Microagglutination Procedures for Febrile Agglutination Tests

Febrile agglutination tests were done by using as antigens Brucella abortus, Salmonella group D, Proteus OX19, and Pasteurella tularensis. Comparison of results from 23 sera showed that the microtechnique, rapid slide, and test tube methods gave similar titers, although those from the microtechnique were generally higher. The sensitivity of the microtechnique depended upon the concentration of antigen, and, to obtain reproducible results, the optimal concentration of antigens had to be determined by preliminary titrations against specific, positive control antisera. Readability of reactions in the microtechnique was enhanced by adding the dye Safranin O to diluent for antigen and by use of V-type, rather than U-type, microtiter plates. Tests were also done to determine the effects of dye and salt concentrations, pH, and temperature of incubation upon the titer of agglutinations by the microtechnique. Our results indicated that the microtechnique could be used for agglutination tests involving febrile antigens. The procedure is less time-consuming than the tube method and requires less antigen and serum than the latter method or the rapid slide method.     

Long-distance dispersal of the gypsy moth (Lepidoptera: Lymantriidae) facilitated its initial invasion of Wisconsin

Gypsy moth (Lymantria dispar L.) spread is dominated by stratified dispersal, and, although spread rates are variable in space and time, the gypsy moth has invaded Wisconsin at a consistently higher rate than in other regions. Allee effects, which act on low-density populations ahead of the moving population that contribute to gypsy moth spread, have also been observed to be consistently weaker in Wisconsin. Because a major cause of an Allee effect in the gypsy moth is mate-finding failure at low densities, supplementing low-density populations with immigrants that arrive through dispersal may facilitate establishment and consequent spread. We used local indicator of spatial autocorrelation methods to examine space-time gypsy moth monitoring data from 1996 to 2006 and identify isolated, low-density colonies that arrived through dispersal. We measured the distance of these colonies from the moving population front to show that long-distance dispersal was markedly present in earlier years when Wisconsin was still mainly uninfested. Recently, however, immigrants arriving through long-distance dispersal may no longer be detected because instead of invading uninfested areas, they are now supplementing high-density colonies. In contrast, we observed no temporal pattern in the distance between low-density colonies and the population front in West Virginia and Virginia. We submit that long-distance dispersal, perhaps facilitated through meteorological mechanisms, played an important role in the spread dynamics of the initial Wisconsin gypsy moth invasion, but it currently plays a lesser role because the portion of Wisconsin most susceptible to long-distance immigrants from alternate sources is now heavily infested.     

Effect of glucose and amino acids on expression of K99 antigen in Escherichia coli

K99 antigen production by enterotoxigenic Escherichia coli strains of bovine origin was investigated by slide agglutination and in vitro attachment to intestinal villi. Work with two strains (B41 and B44) showed that on minimal medium M2, K99 antigen was not repressed by a high concentration of glucose (2%, w/v). Growth on synthetic or complex medium did not affect K99 antigen detection, which was independent of capsular antigens, and its synthesis was not repressed by Casamino acids or glucose. A survey of 12 strains revealed two groups: in one group K99 antigen production was constitutive on basal medium without glucose, and in the second group K99 antigen was produced only in the presence of glucose. Immunoelectrophoresis patterns, and the results of slide agglutination and attachment tests, were dependent upon K99 type, whereas haemagglutination patterns were not.     

Climate suitability and management of the gypsy moth invasion into Canada

The gypsy moth has become established throughout southern Canada east of Lake Superior where the climate is suitable for the completion of its univoltine life cycle. The spread of the gypsy moth to the north and west in Canada has so far been prevented by climatic barriers and host plant availability as well as by aggressive eradication of incipient populations. Climate change is expected to increase the area of climatic suitability and result in greater overlap with susceptible forest types throughout Canada, especially in the west. At the same time, the gypsy moth is spreading west in the USA into states bordering western Canadian provinces. These circumstances all lead to a greatly increased risk of further invasion into Canadian forests by the gypsy moth. Management actions need to be intensified in different ways in different parts of the country to reduce the impacts of spread in eastern Canada and to prevent the gypsy moth from invading western regions.     

Isolation and characterization of an asthma-inducing sea-squirt antigen

An antigen capable of inducing asthmatic attack in patients with sea-squirt allergy has been isolated from body fluid of sea-squirt, Styela plicata, and designated as DIIIa. The chemical composition and the behavior in anion-exchange chromatography showed that the antigen was a weakly acidic glycoprotein. The weight-average molecular weight of DIIIa was estimated to be 9,880 by the sedimentation equilibrium method. This antigen was apparently discriminated from the previously isolated sea-squirt antigens, Gi-rep and Ei-M, by its activity to induce asthmatic attack and conjunctival congestion, though it gave a strongly positive reaction in skin tests against allergic patients, similarly to the previous antigens. From the cross-reaction of DIIIa to rabbit anti-Gi-rep and anti-Ei-M sera in vitro, it was confirmed that the antigen carried essentially the same antigenic determinant as Gi-rep and Ei-M, and this was termed type alpha determinant. Furthermore, the allergenic activities detectable in vivo and the reactivity to the rabbit antisera in vitro were simultaneously absorbed by immobilized immunoglobulin from anti-Gi-rep serum. Thus, it was suggested that the antigenic determinant responsible for the allergic reactions in vivo corresponded to that specified as type alpha in vitro on the basis of the reactivity against the rabbit anti-Gi-rep serum.     

Tumor-associated antigens are cytokine inducers and hyporeactivity factors to the immune system

We investigated possible mechanisms leading to the inhibition of theimmune system in people with chronic disorders. Tumor cell produce proteinreleased into the circulation, such as tumor associated antigens, may playan important role in processes preceding paralysis of the immune system. Totest this hypothesis the following tumor associated antigens were used: AFP,OFP, CA-125, CA-50 and CA-19-9. Their role was assessed by modulatingcytokine production in cord blood lymphocytes and peripheral white bloodcells obtained from grown population of patients treated with colostrinin,an cytokine inducer. PHA, LPS and colostrinin were used as positive controlin those essays. Each antigen tested individually induced IFN, TNFand IL-6 in dose dependent fashion. None of the tested cytokines werespontaneously released by the cells. Data generated from these experimentsindicated that tumor associated antigens are inducing type 1 cytokines insimilar fashion as LPS or colostrinin. However, lymphocytes taken frompatients undergoing therapy with colostrinin revealed progressive losscapability to produce type 1 cytokines as they did in case of colostrinin.The loss of the capability to respond to antigen may represent phenomenonleading to immune tolerance.     

Trichophytin Extraction: Biological Comparison of Trichophytin Extracted from Trichophyton mentagrophytes Grown in a Complex Medium and a Defined Medium

The extraction of antigens (trichophytin) from Trichophyton mentagrophytes var. asteroides grown in both a complex medium and a defined medium proved to be reproducible and feasible. These extracts were first evaluated by skin testing in both infected and noninfected guinea pigs. Further evaluation and standardization was accomplished in selected humans by comparing these antigens to a trichophytin known to elicit allergic reactions to the dermatophytes. The trichophytins produced by this extraction procedure gave no false positive (or negative) skin test reactions. These antigens did, however, reliably elicit allergic sensitivity of both the immediate and delayed type. The potency of both complex and defined media antigen was shown to be equal to that of the standard trichophytin.     

Interleukin-13 induces thymus and activation-regulated chemokine (CCL17) in human peripheral blood mononuclear cells

BACKGROUND: In allergic inflammation involving allergic rhinitis, the predominance of Th(2) lymphocytes is one of the primary causal agents in promotion of the allergic condition. Thymus and activation-regulated chemokine (TARC/CCL17) is a recently identified chemokine that induces the development of Th(2) lymphocytes. One of the sources of TARC has been reported to be peripheral blood mononuclear cells (PBMCs). OBJECTIVE: We investigated TARC production from PBMCs by the stimulation of specific antigens and Th(2) type cytokines. METHOD: PBMCs were isolated from both allergic rhinitis patients and healthy volunteers. PBMCs were incubated with cytokine. TARC mRNA expression was examined by real time PCR methods and the amount of TARC production was examined by ELISA. RESULTS: IL-13 was found to be the most potent inducer for TARC mRNA expression and protein production in PBMCs. Furthermore, tumour necrosis factor alpha and IL-13 synergistically induce TARC. The amount of TARC from allergic rhinitis patients was significantly larger than that from healthy volunteers. Moreover, TARC was induced by a specific antigen, and was 35% inhibited by an anti-IL-13 neutralizing antibody. CONCLUSION: These results indicate that IL-13 is important in TARC mediated Th(2) lymphocytes infiltration in the nasal mucosa.     

A Romani mitochondrial haplotype in England 500 years before their recorded arrival in Britain

The nomadic Romani (gypsy) people are known for their deep-rooted traditions, but most of their history is recorded from external sources. We find evidence for a Romani genetic lineage in England long before their recorded arrival there. The most likely explanations are that either the historical record is wrong, or that early liaisons between Norse and Romani people during their coincident presence in ninth to tenth century Byzantium led to the spread of the haplotype to England.     

Hyper‐reactive cloned mice generated by direct nuclear transfer of antigen‐specific CD4+ T cells

T‐cell receptor (TCR)‐transgenic mice have been employed for evaluating antigen‐response mechanisms, but their non‐endogenous TCR might induce immune response differently than the physiologically expressed TCR. Nuclear transfer cloning produces animals that retain the donor genotype in all tissues including germline and immune systems. Taking advantage of this feature, we generated cloned mice that carry endogenously rearranged TCR genes from antigen‐specific CD4⁺ T cells. We show that T cells of the cloned mice display distinct developmental pattern and antigen reactivity because of their endogenously pre‐rearranged TCRα (rTα) and TCRβ (rTβ) alleles. These alleles were transmitted to the offspring, allowing us to establish a set of mouse lines that show chronic‐type allergic phenotypes, that is, bronchial and nasal inflammation, upon local administrations of the corresponding antigens. Intriguingly, the existence of either rTα or rTβ is sufficient to induce in vivo hypersensitivity. These cloned mice expressing intrinsic promoter‐regulated antigen‐specific TCR are a unique animal model with allergic predisposition for investigating CD4⁺ T‐cell‐mediated pathogenesis and cellular commitment in immune diseases.     

How many signals are enough?

The many signals that control the progress of various immune responses to both foreign and self antigens can be divided into no less than three major groups. The first group is the initial positive stimulus, associated with activation events through antigen receptors and their associated proteins. These signals launch lymphocytes in their response to antigen, either foreign or self. The second group of signals is negative and involves various end products and interactions between cells, all recognizing antigen. These signals are endogenous to the reacting cell, or nearly so (two interacting cells from the same clone, daughter cells, which are in the same locale and bind to the same ligand). The third group (the prevention of end product feedback, involving various forms of antigen presentation, T cell contributions, rheumatoid factor activity, and other mechanisms) is more likely to occur with nonself antigens, which are temporally and spatially more restricted than self antigens. Experimental evidence for this immunological schema is summarized and clarified in its relationship to the Bretscher-Cohn theory of self-nonself recognition and to suppressor cell and idiotype-antiidiotypic theories.     

Comparison of methods for estimating the spread of a non-indigenous species

Aim  To compare different quantitative approaches for estimating rates of spread in the exotic species gypsy moth, Lymantria dispar L., using county-level presence/absence data and spatially extensive trapping grids.
Location  USA
Methods  We used county-level presence/absence records of the gypsy moth's distribution in the USA, which are available beginning in 1900, and extensive grids of pheromone-baited traps, which are available in selected areas beginning in 1981. We compared a regression approach and a boundary displacement approach for estimating gypsy moth spread based on these sources of data.
Results  We observed relative congruence between methods and data sources in estimating overall rates of gypsy moth spread through time, and among regions.
Main conclusions  The ability to estimate spread in exotic invasive species is a primary concern in management programmes and one for which there is a lack of information on the reliability of methods. Also, in most invading species, there is generally a lack of data to explore methods of estimating spread. Extensive data available on gypsy moth in the USA allowed for such a comparison. We show that, even with spatially crude records of presence/absence, overall rates of spread do not differ substantially from estimates obtained from the more costly deployment of extensive trapping grids. Moreover, these methods can also be applied to the general study of species distributional changes, such as range expansion or retraction, in response to climate change or other environmental effects.     

用免疫金颗粒标记鉴定舞毒蛾核型多角体病毒的抗原

用免疫电镜金颗粒标记技术准确、快速地对舞毒蛾(Lymantria dispar)核型多角体病毒抗原进行了定位和鉴定.舞毒娥病毒的多克隆抗体与同源的多角体抗原之间存在着强烈的亲和性,但与不同源的松柏锯角叶蜂(Neodiprion sertifer)病毒多角体抗原仅有极微弱的交叉反应.舞毒蛾病毒粒子和核衣壳抗原也能与同源的多克隆抗体作用.在被病毒感染的舞毒蛾脂肪体细胞核中,成熟的多角体被金颗粒重重标记,其外缘的游离病毒粒子和核衣壳亦被标记,但亲和力较弱.在被感染的脂肪体细胞核和质内发现一种与多角体蛋白晶体不同源的菱形结晶体.     

The Content of Antigens 1, 2 and 3 in Strains of Bordetella pertussis and in Vaccines

A comparison was made of the content of heat-labile antigens 1, 2 and 3 in strains of Bordetella pertussis isolated in Toronto, and the content of the same antigens in vaccines being used in that community. Antisera were prepared in rabbits and made monospecific by absorptions. Antigens were detected by slide agglutination tests. Of the 58 strains examined, 56 contained antigens 1 and 3 but no antigen 2 was detected. Nine of these strains were isolated from children who had received a full course of vaccination and had subsequently developed pertussis. All 10 vaccines examined contained antigen 1; two, possibly four, contained no antigen 2; and seven of the 10 vaccines were weak or deficient in antigen 3. This work suggests that improved protection might be attained by the use of vaccines containing adequate amounts of all three antigens.     

Persistence of invading gypsy moth populations in the United States

Exotic invasive species are a mounting threat to native biodiversity, and their effects are gaining more public attention as each new species is detected. Equally important are the dynamics of exotic invasives that are previously well established. While the literature reports many examples of the ability of a newly arrived exotic invader to persist prior to detection and population growth, we focused on the persistence dynamics of an established invader, the European gypsy moth (Lymantria dispar) in the United States. The spread of gypsy moth is largely thought to be the result of the growth and coalescence of isolated colonies in a transition zone ahead of the generally infested area. One important question is thus the ability of these isolated colonies to persist when subject to Allee effects and inimical stochastic events. We analyzed the US gypsy moth survey data and identified isolated colonies of gypsy moth using the local indicator of spatial autocorrelation. We then determined region-specific probabilities of colony persistence given the population abundance in the previous year and its relationship to a suite of ecological factors. We observed that colonies in Wisconsin, US, were significantly more likely to persist in the following year than in other geographic regions of the transition zone, and in all regions, the abundance of preferred host tree species and land use category did not appear to influence persistence. We propose that differences in region-specific rates of persistence may be attributed to Allee effects that are differentially expressed in space, and that the inclusion of geographically varying Allee effects into colony-invasion models may provide an improved paradigm for addressing the establishment and spread of gypsy moth and other invasive exotic species.     

Immunological tolerance using synthetic peptides--basic mechanisms and clinical application

Dysregulation of T lymphocyte function underpins the development of autoimmune and allergic diseases. These autoantigen-, or allergen-reactive pathogenic T cells are rare within the entire immune repertoire and it is therefore desirable to develop more specific therapies than are currently in use to directly target these cells and avoid adverse side effects. The obvious approach is to use the antigens that are recognized to impose a state of T cell tolerance. T cells recognize antigens as peptide fragments and we can therefore produce the relevant antigens as synthetic peptides. It has been known for many years that the decision of the T cell to mount a productive response (immunity) or to remain silent (tolerance) is controlled by the form in which the antigen is administered. Antigen with adjuvant leads to immunity, whereas soluble antigen without adjuvant leads to tolerance. This paradigm has been used successfully to induce tolerance with soluble peptides, preventing several animal models of autoimmune and allergic disease. These findings obviously have exciting potential for translation to human diseases. However, the basic immune mechanisms that lead to tolerance versus immunity are only beginning to be unravelled. The "effector" phase of tolerance also remains controversial with evidence for T cell death, anergy and the development of immunoregulatory function. This latter possibility of specifically generating autoantigen- or allergen-reactive regulatory T cells is particularly attractive. Here we review recent advances in our understanding of the requirements for tolerance induction and the potential for establishing dominant immune-regulation with peptide therapy.     

The filaggrin story: novel insights into skin-barrier function and disease

Recent reports have uncovered the key role of the protein filaggrin in maintaining an effective skin barrier against the external environment. Loss-of-function mutations in the profilaggrin gene (FLG) are common and are present in up to 10% of the population. These mutations are the cause of the semi-dominant skin-scaling disorder ichthyosis vulgaris and are a major risk factor for the development of atopic dermatitis. The discovery of these mutations also provides new data concerning the genetics of atopic asthma as well as intriguing insight into disease mechanisms of systemic allergies involving antigen exposure in skin with defective barrier function. Collectively, these novel findings have significant implications for the classification and future clinical management of patients with atopic and allergic diseases.     

Physico-chemical properties of microbial antigens and the immuno-allergic tests]

Experiments were conducted on guinea pigs sensitized by the delayed and immediate types of allergy. Different antigens obtained from the strain of Brucella abortus BA-19 were used for sensitization and the resolving action. Comparison of the resolving properties of the corpuscular, soluble (ultrasound treated) antigens and purified protein fractions, polysaccharide and RNA was carried out in the skin reactions of the immediate and delayed type, passive skin anaphylaxis, acute anaphylactic shock, and the Schults-Dale test. Immediate reactions to the purified protein fraction were weaker than those to the whole soluble antigen, by which the animals were sensitized. Polysaccharide and the RNA-fractions proved to be inactive in the allergic reactions.     

Breast milk-mediated transfer of an antigen induces tolerance and protection from allergic asthma

Allergic asthma is a chronic disease characterized by airway obstruction in response to allergen exposure. It results from an inappropriate T helper type 2 response to environmental airborne antigens and affects 300 million individuals. Its prevalence has increased markedly in recent decades, most probably as a result of changes in environmental factors. Exposure to environmental antigens during infancy is crucial to the development of asthma. Epidemiological studies on the relationship between breastfeeding and allergic diseases have reached conflicting results. Here, we have investigated whether the exposure of lactating mice to an airborne allergen affects asthma development in progeny. We found that airborne antigens were efficiently transferred from the mother to the neonate through milk and that tolerance induction did not require the transfer of immunoglobulins. Breastfeeding-induced tolerance relied on the presence of transforming growth factor (TGF)-beta during lactation, was mediated by regulatory CD4+ T lymphocytes and depended on TGF-beta signaling in T cells. In conclusion, breast milk-mediated transfer of an antigen to the neonate resulted in oral tolerance induction leading to antigen-specific protection from allergic airway disease. This study may pave the way for the design of new strategies to prevent the development of allergic diseases.