Antigens Of Other Species Research Articles

Evaluation of the immunogenic properties of the recombinant Histophilus somni outer membrane protein 40 kDa (rOMP40)

BackgroundGram-negative bacterial infections are a serious problem in beef and dairy cattle. Bacterial outer membrane proteins (OMPs) play a pivotal role in cellular survival and the host-bacterium interaction. Histophilus somni OMP40 was identified as a porin with homology between its N-terminal amino acid sequence and the sequences of porins of other gram-negative bacteria The aim of this study was to produce recombinant H. somni OMP40 (rOMP40), optimize its production and evaluate its immunogenic properties in calves. The cross-reactivity of anti-rOMP40 antibodies were also checked.ResultsThe highest overexpression of rOMP40 was demonstrated by Escherichia coli C41 using the autoinduction process. Double immunization of calves (20 μg rOMP40 per animal) induced a significant increase of anti-rOMP40 antibodies in the IgG1 (P ≤ 0.01) and IgG2 (P ≤ 0.01, after first immunization only) subclasses, but not IgM. ELISA revealed increased reactivity of the IgG against surface antigens of E. coli and Pasteurella multocida after the second immunization (P < 0.01). Cross reactivity of anti-rOMP40 antibodies with ~ 40 kDa antigens of most common gram-negative pathogens was shown by Western blotting.ConclusionImmunization with H. somni rOMP40 induced a humoral response in cattle with broad cross-reactivity with similar antigens of other species of Pasteurellaceae and Enterobacteriaceae families and the delayed-type hypersensitivity reaction. The obtained results encourage further study to evaluate the protective effect of the produced protein as a subunit vaccine in cattle.

Conserved Gammaherpesvirus Protein Kinase Selectively Promotes Irrelevant B Cell Responses.

Gammaherpesviruses are ubiquitous pathogens that are associated with B cell lymphomas. In the early stages of chronic infection, these viruses infect naive B cells and subsequently usurp the B cell differentiation process through the germinal center response to ensure latent infection of long-lived memory B cells. A unique feature of early gammaherpesvirus chronic infection is a robust differentiation of irrelevant, virus-nonspecific B cells with reactivities against self-antigens and antigens of other species. In contrast, protective, virus-specific humoral responses do not reach peak levels until a much later time. While several host factors are known to either promote or selectively restrict gammaherpesvirus-driven germinal center response, viral mechanisms that contribute to the irrelevant B cell response have not been defined. In this report we show that the expression and the enzymatic activity of the gammaherpesvirus-encoded conserved protein kinase selectively facilitates the irrelevant, but not virus-specific, B cell responses. Further, we show that lack of interleukin-1 (IL-1) receptor attenuates gammaherpesvirus-driven B cell differentiation and viral reactivation. Because germinal center B cells are thought to be the target of malignant transformation during gammaherpesvirus-driven lymphomagenesis, identification of host and viral factors that promote germinal center responses during gammaherpesvirus infection may offer an insight into the mechanism of gammaherpesvirus pathogenesis.IMPORTANCE Gammaherpesviruses are ubiquitous cancer-associated pathogens that usurp the B cell differentiation process to establish life-long latent infection in memory B cells. A unique feature of early gammaherpesvirus infection is the robust increase in differentiation of B cells that are not specific for viral antigens and instead encode antibodies that react with self-antigens and antigens of other species. Viral mechanisms that are involved in driving such irrelevant B cell differentiation are not known. Here, we show that gammaherpesvirus-encoded conserved protein kinase and host IL-1 signaling promote irrelevant B cell responses and gammaherpesvirus-driven germinal center responses, with the latter thought to be the target of viral transformation.

Infection with Paragonimus westermani of boar-hunting dogs in Western Japan maintained via artificial feeding with wild boar meat by hunters.

Infection of boar-hunting dogs with Paragonimus westermani was investigated in Western Japan. Blood and rectal feces were collected from 441 dogs in the three districts (205 in Kinki, 131 in Chugoku and 105 in Shikoku District). In a screening ELISA for serum antibody against P. westermani antigen, 195 dogs (44.2%) showed positive reaction. In the 195 dogs, 8 dogs were found excreting P. westermani eggs after molecular analysis of fecal eggs, and additional 7 were identified serologically for the parasite infection because of their stronger reactivity against P. westermani antigen than against antigens of other species of Paragonimus. A spatial analysis showed that all of the P. westermani infections were found in Kinki and Chugoku Districts. In this area, dogs’ experience of being fed with raw boar meat showed high odds ratio (3.35) to the sero-positivity in the screening ELISA, and the frequency of such experiences was significantly higher in sero-positive dogs. While clear relationship was not obtained between predation of boars by dogs during hunting and their sero-positivity. Therefore, it is suggested that human activity of feeding with wild boar meat is the risk factor for P. westermani infection in boar-hunting dogs. Considering that hunting dogs could play as a major definitive host and maintain the present distribution of P. westermani in Western Japan, control measures for the infection in hunting dogs, such as prohibition of raw meat feeding and regular deworming, should be undertaken.

The Elephant Interferon Gamma Assay: A Contribution to Diagnosis of Tuberculosis in Elephants

Mycobacterium tuberculosis (M. tb) has been shown to be the main causative agent of tuberculosis in elephants worldwide. M. tb may be transmitted from infected humans to other species including elephants and vice versa, in case of prolonged intensive contact. An accurate diagnostic approach covering all phases of the infection in elephants is required. As M. tb is an intracellular pathogen and cell-mediated immune (CMI) responses are elicited early after infection, the skin test is the CMI assay of choice in humans and cattle. However, this test is not applicable in elephants. The interferon gamma (IFN-γ) assay is considered a good alternative for the skin test in general, validated for use in cattle and humans. This study was aimed at development of an IFN-γ assay applicable for diagnosis of tuberculosis in elephants. Recombinant elephant IFN-γ (rEpIFN-γ) produced in eukaryotic cells was used to immunize mice and generate the monoclonal antibodies. Hybridomas were screened for IFN-γ-specific monoclonal antibody production and subcloned, and antibodies were isotyped and affinity purified. Western blot confirmed recognition of the rEpIFN-γ. The optimal combination of capture and detection antibodies selected was able to detect rEpIFN-γ in concentrations as low as 1 pg/ml. The assay was shown to be able to detect the native elephant IFN-γ, elicited in positive-control cultures (pokeweed mitogen (PWM), phorbol myristate acetate plus ionomycin (PMA/I)) of both Asian and African elephant whole-blood cultures (WBC). Preliminary data were generated using WBC from non-infected elephants, a M. tb infection-suspected elephant and a culture-confirmed M. tb-infected elephant. The latter showed measurable production of IFN-γ after stimulation with ESAT6/CFP10 PPDB and PPDA in concentration ranges as elicited in WBC by Mycobacterium tuberculosis complex (MTBC)-specific antigens in other species. Hence, the IFN-γ assay presented potential as a diagnostic tool for the detection of elephant tuberculosis. Validation of the assay will require its application in large populations of non-infected and infected elephants.

A novel tegumental protein 31.8 kDa of Clonorchis sinensis: sequence analysis, expression, and immunolocalization

The complementary DNA (cDNA) plasmid libraries of adult worm, metacercariae and egg of Clonorchis sinensis (C. sinensis) were constructed for researches on genomics and proteomics of C. sinensis. The full-length cDNA sequence encoding tegumental protein 31.8 kDa (CsTP31.8) was identified from the adult cDNA library. The cDNA sequence has been submitted to the GeneBank Database with accession number ABK60086. This novel cDNA sequence contains 828 bp with a putative open reading frame of 275 amino acids. The deduced amino acid sequence shows identity to membrane-associate antigens or tegumental antigens of other species. There were conserved calcium-binding EF hand and dynein light chain type 1 in the sequence. CsTP31.8 transcripts were detected in cDNA libraries of adult worm and metacercariae but not in that of egg. Recombinant CsTP31.8 was expressed and purified from Escherichia coli BL21 (DE3). CsTP31.8 was immunolocalized at the tegument of adult C. sinensis by using antirecombinant CsTP31.8 sera.

Occurrence and specificity of human natural and in vitro induced antibodies to Nocardia opaca antigens

Nocardia opaca, a Gram-positive bacterium, is a potent source of immunostimulatory substances. Screening of sera of adult human donors revealed that all sera tested contained antibodies reactive with isolated Nocardia fractions (Nocardia delipidated cell mitogen, NDCM; Nocardia lysozyme digest, NLD; Nocardia water-soluble mitogen, NWSM; and fraction B). The respective values of reciprocal titres for IgM and IgG were in the range of 100 to 12 800, and 10 to 320 for IgA antibody isotypes, when NLD or fraction B were used as antigens in enzyme-linked immunosorbent assay (ELISA) tests. The level of antibodies directed to NDCM, a potent polyclonal B cell activator, was found to be the lowest. In ritro spontaneous as well as NDCM-induced production of antibodies to NDCM by human peripheral blood lymphocytes involved mainly the IgM class. Western-blot analysis demonstrated that antibodies in normal human sera react with nocardial antigens of molecular mass ∼60, 40, 20 and 15–10 kDa. The same antigens were also recognized by rabbit and mouse hyperimmune sera, also confirming the immundominancy of these nocardial antigens in other species. The presence of anti-nocardia antibodies in human sera and their production by both stimulated and non-stimulated lymphocytes points to the natural sensitization of humans either by ubiquitous nocardial components or by cross-reactive bacterial or food antigens.

Identification and partial characterization of a parasite antigen in sera from humans infected with Wuchereria bancrofti.

The purpose of this study was to identify and characterize soluble parasite antigens present in sera from humans infected with the filarial nematode Wuchereria bancrofti. Affinity chromatography and immunoblot methods were used to demonstrate a 200,000 m.w. circulating parasite antigen in sera from infected humans which corresponded to an antigen released by adult W. bancrofti during in vitro culture. Two monoclonal antibodies were produced to this antigen by immunizing mice with antigens from Dirofilaria immitis, a filarial parasite that is closely related to W. bancrofti, and screening cell fusion supernatants by enzyme immunoassay and counterimmunoelectrophoresis inhibition. The antibodies bound to a single repeated epitope (not phosphorylcholine) that was resistant to heat, acid, and protease treatments but sensitive to periodate oxidation. Immunoperoxidase studies showed that the epitope was concentrated in the cuticle and reproductive organs in D. immitis, and it was released in relatively large amounts by adult female D. immitis in vitro. The epitope is also present in antigens of other species of filarial and nonfilarial nematodes, but on the basis of preliminary studies, its presence in human serum appears to be specific for W. bancrofti infection.

Sharing of Ia antigens between species. IV. Interspecies cross-reactivity of monoclonal antibodies directed against polymorphic mouse Ia determinants.

The specificity of interspecies Ia cross-reactions has been analyzed by testing a panel of monoclonal antibodies (mAb) to mouse I-E and I-A antigens for reactivity with pig Ia antigens. Our earlier studies showed that mouse anti-I-E alloantisera recognized common determinants on Ia antigens of other species, whereas anti-I-A alloantisera showed much more limited cross-reactivity. These results were confirmed using a panel of 17 anti-I-E mAb, 10 of which were cytotoxic to pig cells. 2D gel electrophoretic analyses of precipitates with these mAb of 35S-labeled, NP40 solubilized pig cells revealed a limited set of protein spots that appeared to be identical to the subset of pig Ia antigens precipitated by A.TH anti-A.TL alloantiserum. Because the cross-reactive mouse sera were produced in mouse strains that do not express an I-E molecule (H-2b and H-2s), it was anticipated that the cross-reacting antibodies would be reactive with the monomorphic determinant of the I-E molecule, Ia.7. However, comparison of the reactivity of these mAb with pig cells and mouse cells revealed that the cross-reactivity on pig cells correlated not with Ia.7 but rather with detection of epitope(s) of the I-E molecule associated with inter-strain polymorphism. Anti-I-A cross-reactions were also detected, but were weaker and more limited. These findings may have implications for the evolution of Ia antigens in mammalian species.

Serological and genetic identification of a bovine B lymphocyte alloantigen system.

A two-colour fluorescence microcytotoxicity test was used to screen antisera for antibodies specific for bovine B lymphocytes. A total of 114 cattle alloantisera were screened against peripheral blood lymphocytes from 100 unrelated individuals. Anti-B lymphocyte activity was detected in 47 antisera. Cytotoxic antibodies to antigens other than B lymphocyte specific antigens were removed by absorbing the antisera with buffy coat cells or platelets isolated from whole blood. Selected antisera were used to type paternal half-sib families. The presence of a polymorphic, MHS-linked antigen system on B lymphocytes was demonstrated. The tissue distribution and MHS linkage of these antigens suggests this system is analogous to the class II or Ia antigens of other species.

Biochemistry of the Variant Surface Glycoproteins of Salivarian Trypanosomes

This chapter describes the progress made over the past years in characterizing antigens of the principal pathogenic salivarian trypanosomes. Although bulk of information comes from work on the variant antigens of T. b. brucei in particular, much interesting work has been done on the variant antigens of other species and is discussed in the chapter. Future perspectives for immunoprophylaxis, serodiagnosis, or chemotherapy of trypanosome infections are also presented. The mammalian trypanosomes have been divided into seven subgenera that are grouped into two sections based on the site of the production of infective metacyclic trypanosomes in the invertebrate host. The subgenera are Trypanosoma brucei brucei , Trypanosoma brucei gambiense , Trypanosoma brucei rhodesiense , Trypanosoma evansi , Trypanosoma equiperdum Trypanosoma ( Nannomonas ) congolense , and Trypanosoma ( Duttonella ) vivax . The chapter presents the systematic position of the genus Trypanosoma and the seven subgenera that infect mammals and summarizes the main features of each subgenus. Antigenic variation in the mammalian host is the most striking characteristic of salivarian trypanosomes. Variant surface glycoproteins (VSGs) are synthesized with a hydrophobic C-terminus absent from purified VSG. Little work has been done on the biochemistry of antigens expressed by metacyclic trypanosomes of any species, because the amount of material available is limited. There is interest in the antigenic composition of metacyclic trypanosomes because if the repertoire is restricted, vaccination could be a possibility. The serological evidence suggests that the repertoire of metacyclic variant antigen types (VATS) is restricted, and there is limited overlap between VATS present in metacyclic and in bloodstream populations.

Interspecies exchange of beta 2-microglobulin and associated MHC and differentiation antigens.

Radiolabeled human beta 2-microglobulin (beta 2m) can bind to mouse histocompatibility (H-2) antigens on the cell surface or to partially purified H-2 antigens in solution. The complexes containing human beta 2m and H-2 antigens from C3H (H-2k) mice could be immunoprecipitated specifically with alloantisera, rabbit anti-H-2 xenoantisera, and with monoclonal H-2-specific antibodies. Specific association with H-2 antigens was also observed with other haplotypes. The only exception was B10.D2 (H-2d) from which complexes containing human beta 2m could only be precipitated with anti-H-2-xenosera. Thus radiolabeled human beta 2m can be used as a specific label for mouse H-2 antigens in precipitation and radioimmunoassays. The application of this finding extends to major histocompatibility complex antigens of other species, and to differentiation antigens with primary association with beta 2m.

Precipitating and Agglutinating Activity in Duck Anti-Soluble Protein Immune Sera

Ducks were inoculated parenterally, up to 4 times, with bovine serum albumin (BSA), B gamma globulin (BGG), or horse gamma globulin (HGG). Their sera were tested in immunodiffusion tests for precipitins. Only BSA and BGG induced precipitins. Only 7 of 443 sera tested, obtained from 3 of 62 inoculated ducks, developed a precipitin line with homologous antigen. All 7 sera were obtained from the earliest bleeding (7 days) after inoculation. Sera were also tested for agglutinins in direct passive hemagglutination (DPHA) and direct red-cell-linked antigen tests (DRCLAT) and for nonagglutinating antibodies in indirect red-cell-linked antigen tests (IRCLAT). No duck had a passive-hemagglutination-demonstrable primary immune response. Demonstrable DPHA titers after subsequent inoculations were very low compared with responses to these antigens of other species noted by other workers. Duck immune response was greatly diversified: all inoculation regimens that induced agglutinins in some ducks left others completely unstimulated. Precipitins and agglutinins of the same ducks correlated well: those 7 sera that had precipitins also had the highest DPHA titers for the 3 donor ducks. However, hemagglutinating titer and presence of precipitins in sera of different ducks correlated poorly. Increasing the age of the ducks at the first inoculation from 6 to 10 weeks increased the number of responders and DPHA titers of their sera. Non-agglutinating antibodies were demonstrated in anti-BSA sera: in IRCLAT, the sera had titers 2-to-256-fold higher than the sera in DPHA or DRCLAT had. Duck immunoglobulins were deficient in those immunological reactions (precipitation and agglutination) that require functional bivalency.

Partial N-terminal amino acid sequences of guinea pig classic histocompatibility antigens.

Abstract Partial N-terminal amino acid sequences were determined by microsequencing for three of the guinea pig GPLA-B classic histocompatibility antigens, which are presumably allelic products. The sequences of GPLA-B.1 derived from the noncongenic inbred strain 2 and strain 13 animals showed no differences a their N-termini, suggesting that these molecules are chemically as well as serologically identical. Two residues, isoleucine and leucine, were found at position 5 of the sequence of GPLA-B.1, suggesting that GPLA-S, the product of the second classic histocompatibility locus, was being cosequenced. GPLA-B.2 and GPLA-B.3 antigens showed distinct sequence differences from GPLA-B.1 and from each other. The sequences of GPLA-B.3 derived from a random-bred guinea pig and from the inbred DHCBA strain showed no differences and showed a possible species specific residue. All sequences showed significant homology with histocompatibility antigens of other species, suggesting a common evolutionary origin.

Structural characterization of antigens encoded by rabbitRLA-11 histocompatibility genes

Products of rabbitRLA-11 histocompatibility genes were isolated from rabbit lymphoid tumor cells (RL-5) and characterized. The tumor cells were grown in culture with radioactive amino acids and were lysed by treatment with the detergent NP-40. Glycoprotein molecules were isolated by affinity chromatography using immobilized lentil lectin. Chromatography on purified sheep anti-rabbit beta-2-microglobulin yielded a 43,000 dalton glycoprotein fraction, designated RLA-11gp, which was noncovalently associated with beta-2-microglobulin. N-terminal amino acid sequence analysis using radiochemical methods allowed assignment of 28 of the N-terminal 35 residues. The data revealed 89% homology of the RLA-11gp N-terminus with that of the human HLA-B7 and 82% with the mouse H-2Kb histocompatibility antigens. Comparison of the RLA-11 gp N-terminal sequence data obtained in this work to sequence data reported for major histocompatibility complex antigens of other species revealed no amino acid substitutions unique to the rabbit antigens.

Identification of H-2 and Ia-antigen analogues in several species by immunological crossreactions of xenoantisera.

Rabbit antisera against H-2K and D antigens react with molecules composed of 12,000 and 45,000 dalton subunits derived from human, monkey, rat, guinea-pig, cow and pig lymphocytes. The antisera failed, however, to react with similar type molecules from chicken. A rabbit antihuman beta 2-microglobulin serum reacted with chicken molecules comprising 12,000 and 45,000 dalton polypeptide chains. An antiserum against HLA-DR antigens reacted with Ia-antigen-like molecules from monkey, rat, mouse, guinea-pig, cow, pig and chicken lymphocytes. All Ia-antigen-like molecules displayed two dissimilar subunits. The present data suggest that xenoantisera directed against highly purified MHC antigens from one species may be useful tools in elucidating the structure of similar antigens in other species where alloantisera are not available.

Gene dosage effect in expression of blood group antigens.

AbstractChicken blood cells were investigated with respect to the distribution of A blood group antigens on the cell membrane surface via the fluorescent antibody technique. A direct gene‐blood group antigen relationship was found. Erythrocytes from chickens homoqygous for A6 antigen fluoresced more intensely than those from birds heterozygous for the antigen. The respective intensities, approximating a 2:1 ration, idicate a direct gene dosage effect in the expression of at least these antigens. This novel finding disccussed with respect to the possible molecular constitution of and blodd group antigens in other species (e.g., human). This finding has important iplications leading to the understanting of the nature of blood group antigens and the specific role they play in reconstructive surgery.

Serological diagnosis of scrub typhus by indirect immunofluorescence.

The indirect fluorescent antibody test, in which dilutions of sera from patients with scrub typhus were tested on smears of Rickettsia tsutsugamushi, gave results which were serologically diagnostic of the disease in each instance. Non-specific reactions did not occur when the scrub typhus sera were exposed to antigens of other species of Rickettsia. Likewise, there was no reaction between R. tsutsugamushi smears and sera from other rickettsial infections, or with sera from patients with Proteus OXK agglutinins of non-rickettsial origin with one exception, i.e., a case of leptospirosis from Puerto Rico.

Natural occurrence of delayed-type iso-hypersensitiveness.

EVIDENCE has been obtained for the existence, in certain normal guinea pigs, of a state of delayed iso-hypersensitiveness to a heritable factor present in the serum of other normal guinea pigs. This condition, which arises at about 2 months of age, appears analogous to the long known1 natural existence of circulating iso-antibodies to heritable blood group antigens in other species. Delayed-type hypersensitivity to isophile antigens may, of course, be experimentally induced (cf. ref. 2), but its spontaneous occurrence seems not to have been suspected.