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Background: Cannabis allergy (CA) has mainly been attributed to Can s 3, the nsLTP (non-specific lipid transfer proten) of Cannabis sativa. Nevertheless, standardized diagnostic tests are lacking and research on CA is scarce. Objective: To explore the performance of five cannabis diagnostic tests and the phenotypic profile of CA. Methods: 120 CA patients were included and stratified according to the nature of their cannabis-related symptoms, 62 healthy and 189 atopic controls were included. Specific (s)IgE hemp, sIgE and BAT rCan s 3, BAT with a crude cannabis extract and a skin prick test (SPT) with a nCan s 3-rich cannabis extract were performed. Clinical information was based on patient-history and a standardized questionnaire. Results: Firstly, up to 72% of CA reporting likely-anaphylaxis (CA-A) are Can s 3 sensitized. Actually, the Can s 3-based diagnostic tests show the best combination of positive and negative predictive values; 80% and 60%, respectively. sIgE hemp displays 82% sensitivity but only 32% specificity. Secondly, Can s 3+CA reported significantly more cofactor mediated reactions and displayed significantly more sensitizations to other nsLTPs than Can s 3-CA. Finally, the highest prevalence of systemic reactions to plant-derived foods was seen in CA-A, namely 72%. Conclusions: The most effective and practical tests to confirm CA are the SPT with a nCan s 3-rich extract and the sIgE rCan s 3. Can s 3 entails a risk of systemic reactions to plant-derived foods and cofactor-mediated reactions. However, as Can s 3 sensitization is not absolute, other cannabis allergens probably play a role.
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Original Article
Exploring the Diagnosis and Profile of Cannabis
Allergy
Ine Ilona Decuyper, MD
a,b
, Athina Ludovica Van Gasse, MD
a
, Margaretha A. Faber, MD, PhD
a
, Jessy Elst, MSc
a
,
Christel Mertens, MLT
a
, Hans-Peter Rihs, PhD
c
, Margo M. Hagendorens, MD, PhD
a,b
, Vito Sabato, MD, PhD
a
,
Hilde Lapeere, MD, PhD
d
, Chris H. Bridts, MLT
a
, Luc S. De Clerck, MD, PhD
a
, and Didier Gaston Ebo, MD, PhD
a
Antwerp, Ghent, Belgium; and Bochum, Germany
What is already known about this topic? Cannabis allergy, although rare, can manifest with severe and generalized
symptoms and has been linked to Can s 3, the nonspecic lipid transfer protein present in Cannabis sativa.
What does this article add to our knowledge? This article compares the performance of multiple cannabis diagnostic
methods and explores clinical and in vitro characteristics of cannabis allergy in one of the largest cannabis allergic
populations described up until now.
How does this study impact current management guidelines? There are no guidelines available on cannabis allergy
diagnosis or management. This articles perspective on diagnostic performances could aid in accurately approximating
post-test probabilities and gives insight into the prole of Western European cannabis allergic patients.
BACKGROUND: Cannabis allergy (CA) has mainly been
attributed to Can s 3, the nonspecic lipid transfer protein
(nsLTP) of Cannabis sativa. Nevertheless, standardized
diagnostic tests are lacking and research on CA is scarce.
OBJECTIVE: To explore the performance of 5 cannabis
diagnostic tests and the phenotypic prole of CA.
METHODS: A total of 120 patients with CA were included and
stratied according to the nature of their cannabis-related
symptoms; 62 healthy and 189 atopic controls were included.
Specic IgE (sIgE) hemp, sIgE and basophil activation test
(BAT) with a recombinant Can s 3 protein from Cannabis sativa
(rCan s 3), BAT with a crude cannabis extract, and a skin prick
test (SPT) with an nCan s 3-rich cannabis extract were
performed. Clinical information was based on patient history
and a standardized questionnaire.
RESULTS: First, up to 72% of CA reporting likely-anaphylaxis
(CA-A) are Can s 3 sensitized. Actually, the Can s 3-based
diagnostic tests show the best combination of positive and
negative predictive values, 80% and 60%, respectively. sIgE
hemp displays 82% sensitivity but only 32% specicity.
Secondly, Can s 3DCA reported signicantly more cofactor-
mediated reactions and displayed signicantly more sensitiza-
tions to other nsLTPs than Can s 3-CA. Finally, the highest
prevalence of systemic reactions to plant-derived foods was seen
in CA-A, namely 72%.
CONCLUSIONS: The most effective and practical tests to
conrm CA are the SPT with an nCan s 3-rich extract and the
sIgE rCan s 3. Can s 3 sensitization entails a risk of systemic
reactions to plant-derived foods and cofactor-mediated reactions.
However, as Can s 3 sensitization is not absolute, other cannabis
allergens probably play a role. Ó2018 Published by Elsevier
Inc. on behalf of the American Academy of Allergy, Asthma &
Immunology (J Allergy Clin Immunol Pract 2018;-:---)
Key words: Cannabis allergy; Diagnosis; BAT; Specic IgE; Skin
prick test; Can s 3; nsLTP; Cofactor; Basophil; Anaphylaxis;
Hemp
Cannabis is one of the most consumed drugs worldwide.
1
Despite its widespread use, reports on cannabis allergy (CA)
remain rare and generally deal with relatively small numbers of
cases.
2-6
Nevertheless, from these reports evidence is accumu-
lating that CA can manifest with severe and generalized
a
Department of Immunology-Allergology-Rheumatology, University Hospital of
Antwerp, University of Antwerp, Antwerp, Belgium
b
Department of Pediatrics, University Hospital of Antwerp, University of Antwerp,
Antwerp, Belgium
c
IPAInstitute for Prevention and Occupational Medicine, German Social Accident
Insurance, Ruhr-University Bochum, Bochum, Germany
d
Ghent University Hospital, University of Ghent, Ghent, Belgium
This work was supported by the Agency for Innovation by Science and Technology
(grant number 140185). Furthermore, V. Sabato is a senior clinical researcher of
the Research Foundation Flanders/Fonds Wetenschappelijk Onderzoek (FWO:
1804518N). D. G. Ebo is a senior clinical researcher of the Research Foundation
Flanders/Fonds Wetenschappelijk Onderzoek (FWO: 1800614N). A. L. Van
Gasse is a fellow of the Research Foundation Flanders/Fonds Wetenschappelijk
Onderzoek (FWO: 1113617N).
Conicts of interest: The authors declare that they have no relevant conicts of
interest.
Received for publication July 17, 2018; revised August 17, 2018; accepted for
publication September 15, 2018.
Available online --
Correspondence author: Didier Gaston Ebo, MD, PhD, Department of Immunology,
Allergology, Rheumatology, Faculty of Medicine and Health Science, University
of Antwerp, Campus Drie Eiken T5.95, Universiteitsplein 1, 2610 Antwerp,
Belgium. E-mail: immuno@uantwerpen.be.
2213-2198
Ó2018 Published by Elsevier Inc. on behalf of the American Academy of Allergy,
Asthma & Immunology
https://doi.org/10.1016/j.jaip.2018.09.017
1
Abbreviations used
BAT- Basophil activation test
CA- Cannabis allergy
CA-A- Likely-anaphylaxis to cannabis
CA-C- Cutaneous symptoms to cannabis
CA-R- Respiratory symptoms to cannabis
CA-RC- Localized respiratory and cutaneous symptoms to
cannabis
CBA- Cytometric bead array
CS- Cannabis sativa
HC- Healthy controls
NPV- Negative predictive value
nsLTP- Nonspecic lipid transfer protein
PþLTP- Atopic pollen-sensitized participants without an nsLTP
sensitization
PþLTPþ- Atopic pollen and nsLTP-sensitized participants
PPV- Positive predictive value
rCan s 3- Recombinant Can s 3 protein from CS
sIgE- Specic IgE
SPT- Skin prick test
symptoms and a variety of cross-reactive plant-derived food
allergies, mainly attributed to a Can s 3 sensitization, the
nonspecic lipid transfer protein (nsLTP) from Cannabis sativa
(CS). As a matter of fact, in some European surveys, Can s 3 has
been demonstrated to be a major allergen.
7-9
NsLTPs are heat
stable allergens widely distributed throughout the plant kingdom
and showing extensive in vitro and in vivo cross-reactivity.
10
Both
the severe phenotype and the extensive cross-reactivity associated
with CA can be attributed to the physiochemical properties of
Can s 3. Other putative cannabis allergens are ribulose-1,5-
bisphosphate carboxylase/oxygenase, oxygen-evolving enhancer
protein 2, and a thaumatin-like protein.
2,4
However, unlike Can
s3,
3
these allergens have not yet been successfully isolated nor
expressed as a recombinant protein and are currently unavailable
for diagnosis.
So far, in the majority of studies on CA, diagnosis is docu-
mented by prick-prick tests with buds or leaves
4-6,9
and therefore
are difcult to standardize, because of the heterogeneous
composition of the different source materials. The clinical
severity and cross-reactivity of CA together with the unpredict-
ability of the source materials used for skin testing constitute
strong incentives for more reliable cannabis diagnostic tests,
in vitro or in vivo.
In 2 preliminary studies, we have standardized and presented
initial performance results using 4 different cannabis diagnostic
tests, namely, a basophil activation test (BAT) with recombinant
Can s 3 protein from CS (rCan s 3), a BAT with a crude CS
extract, a skin prick test (SPT) with an nCan s 3-rich extract, and
nally, a specic IgE (sIgE) rCan s 3 assay using a cytometric
bead array (CBA) technique. These diagnostic tests were
compared with sIgE industrial hemp by uorescence enzyme
immunoassay (FEIA) ImmunoCAP. All 4 of our diagnostic tests
have been found reliable in diagnosing CA
7,8
and revealed Can s
3 sensitization in up to 75% of patients with CA with an
anaphylaxis-like phenotype. Alternatively, the sIgE hemp assay,
albeit displaying an excellent sensitivity, was shown to be poorly
reliable because of an important proportion of clinically irrele-
vant positive results in cannabis-tolerant individuals sensitized to
pollen and/or nsLTPs.
Importantly, for robust validation purposes, our recent study
8
was restricted to patients with an anaphylaxis-like phenotype on
cannabis exposure. However, in general practice, physicians
might frequently encounter patients with less compelling
histories such as isolated respiratory symptoms and in whom Can
s 3 sensitization seems less predominant.
3
Therefore, this study
investigates the diagnostic test performances and intertest
differences between these 5 diagnostic tests in a larger study
population expressing distinct clinical phenotypes on cannabis
exposure. Secondly, this study explores the clinical and molecular
characteristics of CA; the sensitization proles, the severity of
cross-reactivities with other plant-derived foods, and the signi-
cance of cofactors, as patients presenting with nsLTP-related
allergies have frequently been reported to necessitate a cofactor
to become symptomatic.
11,12
METHODS
Inclusion
Patients and controls were included through the outpatients
clinic of Allergology at the Antwerp University Hospital and the
Dermatology department of the Ghent University Hospital,
Belgium. The local ethics committees of both hospitals approved this
study (B300201524055), and patients or their representatives signed
an informed consent in accordance with the Declaration of Helsinki.
Patients with respiratory, gastrointestinal, cardiovascular, and/or
cutaneous symptoms on exposure to cannabis were included.
Exposure to cannabis was dened as active smoking, ingestion, and/
or direct cutaneous contact with cannabis. Patients with generalized
symptoms in 2 or more organ systems were categorized as likely-
anaphylactic according to the criteria dened by Sampson et al.
13
Furthermore, 2 distinct control groups were included: rst,
healthy controls (HC) without pollen or nsLTP sensitization;
secondly, a so-called atopic control group comprising patients with a
documented pollen allergy with (PþLTPþ) or without nsLTP
(PþLTP) sensitization. Controls were further stratied according
to exposure and tolerance to cannabis, that is, uneventful exposure.
Denitions of pollen and nsLTP sensitizations are shown in this
articles Online Repository at www.jaci-inpractice.org.
Information on CA, cofactor-associated reactions (reported plant-
derived food allergies with a history of overt or more severe/gener-
alized reactions in the presence of nonsteroidal anti-inammatory
drugs, alcohol, or physical exercise than when the reaction
occurred in the absence thereof), and severity of plant-derived food-
associated reactions was gathered by history taking and a standard-
ized questionnaire. Three cofactors were dened in this study: the
use of alcoholic beverages, nonsteroidal anti-inammatory drugs,
and/or the performance of exercise within 3 hours preceding
occurrence of an allergic reaction. A systemic reaction was dened as
grade 1 or higher as dened by the World Allergy Organization
criteria of systemic allergic reactions.
14
Patients with chronic
spontaneous urticaria, uncontrolled asthma, eosinophilic esophagi-
tis/colitis, or systemic mastocytosis were excluded.
Diagnostic tests
Basophil activation test. BAT with rCan s 3 and a crude CS
extract were performed, as detailed in this articles Online Repository
at www.jaci-inpractice.org and previously validated as described in
detail elsewhere.
8
Results were expressed as net percentages of
CD63
þ
basophils, calculated by subtraction of the spontaneous
expression from the allergen-induced CD63 expression. A result
J ALLERGY CLIN IMMUNOL PRACT
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2DECUYPER ET AL
>5% CD63
þ
basophils was considered positive as dened by
previous validation.
8
Total and specific IgE. Total and sIgE to industrial hemp,
rBet v 1 and rBet v 2 from birch (Betula verrucosa), rPhl p 1 and rPhl
p 5b from timothy grass (Phleum pratense), nArt v 1 and nArt v 3
from mugwort (Artemisia vulgaris), rAra h 9 from peanut (Arachis
hypogeae), rCor a 8 from hazelnut (Corylus avellana), rMal d 3 from
apple (Malus domesticus), rJug r 3 from walnut (Juglans regia), rPru
p 3 from peach (Prunus persica), rPar j 2 from wall pellitory (Pari-
etaria judaica), and nAna 2 c from bromelain (Ananas comosus), as a
marker for sensitization to cross-reactive carbohydrate determinants,
were quantied by the FEIA ImmunoCAP technique (Thermo-
Fisher Scientic, Uppsala, Sweden) according to the manufacturers
instructions. All sIgE assays are readily available, except for industrial
hemp, which is available for research use only and was kindly pro-
vided by ThermoFisher Scientic. Specic IgE to rCan s 3 was
quantied using a ow CBA technique (BD Biosciences, Franklin
Lakes, NJ). The method was validated as previously described.
8
Results 0.10 kU
A
/L were considered positive.
Skin prick tests. SPT implied an nCan s 3-rich CS extract that
was prepared as described elsewhere.
7
SPT responses were read after
15 minutes and considered positive when the wheal exceeded 3 mm
(largest diameter). A positive control with histamine (10 mg/mL)
and a negative saline control without allergen (ALK-Abello Ltd,
Berkshire, United Kingdom) were performed to rule out non-
responsiveness or dermographism of the skin, respectively.
Statistical analysis
IBM SPSS version 24.0 (IBM, Chicago, Ill) software was used for
data analysis. Data are expressed as medians and interquartile ranges.
Nonparametric tests and
c
2
analysis were used where appropriate.
Test performances were compared by using McNemars test. Where
needed, missing values were imputed by using a multiple-imputation
model with 5 imputations based on all available information that
were subsequently pooled in SPSS. Signicance levels for the pooled
imputed data were calculated according to the method described by
Schafer et al.
15
APvalue of <.05 was regarded as statistically
signicant.
RESULTS
Demographics
As shown in Figure 1, a total of 371 individuals were
included; 120 patients with symptoms on cannabis exposure
(CA) of which 21% (n ¼25) were classied as likely-
anaphylactic (CA-A), 19% (n ¼23) presented with mild and
localized respiratory and cutaneous symptoms (CA-RC), 51%
reported isolated respiratory symptoms (CA-R), and 9% report
isolated cutaneous symptoms (CA-C). The remaining 251 par-
ticipants were control individuals, either HC or atopics with a
pollen sensitization (PþLTPþ), with or without nsLTP sensi-
tizations (PþLTP). As displayed in Figure 1, 50% to 60% of
each control group reported the regular use of cannabis in the
past 12 months without any symptoms apart from the known
psychoactive effects; the other half reported no previous contact
with cannabis. All patients with CA displayed symptoms during
active smoking, except for 3 patients denying any previous direct
contact with cannabis (no active smoking, ingestion, or cuta-
neous contact) but who had experienced symptoms on passive
exposure to cannabis smoke. Furthermore, in total 34 patients
with CA reported respiratory and/or cutaneous symptoms on
n = 371
CA
n = 120
likely-
anaphylaxis
n = 25
respiratory
symptoms
n = 61
cutaneous
symptoms
n = 11
localized
respiratory and
cutaneous symptoms
n = 23
HC
n = 62
CS exposed
n = 37
not CS exposed
n = 25
P+
n = 90
CS exposed
n = 47
not CS exposed
n = 43
P+LTP+
n = 99
CS exposed
n = 49
not CS exposed
n = 50
FIGURE 1. Inclusion overview. CA, Cannabis allergic patients; CS,Cannabis sativa;HC, healthy controls; PþLTP, pollen-sensitized
controls without an nsLTP sensitization; PþLTPþ, pollen and nsLTP-sensitized controls.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 3
isolated passive exposure to cannabis smoke apart from symp-
toms on active smoking. Finally, 4 patients also reported
symptoms on ingestion of cannabis processed as space cake,
cannabis seeds or oil, resulting in anaphylaxis in 2 of the cases.
The individual symptoms reported by CA-A are shown in
Table E1 in this articles Online Repository at www.jaci-
inpractice.org. In summary, 23 of 25 reported respiratory
symptoms and/or cutaneous symptoms, 4 patients also
mentioned cardiovascular symptoms comprising palpitations
and/or hypotension, and nally, 5 patients additionally reported
gastrointestinal symptoms comprising abdominal pain, nausea,
and vomiting.
Table I displays demographic data of the different study
groups revealing similar age, sex ratios, and asthma prevalence in
all groups. In contrast, atopic dermatitis and elevated total IgE
values were signicantly more prevalent in the PþLTPþgroup
than in the CA group and in the PþLTPgroup. Total IgE was
also signicantly higher in the PþLTPgroup compared with
the CA group. Finally, importantly, 84% of patients with CA
showed a pollen sensitization and 72% an nsLTP sensitization.
It is important to note that pollen sensitization was predomi-
nated by Bet v 1; 72% of CA sensitized) and 79% of PþLTPþ
exhibited a Bet v 1 sensitization.
Performance of cannabis diagnostic tests
Figure 2 shows the individual results of 5 different cannabis
diagnostic tests: the sIgE industrial hemp, sIgE rCan s 3 CBA,
SPT with an nCan s 3-rich extract, and the BAT with both rCan
TABLE I. Demographic data
HC [62 CA [120 PDLTPL[90 PDLTPD[99
Age (y), median (Q
25
-Q
75
) 28.3 (24.8-36.1) 29.2 (25.1-35.2) 28.8 (22.9-37.7) 29.9 (20.1-37.1)
Sex
(% male) 42% 48% 37% 49%
Eczema*0% 37% 37% 54%
Asthma5% 30% 28% 39%
Total IgE (kU/L), median (Q
25
-Q
75
) 16.7 (6.0-46.5) 247.4 (83.0-495.0) 126.0 (65.0-314.0) 424.5 (147.0-1054.0)
Pollen sensitizationz0% 84% 100% 100%
NsLTP sensitizationx0% 72% 0% 100%
CA, Cannabis allergy; CS,Cannabis sativa;HC, healthy controls; PþLTPþ, pollen and nsLTP-sensitized controls; PþLTP, pollen-sensitized controls without an nsLTP
sensitization; nsLTP, nonspecic lipid transfer protein.
*According to patient recollection and recent use of topical CS.
According to patient recollection.
zAt least one of the following sIgEs 0.1 kU
A
/L: rBet v 1, rBet v 2, nArt v 1, rPhl p 1, rPhl 5b.
xAt least one of the following sIgEs 0.1 kU
A
/L: rPru p 3, rMal d 3, rJug r 3, rAra h 9, rCor a 8, nArt v 3, rPar j 2, rTri a 14.
FIGURE 2. Individual test results. A, Dot plots showing healthy controls (HC), pollen and nsLTP-sensitized controls (PþLT P þ), pollen-
sensitized controls without an nsLTP sensitization (PþLT P ), and cannabis allergy (CA). B, Dot plots for the different CA groups:
CA-A, CA-RC, CA-R, and CA-C. Percentages reflect the proportion of positive results and horizontal lines represent group mean.
Dindicates patients with <15% response to anti-IgE stimulation (nonresponders). Of 371, 55 (15%) were classified as nonresponders:
15 HC, 12 PþLT P ,14PþLT P þ, and 14 CA. BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CA-C, cutaneous
symptoms to cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and cutaneous symptoms to cannabis;
sIgE, specific IgE; SPT, skin prick test.
J ALLERGY CLIN IMMUNOL PRACT
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4DECUYPER ET AL
s 3 and a crude cannabis extract. Table II compares the test
performances. For more details on the difference in test perfor-
mance for sIgE rCan s 3 and sIgE hemp (considering 0.10 or
0.35 kU
A
/L cutoff), the reader is referred to Figure E1 and
Table E2 in this articles Online Repository at www.jaci-
inpractice.org.
First of all, test performances showed important variances
between the different clinical CA groups. The 3 Can s 3-based
diagnostic methods (BAT, sIgE, and SPT) displayed a similar
sensitivity, 63% to 72% in CA-A (45% to 58% in the total CA
group), and a similar specicity (81% to 87% in the total CA
group). However, up to 37% (n ¼34) of PþLTPþshowed
clinically irrelevant Can s 3 sensitizations (measured by BAT,
sIgE, or SPT): 20 of 34 reported tolerance to active cannabis use
and 14 of 34 reported no previous cannabis contact. In com-
parison, the sIgE rCan s 3 and BAT rCan s 3 showed no clini-
cally irrelevant positive results in pollen-sensitized individuals
without nsLTP sensitizations (PþLTP).
Secondly, the sIgE industrial hemp displayed a signicantly
higher sensitivity, up to 82% (P<.01) in the total CA group
compared with the Can s 3-based diagnostic tests (45% to 58%).
However, sIgE hemp also demonstrated a signicantly higher
number of clinically irrelevant positive results in PþLTPand
PþLTPþ, that is, 51% to 82% respectively compared with 0%
to 25% for the Can s 3 diagnostic tests (all P<.01). Interest-
ingly, an increase in sensitivity as seen in the sIgE hemp was not
found in the BAT with a crude cannabis extract. The latter
reached an overall sensitivity of 49% in the total CA group that
was not superior to the Can s 3-based assays. In addition, the
BAT with the crude extract was not superior to the Can s 3
diagnostic tests in terms of specicity either, showing 19% to
38% of clinically irrelevant positive results in PþLTPand
PþLTPþ. Collectively, for all diagnostic techniques, the
majority of clinically irrelevant results were seen in the PþLTPþ
group.
In summary, when all different clinical CA groups are
considered (analysis B in Table II), it appears that the 3 Can s
3-based diagnostic tests did not signicantly differ in perfor-
mance and had the best combined positive predictive value
(PPV) and negative predictive value (NPV) of around 80% and
60%, respectively. The sIgE industrial hemp lacked specicity,
whereas the BAT crude CS extract showed no advantage over the
Can s 3-based diagnostic tests.
The clinical and molecular characteristics of
CA. Figure 3 compares different clinical and in vitro charac-
teristics for the different CA proles and the control groups. The
most prominent differences were found between CA-A and
CA-R with signicantly higher numbers of Pru p 3, Mal d 3, Cor
a 8, Jug r 3, Tri a 14, Art v 3 sensitizations (all P<.01) in CA-A
than in CA-R. Furthermore, CA-A showed a higher prevalence
of systemic reactions to plant-derived foods (72% compared with
40%, P¼.02) and cofactor-mediated allergic reactions (50%
compared with 18%, P¼.01) compared with CA-R. In addi-
tion, CA-C and CA-RC showed a single difference from CA-A,
namely a considerably lower prevalence of systemic reactions to
plant-derived foods (71% in CA-A compared with 43% in
CA-RC [P<.01] and 18% in CA-C [P¼.08]). It appears that
none of the clinical nor in vitro parameters displayed signicant
differences between CA-R, CA-C, and CA-RC.
Regarding the comparison of Can s 3-sensitized and non-
sensitized CA (as demonstrated in Table E3 in this articles
Online Repository at www.jaci-inpractice.org), it became clear
that Can s 3þCA had a signicantly higher prevalence of other
nsLTP sensitizations (92%) than Can s 3-CA (39%) with higher
frequencies of all measured nsLTPs (all P<.01), except for Par j
2. Also, Can s 3þCA displayed higher frequencies of pollen
sensitizations than Can s 3-CA (92% compared with 74%) with
signicantly more Bet v 1 sensitizations in the Can s 3-sensitized
population. In addition, Can s 3þCA showed a considerably
higher prevalence of cofactor-mediated allergic reactions when
compared with Can s 3-CA (41% vs 12%; P<.01).
In a further analysis, the complete CA group was compared
with the PþLTPþgroup (as demonstrated in Table E4 in this
TABLE II. Test performance
sIgE hemp sIgE rCan s 3 BAT rCan s 3 BAT crude CS extract SPT nCan s 3-rich extract
A
Sensitivity 86% (66-97) 63% (41-81) 71% (48-89) 63% (38-84) 72% (51-89)
Specicity 32% (20-45) 87% (78-93) 85% (76-92) 67% (55-78) 81% (71-88)
PPV 33% (28-38) 56% (40-70) 54% (39-67) 35% (25-47) 51% (39-63)
NPV 86% (66-95) 90% (84-94) 93% (86-96) 87% (78-92) 91% (84-95)
LHRþ1.3 (1.0-1.6) 4.7 (2.6-8.7) 4.8 (2.7-8.6) 1.9 (1.2-3.1) 3.7 (2.3-6.0)
LHR0.4 (0.1-1.3) 0.40 (0.3-0.7) 0.3 (0.2-0.7) 0.6 (0.3-1.0) 0.4 (0.2-0.7)
B
Sensitivity 82% (74-89) 47% (38-56) 45% (35-55) 49% (37-60) 58% (49-67)
Specicity 32% (20-45) 87% (78-93) 85% (76-92) 67% (55-78) 81% (71-88)
PPV 70% (66-74) 82% (72-89) 78% (67-86) 64% (54-73) 80% (72-86)
NPV 47% (34-61) 56% (51-60) 57% (52-62) 52% (46-59) 58% (53-64)
LHRþ1.2 (1.0-1.5) 3.5 (2.0-6.2) 3.0 (1.8-5.2) 1.5 (1.0-2.2) 3.0 (1.9-4.7)
LHR0.6 (0.3-1.0) 0.60 (0.5-0.7) 0.7 (0.5-0.8) 0.8 (0.6-1.0) 0.5 (0.4-0.7)
A: calculations based on CA-A group vs cannabis-tolerant PþLTPand PþLTPþ.
B: calculations based on the whole CA group (respiratory and/or cutaneous symptoms) vs cannabis-tolerant PþLTPand PþLTPþ. Test performance for both BATs was
calculated by considering both responders and nonresponders to anti-IgE.
BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CS,Cannabis sativa;NPV, negative predictive value; PPV, positive predictive value; LHRþ, positive
likelihood ratio; LHR, negative likelihood ratio; PþLTPþ, pollen and nsLTP-sensitized controls; PþLTP, pollen-sensitized controls without an nsLTP sensitization; sIgE,
specic IgE; SPT, skin prick test.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 5
articles Online Repository at www.jaci-inpractice.org). This
exploration revealed a signicantly (P<.01) higher prevalence of
Can s 3 sensitizations in CA (63%) compared with PþLTPþ
(35%). Furthermore, a signicantly lower prevalence of Pru p 3,
Mal d 3, Jug r 3, Par j 2 (all P<.01) but also bromelain (P¼
.02) and Phl p 1 (P<.01) sensitizations was seen in the CA
group compared with PþLTPþ. Finally, as already mentioned in
the demographic paragraph, signicantly (P<.01) more eczema
was reported in the PþLTPþgroup than the CA group and
subsequently total IgE values were also signicantly higher in
PþLTPþthan in the CA group (P<.01). Although there was
no signicant difference between CA and PþLTPþconcerning
the frequency of systemic reactions to plant-derived foods (P¼
.11), CA-A did show double the frequency of systemic reactions
to plant-derived foods than PþLTPþ(71% vs 35%, P<.01).
DISCUSSION
To the best of our knowledge, this is the largest survey
exploring diagnostic performances in different clinical pheno-
types of CS allergy. Along with the observation that the diag-
nostic utilities of our tests depend on the clinical presentation, it
appears that the CA prole in this study population has the
following peculiarities.
Primarily, in terms of practicality, efciency, and standardi-
zation, the SPT with an nCans 3-rich extract and the sIgE rCan s
3 are the easiest and fastest tests to conrm a clinical suspicion of
CA, both equally reliable. However, because of unavailability, in
clinical practice, physicians will need to rely on other tests to
screen patients with a convincing history. As a matter of fact,
according to our data, it seems that the sIgE hemp assay (avail-
able on request from ThermoFisher) could serve as a suitable
diagnostic in central Europe to exclude CA, because a negative
test result reduces the risk of CA considerably (only 18% of CA
have negative sIgE hemp results). Alternatively, patients with a
convincing history together with a positive sIgE hemp should
undergo additional testing to elucidate the clinical signicance of
the hemp solid phase assay. In addition, exploration of different
cutoffs for the sIgE rCan s 3 and hemp shows that, when
considering the 0.35 kUa/L cutoff, sensitivity of both tests de-
creases by around 10% in the total CA population. Nevertheless,
sensitivity to detect CA-A remains the same for both. Even
though the specicity of sIgE hemp almost doubles, it still only
reaches a maximum of 60%, which is not ideal.
However, none of our diagnostic tests appear absolutely pre-
dictive for the clinical outcome. Nevertheless, for the time being,
on the basis of our ndings, we propose to perform the SPT with
an nCan s 3-rich extract or quantify sIgE rCan s 3 keeping in
mind that Can s 3 does not cover the entire IgE sensitization
prole, particularly in patients with a less severe/pronounced
phenotype. In addition, it could be questioned whether Can s 3-
negative patients, especially if reporting only milder symptoms to
cannabis, should effectively be categorized as CA, because their
symptoms could result from nonspecic skin or airway irritation.
Furthermore, because of ethical and legal limitations, it is
impossible to conrm CA by an oral or respiratory challenge.
Considering this hypothesis, it follows that the actual test per-
formances are possibly underestimated in this study and that Can s
3 might even play a more prominent role than already suspected.
Furthermore, it is likely that performances of a Can s 3 assay
display regional differences due to geographic differences in IgE
reactivity proles. The reason(s) why Can s 3 negative CA pa-
tients go undetected in the BAT with the full CS extract re-
main(s) elusive but could relate to a sensitization to allergens that
are poorly present in our crude extract or do not resist our
current extraction procedure. Moreover, the low presence and
the physicochemical properties of the constituent allergens might
also explain the different sensitization proles in the distinct
phenotypes, namely, the lower prevalence of nsLTP sensitiza-
tions in CA-R compared with CA-A.
Eczema
Asthma
Cofactor
SR to plant-foods
Pollen sensitization
Bet v 1
Bet v 2
Phl p 1
Phl p 5b
Art v 1
LTP sensitization
Pru p 3
Mal d 3
Cor a 8
Ara h 9
Jug r 3
Tri a 14
Art v 3
Par j 2
Can s 3*
Bromelain
P+LTP-
P+LTP+
CA total
CA-A
CA-R
CA-RC
CA-C
SCALE
0% 0%
10% 10%
20% 20%
30% 30%
40% 40%
50% 50%
60% 60%
70% 70%
80% 80%
90% 90%
100% 100%
FIGURE 3. Overview of clinical and in vitro parameters. Color variations represent increasing frequencies of positive results for the shown
variable. For example, frequency of asthmatics (specific IgE [sIgE] measurements are shown as percentage sensitized/not sensitized).
*Measured by BAT or sIgE rCan s 3.BAT, Basophil activation test; CA-A, likely-anaphylaxis to cannabis; CA-C, cutaneous symptoms to
cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and cutaneous symptoms to cannabis; LTP, lipid transfer
protein; OAS, oral allergy syndrome defined as localized and mild oropharyngeal symptoms without generalization; PþLTP, pollen-
sensitized controls without an nsLTP sensitization; PþLTPþ, pollen and nsLTP-sensitized controls; SR, systemic reaction defined by
generalized and severe symptoms in at least 1 organ system
14
;TOL, tolerant.
J ALLERGY CLIN IMMUNOL PRACT
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6DECUYPER ET AL
Secondly, although historically sensitization to nsLTP has
mainly been recognized to occur in the Mediterranean region,
characterized by severe reactions and governed by peach,
10,16
more
recent data have accumulated showing that sensitization to nsLTP
might also occur in other European regions and frequently remain
asymptomatic with uncertainties about the route(s) of sensitiza-
tion.
17-19
In this survey, we conrm that nsLTP sensitization
occurs frequently in CA and Can s 3 is a major allergen in patients
with CA-A, but CA also implies a risk of systemic reactions to
plant-derived foods and cofactor-mediated reactions. Furthermore,
Can s 3 sensitization can occur as a result of in vitro cross-reactivity
to nsLTPs from taxonomically related or more distant sources
such as pollen and/or plant-derived foods as suggested by the Can
s 3 positive PþLTPþpatients without any previous cannabis
contact. On the other hand, it seems that a Can s 3 sensitization in
patients with CA might also mirror a primary sensitization instead
of only in vitro cross-reactivity as indicated by the signicantly
higher prevalence of Can 3 and lower prevalence of Pru p 3, Mal
d 3, Jug r 3, and Par j 2 sensitizations in CA compared with
PþLTPþ.
Another important fact to highlight is that, because of the lack
of data on the true prevalence of CA, it is likely that the number
of patients per study group in this survey do not necessarily
reect the true prevalence of CA. Therefore, the test perfor-
mances would differ depending on the characteristics of the
tested population and the geographic prevalence of CA itself.
Finally, this study was not designed to explore the different
individual types of plant-derived food allergies, as symptoms to
different plant-derived foods were only assessed by a standardized
questionnaire complemented with a history taking without sys-
tematic conrmatory testing. However, it would be interesting to
further explore the actual differences in individual plant-derived
food allergies within CA such as the differences in symptom
severity with and without peel, the types of plant-derived foods
eliciting allergic symptoms but also the comparison of these
factors between CA and other nsLTP-sensitized individuals.
In conclusion, this study is the largest study exploring diagnostic
test performance, clinical phenotypes, and biological proles of
CA. It shows that the most effective and practical tests to conrm a
clinical suspicion of CA are the SPT with an nCan s 3-rich extract
and sIgE rCan s 3. Both tests display a PPV and NPV of approx-
imately 80% and 60%, respectively. However, because of current
unavailability, screening with sIgE hemp could be a suitable tool in
symptomatic cannabis users, because a negative result considerably
reduces the likelihood of CA. Alternatively, we dissuade the general
use of sIgE hemp to diagnose CA, mainly because of its limited
PPV. Furthermore, we show that Can s 3 is a major allergen in
patients with a history of likely-anaphylaxis on cannabis exposure
and, like other nsLTP-associated allergies, CA might indicate a risk
of systemic reactions to plant-derived foods and cofactor-mediated
reactions. Because around 30% of CA-A and even higher pro-
portions in other, milder CA groups are not sensitized to Can s 3, it
is likely that other cannabis allergens might play a role in CA.
Further studies are thus warranted to identify and express other CA
allergens that could then be applied to spike natural extracts or to
compose mixtures of allergens. Lastly, additional research should
further explore the nature of plant-derived food allergies in CA as
this study was not designed to evaluate specic plant-derived food
allergies in CA.
Acknowledgments
We thank B. Van Camp, head of the Unit drug production,
Central Drug Department, Directorate of organized crime,
Belgian Federal Judicial police for his help with providing the
necessary plant materials. We would also like to thank Mrs. K.
Vandebos and N. Maes, our study nurses, for their help in the
performance of the skin prick tests and blood sampling. Finally,
our gratitude goes out to Mrs. K Wouters, for her help and
statistical advice.
REFERENCES
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from: www.unodc.org/wdr2017. Accessed June 7, 2018.
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acterization of Cannabis sativa allergens. Ann Allergy Asthma Immunol 2013;
111:32-7.
3. Rihs HP, Armentia A, Sander I, Bruning T, Raulf M, Varga R. IgE-binding
properties of a recombinant lipid transfer protein from Cannabis sativa. Ann
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4. Larramendi CH, López-Matas M, Ferrer A, Huertas AJ, Pagán JA, Navarro LÁ,
et al. Prevalence of sensitization to Cannabis sativa. Lipid-transfer and
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Cannabis allergy: a diagnostic challenge. Allergy 2018;73:1911-4.
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Quesada J. Molecular diagnosis in cannabis allergy. J Allergy Clin Immunol
Pract 2014;2:351-2.
10. Pastorello EA, Robino AM. Clinical role of lipid transfer proteins in food
allergy. Mol Nutr Food Res 2004;48:356-62.
11. Pascal M, Munoz-Cano R, Reina Z, Palacin A, Vilella R, Picado C, et al. Lipid
transfer protein syndrome: clinical pattern, cofactor effect and prole of molecular
sensitization to plant-foods and pollens. Clin Exp Allergy 2012;42:1529-39.
12. González-Mancebo E, González-de-Olano D, Trujillo MJ, Santos S, Gandolfo-
Cano M, Meléndez A, et al. Prevalence of sensitization to lipid transfer proteins
and prolins in a population of 430 patients in the south of Madrid. J Investig
Allergol Clin Immunol 2011;21:278-82.
13. Sampson HA, Munoz-Furlong A, Campbell RL, Adkinson NF Jr, Bock SA,
Branum A, et al. Second symposium on the denition and management of
anaphylaxis: summary reportsecond National Institute of Allergy and Infec-
tious Disease/Food Allergy and Anaphylaxis Network symposium. Ann Emerg
Med 2006;47:373-80.
14. Cox LS, Sanchez-Borges M, Lockey RF. World Allergy Organization systemic
allergic reaction grading system: is a modication needed? J Allergy Clin
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15. Schafer JL. Analysis of Incomplete Multivariate Data. London: Chapman &
Hall; 1997.
16. Fernandez-Rivas M. Fruit and vegetable allergy. Chemical Immunol Allergy
2015;101:162-70.
17. Pascal M, Vazquez-Ortiz M, Folque MM, Jimenez-Feijoo R, Lozano J,
Dominguez O, et al. Asymptomatic LTP sensitisation is common in plant-food
allergic children from the Northeast of Spain. Allergol Immunopathol (Madr)
2016;44:351-8.
18. Faber MA, Van Gasse AL, Decuyper II, Uyttebroek A, Sabato V,
Hagendorens MM, et al. IgE-reactivity proles to nonspecic lipid transfer
proteins in a northwestern European country. J Allergy Clin Immunol 2017;139:
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Immunol 2016;169:181-8.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 7
ONLINE REPOSITORY
METHODS
Patient stratification. Pollen sensitization was dened by at
least 1 positive specic IgE (sIgE) result to the following re-
combinant (r) or native (n) allergens: rBet v 1 and rBet v 2 from
birch (Betula verrucosa), rPhl p 1 and rPhl p 5b from timothy
grass (Phleum pratense), and/or nArt v 1 of mugwort (Artemisia
vulgaris). Nonspecic lipid transfer protein (nsLTP) sensitization
was documented by at least 1 positive sIgE result to the following
nsLTP allergens: rAra h 9 from peanut (Arachis hypogeae), rCor a
8 from hazelnut (Corylus avellana), rMal d 3 from apple (Malus
domesticus), rJug r 3 from walnut (Juglans regia), rPru p 3 from
peach (Prunus persica), nArt v 3 from mugwort (Artemisia vul-
garis), and rPar j 2 from wall pellitory (Parietaria judaica).
Diagnostic methods. Briey, prewarmed heparinized blood
samples were stimulated with rCan s 3 (expressed in Escherichia
coli
E1
) and a crude Cannabis sativa extract (extraction method as
described elsewhere,
E2
obtained by solvent precipitation in the
presence of enzyme inhibitors). Antihuman IgE served as a
positive control (10
m
g/mL, clone G7-18, BD Biosciences,
Erembodegem, Belgium) and stimulation buffer was used to
measure spontaneous CD63 expression by quiescent cells.
Basophil activation test was performed using side scatter, anti-
IgE, and anti-CD203c (clone NP4D6, BD Biosciences) to
characterize the basophils. Subsequently, within this gate, baso-
phil activation was quantied as the percentage
CD203cþþCD63þexpressing basophils using anti-CD63
(clone H5C6, BD Biosciences). Results were expressed as net
percentages of CD63
þ
basophils, calculated by subtraction of the
spontaneous expression from the allergen-induced CD63
expression.
Results
Cannabis allergyerelated symptoms
Performance of cannabis diagnostic tests. As shown
in Figure E1 and Table E2, sIgE rCan s 3s specicity does not
change when the cutoff is increased from 0.10 to 0.35 kU
A
/L.
Specicity of sIgE hemp rises almost 2-fold (32% to 60%)
when a cutoff of 0.35 kU
A
/L is considered.
When only considering CA reporting likely-anaphylaxis (CA-
A), there is no difference in sensitivity between a cutoff of 0.10 vs
0.35 kU
A
/L for either sIgE rCan s 3 or sIgE hemp. In other
words, all sensitized CA-A have an sIgE result 0.35 kU
A
/L for
both tests. When the complete CA group is considered, sensi-
tivity decreases around 10% for both tests: sIgE hemp (82% to
72%; P<.001), sIgE rCan s 3 (47% to 38%; P¼.002).
J ALLERGY CLIN IMMUNOL PRACT
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7.e1 DECUYPER ET AL
FIGURE E1. sIgE results for sIgE hemp and sIgE rCan s 3 showing both cutoff 0.10 kU
A
/L and 0.35 10 kU
A
/L. CA-A, CA reporting likely-
anaphylaxis; CA-C, cutaneous symptoms to cannabis; CA-R, respiratory symptoms to cannabis; CA-RC, localized respiratory and
cutaneous symptoms to cannabis; HC, healthy controls; PþLTP, pollen-sensitized controls without an nsLTP sensitization; PþLTPþ,
pollen and nsLTP-sensitized controls; sIgE, specific IgE.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 7.e2
TABLE E1. Symptoms reported on cannabis exposure by CA-A
Patient Symptoms on cannabis exposure
PT1 AE, D, RC
PT2 AE, AP, C, U, W
PT3 AE, D, P
PT4 D, P, RC, U
PT5 AE, D, RC, U
PT6 AD, D, P, RC, U
PT7 AE, D, P, U, W
PT8 AE, C, D, RC
PT9 BP, OAS, RC, U
PT10 D, RC, U
PT11 AE, D, N, V
PT12 D, RC, U
PT13 D, RC, U
PT14 D, OAS, RC, U
PT15 D, RC, U
PT16 AE, D, P, PL, U
PT17 AE, D, OAS
PT18 AE, D, U
PT19 D, RC, U
PT20 OAS, V, U
PT21 D, N, PL, W
PT22 D, P, RC, U
PT23 N, U, V
PT24 AE, D, P, PL, RC, U
PT25 D, RC, P, U
AD, Atopic dermatitis air; AE, angioedema; AP, abdominal pain; BP, blood pres-
sure drop; C, cough; CA-A, CA reporting likely-anaphylaxis; D, dyspnea; N, nausea;
OAS, oral allergy syndrome; P, pruritus; PL, palpitations; RC, rhinoconjunctivitis; U,
urticaria; V, vomiting; W, wheezing.
J ALLERGY CLIN IMMUNOL PRACT
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7.e3 DECUYPER ET AL
TABLE E2. Differences in sIgE results between 0.1 and 0.35 kU
A
/L cutoffs
Cutoff
sIgE hemp
Pvalue
sIgE rCan s 3
Pvalue0.10 kU
A
/L 0.35 kU
A
/L 0.10 kU
A
/L 0.35 kU
A
/L
Sensitivity*CA-A 86 (66-97) 77 (55-92) .5 63 (41-81) 63 (41-81) 1
Sensitivity CA82 (74-89) 72 (63-80) .001 47 (38-56) 38 (29-48) .002
Specicityz32 (20-45) 60 (49-70) <.001 87 (78-93) 89 (83-93) 1
CA, Cannabis allergy; CA-A, CA reporting likely-anaphylaxis; PþLTP, pollen-sensitized controls without an nsLTP sensitization; PþLTP þ, pollen and nsLTP-sensitized
controls; sIgE, specic IgE.
*Calculations based on the CA-A group vs cannabis-tolerant PþLTPand PþLTPþ.
Calculations based on the whole CA group (respiratory and/or cutaneous symptoms) vs cannabis-tolerant PþLTPand PþLTPþ.
zCalculations based on the PþLTPand PþLTPþgroups. Pvalues calculated by McNemar analyses.
TABLE E3. Comparison of Can s 3-sensitized and nonsensitized
CA patients
Can s 3DCA Can s 3LCA Pvalue
Eczema 31% 47% .08
Asthma 30% 35% .42
Cofactor*42% 15% <.01
Systemic reaction to plant foods52% 35% .14
Pollen-sensitized individuals*92% 74% .03
Bet v 1*82% 56% <.01
Bet v 2 14% 20% .46
Phl p 1 54% 62% .45
Phl p 5b 43% 43% 1
Art v 1 17% 8% .15
nsLTP-sensitized individuals*92% 39% <.01
Pru p 3*86% 35% <.01
Mal d 3*87% 30% <.01
Cor a 8*80% 16% <.01
Ara h 9*76% 18% <.01
Jug r 3*80% 28% <.01
Tri a 14*55% 19% <.01
Art v 3*70% 10% <.01
Par j 2 9% 20% .11
Bromelain 37% 20% .07
CA, Cannabis allergy; nsLTP, nonspecic lipid transfer protein; WAO, World
Allergy Organization.
*Signicant differences as measured by P<.05.
Dened as grade 1 or higher as described by the WAO criteria of systemic allergic
reactions.
E3
TABLE E4. Comparison of CA and PþLT P þpatients
CA PDLTPDPvalue
Eczema*37% 54% .01
Asthma 30% 39% .14
Cofactor*31% 19% .05
Systemic reaction to plant foods45% 33% .11
Total IgE (mean)*247.4 424.5 <.01
Pollen-sensitized individuals*84% 100% <.01
Bet v 1 72% 79% .27
Bet v 2 16% 23% .17
Phl p 1*57% 79% <.01
Phl p 5b 43% 54% .09
Art v 1 14% 19% .32
nsLTP-sensitized individuals*72% 100% <.01
Pru p 3*66% 88% <.01
Mal d 3*65% 82% <.01
Cor a 8 56% 61% .50
Ara h 9 54% 62% .23
Jug r 3*60% 79% <.01
Tri a 14 41% 45% .54
Art v 3 47% 42% .45
Par j 2*14% 38% <.01
Can s 3*z63% 35% <.01
Bromelain*30% 45% .02
BAT, Basophil activation test; CA, cannabis allergy; nsLTP, nonspecic lipid transfer
protein; PþLTPþ, pollen and nsLTP-sensitized controls; sIgE, specic IgE; SPT,
skin prick test; WAO, World Allergy Organization.
*Signicant differences as measured by P<.05.
Dened as grade 1 or higher as described by the WAO criteria of systemic allergic
reactions.
E3
zMeasured by sIgE, BAT, or SPT.
J ALLERGY CLIN IMMUNOL PRACT
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DECUYPER ET AL 7.e4
REFERENCES
E1. Rihs HP, Armentia A, Sander I, Bruning T, Raulf M, Varga R. IgE-binding
properties of a recombinant lipid transfer protein from Cannabis sativa. Ann
Allergy Asthma Immunol 2014;113:233-4.
E2. Decuyper II, Faber MA, Lapeere H, Mertens C, Rihs HP, Van Gasse AL, et al.
Cannabis allergy: a diagnostic challenge. Allergy 2018;73:1911-4.
E3. González-Mancebo E, González-de-Olano D, Trujillo MJ, Santos S, Gandolfo-
Cano M, Meléndez A, et al. Prevalence of sensitization to lipid transfer proteins
and prolins in a population of 430 patients in the south of Madrid. J Investig
Allergol Clin Immunol 2011;21:278-82.
J ALLERGY CLIN IMMUNOL PRACT
MONTH 2018
7.e5 DECUYPER ET AL
... Sensitization to C. sativa can lead to cross-reactivity with a vaierty of foods, beverages, latex and testing, most case reports in the literature use prick-prick tests with crude cannabis or non-standardized cannabis extracts for diagnosis. 30 This approach could potentially be confounded by variable results depending on the composition of the source material or varieties of C. sativa used. A recent European study showed that Can s 3-based testing is the most effective and reliable. ...
... A recent European study showed that Can s 3-based testing is the most effective and reliable. 30 Although not commercially available, skin prick testing with a Can s 3 enriched extract and specific IgE testing to recombinant Can s 3 were both demonstrated to have positive and negative predictive value of around 80% and 60%, respectively. 30 However, whether these results apply to the North American population, which seems to have different pattern of sensitization, remains unknown. ...
... 30 Although not commercially available, skin prick testing with a Can s 3 enriched extract and specific IgE testing to recombinant Can s 3 were both demonstrated to have positive and negative predictive value of around 80% and 60%, respectively. 30 However, whether these results apply to the North American population, which seems to have different pattern of sensitization, remains unknown. Specific IgE testing to hemp, which is commercially available, can be considered a proxy although it lacks specificity (32%). ...
Article
Cannabis refers to a genus of annual, herbaceous, dioecious flowering plants that are members of the family Cannabaceae, which include about 102 plant species.Although there is much debate, the most common taxonomy is that the genus Cannabis comprises one species, Cannabis sativa L (C. sativa)., which includes the highly polymorphic subspecies sativa, and indica. Hemp and cannabis both refer to the same species C. sativa; however, there is important distinction between the two. Whereas hemp (fiber-type) is grown for its cellulose-rich fiber in the stem, cannabis (drug-type) is cultivated for its flowers where the glandular trichomes produce the psychoactive delta-9-tetrahydrocannabinol (THC) and cannabidiol (CBD). THC provides the analgesic and relaxing effects of cannabis, whereas CBD produces other effects such as antiemetic and soporific properties. Researchers have used the THC content to define C. sativa subspecies sativa as containing less than 0.3% THC in dried flowering tops of female plants and C. sativa subspecies indicaas containing ≥0.3% THC.This threshold has been used by regulatory bodies to legally differentiate hemp plants (<0.3% THC) and cannabis plants (≥0.3% THC). Canada legalized the production, distribution, sale and non-medical use of cannabis for adults in October 2018. Recent data from nation-wide surveys show that approximately 6.2 million people aged 15 or older, or 20% in this age group, reported using cannabis in the past 3 months. which represents an increase from 14% before legalization. Cannabis can be used and/or ingested in a variety of forms including capsules, oils, dried flower, vaporization and through the consumption of edibles. With the increased use of recreational cannabis in Canada, it is expected that there will be a concomitant increase in cases of cannabis hypersensitivity.
... Rhinoconjunctivitis, pruritus and contact urticaria are the most commonly reported symptoms. However, IgE-mediated CA can present with the full clinical spectrum of mast cell and/or basophil activation, including angioedema, bronchospasm, gastrointestinal symptoms and life-threatening or fatal anaphylaxis [18,19]. Recently, an employee at a cannabis processing facility packaging ground cannabis into pre-rolls suffered an asthma attack and later died in the hospital (Occupational Safety and Health Administration (OSHA); Inspection detail: 1572011.015: ...
... Can s 3, a nsLTP, is one of the most intensively studied cannabis allergens. Almost a decade ago, Can s 3 gained prominence in Europe and more recently also in the USA, where several independent studies have identified it as a major allergen [8,19,[25][26][27][28]. Furthermore, the European Academy of Allergy and Clinical Immunology (EAACI) Task Force on non-specific lipid transfer protein allergy in Europe recently reported on cannabis as a potential primary source of nsLTP sensitization causing cannabis-related allergies [12]. ...
... They may fear stigma or legal consequences, depending on local legislation. They may also find it difficult to distinguish nonallergic symptoms associated with cannabis from those due to underlying atopic status, which is present in the majority of our cases [8,19]. Although a controlled challenge test is considered the reference confirmatory diagnostic test, it can present major difficulties for obvious reasons. ...
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Purpose of the review: With increased access and decriminalization of cannabis use, cases of IgE-dependent cannabis allergy (CA) and cross-reactivity syndromes have been increasingly reported. However, the exact prevalence of cannabis allergy and associated cross-reactive food syndromes (CAFS) remains unknown and is likely to be underestimated due to a lack of awareness and insufficient knowledge of the subject among health care professionals. Therefore, this practical roadmap aims to familiarize the reader with the early recognition and correct management of IgE-dependent cannabis-related allergies. In order to understand the mechanisms underlying these cross-reactivity syndromes and to enable personalized diagnosis and management, special attention is given to the molecular diagnosis of cannabis-related allergies. Recent findings: The predominant signs and symptoms of CA are rhinoconjunctivitis and contact urticaria/angioedema. However, CA can also present as a life-threatening condition. In addition, many patients with CA also have distinct cross-reactivity syndromes, mainly involving fruits, vegetables, nuts and cereals. At present, five allergenic components of Cannabis sativa (Can s); Can s 2 (profilin), Can s 3 (a non-specific lipid protein), Can s 4 (oxygen-evolving enhancer protein 2 oxygen), Can s 5 (the Bet v 1 homologue) and Can s 7 (thaumatin-like protein) have been characterized and indexed in the WHO International Union of Immunological Sciences (IUIS) allergen database. However, neither of them is currently readily available for diagnosis, which generally starts by testing crude extracts of native allergens. The road to a clear understanding of CA and the associated cross-reactive food syndromes (CAFS) is still long and winding, but well worth further exploration.
... Cannabis may also be a cofactor of the generalized hypersensitivity reaction [15]. This raises a reverse question-what may be the cofactors causing allergic reactions to cannabis, as shown in case 5? Various factors can be considered in this context including physical exercise, alcohol, and drugs (NSAIDs, oral contraceptives). ...
... However, this does not mean that all CA patients are sensitized to Can s 3. In a study by Decuyper et al., 42 of 120 patients did not show this sensitization [15]. In this context, our case no. 1 supports the theory that other cannabis allergens may lead to sensitization. ...
... Most of the described symptoms of CA are based on case reports in the literature, and thus Table 3 summarizes the occurrence of symptoms along with their severity and treatment in patients diagnosed with CA [15,25,26,29,30,32,33]. Since the case reports were published over a few decades and different grading systems were used for the assessment of symptoms, we have set to employ two different current grading scales to unify and compare the clinical picture of anaphylaxis described in different reports. ...
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Cannabis allergy is a relatively new phenomenon described in the 1970s. Its increased frequency has been observed over the last years due to the increasing therapeutic and recreational use of cannabis-based products. Sensitization possibly leading to allergy symptoms can occur not only through the smoking of cannabis, but also through ingestion, the inhalation of pollen, or direct contact. The severity of symptoms varies from benign pruritus to anaphylaxis. There is scant information available to support clinicians throughout the entire therapeutic process, starting from diagnosis and ending in treatment. In this review, we present six cases of patients in whom molecular in vitro testing revealed sensitization to cannabis extract and/or cannabis-derived nsLTP molecules (Can s 3). Based on these cases, we raise important questions regarding this topic. The article discusses current proposals and highlights the importance of further research not only on cannabis allergy but also on asymptomatic sensitization to cannabis allergens, which may be ascertained in some percentage of the population.
... Patients with a history of immediate respiratory and/or cutaneous symptoms on cannabis exposure (CA), asymptomatic atopic cannabis users (henceforth designated as exposed atopic controls (EAC)) and asymptomatic exposed healthy controls (EHC) were included as described previously. 1 Total IgE and specific (s)IgE to hemp and recombinant (r) pollen components were quantified by ImmunoCAP (Thermo Fisher Scientific) as described elsewhere. 1 Results were considered positive if ≥0.10 kU A /L. ...
... Patients with a history of immediate respiratory and/or cutaneous symptoms on cannabis exposure (CA), asymptomatic atopic cannabis users (henceforth designated as exposed atopic controls (EAC)) and asymptomatic exposed healthy controls (EHC) were included as described previously. 1 Total IgE and specific (s)IgE to hemp and recombinant (r) pollen components were quantified by ImmunoCAP (Thermo Fisher Scientific) as described elsewhere. 1 Results were considered positive if ≥0.10 kU A /L. To depict sensitization to cannabis TLP, sera were analyzed for IgE-reactivity towards rCan s-TLP (rCs-TLP) by using ELISA as described in the Online Repository. ...
... ). As described in the Online Repository, in the pMAT, mast cells (MCs) were passively sensitized with serum from CA patients or controls (both EAC and EHC) and subsequently incubated with rCs-TLP.All participants had skin prick tests (SPTs) with aeroallergens and cannabis extract as prepared in.1 SPTs were read after 15 min and considered positive when the largest wheal diameter exceeded 3 mm. ...
... Thus, geography may dictate whether pollen LTP are important sensitising allergens in individuals sensitised and reactive to LTP in foods but evaluating sensitisation to Art v 3 and/or Pla a 3 might help to support a diagnosis of LTP allergy in some populations [23]. In addition to pollens, another potential sensitising allergen is Can s 3, the nsLTP in cannabis sativa, with resulting reactions to LTP in foods without the need for Pru p 3 sensitisation [4,36]. Sensitisation to Can s 3 seems to be a common feature of individuals with cannabis-related LTP allergy, including a cohort from North America, a continent where LTP sensitisation or allergy has not hitherto been recognised [7]. ...
... Alcohol such as wine and beer can also contain nsLTP allergens, and so may have a dual effect as both a trigger of symptoms as well as a co-factor. The same is also true for cannabis [36,40]. Other potential co-factors include sleep deprivation, fasting and infections [41]. ...
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Purpose of Review To provide an update on the diagnosis of non-specific Lipid Transfer Protein (nsLTP) allergy. Recent Findings More publications report the presence of nsLTP allergy in Northern European countries and nsLTP sensitisation in children. Individuals are more likely to have severe reactions if there is recognition of increasing numbers of LTP components. Diagnosis is problematic; not all those with nsLTP allergy will have a positive test to a peach extract containing Pru p 3, the peach nsLTP. Summary Sensitisation to nsLTP is being reported in more countries, including to the nsLTP in Cannabis Sativa in North America. Meals containing multiple nsLTP foods are more likely to be involved in co-factor reactions. Component-resolved diagnostics are superior to skin prick tests, to determine sensitisation to the individual nsLTP allergens causing symptoms and, in the future, the Basophil Activation test may best discriminate between sensitization and clinical allergy.
... This does not appear to be the case in the three cases described who all tolerated peach. Moreover, it has already been demonstrated that in a group of 120 people with Cannabis allergy, only 66% were sensitized to Pru p 3; that means there is also a percentage of allergic patients primarily sensitized to Can s 3, with absence of sIgE for Pru p 3. (30) In conclusion, these clinical cases show that in the Mediterranean region, also Can s 3 could represent the primary sensitizing allergen in a nsLTP syndrome in absence of laboratory and/or clinical manifestations of IgE-mediated reactions to peach, as already extensively reported in Northern-European Countries. ...
... Sono stati recentemente identificati alcuni pazienti con un'allergia al lattice IgE-mediata apparentemente associata alla mono-sensibilizzazione per Hev b 12 (la proteina di trasporto dei lipidi non specifica del lattice). Le ragioni principali della monosensibilizzazione all'Hev b 12 rimangono indefinite, ma potrebbero in qualche misura riguardare una sottostante allergia alla Cannabis sativa 59,60 . La diagnostica molecolare, quindi, ci aiuta, non solo a stabilire gli allergeni rilevanti nei casi di reazione allergica, ma anche le cross-sensibilizzazioni che non hanno un reale riscontro clinico. ...
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Nonostante gli sforzi profusi negli ultimi decenni per mitigare le conseguenze di questa malattia, l’allergia al lattice di gomma naturale continua a essere un problema sanitario rilevante, oltre che una delle principali preoccupazioni nell’ambiente di lavoro in molti paesi del mondo, soprattutto in quelli in via di sviluppo. Le categorie di pazienti con maggiore e frequente esposizione al lattice (come quella sanitarie e, in ambito pediatrico, i soggetti sottoposti a ripetuti interventi chirurgici, ad esempio coloro che sono affetti da spina bifida e malformazioni uro-genitali) sono a maggior rischio di sviluppare sensibilizzazione e allergia. Lo scopo di questa revisione è fornire un aggiornamento delle attuali conoscenze e raccomandazioni pratiche in termini epidemiologici, diagnostici e gestionali (incluse prevenzione e terapia) al fine di guidare un corretto riconoscimento e contenimento di questa condizione potenzialmente fatale.
... Crude cannabis extracts are often used in skin prick tests to determine cannabis-specific allergic sensitization; however, these cannabis extracts are not strictly reproducible, can be difficult to procure in many states, and positive results do not identify the causal allergen(s) involved in IgE-mediated responses. There are Can s 3-based diagnostic tests that detect specific anti-Can s 3 IgE antibodies or patient basophil activation following exposure to purified Can s 3; however, to date, there are no clinically available tests that investigate sensitization to the other four identified cannabis allergens [42,85]. ...
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Purpose of Review This review investigates occupational inhalation hazards associated with biologically derived airborne particles (bioaerosols) generated in indoor cannabis cultivation and manufacturing facilities. Recent Findings Indoor cannabis production is growing across the US as are recent reports of respiratory diseases among cannabis workers, including occupational asthma morbidity and mortality. More information is needed to understand how bioaerosol exposure in cannabis facilities impacts worker health and occupational disease risk. Summary Preliminary studies demonstrate a significant fraction of airborne particles in cannabis facilities are comprised of fungal spores, bacteria, and plant material, which may also contain hazardous microbial metabolites and allergens. These bioaerosols may pose pathogenic, allergenic, toxigenic, and pro-inflammatory risks to workers. The absence of multi-level, holistic bioaerosol research in cannabis work environments necessitates further characterization of the potential respiratory hazards and effective risk prevention methods to safeguard occupational health as the cannabis industry continues to expand across the US and beyond.
... 36,38,39 Symptoms during such activities can also reflect exposure to mold and endotoxin, which complicate diagnosis. 35,36,[40][41][42] A study of nearly 800 cases of work-related asthma in Washington state found 10 that were associated with marijuana production or law enforcement. 41 Aside from dusts and pollen from the plants, Cannabis volatile organic compounds (VOCs), described as irritants, may be part of the health response in production or handling of these plants indoors. ...
Chapter
Allergens that are important in some industrial workplaces including cannabis production and laboratory animal research units.
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Background: The sensitisation profile at molecular level in plant-food allergy is complex. Several allergens may be involved, with different potential for severe reactions. lipid transfer proteins (LTP) are considered the most relevant plant-food allergens in adults in Mediterranean countries, but less is known in children. Aim: To describe the clinical pattern and sensitisation profile of children with plant-food allergy and LTP sensitisation from Northeast Spain. Methods: Children with history of immediate reaction to plant-food(s), positive skin-prick-test to the culprit plant-food(s) and specific-IgE to plant-food LTPs were analysed. Results: 130 children were included. 69.2% (90/130) had reacted to ≥2 taxonomically unrelated plant-foods. Peach, walnut, hazelnut and peanut were most frequently involved. Reactions severity ranged from anaphylaxis (45.4%, 59/130) to oral symptoms only. Sensitisation to a particular plant-food LTP not always caused clinical symptoms with that plant-food; 69% (40/58) and 63% (17/27) of peach- and walnut-tolerant subjects had positive rPru p 3 and nJug r 3 specific IgE, respectively. 65.4% (85/130) of children were also sensitised to storage proteins, which was associated to anaphylaxis and nut allergy. However, 60% of patients without nuts/seeds allergy were sensitised to storage proteins. Specific-IgE levels to LTPs and/or storage proteins were not useful to predict allergy (vs. tolerance) to peach, walnut, peanut or hazelnut. Conclusions: Sensitisation to LTP and/or storage proteins without clear clinical significance is relatively common. Prospective longitudinal studies are required to evaluate the relevance of these silent sensitisations over time. Caution is required when interpreting the results of molecular-based diagnostic tools in clinical practice.
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Background Due to the increasing social, medical and occupational exposure to Cannabis sativa, allergic reactions grow in frequency but little is known about the IgE-reactivity of single Cannabis allergens. Methods To identify the mature peptide sequence of the lipid transfer protein (LTP) and to study the IgE-binding reactivity of a recombinant Can s 3 (rCan s 3), a cDNA was synthesized from total RNA of Cannabis sativa L. ssp. sativa cv. Kompolti leaves obtained from the botanical garden of the University Bonn. The LTP gene was amplified with a primer mix deduced from published amino acid sequences. The LTP variant was identified in five independent clones by sequencing in the pDrive vector system followed by the expression of the mature LTP in the pMAL-vector. After isolation of a soluble recombinant maltose-binding protein (MBP)-Can s 3 fusion protein, aliquots were biotinylated and coupled to Streptavidin-ImmunoCAPs. Results Sera of 16 (6 Spanish and 10 German) subjects with allergic symptoms to Cannabis were tested. Specific IgE (sIgE) values >0.35 kUA/L were regarded as positive. Twelve out of 16 sera (75%) showed sIgE to Cannabis (range: 0.42-31.80 kUA/L). Five of them (31%) displayed sIgE to rCan s 3 (range: 0.40-14.10 kUA/L) but no sIgE to MBP. Specific IgE-reactivity to Pru p 3 was detected in all Cannabis-positive sera from Spain but only in 3 out of 6 German sera. All sera with sIgE to rCan s 3 showed also sIgE to Pru p 3. Conclusions These results show for the first time IgE-binding of a recombinant Cannabis allergen (rCan s 3). Due to the 60% amino acid identity between Can s 3 and Pru p 3 a cross-reactivity is possible, but also to the LTPs of other plants. Since rCan s 3 is now available for sIgE diagnostics, implementation in larger studies may help to further elucidate the impact of this allergen.
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There is no universally accepted grading system to classify the severity of systemic allergic reactions (SARs), including anaphylaxis. Although a consensus definition for anaphylaxis was established in 2005, the signs and symptoms required to define a reaction as anaphylaxis are inconsistently applied in research and clinical practice. As a result, it is difficult to compare and evaluate safety outcomes in surveys, clinical practice and trials, and pharmacovigilance data. In 2010, the World Allergy Organization (WAO) proposed a uniform grading system to classify allergen immunotherapy SARs. The basis of the grading system is the organ system(s) involved and reaction severity. The final grade is determined by the physician/health care professional after the event is over. Although the 2010 WAO grading system was developed to classify allergen immunotherapy SARs, with appropriate modifications, it can be used to classify SARs from any cause. The purpose of this Rostrum is to present a proposed modification of the 2010 WAO SAR grading system that will make it applicable to all SARs due to any cause. The modified grading system allows for classification of less severe SARs, which may be underreported or overreported in clinical trials and surveillance studies, depending on the criteria specified for adverse event reporting. The universal use of the proposed modified SAR grading system will allow for better safety comparisons across different venues and treatment protocols.
Article
Background: Plant food allergies associated with lipid transfer protein (LTP) have been widely described in the Mediterranean Basin. Objective: The aim of this work was to describe the clinical profile and pollen sensitization of plant food- allergic patients sensitized to LTP in a non-Mediterranean area. Methods: Patients with clear IgE-mediated symptoms associated with plant foods and a positive skin prick test (SPT) to Pru p 3 were included in a prospective study in the north of Spain. Reported symptoms were analyzed together with a battery of food and pollen SPTs and specific IgE components by ISAC microarray. Cross-inhibition studies were performed by ImmunoCAP with plane tree, mugwort and rPru p 3. Results: Among the 72 patients included, the most frequent food allergy reported was to peaches (69%) followed by nuts (walnuts 55%, peanuts 54% and hazelnuts 43%). Most patients suffered from symptoms with multiple plant foods (a median of 6 foods per patient). Regarding the patients' pollen sensitization, 36% were sensitized to mugwort pollen (72% showing sIgE to Art v 3), 33% to grass pollen and 24% to plane tree pollen (94% with sIgE to Pla a 3). Inhibition studies showed that specific IgEs against mugwort and plane tree pollen are inhibited by Pru p 3 in a strong manner, whereas Pru p 3 was less inhibited by pollen extracts. Conclusions: LTP syndrome occurs in a non-Mediterranean area and is related to multiple sensitizations to foods and pollens such as plane tree and mugwort. In these pollen sensitizations, Pru p 3 seems to be the primary sensitizer.
Article
Fruit and vegetable allergies are the most prevalent food allergies in adolescents and adults. The identification of the allergens involved and the elucidation of their intrinsic properties and cross-reactivity patterns has helped in the understanding of the mechanisms of sensitisation and how the allergen profiles determine the different phenotypes. The most frequent yet contrasting fruit and vegetable allergies are pollen-food syndrome (PFS) and lipid transfer protein (LTP) syndrome. In PFS, fruit and vegetable allergies result from a primary sensitisation to labile pollen allergens, such as Bet v 1 or profilin, and the resulting phenotype is mainly mild, consisting of local oropharyngeal reactions. In contrast, LTP syndrome results from a primary sensitisation to LTPs, which are stable plant food allergens, inducing frequent systemic reactions and even anaphylaxis. Although much less prevalent, severe fruit allergies may be associated with latex (latex-fruit syndrome). Molecular diagnosis is essential in guiding the management and risk assessment of these patients. Current management strategies comprise avoidance and rescue medication, including adrenaline, for severe LTP allergies. Specific immunotherapy with pollen is not indicated to treat pollen-food syndrome, but sublingual immunotherapy with LTPs seems to be a promising therapy for LTP syndrome. © 2015 S. Karger AG, Basel.
Article
Background: Although allergy to Cannabis sativa was first reported over 40 years ago, the allergenicity has scarcely been studied. The objectives of this study were to investigate the frequency of sensitization to this plant, to analyze the clinical characteristics and allergenic profile of sensitized individuals and to identify the allergens involved. Methods: Five hundred and forty-five individuals in Spain attending allergy clinics with respiratory or cutaneous symptoms underwent a skin-prick test (SPT) with C. sativa leaf extract. The extract was characterized by SDS-PAGE and 2-dimensional electrophoresis. Specific IgE to C. sativa was measured in positive SPT individuals. The clinical and allergenic profiles of sensitized individuals were investigated and the most-recognized allergens sequenced and characterized by liquid chromatography-mass spectrometry/mass spectrometry. Results: Of this preselected population, 44 individuals had positive SPT to C. sativa (prevalence 8.1%). Prevalence was higher in individuals who were C. sativa smokers (14.6%). Two individuals reported mild symptoms with C. sativa. Twenty-one individuals from 32 available sera (65.6%) had positive specific IgE to C. sativa. Twelve sera recognized at least 6 different bands in a molecular-weight range of between 10 and 60 kDa. Six of them recognized a 10-kDa band, identified as a lipid transfer protein (LTP) and 8 recognized a 38-kDa band, identified as a thaumatin-like protein. Conclusions: There is a high prevalence of sensitization to C. sativa leaves. The clinical symptoms directly attributed to C. sativa were uncommon and mild. The sensitization profile observed suggests that C. sativa sensitization may be mediated by two mechanisms, i.e. cross-reactivity, mainly with LTP and thaumatin-like protein, and exposure-related 'de novo' sensitization.