Donald H. Beezhold

Latex allergy can induce clinical reactions to specific foods.

Objective The purpose of this study was to investigate crossreactivity between latex and foods, to identify crossreacting IgE binding proteins, and to assess the clinical significance.

Allergy to latex rubber.

Latex gloves are indispensable in todays health care practice. The comfort, barrier, and tactile properties of powdered latex gloves have been thought to be ideal. Between 1988 and 1992, an estimated 11.8 billion examination gloves and 1.8 billion surgical latex gloves were used in the United States. Recent reports of allergic responses to latex have led some authors to re-examine the safety of latex medical devices. Although many health care workers report allergic reactions to gloves, most are not serious. The predominant immunologic response (82%) to natural rubber latex is type IV delayed hypersensitivity to rubber additives, which presents clinically as contact dermatitis [1]. However, between 1988 and 1992, the FDA received reports of more than 1000 systemic allergic reactions to latex, 15 of which were fatal [2]. These systemic reactions are type I immunologic responses that are mediated by IgE antibody to residual rubber tree proteins in latex medical devices. Health care workers develop sensitization from regular latex exposure: wearing latex gloves or inhaling aerosolized latex in the workplace [3]. Re-exposure in a sensitized patient may result in contact urticaria, allergic rhinoconjunctivitis, asthma, or anaphylaxis [4]. Epidemiology There are several explanations for the recent epidemic of latex allergy. After the introduction of universal precautions for human immunodeficiency virus, use of natural rubber latex gloves increased. The increased demand for gloves may have temporarily changed manufacturing procedures, resulting in a poor-quality, highly allergenic product. Increased awareness of latex allergy has resulted in more numerous reports [5] of this allergy during the last 6 years. Presently, the incidence of latex sensitization is unknown. The prevalence of latex allergy in the general nonatopic population is believed to be less than 1%. Latex sensitivity has been reported in 1 of 800 patients (0.125%) before surgery [6]. However, the prevalence of latex allergy in children with spina bifida ranges from 28% to 67% [5]. Mucosal absorption of latex allergens during multiple surgical procedures done early in life may have sensitized these children. The prevalence of latex allergy in health care workers is between 7% and 10% [7, 8]. Health care workers with atopic allergy have been reported [8] to have a 24% prevalence of a positive result from latex skin-prick testing. Fifty percent (5 of 10) of patients in this study [8] were clinically asymptomatic. The risk for subsequent development of latex allergy in this group is unknown. Eczematous hand dermatitis disrupts the skin barrier and may predispose persons to latex allergy. A progression may occur from localized (hand) to generalized (anaphylactic) allergic responses [9]. Areas with significant airborne latex allergens (operating rooms, intensive care units, and dental suites) may sensitize health care workers who inhale allergenic proteins [10-12]. Clinical Latex Allergy Irritant contact dermatitis, the most common clinical manifestation of latex allergy, is a nonallergic cutaneous response that manifests as dry, crusted lesions in latex glove-exposed areas. Prolonged and repeated latex exposure is aggravated by sweating and rubbing under the glove, leading to papular and ulcerative lesions [1]. Allergic contact dermatitis is a delayed hypersensitivity reaction to rubber additives (thiurams, mercaptobenzothiazole, carbamates) [13]. The acute phase of the reaction occurs 48 to 96 hours after exposure, affects the dorsum of the hands, and is characterized by vesicular skin lesions. With continued latex exposure, these skin lesions develop a crusted, thickened appearance. Immunoglobulin E-mediated latex allergy may be locally visible as contact urticaria [14] or may present as occupational rhinoconjunctivitis or asthma [3, 12]. Anaphylactic reactions have most often been caused by exposure to the surgeons latex gloves during abdominal or genitourinary surgery or by other sources of mucosal exposure to latex (barium enema, dental procedures) [3-5]. Anaphylaxis has been reported less often from latex exposure that occurs when health care workers put on gloves or work in a latex-laden environment. Identification and Management of Latex Allergy The prevention of adverse latex reactions depends on identification of patients who are allergic. A careful and complete history will not identify all persons at risk for latex allergy. The latex skin-prick test is a sensitive indicator of IgE sensitization, but a standardized latex extract that can be used in this test is presently not available. Latex skin-prick testing of extremely allergic persons using commercially available extracts has been safe [3, 6-8]. However, five [15, 16] anaphylactic episodes occurring after the use of extracts prepared from latex gloves or after the use of certain skin test reagents have been reported. Extracts prepared from latex gloves have variable allergenic potency, and skin-prick testing appears to have an increased risk for adverse reactions. Latex skin-prick tests should therefore be done cautiously starting with very diluted (1:1 million) extracts of the stock testing solution. This should only be done in an allergy center familiar with the test. Full emergency equipment must be available to treat possible systemic reactions. In vitro testing for latex protein-specific IgE antibodies (using radioallergosorbent tests, enzyme-linked immunosorbent assays, and Western blots) reportedly identifies 50% to 60% of persons with IgE sensitivity [17]. The American Academy of Allergy and Immunology has published guidelines [18] for providing care to persons with latex allergy. A flow chart for management of latex aller-gy is shown in Figure 1. Key points include the following: 1. All persons at risk for latex allergy should have a careful history and should complete a standardized latex allergy questionnaire (Table 1). Table 1. Questionnaire for Identification of Possible Latex Allergy* Figure 1. Overview of the identification and management of persons with latex allergy. 2. High-risk patients should be offered clinical testing for latex allergy. This includes children with spina bifida and health care workers with atopy (type I allergic reactions) (Table 1). 3. A latex-free environment is defined as one in which there is no latex glove use by any personnel. In addition, no direct patient contact should occur with other latex devices (catheters, condoms, adhesives, tourniquets, and anesthetic equipment). 4. Procedures on children with spina bifida should be done in a latex-free environment (Table 2). Table 2. Nonlatex Alternatives for Patients Allergic to Latex* 5. Procedures on all patients with positive skin test results should be done in a latex-free environment. Treatment of Latex Allergy Patients with an irritant latex reaction should eliminate unnecessary glove use. Cotton liners or barrier creams can be effective treatments. Patients who have contact dermatitis (type IV delayed hypersensitivity reaction) to latex additives should be appropriately diagnosed by patch testing. The implicated allergen should be avoided by changing to a different glove [1]. Patients with a negative history for latex allergy but with a positive result from latex skin testing or radioallergosorbent testing (RAST) or both should have a latex glove challenge test called a use test [19]. Patients with a positive latex challenge result should wear nonlatex gloves (Table 2). Patients with a negative challenge test result who have a positive result from latex skin testing or radioallergosorbent testing may tolerate low-protein latex glove use, but these patients should not have mucosal latex exposure during surgery or medical procedures. Patients with symptomatic latex allergy often present with severe allergic rhinoconjunctivitis and asthma that require them to leave their workplace. Although it is known that latex proteins are the responsible allergens, cornstarch glove powder has an important role. Latex proteins are easily absorbed by glove powder and are aerosolized at levels similar to those of other occupational respiratory allergens [10-12]. Complete removal of powdered latex gloves has resulted in undetectable levels of airborne latex particles. When all their coworkers switch to using powder-free latex gloves, health care workers with latex allergies have been able to return to their workplace [12]. In 1991, latex and banana were reported to be cross-reactive. Patients with latex allergy have subsequently presented with allergies to avocado, kiwi, and chestnut [20]. Clinically, these patients often have perioral itching and local urticaria, and they occasionally have been reported to have life-threatening, food-induced anaphylactic shock. The observed cross-reactivity of latex with avocado, kiwi, and chestnut probably occurs because latex proteins are structurally homologous with other plant proteins. Latex is ubiquitous in the medical environment, and health care workers encounter these allergens by multiple routes, including compromised skin and mucous membranes of the respiratory tract. Once sensitized, these health care workers are at risk for severe systemic allergic reactions. The antigenic protein level on latex rubber devices should be reduced to prevent further sensitization and allergic reactions [21]. Low-allergen latex gloves are available [22], and most manufacturers are working to lower protein levels. As cleaner products are brought to market, the incidence of new sensitization and adverse reactions is expected to decrease.

Airborne fungal fragments and allergenicity

Exposure to fungi, particularly in water damaged indoor environments, has been thought to exacerbate a number of adverse health effects, ranging from subjective symptoms such as fatigue, cognitive difficulties or memory loss to more definable diseases such as allergy, asthma and hypersensitivity pneumonitis. Understanding the role of fungal exposure in these environments has been limited by methodological difficulties in enumerating and identifying various fungal components in environmental samples. Consequently, data on personal exposure and sensitization to fungal allergens are mainly based on the assessment of a few select and easily identifiable species. The contribution of other airborne spores, hyphae and fungal fragments to exposure and allergic sensitization are poorly characterized. There is increased interest in the role of aerosolized fungal fragments following reports that the combination of hyphal fragments and spore counts improved the association with asthma severity. These fragments are particles derived from any intracellular or extracellular fungal structure and are categorized as either submicron particles or larger fungal fragments. In vitro studies have shown that submicron particles of several fungal species are aerosolized in much higher concentrations (300-500 times) than spores, and that respiratory deposition models suggest that such fragments of Stachybotrys chartarum may be deposited in 230-250 fold higher numbers than spores. The practical implications of these models are yet to be clarified for human exposure assessments and clinical disease. We have developed innovative immunodetection techniques to determine the extent to which larger fungal fragments, including hyphae and fractured conidia, function as aeroallergen sources. These techniques were based on the Halogen Immunoassay (HIA), an immunostaining technique that detects antigens associated with individual airborne particles >1 microm, with human serum immunoglobulin E (IgE). Our studies demonstrated that the numbers of total airborne hyphae were often significantly higher in concentration than conidia of individual allergenic genera. Approximately 25% of all hyphal fragments expressed detectable allergen and the resultant localization of IgE immunostaining was heterogeneous among the hyphae. Furthermore, conidia of ten genera that were previously uncharacterized could be identified as sources of allergens. These findings highlight the contribution of larger fungal fragments as aeroallergen sources and present a new paradigm of fungal exposure. Direct evidence of the associations between fungal fragments and building-related disease is lacking and in order to gain a better understanding, it will be necessary to develop diagnostic reagents and detection methods, particularly for submicron particles. Assays using monoclonal antibodies enable the measurement of individual antigens but interpretation can be confounded by cross-reactivity between fungal species. The recent development of species-specific monoclonal antibodies, used in combination with a fluorescent-confocal HIA technique should, for the first time, enable the speciation of morphologically indiscernible fungal fragments. The application of this novel method will help to characterize the contribution of fungal fragments to adverse health effects due to fungi and provide patient-specific exposure and sensitization profiles.

Measurements of Airborne Influenza Virus in Aerosol Particles from Human Coughs

Influenza is thought to be communicated from person to person by multiple pathways. However, the relative importance of different routes of influenza transmission is unclear. To better understand the potential for the airborne spread of influenza, we measured the amount and size of aerosol particles containing influenza virus that were produced by coughing. Subjects were recruited from patients presenting at a student health clinic with influenza-like symptoms. Nasopharyngeal swabs were collected from the volunteers and they were asked to cough three times into a spirometer. After each cough, the cough-generated aerosol was collected using a NIOSH two-stage bioaerosol cyclone sampler or an SKC BioSampler. The amount of influenza viral RNA contained in the samplers was analyzed using quantitative real-time reverse-transcription PCR (qPCR) targeting the matrix gene M1. For half of the subjects, viral plaque assays were performed on the nasopharyngeal swabs and cough aerosol samples to determine if viable virus was present. Fifty-eight subjects were tested, of whom 47 were positive for influenza virus by qPCR. Influenza viral RNA was detected in coughs from 38 of these subjects (81%). Thirty-five percent of the influenza RNA was contained in particles >4 µm in aerodynamic diameter, while 23% was in particles 1 to 4 µm and 42% in particles <1 µm. Viable influenza virus was detected in the cough aerosols from 2 of 21 subjects with influenza. These results show that coughing by influenza patients emits aerosol particles containing influenza virus and that much of the viral RNA is contained within particles in the respirable size range. The results support the idea that the airborne route may be a pathway for influenza transmission, especially in the immediate vicinity of an influenza patient. Further research is needed on the viability of airborne influenza viruses and the risk of transmission.

Discrimination of Aspergillus isolates at the species and strain level by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry fingerprinting.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to generate highly reproducible mass spectral fingerprints for 12 species of fungi of the genus Aspergillus and 5 different strains of Aspergillus flavus. Prior to MALDI-TOF MS analysis, the fungi were subjected to three 1-min bead beating cycles in an acetonitrile/trifluoroacetic acid solvent. The mass spectra contain abundant peaks in the range of 5 to 20kDa and may be used to discriminate between species unambiguously. A discriminant analysis using all peaks from the MALDI-TOF MS data yielded error rates for classification of 0 and 18.75% for resubstitution and cross-validation methods, respectively. If a subset of 28 significant peaks is chosen, resubstitution and cross-validation error rates are 0%. Discriminant analysis of the MALDI-TOF MS data for 5 strains of A. flavus using all peaks yielded error rates for classification of 0 and 5% for resubstitution and cross-validation methods, respectively. These data indicate that MALDI-TOF MS data may be used for unambiguous identification of members of the genus Aspergillus at both the species and strain levels.

Skin Prick Test Reactivity to Recombinant Latex Allergens

Background: Allergy to latex has become a serious and increasingly common health problem, particularly for healthcare workers and patients who undergo frequent surgical procedures. Testing for latex allergy currently involves in vitro tests and skin prick testing using crude preparations of natural rubber latex (NRL). To date, 10 latex proteins have received designation as allergens (Hev b 1 to Hev b 10) and, except for Hev b 4, have been cloned as recombinant proteins. Our aim was to compare the skin prick test (SPT) reactivity of six recombinant latex allergens with SPT reactivity to natural rubber latex proteins in known latex-allergic individuals. Methods: Six recombinant proteins were expressed in Escherichia coli, and tested as the intact fusion proteins (Hev b 2, 5, 6, 8) or as purified proteins (Hev b 3 and 7). SPT with the six recombinant latex allergens was performed using 10-fold serial dilutions on 31 latex-allergic subjects to determine the level of reactivity to each recombinant allergen. Latex-specific IgE was determined using the AlaSTAT assay. Results: All six recombinant allergens were reactive by SPT in at least 1 latex-allergic patient but not in any of the control patients. The frequency of sensitization to the various recombinant allergens was similar to previous studies using the native proteins isolated from NRL. The minimal level of protein for a positive skin test was 70 pg/ml for NRL and 1 ng/ml for one recombinant allergen (Hev b 7). In our patients, the use of a combination of recombinant latex allergens Hev b 5, 6 and 7 diagnosed latex allergy with 93% sensitivity and 100% specificity. Conclusion: Recombinant latex allergens are clinically reactive, can be produced in a standardized manner, and could potentially provide safe, sensitive and specific reagents for the diagnosis of latex allergy.

Hev b 8, the Hevea brasiliensis latex profilin, is a cross-reactive allergen of latex, Plant foods and pollen

Background: Plant profilins are important pan-allergens. They are responsible for a significant percentage of pollen-related allergies. Limited information is available about their involvement in the latex-fruit syndrome and the cross-reactivities between latex and pollen. We aimed to clone and express the Hevea brasiliensis latex profilin to investigate its allergological significance and serological cross-reactivities to profilins from plant foods and pollens. Methods: A DNA complementary to messenger RNA (cDNA) coding for the Hevea latex profilin, Hev b 8, was amplified by polymerase chain reaction from latex RNA. Recombinant (r)Hev b 8 was produced in Escherichia coli and used to screen sera from 50 latex- allergic health care workers (HCWs) with well-documented histories of food and pollen allergy and 34 latex-allergic spina bifida (SB) patients. The cross-reactivity of natural Hev b 8 and rHev b 8 with other plant profilins was determined by ELISA inhibition assays. A three-dimensional homology model of Hev b 8 was constructed based on known profilin structures. Results: The cDNA of Hev b 8 encoded a protein of 131 amino acids with a predicted molecular mass of 14 kD. Twelve of the 50 HCWs and 2 of the 34 SB patients were sensitized to Hev b 8. All Hev b 8-sensitized patients showed allergic symptoms to pollen or plant foods. Cross-reactivities between profilins of latex, pollen and plant food were illustrated by their ability to inhibit IgE binding to rHev b 8. Homology modeling of Hev b 8 yielded a structure highly similar to Bet v 2, the birch pollen profilin, with the most distinct differences located at the N-terminus. Conclusions: We conclude that primary sensitization to latex profilin in the majority of cases takes place via pollen or food profilins. Additionally, pollinosis and food-allergic patients with profilin-specific IgE can be at risk of developing latex allergy.

Detection of Infectious Influenza Virus in Cough Aerosols Generated in a Simulated Patient Examination Room

BACKGROUND The potential for aerosol transmission of infectious influenza virus (ie, in healthcare facilities) is controversial. We constructed a simulated patient examination room that contained coughing and breathing manikins to determine whether coughed influenza was infectious and assessed the effectiveness of an N95 respirator and surgical mask in blocking transmission. METHODS National Institute for Occupational Safety and Health aerosol samplers collected size-fractionated aerosols for 60 minutes at the mouth of the breathing manikin, beside the mouth, and at 3 other locations in the room. Total recovered virus was quantitated by quantitative polymerase chain reaction and infectivity was determined by the viral plaque assay and an enhanced infectivity assay. RESULTS Infectious influenza was recovered in all aerosol fractions (5.0% in >4 μm aerodynamic diameter, 75.5% in 1-4 μm, and 19.5% in <1 μm; n = 5). Tightly sealing a mask to the face blocked entry of 94.5% of total virus and 94.8% of infectious virus (n = 3). A tightly sealed respirator blocked 99.8% of total virus and 99.6% of infectious virus (n = 3). A poorly fitted respirator blocked 64.5% of total virus and 66.5% of infectious virus (n = 3). A mask documented to be loosely fitting by a PortaCount fit tester, to simulate how masks are worn by healthcare workers, blocked entry of 68.5% of total virus and 56.6% of infectious virus (n = 2). CONCLUSIONS These results support a role for aerosol transmission and represent the first reported laboratory study of the efficacy of masks and respirators in blocking inhalation of influenza in aerosols. The results indicate that a poorly fitted respirator performs no better than a loosely fitting mask.

Human IgE-binding epitopes of the latex allergen Hev b 5

BACKGROUND Hev b 5 is an acidic protein (isoelectric point, 3.5) rich in glutamic acid with 9 repeated amino acid (AA) sequences of XEEX or XEEEX. Although its function in Hevea brasiliensis is unknown, Hev b 5 has been identified as a major latex allergen. Immunoblot inhibition studies suggest Hev b 5 exists as multiple isoforms or contains a common epitope found in several other proteins. OBJECTIVE The purpose of this study was to further characterize Hev b 5 and to identify linear IgE-binding epitopes. METHODS Octapeptides spanning the entire Hev b 5 protein were synthesized on a derivatized cellulose membrane. The membrane was reacted with sera pooled from health care workers allergic to latex or rabbits immunized with latex proteins. B-cell epitopes were identified by subsequent incubations with the appropriate secondary antibodies and detected by using chemifluorescence. RESULTS Sera from patients allergic to latex recognized 6 IgE-binding regions located throughout the molecule. Two epitopes (2 and 4) had the common AA sequence of KTEEP. Epitopes 3 and 5 had a similar AA sequence of EEXXA, where X was P, T, or K. Epitopes 1 and 6 appeared to be unrelated to the other epitopes. Database analysis could not identify other proteins with similar sequences. Neither of the XEEEX sequences bound IgE. Control sera failed to react to any peptides. CONCLUSIONS Hev b 5 exists as multiple isoforms, but only small amounts are present in the nonammoniated latex preparations, such as those used for diagnostic tests, and this may help to explain the relatively poor sensitivity of some in vitro tests.

Fungal pigments inhibit the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of darkly pigmented fungi

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used to discriminate moniliaceous fungal species; however, darkly pigmented fungi yield poor fingerprint mass spectra that contain few peaks of low relative abundance. In this study, the effect of dark fungal pigments on the observed MALDI mass spectra was investigated. Peptide and protein samples containing varying concentrations of synthetic melanin or fungal pigments extracted from Aspergillus niger were analyzed by MALDI-TOF and MALDI-qTOF (quadrupole TOF) MS. Signal suppression was observed in samples containing greater than 250ng/μl pigment. Microscopic examination of the MALDI sample deposit was usually heterogeneous, with regions of high pigment concentration appearing as black. Acquisition of MALDI mass spectra from these darkly pigmented regions of the sample deposit yielded poor or no [M+H](+) ion signal. In contrast, nonpigmented regions within the sample deposit and hyphal negative control extracts of A. niger were not inhibited. This study demonstrated that dark fungal pigments inhibited the desorption/ionization process during MALDI-MS; however, these fungi may be successfully analyzed by MALDI-TOF MS when culture methods that suppress pigment expression are used. The addition of tricyclazole to the fungal growth media blocks fungal melanin synthesis and results in less melanized fungi that may be analyzed by MALDI-TOF MS.