Duration of serum antibody responses following vaccination and revaccination of cattle with non-living commercial Pasteurella haemolytica vaccines

This study was designed to determine the duration of serum antibody responses to Pasteurella haemolytica whole cells (WC) and leukotoxin (LKT) in weanling beef cattle vaccinated with various non-living P. haemolytica vaccines. Serum antibodies to P. haemolytica antigens were determined periodically through day 140 by enzyme-linked immunosorbent assays. At day 140, cattle were revaccinated, and antibody responses periodically determined through day 196. Three vaccines were used in two experiments (A and B), OneShot, Presponse HP/tK, and Septimune PH-K. In general, all three vaccines between 7 and 14 days induced antibody responses to WC after vaccination. Antibodies to LKT were induced with OneShot and Presponse. Revaccination at days 28 and 140 usually stimulated anamnestic responses. Serum antibodies to the various antigens remained significantly increased for up to 84 days after vaccination or revaccination. The intensity and duration of antibody responses were variable depending on the experiment and vaccines used. Vaccination with OneShot usually stimulated the greatest responses to WC. Vaccination with OneShot or Presponse resulted in equivalent primary anti-LKT responses. In experiment B, spontaneous seroconversion was found in numerous calves on day 112. Revaccination of those cattle at day 140 resulted in markedly variable antibody responses such that several groups had no increase in antibody responses.     

Differences between intraocular and serum antibody responses in patients with ocular toxoplasmosis

The neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) is present in many regions of the adult and developing brain as are receptors for PACAP. PACAP stimulates different signalling cascades in neurons, involving cAMP, MAP kinase, and calcium. These characteristics suggest that PACAP may influence neuronal development. Here we have studied the effects of PACAP on mesencephalic dopaminergic neurons using primary cultures from embryonic rats. PACAP increased the number of tyrosine hydroxylase (TH)-immunoreactive neurons, elevated TH protein, and enhanced tritiated dopamine uptake in these cultures. Moreover, PACAP counteracted the effects of 6-hydroxydopamine treatments, which induce cell death of dopaminergic neurons. In situ hybridisation showed that both PACAP and PACAP receptor type 1 are present in developing and adult rat mesencephalon. These results show that PACAP has a neurotrophic action on dopaminergic neurons and partially protects them against 6-OHDA induced neurotoxicity.     

Bovine mastitis caused by Listeria monocytogenes: kinetics of antibody responses in serum and milk after experimental infection

The kinetics of antibodies in serum and milk directed against proteins from Listeria monocytogenes were studied using 4 lactating cows after infection was experimentally induced in the udder with four strains of serotypes 4b or 1/2a. Antibodies (IgG and IgA) in samples of composite quarter milk and serum of the cow were measured by indirect ELISA. Microtiter plates were coated with proteins obtained from the culture supernatant of L. monocytogenes 4b. After challenge, an IgG response in serum and milk to listerial infections in the udder occurred for all cows, although the response varied among cows. In sera, the IgG titers reached a peak at 9 to 13 wk after challenge and remained elevated until 21 to 33 wk after challenge. In milk, the IgG titer increased significantly 3 wk after the challenge for all cows. A weak and nonpersistent increase in IgA antibodies also occurred. These results indicate that IMI by L. monocytogenes induced an increase of antibodies in milk, which could be detected with an ELISA test using our antigenic preparation. Therefore, this antigenic preparation could be used for the evaluation of a new method of diagnosis for bovine mastitis caused by L. monocytogenes.     

Adverse impact of infections on antibody responses to measles vaccination

Allelic frequencies for up to five short tandem repeat systems (HumTH01. HumVWA, HumF13B, HumCD4, HumD2111) were analyzed in seven population samples from Asia using the polymerase chain reaction and gel electrophoresis. No deviations from Hardy-Weinberg equilibrium were observed. Two new alleles of the CD4 and TH01 loci were detected, and sequenced and their molecular structure is presented. A phylogenetic tree based on Thai, Han Chinese (from the northeast of China), Japanese, German and Ovambo allelic frequencies was constructed and demonstrates the close relationship of the Asian populations. Additionally, allelic frequency data for the VWA and TH01 systems were determined for the south Chinese minorities Bai, Dai and Qiang and for Koreans and compared with the above data. The Bai and Dai populations were clear outliers of the cluster of all other Asians, indicating an unexpected pattern of genetic heterogeneity of the Chinese nation. Two clusters of Asian populations could be established: the Koreans and Japanese together with the Han and Qiang Chinese, and, forming a separate cluster, the Bai and Dai populations.     

Cytokine and antibody responses in women infected with Neisseria gonorrhoeae: effects of concomitant infections

The levels of interleukin (IL)-1, IL-6, IL-8, IL-10, and transforming growth factor-beta in sera and genital tract secretions from women with gonococcal cervicitis and other genital infections were examined. Cytokines were not elevated in genital secretions from gonococcus-infected compared with uninfected patients. The level of serum IL-6 was higher in gonococcus-infected than in uninfected patients at recruitment. Serum, but not local, IL-1 and IL-6 levels were elevated in patients concomitantly infected with Trichomonas vaginalis or Chlamydia trachomatis in addition to Neisseria gonorrhoeae compared with levels in patients infected with any single organism. Concomitant infection altered neither the total immunoglobulin concentrations nor the levels of antigonococcal antibodies in serum or local secretions. The results suggest that N. gonorrhoeae induces only a limited cytokine and antibody response during uncomplicated cervical infections; however, the presence of other sexually transmitted disease-causing organisms can alter the systemic cytokine but not the antigonococcal antibody levels.     

Evaluation of humoral immune responses in cattle grazing endophyte-infected or endophyte-free fescue

Anecdotal reports suggest cattle with fescue toxicosis may not respond to vaccination and thus, experience increased incidence of Bovine Respiratory Disease Complex (BRDC) when shipped to feedlots. Fescue toxicosis causes hypoprolactemia in cattle. Hypoprolactemia decreases humoral immune responses in mice. Therefore, a study was conducted to compare the magnitude of primary and secondary humoral immune responses against specific antigens in cattle grazing endophyte-infected or endophyte-free fescue. Angus steers were blocked by weight and allocated into four groups. Two groups grazed endophyte-infected (EI) fescue and the other two groups grazed endophyte-free (EF) fescue. All steers were injected IM on d 0 and 21 with lysozyme without adjuvant and concanavalin. A (Con A) with sheep red blood cells (SRBC) in incomplete adjuvant of Freund. Steers were bled on days 0, 21 and 35 post-vaccination. Average daily gains (ADG), alkaline phosphatase (ALP) activity, cholesterol concentrations, rectal temperatures, and serum prolactin concentrations were measured to confirm fescue toxicosis in steers grazing EI fescue. Antibodies to Con A and SRBC were determined by ELISA and hemagglutination assay, respectively. The ADG were decreased for the EI group during the first month. Rectal temperature were elevated and serum prolactin concentrations were decreased in the EI group. Cholesterol and ALP concentrations also were decreased in the EI group. Primary and secondary immune responses against Con A tended to be increased and were increased against SRBC in the EI group. Antibodies against lysozyme were not induced in either group. In conclusion, cattle grazing EI fescue mounted similar humoral immune responses to vaccination, despite hypoprolactemia, as cattle grazing EF fescue. Increases in bovine respiratory disease in cattle maintained on EI fescue probably is not associated with lack of humoral immune response to vaccination protocols as a result of fescue toxicosis.     

Serological survey of antibody to Neospora caninum in Japanese dogs

Prevalence of antibody to Neospora caninum (NC) in Japanese dogs were examined. The antibody was positive in 15 of 48 dogs (31.3%) reared in the dairy farms that had case of the abortions due to NC infection or had the cattle seropositive to NC, whereas the prevalence was 7.1% (14 of 198 dogs) among the dogs kept in urban areas. In one dog breeder, all 17 Shetland sheepdogs older than 7 months were seropositive, and one pup was diagnosed as neosporosis 2 months before the first serological examination. The antibody titers of the dogs kept at this breeder were almost unchanged for 1.5 years. Serological evidence of the dogs in the dairy farms and urban areas indicates the transmission of NC between dogs and cattle. Also serological results of the dogs in one breeder may suggest the potential horizontal transmission among dogs.     

Detection of mutant K-ras DNA in plasma or serum of patients with colorectal cancer

Increased understanding of the molecular basis of colorectal cancer and recognition that extracellular DNA circulates in the plasma and serum of cancer patients enables new approaches to detection and monitoring. We used a polymerase chain reaction (PCR) assay to demonstrate mutant K-ras DNA in the plasma or serum of patients with colorectal cancer. Plasma or serum was fractionated from the blood of 31 patients with metastatic or unresected colorectal cancer and from 28 normal volunteers. DNA was extracted using either a sodium chloride or a gelatin precipitation method and then amplified in a two-stage PCR assay using selective restriction enzyme digestion to enrich for mutant K-ras DNA. Mutant K-ras DNA was detected in the plasma or serum of 12 (39%) patients, all confirmed by sequencing, but was not detected in any of the normal volunteers. K-ras mutations were detected in plasma or serum regardless of sex, primary tumour location, principal site of metastasis or proximity of chemotherapy and surgery to blood sampling. Tumour specimens available for 19 of the patients were additionally assayed for ras mutations and compared with blood specimens. Our results indicate mutant K-ras DNA is readily detectable by PCR in the plasma or serum of patients with advanced colorectal cancer. Thus, plasma- or serum-based nucleic acid amplification assays may provide a valuable method of monitoring and potentially detecting colorectal cancer.     

Chronic Sarcocystis infections in slaughtered cattle

Parasitological, histopathological and immunohistochemical examinations were carried out on three slaughtered cattle with many nodules in all the striated muscles. At necropsy, many yellowish green rice-grain sized nodules including cheesy contents were observed in all the striated muscles. Histopathologically the nodules were granuloma principally consisting of eosinophiles. No Sarcocystis cysts nor bradyzoites were found in the nodules, but intact sarcocysts were found in the normal tissues surrounding the nodules. The central necrotic focus of nodules showed intense positive responses against anti-Sarcocystis cruzi rabbit serum by immunohistochemical examination. From the above findings the slaughtered cattle were diagnosed as chronic sarcocystiasis.     

Neospora caninum causes abortions in a cattle herd in North Rhine Westphalia]

In a cattle herd in North Rhine-Westphalia a series of eight abortions occurred between 15th August and 14th October, 1996. Four of the aborted fetuses were histopathologically, virologically, bacteriologically and parasitologically examined. A multifocal necrotising encephalitis was observed in two fetuses. In addition, the liver of two fetuses and the placenta of one fetus contained infected foci. In three fetuses protozoan stages were detected which were identified as N. caninum by immunohistochemistry. A polymerase chain reaction conducted with histological material revealed N. caninum-DNA in the placenta of an aborted fetus. When seven cows that had aborted were tested for antibodies directed against N. caninum, six animals were positive by immunofluorescence and all seven by immunoblotting. The serological examination of the entire herd revealed a high herd seroprevalence of antibodies directed against N. caninum. Potential modes of infection with N. caninum in the herd are discussed, in particular the possibility of exposure to a point source (definitive host).     

Detection of thioredoxin in human serum and biological samples using a sensitive sandwich ELISA with digoxigenin-labeled antibody

Currently, recombinational cloning procedures based upon methods developed for yeast, Saccharomyces cerevisiae, are being exploited for targeted cloning and in-vivo modification of genomic clones. In this review, we will discuss the development of large-insert vectors, homologous recombination-based techniques for cloning and modification, and their application towards functional analysis of genes using transgenic mouse model systems.     

Detection of altered adhesion molecule expression in experimental tumors by a radiolabeled monoclonal antibody

Adhesion molecules play a major role in the processes of invasion and metastasis of malignant tumors. Their expression within tumors has been reported to be quantitatively and qualitatively altered according to the invasiveness and metastatic potential of the tumor. The present study tested whether the intratumoral expression of integrin alpha 3 can be detected by a radiolabeled monoclonal antibody. The in vitro binding study with four different human cancer cells showed that radioiodinated GA17 antibody recognizing integrin alpha 3 bound specifically to these cells to varying degrees, according to the antigen density on each cell. The biodistribution study with 125I- and 111In-labeled antibodies showed specific localization of radiolabeled GA17 to the xenografts. However, the in vivo tumor localization was not proportional to the antigen density calculated in vitro, and antibody metabolism varied among the tumors, as was also confirmed by in vitro radionuclide retention assay. The intratumoral distribution of radioactivities varied reflecting the antigen expression within the tumor. These results indicate that 1) integrin alpha 3 was expressed in various kinds of tumors and could be localized by the radiolabeled antibody, and 2) the expression of integrin alpha 3 and the metabolism of the radiolabeled antibody after binding to the antigen within the tumor were variable among the tumors, which affected the radionuclide distribution characteristics. The expression of adhesion molecules within these tumors was noninvasively detected by a radiolabeled antibody. It may be possible to use integrin alpha 3, when it is overexpressed, as a target of therapy with antibodies radiolabeled with alpha or beta emitters.     

Isotype-specific antibody responses to foot-and-mouth disease virus in sera and secretions of "carrier' and "non-carrier' cattle

Isotype-specific antibody responses to foot-and-mouth disease virus (FMDV) were measured in the sera and upper respiratory tract secretions of vaccinated and susceptible cattle challenged with FMDV by direct contact or by intranasal inoculation. A comparison was made between cattle that eliminated FMDV and those that developed and maintained a persistent infection. Serological and mucosal antibody responses were detected in all animals after challenge. IgA and IgM were detected before the development of IgG1 and IgG2 responses. IgM was not detected in vaccinated cattle. Challenge with FMDV elicited a prolonged biphasic secretory antibody response in FMDV "carrier' animals only. The response was detected as FMDV-specific IgA in both mucosal secretions and serum samples, which gained statistical significance (P < 0.05) by 5 weeks after challenge. This observation could represent the basis of a test to differentiate vaccinated and/or recovered convalescent cattle from FMDV "carriers'.     

Heterogeneous antibody responses in tuberculosis

Antibody responses during tuberculosis were analyzed by an enzyme-linked immunosorbent assay with a panel of 10 protein antigens of Mycobacterium tuberculosis. It was shown that serum immunoglobulin G antibodies were produced against a variety of M. tuberculosis antigens and that the vast majority of sera from tuberculosis patients contained antibodies against one or more M. tuberculosis antigens. The number and the species of serologically reactive antigens varied greatly from individual to individual. In a given serum, the level of specific antibodies also varied with the antigen irrespective of the total number of antigens recognized by that particular serum. These findings indicate that person-to-person heterogeneity of antigen recognition, rather than recognition of particular antigens, is a key attribute of the antibody response in tuberculosis.     

Immunoepidemiology of Dracunculus medinensis infections II. Variation in antibody responses in relation to transmission season and patency

Recent studies have shown that mutations in the hepatocyte nuclear factor (HNF)-4alpha gene give rise to maturity-onset diabetes of the young, type 1 (MODY1). HNF-4, an orphan member of the nuclear receptor superfamily, contains a DNA-binding domain (DBD) and a putative ligand-binding domain (LBD) that can act independently of each other. The first MODY1 mutation identified creates a stop codon at amino acid 268 in the LBD of HNF-4 (Q268X) that leaves the DBD intact, suggesting that the mutant protein may retain some of the properties of the wild-type protein. To determine the functional properties of this mutant, we constructed HNF4.Q268X and tested it in vitro and in vivo for DNA binding, protein dimerization, and transactivation activity. Results of an electrophoretic mobility shift assay showed that HNF4.Q268X neither binds DNA alone nor binds it as a dimer with wild-type HNF-4 (HNF4.wt). In contrast, a co-immunoprecipitation assay showed that HNF4.Q268X is capable of dimerizing in solution with HNF4.wt. Transient transfection assays, however, indicated that HNF4.Q268X does not affect transactivation by HNF4.wt in vivo, supporting the argument against a dominant negative effect. Additional results suggest that the lack of a dominant negative effect could be due to a striking differential subcellular localization of the HNF4.Q268X protein: HNF4.Q268X could be extracted from transfected cells only when treated with SDS. Taken together, our results suggest that the MODY1 phenotype is due to a loss of functional HNF-4 protein that is aggravated in tissues that express relatively low amounts of HNF-4, such as pancreas.     

Local immune responses in colon from cattle infected with Fasciola hepatica

Colonic mucosae isolated from eight Charolais heifers 14 weeks after oral infection with Fasciola hepatica metacercariae were voltage clamped in Ussing chambers. Antigen challenge provoked a rapid-onset, inward short circuit current in infected but not in control tissue preparations. Chloride secretion accounted for part of the response as the loop diuretic bumetanide attenuated the response to antigen by 89%. The bacterial cell wall component n-formyl-methionyl-leucyl-phenylalanine induced a similar response to antigen and partly desensitised tissues to subsequent antigen challenge, consistent with the involvement of immunocytes in the response to antigen. Infection induced an elevation in the number of tissue eosinophils compared with control colon.     

Cytotoxic T-lymphocyte responses in cattle infected with bovine viral diarrhea virus

CRP1, a Drosophila nuclear protein that can catalyze decondensation of demembranated Xenopus sperm chromatin was cloned and its primary structure was deduced from cDNA sequence. Alignment of deduced amino acid sequence with published sequences of other proteins revealed strong homologies to Xenopus nucleoplasmin and NO38. CRP1 is encoded by one or several closely related genes found at a single locus, position 99A on the right arm of chromosome 3. CRP1 mRNA is expressed throughout Drosophila development; it is highest during oogenesis and early embryogenesis. mRNA levels correlate closely with levels of protein expression measured previously. Results of chemical crosslinking indicate that CRP1 is either tetrameric or pentameric; similar ambiguity was revealed by direct visualization using scanning transmission electron microscopy. Consistent with previously published results, parallel crosslinking studies of Xenopus nucleoplasmin suggested a pentameric structure. Scanning transmission electron microscopic examination after negative staining revealed that CRP1 and Xenopus nucleoplasmin are morphologically similar. CRP1 is able to substitute for nucleoplasmin in Xenopus egg extract-mediated sperm chromatin decondensation. In vitro, CRP1-induced decondensation is accompanied by direct binding of CRP1 to chromatin.     

Dipstick enzyme immunoassay for rinderpest antibody in cattle

A dipstick enzyme immunoassay (ELISA) has been standardized for the detection of rinderpest antibodies. One hundred and thirty bovine serum samples were analysed by the dipstick ELISA method and the results compared with the conventional plate ELISA method. The sensitivity was found to be similar in both methods. The dipstick ELISA does not require expensive micro-plates and an ELISA reader, and is recommended for use in field laboratories where the qualitative detection of rinderpest antibodies is required.     

Detection of serum interferon-alpha by dissociation-enhanced lanthanide fluoroimmunoassay. Studies of patients with acute viral and bacterial infections

A sensitive dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) was evaluated for ability to detect interferon-alpha (IFN-alpha) in serum of patients with acute infectious disease of less than one week's duration and a fever of > 38 degrees C. None of 36 patients with confirmed or probable bacterial disease was IFN-alpha positive. In contrast, 13/26 patients with viral infections had detectable levels of IFN-alpha in serum, all clearly positive (> or = 10 U/ml). The IFN-alpha positive serum samples were obtained early after onset of clinical disease, after a mean of 2.4 days. The IFN-alpha positive samples were obtained from 10 of the 12 patients with influenza or flu-like infection, and 3 of the 5 patients with varicella or herpes zoster. The IFN-alpha negative patients with viral disease (n = 9) included five patients with mononucleosis. The DELFIA should be useful in further studies of the value of IFN-alpha determinations in the identification of acute viral infections.     

The effect of immunization route on rat serum and tear antibody responses to Chlamydia trachomatis

Using reliable displacement radiobinding assay (RBA) and ELISA, the existence of anti-human growth hormone autoantibodies (hGHAA) was confirmed in idiopathic hypopituitary patients with growth impairment. Six of 35 hypopituitary patients (17.1%) and 1/85 (1.2%) control children proved positive for hGHAA by RBA (>control mean + 3 SD). IgG isotype-hGHAA by ELISAIgG (> control mean + 3 SD) were positive for 6/34 (17.7%) and 3/85 (3.5% hypopituitary and control children, respectively. Due to an asymmetry to the right of the ELISAIgG distribution, an alternative cutoff based on a nonparametric method was obtained, and positive results for hypopituitary children increased to 10/34 (29.4%). Three of 34 hypopituitary patients but no control children were positive for hGHAA of IgM isotype. The hGHAA were detected in children with or without perinatal problems. These autoantibodies may represent markers of a major autoimmune process involving a portion of the anterior pituitary and may contribute to the development of hypopituitarism in over 15% of hypopituitary children.