f443 Ana o 3

Summary

Ana o 3 is a water-soluble, 2S albumin, seed-storage protein in cashew nuts (Anacardium occidentale), having a molecular weight of 14 kDa. The prevalence of cashew nut allergy is widespread across the globe, with Ana o 3 being the major allergen component of cashew nut. The sensitization of Ana o 3 is responsible for severe systemic reactions and anaphylaxis in cashew-allergic individuals. This protein is resistant to gastric and intestinal digestion by pepsin and trypsin. The immunoreactivity of Ana o 3 is reduced by thermal processing methods, like blanching, autoclaving or roasting (10 min), however it may increase when subjected to high temperature processing (170℃ for 20 min). Further, Ana o 3 can serve as a marker for primary sensitization to cashew nuts. Also, monosensitization to Ana o 3 without sensitization to other food allergens could be the most potent predictor of severe anaphylaxis. Extensive cross-reactivity is reported among cashew nuts and pistachio due to cross-reactivity among their 2S albumins i.e. Ana o 3 and Pis v 1 (pistachio). Besides, Ana o 3 is also cross-reactive with Jug r 1 (walnut) and to some extent with Cor a 14 (hazelnut). 

Epidemiology

Worldwide distribution

Allergy to cashew nuts (Anacardium occidentale) is a typical immunoglobulin (IgE)-mediated allergic disorder, being reported from the United States, Australia and Europe and sparsely from Asia and Africa. Ana o 3 is considered as one of the major allergens of cashew nut belonging to the prolamin superfamily of proteins (1).

A study conducted in Germany found that 81% of sera obtained from cashew-allergic patients were reactive to Ana o 3 on immunoblot analysis (2). Another study performed in Germany reported increased levels of serum-specific immunoglobulin E (sIgE) towards Ana o 3 in 69% of cashew-allergic patients (42 out of 61), while in only 5% of cashew sensitized but tolerant patients (1 out of 19) (3). Furthermore, Ana o 3-IgE sensitization was found in 27% of cashew and peanut-sensitized individuals in a study in Finland (26 out of 98) (4).

In a Japanese study conducted on 95 patients with suspected cashew allergy, sensitization to Ana o 3 (based on sIgE levels) was found in 100% (n=26) of cashew-allergic patients and in 51% (n=69) of tolerant, but cashew-sensitized patients (5). 

Environmental Characteristics

Source and tissue

Ana o 3 is a major allergen component extracted from the seed of cashew tree. It is mainly a water-soluble, seed storage protein (1).

The sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) or high- performance liquid chromatography (HPLC) are some methods used for purification of natural Ana o 3 from defatted cashew nut extract (6). It was further characterized by mass spectrometry, followed by circular dichroism (7). The molecule can also be generated by recombinant systems in Escherichia coli (2). 

Clinical Relevance

Disease severity

Ana o 3 is one of the major allergens of cashew nut and is responsible for severe systemic reactions and anaphylaxis in allergic individuals (1). In a study conducted on 227 children with severe allergic reactions or anaphylaxis, Ana o 3 (Ana o 3) was found as the most frequent cause of severe anaphylaxis (14.3%). Furthermore, the severe anaphylaxis rate with systemic reactions was found to be 84.6%, due to Ana o 3 (8).

A study performed among 100 Greek children reported recombinant Ana o 3-sIgE sensitization in 93% of cashew-allergic children and in only 6% of cashew-tolerant children (9).

Sensitization to Ana o 3 was found in all (100%) of the cashew-allergic patients based on oral food challenge (OFC), while it was found in only 51% of cashew-tolerant patients. The OFC-positive patients depicted moderate-to-severe reactions, which included gastrointestinal reactions (73%) as most common, followed by oral mucosa (69%), skin reactions (50%), respiratory reactions (38%) and cardiovascular reactions (15%) (5).

Cross-reactive molecules

Cross-reactivity between cashew and pistachio has been widely reported and this is attributed to their homologous seed storage proteins. A study conducted on 2 groups of patients evaluated the cross-sensitization profiles of the cashew-sensitized patients among Anacardiaceae family members (pistachio, mango and pink peppercorn). The first group (n=7) consisted of patients having co-sensitization to only Anacardiaceae species, while the second group (n=11) consisted of patients with co-sensitization to other tree nuts in addition to Anacardiaceae species. It was reported that, in both the patient groups, the 2S albumins of cashew nut (Ana o 3) and pistachio (Pis v 1) were identified on the western blots (10).

A study conducted in Germany reported that 87% of cashew-allergic patients sensitized to Ana o 3 also suffered from walnut allergy (13 out of 15), and this may be due to cross-reactivity between Ana o 3 and Jug r 1 (2).

Co-sensitization between cashew nut and hazelnut allergy has been reported in a large study conducted in the US, however, the correlation between the sIgE values of Ana o 3 and Cor a 14 was found to be low (correlation coefficient=0.43). Furthermore, a low-to-moderate correlation was reported between the 2S albumin of cashew nut (Ana o 3) and 11S globulin of hazelnut (Cor a 9) (correlation coefficient=0.50) (11). Another study observed that the cross-reactivity was found to be rare between the 2S albumins of cashew nut and hazelnut (Ana o 3 and Cor a 14 respectively) (12).

Molecular Aspects

Biochemistry

Ana o 3 is part of 2S albumin of the prolamin superfamily of proteins. These 2S albumins are small, water-soluble, globular proteins that consist of 2 subunits: large - 8 to 10 kDa and small -3 to 4 kDa, connected via four disulfide linkages. They possess large content of asparagine, arginine, cysteine and glutamine residues. Furthermore, they are found in abundance in the seed cells and are helpful during germination due to their ability to donate nitrogen and sulfur (1).

Ana o 3 has a primary structure of 138 amino acid with a molecular weight of 14 kDa (1). The recombinant protein may not be able to properly replicate the microheterogeneity and post-translational modifications of native protein (6).

Ana o 3 is comparatively stable within a pH of 1-11, while it becomes completely undetectable at pH 13. Moreover, reduced immunoreactivity is observed when it is subjected to thermal processing methods, like blanching, autoclaving or roasting (10 min), however, the allergen has shown to have resistance to high-temperature processing (170℃ for 20 min). The immunoreactivity may increase rather than reducing through such processing methods (1). Ana o 3 is also resistant to gastric and intestinal digestion by pepsin and trypsin (7). However, treatment with high concentrations of sodium sulphite (≥50 mM) and high temperature (≥100℃) could reduce the immunoreactivity of the allergen Ana o 3 (1). It was also found to resist lactic fermentation used during the process of preparing yogurt (13).

Isoforms, epitopes, antibodies

Ana o 3.0101 is the major isoform Ana o 3 (1).

Cross-reactivity

A high degree of cross-reactivity has been observed between cashew nuts and pistachios, due to structural similarity between their homologous proteins. A similarity in amino acid sequences of Ana o 3 with its homologous 2S albumin protein i.e. Pis v 1 (pistachio) has been found to be 66% (14).

The sequence identity between 2S albumin of cashew (Ana o 3) and hazelnut (Cor a 14) has been reported to be 43% (12). Further, Ana o 3 IgE-epitope was found to overlap with the corresponding epitope of walnut allergen Jug r 1, that showed 81% sequence homology with Ana o 3 (2).

Diagnostic Relevance

Disease Severity

A study conducted on 227 children with systemic allergic reactions or anaphylaxis reported that sensitization to Ana o 3 can display good predictive ability for identifying cashew nut allergies, however this sensitization has shown to increase the risk of severe anaphylaxis in cashew-allergic individuals. The study further suggested that monosensitization to Ana o 3 without sensitization to other food allergens could be the most potent predictor of severe anaphylaxis (8).

In a study conducted on 100 Greek cashew-allergic children, Ana o 3-sIgE levels of 0.1 kUA/l displayed a diagnostic sensitivity and specificity of 98% and 94%, respectively, thus proving it to be of high diagnostic value (9). Further, a study performed on 61 cashew-sensitized subjects in Germany, reported differentiating ability of Ana o 3-sIgE between clinically allergic and tolerant children to cashew nuts. The predictive ability of Ana o 3-sIgE (95% at 2.0 kUA/l) for clinically relevant cashew nut allergy was found to be significantly higher (p<0.005) than sIgE for whole cashew extract having AUC (area under the curve) value of 0.9 as compared to 0.7 for cashew extract (3).

In a prospective, multicenter study performed on 173 children with suspected cashew nut allergy, sIgE to Ana o 3 was able to better discriminate between clinically allergic or tolerant individuals in comparison with sIgE to whole cashew extract or skin prick test (15).

Further, a Japanese study conducted on 95 cashew-sensitized children also confirmed the predictive ability of Ana o 3 sensitization for clinically relevant cashew nut allergy based on OFC. The study reported a clinical sensitivity and specificity of 69% and 96%, respectively for Ana o 3-sIgE levels of 2.2 kUA/l (5).

Cross-reactivity

A study conducted to isolate and purify cashew allergens: Ana o 1, Ana o 2 and Ana o 3, have reported cross-reactivity between Ana o 1 and Ana o 3. Ana o 1 was able to inhibit the IgE-binding to Ana o 3, and to lesser extent with Ana o 2 (6).

Exposure

The main route of exposure is through ingestion (1).

Compiled By

Author: Turacoz Healthcare Solutions

Reviewer: Dr. Fabio Iachetti

Last reviewed: January 2021