IgE against bed bug (Cimex lectularius) allergens is common among adults bitten by bed bugs
2012; Elsevier BV; Volume: 129; Issue: 3 Linguagem: Inglês
10.1016/j.jaci.2012.01.034
ISSN1097-6825
AutoresJason B. Price, Adnan Divjan, W.R. Montfort, Kirstie H. Stansfield, Greg A. Freyer, Matthew S. Perzanowski,
Tópico(s)Allergic Rhinitis and Sensitization
ResumoDiscuss this article on the JACI Journal Club blog: www.jaci-online.blogspot.com. Cimex lectularius, the bed bug, is found worldwide and likely has been sharing human dwellings dating back to prehistoric times. With increased use of pesticides after World War II, the reports of bed bugs in developed countries decreased significantly.1Boase C. Bedbugs back from the brink.Pestic Outlook. 2001; 12: 159-162Crossref Scopus (107) Google Scholar Recently, C lectularius has made a resurgence, including in New York City (NYC) where housing violations for bed bugs rose from 82 to 4084 between 2005 and 2009.2Recommendations for the management of bed bugs in New York City: New York City Bed Bug Advisory Board Report to the Mayor and City Council. New York, NY; 2010. Available at: http://council.nyc.gov/downloads/pdf/bed_bugs_report_2010.pdf. Accessed January 4, 2012.Google Scholar The potential health implications of this increase in domestic exposure are not known; however, there is some evidence that exposed individuals can make a type 1 allergic response to C lectularius bites (see Table E1 in this article's Online Repository at www.jacionline.org).3Parsons D.J. Bedbug bite anaphylaxis misinterpreted as coronary occlusion.Ohio Med. 1955; 51: 669PubMed Google Scholar In the 1990s, researchers in Egypt reported skin test positivity to crude C lectularius extract among asthmatic patients.4Abou Gamra E.M. el Shayed F.A. Morsy T.A. Hussein H.M. Shehata E.S. The relation between Cimex lectularius antigen and bronchial asthma in Egypt.J Egypt Soc Parasitol. 1991; 21: 735-746PubMed Google Scholar Another case report identified C lectularius nitrophorin protein (cNP), a nitric oxide–carrying protein found in the injected saliva, that elicited an IgE antibody response in 1 patient.5Leverkus M. Jochim R.C. Schad S. Brocker E.B. Andersen J.F. Valenzuela J.G. et al.Bullous allergic hypersensitivity to bed bug bites mediated by IgE against salivary nitrophorin.J Invest Dermatol. 2006; 126: 91-96Crossref PubMed Scopus (43) Google Scholar, 6Walker F.A. Nitric oxide interaction with insect nitrophorins and thoughts on the electron configuration of the {FeNO}6 complex.J Inorg Biochem. 2005; 99: 216-236Crossref PubMed Scopus (140) Google Scholar, 7Weichsel A. Maes E.M. Andersen J.F. Valenzuela J.G. Shokhireva T. Walker F.A. et al.Heme-assisted S-nitrosation of a proximal thiolate in a nitric oxide transport protein.Proc Natl Acad Sci U S A. 2005; 102: 594-599Crossref PubMed Scopus (141) Google Scholar The uniqueness of the predicted cNP sequence8Valenzuela J.G. Ribeiro J.M. Purification and cloning of the salivary nitrophorin from the hemipteran Cimex lectularius.J Exp Biol. 1998; 201: 2659-2664PubMed Google Scholar suggests that IgE to cNP would indicate a hypersensitivity response that is specific to C lectularius. Aside from the single case described above,5Leverkus M. Jochim R.C. Schad S. Brocker E.B. Andersen J.F. Valenzuela J.G. et al.Bullous allergic hypersensitivity to bed bug bites mediated by IgE against salivary nitrophorin.J Invest Dermatol. 2006; 126: 91-96Crossref PubMed Scopus (43) Google Scholar the development of IgE antibodies to cNP among individuals bitten by bed bugs has not been reported. Our goal was to develop assays for measuring IgE antibodies against both the C lectularius extract and the cNP protein in order to determine the prevalence of sensitization to bed bug allergens among adults with reported bed bug bites. Thirty NYC residents who reported being bitten by bed bugs were recruited through Web advertisement (www.craigslist.org), fliers, and physician referral. Entry criteria for the study included report of having been bitten by a bed bug resulting in an itchy raised bump within the past year. Qualifying consenting participants donated serum and were queried about bed bug exposure and respiratory and allergic symptoms (wheezing, coughing, pruritic rash, itchy eyes, or runny nose) at the time of being bitten. Columbia University's Institution Review Board approved this study. The C lectularius extract was prepared from 160 mg of dried C lectularius (generously donated by Louis Sorkin, Museum of Natural History and George Keeney, Ohio State University) by freezing it in liquid nitrogen and then grinding it to a powder by using mortar and pestle. The powder was incubated for 2 hours in 600 μL of PBS with Tween 20 at 30°C. After 10 minutes of centrifugation, the supernatant was dialyzed against PBS to remove Tween 20. Recombinant cNP (prepared from Escherichia coli as described previously)7Weichsel A. Maes E.M. Andersen J.F. Valenzuela J.G. Shokhireva T. Walker F.A. et al.Heme-assisted S-nitrosation of a proximal thiolate in a nitric oxide transport protein.Proc Natl Acad Sci U S A. 2005; 102: 594-599Crossref PubMed Scopus (141) Google Scholar was dialyzed in PBS and diluted to 1.7 μg/mL. Both recombinant cNP and the C lectularius extracts were biotinylated (separately) by using EZ-Link Sulfo-NHS-LC-Biotin (Pierce, Rockford, Ill) in a 10 mM solution of the Sulfo-NHS-LC-Biotin reagent. The biotinylated cNP and C lectularius extracts were bound separately to streptavidin ImmunoCAPs as described previously (Phadia, Portage, Mich).9Erwin E.A. Custis N.J. Satinover S.M. Perzanowski M.S. Woodfolk J.A. Crane J. et al.Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase.J Allergy Clin Immunol. 2005; 115: 1029-1035Abstract Full Text Full Text PDF PubMed Scopus (87) Google Scholar Serum from the subjects was then incubated separately with C lectularius and cNP ImmunoCAPs for 30 minutes at 37°C. IgE concentrations were measured by using standard ImmunoCAP methods.9Erwin E.A. Custis N.J. Satinover S.M. Perzanowski M.S. Woodfolk J.A. Crane J. et al.Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase.J Allergy Clin Immunol. 2005; 115: 1029-1035Abstract Full Text Full Text PDF PubMed Scopus (87) Google Scholar Optimum C lectularius and cNP coating dilutions were determined to be 67 and 264 μg/mL, respectively, on the basis of maximal IgE binding experiments with serum samples identified as positive in our initial screening. IgE levels against cockroach and dust mite, 2 major allergens in NYC, and the total IgE level were also measured to assess seroatopy. Inhibition of IgE binding to C lectualrius experiments were conducted with dust mite and cockroach allergen extracts (see the Methods section in this article's Online Repository at www.jacionline.org). Seventeen (57%) subjects had detectable IgE levels (≥0.1 IU/mL) against the C lectularius extract (Fig 1). Of these 17, 9 subjects had detectable IgE level against cNP, while 8 did not. All subjects with IgE against cNP had measureable levels of IgE against the C lectularius extract. The presence of cNP in the C lectularius extract was confirmed by Western blot (Fig 2; see the Methods section in this article's Online Repository at www.jacionline.org). While there was a correlation between IgE to C lectularius and cNP (R = 0.64, P = .061) among those with measurable IgE levels to both, there was a wide range in the anti-cNPIgE/anti-C lectularius IgE ratios (0.05-3.2). The mean total IgE level was higher among subjects with than among subjects without IgE against the C lectularius extract (117 vs 28.8 IU/mL; P = .002) and cNP (125 vs 47.6 IU/mL; P = .057). The prevalence of IgE against cockroach was more common among subjects with than among subjects without IgE against the C lectularius extract (65% vs 35%; P = .024). Among the 8 subjects with measurable IgE levels against the C lectularius extract but not cNP, most had IgE against dust mite (6 of 8 subjects) or cockroach (7 of 8 subjects). Among the subjects with IgE to dust mite and cockroach, dust mite and cockroach allergen partially (and in 1 case, completely) inhibited the IgE response to C lectularius, while inhibition was minimal among the subjects without IgE to cockroach or dust mite (Table I). Reports of symptoms of wheeze, rhinitis, or cough on the day of or the day after the bite were similar among those without (≥0.1 IU/mL) than among those with IgE to either the C lectularius extract (54% vs 41%; P = .49) or cNP (52% vs 33%; P = .34).Fig 2Western blot of human (subject 23) IgE binding to cNP and C lectularius extract. cNP is a 32-kDa protein. A similar sized band can be seen in the extract. The slight difference in size likely is due to posttranslational modifications of the cNP.View Large Image Figure ViewerDownload Hi-res image Download (PPT)Table IPercent inhibition∗Percent inhibition was calculated by subtracting the IgE level measured after preincubation with allergen extract from the IgE level measured after incubation with PBS and dividing by the latter. of IgE binding with dust mite and cockroach extractsA. Inhibition with dust mite extractSubjectCosensitized to dust miteInhibition of IgE against†IgE against dust mite, C lectularius, or cockroach measured after inhibition.Dust miteC lectularius25Yes97%>77%‡After incubation with dust mite extract, IgE level to C lectularius was below the limit of detection (<0.1 IU/mL); however, the uninhibited sample was also low (0.43 IU/mL).27Yes95%88%8No—14%23No—4%B. Inhibition with cockroach extractSubjectCosensitized to dust miteInhibition of IgE against†IgE against dust mite, C lectularius, or cockroach measured after inhibition.CockroachC lectularius3Yes89%79%27Yes93%58%8No—14%23No—4%∗ Percent inhibition was calculated by subtracting the IgE level measured after preincubation with allergen extract from the IgE level measured after incubation with PBS and dividing by the latter.† IgE against dust mite, C lectularius, or cockroach measured after inhibition.‡ After incubation with dust mite extract, IgE level to C lectularius was below the limit of detection (<0.1 IU/mL); however, the uninhibited sample was also low (0.43 IU/mL). Open table in a new tab In summary, we have developed assays for measuring IgE antibodies to both the C lectularius extract and a unique protein from C lectularius (cNP). IgE antibodies to C lectularius and cNP were common among adults who reported being bitten and having a visible response to bed bugs. That many (30%) individuals had IgE to cNP, which has limited homology to proteins from other species, suggests a response specific to bed bug exposure. That there was variability in the correlation between concentrations of IgE to cNP and the C lectularius extract (including individuals with measurable IgE levels to the C lectularius extract, but not cNP) suggests that sensitized individuals were exposed to other allergens found in the C lectularius extract in addition to cNP. Many subjects who had IgE to the C lectularius extract also had IgE to dust mites and/or cockroaches and among these individuals, partial inhibition of binding to C lectularius was observed with dust mite and cockroach extracts. This indicates the presence of cross-reactivity with allergens from cockroaches and/or dust mites. While we were unable to demonstrate an association between allergic symptoms and IgE to either the C lectularius extract or cNP, this may be a function of our study's relatively small sample size and the limitations of retrospective reporting of symptoms. Given the large increase in human exposure to bed bugs in NYC and elsewhere and the demonstrated IgE response to allergens from C lectularius, it is clear that future studies need to examine the clinical relevance of IgE responses to bed bug allergens on allergic symptoms. From subjects with IgE to C lectularius, 2 subjects with IgE to dust mite, 2 subjects with IgE to cockroach, and 2 subjects without IgE to dust mite or cockroach were selected for inhibition experiments. Skin test extracts for Dermataphagoides farina (dust mite) and Blatella germanica (cockroach) (Greer, Lenoir, NC) were buffer exchanged with PBS in order to remove glycerin by using the Amicon Ultra-0.5 centrifugal filter unit with a molecular cutoff of 10 kDa (Millipore, Billerica, Mass). Serum (25 μL) was incubated at 37°C with either cockroach or dust mite extract or PBS (25 μL) for 2 hours. Samples were then assayed for IgE against the C lectuarius extract, cockroach, or dust mite as described for the undiluted samples. Percent inhibition was calculated by subtracting the IgE level measured after incubation with allergen extract from the IgE level measured after incubation with PBS and dividing by the latter concentration. Approximately 25 μg of cNP and 240 μg of the C lecularius extract in sample buffer (6% glycerol [vol/vol]; 30 mM Tris, pH 6.8; 1.2 mM EDTA; 1.2% SDS [wt/vol]; and 8.64 mM β-mercaptoethanol) were loaded in different wells of a 12 + 2 well precast Tris-HCl gel (4%-15%) (Criterion; Bio-Rad, Hercules, Calif). Gels were run at 50 mA for 1.5 hours and then protein was transferred onto a nitrocellulose membrane (500 mA for 1 hour). The membrane was blocked for 1 hour with Odyssey Blocking Buffer (Licor, Lincoln, Neb) before being incubated overnight at 4°C with subject 23's serum (1:10). Membranes were then incubated with a goat antihuman IgE epsilon DyLight 800 (1:1000, Antibodies Online, Atlanta, Ga) before being visualized by using the Odyssey imaging system (Licor).Table E1Previous reports of clinical reactions to bed bugsAuthor (year)No. of patientsPrimary clinical presentationSecondary reactionKinnear (1948)E1Kinnear J. Epidemic of bullous erythema on legs due to bed-bugs.Lancet. 1948; 2: 55Abstract PubMed Scopus (17) Google Scholar6Erythematous bullae on calvesNone reportedParsons (1955)E2Parsons D.J. Bedbug bite anaphylaxis misinterpreted as coronary occlusion.Ohio Med. 1955; 51: 669PubMed Google Scholar1Itchy lesions and mild neck swelling within 2 h after bitesUpper extremity, neck, and perioral edema, severe dyspnea within 3.5 h of bites; diagnosed as anaphylaxisSansom et al (1992)E3Sansom J.E. Reynolds N.J. Peachey R.D. Delayed reaction to bed bug bites.Arch Dermatol. 1992; 128: 272-273Crossref PubMed Scopus (37) Google Scholar2Patient 1 developed diffuse intensely pruritic, papular lesions 60 h after bites; patient 2 developed nonpruritic, erythematous, papular lesionsPatient 1 developed florid, hemorrhagic, bullous rash with edema 6 d after bitesTharakaram (1999)E4Tharakaram S. Bullous eruption due to Cimex lecticularis.Clin Exp Dermatol. 1999; 24: 241-242Crossref PubMed Scopus (25) Google Scholar1Pruritic, erythematous rash that evolved into a vesiculobullous eruption over 3 wkNone reportedFletcher et al (2002)E5Fletcher C.L. Ardern-Jones M.R. Hay R.J. Widespread bullous eruption due to multiple bed bug bites.Clin Exp Dermatol. 2002; 27: 74-75Crossref PubMed Scopus (34) Google Scholar1Extremely pruritic widespread bullous eruption; urticarial papules and plaquesNone reportedLiebold et al (2003)E6Liebold K. Schliemann-Willers S. Wollina U. Disseminated bullous eruption with systemic reaction caused by Cimex lectularius.J Eur Acad Dermatol Venereol. 2003; 17: 461-463Crossref PubMed Scopus (48) Google Scholar1Papular pruritic rashSecond exposure 4 wk later developed diffuse pruritic, erythematous large tense bullae with mild pyrexiaTer Poorten et al (2005)E7Ter Poorten M.C. Prose N.S. The return of the common bedbug.Pediatr Dermatol. 2005; 22: 183-187Crossref PubMed Scopus (84) Google Scholar1Diffuse intensely pruritic erythematous macules and papules; few discrete urticarial papulesNone reportedLeverkus et al (2006)E8Leverkus M. Jochim R.C. Schad S. Brocker E.B. Andersen J.F. Valenzuela J.G. et al.Bullous allergic hypersensitivity to bed bug bites mediated by IgE against salivary nitrophorin.J Invest Dermatol. 2006; 126: 91-96Crossref PubMed Scopus (78) Google Scholar1Multiple pruritic, erythematous papules, nodules, and blisters; several nodules with underlying erythema and moderate peripheral whealingNone reportedScarupa and Economides (2006)E9Scarupa M.D. Economides A. Bedbug bites masquerading as urticaria.J Allergy Clin Immunol. 2006; 117: 1508-1509Abstract Full Text Full Text PDF PubMed Scopus (40) Google Scholar17Papular urticaria–like dermatitisNone reportedMasetti and Bruschi (2007)E10Masetti M. Bruschi F. Bedbug infestations recorded in Central Italy.Parasitol Int. 2007; 56: 81-83Crossref PubMed Scopus (47) Google Scholar2Patient 1 developed pruritic erythematous macules and papules; patient 2 developed severe diffuse pruritic bullous eruptionsNone reportedGoddard and deShazo (2008)E11Goddard J. de Shazo R. Rapid rise in bed bug populations: the need to include them in the differential diagnosis of mysterious skin rashes.South Med J. 2008; 101: 854-855Crossref PubMed Scopus (11) Google Scholar1Pruritic, erythematous “bumps” 1 d after biteTwo days after initial bites developed erythematous papules; 12 d after initial bites rash improved but developed worsening pruritisStucki and Ludwig (2008)E12Stucki A. Ludwig R. Images in clinical medicine: bedbug bites.New Engl J Med. 2008; 359: 1047Crossref PubMed Scopus (19) Google Scholar1Multiple pruritic erythematous papules 1 d after bitesNone reportedMumcuoglu (2008)E13Mumcuoglu K.Y. A case of imported bedbug (Cimex lectularius) infestation in Israel.Isr Med Assoc J. 2008; 10: 388-389PubMed Google Scholar1Pruritic erythematous “spots”None reportedLevy Bencheton et al (2011)E14Levy Bencheton A. Berenger J.M. Del Giudice P. Delaunay P. Pages F. Morand J.J. Resurgence of bedbugs in southern France: a local problem or the tip of the iceberg?.J Eur Acad Dermatol Venereol. 2011; 25: 599-602Crossref PubMed Scopus (36) Google Scholar2Patient 1 developed diffuse pruritic erythematous maculopapular lesions with purpuric centers; patient 2 developed pruritic rash of several monthsNone reported Open table in a new tab
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