Phytolacca tetramera, an Ecological Anachronism from the Pleistocene Surviving in the Pampean Grasslands

Abstract

The Dwarf Ombú, Phytolacca tetramera, is a rare and highly unusual plant endemic to the northeastern Pampean grasslands of Argentina and is currently considered of high conservation priority. In order to better understand its biology, ecology, and conservation requirements, we studied its anatomy, reproductive traits, life history, and distribution based on field observations and herbarium material. Our results show that P. tetramera possesses a combination of traits consistent with the concept of ecological anachronism. The species produces large fleshy fruits whose size and shape are comparable to those interpreted as adapted for dispersal by extinct megafauna. In addition, the plant exhibits morphological and ecological adaptations associated with intense grazing, trampling, and drought tolerance, including robust underground structures and a growth pattern comparable to underground trees from seasonally dry open habitats. These findings suggest that P. tetramera evolved under ecological conditions markedly different from those existing today, including megafaunal disclimax environments that disappeared after the late Pleistocene extinctions. This ecological mismatch may help to explain its present rarity, fragmented distribution, and low population numbers. Our results also indicate that current conservation strategies for P. tetramera should consider the role of disturbance regimes and extinct ecological interactions in shaping the biology of this species.

1. Introduction

The recognition of features developed under past ecological pressures that no longer exist in extant plants (known as anachronic traits) has become a hotly debated topic in botanical and ecological studies since Janzen and Martin’s seminal 1982 paper [1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17]. Janzen and Martin [18] defined seed dispersal anachronisms as those syndromes in which fruit traits are better explained by interactions with extinct animals, especially with big herbivores (weighing more than 100 kg) and megafauna (exceeding 1000 kg) in places where this fauna became extinct near the Pleistocene–Holocene boundary. Anachronic plants are often mechanically protected by exceedingly large epidermal or branch excrescences (i.e., spines, thorns, and thick cork) and bear fruits that are unattractive to extant native vertebrates, and due to that, seed crops rot on the ground beneath fruiting plants [3,18,19]. Beyond their anomalous fruits, these plants also exhibit atypical life-history and ecological attributes, and thus cannot be fully understood without considering the relatively recent extinction of their primary dispersers and the dramatic climatic and ecological changes that occurred in the last thousands of years [20].

When identifying anachronic plants, population dynamics and environmental characteristics should also be considered along with the characteristics of the plant itself. Populations of anachronic plants are usually homogeneous and scattered, and a high number of these plants are facing some degree of extinction risk as a consequence of the loss of their primary dispersers [7]. For this reason, the identification and study of anachronic species and their environmental conditions is an urgent need for the planning of conservation actions. The main problem is that the data about current interactors with plants and their feeding behavior remain incomplete in many cases [21]. Zaya and Howe [22] stated that the cradle of evolution of the anachronic plants was highly disturbed habitats in a permanent state of successional flux due to the highly destructive herds of megamammals, which they called “megafaunal disclimax”. Natural habitats of these plant species today are frequently disturbed, like alluvial bottoms on gentle slopes, forest–grassland transitions, roadsides, or crops, where the perturbations create similar conditions to the megafaunal disclimax [22,23].

The objective of this work is to discuss whether a rare and endangered plant of the Pampean grassland may be recognized as an ecological anachronism. The selected species, Phytolacca tetramera Hauman, is a dioecious rhizomatous geophyte with one-meter-tall stems, pulpy yellow berries [24], and many toxic compounds [25,26], that is very scarce in the field [27]. Morphologically, it resembles the gigantic “ombúes” P. dioica L. and P. weberbauerii H. Walter, for which it is locally known as “ombusillo”, or dwarf ombú, for its smaller size (Figure 1). It is similar in size and habitat to Phytolacca americana, the North American Pokeweed, which was previously reported as a part of the diet of the extinct Mammut americanum [28]. With its big, pulpy and smelly infructescence, P. tetramera is one of the most conspicuous plant species in the south of the Río de la Plata coast. However, important data, including the feminine reproductive structure of Phytolacca tetramera, remain insufficiently described in previous works [24,29,30,31]. Previous authors do not state the color, smell, size, and mass of the infructescences, nor how they are arranged in the plant, and without this essential data, the megafaunal dispersal syndrome cannot be confirmed. The origin of the common name of P. tetramera (Dwarf Ombú in English/Ombusillo in Spanish) is the point of the iceberg of a series of remarkable features related to its phylogenetic placement and the possible environmental adaptations of the clade.

P. tetramera is an endemic plant with a small and patchy distribution that is restricted to about 100 km² at the northeastern coast of Buenos Aires province, Argentina [32] (Figure 2). Some evidence, like the existence of an old herbarium specimen collected 50 km north of its known populations, and the vast zones without specimens between the subpopulations, indicates that the past area of distribution and the low number of individuals of P. tetramera are the result of a historical decline. It was categorized as EN Endangered following the UICN criteria because it grows within a menaced productive territory dedicated to cattle raise, agriculture, and urban settlements, and considering its restricted distribution and the low number of individuals [32].

The published information is congruent with an alternative hypothesis to the anthropic-caused threat of P. tetramera. This species shows traits that may have been developed in a dry environment with high grazing pressure, and fruits attractive to large mammals, but, contrarily, it nowadays grows in a humid region where drought and fires are extremely infrequent and lacks native big herbivores. Hence, this paper has two main goals. In the first place, we provide new data to evaluate whether P. tetramera has a fruit megafaunal dispersal syndrome. In the second place, we explore if the kind of growth, disturbance resistance, and habitat preferences of this species could be adaptations to Pleistocene environmental forces, which may be the main driver of the evolution of this plant. If P. tetramera is an anachronic species, this will impact the conservation plans of this flagship species of Buenos Aires province. The survival of this endangered species depends on the conservation practices that will be taken forward: the search for pristine temperate–humid grasslands where the species may survive untouched or the ex situ reproduction to reinforce the current populations with new individuals, even when they live in menaced or disturbed places.

Accordingly, this study addresses three main questions: (1) Does Phytolacca tetramera exhibit reproductive traits consistent with a megafaunal dispersal syndrome? (2) Do its vegetative and ecological traits suggest adaptation to disturbance regimes compatible with megafaunal disclimax environments? (3) Can these putative anachronic traits help explain the current rarity and conservation status of the species?

2. Materials and Methods

The present contribution followed a comparative observational approach, focusing on morphological and ecological traits present in Phytolacca tetramera, which were previously proposed in the literature as being indicative of megafaunal dispersal syndromes and megafaunal disclimax adaptations.

Morphological assessment. We assessed the morphological characteristics of Phytolacca tetramera based on both living and collected specimens, as well as available bibliography [24,25,27,29,31]. Dried specimens from the herbaria of the Museo de La Plata (LP, 38 specimens), Facultad de Ciencias Agrarias y Forestales de la Universidad Nacional de La Plata (LPAG, 8 specimens), and Museo Argentino de Ciencias Naturales (BA, 5 specimens) [33] were examined.

Field surveys included visits to all known occurrence sites of P. tetramera, where all the 74 living individuals were observed (see details in [32]). Additionally, specimens were cultivated under controlled conditions at the nursery of the Facultad de Ciencias Naturales y Museo, Universidad Nacional de La Plata. In the discussion, characters were analyzed using a phylogenetic bracketing [34], for which we also examined living specimens of the closely related species Phytolacca dioica from the Paseo del Bosque de La Plata Park (Buenos Aires, Argentina) and P. weberbaueri from the Natural History Museum of the National University of San Marcos, Botanical Garden (Lima, Perú). We also consulted herbarium specimens of P. dioica from LP (24 specimens) and P. weberbaueri from USM (Universidad Nacional Mayor de San Marcos, Perú, 26 specimens).

Given that several aspects of the growth habit and reproductive structures of P. tetramera remain poorly documented, we recorded detailed information on plant size, underground stems, and reproductive structures based on both field observations and herbarium material.

Reproductive structures. Clusters of infructescences located at the apex of epigeal stems were treated as single reproductive units. We measured the weight and length of individual infructescences and the total fruit mass of these clusters. A total of 82 infructescences and infructescence clusters from 14 shoots belonging to four individuals from the La Plata population were measured and their fresh mass was recorded (reference specimens in LP: Guerrero E.L. Nº 714, Villa Garibaldi, Road to I. Correas, 8 May 2018; Guerrero E.L. Nº 788, Villa Garibaldi, Road to I. Correas, 19 December 2019; Guerrero E.L. Nº 824, Villa Garibaldi, 661 and 22 corner, 16 January 2021). Infructescence length was measured from the first basal fruit to the apex, without the stalk, and then was weighed. Subsequently, the infructescence groups associated with each stem were weighed to obtain the total fruit mass of each cluster. These measurements were used to characterize the main features of the reproductive units.

Vegetative growth and plant size. To evaluate plant size, defined as the horizontal area occupied by each individual, we mapped the extreme points of each specimen using Google Earth and constructed polygons to calculate surface area [32].

Individuals were considered discrete only when their underground stems and aligned shoots could be clearly traced. Subterranean structures were excavated at two localities within La Plata city to document rhizome-like stems and growth patterns.

Habitat characterization. To assess habitat preferences, all known occurrence sites of P. tetramera were surveyed [32]. Two environmental variables were recorded: 1—vegetation physiognomy (grassland or forest); 2—disturbance regime (disturbed or non-disturbed). Non-disturbed sites were defined as native environments with no severe or recurrent disturbances [32]. Disturbed sites included anthropically modified environments, heavily grazed areas, weed-invaded fields, and roadsides. Habitat preferences were expressed as the proportion of individuals occurring under each condition.

Comparative framework. For comparative purposes, we examined living specimens of closely related species, including Phytolacca dioica (Paseo del Bosque, La Plata, Argentina) and P. weberbaueri (Botanical Garden, Universidad Nacional Mayor de San Marcos, Lima, Perú). Herbarium specimens of these taxa from LP and USM were also consulted.

Experimental considerations. No feeding experiments were conducted in the present study because the toxic compounds present in Phytolacca species could interfere with experimental trials involving vertebrate consumers.

3. Results

3.1. Reproductive Units and Growth of Phytolacca tetramera

Fruits of Phytolacca tetramera are greenish-yellow, fleshy, and smelly berries of 1 cm with 6–10 seeds. During ripening, smell varies from a less intense, almost imperceptible, fragrance (like the flower scent) to a highly intense sweet odor. They are densely aggregated (up to 55 berries) in axillary semi-erect infructescences, except for the upper one, which is erect and terminal (Figure 3a–d), of 1.8–3 cm wide (mean 2.36 cm), 2–12 cm long (mean 7.26 cm), and 3–28 g weight (mean 10.28 g). Normal ramets (of more than 30 cm high) have three to eight infructescences closely grouped in the upper part of the aerial stem, and sometimes up to 67 infructescences divided into several secondary stems in bigger plants. These groups of infructescences placed together work as single reproductive units. The whole groups in the studied plants reached 10–15 cm wide (mean 11.25), 13.5–22 cm long, and weighed an average of 60.21 g (between 46 and 74 g). Nevertheless, smaller ramets sometimes have only one infructescence of up to 28 g.

We found that at the end of the season, the unconsumed infructescences still hang from the parental plants (Figure 3d). As unconsumed fruits increase in mass, the plants bend down, and after the first winter freeze, the plants with their rot fruits decay and dry up on the ground beneath the plants (Figure 3e) or are attacked by fungi, snails, and insects (we registered Bulimulus bonariensis, Acromyrmex lundi and an indeterminate moth larvae; Figure 3f). We have never found renewals of P. tetramera in the field. The seeds of this species are outlined by a fine tissue (endocarp) that should be damaged or removed to promote germination. If the fruits are not mechanically treated (or consumed), the endocarp avoids seed activation (pers. obs.).

3.2. Size, Habit, and Survival to Disturbances

P. tetramera has an enormous subterranean system in relation to the modest aerial development. Rhizomes of P. tetramera are perennial, plagiotropic underground stems with successive cambia that develop many active rings of vascular tissue (Figure 4a–c; see [35]) and an elevated water storage capability (Figure 4d). These rizome-like subterranean branches are comparable to the structural trunks of the tree species of the subgenus Pircuniopsis. Thanks to these rhizomes, P. tetramera plants develop their fleshy aerial shoots during summer, when other plants are under water stress. Rhizomes grow almost superficially to more than 1 m in depth. Their diameter varied between 2 and 25 cm (Figure 4a–c) (Supplementary Information Table S2). The biggest P. tetramera that we measured is a masculine specimen of approximately 35 × 23 m maximum length in La Plata covering 608 m² (Supplementary Information Table S2), whose rhizomes are visibly crossing the street, peeking out in the middle of the street, on both sides of the road, and into the surrounding house yards.

3.3. Habitat Preferences

Most Phytolacca tetramera specimens (77.46%) live in altered and periodically disturbed sites like roadsides and weed-invaded fields and are less common in well-preserved grassland or forest sites (22.53%) (Supplementary Information, Table S3). They are tolerant of heavy disturbances. P. tetramera specimens do not die after municipality road-workers cut off the road vegetation, pruning them to the base, when motorized vehicles run over them, or when gardeners cut off the plants from their parks, etc. In Pipinas locality (Punta Indio County), horses grazed upon P. tetramera leaves and aerial stems down to the base, but after a few days, the plants resprout.

The rhizomes resprout vigorously after the disturbances (Figure 4c), for which the plants seem to be immortal. For example, in the classic collection site of P. tetramera “Road to Arditi and Route 11 crossroad”, visited each year by many students from La Plata University during botanical field trips since the 1990s, plants still regrow despite being severely pruned several times every summer by province road-workers.

No attempts to know the age of any specimen have been conducted, but we confirmed that P. tetramera specimens can live for more than a century, because the specimen planted by Karl Wolffhuegel in 1906 from seeds that Lucièn Hauman collected still lives in the Facultad de Agronomía (Buenos Aires National University), where it was transplanted before 1913 [25].

P. tetramera is a heliophyte because 80.28% of the studied sites are grasslands under direct sunlight [32]. Also, some differences were observed between specimens from exposed and shaded sites: specimens under direct sunshine are densely covered by thick spatulate leaves that lack petioles; plants from shaded sites have slender stems with long internodes, petioles present, and ovate leaves, and they produce few inflorescences later than plants that grow under direct sunlight. This kind of shade intolerance is a common feature in plants from periodically disturbed environments.

In sum, we found that Phytolacca tetramera has consistent attributes with megafaunal dispersal syndrome, resistance to megafaunal stomp and herbivory, and seasonally dry climate adaptations (

Table 1. List of traits characteristic of anachronic plants. 1, present in Phytolacca tetramera; -, absent in Phytolacca tetramera; *, data obtained in this study, otherwise obtained from the literature.

Megafaunal dispersal syndrome:
Having a large and indehiscent structure	1 *
Inflorescences erect, visible upon the grassland	1 *
Up to 50–1000 g total fruit mass	1 *
Sugar, oil, or nitrogen-rich pulp	1
Similarity to Old World fruits dispersed by megafauna	1
Color attractive to megafauna	1 *
Fruits do not attract native vertebrates	1 *
Undispersed seed crops that rot on the ground beneath fruiting plants	1 *
Fruits attract exotic vertebrates (surrogates)	-
Nuts/seeds are protected mechanically	1 *
Low seedling mortality near the parent tree to persist	-
Other characteristics associated with the megafaunal dispersal syndrome:
Dioic reproductive system	1
Long life cycle	1 *
Current restricted distribution	1
Low population number	1
Traits related to megafaunal disclimax:
Stomp and pruning resistance. Compensatory growth.	1 *
Currently grows in altered sites, like roadsides	1 *
Chemical defenses in leaves and branches.	1
Shade intolerance	1 *
Traits expected for a seasonally dry and cooler climate:
Seasonal life cycle, underground resistance organs	1
Succulence	1
Phylogenetic relationships with other seasonal-climate-adapted taxa	1 *

).

4. Discussion

Phytolacca tetramera and other members of the subgenus Picuniopsis. Phytolacca tetramera’s nearest relatives are the gigantic “ombúes” (singular: ombú) P. dioica L. and P. weberbauerii H. Walter (Figure 1). These three species constitute a robust clade called Phytolacca subgen. Pircuniopsis [P. weberbauerii [P. dioica, P. tetramera]] defined both by morphology and molecular data [30,36,37]. P. weberbaueri and P. dioica are tall trees (up to 20 m), with a swollen caudex and palisade adventitious roots from which many trunks develop [36,38]. For example, a single specimen of P. dioica from Buenos Aires has a circumference of more than twenty meters [39]. In contrast, P. tetramera is a 0.3–1.7 m tall plant, with its major biomass located underground [31]. The obvious size differences led to the popular name “Ombusillo” (diminutive of Ombú), or Dwarf Ombú, for their little brother P. tetramera. Fruit structures of P. tetramera are like those of the ombúes, but the main difference is that the infructescences are erect [29].

In forested areas, the pendant yellow smelly infructescences of the Ombúes are consumed by monkeys [40,41], which disperse the seeds [42] and enhance their germination capability [43]. Other mammals, like the spectacled bear (Tremarctos ornatus) in Ecuador, also consume Ombú fruits [44], and peccaries in Argentina occasionally eat fallen fruits [45]. Some birds also consume P. dioica fruits [46,47,48], but most species systematically avoid them [49], leaving an enormous number of unconsumed fruits near the parental trees (Hauman 1913 [25]), which suggests that they may not be the direct dispersers of this plant, but a secondary one.

4.1. Phytolacca tetramera Anachronic Fruits

If Phytolacca tetramera is a megafauna-dispersed species that survived until our times, we should expect to find classic megafaunal dispersal syndrome traits, but also other traits related to the presence of the megafauna and specialized grazers (i.e., characters avoiding big grazers’ herbivory, compensatory mechanisms to resprout after grazing, or stomp resistance). The following list summarizes the traits that make an ecological anachronism, based on previous studies [1,3,6,18,19,22,23,50,51,52,53,54]:

Megafaunal dispersal syndrome: having a large and indehiscent structure; fruit structures erect, visible upon the grassland; up to 50–1000 g total fruit mass; sugar, oil, or nitrogen-rich pulp; similarity to Old World fruits dispersed by megafauna; color attractive to megafauna; fruits do not attract native vertebrates; undispersed seed crops that rot on the ground beneath fruiting trees; fruits attract exotic vertebrates (surrogates); nuts/seeds are protected mechanically; and low seedling mortality near the parent tree to persist.

Other characteristics associated with the megafaunal dispersal syndrome: dioic reproductive system; long life cycle; current restricted distribution; and low population number.

Traits related to the megafaunal disclimax: stomp and pruning resistance; compensatory growth; chemical defenses in leaves and branches; and shade intolerance.

Traits expected for a seasonally dry and/or cooler climate: Seasonal life cycle with underground resistance organs; succulence; and phylogenetic relationships with seasonal-climate-adapted plants. In addition, as the climate shifted drastically during the initial stages of the Holocene (see above), we should expect to find adaptations in P. tetramera to the Pleistocene drier climate.

All the members of the genus Phytolacca, except for three species, have green, reddish, brown, black, or purple fruits, smelly or not, and loosely arranged (with short petioles) in the infructescences [30]. The abovementioned colors are appropriate to attract birds [55], which are their main dispersers [56,57,58,59,60,61], and the separate fruits can be easily individualized and eaten by birds with a small gape-width [62]. The three exceptions are the species of Phytolacca subgen. Pircuniopsis (P. tetramera, P. dioica and P. weberbauerii), which have yellow fruits in dense infructescences with a strong odor that increases during the ripening process. These fruit characters, that passed unadvertised by taxonomists, are compatible with mammal dispersal syndrome [3,63].

The fruit dispersal syndrome of P. tetramera is one step beyond. Differently from the closely related ombúes, P. tetramera infructescences do not hang solitarily from lateral branches; on the contrary, they concentrate in the upper part of the plant (Figure 3a). Fruit structures are evident in the upper layer of the grassland, as it is expected in a plant that “offers” its fruits to a large-sized grassland frugivore [1]. And despite the size of the individual infructescences being like that of the ombúes, the size of the whole aggregates of infructescences of P. tetramera fit well with Guimaraes et al. [3]’s “Type II” of megafaunal fruits (fleshy fruits of more than 10 cm diameter with more than 100 small seeds) and exhibits the appropriate mass to be a megafaunal fruit candidate.

Although it has pulp-rich and smelly yellow visible fruits, nobody has registered any native or introduced vertebrates that feed upon its fruits and disperse their seeds. After twelve years of field study on this plant, we have never seen any signal of feeding behavior of vertebrates on the fruits of this species (i.e., lack of some infructescence, grabbing marks, or biting marks). Until now, we have not observed fruit consumption by introduced animals, so it appears that there are no surrogates for this plant. Which animals eat the fruits of P. tetramera and disperse the seeds remains an unsolved question. The scarcity of the plants [32] and the fact that we have seen only a few seedlings in ten years of field study also suggest that it may lack an effective disperser.

A study released by Di Sallo et al. [64] aimed to determine if birds (which constitute the main frugivores in the area of distribution of the plant) consume the fruits of P. tetramera. They found that neither birds nor any other diurnal vertebrate visits P. tetramera, and that all the fruits rot while still hanging from the plants. The current mammalian fauna of Buenos Aires is composed of medium-sized species with wide areas of distribution (i.e., Didelphis, Lutreolina, Chaetophractus, Dasypus, Lagostomus, Lyncodon, and Lycalopex), with Ozotoceros bezoarticus being the only mid- to big-sized mammal [65,66]. Based on the scarce bibliography, none of them seems to feed upon Phytolacca fruits. There is a team working with camera traps in the field to search for possible natural dispersers or exotic surrogates that feed on Phytolacca tetramera (A. Abba and N. Pagnutti com pers.).

4.2. An Underground Tree in the Pampean Grassland

Both ombúes, Phytolacca dioica and P. weberbauerii, are adapted to a warm and seasonal-rainfall climate. They survive droughts in seasonal environments thanks to a combination of vegetative morphological structures like the crass twigs and petioles, the thick succulent trunks emerging from a swollen caudex with adventitious palisade roots, and enormous deep primary roots (see Guaglione 1987 [38]). Their trunks can resprout after severe disturbances like fires, pruning or when the wind tumbles them down, but do not tolerate floods (obs. pers.). The xeromorphic traits of the ombúes allow them to inhabit the Seasonally Dry Forests of the Pacific Coastal of Peru–Ecuador and the Misiones-Low Paraguay and Paraná nuclei, occupying dry zones, with no severe frosts, although they reach wetter and cooler places marginally [67,68,69]. In fact, P. dioica is a plant recommended for gardens in arid regions for its drought-resistance [70,71]. The anomalous growth with numerous cambium rings of P. dioica, also present in P. weberbauerii and P. tetramera [35], is considered an anatomical key feature for surviving in harsh seasonally dry environments [72,73]. P. tetramera is derived from a seasonal-climate-adapted group of species and has the appearance of an underground ombú, which may be interpreted as an additional adaptation to survive drought seasons. Like other geophytes [74], the position of the trunks of P. tetramera below the soil surface can also be an adaptation of the plant to survive fires in a seasonally dry climate. Instead, it lives in a place without rainfall seasonality, where hydric deficiency is neither severe nor frequent.

With underground trunks and only the reproductive shoots over the surface in the flowering season, P. tetramera looks like the famous underground trees, or geoxylic suffrutices of subtropical and tropical savannas of South America and Africa [75,76]. Geoxyle habit is characterized by massive structural trunks under the soil surface, much higher in biomass in comparison with the small epigeal and usually seasonal shoots, and close relatedness with large tree species [75,76,77,78]. This kind of growth evolved in many lineages in ecosystems with seasonal rainfall and fire regime pressures [76,79], but probably, herbivory was also an important driver of their evolution [80,81,82].

Simon and Pennington [77] stated that extreme geoxyles have massive (more than 8 cm in diameter) belowground systems that can radiate for several meters, with some individuals forming patches up to 10 m in total diameter [80]. P. tetramera reaches these “giant” dimensions, and although not woody at all, they are very similar to the typical geoxylic suffrutices. They are not the only exception of a woodless underground tree, with Erythrina zeyheri, with corky subterranean stems, being another example of a non-woody underground tree [83].

The kind of growth of Phytolacca tetramera in relation to its phylogenetic placement and possible environmental adaptations is remarkable. The fact that current geoxylic suffrutices are predominately found in tropical to temperate seasonally hydric grassland ecosystems leads us to the possibility that P. tetramera could have differentiated from closely related trees during an interglacial cycle of the late Pleistocene, when these climatic conditions promoted the development of savannas in the Buenos Aires plains [84,85,86].

Current climate in the distribution area of P. tetramera is “temperate without dry season” according to the Köppen–Geigen classification [87]. In the Thorntwhaite classification, seasonal variations in water balance in northern Buenos Aires qualify as “moist mesothermal with scarce seasonal hydric efficiency variation and low summer concentration of thermal efficiency” [88]. The evapotranspiration/precipitation ratio is positive [89], with an annually averaged runoff of 200 mm [90]. Modern annual climate variability is moderate and caused by temperature [91]. Although there is a cold phase in winter and a dry-warm phase during summer, grasslands’ rest periods are subtle in northern Buenos Aires. However, the modern aspect of the Pampean grasslands is an extensive loess plain nowadays covered by prairies and steppes, and few thorny forests appear to be very recent, having, in all probability, an antiquity of a few thousand years [92]. When compared with today’s climate, during the Pleistocene, temperature in this territory was markedly lower, continentality was higher, and rainfall was possibly lower and/or seasonal during glacial events, similar to the desertic Patagonia [85,92,93].

Thus, it is probable that the adaptations of P. tetramera to overpass dry seasons in this humid region are anachronic traits inherited from another time. The massive subterranean trunks of P. tetramera could be an advantage in overcoming dry seasons during interglacial cycles, and subsequent glacial cycles and winter frosts.

4.3. Pleistocene Megafauna-Dominated Environment for Phytolacca tetramera

During the Pleistocene, Pampas grasslands were populated by herds of gigantic mammals that paced until the end of the Pleistocene and during the early to middle Holocene. These include big mammalian herbivores (more than 100 kg) like horses, deer and camelids (e.g., Equus, Morenelaphus, Antifer, Hippidion, and Hemiauchenia) and megamammals (more than 1000 kg) like glyptodonts, ground sloths, ungulates and mastodonts (e.g., Glyptodon, Doedicurus, Lestodon, Megatherium, Macrauchenia, Toxodon, and Notiomastodon) [66,94,95]. The extinction of most members of the megafauna in the Pleistocene/Holocene boundary (some taxa persisted during the early Holocene) matches with a sudden increase in temperature and rainfall values, but also with the dispersal and increase in human populations, suggesting that humans themselves could have been another ecological pressure that contributed towards their extinction [94,96,97]. After the megamammal extinction, humans became the largest omnivores in the Pampas, and only two medium-sized herbivores of up to 100 kg remained in the study area, the not-specialized grazer-browsers Ozotoceros bezoarticus and Rhea americana [66].

If P. tetramera is an anachronism, it should retain features related to support its coexistence with large mammals. In fact, P. tetramera exhibits traits related to support intense herbivory, trample on, and stomping disturbances typical of plants inhabiting Pleistocene megafaunal disclimax. While pruning, grazing, and other disturbances affect its aerial organs, the underground stems remain intact, enabling the plant to resprout vigorously. The numerous cambium rings found in the rhizome of P. tetramera (Figure 4b) can develop new shoots shortly after disturbances, as P. dioica do [98]. Such a rapid reaction to the cut-off is usually regarded as a response to high grazing pressures [99], like those inferred to have taken place during the Pleistocene, when the megafauna had its climax. The resistance to commonly disturbed areas is also considered an anachronic adaptation for several plants [22,23].

It should be mentioned that, as well as the rhizome-like stems, the aerial stems and leaves’ middle veins are succulent, like those of most Phytolacca species [30]. At first sight, the fleshy twigs and leaves of Phytolacca tetramera seem to be a suitable resource for cattle during the hot summer season. However, we have observed that cattle do not consume dwarf ombúes as the plants contain toxic compounds [100] and calcium oxalate needles [25], although bite marks from horses were registered. Rhinoceros and elephants also browse leaves of P. dodecandra in Africa [101], which may indicate that megafauna can tolerate the toxicity present in the vegetative parts of this genus.

4.4. How Could Phytolacca tetramera Have Persisted Thousands of Years Without a Disperser in a Suboptimal Climate?

Some key traits to understand the persistence of this species in a supposed suboptimal climate are hidden below the thick grass carpet. Vegetative reproduction in Phytolacca tetramera is possibly the main source of individual multiplication. Since the multiplication of this species depends on the fragmentation of its underground stems, which are rarely extracted from the soil, this system does not contribute to dispersal (nor does it provide variability) for the species. On the other hand, the enormous fleshy rhizomes of P. tetramera allow the plant to have independence from the surface conditions. We think that with such long-living structures underground, P. tetramera may wait for decades to have a “good year” when a lucky seed in the soil germinates. Even that modest renewal can maintain the population for centuries. In this regard, some Phytolaccaceae are characteristic of early successional stages, with seeds that remain viable in the soil for decades [102,103]. This so-called “indestructible offspring” is characteristic of evolutionary anachronic species [18,52]. The seeds of P. tetramera probably have a long-term persistence too. Although Hernández et al. [104] reported that, after one year, only 60% of the P. tetramera germinated, the season (autumn) in which the experiments were conducted could have biased their results [105]. As an example, from a sample of six seeds recollected in 2014, kept in a paper envelope without any treatment (no chemical, temperature, or humidity control) for five years, we observed that all germinated when they were sown in 2019 spring in a plant nursery (Unidad Vivero, FCNyM, UNLP). In spite that this may be considered merely anecdotal, it reinforces the idea that P. tetramera may have an “indestructible offspring”. Currently, experimental studies on the germination of Phytolacca tetramera seeds of different ages are being conducted under different conditions in the Unidad Vivero, FCNyM, UNLP (J. Ruiz Díaz com. pers.)

4.5. The Future of Phytolacca tetramera

The present study highlights the possibility that the original seed dispersers of P. tetramera vanished from the study area. The dispersal syndrome of Phytolacca tetramera indicates that a large-sized mammal might have been its original seed disperser, and other features of P. tetramera would place it in a time when grazing pressure, stomp and rainfall seasonality were higher than today. These contrasts with the conditions that are prevalent in northeastern Buenos Aires province nowadays but are congruent with the hypothesis proposing that P. tetramera may be regarded as an anachronic plant. Based on that possibility, the restricted and patchy distribution of extant populations of Phytolacca tetramera may be explained by its anachronic condition (the lack of effective seed dispersers and the disappearance of its optimal ecosystem qualities) and anthropic threats (urban growth, high grazing pressures, natural invasions, and other human activities).

Understanding the causes of the local extinction, patchy distribution, and low population size of a species is a basic prerequisite for planning future conservation strategies. Previous authors believed that human activities are pushing P. tetramera to extinction [106,107,108]. Urban growth is an obvious menace to P. tetramera [32], for which the human-mediated extinction is, at least in part, a serious risk. But why is this plant still so rare, even when the cattle activities in the rest of the region have left enormous grassland zones little altered? The premise of a risk of extinction caused by humans can be challenged or complemented by our hypothesis of the anachronic condition of P. tetramera. If we consider that it is an anachronic species, well-adapted to periodically disturbed zones, pristine habitats are not good areas for its conservation. A striking fact about this plant is that it often grows by the side of roads. A possible explanation is that cars or the weeding during road maintenance works destroy the fruits (damaging the endocarp), and that these actions have become a new form of seed dispersal for the plant. In the literature, these procedures were considered as having a negative effect on the plant populations [107], because they damage the existing plants, but otherwise may promote the dispersal and growth of new individuals.

The only “field action” carried out to date to preserve P. tetramera populations was the search for individuals in pristine (or not-so-altered) grasslands, with the assumption that the plant would prefer that condition. Although well-preserved grasslands are not uncommon in northeastern Buenos Aires province, the presence of P. tetramera there is very unusual [108]. The study of the anachronic features of Phytolacca tetramera shows that this species probably does not need a pristine natural area for its conservation because it takes advantage of sites with frequent disturbances. Urban natural reserves or protected roadsides can be adequate solutions for allowing the survival of the plant. The future planning of protected areas should focus on these zones, potentially incorporating artificial disturbances such as pruning or controlled burning experiments. The introduction of new plants produced in plant nurseries in places where P. tetramera plants are scarce (i.e., in sites with only one sex represented) and connecting isolated occurrence sites is a reasonable option. The most recent conservation effort was carried out by a team including one of the authors (EG), founded by the Ministerio de Ambiente de la provincia de Buenos Aires, with the aim of understanding the genetic variability of P. tetramera subpopulations, and trying to multiply them in nurseries, and reintroduce them in the field. Unfortunately, the project is currently unfunded since the dissolution of the Ministry of Science, Technology and Innovation under the current national administration.

5. Conclusions

Anachronism studies have been classically focused on tropical or temperate forests, and mainly on the evolutionary responses to extinct megafaunal pressures (e.g., herbivory and seed dispersal). The present work suggests that anachronic plant species may still persist in the Pampean grasslands, an ecosystem that was historically subjected to intense grazing by large herbivores. Examples of big fruits in temperate grasslands or deserts are scarce, and in the Pampean grasslands, only the wild bitter pumpkin Cucurbita maxima subsp. andreana (Naudin) Filov was previously considered an anachronic species with fruits of more than 5 cm in diameter [9]. Other classic anachronic genera like the cacti Opuntia and Cereus grow in the thorny Prosopis forests that surround the grasslands [109], thus not being strictly Pampean species [9]. To this meager list, we add the Dwarf Ombú (Phytolacca tetramera).

P. tetramera shows a combination of reproductive, ecological, and vegetative traits consistent with morphology and ecological attributes previously associated with ecological anachronisms and megafaunal dispersal syndromes. The species also exhibits adaptations compatible with persistence under recurrent disturbance regimes and seasonal environmental stress. Although direct evidence of interactions with extinct megafauna is necessarily unavailable, the available data support the hypothesis that P. tetramera represents a relic linked to late Pleistocene environments. These results have direct implications for conservation planning, suggesting that disturbance-mediated habitats may play an important role in the persistence of extant populations.