Weaning, food allergens and potential risk factors for delayed introduction: a cross-sectional study

Abstract Much evidence supports that the early introduction of allergenic foods in weaning is useful to prevent food allergies later in life. Real life is often different, with factors related to mothers and infants. Our study aimed to deepen the timing of introducing the foods responsible for most allergic reactions during the weaning and why parents delay their introduction. 110 mothers participated in the study, compiling a questionnaire. Exclusive breastfeeding was associated with a delayed introduction of allergenic foods at 4 months (r = 0.433, p < 0.01) and 1 year (r = 0.486, p < 0.01). Large-for-gestational age at birth was inversely associated with a delayed introduction of allergenic foods (r=-0.204, p < 0.05). This study demonstrates that introducing many allergens is delayed during the weaning. Parents with infants fed with exclusive breastfeeding could need more information about the correct time of introduction of potential allergens in the weaning.


Introduction
The prevention of food allergy in the paediatric age consisted of delaying the introduction of allergenic foods until 2008 (American Academy of Pediatrics 2000).Suddenly, no studies demonstrated the effectiveness of this preventive technique (de Silva et al. 2014), and this statement was not confirmed (Greer et al. 2019).Many studies support the early introducing of allergenic foods in weaning to prevent food allergies later in life (Joshi et al. 2019;Brough et al. 2022).Du Toit et al. (2016) and Perkin et al. (2016) demonstrated that the introduction to the diet of peanuts and eggs before 11 months of life in children affected by a high risk of food allergy was related to a reduction in the incidence of peanut-and egg-allergy later.According to these results, many scientific societies (i.e.American Academy of Paediatrics, Asia Pacific Association of Paediatric Allergy, Respirology & Immunology, European Academy of Allergy and Immunology, German Society for Allergology and Clinical Immunology, and National Institute of Allergy and Infectious Diseases) advocated the possibility of an early introduction of potential food allergens during the weaning.(Schäfer et al. 2014;Togias et al. 2017;Tham et al. 2018;Greer et al. 2019;Halken et al. 2021).Moreover, a recent randomised trial conducted by Skjerven et al. (2022) demonstrated that exposure to allergenic foods from 3 months of age reduced the incidence of food allergy at 36 months.(McWilliam et al. 2022) In addition, Venter et al. (2022) estimated that for every allergen introduced before the age of one, the probability of developing an allergy in the following ten years is reduced by 33%.The pathophysiological basis seems to consist of developing the primary oral exposure of antigens through a well-developed mucosal immune system rather than an altered skin epidermal barrier.(Brough et al. 2020) Although indications in the day-by-day clinical work from paediatric allergologists addressed an early introduction of potentially allergenic foods, real life is often different.This event could be explained by many reasons, such as parental fear, low child cooperation, demand for access to health services, and practical difficulties (Fisher et al. 2018).Moreover, Almutairi et al. (2021) demonstrated that many mothers had limited knowledge regarding the beneficial health-related outcomes of this practice.Finally, non-white ethnicity, advanced maternal age, low parental quality of life, atopic dermatitis, and feeding difficulties of the infant could represent further causes explaining the delay of introducing the potential food allergens (Perkin et al. 2016;Perkin et al. 2019;Trogen et al. 2022).Thus, this study aimed to deepen the timing of introducing the eight foods responsible for most allergic reactions (milk, egg, wheat, fish, soy, peanut, nuts, shellfish) during the weaning.Moreover, it deepens why parents delay introducing one or more specific potentially allergenic foods.Specifically, the number and the time of the introduction of solid foods during the weaning represent the first outcome.The secondary outcome was the association between the introduction of food allergens in weaning and demographic features, breastfeeding (duration and frequency), milk replacement formulas (breast milk replacement), factors that determine the introduction of complementary feeding, knowledge about the practice of the weaning, and maternal anxiety.

Study design and participants
This cross-sectional study was conducted between April 2021 and November 2021 in Mantua, Lombardia, Italy.All family paediatricians working in the province of Mantua were requested to participate in the study.Five paediatricians agreed to be involved and invited to participate the mothers of children aged 2-14 years attending their office.They administered a self-reported questionnaire that included 29 self-reported questions deepening the time of introduction of the most involved allergens in their infants' diet and the factors potentially related to a delay in their introduction.The inclusion criteria were represented by the condition of a healthy child without food allergy, feeding difficulties, or other disorders or choices that required a specific diet.Mothers completed a questionnaire, and paediatricians verified for completeness to ensure they met the required quality standards.

Data collection
All questionnaires were fully and correctly completed.The research tool was an original questionnaire developed by the study's authors based on the Complementary Food Frequency Questionnaire (Mejía-Rodríguez et al. 2014) Selected questions from this questionnaire were translated into Italian and modified to include the time of introduction of the most involved allergens commonly present in the diet of the infants.In addition, the factors potentially related to a delay in introducing allergenic foods were deepened.Specifically, the family history, demographic features, pregnancy, birth, breastfeeding, milk replacement formulas, factors related to the introduction of complementary weaning, knowledge about the weaning practice, and maternal anxiety.The local Ethical C om m itte e approve d t he stu dy ( c o d e 9-2022-OSS_ALTRO-MN4), and the family paediatrician took the parents' informed consent.

Statistical analysis
Categorical variables are presented as the percentage of samples (%).Continuous variables are reported as the median and the interquartile range (IQR) or mean and standard deviation as appropriate.The normality of the distribution of the variables is checked with a Kolmogorov-Smirnov test.The Spearman correlation coefficient for non-parametric data was used.p < 0.05 was considered as statistically significant for all tests, MedCalc Statistical Software version 17.6 (MedCalc Software bvba, Ostend, Belgium; http://www.medcalc.org) and GraphPad Prism version 6.00 for Mac (GraphPad Software, La Jolla California USA, http:// www.graphpad.com)were used to perform the analyses.

Results
One hundred ten mothers participated in the study, compiling the questionnaire.Characteristics of the population are shown in Table 1 and Table A (supplementary material).The average age of mothers at birth was 31 years, and most were European (97.2%).The average gestational age of their children was 39 weeks.Most mothers had no history of family allergies or anaphylaxis (70% and 95.5%, respectively).Few mothers referred to pregnancy-related anxiety (12.7%) or weaning (15.5%).Most mothers declared to know correct information about food composition and weaning practices (90.9%) and had adequate levels of instruction (92.7%).Exclusive breastfeeding, combination feeding and exclusive use of baby formula were adopted at 4 months by 41.8%, 23.6%, and 34.5% of mothers.About the questionnaires received, all were included as complete and legible.Regarding the introduction of food allergens in weaning (Figure 1 and Table 2), fish, wheat, egg and soy are mainly introduced during the first year of age.Cow milk was introduced almost exclusively at 12 months.Soybean was introduced mainly at 12 months, but fewer cases were introduced earlier (around 6 months).Shellfish, hazelnuts, walnuts and peanuts were often introduced after 2 years of age.Regarding the deepening of the  factors potentially related to a delay in introducing allergenic foods in weaning, exclusive breastfeeding during the first year of life was related to a delayed introduction of allergenic foods (r = 0.433; p < 0.01).Specifically, the mothers who feed infants exclusively through breastfeeding at four months delay the introduction of allergenic foods (r = 0.486; p < 0.01).On the contrary, large-for-gestational age (LGA) was inversely associated with a delayed introduction of allergenic foods (r = 0.204; p < 0.05), appearing as a potential protective factor.No statistical association was found when considering the time of introducing the foods into the weaning and family history, demographic features, pregnancy, solid foods, factors related to the introduction of complementary feeding, knowledge about the weaning, and maternal anxiety (Table 3).

Discussion
It is demonstrated that early introduction of potential food allergens during the weaning is helpful advice to reduce the incidence of food allergy later in life (McWilliam et al. 2022).Many scientific societies advocated the possibility of an early introduction of potential food allergens during the weaning.In clinical practice, it is reasonable to start introducing all allergenic foods at the same time as other solid ones at around six months, but not before four months, while continuing breastfeeding.(Schäfer et al. 2014;Togias et al. 2017;Tham et al. 2018;Greer et al. 2019;Halken et al. 2021).The results of our study showed that the introduction of many potential food allergens during the weaning is delayed.Specifically, eggs, milk, and soy are introduced between 9 and 12 months.In addition, nuts, shellfish, and peanuts are introduced at 2 years old.This parental behaviour confirmed the evidence demonstrated in previous studies (Venter et al. 2022).
A similar study conducted by Almutairi et al. (2021) demonstrated similar results for the introduction of eggs (9.84 ± 3.16 months) but was earlier for peanuts (14.6 ± 5.78 months) and fish (13.6 ± 5.37 months).Schiess et al. (2011) estimated that introducing egg, fish, and soya in weaning was rare before six months.Many factors could explain this parental behaviour.Specifically, parental fear, low child cooperation, demanding access to health services, practical difficulties, limited knowledge, non-white ethnicity, advanced maternal age, low parental quality of life, atopic dermatitis, and feeding difficulties of the infants were demonstrated risk factors (Perkin et al. 2016;Fisher et al. 2018;Almutairi et al. 2021;Perkin et., 2019;Trogen et al. 2022).In addition, our study highlighted an additional risk factor that was quite peculiar and unproven before.Starting from the statement that breastfeeding should always be encouraged, we unexpectedly found a positive association between exclusive breastfeeding during weaning and the delayed introduction of allergenic foods.It is impossible that teasing a part cause an effect, although mothers who have not yet introduced any other food (i.e.milk formula) could be more reluctant to introduce new elements into their infants' diet, continuing a pattern based on their resources.This result is similar to the evidence demonstrated by Schiess et al. (2011), where formula-fed infants received potentially allergenic foods earlier than breastfed infants.On the other hand, this study showed another unexpected result: LGA seems better to respect the timing of the correct introduction of potential allergens in weaning.Specifically, there is an inverse correlation between high birth weight and delayed introduction of allergenic foods.Anyway, this finding should be interpreted with a precautionary approach.
It is known that LGAs are at greater risk of developing endocrine-metabolic diseases later in life, such as diabetes, metabolic syndrome, hypertension and obesity (Derraik et al. 2020;Zhang et al. 2023).Starting from this statement, it is possible to speculate that an LGA newborn could be a high-weight infant, appearing healthier in the eyes of the parents, who could be reassured not to delay the introduction of foods in weaning.Consequently, this result has only a descriptive value and was not demonstrated before.We did not find any relationship between the time of introducing allergenic foods and the other risk factors for delayed weaning, maybe related to the small sample size.All risk factors are known and seem related to the parents' fear of the onset of allergic reactions in their children and the potential difficulty that infants could manage the consistency of these foods (Perkin et al. 2016;Fisher et al. 2018;Almutairi et al. 2021;Perkin et., 2019;Trogen et al. 2022).Some studies experimented with using food products containing a mixture of allergens in small doses to be consumed daily (Holl et al. 2020), (Quake et al. 2022).Our study is subjected to multiple limitations.First, it is monocentric.Second, it involved a limited number of paediatricians and mothers.Third, data were collected in a single province.Fourth, the anamnestic information is self-reported by the parents of children.

Conclusions
This study shows that most parents do not follow a correct approach in introducing potential allergens in weaning.This statement is especially true for peanuts, nuts and shellfish.Starting from the known evidence of an "at-risk population" suffering from parental fear, low child cooperation, demanding access to health services, practical difficulties, limited knowledge, non-white ethnicity, advanced maternal age, low parental quality of life, atopic dermatitis, and feeding difficulties, our study demonstrates that parents who have children born as LGA and fed with exclusive breastfeeding could need more information about the correct time of introduction of potential allergens in the weaning.Future studies are needed to confirm or exclude this very peculiar and surprising finding.Finally, our study could help paediatricians identify a specific subgroup of parents that could benefit from tailored information about introducing potential allergen foods in weaning.

Figure 1 .
Figure 1.age of introduction of allergens during the weaning.Values are expressed as a median.

Table 1 .
Study population and main questions and answers by the questionnaire.

Table 2 .
age of introduction of allergens during the weaning, represented through median age and IQr.

Table 3 .
relationship between the time of introduction of potential food allergens and factors potentially involved in the delay of their introduction.