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\begin{document}

\title{Pinus halepensis (Aleppo pine) nuts induced anaphylaxis: a case series}



\author[1]{Gabrielle Doré-Brabant}%
\author[1]{Joëlle Bouchard}%
\author[1]{Louis Marois}%
\author[1]{Aubert Lavoie}%
\affil[1]{Centre hospitalier universitaire de Quebec-Universite Laval}%


\vspace{-1em}



  
  \date{June 14, 2024}


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\emph{Pinus halepensis} \textbf{(Aleppo pine) nuts induced anaphylaxis:
a case series}

Gabrielle Dor\selectlanguage{ngerman}é-Brabant\textsuperscript{1}, Joëlle
Bouchard\textsuperscript{2}, Louis Marois\textsuperscript{2} and Aubert
Lavoie\textsuperscript{2}

\textsuperscript{1} Department of Pediatrics, CHU de Québec - Université
Laval, Québec, Canada.

\textsuperscript{2} Department of Allergy and Immunology, CHU de Québec
- Université Laval, Québec, Canada.

\textbf{Running title: Anaphylaxis to Aleppo pine nuts}

\textbf{Corresponding author}

Gabrielle Doré-Brabant

Department of Pediatrics, CHU de Québec, Université Laval, 2705 boul.
Laurier, G1V 4G2, Québec (Qc), Canada

E-mail: gabrielle.dore-brabant.1@ulaval.ca

\textbf{Word count:} 1072

\textbf{Number of tables} : 1

\textbf{Conflicts of interest}

All authors declare that there is no conflict of interest.

\textbf{Financial disclosure}

This work was not supported by any funding source. All authors declare
that they have no commercial and/or organizational relationships.

\textbf{Key words:} Food allergy, Anaphylaxis, Aleppo pine nuts, White
pine nuts, \emph{Pinea} nuts.

\textbf{Main text}

\emph{Pinus halepensis} , commonly known as Aleppo pine, is an evergreen
perennial tree distributed in the Northern hemisphere, especially in the
Mediterranean area. This pine, which is a medicinal plant with numerous
traditional applications, also produces seeds that can be used for
culinary purposes\textsuperscript{1}. A fermented matrix, popularly
known as ``zgougou'' and consisting of a watery mixture of ground Aleppo
pine seeds, is traditionally consumed once a year during religious
celebrations, particularly in Tunisia\textsuperscript{2}.\emph{Pinus
pinea} , a member of the same subfamily of pine as \emph{Pinus
halepensis} (both are in the Gymnosperms category) produces the pignoli
nut, or white pine nut\textsuperscript{1}, an edible pine seed known for
its various uses and commonly used in numerous countries. Allergic
reactions to white pine nuts are well documented as mostly severe
allergic reactions and high monosensitization
rates\textsuperscript{3,4}. Surprisingly, allergy to Aleppo pine nuts,
or zgougou, has never been described, either in monosensitization or in
cross-reactivity with white pine nuts. In this article, we present a
case series of three Tunisian immigrant patients living in Québec
(Canada) who presented anaphylaxis to Aleppo pine nuts.

Patient one is a five-year-old boy who developed rhinorrhea, angioedema,
generalized urticaria, rapidly progressing respiratory difficulties and
vomiting after his second spoonful of zgougou soup. He was treated with
intramuscular epinephrine in an emergency department, leading to
symptoms resolution within a few hours. All ingredients of the home-made
soup, except for zgougou, and all nuts other than pine nuts were
tolerated after this reaction. A year earlier, the last time the boy ate
zgougou, he experienced swollen lips, conjunctival erythema, and
urticaria. His symptoms were relieved with cetirizine alone. Skin
prick-to-prick testing showed a positive response to pure zgougou paste
and ground white pine nuts and serum-specific IgE for white pine nuts
were also positive {[}Table~1{]}. The parents were almost certain their
boy had tolerated white pine nuts between the two anaphylactic reactions
to zgougou. Considering the discrepancy between the history and the
positive skin test, an oral food challenge with white pine nuts was
conducted and resulted in an anaphylactic reaction (urticaria, sneezing,
nasal discharge, vomiting), which was treated with epinephrine.

Patient two is a 16-month-old girl who developed cough, dyspnea and
vomiting within 10 minutes following ingestion of mixed nuts including
white pine nuts. Minutes after, she developed swollen lips and
urticaria. At the emergency department, she received two doses of
epinephrine, diphenhydramine and prednisolone for recurrence of rash and
dyspnea 30 minutes after the first dose of epinephrine. Two months
later, she presented dyspnea and vomiting twice within 10 minutes of
zgougou ingestion. Angioedema and urticaria occurred minutes later. She
received one dose of epinephrine, which resulted in rapid resolution of
symptoms. Skin prick and prick-to-prick testing {[}Table 1{]} showed a
positive response to white pine nut extract, ground white pine nuts and
pure zgougou paste. As well, skin prick testing showed a response of 4
mm for cashews extract, 2 mm for pistachios extract and negative for
other nuts extracts (walnut, pecan, almond and hazelnut). Serum-specific
IgE for white pine nuts were positive {[}Table 1{]}. Serum-specific IgE
for almonds, hazelnuts, cashews, Brazil nuts, macadamia nuts, pecans,
and pistachios were all negative. An oral food challenge with cashews
was conducted and was well-tolerated, confirming an isolated allergy to
white pine nuts.

Patient three is a four-year-old girl who developped swollen lips,
peribuccal erythema, sneezing, vomiting, urticaria and dysphagia within
minutes of ingestion of pesto (pistachio and white pine nuts). She was
rapidly treated with diphenhydramine and her symptoms were resolved on
arrival at the emergency room, except for persistant sneezing. She did
not receive epinephrine. Two months later, she ate two spoonfuls of
zgougou and presented cough, respiratory distress and dysphagia. She
received a total of four doses of epinephrine, methylprednisolone,
diphenhydramine and was observed overnight in the emergency room. Skin
prick and prick-to-prick testing showed a positive response to white
pine nut extract and pure zgougou paste and serum-specific IgE for white
pine nuts were positive {[}Table 1{]}.

These three cases involved IgE-mediated severe anaphylactic reaction to
Aleppo pine nuts, which prompted an allergological assessment. All three
children were Tunisian immigrants and presented with typical clinical
manifestations of anaphylaxis and showed a positive skin test to the
zgougou product. To our knowledge, no other cases of Aleppo pine nut
allergy have been published. In addition, severe anaphylaxis reaction
has been described in relation to other pine nuts from the same
subfamily, namely \emph{Pinus pinea} \textsuperscript{3,4}.

Each patient also had an anaphylactic reaction to white pine nuts, with
positive skin test, and positive serum-specific IgE for white pine nuts
suggesting a cross-reactivity between Aleppo pine nuts and white pine
nuts. We hypothesize that this immunological cross-reactivity is due to
the substantial homology these two nuts share, being from the same pine
subfamily. In fact, pine seed allergens have not been well described
and, specifically, allergenic proteins from \emph{Pinus halepensis} have
never been studied. So far, only Pin p 1 from the white pine nut has
been identified. This allergen is recognized by 75\% of patients
presenting with an allergic reaction to \emph{Pinus pinea} , indicating
that other proteins might be implicated\textsuperscript{5}. In addition,
cross-reactivity has been found between different pine nuts, namely, the
Korean pine nut (\emph{Pinus koraiensis)} \textsuperscript{6} and the
Swiss stone pine nut (\emph{Pinus cembra)} \textsuperscript{7}. Of note,
only 29\% of the serum of patients allergic to white pine nuts
recognizes the Korean pine vicilin, Pine k 2\textsuperscript{6}.
Cross-reaction with other tree nuts (angiosperms) is normally low or
absent, showing that pine nut allergy is characterized by a high degree
of monosensitization. All three patients show monosensitization to pine
nuts, which is consistent with the results presented in other
publications\textsuperscript{3,4}.

In summary, this is the first report of IgE-mediated allergy to Aleppo
pine nuts. Further studies to characterize pine nut allergens,
especially Aleppo pine nut allergens, would be important to better
understand cross-reactivity among the different pine nuts and their
distinct clinical presentation. Allergists and patients must be aware
that Aleppo pine nut allergy exists, and we believe that other pine
nuts, if part of the local diet, should be tested to assess possible
cross-reactivity. This article also stresses the importance for
allergists to be sensitive to cultural differences in nutrition and to
look for hidden allergens to counsel their patients. Finally, it would
be interesting to compare the prevalence of this allergy among immigrant
and local populations, to assess the impact of diet on the development
of pine nut allergy.

Gabrielle Doré-Brabant, MD,

Joëlle Bouchard, MD,

Louis Marois, PhD, MD, FRCPC and

Aubert Lavoie, MD, FRCPC

CHU de Québec - Université Laval, Québec, Canada

\textbf{Acknowledgements}

We are grateful to the patients' parents for authorizing use of the data
related to this article.

\textbf{Impact statement}

Our article presents the first cases of IgE-mediated allergy to Aleppo
pine nuts with cross-reactivity with white pine nuts. We believe that
this article should be of interest to both current and future
generations of allergists, firstly to acknowledge the existence of this
allergy and secondly to consider the cross-reactivity between Aleppo
pine nuts and white pine nuts.

\textbf{References}

1. El Omari N, Fatima Ezzahrae G, El Menyiy N, Benali T, Aanniz T,
Chamkhi I, et al. Phytochemical and biological activities of Pinus
halepensis mill., and their ethnomedicinal use. J Ethnopharmacol
{[}Internet{]}. 2021;268(August 2020):113661. Available from:
https://doi.org/10.1016/j.jep.2020.113661

2. Missaoui J, Saidane D, Mzoughi R, Minervini F. Fermented seeds
(``Zgougou'') from aleppo pine as a novel source of potentially
probiotic lactic acid bacteria. Microorganisms. 2019;7(12).

3. Cabanillas B, Cheng H, Grimm CC, Hurlburt BK, Rodríguez J, Crespo JF,
et al. Pine nut allergy: Clinical features and major allergens
characterization. Mol Nutr Food Res. 2012;56(12):1884--93.

4. Cabanillas B, Novak N. Allergic reactions to pine nut: A review. J
Investig Allergol Clin Immunol. 2015;25(5):329--33.

5. Cabanillas B, Crespo JF, Maleki SJ, Rodriguez J, Novak N. Pin p 1 is
a major allergen in pine nut and the first food allergen described in
the plant group of gymnosperms. Food Chem. 2016;210:70--7.

6. Zhang Y, Du W-X, Fan Y, Yi J, Lyu S-C, Nadeau KC, et al.
Identification, characterization, and initial epitope mapping of pine
nut allergen Pin k 2. Food Res Int. 2016 Dec;90:268--74.

7. Jansen A, Vermeulen A, Dieges PH, van Toorenenbergen AW. Allergy to
pine nuts in a bird fancier. Allergy. 1996 Oct;51(10):741--4.



\textbf{Table}\selectlanguage{english}
\begin{longtable}[]{@{}lllll@{}}
\toprule
\begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Table 1 Investigation of patients\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Table 1 Investigation of patients\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Table 1 Investigation of patients\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
Table 1 Investigation of patients\strut
\end{minipage} & \begin{minipage}[b]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\midrule
\endhead
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Patient\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{One}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Two}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Three}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\textbf{Three}\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
Skin prick-to-prick test, mm Zgougou paste\\
Ground white pine nut\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
11 7\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
8 10\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
19 NA\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
19 NA\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Skin prick test, mm White pine nut extract\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
NA\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
10\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
14\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
14\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
Total IgE, kUI/L N\textless{}60\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
157\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
587\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
581\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
581\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
White pine nut specific IgE, kUA/L N\textless{}0.35\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
1.73\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
1.07\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
49\strut
\end{minipage} & \begin{minipage}[t]{0.19\columnwidth}\raggedright\strut
49\strut
\end{minipage}\tabularnewline
\begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\emph{Skin test readings taken at 15 min were evaluated by comparing the
wheal induced by the offending food and by a positive control of 10
mg/ml of histamine and a negative control of saline. A positive response
was interpreted when the induced wheal was [?] 3 mm compared to the
negative control.} \emph{NA : not available}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\emph{Skin test readings taken at 15 min were evaluated by comparing the
wheal induced by the offending food and by a positive control of 10
mg/ml of histamine and a negative control of saline. A positive response
was interpreted when the induced wheal was [?] 3 mm compared to the
negative control.} \emph{NA : not available}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\emph{Skin test readings taken at 15 min were evaluated by comparing the
wheal induced by the offending food and by a positive control of 10
mg/ml of histamine and a negative control of saline. A positive response
was interpreted when the induced wheal was [?] 3 mm compared to the
negative control.} \emph{NA : not available}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\emph{Skin test readings taken at 15 min were evaluated by comparing the
wheal induced by the offending food and by a positive control of 10
mg/ml of histamine and a negative control of saline. A positive response
was interpreted when the induced wheal was [?] 3 mm compared to the
negative control.} \emph{NA : not available}\strut
\end{minipage} & \begin{minipage}[t]{0.17\columnwidth}\raggedright\strut
\strut
\end{minipage}\tabularnewline
\bottomrule
\end{longtable}

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