Introduction

The aromatic component of wine is an important element in consumer wine appreciation, as olfactory preferences are generally driven by the intensity and the dominant family of aroma perceived at tasting (Lesschaeve, 2008). In the 1980s, Noble et al. (1987) undertook to classify wine aroma into families leading to the development of a “wine aroma wheel”, which became a reference for wine writers, consumers, producers and researchers. From this work twelve distinct aroma families were identified, including a “vegetative” family related to sensory attributes, such as “grass cut green”, “eucalyptus”, “mint” and “bell pepper”.

The molecular determinism of these “vegetative” notes in wine has been widely studied and is currently well-understood. 3-Isobutyl-2-methoxypyrazine (IBMP) and 3-isopropyl-2-methoxypyrazine (IPMP) are two potent grape-derived molecules contributing to the bell pepper and asparagus character respectively (Bayonove et al., 1975). 3-alkyl-2-methoxypyrazines have been among the most studied aroma compounds since their first discovery in Cabernet Sauvignon (Bayonove et al., 1975), and some years later in Sauvignon (Augustyn et al., 1982). 1,8-cineole (also called eucalyptol) and its isomer 1,4-cineole, which impart eucalyptus and hay/dried herbs aromas respectively, have received a lot of attention from researchers in the past few years (Antalick et al., 2015; Capone et al., 2012; Poitou et al., 2017). Although these monoterpenes can have an endogenous origin, it is also possible for them to be introduced to grapes and wines from neighbouring plants (Capone et al., 2012; Poitou et al., 2017). C6-compounds (i.e., trans-2-hexenal or hexanal), which are formed in the early winemaking stages through enzymatic oxidation of lipidic precursors, are responsible for the smell of cut grass in wines (Cordonnier and Bayonove, 1981). During alcoholic fermentation, a part of these compounds is reduced by yeast to an alcohol form or, for trans-2-hexenal, is involved in the synthesis of 3-mercaptohexanol (Roland et al., 2010), which generally limits their sensory contribution to finished wines. Another monoterpene known as piperitone was recently identified as the main contributor of the mint nuances perceived in aged Bordeaux wines (Picard et al., 2016; Pons et al., 2016). It was also highlighted that p-menthane lactones can enhance the freshness and the mint odours perceived in these wines (Picard et al., 2017).

However, few research studies have been conducted to investigate consumer appreciation of the molecules responsible for these “vegetative” characters. The most significant study demonstrated that 1,8-cineole became undesirable in red wines above a concentration of 27.5&nbspμg/L, thus encouraging winemakers to produce wines with moderate levels of this compound (Saliba et al., 2009). Without performing a formal consumer study, piperitone was considered a positive aroma compound, as the molecule was positively correlated with aging-bouquet typicality scores (Picard et al., 2016). 3-alkyl-2-methoxypyrazines, and notably IBMP, are usually seen as highly undesirable compounds by wine professionals and researchers. However, while the appreciation of methoxypyrazines has not yet been studied in red wines, moderate levels of IBMP can have a positive contribution to the aroma of Sauvignon wines (King et al., 2011; Rajchl et al., 2009).

The aim of this study was to formally assess the appreciation of IBMP in white and red wines by applying the consumer rejection threshold methodology (CRT) (Prescott et al., 2005). This valuable approach, based on the application of the paired preference test, allows the concentration of a molecule that gives a significantly lower preference to be determined. It has been previously applied to several compounds or flavours, including those that cause wine faults, such as 2,4,6-trichloroanisole (Prescott et al., 2005), 1,1,6-trimethyl-dehydronaphthalene also known as TDN (Ross et al., 2014), ethyl phenyl-acetate and phenyl acetic acid (Campo et al., 2012), 1,8-cineole (Saliba et al., 2009) and rotundone (Geffroy et al., 2018).

Sauvignon and Fer are two grape cultivars grown in the southwest of France within the Protected Designation of Origin (PDO) Gaillac, which cover 500 and 1000&nbspha respectively, according to the local winegrower association (www.vins-gaillac.com). Wines made from these two grape cultivars, which are known to be regular IBMP producers (Allen et al., 1991; Davaux, 2005; Geffroy et al. 2019; Lacey et al., 1991; Poitou et al., 2017), were used as base wines for this study. In both base wines, the best estimate threshold (BET) for olfactive detection of IBMP was also determined to find out the concentration at which the addition of IBMP can be perceived by panelists.

Materials and methods

Results and discussion

The panel BET was estimated as being 5.5&nbspng/L and 16.8&nbspng/L for Sauvignon and Fer respectively. These results are in accordance with previous studies, emphasising an odour threshold for IBMP of 8&nbspng/L in Sauvignon (Allen et al., 1991), and 15&nbspng/L in Cabernet Sauvignon, Cabernet Franc and Merlot (Roujou de Boubée et al., 2000). The importance of the matrix (i.e., the base wine) on aroma perception in wine has already been highlighted (Robinson et al., 2009; Villamor et al., 2013). These latter reported that increased contents in ethanol, tannin and sugar could reduce the headspace concentration of volatile aroma compounds and threshold values by between 2- and 10,000-fold. As the two studied matrices contained similar levels of ethanol and sugar (Table 1), we can suppose that the larger amount of tannins found in the Fer wine, as reflected by TPI values, is mainly responsible for the BET differences observed between the two wines. The differences in aroma composition between the two wines might also have played a role via mechanisms related to perception interactions (McKay et al., 2020).

As the panel was only composed of young panelists (<&nbsp25 years old), we cannot exclude the fact that slightly higher BETs would have been observed for a more age-balanced population. Indeed, it is well known that sensitivity for detecting odour usually declines with age (Griep et al., 1995). In the same way, as experience is known to facilitate the ability of panelists to discriminate wines when conducting triangular tests (Solomon, 1990), the fact that a less trained panel would have led to a higher BET cannot be rejected.

The distribution of best estimate threshold concentration showed a large variability between panelists and did not follow a normal law (Figure 1). For Sauvignon, one panelist succeeded at the lowest concentration and three panelists missed at the highest concentration. According to the Mann-Whitney test, no significant differences in BET was observed at P&nbsp<&nbsp0.05 between female and male panelists for Sauvignon (P&nbsp=&nbsp0.067) and Fer (P&nbsp=&nbsp0.302). However, BETs tended to be lower for female panelists (3.5&nbspng/L for Sauvignon and 14.6&nbspng/L for Fer) in comparison to male panelists (21.6&nbspng/L for Sauvignon and 21.6&nbspng/L for Fer). Actually, most of the research investigating sex differences in terms of olfactory threshold have reported either greater female sensitivity or no sex differences in sensitivity, depending on the odorant (Doty and Cameron, 2009). Consequently, the fact that the greater proportion of female panelists within the panel might have contributed to lower the BETs for both wines cannot be completely discarded.

Figure 1. Distribution of best estimate threshold (BET) concentration (ng/L) for IBMP in (A) Sauvignon wine (n = 20 panelists) with a global BET at 5.5 ng/L, and (B) Fer wine (n = 19 panelists) with a global BET at 16.8 ng/L, according to the ASTM method E 679 (see Mat. & Met. section).

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Following these results, the base wines were spiked with IBMP for the CRT study at 7.5, 15, 30, 60, and 120&nbspng/L for Sauvignon, and at 15, 30, 60, 120, and 240&nbspng/L for Fer, which, for the latter variety, corresponds to total IBMP concentrations of 18, 33, 63, 123 and 243 ng/L. These concentrations were chosen on the basis that IBMP range should start at levels which are detectable by panelists. For both wines, CRTs were established, at 50&nbspng/L and 30&nbspng/L for Sauvignon and Fer respectively (Figure 2).

Figure 2. Proportion of panelists preferring the control sample in A) Sauvignon wine and B) Fer wine. Horizontal lines show “no preference” (–––) and the “control is significantly preferred” at 5 % risk using paired comparison tests (– – –), n = 48 panelists. CRT stands for Consumer Rejection Threshold according to Prescott et al. (2005).

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The difference in CRT concentrations between the two matrices indicates that IBMP is more acceptable in Sauvignon wine than in Fer wine. Although the preference was not significant, the Sauvignon wine spiked with 7.5&nbspng/L of IBMP was preferred to the control (Figure 2A), which is consistent with previous research conducted with Czech panelists (Rajchl et al., 2009). Such IBMP concentration is likely to enhance the freshness of a white wine, a character usually desirable for this kind of product (IFV, 2013). This refreshing effect must have been particularly pronounced as the base wine was made from fully ripe grapes. The results obtained might have been different if other styles of Sauvignon had been used, notably those with high level of varietal thiols, potent aroma compounds that can also contribute to the typicality of wines made from this cultivar (Tominaga et al., 1998). For this latter style of Sauvignon wines, it has been shown that perceptual interactions are likely to reduce the green perception of IBMP (van Wyngaard et al., 2014), which could have led to higher BET and CRT values. However, other studies conducted in Australia on a base wine spiked with high thiol concentrations also demonstrated that the largest cluster of consumers, representing 43&nbsp% of the whole panel, preferred Sauvignon blanc wine containing moderate (10&nbspng/L), rather than high (20&nbspng/L) levels of methoxypyrazines (King et al., 2011).

It should be noted that the CRT in Sauvignon may not be applicable to all other countries, in particular the countries in which the consumers are familiar with wines containing great concentrations of IBMP, such as those from New Zealand (Lund et al., 2009). As familiarity is known to positively affect the appreciation of a given aroma by consumers (Saliba et al., 2009), it is possible that such consumers would have rejected spiked samples at higher IBMP concentration.

The lower CRT observed for Fer wine is somehow consistent with research highlighting that French consumers tend to have a short-term preference for Bordeaux red wines with a fully ripe/jammy fruit aroma (for which the sensory contribution of IBMP is likely to be negligible), to the detriment of wines with fresh fruit notes (Tempère et al., 2019). While we targeted red wines made with Fer grapes, our results might be transposable to other red grape cultivars, such as Cabernet Sauvignon, Cabernet Franc and Merlot, for which the odour threshold of IBMP has been reported at a similar range of concentration (Roujou de Boubée, et al. 2000).

For both wines, the proportion of panelists preferring the control wine slightly decreased at between 60&nbspng/L and 120&nbspng/L for Sauvignon, and 123&nbspng/L and 243&nbspng/L for Fer, which may reflect a saturation of olfactive receptors as a result of the large quantity of aroma compounds inhaled by panelists.

Non-parametric statistical tests on our data showed that the age of panelists had an impact on the appreciation of Sauvignon wines spiked with 120&nbspng/L of IBMP, P&nbsp=&nbsp0.038 (Table 2). For this IBMP concentration, younger panelists aged between 18 and 24 preferred the control wine in comparison with the other age groups. Although not statistically significant, the same trend was observed for Fer wines spiked with 240&nbspng/L, P&nbsp=&nbsp0.082. As our panel had a larger proportion of young panelists, the proportion of panelists which preferred the control might have been overestimated for the highest level of IBMP concentration. For Sauvignon and Fer with 15&nbspng/L and 243&nbspng/L of IBMP, the panelists’ preference was affected by the self-reported liking of vegetative notes in wines. The wines enriched with IBMP were preferred by the consumers, who stated that they enjoyed the vegetative notes. This coherence between the real appreciation of vegetative notes and consumers’ self-reported liking indicates that panelists recruited during a wine fair have good knowledge of wine and are able to analyse their olfactory sensations. It is not clear why this observation was not confirmed for Sauvignon at 30, 60, and 120&nbspng/L.

Finally, it should be pointed out that previous IBMP concentrations found in Sauvignon wines have never exceeded the CRT values of 50&nbspng/L determined in our study. Indeed, a couple of studies have investigated IBMP levels in commercial Sauvignon wines sourced from various countries throughout the world (Lacey et al., 1991; Lund et al., 2009). Outside New Zealand, or other cool climate vineyards such as the Loire valley in France, levels of IBMP rarely exceed 20&nbspng/L. The highest IBMP concentration of 47&nbspng/L was recorded in a wine from the Wairarapa region in New Zealand (Lund et al., 2009). As an autochthonous cultivar only grown in the southwest of France, data concerning IBMP in Fer wines are scarce. However, the usual IBMP concentrations also tend to be below the CRT concentration of 30&nbspng/L, but very high IBMP levels have been previously found in Fer wines; for example, two decades ago in cool climate vineyards (PDO Marcillac) with concentrations of up to 110&nbspng/L (Davaux, 2005). Indeed, as a result of the expected rise in temperature during the berry development stage, climate change is already decreasing IBMP levels in finished wines (Pons et al., 2017). To our knowledge, in experimental Fer wines made during the last decade without the aim of minimising IBMP content, concentrations did not exceed 27&nbspng/L, including in those from cool climate vineyards (Geffroy et al., 2015; Geffroy et al., 2019; Poitou et al., 2017). These data from the literature indicate that IBMP is neutrally perceived by consumers in Sauvignon and Fer wines at the usual concentration levels.

Conclusion

In our study, BET and CRT values for 3-isobutyl-2-methoxypyrazine in Sauvignon and Fer wines from Gaillac were estimated. For Sauvignon, these BET and CR thresholds were established as being 5.5&nbspng/L and 50&nbspng/L respectively, while for Fer they were 16.8 ng/L and 30 ng/L.

These findings suggest that IBMP is much more desirable in white wine than in red wines. While the IBMP levels reported in the literature for Sauvignon are always below the CRT value estimated for this cultivar, those recorded for Fer during the last decade are also below its respective CRT value. These observations suggest that IBMP is neutrally perceived by consumers at the usual concentrations in Sauvignon and Fer wines from Gaillac, and this contradicts the belief in the wine industry that consumers find the ‘vegetative' styles of wine undesirable, and that IBMP levels should be lowered by any means. Climate change is likely to negatively affect IBMP concentrations in finished wines, with an even more pronounced effect expected over the coming decades (Pons et al., 2017). Therefore, our findings do not necessarily encourage local winegrowers and winemakers to implement production practices to minimise IBMP concentrations in wine and thus reduce the impact of this molecule on the wine sensory profile.

Acknowledgments

We are grateful to Domaine Mas Pignou and Domaine Gayrard for supplying the wines, and to the panelists from the Ecole d’Ingénieurs de PURPAN involved in the BET study. We would also like to thank all the consumers who participated in the CRT study.

References

• ASTM (2004). Standard practice E 679–04. Standard practice for determination of odor and taste thresholds by a forced-choice ascending concentration series method of limits. Philadelphia, PA: American Society for Testing and Materials.
• Allen, M.S., Lacey, M.J., Harris, R.L., & W.V., Brown (1991). Contribution of methoxypyrazines to Sauvignon blanc wine aroma. American Journal of Enology and Viticulture 42, 109-112.
• Antalick, G., Tempère, S., Šuklje, K., Blackman, J.W., Deloire, A., de Revel, G. & L.M., Schmidtke (2015). Investigation and sensory characterization of 1, 4-Cineole: a potential aromatic marker of Australian Cabernet-Sauvignon Wine. Journal of Agricultural and Food Chemistry 63, 9103-9111. doi:10.1021/acs.jafc.5b03847
• Augustyn, P., Rapp, A. & C., van Wyk (1982). Some volatile aroma components of Vitis vinifera L. cv. Sauvignon blanc. South African Journal of Enology and Viticulture 3, 52-60. doi:10.21548/3-2-2382
• Bayonove, C., Cordonnier, R., & P. Dubois (1975). Study of an aromatic characteristic fraction of Cabernet-Sauvignon grape variety, identification of 2-methoxy-3-isobutyl-pyrazine. Comptes rendus hebdomadaires des séances de l'Académie des Sciences Série D 281, 75-78.
• Campo, E., Sáenz-Navajas, M., Cacho, J., & V., Ferreira (2012). Consumer rejection threshold of ethyl phenylacetate and phenylacetic acid, compounds responsible for the sweet‐like off odour in wines made from sour rotten grapes. Australian Journal of Grape and Wine Research 18, 280-286. doi:10.1111/j.1755-0238.2012.00198.x
• Capone, D.L., Jeffery, D.W., & Sefton, M.A. (2012). Vineyard and fermentation studies to elucidate the origin of 1, 8-cineole in Australian red wine. Journal of Agricultural and Food Chemistry 60, 2281-2287. doi:10.1021/jf204499h
• Cordonnier, R. & Bayonove, C.L. (1981). Etude de la phase préfermentaire de la vinification: extraction et formation de certains composés de l'arôme; cas des terpenols, des aldehydes et des alcools en C 6. OENO One 15, 269-286. doi:10.20870/oeno-one.1981.15.4.1365
• Doty, R.L. & Cameron, E.L. (2009). Sex differences and reproductive hormone influences on human odor perception. Physiology & Behavior 97, 213-228. doi:10.1016/j.physbeh.2009.02.032
• Geffroy, O., Descôtes, J., Serrano, E., Li Calzi, M., Dagan, L., & R. Schneider (2018) Can a certain concentration of rotundone be undesirable in Duras red wine? A study to estimate a consumer rejection threshold for the pepper aroma compound. Australian Journal of Grape and Wine Research 24, 88-95. doi:10.1111/ajgw.12299
• Geffroy, O., Li Calzi, M., Ibfelt, K., Yobrégat, O., Feilhès, C., & T., Dufourcq (2019). Using common viticultural practices to modulate the rotundone and 3-isobutyl-2-methoxypyrazine composition of Vitis vinifera L. cv. Fer red wines from a temperate climate wine region with very cool nights. OENO One 53, 581-595. doi:10.20870/oeno-one.2019.53.4.2459
• Geffroy, O., Lopez, R., Serrano, E., Dufourcq, T., Gracia-Moreno, E., Cacho, J., & V., Ferreira (2015) Changes in analytical and volatile compositions of red wines induced by pre-fermentation heat treatment of grapes. Food Chemistry 187, 243-253. doi:10.1016/j.foodchem.2015.04.105
• Griep, M.I., Mets, T.F., Vercruysse, A., Cromphout, I., Ponjaert, I., Toft, J., & D.L., Massart (1995) Food Odor Thresholds in Relation to Age, Nutritional, and Health Status. The Journals of Gerontology: Series A 50A, B407-B414. doi:10.1093/gerona/50A.6.B407
• King, E., Osidacz, P., Curtin, C., Bastian, S., & I., Francis (2011) Assessing desirable levels of sensory properties in Sauvignon blanc wines–consumer preferences and contribution of key aroma compounds. Australian Journal of Grape and Wine Research 17, 169-180. doi:10.1111/j.1755-0238.2011.00133.x
• Lacey, M.J., Allen, M.S., Harris, R.L.N., & W.V., Brown (1991) Methoxypyrazines in Sauvignon blanc Grapes and Wines. American Journal of Enology and Viticulture 42, 103-108.
• Lesschaeve, I. (2008) Wine consumer flavour preferences. Wine Active Compounds proceedings p71-74, Beaune, Edited by David Chassagne.
• Lund, C.M., Thompson, M.K., Benkwitz, F., Wohler, M.W., Triggs, C.M., Gardner, R., Heymann, H., & L., Nicolau  (2009). New Zealand Sauvignon blanc distinct flavor characteristics: sensory, chemical, and consumer aspects. American Journal of Enology and Viticulture 60, 1-12.
• McKay, M., Bauer, F.F., Panzeri, V., & A., Buica (2020). Perceptual interactions and characterisation of odour quality of binary mixtures of volatile phenols and 2-isobutyl-3-methoxypyrazine in a red wine matrix. Journal of Wine Research, 31, 49-66. doi:10.1080/09571264.2020.1723069
• Noble, A.C., Arnold, R.A., Buechsenstein, J., Leach, E.J., Schmidt, J.O., & P.M., Stern (1987) Modification of a standardized system of wine aroma terminology. American Journal of Enology and Viticulture 38, 143-146.
• Picard, M., de Revel, G., & S., Marchand (2017) First identification of three p-menthane lactones and their potential precursor, menthofuran, in red wines. Food Chemistry 217, 294-302. doi:10.1016/j.foodchem.2016.08.070
• Picard, M., Lytra, G., Tempère, S., Barbe, J.-C., de Revel, G., & S., Marchand (2016). Identification of piperitone as an aroma compound contributing to the positive mint nuances perceived in aged red bordeaux wines. Journal of Agricultural and Food Chemistry 64, 451-460. doi:10.1021/acs.jafc.5b04869
• Poitou, X., Thibon, C., & Darriet P. (2017) 1,8-Cineole in French red wines: Evidence for a contribution related to its various origins. Journal of Agricultural and Food Chemistry 65, 383–393. doi:10.1021/acs.jafc.6b03042
• Pons, A., Allamy, L., Schüttler, A., Rauhut, D., Thibon, C., & Darriet P. (2017). What is the expected impact of climate change on wine aroma compounds and their precursors in grape? OENO One 51, 141-146. doi:10.20870/oeno-one.2017.51.2.1868
• Pons, A., Lavigne, V., Darriet, P., & Dubourdieu, D., (2016). Identification and analysis of piperitone in red wines. Food Chemistry 206, 191-196. doi:10.1016/j.foodchem.2016.03.064
• Prescott, J., Norris, L., Kunst, M., & S., Kim (2005). Estimating a “consumer rejection threshold” for cork taint in white wine. Food Quality and Preference 16, 345-349. doi:10.1016/j.foodqual.2004.05.010
• Rajchl, A., Čížková, H., Voldřich, M., Lukešová, D., & Z., Panovska (2009). Methoxypyrazines in Sauvignon blanc wines, detection of addition of artificial aroma. Czech Journal of Food Sciences 27, 259-266. doi:10.17221/4/2009-CJFS
• Robinson, A.L., Ebeler, S.E., Heymann, H., Boss, P.K., Solomon, P.S., & R.D., Trengove (2009) Interactions between wine volatile compounds and grape and wine matrix components influence aroma compound headspace partitioning. Journal of Agricultural and Food Chemistry 57, 10313-10322. doi:10.1021/jf902586n
• Roessler, E., Pangborn, R., Sidel, J., & H., Stone (1978) Expanded statistical tables for estimating significance in paired-preference, paired–difference, duo–trio and triangle tests. Journal of Food Science 43, 940-943. doi:10.1111/j.1365-2621.1978.tb02458.x
• Roland, A., Vialaret, J., Razungles, A., Rigou, P., & R., Schneider (2010). Evolution of S-Cysteinylated and S-Glutathionylated thiol precursors during oxidation of Melon B. and Sauvignon blanc musts. Journal of Agricultural and Food Chemistry 58, 4406-4413. doi:10.1021/jf904164t
• Ross, C., Zwink, A., Castro, L., & Harrison, R. (2014). Odour detection threshold and consumer rejection of 1, 1, 6-trimethyl-1, 2-dihydronaphthalene in 1-year-old Riesling wines. Australian Journal of Grape and Wine Research 20, 335-339. doi:10.1111/ajgw.12085
• Roujou de Boubée, D., van Leeuwen, C., & D., Dubourdieu (2000). Organoleptic impact of 2-methoxy-3-isobutylpyrazine on red Bordeaux and Loire wines. Effect of environmental conditions on concentrations in grapes during ripening. Journal of Agricultural and Food Chemistry 48, 4830-4834. doi:10.1021/jf000181o
• Saliba, A.J., Bullock, J., & W.J, Hardie (2009). Consumer rejection threshold for 1,8-cineole (eucalyptol) in Australian red wine. Food Quality and Preference 20, 500-504. doi:10.1016/j.foodqual.2009.04.009
• Solomon, G.E. (1990). Psychology of novice and expert wine talk. The American Journal of Psychology, 103, 495-517. doi:10.2307/1423321
• Tempère, S., Pérès, S., Espinoza, A.F., Darriet, P., Giraud-Héraud, E., & A., Pons (2019). Consumer preferences for different red wine styles and repeated exposure effects. Food Quality and Preference 73, 110-116. doi:10.1016/j.foodqual.2018.12.009
• Tominaga, T., Peyrot des Gachons, C., & Dubourdieu, D. (1998). A new type of flavor precursors in Vitis vinifera L. cv. Sauvignon blanc: S-Cysteine Conjugates. Journal of Agricultural and Food Chemistry, 46(12), 5215–19. doi:10.1021/jf980481u
• Villamor, R.R., Evans, M.A., Mattinson, D.S., & C.F., Ross (2013). Effects of ethanol, tannin and fructose on the headspace concentration and potential sensory significance of odorants in a model wine. Food Research International 50, 38-45. doi:10.1016/j.foodres.2012.09.037
• van Wyngaard, E., Brand, J., Jacobson, D., & W.J., Du Toit (2014). Sensory interaction between 3‐mercaptohexan‐1‐ol and 2‐isobutyl‐3‐methoxypyrazine in dearomatised Sauvignon blanc wine. Australian Journal of Grape and Wine Research, 20, 178-185. doi:10.1111/ajgw.12082