunotherapy. Venom allergy will be an active area of research for many years. The prevalence of severe reactions to insect stings is unknown. Studies performed mainly by tabulating questionnaires filled out by parents and phy