Cockroaches have been implicated as a cause of respiratory allergy in urban areas worldwide. IgE-reactive German cockroach proteins were identified with molecular weights (MWs) of 90, 66, 50, 43 and 36 KD by immunoblot analysis in both immune BALB/c mice and sensitized humans. Prominent IgE-reactive proteins were purified using FPLC by ion-exchange chromatography, gel filtration and hydrophobic chromatography. The N-terminal amino acid sequence of a purified protein with a MW of 66 KD on SDS-PAGE was Val-Thr-Leu-Lys-Lys(Val)-Met-Ile-Lys-Thr-Phe-Tyr. No homologous protein was found through a search of GenBank that indicated a novel IgE-reactive protein in German cockroach extract. Another purified protein with a MW of 36 KD reacted strongly with a monoclonal antibody against Bla g 2.
Amino Acid Sequence/genetics ; Animal ; Cockroaches/chemistry* ; Human ; IgE/immunology* ; Insect Proteins/isolation & purification* ; Insect Proteins/immunology* ; Insect Proteins/genetics ; Mice ; Mice, Inbred BALB C ; Tissue Extracts/chemistry*

We investigated the induction of resistance to Haemaphysalis longicornis infestation in rabbits that had been immunized with recombinant H. longicornis P27/30 protein. The success of immunological control methods is dependent upon the use of potential key antigens as tick vaccine candidates. Previously, we cloned a gene encoding 27 kDa and 30 kDa proteins (P27/30) of H. longicornis, and identified P27/30 as a troponin I-like protein. In this study, rabbits that were immunized with recombinant P27/30 expressed in Escherichia coli showed the statistically significant longer feeding duration for larval and adult ticks (P< 0.05), low engorgement rates in larval ticks (64.4%), and an apparent reduction in egg weights, which suggest that H. longicornis P27/30 protein is a potential candidate antigen for a tick vaccine. These results demonstrated that the recombinant P27/30 protein might be a useful vaccine candidate antigen for biological control of H. longicornis.
Animals ; Antibodies/blood ; Escherichia coli/genetics ; Female ; Gene Expression ; Insect Proteins/immunology ; Ixodidae/*immunology ; Microfilament Proteins/*immunology ; Rabbits ; Recombinant Proteins/*immunology ; Research Support, Non-U.S. Gov't ; Tick Infestations/*immunology/prevention & control

This study investigated whether elevated host immune capacity can inhibit T. gondii infection. For this purpose, we used silk protein extracted from Bombyx mori cocoons as a natural supplement to augment immune capacity. After silk protein administration to BALB/c mice for 6 weeks, ratios of T lymphocytes (CD4+ and CD8+ T-cells) and splenocyte proliferative capacities in response to Con A or T. gondii lysate antigen (TLA) were increased. Of various cytokines, which regulate immune systems, Th1 cytokines, such as IFN-gamma, IL-2, and IL-12, were obviously increased in splenocyte primary cell cultures. Furthermore, the survival of T. gondii (RH strain)-infected mice increased from 2 days to 5 or more days. In a state of immunosuppression induced by methylprednisolone acetate, silk protein-administered mice were resistant to reduction in T-lymphocyte (CD4+ and CD8+ T-cells) numbers and the splenocyte proliferative capacity induced by Con A or TLA with a statistical significance. Taken together, our results suggest that silk protein augments immune capacity in mice and the increased cellular immunity by silk protein administration increases host protection against acute T. gondii infection.
Animals ; Bombyx/*chemistry ; CD4-CD8 Ratio ; Cell Proliferation ; Cells, Cultured ; Cytokines/secretion ; Insect Proteins/*immunology ; Leukocytes, Mononuclear/immunology ; Male ; Mice ; Mice, Inbred BALB C ; Silk/immunology ; Spleen/immunology ; Survival Analysis ; Toxoplasma/*immunology/pathogenicity ; Toxoplasmosis, Animal/immunology/*prevention & control

Many allergists are currently focusing on the development of new diagnostic tools, and are attempting to improve both the sensitivity and specificity. A multiple allergen simultaneous test-chemiluminescent assay (MAST-CLA) is one of the most popular diagnostic tools used in the Republic of Korea. However, there remains controversy among allergists with regard to the cut-off point for a positive result. The present study was conducted in order to determine the validity of MAST-CLA as compared with that of the skin prick test, with particular emphasis on arthropod allergens, on the basis of percentage agreement rates and k-values, and also to suggest the optimal positive cutoff points using receiver operating characteristic (ROC) curves. The study was conducted with 97 subjects (54 men, 43 women). Optimal individual cut-off points were calculated as follows; class II for Dermatophagoides farinae, class I for Dermatophagoides pteronyssinus, and trace for a cockroach mix. These findings suggest that attempting to apply optimal individual cut-off points will be a good way of improving diagnostic tests, particularly MAST-CLA.
Adult ; Allergens/*immunology ; Animals ; Antigens, Dermatophagoides/*immunology ; Chemiluminescent Measurements/*methods/standards ; Cockroaches/chemistry ; Dermatophagoides farinae/chemistry ; Dermatophagoides pteronyssinus/chemistry ; Female ; Humans ; Hypersensitivity/*diagnosis/immunology ; Insect Proteins/*immunology ; Male ; *ROC Curve ; Skin Tests/methods

OBJECTIVETo transform the antimicrobial peptide fusion gene of cecropin B and rabbit NP-1(CN) into Houttuynia cordata to improve its antimicrobic capability.

METHODThe fusion gene of CN designed and synthesized artificially was recombined with expression vector pBI121. The recombined vector was transformed to Agrobacterium tumefaciens LBA4404, by which CN gene was transformed to the explants of H. cordata. The transgenic regeneration plantlets were selected by kanamycin and rapid screening PCR. The transgenic plants were identified by PCR-Southern of genomic DNA and RT-PCR. The disease resistances were detected by antibacterial zone trail of leaf extracts to E. coli K12 and infection by Rhizoctonia solani.

RESULTGene of interesting CN was inserted into genomic DNA and expressed in transformed H, cordata, whose resistance to E. coli K12 and Rh. solani was stronger than that of the non-transformed control.

CONCLUSIONThe fusion gene CN can improve antimicrobic capability of transformed H. cordata.

Animals ; Anti-Bacterial Agents ; immunology ; pharmacology ; C-Reactive Protein ; genetics ; metabolism ; pharmacology ; Houttuynia ; genetics ; immunology ; microbiology ; Immunity, Innate ; Insect Proteins ; genetics ; immunology ; pharmacology ; Nerve Tissue Proteins ; genetics ; metabolism ; pharmacology ; Plant Diseases ; immunology ; microbiology ; Plants, Genetically Modified ; genetics ; immunology ; microbiology ; Rabbits ; Recombinant Fusion Proteins ; genetics ; immunology ; pharmacology ; Rhizoctonia ; physiology ; Transformation, Genetic

Cockroaches have been recognized as a major cause of asthma. Bla g 4 is one of the most important German cockroach allergens. The aim of this study is to investigate IgE reactivity to the recombinant Bla g 4 (rBla g 4) in the sera of allergic patients and identify linear IgE binding epitope. For protein expression, full-length Bla g 4 (EF202172) was divided into 5 overlapping peptide fragments (E1: aa 1-100, E2: aa 34-77, E3: aa 74-117, E4: aa 114-156, and E5: aa 153-182). The full-length and 5 peptide fragments of Bla g 4 was generated by PCR and over-expressed in E. coli BL21 (DE3). The IgE binding reactivities of the full-length and peptide fragments were measured by ELISA using 32 serum samples of cockroach allergy. The sera of 8 patients (25%) reacted with rBla g 4. Four sera (100%) showed IgE-binding reactivity to full-length and peptide fragment 4, and 2 sera (50%) reacted with peptide fragment 2. One (20%) serum reacted with peptide fragment 3. The results of ELISA using overlapping recombinant fragments indicated that the epitope region was located at amino acid sequences 34-73 and 78-113, and major IgE epitope of Bla g 4 was located at amino acid sequences 118-152 of C-terminal. B-cell epitope analysis of German cockroach allergen Bla g 4 could contribute to the strategic development of more specific and potentially efficacious immunotherapy.
Adolescent ; Adult ; Allergens/chemistry/genetics/*immunology ; Amino Acid Sequence ; Animals ; Child ; Cockroaches/*immunology ; *Epitope Mapping ; Escherichia coli/genetics/metabolism ; Female ; Humans ; Hypersensitivity/*immunology ; Immunoglobulin E/*immunology ; Insect Proteins/chemistry/genetics/*immunology ; Male ; Middle Aged ; Molecular Sequence Data ; Sequence Alignment ; Young Adult