# paper.pdf ## Metadata - PDFFormatVersion=1.7 - Language=en - IsLinearized=true - IsAcroFormPresent=false - IsXFAPresent=false - IsCollectionPresent=false - IsSignaturesPresent=false - Custom.CrossMarkDomains[2]=elsevier.com - Custom.CrossmarkMajorVersionDate=2010-04-23 - Custom.EPSprocessor=PStill version 1.84.42 - Custom.ElsevierWebPDFSpecifications=7.0.1 - Custom.CrossmarkDomainExclusive=true - Custom.robots=noindex - Custom.doi=10.1016/j.waojou.2026.101391 - Custom.CrossMarkDomains[1]=sciencedirect.com - CreationDate=D:20260429042557+05'30' - Subject=World Allergy Organization Journal, 19 (2026) 101391. doi:10.1016/j.waojou.2026.101391 - Author=Jingping Fan MMSc - Creator=Elsevier - Producer=PDFlib+PDI 10.0.2p2 (C++/Win64) - ModDate=D:20260429043017+05'30' - Title=Evolution of IgE sensitization profiles to Artemisia pollen allergens in the allergic pediatric population - dc:format=application/pdf - dc:identifier=doi:10.1016/j.waojou.2026.101391 - dc:title=Evolution of IgE sensitization profiles to Artemisia pollen allergens in the allergic pediatric population - dc:creator=Jingping Fan MMSc,Huifang Song MMSc,Chen Zhao MMSc,Xiaomin Wang MMSc,Qianwei Zhao MMSc,Xiaoran Wang MMSc,Yumin Wang MMSc,Yuan Xia PhD - dc:subject=Artemisia pollen allergy,IgE sensitization profiles,Molecular spreading,Pediatric allergic rhinitis - dc:description=World Allergy Organization Journal, 19 (2026) 101391. doi:10.1016/j.waojou.2026.101391 - dc:publisher=The Author(s) - prism:aggregationtype=journal - prism:publicationname=World Allergy Organization Journal - prism:copyright=© 2026 The Author(s). 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World Allergy Organization Journal (2026) 19:101391 Open Access # Evolution of IgE sensitization profiles toArtemisia pollen allergens in the allergicpediatric population Jingping Fan, MMSca,c, Huifang Song, MMScb,c, Chen Zhao, MMSca,c, Xiaomin Wang, MMScaQianwei Zhao, MMSca, Xiaoran Wang, MMSca, Yumin Wang, MMScb and Yuan Xia, PhDa , ## ABSTRACT Objective: Allergic rhinitis prevalence has steadily risen among Hohhot’s pediatric andadolescent populations. Local epidemiological studies confirm Artemisia pollen as the primaryaeroallergen causing allergic rhinitis in summer and autumn. This study aimed to investigate howmultiple Artemisia pollen allergens influence immunological sensitization in children with allergicrhinitis. Methods: This study systematically investigated IgE sensitization profiles of 196 diagnosedpediatric allergic rhinitis patients with Artemisia pollen allergy. Obtained corresponding serumsamples, and comprehensive allergen profiling was conducted through enzyme-linked immunosorbent assay and immunoblot analysis. The immunodominant fractions of Artemisia pollenallergens were identified using mass spectrometry, with subsequent validation through recombinant allergen purification. Comprehensive pattern analysis revealed distinct sensitization profiles within the study cohort. Results: This study established Artemisia pollen as the predominant summer-autumn aeroallergen in Hohhot, with major allergenic components Art an 7 (69.01%), Art an 4 (56.14%), Art an 1 (18.71%), and Art an 3 (13.45%). Pediatric IgE sensitization profiles demonstrated markedheterogeneity. Monomolecular analysis revealed age-stratified dominance: 94.29% of childrenaged ≤6 years and 85.71% of those aged 7–14 years exhibited primary sensitization to either Artan 7 or Art an 4. Moreover, both dual and multi-component sensitization patterns predominantlyinvolved co-recognition of Art an 7 and/or Art an 4. Conclusions: We propose 2 distinct sensitization trajectories: (A) initial IgE priming by Art an 7and/or Art an 4 in early childhood, and (B) subsequent molecular spreading to Art an 1 and/or Artan 3 with immunological maturation. These findings established optimal therapeutic window forimmune tolerance. Keywords: Artemisia pollen allergy, IgE sensitization profiles, Molecular spreading, Pediatricallergic rhinitis aInner Mongolia Medical University, Hohhot, 010110, Inner Mongolia,China*Corresponding author. E-mail:**Corresponding author. School of public health, Inner Mongolia MedicalUniversity, Hohhot, 010110, Inner Mongolia, China. Email:c Jingping Fan, Huifang Song, and Chen Zhao are co-first authors.Full list of author information is available at the end of the article Received 18 July 2025; Received in revised from 22 March 2026; Accepted 9 April 2026Online publication date xxx1939-4551/© 2026 The Author(s). Published by Elsevier Inc. on behalf ofWorld Allergy Organization. This is an open access article under the CC BYlicense (). ### Page 2 2 Fan et al. World Allergy Organization Journal (2026) 19:101391 ## INTRODUCTION Allergic diseases arise from genetic predisposition and aberrant immune responses to allergens. Respiratory allergy (RA), primarilymanifesting as allergic rhinitis (AR) and asthma(AA), exemplifies the “united airway disease”concept due to shared upper/lower airway pathophysiology.1 AR is an IgE-mediated reactiontriggered by aeroallergens like pollen, dust mites,and animal dander.2 Clinically presenting as asymptomatic triad of nasal mucosal edema(congestion), paroxysmal sneezing, and itchyeyes. These symptoms typically persist for morethan 60 min per episode and recur on 2 or moreconsecutive days, thereby fulfilling the temporaldiagnostic criteria outlined in the ARIA (AllergicRhinitis and its Impact on Asthma) guidelines.3,4 AR affects 10–30% of adults and 40% of childrenglobally,5 with early onset evidenced by over 5%prevalence in 3-year-olds and over 14% in adolescents.6,7 Its comorbidity with allergic asthma(AA, prevalence 5–10%8) reflects sharedimmunopathology under the “united airway”paradigm. In China, AR/AA prevalence surgeddramatically, driven by Artemisia pollensensitization.4,9 Between 2008 and 2018,northern China’s Artemisia vulgaris sensitizationtripled to 30.3%, correlating with expandedcultivation of cross-reactive Artemisia desertorumwindbreaks.10 Artemisia pollen dominatesautumn aeroallergens in Inner Mongolia,11–14with regional AR prevalence rates of 4.5%among children aged 6–13 years15 and 18.5% inthe Inner Mongolia grassland area.13 The World Health Organization and International Union of Immunological Societies (WHO/IUIS) Allergen Nomenclature Subcommittee() has identified 7 allergens inArtemisia pollen.16 In China, Art v 1, Art v 3, andArt an 7 are major allergens.17 Currently, onlyArt v 1 and Art v 3 are accessible for componentanalysis diagnostics (CRDs).18 Moreover,emerging evidence suggests that IgEsensitization profiles to mugwort pollen allergensexhibit familial aggregation, with significantconcordance observed between Chinese parentsand their offspring, further underscoring theimportance of understanding componentresolved sensitization patterns in pediatric populations.19 Despite rising pediatric AR cases,Artemisia sensitization research in InnerMongolia remains adult-focused. This study investigates multiple allergenic proteins’ immunological effects on children’s sensitization profilesand their molecular spreading patterns, aiming toimprove diagnostic accuracy and advancepersonalized immunotherapy. METHODS Allergic pediatric subjects Patients with allergic symptoms were tested atInner Mongolia Maternity and Child Health CareHospital during August–September 2021–2022without specialized recruitment. All these patientshave been confirmed to have AR with or withoutAA based on the guideline for diagnosis andtreatment of pediatric allergic rhinitis and guideline for the diagnosis and optimal management ofasthma in children,20 with non-atopic children ascontrols. All patients have the following characteristics: (a) presence of 2 or more local symptoms(nasal obstruction, rhinorrhea, nasal pruritus, orparoxysmal sneezing) with daily symptom duration exceeding 1 h; (b) Artemisia pollen allergenspecific IgE (sIgE) antibodies (sIgE ≥0.35 IU/mL); (c) with or without AA, defined as a history ofdyspnea, wheezing, and/or coughing episodes,and confirmed by a positive bronchial dilationtest; (d) aged 0–14 years. The study was approved by the ethics committee of the Inner Mongolia Maternity and ChildHealth Care Hospital (No.2020064) and informedconsent was obtained from all patients’ parents orguardians prior to serum collection. Total and specific IgE A retrospective analysis of serum sIgE levelswas conducted using the Allergen-Specific IgEAntibody Detection Kit (HOB Biotech, Suzhou,China) in Hohhot, Inner Mongolia, China. This kit isused to detect total IgE antibodies and allergenspecific IgE antibodies in serum by enzymelinked immunosorbent assay (ELISA) rapid teststrip technology, with each strip containing: 1positive control, 1 negative control, 1 total IgEdetection zone, and 13 allergen-specific IgEpanels. The allergens included House dust mite(House dust mite, Dermatophagoides ### Page 3 Volume 19, No. 5, Month 2026 pteronyssinus), Tree pollen (Cypress, Elm, Sycamore, Willow, Poplar), Artemisia pollen (Ragweed,Artemisia, Wormwood), Mold (Penicillium notatum, Aspergillus fumigatus, Cladosporium, Alternaria, Rhizopus, Mucor), Animal dander (Dog,Cat), Milk, Egg fractions, Marine fish (Cod,Salmon, Bass), Shellfish (Shrimp, Crab, Scallop),Fruit (Mango, Pineapple, Apple, Peach, Strawberry), Nut (Peanut, Pistachio, Cashew, Hazelnut),Beef and Lamb. According to the Clinical Study Guidelines forAllergen-Specific IgE Antibody Test Kits, rate thereactivity of the sIgE tests as: class 0 (<0.35 IU/mL), class 1 (≥0.35 and < 0.70 IU/mL), class 2(≥0.70 and < 3.50 IU/mL), class 3 (≥3.50and < 17.50 IU/mL), class 4 (≥17.50 and < 50.00IU/mL), class 5 (≥50.00 and < 100.00 IU/mL), andclass 6 (≥100.00 IU/mL). The serum sIgE level forpollen above class 0 was considered positive. Extraction of pollen protein The pollen of Artemisia annua was collected inthe suburbs of Hohhot city, Inner Mongolia, duringthe flowering season.The pollen samples were fullyground with liquid nitrogen, dehydrated withacetone, and then stored at 20◦ C for reserve.Thepollen proteins were extracted using the PollenProtein Extraction Kit (Bestbio, Shanghai, China). Purification and recombinant expression ofArtemisia pollen components To construct expression plasmids for the 4Artemisia pollen allergens, their respective mRNAsequences were first comprehensively identifiedand optimized. These sequences were cloned intothe pFastBac1 vector to generate recombinantplasmids containing both a His-tag and a Flag-tag.The plasmids were transformed into DH10Baccompetent cells (General Biologicals, Anhui,China) via ice incubation for 30 min, heat shock at42◦ C for 90 s, and subsequent addition of 500 μLSOC medium (composition: 2% (w/v) tryptone,0.5% (w/v) yeast extract, 0.05% (w/v) NaCl, 2.5 mMKCl, 10 mM MgCl2, 20 mM glucose). After 4 hincubation, cells were plated on triple-antibioticLB agar containing 7 μg/mL gentamicin, 100 μg/mL kanamycin, 10 μg/mL tetracycline, 40 μg/mLIPTG, and 40 μL 2% X-gal. Plates were invertedand incubated at 37◦ C for 48 h. 3 White colonies identified via blue-whitescreening were selected and inoculated into 3 mL of SOC medium supplemented with theappropriate antibiotic. The cultures were incubated overnight at 37◦ C with shaking at 200 rpmto facilitate bacterial growth and plasmid amplification. Bacmids were extracted using the M5HiPer Plasmid Miniprep Kit (Mei5Bio, Beijing,China) and stored at 20◦ C. Recombinant bacmids were transfected into Sf9 insect cells usingFuGENE� HD (Promega, Shanghai, China) togenerate P1 baculovirus, which was amplified toproduce high-titer P2 stocks. Sf9 cells were harvested by centrifugation(4000 rpm, 10 min, 4◦ C), resuspended in lysisbuffer (20 mM Tris-HCl, 150 mM NaCl, 1 mMPMSF, 10 mM imidazole, pH 8.0), and lysed bysonication (300 W, 3 s on/5 s off, 30 min total).Lysates were clarified by centrifugation(18,000 rpm, 15 min, 4◦ C), followed by purification of Art an 1, Art an 3, Art an 4, and Art an 7allergens using Ni-IDA beads (Smart-Lifesciences,Changzhou, China) per manufacturer instructions.SDS-PAGE analysis was performed on samplescollected from each purification step to evaluatethe purity of the target protein. Western blotting The protein concentration for all extracts andpurified allergen components was estimated using the BCA Protein Assay kit (Beyotime,Shanghai, China) with bovine serum albumin as astandard. All protein samples were diluted to thesame total protein concentration. The samples were loaded on 12.5% SDS-PAGEgels for protein separation and subsequentlyelectroblotted onto PVDF membranes (Beyotime,Shanghai, China). After blocking of excessprotein-binding sites with QuickBlock� BlockingBuffer (Beyotime, Shanghai, China) for 30 min, thePVDF membranes were incubated with the primary antibody (patient serum samples, 1:40–1:300 dilution) at 4◦ C overnight. After TBSTwashing, the secondary goat anti-human IgG-HRPantibody (ABclonal, Wuhan, China) was added for 1 h incubation. Lastly, immunoreactions werevisualized by the ECL kit (Affinity Biosciences,Changzhou, China). Densitometric analyses were ### Page 4 4 Fan et al. World Allergy Organization Journal (2026) 19:101391 performed using the GelView 6000Plus system(Tanon Technology Co., Ltd., Shanghai, China). Statistical analysis Recombinant plasmids for Artemisia pollen allergens were constructed using SnapGene 4.2.4,with data plotting performed using Origin 2021and R 4.3.3. Statistical analyses were performedusing SPSS 25.0 and R 4.3.3, with a value ofP < 0.05 considered as significantly different.Continuous variables following normal distribution were expressed as mean ± standard devia tion (x ± s), while non-normally distributed datawere reported as median and interquartile range.For two-group comparisons: independent t-testswere applied to normally distributed variables,and Wilcoxon rank-sum tests to non-parametricdata. Categorical variables were compared usingthe chi-square test or Fisher’s exact test. Multigroup comparisons with normal distributionwere analyzed by ANOVA, followed by pairwisepost-hoc testing with the LSD method. ## RESULTS Demographic and clinical characteristics of thestudy participants A total of 196 children (123 males and 73 females) were enrolled in this study ().Participants aged 2–14 years (mean 5.1 ± 2.6years) were stratified into 3 age groups: 63(32.14%) aged 0–3 years, 84 (42.86%) aged 4–6years, and 49 (25.00%) aged 7–14 years.Comorbid allergic conditions included 31 cases(15.82%) of allergic conjunctivitis, 9 (4.6%) ofurticaria, and 7 (3.6%) of eczema. Based onclinical diagnosis, 153 (78.06%) participants werecategorized into AR and 43 (21.94%) into theAA. No statistically significant difference wasobserved in the prevalence of children acrossdifferent genders and age groups (P > 0.05).Complete demographic characteristics arepresented in. Specific IgE profiles to allergens The median serum total IgE level among participants was 166.47 IU/mL (InterQuartile Range[IQR], 101.96–304.17), with Artemisia pollenspecific IgE (sIgE) levels at 51.79 IU/mL(IQR,10.39–117.01). Children diagnosed with AA demonstrated significantly higher serum levels ofArtemisia pollen-sIgE compared to those with ARalone (P = 0.024) (). While all 196 children exhibited sensitization toArtemisia pollen allergens, 66.33% (130/196)showed co-sensitization to other inhalant allergens, primarily tree pollen (52.04%, 102/196),mold (28.06%, 55/196) and dander (dog and cat)(23.98%, 47/196). 42 cases (21.43%, 42/196) werepositive for ingested allergens, with milk (16.33%,32/196), egg fractions (4.59%, 9/196) and beefand lamb (3.57%, 7/196) being the top 3. The 102(66.67%, 102/153) children with AR were positivefor other inhalant allergens, and 33 (21.57%, 33/153) were positive for ingested allergens. Relatively, the 28 (65.12%, 28/43) children with AAwere positive for other inhalant allergens and 9(20.93%, 9/43) were positive for ingested allergens (a). There was no statistically significantdifference observed in the rate of positivity foreach allergen among children of different ageand gender groups, or in the AA and AR groups(P > 0.05) (). The sensitization profile analysis demonstrateda graded distribution of Artemisia-specific IgE responses, with proportions progressivelyincreasing from class 1 (9.69%, n = 19) to class 6(27.55%, n = 54). Notably, children with highersensitization grades (classes 5–6) collectivelyconstituted 52.04% (102/196) of the study cohort,exceeding the cumulative proportion of lowergrades (classes 1–4, 47.96%). A statistically significant difference was observed between differentsIgE classes (χ2 = 14.855, P = 0.007). Notably, the4-6-year age group demonstrated the highestArtemisia pollen sensitization rate. In childrenaged 7–14 years, high-grade sensitization (sIgEclasses 4–6) predominated over lower grades(classes 1–3) (,b). Both gendersshowed comparable patterns, with Artemisiapollen-sensitized clustering in classes 5–6(c). In the AA and AR groups, the prevalenceof high Artemisia sIgE level was higher in the ARgroup, and the sensitization increased with theseverity of the sIgE level. The proportion ofchildren with Artemisia sIgE level of class 5–6was significantly higher than classes 1–4 in theAA group (d). ### Page 5 Volume 19, No. 5, Month 2026 5 | CharactersGender, no. (%) | AR (n = 153) | AA (n = 43) | P-value | | - | - | - | - | | Males | 92 (60.13) | 31 (72.09) | 0.152 | | Females | 61 (39.87) | 12 (27.91) | | Age, no. (%) | | ≤3 | 54 (35.30) | 9 (20.93) | 0.115 | | 4–6 | 65 (42.48) | 19 (44.19) | | 7–14 | 34 (22.22) | 15 (34.88) | | Total IgE, median, IQR (IU/mL) | 142.38 (101.37–265.93) | 195.57 (109.93–500.00) | 0.070 | | Artemisia-sIgE, median, IQR (IU/mL) | 39.67 (7.86–102.53) | 71.57 (40.99–200.00) | 0.024a | | Allergen positive no. (%) | | Tree pollen (Cypress, Elm, Sycamore, Willow,Poplar) | 78 (50.98) | 25 (58.14) | 0.406 | | Mold (Penicillium notatum, Aspergillusfumigatus, Cladosporium, Alternaria,Rhizopus, Mucor) | 40 (26.14) | 15 (34.88) | 0.260 | | Animal dander (Dog, Cat) | 38 (24.84) | 9 (5.88) | 0.596 | | House dust mite (House dust mite,Dermatophagoides pteronyssinus) | 1 (0.65) | 1 (2.33) | 0.392 | | House dust | 10 (6.54) | 3 (6.98) | 1.000 | | Milk | 28 (18.30) | 4 (9.30) | 0.158 | | Beef and lamb | 7 (4.58) | 0 (0.00) | 0.351 | | Egg fractions | 8 (5.23) | 1 (2.33) | 0.687 | | Nut (Peanut, Pistachio, Cashew, Hazelnut) | 2 (1.31) | 3 (6.98) | 0.071 | | Fruit (Mango, Pineapple, Apple, Peach,Strawberry) | 1 (0.65) | 2 (4.65) | 0.122 | | Shellfish (Shrimp, Crab, Scallop) | 2 (1.31) | 0 (0.00) | 1.000 | | Marine fish (Cod, Salmon, Bass) | 2 (1.31) | 1 (2.33) | 0.526 | | sIgE of Artemisia pollen, no. (%) | | Class 1 | 14 (9.15) | 5 (11.63) | 0.007b | | Class 2 | 19 (12.42) | 2 (4.65) | | Class 3 | 24 (15.69) | 1 (2.32) | | Class 4 | 26 (16.99) | 3 (6.98) | | Class 5 | 32 (20.91) | 16 (37.21) | (continued) ### Page 6 6 Fan et al. World Allergy Organization Journal (2026) 19:101391 | CharactersClass 6 | AR (n = 153) 38 (24.83) | AA (n = 43) 16 (37.21) | P-value | | - | - | - | - | | Sensitized to Artemisia component, n (%) | | Art an 1 | 22 (14.37) | 10 (23.26) | 0.164 | | Art an 3 | 20 (13.07) | 3 (6.98) | 0.273 | | Art an 4 | 79 (51.63) | 17 (39.53) | 0.161 | | Art an 7 | 89 (58.17) | 29 (67.44) | 0.272 | InterQuartile Range; sIgE, specific IgE antibody.aP < 0.05bP < 0.01 ### Page 7 Volume 19, No. 5, Month 2026 Patterns of sensitization to Artemisia allergencomponents among those sensitized to Artemisiapollen To characterize the allergenic profile of Artemisia pollen within the studied population,immunoblot analysis revealed 4 distinct IgEreactive protein bands at molecular weights ofapproximately 65, 28, 15, and 10 kDa ().Subsequent identification via mass spectrometry,aligned with the WHO/IUIS AllergenNomenclature database, confirmed theseproteins as Art an 7, Art an 1, Art an 4, and Artan 3, respectively. Component-resolved diagnostics demonstrated a hierarchical pattern ofIgE reactivity: Art an 7 exhibited the highestsensitization rate at 59.7% (118/196), followed byArt an 4 at 49.0% (96/196), Art an 1 at 16.3% (32/196), and Art an 3 at 11.7% (23/196). These findings underscore the predominance of Art an 7and Art an 4 as major allergens in this cohort,highlighting their relevance in the moleculardiagnosis and management of Artemisia-inducedallergic responses. However, 11.2% (25/196) of subjects showedno detectable IgE reactivity to these components(). Notably, 50.0% (98/196) of participantsdemonstrated a single positive component,26.0% (51/196) demonstrated 2 positivecomponents, 9.7% (19/196) displayed 3 positivecomponents, and only 1.5% (3/196) of childrenexhibited 4 positive components. In almost allchildren, the initial IgE response is directed 7 against Art 7, a sort of initiatory molecule of thisIgE response. Sensitization started against Art an7 and/or Art an 4 (defined as group Amolecules), then involved Art an 1 and Art an 3(group B molecules). This progression could bedescribed as the AB march of Artemisia allergy(). When the data were stratified by age, it wasfound that 47.9% of children aged 6 years andyounger responded to only 1 molecule (monomolecular profile), 29.3% responded to 2 molecules (oligomolecular profile), and 10.9%responded to 3 or more molecules (polymolecularprofile) (c). A comparable pattern emergedamong children aged 7–14 years, with 57.1%responding to only 1 molecule, 16.3%responding to 2 molecules, and 12.2%responding to 3–4 molecules (d). All ofthese children demonstrated a predominantsensitivity to Art an 7 or Art an 4. Theoligomolecular pattern in children under 3 yearsof age was predominantly characterized by “Artan 7-Art an 4” (20.63%, 13/63), with a minor proportion exhibiting “Art an 7-Art an 1” (6.35%, 4/63)or “Art an 4-Art an 3” (4.76%, 3/63) sensitizationpatterns (a). In contrast to children aged 4–6years, the proportion of monomolecularsensitization decreased, while the proportion ofsensitization to 3 allergenic molecules increased,predominantly manifesting as a “Art an 7-Art an4-Art an 1” pattern (b). By the age of 7, theproportion of sensitization to the 3 allergeniccomponents decreased. Post this age, the ### Page 8 8 Fan et al. World Allergy Organization Journal (2026) 19:101391 ### Page 9 Volume 19, No. 5, Month 2026 9 oligomolecular sensitization patternpredominantly manifested as “Art an 7-Art an 4”(12.24%, 6/49), with smaller proportions exhibiting “Art an 4-Art an 1” (4.08%, 2/49) and “Art an 7Art an 4-Art an 1” (4.08%, 2/49). Notably, theprevalence of the “Art an 7-Art an 4-Art an 3”sensitization pattern declined with age (d). A further finding of interest is that Art an 4 andArt an 3 tend to occur together, while Art an 7 andArt an 3 rarely co-occur (e). Moreover, forboth groups, the relationship betweensensitizing allergens and symptoms wasexamined. Patients with AR had a higherpercentage of reactions to Art an 3, Art an 4,and Art an 7 than patients with AA (). Purified allergen analysis The recombinant Artemisia pollen allergenicproteins were obtained by following the methoddescribed in the material. The amino acid sequences of the purified recombinant proteinswere validated by mass spectrometry. Crossreferencing with the WHO/IUIS Allergen Nomenclature database confirmed that these proteinscorrespond to Art an 7, Art an 1, Art an 4, and Artan 3, respectively. Following separation by SDSPAGE (), IgE bindingto purified recombinant allergens wereinvestigated by immunoblotting using serumfrom Artemisia-allergic patients (). All recombinant Artemisia pollen allergenic proteinswere observed to exhibit distinct bands at theircorresponding positions, consistent with thepredicted protein size and exhibiting expressionlevels comparable to that of Artemisia pollenprotein extracts. These findings suggest that theypossess similar immunogenicity. ## DISCUSSION In our cohort of 196 pediatric AR patients,21.94% had comorbid AA, and 15.82% had AC, ### Page 10 10 Fan et al. World Allergy Organization Journal (2026) 19:101391 consistent with epidemiological patterns where70–80% of AR cases manifest as isolated diseaseand 20–30% are associated with comorbidasthma.21 The AA-AR association suggests sharedallergic airway mechanisms.11 Bousquet et al22proposed that the multimorbid allergicphenotype (AA, AR, and AC) may synergisticallyincrease disease burden. Our data didn’t revealsignificant gender-based disparities, with malesubjects predominating in both the AR (60.13% vs39.87%) and AA (72.09% vs 27.91%) cohorts(P = 0.152). Furthermore, age-stratified analysis demonstrated that the 4–6 years subgroup comprised oflargest demographic proportion (42.86%)compared to other developmental stages(χ2 = 4.33, P = 0.115), suggesting potential agedependent immunological susceptibility patterns.Consistent with these findings, a nationwideepidemiological survey by Guo et al23 revealedsignificantly elevated asthma prevalence inpreschool children (4–6 years) compared to otherpediatric age strata. Additionally, following thefindings of Luo et al,24 the prevalence ofArtemisia sensitization increased with age in thehigher age groups. In this study, the Artemisiapollen sensitization rate demonstrated an agedependent epidemiological pattern, peaking inchildren aged 4–6 years. Notably, within the schoolaged cohort (7–14 years), high-grade ArtemisiasIgE reactivity (classes 4–6) exhibited significantquantitative predominance over low-grade sensitization (classes 1–3) (72% vs 28%), with the proportion of severe sensitization cases (≥class 4)showing elevated prevalence compared to both 0–3 years and 4–6 years age groups. This agedependent susceptibility pattern was further verified by Wu et al,25 whose quantitative aeroallergensensitization analysis demonstrated that 7–14 yearsexhibited enhanced sIgE responsiveness (P < 0.05for each) to common airborne allergens (cat anddog) relative to younger children (0–6 years). Thisage-dependent susceptibility pattern reflectsenhanced sIgE responsiveness to aeroallergens inolder children, potentially due to increased outdoor exposure. IgE quantification’s diagnostic role in atopicdisorders is confirmed by current guidelines(EAACI/AAAAI 2022).26 Both total and ArtemisiasIgE levels demonstrated statistically significant elevation (P = 0.024) in children with AAcompared with AR. In both pediatric cohortswith AR and AA, we observed significant inhalantallergen sensitization rates, with 71.43% ofpatients exhibiting multi-allergen sensitization(≥2 allergens). 52.04% of the children demonstrated co-sensitization to Artemisia pollen andtree pollen. The secondary allergens are molds(28.06%) and the dander of dogs and cats(23.98%), with minimal to no allergy to house dustmites. These findings contrasted with those of Maet al,11 who reported that children morefrequently showed a monosensitization pattern.However, it was consistent with the polysensitization patterns of AR patients who weresensitized to 2 or more pollens (79.4%) (includingArtemisia, Chenopodium, and Humulus scandens)in northern China.13 Conversely, only 21.43% ofthe children exhibited an allergy to ingestibleallergens, predominantly to milk (16.33%), eggcomponents (4.59%), and beef and lamb(3.57%). As reported by Li et al,27 theprevalence of food allergies in Beijing was foundto exceed 20%, with a particular prevalence ofallergies to grass pollen and tree pollen. The IgE-binding activity of Artemisia pollencomponents was assessed via IgE immunoblottingusing serum samples from pediatric participants.Subsequently, the 4 recombinant Artemisiaallergen proteins were cloned, expressed, andcharacterized. Immunodominant IgE-reactivecomponents were identified as follows: Art an 1(16.3%), Art an 3 (11.2%), Art an 4 (49.0%), and Artan 7 (59.7%). Among 100 mugwort-sIgE positivepatients in western China, the positivity rates forArt v 1, Art v 3, and Art v 4 were 52.0%, 41.0%, and31.0%, respectively.28 In northwestern China,among 135 patients with Artemisia allergy, thepositivity rates for the Artemisia desertorumallergens Art d 1, Art d 3, and Art d 7 were85.2%, 88.9%, and 82.2%, respectively. Thenumber of allergens and the level of specific IgEto Art d 3 were independently associated withasthma.29 Gao et al17 reported significantdifferences in sensitization rates to Art v 1 (84%),Art an 7 (87%), Art v 3 (66%), and Art ar 2 (48%)among children, teenagers, and adults.Specifically, the adult group demonstratedsignificantly lower serum IgE levels to mugwortand Art an 7 than the child and teenage groups. ### Page 11 Volume 19, No. 5, Month 2026 The discrepancy in these findings indicates thatchildren with AR in Hohhot may exhibit a distinctcomponent sensitization profile compared toadult AR patients, with age likely serving as theprincipal factor underlying this difference. Art an 7, formally characterized as a galactoseoxidase by the IUIS Allergen Nomenclature Subcommittee, demonstrated IgE sensitization rates of73–92% in Artemisia pollen-allergic cohorts acrossEuropean epidemiological studies during 1990–200830–33, and more than 94% of Artemisia pollensensitized patients exhibit specific IgE reactivity torecombinant Art an 7.32 This allergen has beenmechanistically linked to mugwort-food crossreactivity through structural homology withgalactose-containing glycoproteins.34 Art an 4,officially designated a profilin family allergen,demonstrates greater than 70% sequence identityand preserved actin-binding domains with panallergenic profilins,35 presenting a sensitizationprevalence of 34–36% in mugwort allergicpatients.30,36 This structural conservationunderlies its cross-reactive IgE epitopes, as evidenced by immunoblot inhibition when preincubated with Bet v 2 or Man i 4 IgE-positivesera.37 Art an 1 is a two-domain glycoproteincomprising an N-terminal defensin-like domainstabilized by 3 conserved disulfide bonds and a Cterminal hydroxyproline-rich glycodomain. It isestablished as a diagnostic marker allergen forgenuine mugwort pollen sensitization.38Epidemiological studies demonstrate that up to41% of European/North American patients withpollen allergy exhibit sensitization to Art an 1confirmed by component-resolved diagnosis.26,39 Art an 3, an immunodominant nonspecific lipid-transfer protein (nsLTP), is recognized as the prototype allergen of the nsLTP family,which is one of the important allergies in adults inthe Mediterranean region. About 9% of more than23,000 allergy patients in the central and southernregions of Italy are sensitive to nsLTP, much higherthan in the northern regions.40,41 About 12% of2000 Spanish pollen allergy patients were foundto be sensitive to nsLTP,42 and there weredifferences in sensitivity to nsLTPs amongseasonal allergy patients in different regions.43Geographical and climatic factors may also causedifferences in the prevalence of nsLTP allergies indifferent countries.44 11 Children with AR in this region showed differentsensitivities to these 4 allergen components. Thisphenomenon may be related to the natural abundance of different components in pollen. Moraleset al45 demonstrated that Profilins are greatlyexpressed near the pollen tube, making themhighly abundant in pollen, resulting in the actualexposure dose varying between differentchildren. On the other hand, current molecularspreading theory postulates that primarysensitization to an initiating allergen moleculeduring a critical developmental window inducesprogressive IgE epitope spreading, ultimatelyleading to extreme heterogeneity in the IgEsensitization spectrum in allergic populations. Thisprocess manifests as distinct sensitizationtrajectories: monomolecular (1 allergen),oligomolecular (2–4 allergens), or polymolecular(≥5 allergens) profiles, governed by individualatopic endotypes and cumulative allergenexposure dose.46 In this study, sensitization patterns varied acrossage groups, with the monomolecular profile(sensitization to a single allergen) being the mostprevalent, followed by the oligomolecular profile(sensitization to 2 allergens). In contrast, the polymolecular profile (sensitization to 3 or more allergens) exhibited significantly lower occurrencerates. The oligomolecular pattern in children under 3 years of age was predominantly “Art an 7-Artan 4” (20.63%), the proportion of monomolecularprofile decreased in children between 4 and 6years of age (46.4%), and the proportion ofsensitization to 3 allergens increased, with a predominantly “Art an 7-Art an 4-Art an 1” pattern(7.14%), while children aged 7–14 years showed adecrease in sensitization to 3 allergenic molecules, with the predominant polymolecular profileagain being “Art an 7-Art an 4” (12.24%). Art an 7or Art an 4 sIgE was detected in the sera of mostof the children included in the present study, andboth single and multimolecule sensitized childrenshowed co-sensitization to Art an 7 and/or Art an4. Thus, Art an 7 and/or Art an 4 may be the initiating molecules for the IgE response to pratensepollen in children in this region (defined as groupA), which further extends to Art an 1 and/or Art an 3in a subset of children with increasing age (definedas group B), showing “Art an 7-Art an 4-Art an 1”, ### Page 12 12 Fan et al. World Allergy Organization Journal (2026) 19:101391 “Art an 7-Art an 4-Art an 3”. The AB process patternof sensitization to Artemisia allergens in childrenwas delineated, representing a molecularspreading trajectory that has been reported inseveral previous studies. A national cross-sectionalsurvey conducted by the Italian Pediatric AllergyNetwork (IPAN) demonstrated that specific IgE toPhl p 1, Phl p 7, and Phl p 12 can serve as reliablebiomarkers for the diagnosis of Phleum pratensepollen allergy in pediatric populations. Moreover,the IgE sensitization profiles among affected children exhibited marked heterogeneity The IgEresponse to Phleum pratense pollen in children isinitiated by Phl p 1 (the initiating molecule), with theproduction of Phl p 1 IgE in serum generally preceding the onset of clinical symptoms by severalyears. This is followed by a gradual expansion tomolecules such as Phl p 2 and Phl p 4 at the onset ofthe disease, and ultimately covering Phl p 5, Phl p 6,and Phl p 11 after the exacerbation of symptoms.Furthermore, it is postulated that the initial phase ofmolecular diffusion (eg, the monomolecular oroligomolecular sensitization stage) represents theoptimal juncture for immune modulation.47 Anotable similarity exists between the molecularprocesses involved in the context ofDermatophagoides pteronyssinus sensitizationand those involved in the context of allergies toPhleum pratense pollen, as evidenced bymolecular spreading. In particular, the Der p 1,Der p 2 and Der p 23 allergens have beenidentified as playing a pivotal role in the initiationof this process, specifically in the initial stage(defined as group A), which is followed by theprogression to the Der p 4, Der p 5, Der p 7 andDer p 21 (defined as group B) allergens, andultimately, to the Der p 11, Der p 14, Der p 15and Der p 18 (defined as group C) allergens. Thisprogression pattern has been designated theABC progression model of house dust miteallergy. Participants exhibiting the most extensiveABC IgE sensitization stage demonstrated asignificantly increased risk of developing miteassociated AR and AA compared to those withmonomolecular sensitization. Furthermore, IgE sensitization to initiator molecules in otherwise healthy children at ≤5 years ofage was predictive of asthma development duringschool age. IgE sensitization in healthy children toinitiatory molecules at age 5 years or younger predicted the development of asthma at schoolage.46 Consequently, IgE tests employingindividual allergenic molecules are regarded asa more precise and informative option thanthose based on allergenic extracts, particularlyfor patients with polysensitization. In conclusion, this study characterized Artemisiasensitization patterns in AR children from Hohhot,identifying 4 major allergens: Art an 7, Art an 1, Artan 4, and Art an 3. We observed a molecularspreading trajectory where Art an 7 and/or Art an 4typically initiate IgE responses, with subsequentprogression to Art an 1 and/or Art an 3 in olderchildren. These findings reveal cross-reactiveArtemisia components and demonstrate that earlydetection of this sensitization pattern enablestimely immunoprevention and immunotherapyduring the critical immune tolerance window. Abbreviations RA, respiratory allergy; AR, allergic rhinitis; AA, allergicasthma; ARIA, Allergic Rhinitis and its Impact on Asthma;WHO, World Health Organization; IUIS, InternationalUnion of Immunological Societies; CRDs, componentanalysis diagnostics; IgE, Immunoglobulin E; sIgE, specificIgE antibody; PVDF, Polyvinylidene fluoride; TBST, Trisbuffered saline with tween; ECL, enhanced chemiluminescence; AC, allergic conjunctivitis; IQR, InterQuartile Range; EAACI, European Academy of Allergy andClinical Immunology; AAAAI, American Academy of Allergy, Asthma and Immunology; Art, Artemisia; Bet, Betulaverrucosa; Man, Mangifera indica; nsLTP, non-specificlipid-transfer protein; IPAN, Italian Pediatric AllergyNetwork; Phl, Phleum pratense; Der, Dermatophagoidespteronyssinus. ## Acknowledgments We thank the clinical and research staff and the patientparticipants for their contributions to this study. ## Funding This study was partly supported by the Science andTechnology Planning Project of Inner MongoliaAutonomous Region of China (Grant nos. 2021GG0191),Natural Science Foundation of Inner MongoliaAutonomous Region of China (2024MS08034), NaturalScience Foundation of Inner Mongolia Medical Universityof China (YKD2023MS029), Program for Young Talents ofScience and Technology in Universities of Inner MongoliaAutonomous Region (NJYT22012), Science andTechnology Program of the Joint Fund of ScientificResearch for the Public Hospitals of Inner MongoliaAcademy of Medical Sciences (2024GLLH0198). ## Data availability statement The data supporting the findings of this study are availablefrom the corresponding author upon reasonable request. ### Page 13 Volume 19, No. 5, Month 2026 ## Author contributions Jingping Fan, Chen Zhao, and Yuan Xia conceived anddesigned the study. Huifang Song and Yumin Wang wereinvolved in the clinical study of patients and conducted IgEtest. Chen Zhao, Xiaomin Wang, and Yuan Xia collectedpollens and performed the protein purification. JingpingFan, Chen Zhao, Xiaomin Wang, Qianwei Zhao, andXiaoran Wang performed Western Blot experiments anddata analysis. Jingping Fan, Huifang Song, and Chen Zhaodrafted the manuscript in close collaboration with all coauthors. Yumin Wang and Yuan Xia were joint corresponding authors. Authors’ consent for publication All authors have reviewed and approved the final versionof the manuscript and agree to its submission forpublication. Ethics approval The study was approved by the ethics committee of theInner Mongolia Maternity and Child Health Care Hospital(No.2020064) and informed consent was obtained from allpatients’ parents or guardians prior to serum collection. ## Declaration of competing interest The authors have no conflicts of interest to report. Declaration of generative artificial intelligence (AI) andAI-assisted technologies in the writing processNothing to disclose. Appendix A. Supplementary data Supplementary data to this article can be found online at. Author details aInner Mongolia Medical University, Hohhot, 010110, InnerMongolia, China.bInner Mongolia Maternity and ChildHealth Care Hospital, Hohhot, 010020, Inner Mongolia,China. REFERENCES - 1.. - 2.. - 3.. - 4.. - 5.. 13 - 6.. - 7.. - 8.. - 9.. - 10.. - 11.. - 12.. - 13.. - 14.. - 15.. - 16.. - 17.. - 18.. - 19.. - 20.. - 21.. ### Page 14 14 Fan et al. World Allergy Organization Journal (2026) 19:101391 - 22.. - 23.. - 24.. - 25.. - 26.. - 27.. - 28.. - 29.. - 30.. - 31.. - 32.. - 33.. - 34.. - 35.. - 36.. - 37.. - 38.. - 39.. - 40.. - 41.. - 42.. - 43.. - 44.. - 45.. - 46.. - 47..