A latitudinal gradient in Darwin’s naturalization conundrum at the global scale for flowering plants

Domain: earth_science
Published: 2023-10-12
Editorial outcome: minor_revision

Abstract
Darwin’s naturalization conundrum describes two seemingly contradictory hypotheses regarding whether alien species closely or distantly related to native species should be more likely to naturalize in regional floras. Both expectations have accumulated empirical support, and whether such apparent inconsistency can be reconciled at the global scale is unclear. Here, using 219,520 native and 9,531 naturalized alien plant species across 487 globally distributed regions, we found a latitudinal gradient in Darwin’s naturalization conundrum. Naturalized alien plant species are more closely related to native species at higher latitudes than they are at lower latitudes, indicating a greater influence of preadaptation in harsher climates. Human landscape modification resulted in even steeper latitudinal clines by selecting aliens distantly related to natives in warmer and drier regions. Our results demonstrate that joint consideration of climatic and anthropogenic conditions is critical to reconciling Darwin’s naturalization conundrum.

Introduction
Biological invasions have become ubiquitous across ecosystems worldwide and are a defining characteristic of the Anthropocene1,2. So far, over 13,000 plant species have successfully established self-sustaining populations beyond their native ranges (i.e., have become naturalized), and the increase in their numbers worldwide does not show any sign of deceleration3,4. These naturalized aliens have resulted in the homogenization of the world’s flora5,6 and exert major environmental, ecological and socioeconomic impacts7–9. The distribution of naturalized alien plants is highly uneven globally4,10, but the determinants of naturalization success and of global geographic patterns therein are not yet fully understood.

The phylogenetic relatedness between alien species and native species in a recipient region has long been suggested as a predictor of their naturalization success. In 1859, Charles Darwin invoked phylogenetic relationships in two seemingly contradictory hypotheses to explain the naturalization of alien species11. In his first hypothesis, now known as the preadaptation hypothesis, Darwin posited that alien species that are closely related to the native species in a regional flora are more likely to naturalize. This is because they are likely to share similar adaptations to the environments in the respective region with their native relatives11,12. In his second hypothesis, now known as Darwin’s naturalization hypothesis, Darwin posited that the native species in a regional flora could reduce the chances of naturalization for closely related aliens. This is because close relatives should compete more intensely with each other and also because natural enemies of native species might also attack the closely related alien species11,13. Together, these two seemingly opposing hypotheses, emphasizing the dominant roles of environmental filtering and biotic interactions, constitute Darwin’s naturalization conundrum14,15 (Fig. 1).

Solving the conundrum has gained increasing attention in recent years, with accumulating evidence in support of both hypotheses15–20. The apparent contradiction between the two hypotheses has been partly resolved by considering the spatial scale of studies14,21. Evidence for the preadaptation hypothesis is believed to be more detectable at regional scales, where the effects of environmental filtering are more apparent17. Conversely, Darwin’s naturalization hypothesis is thought to be more applicable at the scale of local communities where species directly interact. However, studies at regional scales have also revealed instances where naturalized species are distantly related to natives15,16. These observations could reflect that closely related species are generally more likely to share pathogens, parasites, and herbivores than distantly related species22–25, and these shared enemies can exert their influence over larger spatial scales. Multiple abiotic and biotic processes, including environmental filtering and shared enemies, could act simultaneously in regulating species naturalization, and their relative strengths could vary considerably even at the regional scale. Therefore, reconciling empirical support for Darwin’s two hypotheses across different regional studies remains a major challenge.

We propose that the contradictory findings on Darwin’s naturalization conundrum could be clarified by considering the biogeographical context of the study regions (Fig. 1). Darwin posited that in the temperate zone, adaptations are mostly driven by harsh climate, while towards the equator, the importance of biotic interactions should increase (now known as the latitudinal biotic interaction hypothesis26,27). We, therefore, expect that support for the preadaptation hypothesis should be stronger in high-latitude regions characterized by cold and highly seasonal environments. In contrast, in low-latitude regions, which usually have relatively stable environments, benign climates, and more intense biotic interactions (e.g., resource competition, herbivory, and disease)28,29, we expect a higher likelihood of supporting Darwin’s naturalization hypothesis. In other words, we predict that with increasing latitude, there should be a decrease in phylogenetic distance between native and naturalized alien plant species (Fig. 1). Indirect support for this hypothesis is provided by studies on native plant assemblages, which have observed an increase in phylogenetic clustering of native plants with latitude30. Additionally, some studies found that the phylogenetic structures of native or alien communities varies with climate31–33. These studies, however, did not specifically examine the phylogenetic distance between alien and native species and thus did not directly address Darwin’s naturalization conundrum. A notable exception is Park et al.17, which found that the phylogenetic distance between alien and native plant species increased with temperature and precipitation. However, being restricted to the continental USA, their study did not cover tropical or high-latitude environments and did not explicitly incorporate latitude as a factor. Therefore, a global-scale analysis of regional floras is required to determine whether a latitudinal gradient indeed exists in Darwin’s naturalization conundrum and whether it holds true across continents. Such an analysis could potentially clarify some of the seemingly inconsistent findings of previous studies focusing on single regions15–18.

In this study, to test for latitudinal variation in naturalized-to-native phylogenetic distances, we collated checklists of native and naturalized alien plant species for 487 regions around the world, covering more than 83% of the Earth’s ice-free land area. Our dataset included native distributions for 219,520 angiosperms (i.e., flowering plants), naturalized alien distributions for 9531 angiosperms, and phylogenetic information for the full set of species, which represents about 70% of the known global angiosperm flora. Utilizing this data, we calculated the mean pairwise phylogenetic distances (MPD) between naturalized alien and native plant species in each region (Supplementary Fig. 1). We then assessed how MPD varied with latitude and climate variables. Furthermore, given the strong association between species naturalization and human-induced environmental disturbances34,35, we additionally tested whether the degree of human modification to the landscape influenced the relationship of naturalized-to-native phylogenetic distance with latitude and climate. Our study reveals a clear global latitudinal pattern of Darwin’s naturalization conundrum, and provides evidence that this latitudinal pattern is largely driven by temperature and its seasonality. Moreover, our results demonstrate that these patterns become stronger with increasing human modification of the environment.

Results
We found a strong latitudinal gradient in the mean pairwise phylogenetic distances (MPD) between the naturalized alien and native plant species across the globe (Fig. 2a, c). In line with our prediction, naturalized alien species were more distantly related to natives within equatorial regions, and MPD decreased significantly toward the poles (Fig. 2c; P < 0.001). To quantify how much higher or lower these MPD values were than expected by chance, we calculated ΔMPD values as the deviations of the observed MPD values from expected values derived from a null model (Supplementary Fig. 1). To ensure the robustness of our findings across different null models, we used six different potential source pools from which the naturalized alien plant species in a recipient region may have derived: (1) a global nonnative species pool (Global nonnative flora), (2) the pool of species with economic uses combined with the global naturalized species pool (Econ. use flora), (3) the global naturalized alien species pool (Global nat.), (4) the pool of species that have naturalized in the continent of the recipient region (Continent nat.), (5) the global pool of species with climatic suitability in the recipient region (Climate nat.), and (6) the pool of climatically suitable species that have also naturalized in the continent of the recipient region (Climate continent nat.; see Methods for details; Supplementary Fig. 2). Irrespective of the choice of the species pools, ΔMPD consistently decreased with increasing latitude (Fig. 2d; P < 0.001 for all models). The phylogenetic distances between the naturalized alien and native plant species were significantly higher than expected in some low-latitude regions (e.g., Algeria and some Mexican states; Fig. 2b and Supplementary Figs. 3 and 4), consistent with the prediction of Darwin’s naturalization hypothesis. Conversely, in most mid- and high-latitude regions (e.g., Northern Europe and Canada), MPD was significantly lower than expected (Fig. 2b and Supplementary Figs. 3 and 4), which supports the preadaptation hypothesis. As previous studies have found, native flora usually have a phylogenetic clustering tendency under pronounced environmental filtering at high latitudes30. Our findings of a latitudinal gradient in phylogenetic distances between the naturalized alien and native plant species suggest that similar mechanisms contribute to the successful establishment of closely related alien plants in these environments.

In addition to analyzing the average MPD and ΔMPD values between all naturalized alien and native plant species in a region, similar patterns were obtained when we assessed the relationship between naturalization success (yes, no) versus phylogenetic distances of each of the 9531 naturalized alien plant species in our dataset to the natives in each region where they are not native (Fig. 3). At high latitudes, the naturalization probability of an alien species was higher in regions with closely related native floras, whereas, at low latitudes, the reverse was true (Fig. 3). Therefore, both the analysis at the regional flora and individual species level showed that the probability of observing evidence for the preadaptation hypothesis increases from the equator to the poles.

We further tested whether the geographic patterns were explained by global climatic gradients. We first performed a principal component analysis (PCA) on 19 biologically relevant climate variables36. The first PCA axis (PC Temp), explaining 45.7% of the climatic variation, primarily represented temperature and its seasonality, with high values indicating warm, non-seasonal environments (Supplementary Table 1). Notably, PC Temp exhibited a significant negative correlation with latitude (Pearson’s r = -0.943, P < 0.001, Supplementary Fig. 5a). The second PCA axis (PC Prec), explaining 31.7% of the climatic variation, mainly captured information related to precipitation and its seasonality, with high values representing humid regions with non-seasonal precipitation (Supplementary Table 1). Consistent with our expectations, MPD and the six ΔMPDs significantly increased with PC Temp and PC Prec (Fig. 4). In cold and seasonal regions (such as Northern Europe and Canada) or arid regions (such as Central Asia, Interior Australia, and the Southern Cone of South America), we observed a higher likelihood of naturalization for alien species closely related to natives (Fig. 2b and Supplementary Figs. 3 and 4). This finding aligns with the results of a previous study17, indicating that environmental filtering plays a more important role in determining species' naturalization in harsher climates. In warmer and non-seasonal regions (such as western Mexico), naturalized alien species were generally more distantly related to natives (Fig. 2b and Supplementary Figs. 3 and 4), which may reflect the relatively weak abiotic constraints imposed by benign climates. Instead, intense interactions with enemies and competitors may be more prevalent in these typically biodiverse regions, which tend to hinder the naturalization of closely related species27,37–40.

The latitudinal gradient of naturalized-to-native phylogenetic distances was evident in most continents, including Africa, Asia, Europe, Northern America, and Southern America (Supplementary Fig. 6). However, Australasia showed a reversed latitudinal gradient (Supplementary Fig. 6), which is consistent with its known latitudinal gradient of plant diversity41. This reversal is likely due to the pronounced increase in precipitation with latitude across Australasia compared to the other continents (Supplementary Figs. 5b and 7b). Aridity appears to act as the primary environmental filter for species naturalization in this continent. Consistently, we found that both MPD and the ΔMPDs decreased in regions with low PC Prec across Australasia (Supplementary Fig. 7). This is in line with the idea that Australasia, characterized by the second lowest precipitation levels after Antarctica, exhibits biodiversity patterns strongly shaped by variation in precipitation42. Therefore, the reversed gradient in Australasia highlights the necessity of conducting studies spanning multiple continents, as they not only confirm the general patterns of species naturalization at a macro-ecological scale but also reveal exceptions that can provide insights into underlying mechanisms.

While macroclimatic conditions can strongly constrain naturalization success, it is important to note that these climatic variables, along with other potentially influential environmental factors43, are also subject to modification by human activities in various ways and to different degrees across the globe. Therefore, to investigate the potential influence of human modification on the observed latitudinal and climatic patterns, we further explored the interactions between the human modification index (HM) and latitude, as well as climate variables, on MPD and ΔMPD. The HM index quantifies the extent to which humans have changed land cover44. Our analysis revealed that higher values of HM strengthened the latitudinal pattern (Fig. 2e, f; P < 0.001 for the interaction between HM and latitude in all models) and resulted in steeper gradients of MPD and ΔMPD with PC Temp (Fig. 4b, d). The steeper gradients primarily arose from higher MPD and ΔMPD values at low-latitude, warm regions. This observation could be attributed to the relatively recent intense human modifications in tropical regions that have created many novel environments34,45. Such disturbances may particularly benefit alien species from distantly related temperate clades that have preadapted to anthropogenic landscapes through their long histories in human-modified environments (e.g., in Europe and Asia)46. While the precise mechanisms warrant further investigation, our results suggest that with increasing human modification of environments worldwide, the latitudinal gradient of naturalized-to-native phylogenetic distances might steepen.

We also found that in regions with low levels of human modification, increasingly arid conditions (i.e., lower PC Prec values) were associated with lower naturalized-to-native phylogenetic distances (Fig. 4c). This result is again consistent with environmental filtering as a primary constraint on the naturalization of alien species in harsh climates. However, with increasing human modification, the positive relationship between MPD and PC Prec became shallower and was even reversed to negative in regions with the highest human modification (Fig. 4c, e). Possibly, this reflects that human modifications, such as the construction of artificial water bodies and other forms of water management, as well as irrigation of agricultural land, locally alleviate water limitation in arid regions. These human modifications could facilitate the establishment of distantly related alien species that would otherwise be filtered out by the arid natural conditions.

Our findings resonate with previous studies indicating that both environmental filtering and biotic interactions act simultaneously and thus jointly shape naturalization success15,47. Our analysis revealed that in some tropical regions, the observed naturalized-to-native phylogenetic distance was larger than expected. These results thus support Darwin’s naturalization hypothesis, indicating that in these tropical regions, the influence of biotic interactions may outweigh environmental filtering. The reverse pattern was observed in most other regions and particularly in the cold, seasonal environments found at high latitudes, indicating that environmental filtering predominates in most regions. The prevalence of negative ΔMPD values in our study is likely attributable to our focus on regional floras rather than local communities. Indeed, previous studies have found that negative relationships between alien-to-native phylogenetic distance and naturalization success were more likely found for larger spatial units (e.g., administrative units, like in our study), whereas positive relationships are more likely, but not exclusively, observed at local spatial scales (e.g., within vegetation plots), where alien and native plants are more likely to directly compete16,17. At the spatial resolution of our study, the alien and closely related native plant species may not necessarily directly interact with each other. Nevertheless, it is important to recognize that they can still affect each other indirectly through various biotic interactions, such as shared pathogens, herbivores, pollinators, and seed-dispersers. It should also be noted that the occurrence or absence of species at a regional scale is an emergent outcome of local processes. The regional lists of native species provide an indication of the potential compositions of local communities and the probability of an alien encountering closely or distantly related species. While our data do not allow for direct observation of local interactions, we can still detect a signal of their influence at regional scales. In fact, our results align with the prediction of the latitudinal biotic interaction hypothesis, which suggests that biotic interactions are more pronounced at lower latitudes27.

While our analyses encompassed extensive data covering most of the terrestrial world, we acknowledge the potential for future advancements. Our study encountered limitations in data coverage, particularly in parts of Southeast Asia and Africa (Fig. 2), emphasizing the importance of cautious interpretation to account for potential biases. It is therefore crucial to prioritize efforts in obtaining more comprehensive species lists, particularly for naturalized aliens, in these underrepresented regions. With the continuous expansion of public datasets, there is an increased opportunity to explore latitudinal and climatic patterns of alien-to-native phylogenetic relatedness, not only across regional floras but also across local communities (i.e., vegetation plots) and across various spatial scales. Furthermore, it is worth noting that our phylogenetic tree, although informative, contains several polytomies at the genus level due to the current unavailability of a fully resolved global flora phylogeny. However, it has been shown that the use of genus-level phylogenetic trees instead of fully resolved ones have minimal impact on measuring patterns of phylogenetic relatedness among collections of species that contain different families or genera48–52. This is because the deep branches of the phylogenetic tree have a strong influence on the overall patterns. Nevertheless, we acknowledge that our analyses could be improved with the advent of a more accurate species-level phylogenetic tree.

While Darwin clearly referred to the importance of the phylogenetic relatedness between aliens and native species in a region11, he evoked phylogenetic relatedness as an integrative surrogate for similarity in key functional traits and niche preferences between alien and native species. This assumption is reasonable considering the prevalence of niche conservatism53. The current lack of comprehensive trait data and methods of linking the different traits to niche differences pose limitations54. Future global studies should strive to incorporate functional distances and niche similarities between alien and native species in order to enhance our understanding of Darwin’s conundrum. Moreover, as it was recently shown that the effect of alien-to-native phylogenetic distance might vary among the different stages of the invasion process19,55, future studies should also consider those different stages under varying geographic and environmental contexts. Together with important insights from previous studies, our findings of latitudinal and climatic gradients in the effect of phylogenetic distance on naturalization and the effects of human environmental modification on these relationships will help to solve this 160-year-old Darwin’s naturalization conundrum.

Methods
Native and naturalized alien angiosperm floras
We extracted regional lists of naturalized alien and native angiosperms (i.e., flowering plant species) from the Global Naturalized Alien Flora (GloNAF) database56 and the Global Inventory of Floras and Traits (GIFT) database57,58, respectively. As the most comprehensive global database of naturalized alien plants, GloNAF contains lists of naturalized alien vascular plant species for various regions (e.g., countries, states, provinces, islands) that together cover most of the world’s ice-free terrestrial surface10,56. GIFT is a compilation of floras and checklists of predominantly native plant species with an indication of their native status for more than 300,000 species across nearly 3000 regions with near-global coverage57,58. We selected regions present in both GloNAF and GIFT. Additionally, we merged some GloNAF regions to match the higher-level administrative divisions in GIFT regions and vice versa. To further increase the spatial coverage of our analysis, we also collected checklists of native species for some GloNAF regions from other data sources (see Supplementary Table 2 for details), when the native species lists were not yet included or incomplete in GIFT.

Regions with data on native and naturalized alien plants vary largely in their size. Because the study scale may affect the relationship between phylogenetic relatedness and species naturalization17, we reduced this potential effect by restricting our analyses to regions larger than 5000 km2. Small mainland regions were merged into the country that they belong to, and small oceanic islands were excluded from our analysis. However, to assess the sensitivity of the analyses to the region-size threshold, we also conducted additional tests using thresholds of 0, 1000, and 10,000 km2, which confirmed that patterns were consistent regardless of the threshold (Supplementary Table 3). Furthermore, including region area in our statistical model did not change the statistical significance and signs of the effect sizes of the explanatory variables that we were interested in (Supplementary Fig. 8). We therefore only report results without region area as an explanatory variable in the main text. Our selection criteria based on the 5000 km2 threshold resulted in 487 regions, which in total cover more than 83% of the world’s terrestrial ice-free surface.

Because regional checklists of non-angiosperms are usually not as complete as those of angiosperms, we restricted our analysis to angiosperms. We harmonized all taxonomic names of both naturalized alien and native species using the R package Taxonstand v.2.459, according to The Plant List (http://www.theplantlist.org/), which was used as a taxonomic reference list for both the GloNAF56 and GIFT57,58 databases. We excluded hybrids and assigned infraspecific taxa to the binomial species level. If infraspecific taxa were standardized to two different accepted binomial names, they were considered to be separate species, and their native or alien status was assigned accordingly. If multiple infraspecific taxa were standardized to the same binomial species name, the species was considered native if at least one of the infraspecific taxa was classified as native. Species with an uncertain native/alien status or with a conflicting status (including 1286 of 1,156,207 records in total), i.e., being native to a region according to GIFT but being naturalized to the same region according to GloNAF, were assigned as native species. However, the results remained consistent when we assigned them as naturalized aliens (Supplementary Fig. 9). After cleaning the data, our final dataset encompassed distributions of 9531 naturalized alien species and 219,520 native species across 487 regions, which represent ~70% of the global angiosperm flora (Supplementary Fig. 10).

Phylogenetic analyses
To calculate the phylogenetic distances between species, we constructed a phylogenetic tree for the global flora using accepted names from The Plant List. As a basis for this tree, we used the phylogeny “ALLMB” constructed by Smith and Brown (2018)60, which is to date one of the most comprehensive trees for seed plants. We first standardized the taxon names of this tree with The Plant List using the package Taxonstand59, and pruned the tree to only include taxa with accepted names in The Plant List. Some plants with accepted names in The Plant List were not found in the tree, and we therefore added them manually. Specifically, for species that were missing but had congeneric species present in the tree (7.2% of species), we grafted those species to the roots of the corresponding genera. When the entire genus was missing from the tree (1.3% of species), we grafted the species to the roots of the families they belong to. We then pruned the global flora tree to only include the 224,593 species analyzed in this study (including all native angiosperms, naturalized alien angiosperms, and economically useful angiosperms mentioned below). Some species in our megatree species were added as polytomies of their genera or family, while others are fully resolved at the species level. This approach might introduce potential biases when examining latitudinal and climatic gradients in naturalized-to-native phylogenetic distances. Therefore, we also created a tree with a lower but more even phylogenetic resolution across regions. This was achieved by adding all species to the roots of the corresponding genera or, when the entire genus was missing from the tree, to the roots of the families they belong to. We found that the use of this less-resolved tree yielded similar results (Supplementary Fig. 11). This finding supports recent studies showing that phylogenetic metrics derived from mega-phylogenies resolved only at the genus level and those derived from fully resolved species-level trees are nearly perfectly correlated (Pearson’s correlation coefficients ranged from 0.966 to 1.000)51,52. The reason for this high correlation is that the assembled communities typically contain distantly related species across different families or genera that traverse deep nodes. Thus, fine-tuning the relative distance among species within the same genus has minimal impact on phylogenetic distance metrics.

For each region, the phylogenetic distance between naturalized aliens and native species can be calculated as the mean pairwise phylogenetic distance (MPD) and as the mean nearest taxon distance (MNTD)61,62. MPD represents the average of the pairwise phylogenetic distances between naturalized aliens and the native species in a region (Supplementary Fig. 1), and could therefore capture both shallow and deep branching in the phylogenetic tree62,63. In contrast, MNTD estimates the average of the phylogenetic distances of each naturalized alien species with its closest native relative in the focal region and is more strongly influenced by patterns towards the tips of the tree62,63. As it is difficult to achieve good resolution at the tips of the large megaphylogeny with numerous polytomies, we opted to only use MPD in our analysis.

To assess whether the naturalized alien species were more closely or more distantly related to the native ones than expected by random chance, we adopted a null-model approach. This involved comparing the observed MPD between naturalized alien and native species to the distribution of simulated distances that were drawn randomly from a potential alien species source pool. Specifically, we conducted 1000 random simulations by keeping the native species composition and the number of naturalized alien species in a region unchanged but randomly drawing the identities of the naturalized alien species from the respective source pool. Hence, this approach allowed us to account for the different potential source pools of alien species, the likelihood of introduction to a region, and the probability of successful naturalization based on the climatic suitability of the focal regions. For each region and null model, we initially calculated the standardized effect size of MPD (SES.MPD) as (observed MPD – mean of the expected MPDs) divided by the standard deviation of the expected MPDs. While SES.MPD provides information about whether the observed MPD value of a region deviates significantly from the expected values, it is less suitable for comparisons among regions because of SES.MPD values are sensitive to variation in species richness64,65. To avoid this, it has been suggested to use rarefaction64, but because the variance in MPD values expected from randomizations is higher in species-poor regions, rarefaction unnecessarily reduces the certainty of estimates for many regions65. Therefore, we instead calculated ΔMPD as the difference between observed MPD and the mean of the expected MPDs (Supplementary Fig. 1) and used the inverse of the variance of the distribution of expected MPDs as a weight in the statistical models to account for uncertainty (see the section ‘Statistical analyses’ below). Nevertheless, if we analyzed SES.MPD instead of ΔMPD, the results were very similar (Supplementary Figs. 12 and 13). Positive values of ΔMPD (or SES.MPD) indicate that naturalized alien species are more distantly related to native species in a region than expected by chance, aligning with Darwin’s naturalization hypothesis. Conversely, negative values indicate that naturalized alien species are more closely related to native species, supporting the preadaptation hypothesis. All phylogenetic distance indices were calculated using the R package picante v.1.8.266.

How a potential source pool of naturalized alien species is defined can affect the output of the null models and the interpretation of the results67. To assess whether the patterns we found were robust, we considered six different potential pools of alien species (Supplementary Fig. 2). These pools of potentially introduced species for the focal region differed in size. The first species pool included all species not native to the focal region (i.e., the global nonnative species pool, global nonnative flora). As particularly alien species with economic uses, such as for human food and animal fodder, are likely to be introduced and become naturalized68, we used as a second species pool of a focal region all non-native species that have a known economic use. However, because that would exclude naturalized species without known economic use, we also included in this species pool all other species from the global naturalized flora (i.e., the pool of economic-use species combined with the global naturalized species pool, econ. use flora). Data on species with economic uses was extracted from the World Checklist of Useful Plant Species69. As a third species pool, we used all species that have become naturalized in at least one region worldwide (i.e., the global naturalized alien species pool, global nat.). While it is likely that some naturalized alien species have been introduced into the great majority of the world, this might not apply to all naturalized alien species. Therefore, we used, as a fourth species pool, all species that have naturalized in at least one region of the biogeographical continent that the focal region is part of (i.e. the pool of species that have naturalized in the same continent as a given recipient region, continent nat.). This assumes that when a species is naturalized in at least one region of a continent, it is more likely to have also been introduced to all other regions of that continent. For this null model, we divided the world into eight continents using the Taxonomic Databases Working Group continent scheme (TDWG, including Africa, Asia, Europe, South America, North America, Antarctica, the Pacific Islands, and Australasia, https://www.tdwg.org/standards/wgsrpd/). We combined the TDWG continental regions Asia-Temperate and Asia-Tropical into Asia because of the short latitudinal range in Asia-Tropical.

The fifth species pool considered the climatic suitability of the naturalized alien species for the focal region (i.e., the global pool of naturalized species with climatic suitability in a recipient region, climate nat.). This approach accounts for the fact that humans are more likely to intentionally introduce climatically suitable plant species than climatically unsuitable ones70,71. Additionally, this null model controls for the fact that species from across temperate continents tend to be more closely related than those from across tropical continents72. To predict the potential geographic distribution of each species in the global naturalized flora, we employed maximum entropy modeling (MaxEnt v.3.4.1) using the R package dismo v.1.3.573. This modeling was based on bioclimatic variables from both its native and naturalized ranges, and relied on six climate variables extracted from WorldClim 2.5 arc-min resolution. These variables included mean diurnal range, temperature seasonality, the maximum temperature of the warmest month, precipitation seasonality, precipitation of the wettest quarter, and precipitation of driest quarter, which are known to be related to species distributions. To ensure the reliability of our models, we examined pairwise Pearson’s correlations between these climate variables and found them to be below 0.7. This indicates that collinearity among the variables should have a negligible effect on subsequent model estimations and predictions74. Ideally, climatic niche modeling would rely on known point-occurrence data instead of regional occurrence data. Although some widely naturalized alien species (i.e., invasive species) have extensive point-occurrence records, comprehensive point-occurrence data for all naturalized alien species at a global scale is currently unavailable. Consequently, we used coarse occurrence data (i.e., regional occurrence data) to construct species distribution models. To do this, the global map was rasterized into 50 km × 50 km grid cells, with the assumption that a species that occurs in a region resides in each grid cell of that region. Additionally, we upscaled the six bioclimatic variables to match the resolution of the regional occurrence data. To ensure the reliability of our models in predicting global distribution of all naturalized alien species, we firstly focused on species with more than 15 occurrences, as that number is considered to be sufficient for running a species distribution model75. For each naturalized alien species, we then allocated 80% of the distribution data for model training and 20% of the distribution data for testing the model’s performance. We retained models with an area under the curve (AUC) greater than 0.7 (considered a valid model)76. Using these valid models, we proceeded to predict the climatic suitability of each species in every grid cell, based on climatic data in the 487 regions. Subsequently, we extracted the potentially climatically suitable regions for each naturalized alien species by using the maximum training sensitivity plus specificity threshold. Specifically, if a region has at least one grid cell with suitability not lower than the threshold, then we postulate the species to be climatically suitable for this region. Finally, we defined the climatically suitable species pool for each focal region by assuming that only the species whose potential distribution overlaps with the focal region could naturalize there. The sixth and last species pool was even more strict and combined the criteria used to make the fourth and fifth species pools. In other words, the sixth species pool assumed that for a focal region, only the species that are both climatically suitable and have also naturalized in the continent of the focal region have a chance to naturalize there (i.e., the pool of climatically suitable species that have also naturalized in the continent of the recipient region, climate continent nat.).

Climate variables and human modification index
We considered climate variables and human modification of the landscape as potential drivers of global patterns in the naturalized-to-native phylogenetic distance. For climate variables, we used all 19 bioclimatic variables from WorldClim36 at 2.5 arc-min resolution. To account for collinearity among the 19 bioclimatic variables, we performed a PCA on the centered and scaled bioclimatic variables. Due to the skewed distribution of some variables, we used various transformations to approximate normal distributions (see Supplementary Table 1) by using the R package normalizer v.0.1.077. The first two PC axes (PC Temp and PC Prec) accounted for 77.8% of the variation, where PC Temp mainly reflects the temperature and its seasonality, and PC Prec mainly reflects precipitation and its seasonality (Supplementary Table 1). Then, we calculated PC Temp and PC Prec for each region as the mean value across all grid cells in that region. These two variables are also strongly correlated with the global aridity index (AI), and potential evapotranspiration (ET)78 (Supplementary Fig. 14). Therefore, we only considered PC Temp and PC Prec in our analyses, which provide independent indices of temperature and precipitation effects.

To characterize the influence of anthropogenic stressors, we incorporated the global human modification index (HM). HM is strongly positively correlated with human population density79 (Supplementary Fig. 14), but is an index that much more comprehensively quantifies human-caused changes of terrestrial lands (including changes due to human settlement, agriculture, transport, energy production and electrical infrastructure)44. The HM is available at 1-km2 resolution and ranges from 0 (low human impact) to 1 (high human impact)44. The average HM for each region was calculated as the mean value across all grid cells in that region.

Statistical analyses
To describe the general pattern of naturalized-to-native phylogenetic distance along the latitudinal gradient, we first used linear regressions to test whether there were statistically significant relationships (P < 0.05) of latitude with MPD and the six ΔMPDs, each based on a different null model. To explore how human modification of the environment affects the latitudinal gradient of phylogenetic distance, we additionally ran multiple linear regression models of MPD and ΔMPD on latitude, HM, and their interaction. We used the R package visreg v.2.7.080 to visualize the effect of the significant interaction between HM and latitude (P < 0.05; Fig. 2e, f). For plotting, we set the HM values to the 10%, 50%, and 90% quantiles of the HM values across all regions.

We further performed multiple linear mixed-effects models to assess the relative importance of PC Temp, PC Prec and their interactions with HM on MPD, and the six ΔMPDs. To account for differences among the TDWG continents, we included the continent as a random intercept. To account for uncertainty in the estimation of ΔMPD, we used the inverse of the variance of the distribution of expected MPDs as a weight in the ΔMPD models. Linear mixed-effects models were performed in the R package lme4 v.1.1.2881. The standardized effect sizes of the predictor variables in the above models were calculated using the R package effectsize v.0.6.0.182. Again, we used the R package visreg v.2.7.080 to visualize the effect of the significant interaction (P < 0.05) between HM and climate variables (PC Temp and PC Prec) on MPD and ΔMPD based on the global naturalized species pool. Specifically, we plotted the partial relationship of climate variables with MPD and ΔMPD for the 10%, 50%, and 90% quantiles of HM. R2 in all models has been adjusted, and all statistical analyses were conducted in R v.4.0.383.

The analyses described in the preceding section calculate for each focal region the average of the MPD values across the naturalized alien species in that region, which means that region is the unit of analysis. As an alternative analysis, we calculated MPD of each of the 9531 naturalized alien species in our dataset to the natives in each of the 487 regions where the species is not native. Then we ran a binomial generalized linear mixed model, using the R package lme4 v.1.1.2881, across all species and regions to test how the naturalization success of a species in a region relates to its MPD to the natives in that region, the latitude of the region and their interaction. To account for non-independence of observations for the same species and the same region, we included species identity and region identity as random factors. Furthermore, as species might vary in their responses to MPD and latitude, we also included random slopes of species with regard to MPD, latitude, and their interaction. Since the binomial response variable had many more zeros than ones, we used the complementary log-log link function84. For visualizing the statistically significant interactions of MPD and latitude (P < 0.05), we fixed latitude at several fixed values to depict the predicted relationship between species naturalization success and MPD at different latitudes.

Considering the variation in species inventory completeness among regions, with notably better documentation of species in highly developed areas, we conducted additional tests to identify potential biases that this variation might introduce to the latitudinal patterns we found. Specifically, we first focused on the regions (467 regions) where the estimated completeness of naturalized species lists exceeded 50%, based on completeness-estimate scores from the GloNAF database56. In this subset of regions, we re-analyzed the linear relationship between MPD and latitude. Second, we obtained the average native species inventory completeness percentages in 430 regions from Dawson et al. (2017)1. Then we developed the linear regression model of MPD with latitude for these 430 regions, incorporating the average inventory completeness percentage as a weighting variable. This weighting approach assigns greater weight to regions with higher completeness levels. As these additional analyses still revealed a latitudinal cline in naturalized-to-native phylogenetic relatedness (Supplementary Table 4), we only show the results based on all regions in the main text.

Peer review rounds
[{"round": 1, "reviewer_comments": {"reviewer_1": "Review for Nature Communications\nTitle: A latitudinal gradient in Darwin’s naturalization conundrum at the global scale\nGeneral comments\nWith a large-scale, macroecological approach, this study tests whether an important challenge in\ninvasion biology, Darwin’s naturalization conundrum, can be solved when taking into account large-\nscale global differences in latitude, temperature and precipitation, and human land use. The striking\nresult is that indeed, most of the expectations are met: At high latitudes and harsh environments,\nalien and native plant species are closer related to each other, indicating advantages of pre-\nadaptedness, whereas at low latitude and less harsh conditions, aliens and natives are less closely\nrelated, indicating advantages of being dissimilar and thus more novel for competitors and enemies.\nHuman landscape modification even enhances the pattern.\nThis is a remarkable study. The authors report on a very meaningful pattern that strongly aligns with\nconceptual considerations. It seems they found evidence for general rules about naturalization success\nthat are linked to likely mechanistic explanations – a very rare case in ecology. It is remarkable that\neven the exception they found (Australasia) turned out to match the provided explanations very well\nwhen looking closer.\nAs far as I can judge, the data sources are of high quality. It seems the authors took great care in\nassembling a data set that is highly suited to test the research question. The approach they took to\nassembling it is sophisticated and appropriate. The overall approach is well-chosen. I appreciate that\nthe authors test for effects of temperature and precipitation in addition to latitude, and the use of\ndifferent plant species pools for null models. I especially liked the idea of having a species pool that\nincludes economically used species, since this is data I have not very often seen being used for these\nkinds of analyses.\nThe methods and results are presented and visualized in a clear and understandable way. The\npresentation of the results in maps is very appealing and approachable. Also, figures S1 and S2\nincluded in the supplement are quite helpful for clarifying the approach of the analyses. However, the\nunderlying conceptual framework, Darwin’s naturalization conundrum, is pretty complex, and I wonder\nif there would be some space left for a conceptual diagram, helping readers not familiar with these\nideas to understand the underlying considerations.\nAll in all, the study results seem robust, and highly reliable, and the conclusions are considerate and\ndo not contain any over-interpretation. The text is well-written and there is a good line of argument\nguiding the reader.\nTo conclude, this study is a rare example where empirical results neatly match major hypotheses, and\nan impressive demonstration of the power of large-scale analysis of global datasets combined with\nconceptual depth. Usually, I am quite a critical reviewer (and not generally in favor of macroecological\nstudies), but in this case, I did not find any major points of critique. I strongly recommend publication.\nNote: I have reviewed a previous version of this manuscript for a different journal, and this report\nbuilds on my previous review. In my point of view, this current version is much stronger than the\nprevious version I read, with improved analyses and language.\nMinor comments\nLine 82: “be” is missing (“can be clarified”)\nLine 119: I think it is confusing to call the global alien species pool ‘global flora’ because to me this\nsounds as if native species were included.\nLine 197 to 200: Something is wrong with this sentence, please revise.\nLines 440-453: The final data set giving alien and native species per region, compiled and cleaned as\nspecified in the method section, is a highly valuable resource for future studies. Do you plan on\npublishing this as well?\nReviewer: Tina Heger\nORCID https://orcid.org/0000-0002-5522-5632", "reviewer_2": "The authors test Darwin’s naturalization conundrum at a global scale, and find a latitudinal gradient in\nhow closely related naturalized angiosperm species are to native ones. They also examine how this\ntrend may be mediated by human landscape modification. The manuscript is clearly written, but lacks\nsome detail – this may be partially due to the fairly short format it was written in. The study largely\nconfirms previous studies that have found that the relatedness between species is greater in harsher\nclimates (which underlies the overall latitudinal gradient; e.g., Qian et al. 2013; LI et al. 2014;\nEscoriza & Ruhí 2014; Park et al. 2020; Kusumoto et al. 2021; Zu et al. 2023) and albeit at a larger\nscale. Likewise, the effects of human landscape modification on phylogenetic community patterns\nhave been investigated, but at smaller scales and more heterogeneous results (e.g., Cui et al 2019;\nYang et al 2022). The larger scale of this study is both its strength and weakness. It lends support to\nprevious findings at larger scales and provides an interesting, if not surprising, global-scale view of\nhow human dislocated species may integrate into regional flora. However, this comes at a cost – the\nunderlying data are not as detailed/accurate, the coarse spatial resolution cannot account for biotic\ninteractions among taxa, and thus the results do not advance our knowledge as to why we observe\nthese patterns. There is a proliferation of large-scale correlative analyses based on data that others\nhave collected aimed at testing hypotheses on global scales. Though such studies have value, they\noften do not add new knowledge to the field, especially when the gathered data are not made\navailable in their entirety. Along these lines, the study is somewhat descriptive and does not introduce\nnovel insights, new methods, or new data. Again, the strength and novelty of this study are in its\nlarge-scale and the assemble global dataset of native and naturalized plants, but without making the\nunderlying data fully accessible, it is less useful. Please make data openly available and not “by\nrequest”.\nThe examination of human landscape modification effects is a potentially interesting addition that has\nnot been investigated as often. However, I think there needs to be some tests to rule out the\npossibility that the observed patterns are due to lack or resolution and/or biases in the data. For\ninstance, highly developed nations with extensively modified landscapes are often the places where\nboth native and non-native species are best documented. These nations also simultaneously engage in\nlarge amounts of international trade and travel, in effect, increasing propagule pressure of exotic\nspecies and their chances of establishment (Dawson et al 2017). These developed nations mostly\nreside in higher-latitude temperate regions. Similarly, it is possible that higher-latitude regions with\nsimilar floras (e.g., North America and northeast Asia) engage in more biotic exchange than they do\nwith lower-latitude regions, as suggested by some of the authors’ previous work (van Kleunen et al.\n2015).\nSome of the methodological choices need more justification/explanation. For instance, why is >5000\nkm2 the threshold for analysis? Why was The Plant List used for taxonomic harmonization when it has\nbeen static since 2013 and replaced? Several of the 19 bioclimatic variables used to model species\ndistributions are highly correlated and should not be included in the same models. Sampling biases\nassociated with the data sources and their potential effects on the results should be discussed in more\ndetail.\nAlso, while the authors should be commended for their testing of multiple null models, I am curious\nas to why native species were not included as candidates when shuffling taxa. At the end of the\nday Darwin’s naturalization conundrum is a hypothesis of community assembly and thus should consider\nthe insertion of any species that has the means to physically reach a community, regardless of its\nnative status. For example, the fact that native species X is not present in community/region A despite\noccurring nearby for millennia, while non-native species Y from the other side of the world has\ninserted itself over the last 50 years tells us something.\nFigure legends and sizing could be better – for instance, it was very difficult to see the inset map in\nFigure 1. Also, figure captions could be clearer. Individual descriptions of each panel may help. How\nwere the colors binned? The bins do not seem to be equal in size, both in the main figures and the\nsupplementary figures.\nTable S2: Where do the IDs for each region come from?\nLine 82: There’s a word missing. Think it should be “can be clarified”\nLine 92: Another word missing. Should be “would allow us to test”\nLine 106-107: The statement is slightly confusing/misleading. The study examines whether the degree\nof human modification affects the relationship between latitude (and climate) and the relatedness\nbetween native and naturalized species. It does not, however, examine if/how human modification\nalters the biotic/abiotic environment. Also, I do not think the study actually tests “whether the degree\nof human modification of the landscape has changed the latitudinal gradient” – it only shows a\nsignificant interaction term.\nLine 166: “could also” seems strange here. Latitude is often seen as a proxy for climatic factors and\ndoes not generally have a direct effect on biodiversity.\nLine 259: How many taxa had conflicting native statuses? How were native vs non-native subspecies\ntreated? Do the results change if taxa with conflicting status were considered non-native?\nLine 285: Genus-level phylogenies may be sufficient in certain circumstances but such a blanket\nstatement is improper and can be dangerous. Further, the test that was conducted – making all\ngenera polytomies and redoing analyses – does not demonstrate that the lack of phylogenetic\nresolution does not affect the results. What it is showing is that the results from the tree used in\nthe study, which is not fully resolved nor fully supported by molecular data, are not largely different\nfrom a tree with worse resolution. It does not prove that the results from the used tree are congruent\nwith those that would be generated from a more robust, resolved phylogeny supported by molecular data.\nPerhaps the authors could limit their analyses to regions for which they have full phylogenetic\ncoverage and see if the results still stand?\nLine 332: The rationale for limiting a species pool to species with known economic uses is not clear,\nespecially when the focal taxon itself does not have a known use, and needs to be better fleshed\nout/justified. Why not just limit this pool to species that have been introduced somewhere?\nLine 365: The modeling of species distributions with such coarse data are troubling. 250 km2 can\ncomprise extreme amounts of environmental heterogeneity – e.g., coastal invaders can become inland\ninvaders of the desert or alpine invaders. Perhaps some comparisons between these crude models and\nthose fit to actual point occurrence data could be compared for taxa with enough data to support this\napproach – many (if not most) invasive species have large amounts of occurrence data available. Also,\nall model parameters should be provided to ensure reproducibility."}, "author_response": "Responses to the Reviewer 1’s comments\nGeneral comments\nWith a large-scale, macroecological approach, this study tests whether an important challenge in\ninvasion biology, Darwin’s naturalization conundrum, can be solved when taking into account large-\nscale global differences in latitude, temperature and precipitation, and human land use. The striking\nresult is that indeed, most of the expectations are met: At high latitudes and harsh environments,\nalien and native plant species are closer related to each other, indicating advantages of pre-\nadaptedness, whereas at low latitude and less harsh conditions, aliens and natives are less closely\nrelated, indicating advantages of being dissimilar and thus more novel for competitors and enemies.\nHuman landscape modification even enhances the pattern.\nThis is a remarkable study. The authors report on a very meaningful pattern that strongly aligns\nwith conceptual considerations. It seems they found evidence for general rules about naturalization\nsuccess that are linked to likely mechanistic explanations – a very rare case in ecology. It is\nremarkable that even the exception they found (Australasia) turned out to match the provided\nexplanations very well when looking closer.\nRESPONSE: We sincerely thank the reviewer for this very positive evaluation.\nAs far as I can judge, the data sources are of high quality. It seems the authors took great care in\nassembling a data set that is highly suited to test the research question. The approach they took\nto assembling it is sophisticated and appropriate. The overall approach is well-chosen. I appreciate\nthat the authors test for effects of temperature and precipitation in addition to latitude, and the use\nof different plant species pools for null models. I especially liked the idea of having a species pool\nthat includes economically used species, since this is data I have not very often seen being used\nfor these kinds of analyses.\nRESPONSE: We thank the reviewer for the appreciation of the assembled data and the approach\nwe used. We are pleased to see that this reviewer notes the value of including the economic\nspecies pool, as we also discuss this species pool below in response to a comment by Reviewer\n2.\nThe methods and results are presented and visualized in a clear and understandable way. The\npresentation of the results in maps is very appealing and approachable. Also, figures S1 and S2\nincluded in the supplement are quite helpful for clarifying the approach of the analyses. However,\nthe underlying conceptual framework, Darwin’s naturalization conundrum, is pretty complex, and I\nwonder if there would be some space left for a conceptual diagram, helping readers not familiar\nwith these ideas to understand the underlying considerations.\nRESPONSE: This is a very good suggestion, and we now included a diagram illustrating the\nunderlying conceptual framework (see Fig. 1)\nAll in all, the study results seem robust, and highly reliable, and the conclusions are considerate\nand do not contain any over-interpretation. The text is well-written and there is a good line of\nargument guiding the reader.\nTo conclude, this study is a rare example where empirical results neatly match major hypotheses,\nand an impressive demonstration of the power of large-scale analysis of global datasets combined\nwith conceptual depth. Usually, I am quite a critical reviewer (and not generally in favor of\nmacroecological studies), but in this case, I did not find any major points of critique. I strongly\nrecommend publication.\nNote: I have reviewed a previous version of this manuscript for a different journal, and this report\nbuilds on my previous review. In my point of view, this current version is much stronger than the\nprevious version I read, with improved analyses and language.\nRESPONSE: We thank the reviewer for this very positive evaluation.\nMinor comments\nLine 82: “be” is missing (“can be clarified”)\nRESPONSE: Corrected (Line 82)\nLine 119: I think it is confusing to call the global alien species pool ‘global flora’ because to me this\nsounds as if native species were included.\nRESPONSE: We agree, and now refer to this species pool as the ‘global nonnative species pool’.\nLine 197 to 200: Something is wrong with this sentence, please revise.\nRESPONSE: We apologize for this weird sentence. We now revised it to “Our results showed that\nin some of the tropical regions, the observed naturalized-to-native phylogenetic distance was larger\nthan expected. These results thus support Darwin's naturalization hypothesis, indicating that in\nthese tropical regions, the influence of biotic interactions may outweigh environmental filtering”\n(Lines 222-225).\nLines 440-453: The final data set giving alien and native species per region, compiled and cleaned\nas specified in the method section, is a highly valuable resource for future studies. Do you plan on\npublishing this as well?\nRESPONSE: Based on this comment and a similar comment of Reviewer 2, we decided to make\nthe entire dataset available (Lines 508-509). The dataset, encompassing the lists of alien and\nnative species for all 493 regions considered in our analysis, is now openly accessible through\nFigshare (Data.species.distrbution.493.Rdata, https://doi.org/10.6084/m9.figshare.20055611.v4).\nResponses to the Reviewer 2’s comments\nThe authors test Darwin’s naturalization conundrum at a global scale, and find a latitudinal gradient\nin how closely related naturalized angiosperm species are to native ones. They also examine how\nthis trend may be mediated by human landscape modification. The manuscript is clearly written,\nbut lacks some detail – this may be partially due to the fairly short format it was written in.\nRESPONSE: We sincerely appreciate the reviewer's acknowledgment that the manuscript is\nclearly written. In response to the reviewer's valuable feedback provided below, we have carefully\naddressed these specific comments by incorporating additional details into the revised version of\nthe manuscript.\nThe study largely confirms previous studies that have found that the relatedness between species\nis greater in harsher climates (which underlies the overall latitudinal gradient; e.g., Qian et al. 2013;\nLI et al. 2014; Escoriza & Ruhí 2014; Park et al. 2020; Kusumoto et al. 2021; Zu et al. 2023) and\nalbeit at a larger scale. Likewise, the effects of human landscape modification on phylogenetic\ncommunity patterns have been investigated, but at smaller scales and more heterogeneous results\n(e.g., Cui et al 2019; Yang et al 2022).\nRESPONSE: Thank you for bringing up these relevant studies on taxonomic distinctness (Escoriza\net al., 2014) or phylogenetic clustering/dispersion of native species (Zu et al., 2023), alien species\n(Kusumoto et al., 2021), or native and alien species jointly (Qian et al., 2013; Li et al., 2014), and\nwe now cite several of them (Lines 92-99). However, we would like to point out that these studies\nmainly focused on assessing phylogenetic community structures within communities themselves,\nnone of them specifically examined the relatedness of alien species to native floras, which is the\ncore aspect of Darwin's naturalization conundrum. One notable exception is Park et al. (2020),\nwhich directly analyzed alien-native phylogenetic distances within the US and how they relate to\nspatial scale. A novel finding of Park et al. (2020), as we mention in the main text (Lines 98-101,\n170-172), is that alien-native phylogenetic distances were smaller at larger spatial extents and in\nharsher climates. However, the study did not explicitly investigate the latitudinal gradient of\nDarwin's naturalization conundrum, possibly due to the limited latitudinal range covered by the US\nalone. Therefore, to the best of our knowledge, our study is the first to propose the existence of a\nlatitudinal gradient in Darwin's naturalization conundrum and provides empirical evidence for this\nphenomenon at both continental and global scales.\nAnother novel aspect of our study is the consideration of multiple null models (species pools) to\nassess whether the alien-native phylogenetic relatedness is smaller or larger than expected.\nBecause it is impossible to know all species that actually have been introduced into the focal region\nand to recognize that the definition and size of the potential source pool may affect the outcomes,\nwe used multiple species pools based on different assumptions of which species have most likely\nbeen introduced. By using these different species pools, we aim to enhance our understanding of\nthe role of phylogenetic relatedness in naturalization success, while accounting for potential\nphylogenetic biases introduced by specific introduction preferences.\nThe larger scale of this study is both its strength and weakness. It lends support to previous findings\nat larger scales and provides an interesting, if not surprising, global-scale view of how human\ndislocated species may integrate into regional flora. However, this comes at a cost – the underlying\ndata are not as detailed/accurate, the coarse spatial resolution cannot account for biotic\ninteractions among taxa, and thus the results do not advance our knowledge as to why we observe\nthese patterns.\nRESPONSE: We agree with the reviewer that the coarse spatial resolution has its limitations.\nHowever, we respectfully disagree with the assertion that the results therefore do not add new\nknowledge or important insights. Our study reveals a previously unreported pattern—the latitudinal\ngradient in the relatedness between alien and native species at the global scale. This finding, we\nbelieve, significantly contributes to our understanding of the distribution and integration of\nnaturalized species in regional floras.\nIn regard to discerning mechanisms, we point out that this is not only a challenge in large/global\nscale analyses such as ours, but in any observational/correlational study, also the ones that use a\nsmaller spatial resolution (Escoriza et al., 2014; Kusumoto et al., 2021; Park et al., 2020; Qian et\nal., 2013). However, as commended by Reviewer 1, we have clear mechanistically based\nhypotheses on the expected latitudinal and environmental patterns in alien-native phylogenetic\ndistance. While the nature of our data precludes us from definitively concluding that the discussed\nmechanisms drive the patterns, the fact that empirical data clearly support expectations at this\nscale is worth pointing out, and we believe that our results have the potential to prompt future\nstudies to explore these mechanisms using diverse and complementary data and approaches.\nWe sincerely acknowledge that at the spatial resolution of our study, the alien and closely related\nnative plant species do not necessarily interact directly. However, we recognize that even without\ndirect interactions, species can influence each other through various biotic interactions, such as\nshared pathogens, herbivores, pollinators and seed-dispersers. Furthermore, the occurrence or\nabsence of species at a regional scale is influenced by processes playing out at local scales. While\nwe cannot observe local biotic interactions directly in our data, we can detect signals of their\ninfluence at regional scales. In fact, our results align with the prediction of the latitudinal biotic\ninteraction hypothesis, which suggests that biotic interactions are more pronounced at lower\nlatitudes (Schemske et al. 2009). We therefore consider the potential increase in biotic interactions\nat low latitudes as a plausible explanation for the novel latitudinal gradient pattern we identified.\nThank you for the valuable feedback and we have discussed these points more clearly in the\nrevised manuscript (Lines 233-244).\nThere is a proliferation of large-scale correlative analyses based on data that others have collected\naimed at testing hypotheses on global scales. Though such studies have value, they often do not\nadd new knowledge to the field, especially when the gathered data are not made available in their\nentirety.\nRESPONSE: We would like to emphasize that our study adds new knowledge to the field by\nproposing and empirically demonstrating the presence of a latitudinal gradient in Darwin's\nnaturalization conundrum at a global scale. This novel finding offers a potential explanation for the\nmixed results obtained in previous regional-scale studies conducted at different latitudes.\nIn consideration of the comments of both reviewers, we have conferred with contributing authors\n(who have all spent significant time and effort in collecting, gathering, aligning and organizing the\nvarious underlying datasets) and decided to make the data openly available. The dataset includes\nthe lists of both alien and native species for all 493 regions analyzed in our study, and it can be\naccessed through Figshare. (Data.species.distrbution.493.Rdata,\nhttps://doi.org/10.6084/m9.figshare.20055611.v4).\nAlong these lines, the study is somewhat descriptive and does not introduce novel insights, new\nmethods, or new data. Again, the strength and novelty of this study are in its large-scale and the\nassemble global dataset of native and naturalized plants, but without making the underlying data\nfully accessible, it is less useful. Please make data openly available and not “by request”.\nRESPONSE: We respectfully disagree with the notion that our study lacks novel insights, new\nmethods or new data. While it is true that previous studies have reported on phylogenetic\ncommunity structures along latitude, our study goes beyond this by proposing and empirically\ntesting how the relatedness between alien and native species changes along latitudinal gradients.\nTo the best of our knowledge, this is the first study to investigate the latitudinal gradient of Darwin's\nnaturalization conundrum.\nFurthermore, we have introduced new methods in our study, particularly through the utilization of\nmultiple and innovative species pools (e.g., a species pool that includes economically used\nspecies). We believe that these novel approaches will greatly benefit future studies on Darwin's\nnaturalization conundrum, facilitating comparisons and selection of appropriate species pools for\nanalysis.\nMoreover, in response to the suggestions raised by the reviewers, we have made the decision to\nmake our complete dataset openly accessible. By openly sharing our global dataset on native and\nnaturalized plants, we aim to facilitate further research and encourage validation and exploration\nof related topics.\nThe examination of human landscape modification effects is a potentially interesting addition that\nhas not been investigated as often. However, I think there needs to be some tests to rule out the\npossibility that the observed patterns are due to lack or resolution and/or biases in the data. For\ninstance, highly developed nations with extensively modified landscapes are often the places\nwhere both native and non-native species are best documented. These nations also\nsimultaneously engage in large amounts of international trade and travel, in effect, increasing\npropagule pressure of exotic species and their chances of establishment (Dawson et al 2017).\nThese developed nations mostly reside in higher-latitude temperate regions. Similarly, it is possible\nthat higher-latitude regions with similar floras (e.g., North America and northeast Asia) engage in\nmore biotic exchange than they do with lower-latitude regions, as suggested by some of the\nauthors’ previous work (van Kleunen et al. 2015).\nRESPONSE: We agree that species are more likely to be best documented in highly developed\nregions. To address the potential bias introduced by the completeness of species inventories, we\nhave conducted two additional analyses. First, we reanalyzed the data using only regions\nestimated to have over 50% completeness of their naturalized species lists, based on\ncompleteness-estimate scores from the GloNAF database (van Kleunen et al. 2019). The majority\nof regions in our study (470 out of 493) met this criterion, ensuring a reasonable level of\ncompleteness. We reanalyzed the data using only these regions and found that the latitudinal cline\nin alien-native phylogenetic relatedness persisted. Second, for the 437 regions that overlapped\nwith the study by Dawson et al. (2017), we incorporated the average inventory completeness\npercentage as a weighting variable in our analyses. By giving more weight to regions with higher\ncompleteness, we aimed to address potential biases. This analysis also confirmed the presence\nof a latitudinal cline in alien-native phylogenetic relatedness (Response Letter Table 1).\nResponse Letter Table 1 | Results on the linear regression between MPD and latitude (1)\nwithout accounting for data completeness, (2) for the subset of regions for which the naturalized\nspecies lists were judged to have over 50% completeness (the completeness score in the\nGloNAF database; van Kleunen et al. 2019), and (3) for the subset of regions that overlapped\nwith Dawson et al. (2017) and for which the inventory completeness percentage was used as\nweighting variable.\nRegions'\nModels Estimate Std. Error t value Pr(>|t|) Adjusted R-squared\nnumber\n(1) 493 -0.117 0.009 -12.589 <0.0001 0.243\n(2) 470 -0.103 0.009 -11.22 <0.0001 0.210\n(3) 437 -0.109 0.011 -10.193 <0.0001 0.191\nIn van Kleunen et al. (2015), we have indeed shown that certain parts of the world, such as North\nAmerica and Temperate Asia, have exchanged more species with one another than with others.\nThis might partly reflect that some continents are climatically more similar, and we considered this\nby using a global as well as a continental pool of naturalized species that are climatically suitability\nto the focal region (mentioned in Lines 412-413). Moreover, in another paper (van Kleunen et al.\n(2020), we showed that many of the biotic exchange patterns among different parts of the world\ncan be explained by the species that we cultivate for their economic uses. We addressed this by\nalso having a pool of species with economic uses.\nSome of the methodological choices need more justification/explanation. For instance, why\nis >5000 km2 the threshold for analysis? Why was The Plant List used for taxonomic harmonization\nwhen it has been static since 2013 and replaced? Several of the 19 bioclimatic variables used to\nmodel species distributions are highly correlated and should not be included in the same models.\nSampling biases associated with the data sources and their potential effects on the results should\nbe discussed in more detail.\nRESPONSE: According to this thoughtful comment, we carefully revised the method section of our\nmanuscript to address the concerns raised.\n(1) Regarding the threshold of region size, we now clarify more elaborately that we excluded\nregions <5000 km2 to minimize the variation in region size, as smaller regions may be\nsubject to greater variation and potential sampling biases (Lines 291-294). Additionally, we\nconducted sensitivity analyses using different region size thresholds (0, 1000, 5000, and\n10,000 km2) to assess the robustness of our findings. We found that all thresholds\nconsistently showed a negative relationship between phylogenetic distance and latitude\n(Lines 295-298; Supplementary Table 3). In addition, we also tested whether the relationship\nbetween MPD and latitude varied by region sizes. For this, we divided the data into three\nsubsets based on the 33% and 67% quantiles of the region-size distribution. Analyses of\neach of the three resulting subsets shows that they all reveal negative MPD vs latitude\nrelationships (Response Letter Figure 1). These findings indicate the robustness of the alien-\nnative MPD-latitude relationship, regardless of the chosen region-size thresholds.\nResponse Letter Figure 1 | The relationship of MPD with latitude for separate analyses of the one\nthird of regions with the smallest areas (log10(Area) < 4.407), the one third of regions with\nintermediate areas (4.407 ≤ log10(Area) < 5.139) and the one third of regions with the largest areas\n(log10(Area) ≥ 5.139). Pearson correlation coefficient and p value (two sided) are shown.\n(2) Regarding the taxonomic harmonization, we clarified that The Plant List was used because\nboth the GIFT database (Weigelt et al., 2020) and the GloNAF database (van Kleunen et al.,\n2019) were harmonized using The Plant List as a reference (Lines 306-307).\n(3) Regarding the bioclimatic variables, the reviewer is right that some of the 19 variables are\nhighly correlated. Although we believe this should not necessarily be problematic for predictive\nmodels, we now ran the MaxEnt models using only six of the bioclimatic variables (including\nmean diurnal range, temperature seasonality, maximum temperature of warmest month,\nprecipitation seasonality, precipitation of wettest quarter, and precipitation of driest quarter),\nall with correlations <0.7 (details are provided in lines 416-423). We then recalculated the\nexpected MPDs based on the climatically suitable global naturalized alien species pool (climate\nnat.) and the climatically suitable continental naturalized alien species pool (climate continent\nnat.). Although our results show that the results are consistent when including all 19 variables\nor only the six variables with low collinearity (Response Letter Figure 2), we adopted the\nreviewer's suggestion and now included the results based on the six climatic variables in the\nmain text. We updated the results in the manuscript accordingly (Figs. 2d, 4a, and\nSupplementary Figs. 4, 6, 7, 8, 11, 12, 13).\nResponse Letter Figure 2 | Comparison of ΔMPD values calculated using all 19 bioclimatic\nvariables or a subset of six bioclimatic values with r < 0.7 for (a) the climatically suitable global\nnaturalized alien species pool and (b) the climatically suitable continental naturalized alien species\npool.\n(4) Regarding potential biases, we have expanded the discussion to provide a more thorough\nexploration of the limitations of the data (Lines 245-252). While we acknowledge these\nlimitations, we believe that the data sources we utilized are of high quality on a global scale,\nand that these potential limitations are unlikely to significantly impact our main conclusions.\nAccording to the reviewer's suggestion, we highlighted that incorporating well-resolved species\noccurrence data at regional and local scales, as well as species-level molecular phylogenies\nand trait data, would be valuable for future research on Darwin's naturalization conundrum\n(Lines 249-252, 258-261, 265-269). Unfortunately, these approaches are not currently feasible\nwith the available data. We have included these considerations and limitations in the revised\nmanuscript to provide a more comprehensive overview (Lines 233-269)\nAlso, while the authors should be commended for their testing of multiple null models, I am curious\nas to why native species were not included as candidates when shuffling taxa. At the end of the day\nDarwin’s naturalization conundrum is a hypothesis of community assembly and thus should consider\nthe insertion of any species that has the means to physically reach a community, regardless of its\nnative status. For example, the fact that native species X is not present in community/region A despite\noccurring nearby for millennia, while non-native species Y from the other side of the world has\ninserted itself over the last 50 years tells us something.\nRESPONSE: We acknowledge that we may not fully understand the reviewer's comment, but we\ncan see two possible interpretations:\n(1) If the reviewer suggests shuffling both native and alien species in each region, making all\nspecies being randomly selected from the larger species pool, we recognize that such an\napproach would be a conventional test of phylogenetic community structures and community\nassembly. This line of research, as acknowledged by some of the studies cited by this reviewer,\naddresses important questions. However, it is crucial to note that the hypotheses being\ninvestigated in this context are fundamentally different from our study on Darwin's\nnaturalization conundrum. As highlighted by Thuiller et al. (2010) in their influential review\npaper, the null model employed for testing Darwin's naturalization conundrum should\nspecifically break down the phylogenetic relationship between alien and native species, while\npreserving the phylogenetic relationships between native species within the recipient regions.\nIn our study, the question tested by the null model is why a particular alien species successfully\nnaturalized in a recipient region, rather than another alien species from the available pool.\nTherefore, the implementation of the randomization procedure in our study must not change\nthe evolutionary and ecological mechanisms that led to the current structure of native\ncommunities.\n(2) If the reviewer suggests including species native to the continent but not native to the\nfocal region itself in the potential species pool, we already recognize the point made. Such\nspecies are currently already included in the global non-native species pool of a region.\nHowever, to address this more explicitly, we have now created a species pool for each region\nthat encompasses all species native or naturalized to the continent but not native to the specific\nregion. For this null model, like for all our other null models, we still find a negative ∆MPD vs\nlatitude relationship (Response Letter Figure 3). Considering that we are unsure whether the\nreviewer intended this specific type of null model and since such species are already included\nin the global non-native species pool of a region, we did not include this new null model in the\nmanuscript. However, we are willing to incorporate it if the editor believes it is necessary.\nThanks for this comment and please feel free to correct us if we misunderstood any aspects of the\npoint.\nResponse Letter Figure 3 | Latitudinal gradient of ΔMPD based on the null model using a species\npool consisting of all species (native and alien) occurring in the focal region’s continent, but that\nare not native to the focal region.\nFigure legends and sizing could be better – for instance, it was very difficult to see the inset map\nin Figure 1. Also, figure captions could be clearer. Individual descriptions of each panel may help.\nHow were the colors binned? The bins do not seem to be equal in size, both in the main figures\nand the supplementary figures.\nRESPONSE: We thank the reviewer for the useful suggestions regarding the figures. Following\nthe recommendation, we have relocated the inset map from Fig. 1 to Supplementary Fig. 3, and\nwe now provide detailed descriptions for each panel in the figure captions (Lines 734-740, 758). In\nthe revised manuscript, we also clarified that we used bins of equal interval sizes, except for the\nlowest and highest values due to the relatively few observations in these larger bin intervals (Lines\n737-739).\nTable S2: Where do the IDs for each region come from?\nRESPONSE: We have clarified in the revised manuscript that the IDs for each region correspond\nto the IDs used in the open-access GloNAF database (see\nhttps://idata.idiv.de/DDM/Data/ShowData/257).\nLine 82: There’s a word missing. Think it should be “can be clarified”\nRESPONSE: corrected (Line 82).\nLine 92: Another word missing. Should be “would allow us to test”\nRESPONSE: Thank you for catching this. This sentence has been rephrased to “Therefore, a\nglobal-scale analysis of regional floras is required to determine whether a latitudinal gradient\nindeed exists in Darwin’s naturalization conundrum and whether it holds true across continents”\n(Lines 101-103).\nLine 106-107: The statement is slightly confusing/misleading. The study examines whether the\ndegree of human modification affects the relationship between latitude (and climate) and the\nrelatedness between native and naturalized species. It does not, however, examine if/how human\nmodification alters the biotic/abiotic environment. Also, I do not think the study actually tests\n“whether the degree of human modification of the landscape has changed the latitudinal gradient”\n– it only shows a significant interaction term.\nRESPONSE: We agree, and this sentence has been rephrased to “Furthermore, given the strong\nassociation between species naturalization and human-induced environmental disturbances, we\nadditionally tested whether the degree of human modification to the landscape influenced the\nrelationship of naturalized-to-native phylogenetic distance with latitude and climate.” (Lines 114-\n117).\nLine 166: “could also” seems strange here. Latitude is often seen as a proxy for climatic factors\nand does not generally have a direct effect on biodiversity.\nRESPONSE: We agree. This sentence has been removed from the main text.\nLine 259: How many taxa had conflicting native statuses? How were native vs non-native\nsubspecies treated? Do the results change if taxa with conflicting status were considered non-\nnative?\nRESPONSE: In the revised manuscript, we clarified that there were 1,278 taxa-by-region\ncombinations out of 1,177,267 that had conflicting native statuses (Lines 312-313). We also added\na figure to the supplements showing that alien-native phylogenetic distance estimates are not\naffected by whether the species with conflicting status are considered to be natives or aliens\n(Supplementary Fig. 9). Regarding native vs non-native subspecies (or other infraspecific taxa), if\nthey were standardized to different accepted binomial names, they were considered as separate\nspecies, and we considered the native vs non-native status of those species. If the infraspecific\ntaxa were standardized to the same binomial species name, they were treated as native species\nin our study. We now mention this explicitly in the Methods (lines 308-311).\nLine 285: Genus-level phylogenies may be sufficient in certain circumstances but such a blanket\nstatement is improper and can be dangerous. Further, the test that was conducted – making all\ngenera polytomies and redoing analyses – does not demonstrate that the lack of phylogenetic\nresolution does not affect the results. What it is showing is that the results from the tree used in\nthe study, which is not fully resolved nor fully supported by molecular data, are not largely different\nfrom a tree with worse resolution. It does not prove that the results from the used tree are congruent\nwith those that would be generated from a more robust, resolved phylogeny supported by\nmolecular data. Perhaps the authors could limit their analyses to regions for which they have full\nphylogenetic coverage and see if the results still stand?\nRESPONSE: We appreciate the reviewer's concerns regarding the phylogenetic resolution in our\nstudy. We agree that highly resolved species-level phylogenies are extremely important for\nquestions about trait evolution and estimating diversification rates (e.g., Rabosky, 2015). However,\nseveral studies (e.g., Cadotte et al., 2008; Cadotte et al., 2009; Cadotte, 2015; Li et al., 2019;\nQian & Jin, 2021) have previously evaluated the resolution of phylogenies and its ability to detect non-\nrandom community patterns. The consensus is that imperfect resolution below the family level has\na minimal effect on detection of regional patterns and statistical significance. This is because the\nassembled communities usually contain distantly related species across different families or\ngenera that traverse deep nodes. Thus, fine-tuning the relative distance among the small values\nbetween species in the same genus will have a minor impact on the calculation of MPD which\nalways includes distantly related species. There is no current evidence that more precise\nestimations of phylogenetic relationships at the genus level can alter community assembly\ninferences, but there are many analyses showing that results are very robust to changes in\ndistances at the genus level (see Li et al., 2019 and Qian & Jin, 2021 as examples).\nIn the revised manuscript, we now clarify that the reason why we also used a genus-level\nphylogeny with polytomies for all species is that some species in our mega tree species were\nadded as polytomies of their genera or family, while other species are fully resolved at the species\nlevel (Lines 332-336). This could introduce potential biases when examining latitudinal and climatic\ngradients in naturalized-alien-to-native phylogenetic distances. We are open to removing this\nanalysis if the reviewer believes it would improve the clarity and conciseness of the paper.\nIn response to the suggestion of the reviewer, we attempted to perform an analysis using regions\nwith complete molecular phylogeny coverage, but unfortunately we could not find such regions. As\nwe proposed above, using a finer phylogeny might have a minor effect on the calculation of the\nMPD for a certain region, but for studies across regions, we believe that the overall global patterns\nwe observed would not be significantly altered by improved phylogenetic resolution. However, we\nagree with the reviewer that constructing better-resolved phylogenies across a broad latitudinal\ngradient is an important future research direction. We have highlighted this point in the discussion\nsection of the revised manuscript (Lines 252-261).\nLine 332: The rationale for limiting a species pool to species with known economic uses is not clear,\nespecially when the focal taxon itself does not have a known use, and needs to be better fleshed\nout/justified. Why not just limit this pool to species that have been introduced somewhere?\nRESPONSE: Ideally, we would indeed use a pool of species that have been introduced to a region\n(irrespective of whether the species have become naturalized there). As such data unfortunately\ndo not exist, we used different species pools that make different assumptions about which species\nmight have been introduced to a region. For example, the global non-native species pool assumes\nthat all species have been introduced everywhere, whereas the global naturalized species pool\nassumes that all species that have already become naturalized somewhere have been introduced\neverywhere. We also recognize the importance of economic plants, which have a higher likelihood\nof being introduced to other regions for cultivation by humans and often become naturalized. To\ncapture this aspect, we created a species pool that combines all species with known economic\nuses with all species that have become naturalized somewhere. We believe this is a very important\nnull model, as was also pointed out by Reviewer 1, and we therefore would like to keep it in the\nmanuscript. We have now provided a more detailed explanation of the rationale behind this null\nmodel in the manuscript (Lines 388-392).\nLine 365: The modeling of species distributions with such coarse data are troubling. 250 km2 can\ncomprise extreme amounts of environmental heterogeneity – e.g., coastal invaders can become\ninland invaders of the desert or alpine invaders. Perhaps some comparisons between these crude\nmodels and those fit to actual point occurrence data could be compared for taxa with enough data\nto support this approach – many (if not most) invasive species have large amounts of occurrence\ndata available. Also, all model parameters should be provided to ensure reproducibility.\nRESPONSE: We fully agree that these species distribution models (SDM) used in our study are\nvery crude. While it is true that some widely naturalized (i.e., invasive) species have extensive\noccurrence records, this is not the case for all naturalized species at a global scale. Limiting the\nanalysis to well-recorded species could introduce biases when studying the latitudinal gradient on\na global scale. The purpose of our SDMs is not to precisely predict suitable habitats for naturalized\nspecies but to calculate their mean MPD to native species in regions with potentially suitable\nclimates. Therefore, the key question is whether the mean MPD calculated using the coarse data\nwe employed is biased compared to using well-resolved occurrence data.\nTo address this concern, we made a comparison between mean MPD calculated using our ‘rough’\nSDMs and SDMs based on point-occurrence data for a subset of species. We randomly selected\n300 naturalized species from the global naturalized alien flora, and then used MaxEnt to predict\ntheir global potential distribution based on point occurrence data from the Global Biodiversity\nInformation Facility (GBIF; https://www.gbif.org/).\nFirstly, we used the R package rgbif (Chamberlain & Boettiger, 2017; Chamberlain et al., 2023) to\ndownload the point occurrences of these species from GBIF. We kept the occurrences with\ngeographical coordinates. Then, we used the R package CoordinateCleaner (Zizka et al., 2019) to\nclean the geographic coordinates by excluding occurrences that are most likely incorrect. These\ninclude coordinates in the ocean (not on islands), zero coordinates, coordinate-country\nmismatches, coordinates assigned to centroids of countries and provinces, coordinates within\ncities, outlier coordinates, coordinates assigned to biodiversity institutions, and coordinates of\nfossil specimens. Then, we used the same six bioclimatic variables as used for the other analyses\n(including mean diurnal range, temperature seasonality, maximum temperature of warmest month,\nprecipitation seasonality, precipitation of wettest quarter, and precipitation of driest quarter, Lines\n416-423) from WorldClim at 2.5 arc-min resolution. We removed occurrences with a coordinate\nuncertainty greater than 4.5 km (~2.5 arc-min), and removed pseudo-duplicate records, so that\nthere is at most one record per 2.5 arc-min grid cell. Similar to our analysis described in the main\ntext, we first excluded species with fewer than 15 occurrences, and we then ran the MaxEnt model\n(using the R package dismo, Hijmans et al., 2021) separately for each naturalized species. Here,\nwe selected 20% of the data for testing the model, and kept only those models with AUC values\ngreater than 0.7. We then used the maximum training sensitivity plus specificity threshold to test\nwhether a grid cell is suitable for a species. Finally, we obtained the global potential distribution of\n272 of the 300 selected naturalized species.\nWe then compared the mean MPD values based on the SDMs using the coarse distribution data\nand those using point-occurrence data for the 272 species. The analysis revealed a strong\ncorrelation between these MPD values (Pearson’s r = 0.99, Response Letter Figure 4), indicating\nthat our analyses based on the coarse data, available for all naturalized species, provide a reliable\nestimation of MPD.\nFurthermore, we have included all the necessary information on model parameters in the revised\nmanuscript to ensure reproducibility (Lines 413-441). We appreciate the valuable input of the\nreviewer and thank them for bringing these points to our attention.\nResponse Letter Figure 4 | Comparison of phylogenetic distances calculated using two different\ndatasets. For each naturalized species, phylogenetic distances were calculated as the mean value\nof the mean pairwise phylogenetic distances (MPD) of the non-native species to the native species\nin the regions with a suitable climate. These results are based on a subset of 272 species for which\nwe found 15 or more point-occurrences in GBIF (out of an initial subset of 300 species). The value\nalong the x-axis is calculated based on the coarse distribution used in our main text, and the value\nalong the y-axis is calculated based on the point-occurrence data from GBIF."}, {"round": 2, "reviewer_comments": {"reviewer_1": "With this revision, all my comments have been addressed to my complete satisfaction.\nFrom my perspective it also seems that the comments provided by the other reviewer have been\naddressed very thoroughly, with new analyses and major changes in the main text. In cases of\ndisagreement, the authors provided much information and well-founded argument why this is the\ncase.\nI am looking forward to seeing this manuscript published.", "reviewer_2": "I appreciate the authors’ efforts to address reviewer comments. I believe the manuscript is stronger\nwith the added methodological details and discussion of limitations. I also commend the authors'\ndecision to share most of the data underlying their analyses. It is unfortunate that some of the\nsources the authors assemble these data from, however, are not fully open. Though I do not agree\nwith all the methodological choices made here others have employed similar approaches at times. The\n‘regions’ that form the unit of analysis (which are not defined, I think – how were these regions\ndelimited?) here vary greatly in their size and environmental heterogeneity. While I understand this is\nlikely because species checklists are usually only available for countries, this is not ideal and may bias\nthe results. I recommend that the authors include analyses, figures, and tables presented in the\nresponse document as supplements and refer to them in the main text to facilitate understanding of\nhow and why certain choices were made. It may also be good to acknowledge that results could differ\nwith better data/different methods along with this information (e.g., better phylogenetic coverage,\ninclusion of more data from species-rich regions in southeast Asia and Africa, more specific range\ndata) – I do not think such acknowledgments detract from the findings of the study."}, "author_response": "Responses to the Reviewer 1’s comments\nWith this revision, all my comments have been addressed to my complete satisfaction.\nFrom my perspective it also seems that the comments provided by the other reviewer\nhave been addressed very thoroughly, with new analyses and major changes in the main\ntext. In cases of disagreement, the authors provided much information and well-founded\nargument why this is the case.\nI am looking forward to seeing this manuscript published.\nRESPONSE: We express our gratitude to the reviewer for this very positive evaluation\nand valuable feedback throughout the revisions.\nResponses to the Reviewer 2’s comments\nI appreciate the authors’ efforts to address reviewer comments. I believe the manuscript is\nstronger with the added methodological details and discussion of limitations. I also\ncommend the authors' decision to share most of the data underlying their analyses. It is\nunfortunate that some of the sources the authors assemble these data from, however, are\nnot fully open.\nRESPONSE: We are grateful for the reviewer's positive assessment of the manuscript\nand the recognition of the enhancements made through our revisions. Your valuable\nfeedback has undoubtedly played a pivotal role in enhancing the quality of our work.\nWhile a small portion of GIFT database are not publicly available due to data sharing\nrestrictions, we would like to emphasize that all the original data utilized in our study,\nencompassing the comprehensive lists of 219,520 native and 9,531 naturalized alien\nplant species spanning 487 regions, are openly accessible via Figshare\n(https://doi.org/10.6084/m9.figshare.20055611.v5).\nThough I do not agree with all the methodological choices made here others have\nemployed similar approaches at times. The ‘regions’ that form the unit of analysis (which\nare not defined, I think – how were these regions delimited?) here vary greatly in their size\nand environmental heterogeneity. While I understand this is likely because species\nchecklists are usually only available for countries, this is not ideal and may bias the results.\nRESPONSE: In accordance with the reviewer’s insightful input, we have made the\nnecessary clarification that the regions examined in our study pertain to administrative units,\ncomprising countries, states, provinces, and islands (Line 288). We agree with the reviewer\nthat the variation in size and environmental diversity among these regions could introduce\na potential bias to the results. To address this concern, we conducted supplementary\nanalyses by incorporating region area as a covariate in our statistical model. These\nsupplementary analyses unequivocally demonstrated that the variation in region size had\na modest impact on our results and did not compromise the integrity of our primary findings\n(Supplementary Fig. 8) (Lines 304-306).\nI recommend that the authors include analyses, figures, and tables presented in the\nresponse document as supplements and refer to them in the main text to facilitate\nunderstanding of how and why certain choices were made. It may also be good to\nacknowledge that results could differ with better data/different methods along with this\ninformation (e.g., better phylogenetic coverage, inclusion of more data from species-rich\nregions in southeast Asia and Africa, more specific range data) – I do not think such\nacknowledgments detract from the findings of the study.\nRESPONSE: We are glad that the reviewer found these additional analyses helpful. To\naddress this recommendation, we have referenced the completeness analysis in the main\ntext (Lines 514-526) and providing the results in Supplementary Table 4. To ensure the\nsupplementary materials remain focused and easily navigable for readers, we have\nretained Response Letter Figures 1-4 in the response document. As these figures primarily\naddress specific details discussed with the reviewer and might not be of general\nsignificance to all readers, we believe this approach maintains clarity without overwhelming\nthe supplementary materials. These response materials are also fully accessible to the\nreaders, thus ensuring transparency. Furthermore, we have taken your advice and\nexpanded our discussion to acknowledge the potential implications of improved data\nquality on our findings (Lines 253-255, 266-267)."}]

Decision letter