D. Pteronyssinus Research Articles

Immunoglobulin G1 subclass responses can be used to detect specific allergy to the house dust mites Dermatophagoides farinae and Dermatophagoides pteronyssinus in atopic dogs

BackgroundIn dogs with atopic dermatitis, intradermal testing (IDT) or allergen specific IgE serological testing are routinely employed to identify causative allergens. These allergens can then be used for allergen-specific immunotherapy and allergy management. The clinical relevance of this testing is affected by the source of allergen, and other biomarkers that are more related to specific allergens still need to be identified. The aim of this study was to investigate levels of specific IgE, total IgG, and IgG1 and IgG2 subclasses against the local house dust mites (HDM) Dermatophagoides farinae (DF) and D. pteronyssinus (DP) as biomarkers by using in-house ELISAs in healthy (n = 33) and atopic dogs (AD) (n = 44) that were either positive or negative by IDT to HDM.ResultsBeing over 3 years of age was a risk factor for AD (Odds Ratio (OD) = 4.10, 95% Confidence interval (CI) 1.57–10.75, p = 0.0049), but there was no relation to IDT outcomes (OR = 0.9091, 95% CI 0.22–3.74, p = 1.00). High levels of all antibody isotypes (IgE, IgG, IgG1 and IgG2) against HDM were found in aged healthy dogs (> 3 years old). In AD, HDM-IgE and IgG1 levels were higher in dogs that were IDT positive to HDM than in IDT negative animals. Levels of IgE and IgG1 could be used to distinguish the specific allergens, whereas total IgG and IgG2 levels were not different between IDT-positive and IDT-negative AD. By the receiver operating characteristic curve at a false-positive rate = 0.10, both IgE and IgG1 showed better sensitivity than IgG and IgG2. Similar to IgE, serum IgG1 concentration was also relevant to IDT outcomes.ConclusionsOur in-house ELISAs coated with local HDM were useful for evaluating antibody levels, and we propose use of the HDM-specific IgG1 subclass as a biomarker to detect HDM specific allergens in AD, potentially together with an IgE based platform.

Dual systemic allergic rhinitis and local allergic rhinitis

Methods Twenty-nine patients with seasonal SAR (positive SPT to grass and/or olive pollen) with well-defined symptoms during pollen season in addition to nasal symptoms throughout the year were included. Clinical questionnaires, skin tests, serum specific IgE, nasal levels of tryptase and eosinophil cationic protein, and nasal allergen provocation test (NAPT) with grass, olive, D. pteronyssinus (DP), and Alternaria alternata (AA) were performed.

Role Of Basophil Activation Test For Identifying Subjects With Local Allergic Rhinitis

A previous pilot study showed the presence of allergen-specific IgE in the surface of peripheral basophils in 50% of subjects with confirmed local allergic rhinitis (LAR). However, this phenomenon was investigated in a small number of patients. The aim of this study is confirming the presence of specific allergen activation of basophils, by means of basophil activation test (BAT) in a large population of LAR subjects sensitized to D. pteronyssinus (DP).

Subjects with local allergic rhinitis can be identified by basophil activation test

Background Local allergic rhinitis (LAR) is characterized by negative skin testing and serum specific IgE. Diagnosis is based on nasal provocation test (NPT), very sensitive but time-consuming, and/or presence of local synthesis of specific IgE (sIgE) which shows low sensitivity (22%). The aim of the study was to evaluate the presence of specific D. pteronyssinus (DP) activation of basophils by basophil activation test (BAT) in subjects with confirmed LAR. Methods BAT was performed in 43 subjects: 16 with confirmed LAR (positive NPT with DP, negative skin testing and sIgE to DP), 13 with allergic rhinitis (AR)(positive NPT with DP, positive skin testing and sIgE to DP) and 14 healthy controls (negative NPT, negative skin testing and sIgE to DP). To demonstrate a specific IgE mechanism of basophil activation, BAT with wortmannin pre-treatment was performed in four LAR patients. Results BAT with DP was positive in 85% of AR patients and 50% of LAR subjects. BAT showed a substantial correlation with NPT in AR subjects (kappa index: 0.78, p=0.0001). The positive responses of the 4 LAR samples that underwent BAT with wortmannin became negative when this substance was added to the assay. Conclusions

Positive Bronchial Responses To D. Pteronyssinus In Subjects With Confirmed Local Allergic Rhinitis

Local allergic rhinitis (LAR) is characterized by positive responses to nasal challenge with allergens in absence of systemic atopy. Subjects with LAR frequently show other co-morbidities including asthma. The possible role of allergens in the bronchial symptoms of subjects with LAR has not been addressed. Twenty subjects with confirmed LAR with D. Pteronyssinus (DP) and asthmatic symptoms (LARA), 20 subjects with allergic rhinitis with DP and asthmatic symptoms (ARA) and 10 healthy subjects as control group (CG) were recruited. Bronchial challenge was performed with DP extract at 4 μg/ml. Metacholine challenge and induced sputum were performed prior to and 24 hours after challenge. Cell populations and Th1/Th2 cytokines in sputum were evaluated by flow cytometry, and ECP and triptase in sputum by CAP method. Forty percent of LARA and all ARA subjects had positive responses to bronchial challenge with DP. PC20 values decreased in both groups 24 hours after challenge (p<0.05). Sputum ECP values also increased after challenge in both LARA and ARA groups (16.8±14.5 vs. 20.1±21 and 52±69 vs, 95±98, respectively). Also, an increase in sputum eosinophils was observed (LARA: 6.9±11 vs. 14.2±7.3, ARA: 3.7±3 vs. 7.2±6). LARA subjects had higher values of IL5,IL8, IL10, IL12 and IFN-gamma at baseline compared to ARA and controls, but no significant variations after challenge. Positive responses to D. Pteronyssinus were observed in the lower airways of subjects with clear history of asthmatic symptoms. Similar methacholine responses and increases in ECP and eosinophil levels were observed in both LARA and ARA groups.

Toll-like receptor 2 is associated with house dust mite-induced inflammatory response in human bronchial epithelial cells (91.1)

Abstract Exposure of the respiratory tract of an atopic individual to allergens such as D. pteronyssinus (DP) house dust mite is associated with the development of allergic asthma. In the study, we explored the role of Toll-like receptors (TLRs) in allergen-induced inflammatory responses of human bronchial epithelial cells. Our results showed that DP induced production of G-CSF, GM-CSF, CXCL1, IL-6, CXCL8, and CCL2 in BEAS-2B (immortalized human bronchial epithelial cells) in a dose-dependent manner. An inhibitory peptide against MyD88 significantly inhibited the allergen-induced cytokine response, while monoclonal antibodies (mAbs) against TLR2 showed some (but not significant) inhibitory effects. Analysis of mRNA transcripts by qPCR revealed an increased expression of MyD88 at 1 hr post-stimulation with DP. DP stimulation also induced increased expression of TLR1, TLR2, TLR3, TLR5, TLR8, and TLR9, as well as IFNβ. HKLM, a TLR2 agonist, along with DP showed a synergistic effect on cytokine response of BEAS-2B and the response was inhibited by anti-TLR2 mAb. HEK293 cells expressing TLR1/2, TLR2/6 or TLR4/CD14/MD2 showed increased cytokine response to allergens, while anti-TLR2 inhibited the response in HEK293 cells expressing TLR1/2 or TLR2/6. These data demonstrate a link between DP allergen-induced inflammatory response and TLR2-mediated signaling in airway epithelial cells and contribute to our understanding of the role of innate immunity in allergic diseases.

P‐44 The presence of dust mite species in the environment of dust mite‐sensitized atopic dogs

Dust mites (DM), especially Dermatophagoides farinae (DF), are the most common offending aeroallergens in atopic dogs. The aim of this study was to compare the DM species in the households of atopic dogs (Group A, n = 8) that reacted intradermally to DF, D. pteronyssinus (DP), Acarus siro (AS), Lepidoglyphus destructor (LD) and/or Tyrophagus putrescentiae (TP), to DM species in households of nonatopic dogs (Group B, n = 4) and households without pets (Group C, n = 8). Group A dogs exhibited perennial pruritus, were free of pathogenic mites and fleas, did not respond to an elimination diet, and fulfilled the diagnostic criteria for atopic dermatitis. All Group B dogs tested ID negative and had no dermatological problems. Dust samples were vacuum collected in a standardized fashion from two sites of the dog living quarters, one time for each season. DM quantitation was made stereoscopically and their species identification by microscopy. DM median number in Group A was 1.5 for DF, 0.5 for DP and 0 for the other three species. One of eight, one of four and one of four dogs were sensitized to DF, AS and TP, respectively, despite the absence of these species from their environment. No difference could be found in terms of DM species between the three groups (lowest P value = 0.06). In conclusion, cross‐sensitization between DM or irritant intradermal test reactions to their allergenic extracts may be common, and no preferential overpopulation of DF can explain the frequent sensitization of atopic dogs to this mite. Funding: Self‐funded.

Immunological findings in 3 dogs clinically diagnosed with allergic rhinitis.

Three dogs clinically diagnosed with allergic rhinitis (AR) were examined for their immunological findings. House dust mites (HDM) such as Dermatophagoides farinae (DF) and D. pteronyssinus (DP) were identified as positive allergens in the 3 dogs with both intradermal skin test and serum antigen-specific IgE test. Lymphocyte blastogenic response of peripheral blood mononuclear cells (PBMCs) under stimulation with DF antigen in dogs with AR was higher than that in 4 healthy control dogs. Expression level of IL-4 mRNA in PBMCs obtained from the 3 AR dogs was higher than that in PBMCs obtained from 4 healthy control dogs before and after stimulation with DF antigen. Expression level of IFN-gamma mRNA in PBMCs was not different between the AR and control dogs before and after stimulation with DF antigen. These results suggested that allergic reaction to HDM antigen and T(H)2-type immune response were associated with the development of AR in 3 dogs examined in this study.

Comparative incidence of skin reactivity to D. pteronyssinus (DP) and D. farinae (DF) in the San Francisco Bay Area

Comparative incidence of skin reactivity to D. pteronyssinus (DP) and D. farinae (DF) in the San Francisco Bay Area

HLA‐DRB1*07 may have a susceptibility and DRB1*04 a protective effect upon the development of a sensitization to house dust mite Dermatophagoides pteronyssinus

IgE responses to house dust mite-derived allergens seem to be the most important in the development of atopic asthma and rhinitis, but it has been difficult to demonstrate genetic control of the IgE response to the allergens. This study was undertaken to investigate the association between sensitization to house dust mite, D. pteronyssinus (DP), and genotypes of HLA-DRB1 alleles. DNAs were extracted from two groups of unrelated Koreans: (1) 178 with sensitization to DP; and (2) 99 age-matched non-atopic controls. Genotypes of the HLA-DRB1 alleles were determined using polymerase chain reaction (PCR)-based methods. The frequency of HLA-DRB1*07 was significantly increased in the DP-sensitive subjects compared with the controls (15.7% vs 4.0%, P = 0.009). Conversely, the frequencies of HLA-DRB1*04 and *14 were decreased in the DP-sensitive subjects compared with the controls (27.5% vs 45.5%, P = 0.002; 13.5% vs 24.2%, P = 0.02). Of the DRB1*04 alleles, DRB1*0403 was significantly decreased in the DP-sensitive subjects compared with the controls (3.9% vs 13.1%, P = 0.005). No significant differences were found in the distributions of the other HLA-DRB1 alleles between the two groups. HLA-DRB1*07 may have a susceptibility, and DRB1*04 and DRB1*14 may have a protective effect, upon the development of a sensitization to DP.

Serum antibody to Sarcoptes scabiei and house dust mite prior to and during infestation with S. scabiei

In this study, serum antibodies to Sarcoptes scabiei var. canis (SS), Dermatophagoides farinae (DF), and D. pteronyssinus (DP) were determined in 19 healthy, random-source dogs prior to infestation with scabies then again during a primary infestation, cure and challenge infestation with scabies. Prior to scabies infestation, serum of 11 dogs contained faintly detectable amounts of IgE and/or IgG to proteins in SS extract, probably resulting from sensitization to dust mites that share cross-reactive antigenic epitopes with SS. After becoming infested with scabies, the response to SS antigens became stronger with antibodies appearing to more antigens as the scabies infestation progressed. Three of the newly recognized proteins were 170, 155 and 142/133 kD and could be used in a diagnostic test since antibodies to them appeared during the primary infestation. In addition, during the primary infestation, 14 of 15 dogs developed IgE to 1–11 new SS proteins in addition to an increase in IgE binding to those proteins recognized prior to infestation. Overall, the strongest antibody responses (IgE and IgG) were exhibited during cure of the first infestation, when dead mites were still present in the stratum corneum. As expected, the antibody response was strong and rapid during challenge when the infestation self-cured. The immunogenic SS proteins identified by serum antibody binding during challenge, when the hosts self-cured, are candidates for inclusion in a vaccine. These candidate proteins are 200, 185, 170, 155, 142/133, 112, 97, 74, 57, 45/42, 32 and 22 kD. Some of the proteins in SS that exhibited new or increased antibody binding during the experiment also had IgE and IgG binding to proteins with similar molecular weights in DF and DP extracts. These results illustrate the difficulties involved in understanding and interpreting serum antibody for developing a serological test for the diagnosis of scabies, isolating relevant SS antigens that could be included in a vaccine for prevention of scabies, and for understanding the immune response mechanism to scabies.

Cross-reactivity of Tyrophagus putrescentiae with Dermatophagoides farinae and Dermatophagoides pteronyssinus in urban areas

Tyrophagus putrescentiae (TP) have cohabited with D. pteronys-sinus (DP) and D. farinae (DF) in more than 25% of houses in urban areas of Korea, and many atopic subjects have also been cosensitized to TP and Dermatophagoides species. We evaluated the cross-reactivity of TP with DF and DP in atopic subjects of urban inhabitants. The cross-reactivity was evaluated with inhibition ELISA and immunoblotting. Allergenic components of TP were evaluated with IgE immunoblotting of the sera from 25 individual atopics. All enrolled subjects lived in urban areas. In ELISA inhibition with pooled sera, all TP, DP, and DF extract inhibited TP-specific IgE by more than 90%, and the 50% inhibitory concentrations of TP, DP, and DF extract were 0.4 microg/mL, 0.8 microg/mL and 0.8 microg/mL, respectively. The maximum inhibition, however, of DP-specific and DF-specific IgE by TP extracts was 32% and 29%, respectively. With six individual sera, the TP-specific IgE was also inhibited by more than 88% with DF extract in all cases. In inhibition immunoblotting, all of the TP, DP, and DF extracts completely inhibited the TP-specific IgE bands at a concentration of 2.0 microg/mL. Fifteen allergenic components in TP were found. Among them, the 16-kD allergen was most prevalent (52%) and its IgE binding was completely inhibited by 0.1 microg/mL of purified Der f2 and it also bound with 2 different monoclonal antibodies to the group 2 allergen of Dermatophagoides species. Our results suggested considerable cross-reactivity between TP and the two Dermatophagoides species in urban areas where TP and Dermatophagoides species cohabit. The 16-kD allergen, which shared common epitopes with the group 2 allergen of Dermatophagoides, is one of the most prevalent allergens of TP.

IgE sensitivity and cross-reactivity to crude and purified mite allergens (Der f 1, Der f 2, Der p 1, Der p 2) in atopic dogs sensitive to Dermatophagoides mite allergens

The present study investigates IgE-reactivity to crude and purified mite allergens by intradermal skin test (IDST), Immunodot method, and ELISA in atopic dogs sensitive to mite allergens, as well as the allergenic cross-reactivity between Dermatophgoides (D) farinae (DF) and D. pteronyssinus (DP) in dogs by IgE-ELISA inhibition. IDST and Immunodot method for crude mite allergens were performed for atopic dogs and 16 atopic dogs showed sensitivity to mite allergens. Of the 16 dogs, all dogs had anti-DF IgE and 11 had anti-DP IgE. We measured specific IgE to purified major allergens (Der f 1, Der f 2, Der p 1, Der p 2). Of the 16 atopic dogs, six had anti-Der f 1 IgE and seven had anti-Der f 2 IgE. Similarly, of the 16 dogs, six had anti-Der p 1 IgE and seven had anti-Der p 2 IgE. However, eight dogs had no specific IgE to these mite allergens. These dogs may be sensitive to other major mite allergens except Der 1 and Der 2. In the dogs that had both anti-DF and DP IgE, IgE binding to DF was greatly inhibited by DP, and reciprocal inhibition was observed. Based on these data, it appears that there is a strong cross-reactivity between DF and DP in dogs. Similarly, a cross-reactivity between DF and DP in purified allergens was also observed. IDST and Immunodot method are useful methods for the diagnosis of atopic diseases in dogs, and ELISA is a useful method for further investigation of IgE-reactivity for the allergens.

Fauna and geographical distribution of house dust mites in Korea.

During the period of 1933-1994, house dusts were collected from 65 homes at 10 different localities by operating electric vacuum cleaners. House dust mites were isolated from 10 g dust by applying the modified wet sieving method. Total 7,257 mites were collected and 23 species were identified. Among them. Dermatophagoides farinae (DF) was predominant (65.3% of the total), followed by D. pteronyssinus (DP) (20.6%) and Tyrophagus putrescentiae (TP) (6.5%). Rhizoglyphus robini. Sancassania phyllophagianus, Cheyletus traussarti and Scheloribates latipes were the first findings from Korea. DF was predominant in Seoul (66.8%). Kwangju (63.6%), inland of Pusan (79.6%), Inchon (96.5%). Taejon (83.9%), Chonju (87.15) and Chongju (95.2%), whereas DP was predominant in Yongkwang-ub (72.5%) and Yongdo (island) of Pusan (64.9%), and TP in Chunchon (38.2%). The localities where DP and TP were predominant showed higher relative humidity in air (> 73% RH). Among 62 study homes, DF, DP and TP were found in 24.6% of the homes, co-habitat of two species in 48.1% and one species in 27.3%. DF was predominant in 63.5% of the homes studied. DP in 29.6% and TP in 6.9%. In 10 g of the house dust, less than 99 mites were found in 49 homes (70.0%), 100-499 mites in 11 homes (15.7%). 500-999 mites in 3 homes (4.3%) and more than 1,000 mites in 2 homes (2.9%). No mite was found in 5 homes (7.1%). In order to evaluate environmental factors affecting the population density of house dust mites, house type, age of house construction, size of the house, number of the family and frequency of the cleaning were compared with the number of mites, and none of the above factors were statistically correlated with the mite density.

Allergenicity of Euroglyphus maynei and its cross-reactivity with Dermatophagoides species

Background: The house dust mite, Euroglyphus maynei (EM), is common in homes in England, Europe the southern United States, and other parts of the world. It is often present in densities greater than 100 mites per gram of dust. EM usually occurs with Dermatophagoides farinae (DF) and D. pteronyssinus (DP) and it is frequently more abundant than Dermatophagoides species. Therefore the allergenicity of EM and the cross-reactivity between EM and Dermatophagoides species are important considerations for diagnosis of mite-induced allergy and the use of appropriate immunotherapy. Methods: Crossed immunoelectrophoresis revealed that EM was the source of at least 33 antigens. Crossed radioimmunoelectrophorests with 32 different sera from patients with RAST results that were positive to EM and skin test results that were positive to both DF and DP identified 15 allergens. Individual sera recognized two to eight EM antigens as allergens. Results: Ninety-one percent of the patients showed serum IgE directed at three or more allergens. Eighty-four percent of the patients had moderate or strong levels of IgE directed at antigen no. 33. All 15 allergens showed moderate or strong IgE binding by one or more sera. EM shared four and six cross-reacting allergens with DF and DP, respectively. Therefore 11 and 9 allergens of EM were species-specific and not shared by DF and DP, respectively. Conclusions: The results of this study clearly indicate that sensitivity to EM should be considered in geographic areas where this mite is abundant in homes.

QUANTITATIVE MEASUREMENTS OF SKIN PRICK TESTS (SPT) AND SPECIFIC IGE - RELATION BETWEEN VALUES AND DIFFERENT RESPIRATORY ALLERGIC DISEASES

Diagnosis of allergy is supported by “in vivo” (ex: SPT) and “in vitro” methods (ex: specific IgE). SPT is influenciated by a lot of variables, so standardization is essential to obtain precise and reproducible quantitative results. This can be a achieved by a method ol reading using a computerized digitalized graphics tablet with CAD software, created and validated by the authors and published elsewhere. The Pharmacia CAP System allows precise quantification of specific IgE and is more sensitive than RAST. Preliminar data obtained by the authors, showed that for D. Pteronyssinus (D.Pt) the SPT results done with extracts from Bencard (B) and Merck (M) had good correlation with specific IgE determined by CAP System (as quantitative values and as +/-).The aim of the study was to establish ihe possible relations between quantitative results of SPT and CAP and different respiratory allergic diseases with the same severity in a pediatric population sensitized to D.Pt. We include 73 children (47 ♂ 26 ♀,) aged 4 to 15 y. (mean age ± SD:9.70 ± 3.24), 13 with Rhinitis (R), 16 with Asthma (A) and 44 with A+R. The SPT was done with a standardized lancet (DHS) using cut oil +/- 7mm2, with Ihe method cited above and with extracts for D. PI from B and M. The determination of specific IgE was done by Pharmacia CAP System.Results: 1) Mean age ± SD in R group : 11.31 ± 2.72; A group: 9.48 ± 3.50, without statistically significant dilferencies. 2) SPT results (mean areas ± SD): R group (B:43.20 ± 20.69 mm2; M: 36.72 ± 22.18mm2); A group (B:50.48 ± 25.11mm2; M: 50.70 ± 35.24mm2); A+R group (B: 61.93 ± 28.20mm2; M: 47.98 ± 39.34mm2). 3) CAP results (mean values ± SD): R group (13.92 ± 14.44KU/L; A group (27.01 ± 30.26KU/L); A+R group (39.12 ± 29.35KU/L).Conclusions: 1) The results of quantitative SPT for both extracts related lo R, A and A+R don't show statistically signilicant ditterencies. 2) The quantitative values of specilic IgE show that there are differencies between all the entities, with lower results to R and higher to A+R. 3) Based on previous data suggesting that within the same disease, the severity is directed related with higher titers of specific IgE, our data may support that the “severity” of A+R > A > R.

Prevalence of dust mites in the homes of people with asthma living in eight different geographic areas of the United States

The density and species prevalence of dust mites were determined at various times over a 5-year-period in 252 homes of dust mite sensitive people with asthma who lived in eight geographic areas of the United States (Cincinnati, Ohio; New Orleans, La.; Memphis, Tenn.; Galveston, Texas; Greenville, N.C.; Delray Beach, Fla.; San Diego and Los Angeles, Calif.). The most common dust mites found in the homes were Dermatophagoides farinae (DF), D. pteronyssinus (DP), Euroglyphus maynei (EM), and Blomia tropicalis. All homes in all locations contained Dermatophagoides spp. mites, but few homes were populated exclusively by either DF or DP alone. Most homes (81.7%) were coinhabitated by both DF and DP. In coinhabited homes one species was predominant and usually made up at least 75% of the total mite population. Prevalence of the dominant or only species present varied between homes within a geographic area. EM occurred in significant numbers in 35.7% of homes in New Orleans, Memphis, Galveston, Delray Beach, and San Diego. Blomia tropicalis occurred in these same cities but in low densities. For all dust samples, only 13 homes of the 252 sampled had 100 or fewer mites/gm dust, which is considered to be the threshold for sensitivity. Most homes had average mite densities of 500 or more mites/gm dust. The results of the present study suggest a significant and widespread occurrence of both DF and DP. Therefore extracts of both mite species should be considered for diagnostic tests and immunotherapy. Significant levels of EM were present in some areas. Thus sensitivity to EM should be considered in these areas.