Federico Montealegre

Comparative prevalence of sensitization to common animal, plant and mould allergens in subjects with asthma, or atopic dermatitis and/or allergic rhinitis living in a tropical environment

Background and objectives Current information suggests that the expression of allergic diseases is determined by the exposure and nature of the allergen. The objectives of the present study were to determine if the nature of allergenic exposition to animal, plant or fungal allergens influenced the clinical manifestations of atopic dermatitis (AD), allergic rhinitis (AR) or asthma (AS) in patients living in a tropical environment. The prevalence and degree of sensitization to these allergens were analysed by age and gender.

Identification of Blomia tropicalis Allergen Blo t 5 by cDNA Cloning

Identification of Blomia tropicalis Allergen Blo t 5 by cDNA Cloning L.K. Karla Arruda E. Enrique Fernandez-Caldas C.K. Charles K. Naspitz F. Federico Montealegre L.D. Lisa D. Vailes M.D. Martin D. Chapman Asthma and Allergic Diseases Center, Department of Internal Medicine, University of Virginia Health Sciences Center, Charlottesville, Va., University of South Florida College of Medicine, Tampa, Fla., USA; Paulista School of Medicine, Sao Paulo, Brazil; and Ponce School of Medicine, Ponce, Puerto Rico

Cloning and expression of Blo t 1, a novel allergen from the dust mite Blomia tropicalis, homologous to cysteine proteases

Background House dust mite allergens have been shown to be a very important stimulus in the causation of asthma and triggers for the exacerbation of symptoms. Therefore, characterization of mite‐derived allergens at the molecular level is an important step for the development of effective diagnostic and therapeutic approaches, as well as for epidemiological studies.

Exposure Levels of Asthmatic Children to Allergens, Endotoxins, and Serine Proteases in a Tropical Environment

A cross‐sectional study was conducted in Bayamón, Puerto Rico, to identify and quantify indoor allergens, serine proteases, and bacterial endotoxin present in homes of asthmatic children. A total of 126 dust samples from houses were obtained from the entire mattress and bedside floor. Most of the patients had detectable levels of mite, cockroach, cat, and dog allergens. Mold allergens were found only in bedside floor dust samples. Mouse allergens were not detected. Forty‐two percent, 36.5%, and 1.8% of the patients demonstrated exposures to sensitizing levels of mite, Bla g 1 and cat allergens, respectively. The percentage of patients exposed to high levels of allergens capable of triggering asthma symptoms was 33.3% and 26.4% for mite and Bla g 1 allergens. Only dog allergen, bacterial endotoxin, elastase, and trypsin were associated with asthma symptoms. Eighty‐nine percent of the asthmatic children were exposed to endotoxin concentrations greater than 100 EU/mg dust, and more than half of the patients were exposed to high levels of serine proteases. Our study indicates that indoor concentrations of allergens traditionally associated with asthma symptoms and severity may not be applicable in tropical environments and highly ventilated households. In fact, in the study population, endotoxins, dog allergen, and serine proteases may play a dominant role in the induction of asthma symptoms.

Cloning and Molecular Characterization of a cDNA from Blomia tropicalis Homologous to Dust Mite Group 3 Allergens (Trypsin-Like Proteases)

Background: Exposure to the house dust mite Blomia tropicalis is increasingly being implicated as a major risk factor for asthma exacerbations in sensitized individuals. The objective of this study is to clone and characterize B. tropicalis allergens in order to better define their role in allergic asthma. Methods: A λgt22A cDNA library was constructed from B. tropicalis mRNA and screened using specific DNA probes. A full-length cDNA (Bt2-3) was isolated, subcloned, and sequenced. Results: Sequence analysis showed that clone Bt2-3 encodes the full-length serine protease preproenzyme from B. tropicalis. The predicted Bt2-3 protein consists of a 15-amino-acid signal peptide, a 20-amino-acid propeptide and a mature protein of 231 amino residues. BLAST analysis of the deduced amino acid sequence showed high homology (48–54%) between clone Bt2-3 and serine proteases (group 3 allergens) from domestic dust mites. Both the serine active site (DACQGDSGGPVA; amino acids 214–225) and the histidine active site (LTAAHC; amino acids 71–76) of serine proteases were highly conserved. The estimated molecular weight of the preproenzyme is 27.5 kD and its theoretical pI is 8.7. The mature protein contains a putative tyrosine kinase phosphorylation site (amino acids 24–31) and several N-myristoylation sites. Sequence alignment shows that the serine protease active sites are highly conserved among the clinically important mite species, including B. tropicalis. Conclusion: We report the cloning and molecular characterization of a B. tropicalis cDNA clone encoding a trypsin-like protease, with a possible major role as an allergen. Expression and further characterization of the recombinant product will help determine the role of proteolytic enzymes in the pathophysiology of allergic asthma.

Environmental Factors Precipitating Bronchial Asthma Exacerbations in Southern Puerto Rico: A Pilot Study

The purpose of the study was to correlate environmental factors that may play a role in triggering asthma attacks requiring admissions. A retrospective analysis of 320 asthmatic admissions from January 1987 to December 1989 was conducted in two community hospitals in our area. Information from the U.S. Weather Bureau pertaining to temperature, barometric pressure, wind velocity, and precipitation was analyzed. The readings from the Puerto Rican Environmental Agency for Total Suspended Particles (TSP-10) during that period were also reviewed. Our results indicate that the female patients accounted for the majority of the asthmatic admissions: 74.3% versus 26% in males. Eighty-four percent of the patients had a history of asthma attacks and 67% had previous hospitalizations due to asthma exacerbations. Correlation between asthma family history and age was found in only two age groups, 31-40 and 71-80 years. Rhinitis and asthma exacerbations were strongly linked only in the age group of 31-40 years. Active cigarette smoking was strongly associated with asthma exacerbations only in the age group of 61-70 years. Environmental data indicated that there was not a consistent pattern of monthly admissions; however, there was a significant association between admissions and increased precipitation, high counts of annual TSP-10, and decreased wind velocity. In view of our findings, asthmatics living on islands such as Puerto Rico might be affected by an additive effect of pollutants, by local and mainland emissions that are transported by the air currents, leading to a higher incidence of asthma exacerbations.

Detection of serine proteases in extracts of the domestic mite Blomia tropicalis.

Previous studies have shown that the domestic mites Dermatophagoides pteronyssinus and D. farinae contain allergens with serine protease activity. These proteolytic allergens include trypsin, chymotrypsin, elastase, kallikrein, and C3/C5 convertase. However, it is not known whether the domestic mite Blomia tropicalis shares with other mite species the serine protease activities. The enzymatic activity present in extracts obtained from food-free B. tropicalis was investigated using specific substrates and inhibitors. Based upon the concentration response and inhibition profiles, and the digestion of specific substrates our data demonstrate that extracts from B. tropicalis exhibit several serine-protease-like activities. The enzyme activities detected in the B. tropicalis extracts are trypsin, elastase, chymotrypsin, kallikrein, C3/C5 convertase, and mast cell protease. Our results also demonstrate that kallikrein and C3/C5 convertase-like activities were not significantly affected by the α1-antiprotease, a naturally occurring serine protease inhibitor which protects lung mucosa from the enzymatic action. These data strongly suggest that the Echymyopodidae mite B. tropicalis shares at least five serine proteases with members of other mite families, the Glycyphagidae and Pyroglyphidae. In addition, our data demonstrate the potential use of biochemical methods to detect serine proteases for evaluation of mite growth in vitro, or to detect environmental exposures to these enzymes.

Measuring asthma disparities in Hispanics: adherence to the national guidelines for asthma treatment in emergency departments in Puerto Rico

BACKGROUND Puerto Rico has the highest prevalence of asthma in the United States. Currently, there are no data on actual care given to asthmatic patients. OBJECTIVE To determine the prevalence of documented adherence to the 1997 National Asthma Education Prevention Program guidelines regarding care given in emergency departments (EDs) in Ponce, Puerto Rico. METHODS A case series was conducted using 6,002 ED records with a physician-based diagnosis of asthma for 1999 through 2001. RESULTS A history of asthma attack was documented in 82.0% of the cases and in all age groups. In-home beta-agonist use was recorded in only 5.7% of the medical records. Documentation of previous admissions to the ED and the intensive care unit were found in 3.5% and 0.33% of the records, respectively. Nocturnal symptoms before the ED visit were found in only 6.4% of the records, and asthma treatment at home was found in 39.9%. Accessory muscle retraction was documented in 99.1% of the cases, and oxygen saturation was found in 23.2%. Treatment with nebulized beta-agonist was found in 72.1% of the records, and intravenous or oral corticosteroid use was found in 84.1%. Follow-up appointments were detected in 64.8% of the cases, and referrals to specialists were given in only 5.3%. Rate ratios between our data and those of other researchers indicate that there are geographical differences in compliance with the guidelines. CONCLUSION Of the variables tested, only one had acceptable levels of compliance, as evidenced in the patients records, indicating that there are alarming differences in ED evaluation and treatment compared with the 1997 National Asthma Education Prevention Program guidelines.

Use of Recombinant Group 5 Allergens to Investigate IgE-Mediated Sensitization to Blomia Tropicalis and Dermatophagoides Pteronyssinus

In tropical and subtropical areas of the world, mites of the genus Blomia have been recognized as important cause of sensitization and development of asthma. These non-pyroglyphidae mites belong to the Glycyphagidae family, and are currently classified among the “domestic mites”1. IgE antibody responses to Blomia, including B. tropicalis and B. kulagini, have been documented by skin testing and serologie studies2–7. Exposure to B. tropicalis has been reported in houses from different geographic areas, including Hong Kong, Brazil, Colombia, Venezuela, India, Taiwan, Spain, Egypt and other countries1–8. In the United States, B. tropicalis has been found in significant numbers in homes from Tampa FL, New Orleans LA, Memphis TN, Galveston TX, Delray Beach and San Diego CA9,10. In these areas, B. tropicalis was the fourth most frequent mite species found in house dust, in addition to Dermatophagoides pteronyssinus, D. farinae and Euroglyphusmaynei

850 Immunoblot analysis and comparative Ige responses of atopic patients to extracts of the domestic mite Blomia tropicalis

BACKGROUND The domestic mite Blomia tropicalis is found in subtropical and tropical environments, and its clinical importance as a sensitizing agent in allergic disease is widely accepted. OBJECTIVE To investigate the IgE reactivity to allergens present in extracts of the domestic mite B. tropicalis, and compare the IgE responses to these allergens by asthmatics, patients with atopic dermatitis and allergic rhinitis, as well as nonatopic controls. METHODS Extracts from B. tropicalis were used for skin tests. The B. tropicalis specific IgE in the serum were measured using the FAST Plus Test and immunoblot analysis. RESULTS A total of 199 volunteers participated in the study. The data show that 18 out of 29 polypeptide bands present in extracts of this mite species were recognized by the allergic and control sera. Of these allergens, four showed a high IgE binding frequency and had relative molecular weights of 104, 80, 68 and 14 kDa. The 14 kDa allergen demonstrated the highest IgE binding frequency. CONCLUSION Sera from atopic patients reacted to more allergens than sera from patients controls. Extracts from pure bodies of B. tropicalis contain one immunodominant and three important allergens. A common characteristic between all of the sera tested was the high degree of serum IgE reactivity observed to the 14 kDa allergen.