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Allergenicity to worldwide
invasive grass Cortaderia selloana
as environmental risk to public
health
Fernando Rodríguez1*, Manuel Lombardero‑Vega2*, Lucía San Juan3, Leticia de las Vecillas1,
Sofía Alonso4, Eva Morchón1, Diego Liendo5, Marta Uranga6 & Alberto Gandarillas3,7*
Allergies to grass pollen aects about 20% of the population worldwide. In the last few decades, the
South American grass Cortaderia selloana (CS, Pampas grass) has expanded worldwide in a variety
of countries including the USA, Australia and Western Europe. In many of these locations, CS has
strikingly spread and has now been classied an invasive species. Many pernicious consequences of
CS have been reported for local biodiversity, landscape and structures. However, the eect on human
health has not been studied. To investigate this issue, we have chosen a European region on the
northern cost of Spain where CS spread is overwhelming, Cantabria. We obtained CS pollen extract
and analysed the allergenic reaction of 98 patients that were allergic to pollen of local grasses. We
determined the skin reaction and the presence of specic IgE antibodies (sIgE) to CS or to a typical
autochthonous grass, Phleum pratense. We also compared the seasonal symptoms with reported
grass pollen counts in the area. The results strongly suggest that CS can cause respiratory allergies at
a similar extent to the local grasses. Given that CS pollinises later than the local grasses, this would
extend the period of grass allergies in the region for about three months every year, as stated by most
of the patients. This is the rst study reported on the eects of the striking expansion of CS on human
health. Considering the strong impact that respiratory allergies have on the population, our results
suggest that CS can currently constitute a relevant environmental health issue.
Grass pollen is one of the main causes of respiratory allergies worldwide and the rst cause in North America and
Europe, with estimated 20% of the population aected1. Cortaderia selloana (CS) is a grass of the Poaceae family,
of the Danthonioideae subfamily, commonly known as Pampas grass and native to South America. However, in
the last few decades CS was introduced in a wide diversity of countries worldwide including the USA, Australia
and Western Europe2. In these locations, CS has strikingly spread, and it is classied as an invasive species.
Within Europe, France, Great Britain, Portugal and Spain are strongly colonised. e United States Department
of Agriculture, in a report of 2014, stated: ’Cortaderia selloana obtained a relatively high impact potential risk
score because it impacts natural, anthropogenic, and production systems’3. For this reason, it has been forbid-
den to commercialise, plant or maintain in a variety of countries. One of such countries is Spain, where CS has
intensively spread along the northern cost including the regions of Galicia, Asturias, Cantabria and the Basque
Country4,5–7. First report mentioning Cortaderia in Spain are from 1953 in Cantabria8.
e allergic incidence of CS is unknown. CS has been referred to as a danger to autochthonous species,
strongly aecting biodiversity and landscape. Moreover, it is sporadically mentioned in some venues and dis-
cussion groups as a danger to humans, because of material machinery damage and health, such as cuts due to
the sharp nature of its leaves, or allergic reactions in contact with the skin9–11. However, despite the striking
expansion of the grass in regions where it is not autochthonous, there are no studies on the impact on human
health so far reported worldwide.
OPEN
1Allergy Service, Hospital Universitario Marqués de Valdecilla (HUM), Valdecilla 25, 39008 Santander, Spain. 2CMC
R&D Department, ALK-Abelló S.A., Miguel Fleta 19, 28037 Madrid, Spain. 3Institute for Research Marqués de
Valdecilla, Ave Herrera Oria SN, 39011 Santander, Spain. 4Allergy Service, Sierrallana Hospital, Torrelavega,
Spain. 5Department of Plant Biology and Ecology, University of the Basque Country (UPV/EHU), Bilbao,
Spain. 6Asociación Cantabria Sin Plumeros, Liaño, Spain. 7INSERM, Occitanie, Montpellier, France. *email:
fernando.rodriguezf@scsalud.es; Manuel.LombarderoVega@alk.net; agandarillas@idival.org
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CS has strongly colonised extensive areas of Cantabria, a typical Northern Spanish region (Fig.1A,B) of about
500,000 inhabitants, most signicantly during 1990–2008, a period of intensive road and house building6,12. CS
has mainly invaded the coast but it also has reached the inland mountains (Fig.1B–F). e plant has spread
by human activities. It is used in motorways to retain the road slope soil and as a natural barrier13,14 and it is
transported with construction aggregates and gravel from stone quarries. erefore, it is consistently found next
to roads, new buildings or small paths covered with gravel and is abundant around stone quarries (Fig.1C–F).
Plans for limiting and eradicating the growth of this invasive plant have been debated in the local parliament
due to pressure of ecologist organisations although only limited programmes were implemented. Currently,
the European Union is funding a regional network for ghting the inland expansion of the grass and diusing
Figure1. Cortaderia selloana (CS) has strongly invaded northern Spain. (A) Current spread of CS worldwide
(yellow/orange spots). From GBIF.org, GBIF Home Page. Available from: https:// www. gbif. org/ speci es/ 27045
19. (B) Le: location of Cantabria region in Spain (le). Right: current spread of CS in the region is striking, not
only on the coast but also inland (blue line). Source: LIFE Stop Cortaderia, http:// stopc ortad eria. org/ langu age/
en/ early ‐warni ngnet work/. (C–F) Representative photographs of the overwhelming presence of CS in Cantabria
region, on the northern coast of Spain. CS has spread near the coast, next to motorways and new house
buildings (C) but is also notorious inland, next to newly constructed areas (D) and even in discrete locations at
the mountains, where gravel has been used on small paths (E). Stone quarries where the gravel is transported
from, are frequently surrounded by CS (F).
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the rapidly increasing problem among the European society (3.5 million euros for 2018–202215). However, the
presence of CS on the northern cost of Spain is still overwhelming.
Cantabria is a good paradigmatic territory to investigate the allergenic eects of CS on the human population.
In Cantabria, autochthonous grass pollens peak from April to July1,16,17, when they cause a concomitant peak of
hay fever. It is estimated that the percent of the population suering from grass-associated hay fever in Cantabria
is about 19% of patients diagnosed of rhinoconjunctivitis and 14% of asthmatic patients18. In contrast to the
autochthonous grasses, CS in the North of Spain ourishes from mid August to October 5. Grass pollens of the
Pooideae subfamily, the main grasses found in temperate climates of the North Hemisphere, contain proteins
with similarities in their antigens19,20. We questioned whether patients allergic to the autochthonous grass pol-
len (Phleum pratense, Phl, as representative species) might also be allergic to CS pollen. Allergenic molecules
of groups 1 and 5 (Phl p1 and Phl p5) are main antigens inducing allergies due to their high capacity to bind to
immunoglobulin IgE of the human immune system. To investigate this issue, we analysed the skin reaction to
Phl and CS extracts of 98 patients of Cantabria that were allergic to local grass pollen. In addition, we determined
the presence of specic IgE antibodies (sIgE) to Phl and CS pollen extracts and to the single allergens Phl p 1,
Phl p 5, Phl p 7 and Phl p 12 in blood serum. We also compared the seasonal symptoms with reported grass
pollen counts. e results very strongly suggest that CS is a signicant cause of respiratory allergies, at a similar
extent as the local grass. is might thus extend the period of respiratory allergies in the region for more than
three months every year. is is the rst study reported on the eects of the striking expansion of CS on human
health and it has implications in all the regions of the world where CS has become a widespread invasive grass.
Considering the implications that respiratory allergies have on health, not only by the direct eects but also
by allowing opportunist infections, our results suggest that CS can constitute a signicant public health issue.
is risk must be added to the ecological impact, in order to encourage eorts for eradicating CS from invaded,
non-autochthonous regions.
Materials and methods
Setting. is study was conducted in Cantabria, a region of the North coast of Spain.
Design and patients. A cross-sectional study with prospective data collection was performed at the
Allergy Services of the Marqués de Valdecilla University Hospital in Santander and the Sierrallana Hospital in
Torrelavega (Cantabria, Spain).
98 patients diagnosed of rhinoconjunctivitis, asthma or both, caused by sensitization to grass pollen, were
included in a sequential way from October 2015 to March 2016.
Written informed consent was obtained from all patients before entering the study. e study met the prin-
ciples of the 1975 Helsinki declaration and was reviewed and approved by the local Research Committee of
Cantabria (CEIC reference number 2015.207).
A serum sample was obtained from each patient and stored at –20°C until used.
Pollen extract preparation. All methods were performed in accordance with the relevant guidelines and
regulations.
Cortaderia selloana (CS) pollen was obtained commercially (Iber-Polen, Jaén, Spain) and then extracted at a
1:10 (w/v) ratio in PBS pH 6.5 with magnetic stirring for 90min. at 5°C. e soluble fraction was separated by
centrifugation. Aer dialysis against PBS, the extract was ltered through 0, 22µm lters. Protein content was
determined by Bradford method (BioRad, Hercules, CA, USA). Two dierent batches were obtained (07 and
09) with consistent results.
Part of the extract was adjusted to 0.25mg protein/ml and formulated in PBS with 50% glycerol, phenol 0.51%
(SPT buer). e remaining extract was stored in aliquots at −20°C.
Phleum pratense (Phl) pollen extract was made as described for CS. e origin of the pollen in this case was
ALK Source Materials, Post Falls, Idaho, USA.
e protein proles of the CS or the Phl extracts were determined by polyacrylamide electrophoresis in the
presence of sodium dodecyl sulphate (SDS-PAGE) under reducing conditions (Invitrogen-Novex tricine gels
10–20% acrylamide, Fisher Scientic, SL, Madrid Spain).
Skin prick test. Patients were skin prick tested (SPT) with a commercial extract (ALK-Abelló, S.A. Madrid,
Spain) of Phl and the CS extract. Histamine dihydrochloride solution (10mg/ml) and SPT buer were used as
positive and negative control(no reaction), respectively.
e SPT wheal areas were measured by planimetry. A cut-o area of 7 mm2 (about 3mm average diameter)or
higher was considered a positive test result(histamine).
e CS extract was tested in 10 control subjects, that were not sensitised to grass pollen, with negative
result(no reaction).
IgE assays. Serum samples were tested for IgE antibodies against Phleum pratense (Phl) pollen extract and
the allergens Phl p 1, Phl p 5, Phl p 7 (polcalcin) and Phl p 12 (prolin) (ImmunoCap FEIA, ermo Fisher
Scientic, Barcelona, Spain).
In addition, specic IgE against Phl and CS pollen extracts was determined by RAST (Radio Allergo Sorb-
ent Test). Paper discs were activated with CNBr and sensitised with the pollen extracts as described by Ceska
etal.21. Phl and CS discs were incubated overnight with 50 µL of the patient’s serum and aer washing (0.1%
Tween-20 in PBS), with approximately 100,000cpm of the iodine 125–labeled anti-IgE mAb HE-2 for 3h as
described22. Finally, the discs were washed, and their radioactivity was determined in a gamma counter. sIgE
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values in kilounits per litre were determined by interpolating in a standard curve built up with Lolium perenne—
sensitised discs and 4 dilutions of a serum pool from patients with grass allergy, which was previously calibrated
in arbitrary kU/l.
A cut-o value of 0.35 kU/l was considered positive for both ImmunoCap and RAST. ere was a very sig-
nicant correlation between the sIgE against Phl determined by both methods (r Spearman = 0.8874, p < 0.0001).
RAST inhibition assay. Paper discs were sensitised as above in the IgE assays section and then incubated
with 50 µL of a serum pool from all patients combined. 50 µL of (inhibitory) CS extract solution (in serial dilu-
tions) were added onto the paper discs and incubated overnight at room temperature. All other incubations were
performed as indicated above in the IgE assays section. e % of inhibition was determined for each extract
dilution by radioactive counts (cpm) and calculated by means of the following equation:
Cpmx corresponds to the mean radioactivity of the discs incubated with inhibitor at a given X dilution.
cpm100% corresponds to the blank control samples of the assay (no serum pool added). cpm0% corresponds to
the signal obtained with no inhibitor extract added.
Results
To investigate whether patients allergic to the local pollen react to CS pollen, we chose a cohort of 98 patients
from Cantabria. Table1 shows the demographic and clinical characteristics of the patients. All of them were
diagnosed with rhinitis during the spring season and grass pollen sensitisation. In addition to nasal symptoms,
98% had also associated conjunctivitis, 31.6% suered from asthma and 8.2% from urticaria. Only 12.2% had
food allergies and 2 out of 98 drug allergies. 53.06% of the patients underwent grass pollen immunotherapy.
76.5% of the patients referred living in areas with high presence of CS. 78.6% of patients presented a worsening
of their pollen allergic symptoms from August to November (“delayed reactivation”). In addition, 56.12% of the
cohort were polysensitised including other pollens such as Plantago spp. (18/98), trees (9/98), Parietaria spp.
(6/98), animal dander (11/98) or house dust mites (38/98).
CS pollen extract is not commercially available to run skin prick tests or sIgE determination. erefore, we
isolated and prepared a CS pollen extract by a standard extraction protocol used for pollens (see Materials and
Methods). e yield protein/pollen was about 50mg/g, a typical concentration obtained for other grass pollens
(our own unpublished data). Grass-specic ELISA assays showed that the CS extract did not contain group 5
antigen, as expected for a non-Pooideae subfamily grass (< 0.3µg group 5/mL23). e prole of the protein extract
by SDS-PAGE shows a group of 25–37 kD bands with the mobility of the grass group 1 allergens and it might
correspond to the homologous CS group 1 (arrow, Fig.2A;19,24).
Isolated CS pollen extract was used on cutaneous tests on the patient cohort, in parallel with Phl pollen
extract, as a representative of the local autochthonous grass pollens. All 98 patients gave a positive response by
skin prick test to Phl pollen extract and 89% of the patients were also positive to CS pollen extract (Table1).
Moreover, there was a signicant correlation between the area of the papule to Phl and to CS (rPearson = 0.2558,
p = 0.01; Fig.2B). As a control, 10 patients negative for skin reaction to Phl were found also negative for CS
extract. ese results show a strong coincidence in the cutaneous reaction to CS and to the local grass. To further
study the interspecies cross reaction of the patient sera, we run by RAST (radio allergo sorbent test) inhibition
assays. As shown in Fig.2C, Phl extract signicantly competed with CS extract to bind the serum sIgE from the
patients.
Supplementary TableI displays the results of sIgE masurement. We determined sIgE to Phl and to the aller-
gens Phl p 1, Phl p 5, Phl p 7 and Phl p 12 by ImmunoCap (ermo Fisher) and to CS by RAST. All patients had
serum sIgE to Phl by both ImmunoCap and RAST, in agreement with the skin prick test results. We determined
the correlation between both techniques in detecting the sIgE for Ph. e relation was rSpearman = 0.8874,
p < 0.0001. Values obtained by RAST were below those obtained by ImmunoCAP (factor = 0.36) and the linear
range for RAST (0.17–27) was shorter than for ImmunoCAP (0.35–100). Nevertheless, the correlation between
both techniques was good, indicating that the sIgE data obtained by ImmunoCap can be compared with the
sIgE data obtained by RAST (Supplementary Fig.1). All patients but seven contained sIgE specic to CS extract.
Interestingly, within the seven patients with a negative sIgE test to CS, 5 displayed a negative skin response to CS
and the other 2 displayed a weal smaller than 14 mm2. erefore, there was a strong correlation between the skin
response and the sIgE to CS in serum (Fisher’s exact test, p < 0.0001; Supplementary TableII).
We measured the presence of sIgE to the individual allergens Phl p 1, Phl p 5, Phl p 7 and Phl p 12 in the sera
from the patients (Supplementary TableI). For the pan-allergens Phl p 7 (polcalcin) and Phl p 12 (prolin), only
27 patients (27.5%) had sIgE to any of them. Consequently, the patient sensitisation to these allergens cannot
explain the high cross-sensitisation to Phl and CS in this group of patients. e prevalence of sIgE to Phl p 1 was
very high (98%) and only two patients (# 45 and 83) were negative for IgE to Phl p 1. Consistently, these patients
also displayed a negative skin response to CS extract. e prevalence of Phl p 5 was lower but still important
(72%). Twenty-seven patients of the cohort displayed no IgE to Phl p 5 in serum. However, of these, only ve
patients were negative for skin response to the CS extract. ere was a signicant linear regression between the
sIgE to the whole Phl extract and the sIgE to Phl p 1 (Fig.3A) or Phl p 5 (Fig.3B). From the slope of the regression
line, we can conclude that every allergen accounts for about 50% of the total IgE response to the whole extract,
being the IgE-response to Phl p 1 slightly higher. e reaction to Phl p 1 plus Phl p 5 is similar to the reaction to
whole Phl extract (Fig.3C), strongly suggesting that groups 1 and 5 are the main allergens of Phl and they account
for most of the IgE to the whole Phl extract. ere is a signicant correlation between thesIgE to CS extract and
thesIgE to Phl whole extract, toPhl p1 or toPhl p5 (Table2). e correlation is stronger for the whole extract or
100
×
1−
cpmx −cpm100%
/
cpm0% −cpm100%
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Patient Age (years) SexaYears living in
Cantabria ExposurebClinical symptomscMonths with symptoms Other sensitisationsd
Cutaneous reactione
C. selloana P. pr at en s e
E0416001 50 M 10 (*) RC May–Oct HDM/plantago 47 59
E0416002 38 F 38 (*) RCA Mar–Oct –40 24
E0416003 23 M 23 (–) RCAU Mar–Sep HDM/dog 30 21
E0416004 44 10 (*) RCA Apr–Sep HDM 47 45
E0416005 46 M 46 (*) RC Mar–Oct HDM/platanus 10 22
E0416006 49 M 20 (*) RC Jun–Oct – 67 34
E0416007 41 F 41 (–) RCA Mar–Aug Dog 32 113
E0416008 27 F 27 (*) RC May–Aug HDM 31 65
E0416009 27 F 27 (*) RC Mar–Oct –49 76
E0416010 55 M 55 (*) RC Apr/Oct – 30 54
E0416011 50 F 20 (*) RC Jul–Oct – 35 38
E0416012 36 M 36 (*) RC May–Oct – 16 33
E0416013 20 M 20 (*) RC Apr–Aug HDM 39 94
E0416014 42 M 37 (*) RC Mar–Sep – 19 92
E0416015 39 M 12 (*) RCA May–Oct HDM 18 25
E0416016 45 M 45 (*) RC May–Sep – 21 20
E0416017 52 M 31 (*) RCA Mar–Jul –63 92
E0416018 45 M 40 (*) RC May–Sep – 42 48
E0416019 34 M 34 (*) RC May–Oct – 67 78
E0416020 30 M 25 (*) RC Feb–Nov Cat 29 48
E0416021 44 M 21 (*) RCA May–Set – 111 162
E0416022 38 M 38 (*) RC Apr–Aug HDM 737
E0416023 43 M 7 (–) RC Feb–Nov Plantago 14 23
E0416024 50 M 50 (*) RC May–Set – 6(N) 39
E0416025 33 F 33 (*) RC Apr–Oct –49 28
E0416026 29 F 29 (*) RC Apr–Set – 37 77
E0416027 48 M 48 (*) RC May–Oct HDM 68 35
E0416028 41 M 4 (*) RC Apr–Jun – 86 48
E0416029 42 M 14 (*) RCU Apr–Sep HDM 21 48
E0416030 29 M 29 (*) RC May–Sep HDM 48 50
E0416031 42 M 42 (*) RCA Mar–Aug HDM 36 48
E0416032 48 M 15 (–) RC Mar–Aug 0 49 69
E0416033 25 M 25 (–) RC May–Sep HDM 34 169
E0416034 39 F 15 (*) RC Apr–Jul –6(N) 83
E0416035 53 F 17 (*) RC Apr–Nov –27 38
E0416036 48 M 6 (*) RC Apr–Oct HDM 61 33
E0416037 63 F 63 (*) RC Apr–Jul –27 33
E0416038 58 M 58 (*) RC May–Aug – 23 22
E0416039 39 M 39 (*) RC May–Aug HDM 50 114
E0416040 40 F 40 (*) RC May–Oct HDM 18 37
E0416041 31 F 31 (*) RC May–Aug – 1(N) 26
E0416042 29 F 29 (*) RCAU May–Jul – 13 31
E0416043 32 F 18 (*) RC Jul–Sep HDM 24 171
E0416044 42 F 8 (*) RCA Apr–Aug Parietaria 17 22
E0416045 42 M 2 (*) RC Jul–Sep – 1(N) 37
E0416046 22 F 22 (*) RCA Mar–Aug HDM/parietaria 79 64
E0416047 34 M 34 (*) RCA May–Oct HDM/cat 1(N) 18
E0416048 39 M 39 (*) RCA May–Jul – 41 42
E0416049 41 M 15 (*) RC Apr–Oct HDM 57 44
E0416050 28 F 27 (*) RC Apr–Nov HDM 15 23
E0416051 30 M 30 (*) RC Mar–May HDM/plantago/cupresa-
ceous/parietaria 32 128
E0416052 63 M 63 (*) RC May–Oct HDM 40 37
E0416053 22 M 20 (*) RC Apr–Aug Dog 21 75
E0416054 32 F 32 (*) RC May–Oct HDM 22 67
Continued
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Table 1. Demographic and clinical data and SPT results. SPT Skin prick test. a F female, M male. b Exposure
(*) means that the patient lives in an area in which C. selloana plants have been identied. c A athma, C
conjunctivitis, R rhinitis, U urticaria. d HDM House Dust Mites. e Wheal area (mm2). Negative reaction to C.
selloana is highlighted in italic numbers (N). Numbers in bold indicate positive reaction (>6).
Patient Age (years) SexaYears living in
Cantabria ExposurebClinical symptomscMonths with symptoms Other sensitisationsd
Cutaneous reactione
C. selloana P. pr at en s e
E0416055 41 F 7 (*) RCA May–Sep HDM/parietaria/
plantago 15 41
E0416056 44 F 44 (*) RC Apr–Nov HDM/horse/dog/cat 34 79
E0416057 23 F 23 (–) RCA May–Sep HDM 13 49
E0416058 41 F 41 (*) RC Apr–Sep – 38 36
E0416059 31 F 31 (*) RCAU Feb–Nov Cat/dog/plantago/HDM 815
E0416060 41 M 36 (*) RC May–Sep – 37 45
E0416061 29 F 29 (*) RC Apr–Sep – 29 68
E0416062 44 M 43 (–) RCA May–Jul Cat 535
E0416063 50 F 50 (–) RC Jun–Nov HDM 11 21
E0416064 26 F 1.5 (*) RC Mar–Oct –17 46
E0416065 69 M 69 (–) RCA May–Nov – 5(N) 66
E0416066 39 F 31 (*) RCA Apr–Aug –27 47
E0416067 40 M 40 (*) RCA May–Nov Plantago 10 14
E0416068 26 F 26 (*) RC May–Sep Plantago 34 39
E0416069 67 M 67 (–) RC May–Sep – 12 42
E0416070 70 F 70 (*) RC May–Nov HDM 48 46
E0416071 32 F 32 (*) RC Apr–Oct HDM 20 20
E0416072 30 F 30 (*) RC Apr–Jul 130 34
E0416073 18 F 18 (–) R May–Jun HDM/plantago 13 26
E0416074 50 M 24 (*) RC May–Oct – 54 53
E0416075 35 M 35 (–) RC Apr–Aug –75 77
E0416076 23 M 23 (*) RCA Apr–Aug HDM/plantago 71 96
E0416077 38 F 38 (*) RCA May–Oct HDM 38 22
E0416078 34 F 34 (–) RC Apr–Oct Parietaria 37 57
E0416079 23 M 23 (*) RC Apr–Oct HDM 1(N) 24
E0416080 36 M 36 (*) RCA Apr–Sep – 37 36
E0416081 32 F 32 (*) RCA Apr–Jul HDM/parietaria 28 78
E0416082 36 F 9 (–) RC May–Jun – 18 28
E0416083 31 M 31 (–) RC Apr–Jul –329
E0416084 23 M 23 (*) RCA May–Jul – 27 56
E0416085 39 F 39 (*) RC May–Aug HDM 67 55
E0416086 29 F 10 (–) RCAU May–Sep Platanus/cupresaceous/
plantago 11 39
E0416087 18 F 12 (*) RCA Apr–Sep Plantago 16 96
E0416088 46 M 15 (–) RCA Mar–Sep HDM/cat/dog/horse/
platanus/cupresaceous/
plantago 32 32
E0416089 30 F 30 (–) RCAU Mar–Jul Platanus/plantago 33 138
E0416090 23 M 23 (–) RCU Mar–Oct –18 52
E0416091 39 F 39 (*) RCA Mar–Nov HDM/plantago 17 80
E0416092 20 M 20 (*) RCU Apr–Jul –107 65
E0416093 63 F 63 (–) RC Apr–Aug –20 23
E0416094 23 F 23 (–) RC Mar–Sep Plantago/platanus 529
E0416095 44 F 26 (–) RC Apr–Jul Platanus/plantago/
cupresaceous 34 39
E0416096 43 F 43 (–) R May–Jun – 1(N) 89
E0416097 37 F 37 (*) RC May–Oct – 80 27
E0416098 30 M 30 (*) RCA Mar–Jul Dog/cupresaceous/
plantago 20 56
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for Phl p 1 (r = 0.75) than for Phl p 5 (r = 0.55). ese results suggest that the common reaction observed in the
patients to pollen extract of Phl and CS might reside in the antigenic Group 1 that is ubiquitous in all grasses25.
We analysed the measured grass pollen concentration along the year in the region. e regional agency
Health Department of the nearby Basque Country detected a spring main peak of grass pollen around May
and a second, August-to-October peak, in the air of Bilbao, a city 50km o Cantabria with a similar climate
and density of CS. It is interesting that most patients (78.6%) in the study mentioned a second allergic reaction
around September–October (Table1, Supplementary Fig.2). is indicates a timely correlation between grass
pollen and the referred allergic symptoms by the patients.
Discussion
We could not nd in the literatureany report on the impact of CS on human health. is is somehow surpris-
ing and highlights the need of studies on the issue, considering the widespread presence of this invasive plant
worldwide1–5. Concerns about the consequences of CS expansion are evident among professionals regarding the
impact of CS in ecology, industry or health9–12. Our study addresses for the rst time the potential allergenic
eects of CS pollen. Given the wide impact of grass allergy in the population, this constitutes a public health issue.
We here present several lines of evidence strongly suggesting that patients allergic to pollen of northern Span-
ish autochthonous grasses, such as Phl, are also allergic to pollen of CS: (i) 89% of the patients allergic to Phl
were sensitised to CS, as evident both by skin reaction and by sIgE in serum; (ii) the timely coincidence along
the year of allergy symptoms reported by patients, grass pollen counts and ourishing of CS; (iii) the presence
in CS of a protein band with a mobility compatible with grass allergenic group 1 and the strong prevalence of
this group in the sIgE to Phl.
050100 15
0200
0
50
100
150
SPT Phleum
(mm
2
)
SPT Cortaderia
(mm
2
)
BA
C
CS PhL
12M
250
150
100
75
50
37
25
20
15
10
43 kD
G1[ ]G1,5
Figure2. Cortaderia selloana (CS) pollen shares antigens and inmmunogenicity with authoctonous grass
Phleum pratense (Phl). (A) SDS‐PAGE prole of CS (lanes 1, 2) or Phl pollen extract (lanes 3,4) extract. Lane
1 and 2 corresponds to 20µl and 40µl of CS pollen extract, respectively (see also Supplementary Fig.3)
representative of two independent batches. Lane 3 and 4 correspond to 20µl from two dierent batches of Phl
pollen extract. M: the molecular weight markers. Brackets indicate the position of the allergenic groups (G)
according to the documented apparent molecular weights. (B) Correlation between SPT result for PhL and
CS Pearson r: 0.2558; R2: 0.06543; p value : 0.0110 (two‐tailed). (C) IgE Cross‐reactivity of CS and Phl pollen
antigens as measured by radioallergosorbent (RAST) inhibition assays. Note that if there were no cross‐reaction
the Phl plot should be at to cero (broken line).
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020406080100
0
20
40
60
80
100
sIgE Phleum, kU/L
sIgE Phl p 1, kU/L
A
02040608
01
00
0
20
40
60
80
100
sIgE Phleum, kU/L
sIgE Phl p 5, kU/L
B
020406080
100
0
20
40
60
80
100
sIgE Phleum, kU/L
sIgE Phl p 1 + 5, kU/L
C
Figure3. Linear regression of sIgE to Phleum pratense whole extract (ImmunoCap) versus sIgE to Phl p 1 (A),
sIgE to Phl p 5 (B) and sIgE to Phl p 1 + Phl p 5 (C).
Table 2. Correlation between sIgE to C. selloana and to Phl p 1 sIgE, to Phl p 5 sIgE and to P. pratense sIgE.
Phl p: Phleum antigen group.
sIgECS (Ku/L) vs. sIgE Phl p 1 (kU/L) sIgECS (Ku/L) vs. sIgE Phl p 5 (kU/L) sIgECS (Ku/L) vs. sIgE Phl (kU/L)
ImmunoCap
r Spearman 0.755 0.552 0.7476
95% condence interval 0.6513 to 0.8311 0.3920 to 0.6795 0.6400 to 0.8264
P (two‐tailed) < 0.0001 < 0.0001 < 0.0001
Signicant? (alpha = 0.05) Ye s Ye s Yes
Number of XY pairs 98 98 96
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e high cross-sensitisation to Phl and CS pollen in this cohort of patients is not explained by a possible
reaction to pan-allergens, such as prolin (Phl p 12) and polcalcin (Phl p 7), since only 27.5% of the patients
contained serum sIgE against them. Group 1 is a major grass allergen ubiquitous in all grasses in contrast to
group 5 which is absent in non-Pooideae grasses as is the case of CS. e data presented in this study strongly
suggests that grass group 1 might be the culprit of the observed cross-sensitisation between autochthonous
grasses (in this study, Phl) and CS.
Autochthonous grasses in Northern Spain ourish from April to July1,16,17,26, while CS ourishes from August
to October5,7. From a clinical point of view, most patients (78.6%) referred a late allergic symptoms reactivation
around September–October coincident with a second, August-to-October, peak of grass pollen counts in the
air. At present, there are no commercial extracts of CS for immunotherapy. However, the overall improvement
of symptoms usually reported by allergic patients that were treated with conventional grass immunotherapy,
during both pollination peaks, suggests that they might have been protected also to CS pollen. is in addition
holds clinical interest to those regions where CS is autochthonous and possibly allergenic.
e implications of the results into public health-related issues are many and diverse. First, the results encour-
age the international community to run allergenic tests to CS and to biochemically characterise the reaction
to CS. Second, the results suggest that CS might lengthen the grass allergy season in territories where CS has
expanded, by causing a second later peak, additional to the peak due to the autochthonous grasses. To note,
commercially available grass immunotherapy might be benecial to patients allergic to CS worldwide. ird,
given that CS is banned in many countries and states, since it is considered aninvasive species3, a demonstrated
impact on human health would encourage policy makers to run programmes for eradicating this plant in non-
autochthonous areas. e results provide an example of the global eects that alien invasive species can have
on human health.
Received: 5 August 2021; Accepted: 25 November 2021
References
1. García-Mozo, H. Poaceae pollen as the leading aeroallergen worldwide: A review. Allergy Eur. J. Allergy Clin. Immunol. 72(12),
1849–1858 (2017).
2. Lambrinos, J. G. e expansion history of a sexual and asexual species of Cortaderia in California, USA. J. Ecol. 89(1), 88–98
(2001).
3. United State Department of Agriculture. Weed Risk Assessment for Cortaderia selloana. (2014).
4. Global Biodiversity Information Facility. Cortaderia selloana Asch. & Graebn. Syn. Mitteleur. Fl. 2(1): 325 1900. https:// www. gbif.
org/ speci es/ 27045 23. Accessed 2 Mar 2021.
5. Pardo-Primoy, D. & Fagúndez, J. Assessment of the distribution and recent spread of the invasive grass Cortaderia selloana in
Industrial Sites in Galicia, NW Spain. Flora Morphol. Distrib. Funct. Ecol. Plants. 259, 151465. https:// doi. org/ 10. 1016/j. ora. 2019.
151465 (2019).
6. Campos, J. A., Herrera, M., Biurrun, I. & Loidi, J. e role of alien plants in the natural coastal vegetation in central-northern
Spain. Biodivers. Conserv. 13(12), 2275–2293 (2004).
7. Herrera, M. & Campos, J. e reed of the pampas (Cortaderia Selloana) Bizkaia: Practical guide for its control. Inst. Estud. Territ.
Bizcaia Diput. Foral. Bizk. 1, 30 (2006).
8. Guinea E. Geografía Botánica de Santander. [Botanical Geography of Santander]. Publicaciones de la Exma. Diputación Provincial
de Santander (1953).
9. Queensland Government. Cortaderia selloana. https:// keyse rver. lucid centr al. org/ weeds/ data/ media/ Html/ corta deria_ sello ana.
htm. Accessed 2 Mar 2021.
10. Cortaderia selloana risk assessment. http:// www. hear. org/ pier/ wra/ austr alia/ cosel- wra. htm. Accessed 2 Mar 2021.
11. Cortaderia Species. Silver Pampas Grass, Uruguayan Pampas Grass Cortaderia selloana. https:// daves garden. com/ guides/ pf/ go/
1291/. Accessed 2 Mar 2021.
12. Early Warning Network—LIFE Stop Cortaderia—Medidas urgentes de lucha contra el plumero de la Pampa. [Urgent measures
to combat the Pampa]. http:// stopc ortad eria. org/ langu age/ en/ early- warni ng- netwo rk/. Accessed 2 Mar 2021.
13. Ruiz de la Torre, J. et al. Catálogo de especies vegetales a utilizar en plantaciones de carreteras [Catalog of plant species to be used
in roads]. Ministerio de Obras Públicas y Urbanismo SGT, editor. Vol. Primera. EPES lnduslrias Grácasi (2019).
14. Plantaciones|Ministerio de Transportes, Movilidad y Agenda Urbana [Plantations|Ministry of Transport, Mobility and Urban
Agenda]. https:// www. mitma. gob. es/ carre teras/ norma tiva- tecni ca/ 14- plant acion es. Accessed 22 Oct 2021.
15. Background and objectives: LIFE Stop Cortaderia: Medidas urgentes de lucha contra el plumero de la Pampa. [Urgent measures
to combat the Pampas grass]. http:// stopc ortad eria. org/ langu age/ en/ backg round- and- objec tives/. Accessed 2 Mar 2021.
16. Subiza, J. Gramíneas : Aerobiología y polinosis en España. [Grasses: Aerobiology and pollinosis in Spain]. Alergol Inmunol Clin.
18, 7–23 (2003).
17. Jato, V., Rodríguez-Rajo, F. J., Seijo, M. C. & Aira, M. J. Poaceae pollen in Galicia (N.W. Spain): Characterisation and recent trends
in atmospheric pollen season. Int. J. Biometeorol. 53(4), 333–344 (2009).
18. Alergológica-2005. Factores epidemiológicos, clínicos y socioeconómicos de las enfermedades alérgicas en España. [Epidemiologi-
cal, clinical and socioeconomic factors of allergic diseases in Spain]. (SEAIC, 2006).
19. Andersson, K. & Lidholm, J. Characteristics and immunobiology of grass pollen allergens. Int. Arch. Allergy Immunol. 130(2),
87–107 (2003).
20. Bufe, A., Schramm, G., Keown, M. B., Schlaak, M. & Becker, W. M. Major allergen Phl p Vb in timothy grass is a novel pollen
RNase. FEBS Lett. 363(1–2), 6–12 (1995).
21. Ceska, M., Eriksson, R. & Varga, J. M. Radioimmunosorbent assay of allergens. J. Allergy Clin. Immunol. 49(1), 1–9 (1971).
22. Sánchez-Madrid, F., Morago, G., Corbi, A. L. & Carreira, J. Monoclonal antibodies to three distinct epitopes on human IgE: eir
use for determination of allergen-specic IgE. J. Immunol. Methods. 73(2), 367–378 (1984).
23. Ramírez, J. et al. Group 5 determination in Pooideae grass pollen extracts by monoclonal antibody-based ELISA: Correlation with
biologic activity. Allergy Eur. J. Allergy Clin. Immunol. 52(8), 806–813 (1997).
24. WHO/IUIS Allergen Nomenclature Sub-Committee. WHO/IUIS Allergen Nomenclature Home Page. http:// www. aller gen. org/.
Accessed 2 Mar 2021.
Content courtesy of Springer Nature, terms of use apply. Rights reserved
10
Vol:.(1234567890)
Scientic Reports | (2021) 11:24426 | https://doi.org/10.1038/s41598-021-03581-5
www.nature.com/scientificreports/
25. Suphioglu, C. What are the important allergens in grass pollen that are linked to human allergic disease?. Clin. Exp. Allergy. 30(10),
1335–1341. https:// doi. org/ 10. 1046/j. 1365- 2222. 2000. 00955.x (2000).
26. De B enito Rica, V. & Soto Torres, J. Polinosis y aerobiología del polen en la atmósfera de Santander [Pollinosis and aerobiology of
pollen in the atmosphere of Santander]. Alergol. Inmunol. Clin. 16(2), 84–90 (2001).
Acknowledgements
We thank Daniel Liébana Uranga (Cantabria, Spain) for permission to include authorship and data obtained
by Marta Uranga, sadly deceased before preparation of the manuscript. We thank alia Burn (England, UK)
for manuscript text revision and Life Stop Cortaderia (Cantabria, Spain) for image in Fig.2B-right. is work
received nancial support from ALK-Abelló S.A (Madrid, Spain). is article is dedicated to the memory of
Marta Uranga.
Author contributions
F.R.: conceptualization, data curation, investigation, methodology, project administration, supervision, valida-
tion, writing—review & editing. M.L.V.: conceptualization, data curation, formal analysis, investigation, meth-
odology, validation, writing—review & editing. L.d.l.V., S.A., E.M.: data curation, investigation and methodology.
L.S.J.: soware, review & editing. D.L.: review & editing. M.U.: conceptualization, data curation, formal analysis,
investigation. A.G.: conceptualization, investigation, methodology, supervision, validation, writing—review &
editing.
Competing interests
e authors declare no competing interests.
Additional information
Supplementary Information e online version contains supplementary material available at https:// doi. org/
10. 1038/ s41598- 021- 03581-5.
Correspondence and requests for materials should be addressed to F.R., M.L.-V.orA.G.
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