Phylogeny

Thirty species (45 specimens), about a third of the species in the genus, were sampled for molecular phylogenetic inference; samples were obtained from Brazil, Costa Rica and the Dominican Republic (see Supplementary data Tables S1–S3). Sampling was non-random and concentrated on three groups: (1) economically important species valued for human fruit consumption; (2) invasive species that threaten the biodiversity of fragile ecosystems; and (3) taxonomically problematic species that form species complexes, i.e. the Psidium grandifolium complex (Landrum, 2005) and the Psidium salutare complex (Landrum, 2003).

Total DNA was extracted using the cetyltrimethylammonium bromide (CTAB) protocol (Doyle and Doyle, 1987) or the DNeasy kit (Qiagen) in the Laboratório de Biologia Vegetal, Universidade de Brasília or Jodrell Laboratory, Royal Botanic Garden Kew. Two chloroplast (psbA–trnH and ndhF) and two nuclear [external transcribed spacer (ETS) and internal transcribed spacer (ITS) 1 + 5.8S + ITS2] regions were targeted for each accession. Amplification was carried out on a Gene Amp 9700 (Applied Biosystems, CA, USA) thermal cycler under the following conditions: psbA–trnH (Hamilton, 1999), 4 min at 94 °C; 30 cycles of 1 min at 94 °C, 1 min at 48 °C, 2 min and 30 s at 72 °C; ndhF (Biffin et al., 2006), 4 min at 94 °C; 30 cycles of 1 min at 94 °C, 1 min at 55 °C; ETS (Wright et al., 2001; Lucas et al., 2007), 4 min at 94 °C; 30 cycles of 1 min at 94 °C, 1 min at 50 °C, 1 min at 72 °C; and ITS (ITS1 + 5.8S + ITS2; Sun et al., 1994), 2 min at 94 °C, 30 cycles of 1 min at 94 °C, 1 min at 52 °C, 1 min at 72 °C; annealing temperatures were lowered to a minimum value of 48 °C and cycles increased from 30 up to 36 in order to optimize amplification; the standard stabilizing phase was 7 min at 72 °C. PCR products were purified using the QIAquick PCR Purification Kit (Qiagen). Sequencing reactions were conducted using the Taq DyeDeoxy™ Terminator Cycle Sequencing Kit (Applied Biosystems). Sequences were produced on an ABI 3730 Genetic Analyzer (Applied Biosystems). Complementary strands were sequenced, and consensus sequences obtained using Geneious™ version 10. Sequences were aligned using the MUSCLE plug-in implemented in Geneious™ 10 (Edgar, 2004) with default values, and visually inspected and corrected for alignment mismatches. The psbA–trnH inversion flanked by inverted repeats identified by Flickinger et al. (2020) in Eugenia was not found in the dataset; all sequences had the common configuration so no corrections were necessary.

Phylogenetic analyses were run in two separate datasets based on the number of molecular regions successfully sequenced for each specimen. First, we analysed a dataset comprising 30 species and 45 specimens in the ingroup and one specimen each of three species in the outgroup. This included specimens where two, three or four molecular regions could be sequenced; ten single-region species were not included. This inclusive analysis was performed so that more specimens could be included in the phylogenetic tree, even if this was at the cost of reducing the support of some relationships due to the gaps in the matrix. Next, we removed all accessions that were not complete for the four molecular regions in the matrix, leaving only those that could be sequenced for all markers (15 species and 20 accessions in the ingroup; two outgroups). This more restrictive analysis was important to evaluate the support of the main clades in a scenario where the molecular matrix is complete, albeit with fewer samples.

Phylogenetic trees were constructed based on maximum likelihood (ML) using RaxML (default parameters) and Bayesian inference (BI) using MrBayes on the CIPRES (Miller et al., 2011) gateway. ML and BI are phylogenetic inference methods that have become common in the field since the computational advances of the late 20th century allowed calculations based on large datasets. Briefly, ML utilizes sequence data to search for trees that maximize the likelihood of a particular evolutionary history given a molecular substitution model (Huelsenbeck and Crandall, 1997). BI expands the ML assumptions to incorporate prior probabilities and accounts for uncertainty in estimations by using a Markov chain Monte Carlo (MCMC) algorithm to search for a posterior distribution of trees with high probabilities. The way in which trees are summarized differs between each method. Whereas most ML software uses bootstrap (BS) values to indicate support (Felsenstein, 1981), BI usually synthesizes the posterior probability (PP) distributions on the trees by calculating a maximum clade credibility tree (Huelsenbeck et al., 2001). Consequently, metrics used to measure support of relationships are also different, varying from 0 to 100 in ML (with BS values > 70 generally considered high support; Hillis and Bull, 1993) and from 0 to 1 in BI (with values of PP > 0.95 generally considered high support; Huelsenbeck et al., 2001). Metrosideros stipularis Hook.f. was set as the outgroup in the inclusive analysis and Calycolpus goetheanus (Mart. ex DC.) O.Berg in the restricted analysis, based on their placement in the family (Vasconcelos et al., 2017b). For the BI analysis, substitution models were tested using JmodelTest (Posada, 2008) and the models that best fit the individual regions were implemented; these were GTR + G (ETS and ITS1 + 5.8S + ITS2 and psbA–trnH) and GTR + I + G (ndhF). Four MCMC runs of 10 million generations sampling every 1000 were performed and convergence was confirmed in Tracer [effective sample size (ESS) > 200, 0.25 burn-in].

Vegetative ecology

The most common life form is that of a shrub or small tree, but Psidium species can occur as small, recurrent sub-shrubs with well-developed lignotubers (hemixyles that resprout after fire or a prolonged dry season), such as Psidium salutare Sprengel (Landrum, 2017), to large trees such as Psidium myrtoides O.Berg (Tuler et al., 2017a). In fire-prone fields and savannas, Psidium species are small to large shrubs or treelets. In maritime and semi-desertic scrubs or woodlands, such as Caatingas, Restingas and Pinares, they are small to large trees. In densely forested regions, such as the Amazon and in gallery forests, several species are adapted to a riparian habit. Field notes on specimens of Psidium collected in the Amazon (e.g. P. densicomum Mart. ex DC. and P. riparium Mart. Ex DC.) frequently allude to a riparian habitat. Poeppig, collecting P. densicomum Mart. ex DC. on the Amazon River, noted that it grew exclusively along rivers and formed large aquatic populations which resembled Rhizophora mangle L. mangroves, with many fasciculate, red, adventitious roots and decumbent branches (Berg, 1857). Studies of this species (under the synonym P. ovatifolium O.Berg) in a flooded forest on the Mapire River, Venezuela (Fernandez et al., 1999), showed that flooding probably does not cause water stress in that species and that submerged leaves maintained photosynthetic and leaf conductance rates similar to those observed in aerial leaves. Riparian species are not restricted to the Amazon, they also occur in Cuba (P. orbifolium Urban), Paraguay and south-western Brazil (P. striatulum Mart. ex DC.).

Psidium species are thus edaphically very variable, with species that grow on sandy soil, limestone soil, limestone soil with rocky outcrops, mesotrophic soil and water-logged soil (Fernandez et al., 1999; Proença et al., 2013; Landrum and Cornejo, 2016). Myrtaceae are strongly ectomycorrhizal, a rare form of symbiosis estimated to occur in 2 % of land plants (Brundrett and Tedersoo, 2018) and in up to 10 % of tracheophytes, although it can be dominant in certain environmental conditions (Wang and Qiu, 2006). In vitro inoculation of Psidium cattleyanum confirmed that it is ectomycorrhizal (Freire et al., 2018), while experiments with artificial inoculation of P. guajava plantlets have found that species to be highly mycotrophic, with a mycorrhizal dependency index (RDMI) of 103 % (Estrada-Luna et al., 2002); this index compares the dry weight of mycorrhizal plantlets with that of non-mycorrhizal plantlets.

Leaf venation and anatomy

Leaves in Psidium are highly variable in size, shape, morphology, venation and anatomy; if all these parameters are considered, perhaps more so than in any other genus of Tribe Myrteae. Colleters have been sometimes observed in the leaf axils (Landrum, 2017; Tuler et al., 2021). Leaves vary in size from 0.4 cm in diameter in P. nannophyllumLiogier (1973) to 19 cm long in P. oblongatum O.Berg (Tuler et al., 2017a). The midvein can be sunken, flush or prominulous on the upper surface (Landrum, 2017; Oliveira et al., 2017). Secondary veins vary from four to 22 (Landrum, 2017; this study). The secondary venation is predominantly brochidodromous but can be so throughout the leaf or only apically, with the basal veins acrodromous or camptodromous; tertiary venation is admedial reticulate and the ultimate marginal venation can be in complete or incomplete arches, or fimbriate (Cardoso and Sajo, 2006; Fank-de-Carvalho et al., 2007; Oliveira et al., 2017). Considering the high abundance and diversity of Psidium, relatively few anatomical studies of the leaves have been conducted in the genus (see Gomes et al., 2009 for a review; Al-Edany and Al-Saadi, 2012; Oliveira et al., 2017; Endringer, 2020). Amongst leaf anatomical characters common to the genus, we can highlight a uniseriate epidermis and cuticle; hypostomatic leaves (amphistomatic in P. ratterianum Proença and Soares-Silva, Proença et al., 2010); paracytic stomata; and tector trichomes that are abundant on the abaxial surface and rare on the adaxial surface (rarely absent on the adaxial surface). The mesophile is dorsiventral with 1–3 layers of palisade parenchyma on each side and a compact arrangement of spongy parenchyma. The hypodermis is formed by two continuous layers on the adaxial surface. The vascular bundle of the midvein is bicolateral and protected by lignified fibres. Sub-epidermal secretory cavities and prismatic crystals occur throughout the mesophyll (Soares-Silva and Proença, 2008; Gomes et al., 2009; Endringer, 2020).

Reproductive biology

Psidium is mainly bee pollinated and mammal dispersed (Nic Lughadha and Proença, 1996; Gressler et al., 2006). Pollination is by large (Apidae: Bombinae; Anthophoridae: Xylocopinae) or small (Apidae: Ceratini, Meliponinae; Colletidae; Oxaeidae) bees. Phenological strategy is steady state (Gentry, 1974), i.e. a few flowers open per day during a more or less prolonged flowering period (Proença and Gibbs, 1994). Several short, synchronized episodes of flowering per year were recorded for P. guineense Sw. (Suárez and Esquivel, 1987) while P. acidum (DC.) Landrum flowers throughout the year (Falcão et al., 1992). Flowers are white, aromatic, with many anthers and a single style with a punctiform, funnel-shaped or capitate stigma (Proença et al., 2010; Landrum, 2017). In P. cattleyanum, pollen is released as a mixture of monads and tetrads (Patel et al., 1984).

Psidium firmum O.Berg is completely self-compatible, but the PERS (pre-emergent reproductive success; Wiens, 1984) range (4.6–12.7) found does not suggest habitual selfing (Proença and Gibbs, 1994). This study of P. firmum looked at the reproductive biology of eight sympatric species of Neotropical Myrtaceae in six genera and found two unusual characters in P. firmum that were not present in any of the other species: (1) the length of the styles varied within a plant so that some flowers had the stigmas raised 1–2 mm above the anthers, while others had the stigma at the same level as the anthers; and (2) cross-pollen tubes had penetrated ovules 48 and 72 h after pollination while at 72 h self-pollen tubes had still not penetrated the ovules. This suggests two possibilities: fruit set from self-pollinations results from either preferential self-exclusion or facultative apomixis. Preferential self-exclusion has been suggested to occur in Clarkia unguiculata Lindl. (Onagraceae, Myrtales). Clarkia unguiculata is a totally self-compatible species in which the use of a genetic marker (flower colour) showed that 58–100 % of the progeny of mixed self and cross pollen was outcrossed (Bowman, 1987); the author suggested that this was because the cross-pollen tubes grew faster and were responsible for most of the fertilizations. Apomixis has been recently recorded in P. cattleyanum and found to be through diplospory, an unusual and previously unrecorded pathway in Myrtaceae (Souza-Pérez and Speroni, 2017), possibly conforming to the most common type of diplospory, the Antennaria type (Nogler, 1984). Apomixis had been previously reported in Myrtaceae but from non-Neotropical genera and following other pathways. In Syzygium (Tribe Syzygieae), apomixis occurs through adventitious embryony (Thurlby et al., 2012, and references therein) and in Callistemon (Tribe Melaleuceae) through apospory (Rye, 1979). Urquía et al. (2020) suggested that the levels of clonal diversity found in populations of P. galapagaeum Hook.f. are best explained by clonal or asexual reproduction rather than by random mating between related plants. Most species of Psidium are hermaphrodites, but Psidium ovale (Spreng.) Burret is andromonoeicious, with both male and hermaphrodite flowers in the same plant (Soares-Silva and Proença, 2006).

The range of dispersal agents in Psidium was the widest recorded in a review of Neotropical Myrtaceae that included 115 species (Gressler et al., 2006), with nine out of the possible ten classes of dispersers recorded: ants, bats, birds, fish, carnivorous mammals, lizards, marsupials, monkeys and ungulates; their study did not record rodents, but rodent dispersal has also been recorded (Nic Lughadha and Proença, 1996; Alvarenga and Talamoni, 2006). Mammal dispersal is probably dominant, with a few species with small, wine-coloured fruit that suggest bird dispersal, such as P. ovale, P. macahense O.Berg, P. ganevii Landrum & Funch, P. cauliflorum Landrum & Sobral and P. grazielae Tuler & M.C.Souza (Soares-Silva and Proença 2006; Tuler et al., 2017b; Stadnik et al., 2018). In P. guajava, P. cauliflorum and P. grazielae, the fruits are reddish internally. Half-eaten fruits are thus very conspicuous to birds, and one of us (C.E.B.P.) has observed parrots returning to the same half-eaten fruits of P. guajava to feed, while V.G.S. (see also Staggemeier et al., 2017) observed that P. cattleyanum is consumed by large birds such as jays, e.g. Cyanocorax caeruleus (Vieillot, 1818) that remove whole fruits but also by smaller tanagers, e.g. Rhamphocelos carbo (Pallas, 1764), and thrushes, e.g. Turdus rufiventris (Vieillot, 1818), that peck at the fruits, consuming bits of flesh and presumably a few seeds. Psidium cattleyanum showed a keystone functional role in sustaining the fauna in a tropical forest in Southern Brazil (Pizo, 2002; Staggemeier et al., 2017): its many small seeds scattered in the pulp of a medium to large fruit allowed dispersal by a wide range of vertebrate frugivores from small birds to medium sized mammals. Cauliflory has evolved albeit rarely; it is known in two species, P. cauliflorum and P. grazielae. Cauliflory has been suggested to be associated with bat dispersal (van der Pijl, 1982) and also with increased visibility of flowers to pollinators (Wallace, 1878; Warren et al., 1997).

Floral morphology and anatomy

Flowers vary markedly in calyx structure and mode of anthesis, from completely closed buds that open by tearing or by a calyptra, to partially closed lobes, to free, shallow lobes. This plasticity is also found in other genera of Myrtaceae (Vasconcelos et al., 2017a; Giaretta et al., 2019). Besides this, flowers vary in size (4–13 mm long), bud shape and number of stamens (from approx. 80 to 720), number of locules (2–6) and ovules per locule (from approx. 3 to 180). Anthers vary by an order of magnitude in size, from 0.3 to 3 mm, and usually have an apical gland and a few smaller glands scattered along the connective, although some species have no glands and species with long anthers may have many scattered glands (Landrum, 2017; Vasconcelos et al., 2019).

Floral vasculature was found to be of the ‘consistent eight bundle’ type in P. cattleyanum and P. guineense (Pimentel et al., 2014). The presence of sclereids in the pericarp of the fruits has been recorded in several genera of Myrteae (Galan et al., 2016; Pittarelli et al., 2021, and references therein). However, Psidium is apparently the only genus in Tribe Myrteae (based on three studies that together looked at 34 species in 13 genera; Proença, 1991; Pimentel et al., 2014; Pittarelli et al., 2021) to consistently have isodiametric sclereids (stone cells) in the ovarian walls of the flower. Stone cells are found in the ovary of P. cattleyanum, P. guineense, and P. firmum (Proença, 1991; Pimentel et al., 2014); they also occur in three closely related species of Campomanesia of the ‘guazumifolia’ complex, but were absent in nine other members of Campomanesia (Pittarelli et al., 2021). However, these studies did not include all Myrteae genera, or even all nine genera in the subtribe Pimentinae to which Psidium belongs (Lucas et al., 2019): Curitiba Salywon & Landrum, Legrandia Kausel, Mosiera Small and Pimenta Lindl. have not been investigated. Stone cells were hypothesized to offer protection to ovules and seeds (Pimentel et al., 2014), an idea that is compatible with the theory presented by one of us (Proença, 1991) that they act as a mechanical barrier against oviposition by fruit flies (Tephritidae), notorious predators of guavas and other Myrtaceous fruit crops (Malavasi and Morgante, 1980; Burk, 1983; Malavasi et al., 1983). Stone cells in the ovary occur in other fleshy fruited Myrtales, such as Australasian mega-diverse Syzygium (tribe Syzygieae, Myrtaceae; Schmid, 1972; Pimentel et al., 2014, and references therein) and Mouriri Aubl. (Melastomataceae; Morley, 1976).

Placentation and fruit morphology

Although tribe Myrteae is invariably fleshy fruited (Wilson et al., 2005), Psidium is the only genus to have not only fleshy fruit walls but also fleshy, developing placentas. In all other Myrteae genera, fruit fleshiness is exclusively due to the development of ovarian tissue, i.e. the thickness and fleshiness of the fruit wall. In the Myrtales, a fleshy placenta is also found in Melastoma L. and Miconia Ruiz & Pav. (Melastomataceae; Clausing et al., 2000).

The placentation of Psidium is of the (putatively plesiomorphic) carpellate type (Pimentel et al., 2014). It has been described as either axillary or parietal, and also as bilamellate, extrusive, intrusive, lamellate or peltate (Berg, 1857; Landrum and Sobral, 2006; Soares-Silva and Proença, 2008). This can be attributed to the fact that the margins of the carpels can be deeply intrusive into the locule or hardly so, and are sometimes incompletely coalescent, so that, in some cross-sections, placentation appears parietal and in others (at different levels) axillary. The placental lamellae can bear one, two or many series of ovules. The placental lamellae can be linear, bilobed, scutellate, peltate or U-shaped. To complicate matters furthermore, the lamellae can be straight, or curve either outwards or inwards to accommodate the ovules in the locule. If the placental lamellae curve outwards, a tangential cut of the locule will show the ovules, which will appear as one or two neat series or, if the series are more numerous or the lamellae are less developed, a cluster of ovules like a bunch of grapes, sometimes described as mound like (Landrum, 2017). If the placental lamellae curve inwards (sometimes called reflexed), a tangential cut will show the smooth lamellae – only the outer series of ovules will be perceptible; other series, if present, will be hidden from view.

Seed morphology

Psidium seeds can be as few as one per fruit (rarely, and never in most of the fruits in any one species; Tuler et al., 2021) or up to 325 (Landrum, 2017). They are 2–10 mm long and have bony seed coats from (5)8 to 30 cells thick at the narrowest point, in which the cells of the outer layer are pulpy and the inner thick-walled, elongated, somewhat ragged, and superimposed at one of the ends, giving the seed an opaque surface (Landrum and Sharp, 1989). This contrasts with other Myrteae genera with bony seed coats in which the cells are hexagonal and abutting; these genera have shiny seed coats, such as Amomyrtella, Calycolpus, Mosiera, Myrrhinium Schott, Ugni Turcz. (Landrum and Sharp, 1989) and Accara Landrum (Landrum, 1990). In species in which seeds develop imbedded in placental tissue, such as P. guajava and P. guineense, most are reniform and smooth (Landrum et al., 1995); if there are edges, these are rounded and not sharp. If placentas do not develop into fleshy tissue, seeds tend to be more angular probably due to contact during development.

The bony seed coat has a plug-like operculum that in contact with water allows seeds to imbibe water, which is followed by the radicle pushing out the opercular plug and germinating through the aperture. The operculum can be shiny and mammiliform or dull and somewhat sunken (Tuler et al., 2016; Landrum, 2017). Seeds of P. guineense are non-dormant and show a typical three phase imbibition pattern but were able to germinate after 1 year of storage (dos Santos et al., 2015).

Cytogenetics

Psidium has the highest levels of polyploidy of any Neotropical Myrtaceae genus (Costa, 2009). In fact, it is the only Neotropical genus in which the diploid 2n = 22 appears to be rare, although most species have not yet been sampled. A total of 100 counts of 15 species (approx. 15 % of the genus) have been made (Supplementary data Table S2). Three species had a single count that was the diploid number (Moussel, 1965; Tuler et al., 2019a; Silveira et al., 2021), six species had a single count that was a polyploid number, while all other species had more than one count. Total counts were 100, but these were heavily biased (77 %) towards three species: P. cattleyanum (36 counts; exclusively polyploid; Atchison, 1947; Smith-White, 1948; Hirano and Nakazone, 1969; Singhal et al., 1984; Medina, 2014; de Souza et al., 2015), P. guajava (30 counts; mixed ploidy but predominantly diploid; Kumar and Ranade, 1952; Sharma and Majumdar, 1957; D’Cruz and Rao, 1962; Roy and Jha, 1962; Raman et al., 1971; Majumder and Mukherjee, 1972; Srivastava, 1977; Singhal et al., 1980, 1984; Vijayakumar and Subramanian, 1985; Pedrosa et al., 1999; Marques et al., 2016) and P. guineense (11 counts; Hirano and Nakazone, 1969; Srivastava, 1970; Chakraborti et al., 2010; de Souza et al., 2015; Marques et al., 2016; Tuler et al., 2019a; mixed ploidy but predominantly tetraploid). Diploid counts appear to be associated with narrow geographic distributions in general (Dar et al., 2020) and also in Psidium and Eugenia (Silveira et al., 2016; Tuler et al., 2019a). If this hypothesis is true, it suggests that P. guajava might have been originally a rather narrowly distributed species whose range has been expanded by man, while crossing with P. guineense as suggested by Landrum et al. (1995) may have increased its invasive potential. A study of genetic diversity in P. galapagaeum, endemic to the Galapagos Islands, suggests it is an allopolyploid (Urquía et al., 2020).

Importance to humans

The main utility of the genus Psidium to man are the edible fruits of guava (P. guajava). Guava fruits can be very rich in antioxidants, flavonoids and vitamin C (230–1426 mg 100 g^–1 fresh fruit depending upon the cultivar and environmental conditions; Vaughan and Geissler, 1997; Luximon-Ramma et al., 2003), and are consumed as fresh fruit, or industrialized in the form of juice drinks, jams, compotes, sweets, ice creams, fruit salads, biscuit fillings and tarts. The guava is a widely cultivated tropical fruit tree. During the 21st century, major producers for the world market have been Bangladesh, Brazil, China, Colombia, Egypt, India, Indonesia, Mexico, Nigeria, Pakistan, Phillipines, Thailand and the USA (Vaughan and Geissler, 1997; Pariona, 2017).

Also occasionally cultivated are P. cattleyanum, P. guineense (Vaughan and Geissler, 1997) and P. friedrichsthalianum O.Berg (Rojas-Gómez et al. 2020). These species have smaller fruits than the guava but exhibit similar fruit qualities. Psidium cattleyanum has shiny, yellow, deep pink or wine-coloured fruits, and P. guineense has thin-skinned yellow-green fruits which are juicier than guavas, while the fruits of P. friedrichsthalianum are also yellow-green and very tart. These latter species are commercialized on a small scale in local markets in Brazil and Costa Rica. In Cuba, a liqueur is made from the wild fruits of P. salutare (Kunth) O.Berg (Liogier, 1953).

Commercial guava orchards have recently suffered severely from guava decline, a disease caused by co-infection of the roots by species of Meloidogyne enterolobii, a nematode, with Fusarium solani, a fungus (Pereira et al., 2009; Carneiro et al., 2011; Gomes et al., 2011). Investigations into possible sources of resistance to this disease in the P. guajava gene pool have found it to be either absent (Cardoso et al., 2017) or borderline in a few genotypes (Cavalcanti Júnior et al., 2020). Psidium cattleyanum Sabine is resistant to infection by Meloidogyne (de Almeida et al., 2009; Marques et al., 2012), but techniques such as its use as root stock and intraspecific crosses with P. guajava have failed (Cardoso et al., 2017), which the authors attributed to possible differences in ploidy levels between these two species. Psidium myrtoides and P. acidum are also resistant (Marques et al., 2012), and approx. 10 % of genotypes of P. guineense investigated were found to be resistant (Cavalcanti Júnior et al., 2020). Therefore, the phylogenetic affinities of P. guajava are of particular importance to direct research into guava decline.

Invasiveness

Psidium cattleyanum, P. guajava and P. guineense are aggressive, pioneer species that have become naturalized throughout the tropics and are considered globally widespread invaders (Richardson and Rejmánek, 2011). Fragile island ecosystems seem particularly susceptible. In Hawaii, Mauritius, the Phillipines, the Galapagos and the Seychelles, they have become major pests that are successfully outcompeting the native endemic flora. A colonization study in Hawaii has shown that P. cattleyanum was the most abundant understorey tree beneath old Eucalyptus plantations, even though they were surrounded by native rain forest (Harrington and Ewell, 1997); it is listed as one of the 100 worst invasive alien species in the Global Invasive Species Database (2013). In the Galápagos, P. guajava forms large populations (Jackson, 1995) and is considered one of the greatest threats to local biodiversity (Urquía et al., 2019).

However, in some situations, the presence of guavas may be advantageous in secondary vegetation. A study on relative abundance of five species of arboreal mammals (macaque, langur, two flying squirrels and one giant squirrel) in forest fragments in India suggested that the presence of guava trees (that produce fruits that support a wide spectrum of birds and mammals) was beneficial (Umapathy and Kumar, 2000). Another study, in Ecuador, showed that abandoned pastures with guava tree canopies were apparently in a state of active succession to revert to forest, while open pastures were in a state of arrested succession, dominated by a few aggressive herbaceous or shrubby species (Zahawi and Augspurger, 1999).

Phylogenetic structure

Psidium emerges as a monophyletic genus, although with poor support (PP = 0.73) when the sister genus Myrrhinium Schott is included in the analysis. This is congruent with the results of Vasconcelos et al. (2017b) in which five species of Psidium were included in an inclusive phylogeny of Tribe Myrteae. The Psidium ML phylogenetic tree for which sequences of all four regions were available (restricted analysis) was almost fully resolved (Fig. 1). This tree is herein referred to as the backbone tree and major clades are congruent with the more inclusive ML tree (Fig. 2) and with the BI tree (Fig. 3), both of which included 30 species (45 accessions) that corresponds to about a third of the accepted species in the genus. Ten other species (Supplementary data Table S3) are present only in the single region trees that are available as Supplementary data Fig. S1 (ETS), Fig. S2 (ITS), Fig. S3 (ndhF) and Fig. S4 (psbA–trnH).

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Fig. 1.

Maximum likelihood phylogenetic tree of Psidium L. with 16 species (20 accessions). Only species with all four regions (ETS, ITS, ndhF and trnH–psbA) are included (backbone tree). Numbers to the right of nodes are bootstrap values. Section names are to the right of clades. Vouchers are identified by the first letter of the first collector’s surname and number. For complete voucher data, see Supplementary data Tables S1 and S3.

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Fig. 2.

Maximum likelihood phylogenetic tree of Psidium L. with 31 species (45 accessions). Numbers to the right of nodes are bootstrap values. Section names are to the right of clades. Vouchers are identified by the first letter of the first collector’s surname and number. For complete voucher data, see Supplementary data Tables S1 and S3.

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Fig. 3.

Bayesian inference phylogenetic tree of Psidium L. with 31 species (45 accessions). Posterior probability (PP) values are to the right of nodes. PP values are presented with two digits (additional digits removed without approximation). For complete voucher data, see Supplementary data Tables S1 and S3.

We identified four major clades in Psidium (Fig. 4). We propose to treat these clades as formal taxonomic sections with the following structure: section Psidium [section Obversifolia (section Apertiflora + section Mitranthes)]. Section Psidium has two strongly supported subclades that we treat as sub-sections: sub-section Psidium and sub-section Albotomentosa. This structure and the monophyly of these clades are supported by previous phylogenetic analyses in Myrtaceae, all of which included a smaller number of species of Psidium. Previous studies using Sanger sequencing that support the clades were focused on: (1) Tribe Myrteae (five species of Psidium; seven molecular regions; Vasconcelos et al., 2017b); (2) the re-establishment of P. macahense O.Berg [17 species of Psidium (originally 18 but the identity of voucher Pietro s.n. RB595299 has been changed to P. aff. cattleyanum); three molecular regions; Tuler et al., 2019b]; and (3) the Greater Antilles flora [originally seven species of Psidium (eight since we accept Calyptrogenia biflora as a synonym of Psidium amplexicaule; Landrum, 2017); three molecular regions, Flickinger et al., 2020].

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Fig. 4.

Diagnostic characters of Psidium sections. Border colours indicate sections. Pink = section Apertiflora: (A) fissured bark [P. laruotteanum; Proença 3019 (UB)]; (B) venation forming a distinct marginal vein [P. laruotteanum; Kuhlmann 34 (UB)]; (C) auxotelic racemes [P. cupreum; Faria 10009 (UB)]; (D) open floral buds, stamens few and hemispherically distributed, anthers small [P. basanthum; Proença 3022 (UB)]. Yellow = section Obversifolia: (E) large red fruit with thick, torn calyx lobes (P. cattleyanum; photograph Forest & Kim Starr); (F) compressed branches and fleshy leaves [P. gaudichaudiaum; Tuler 637 (RB)]; (G) large yellow fruit (P. ubatubense; photograph Henry M. Alexandre). Green = section Psidium: (H) large fruit with well-developed fleshy placentas [P. guineense; Tuler 625 (RB)]; (I) dichasia, floral buds closed, many stamens at same level, anthers large (Psidium guajava; photograph Amelia C. Tuler); J. brochydodromous venation with arches (P. guajava; photograph Carolyn E.B. Proença); (K). quadrangular branches [Psidium ratterianum; Proença 3016 (UB)]. Purple = section Mitranthes: (L) camptodromous, inconspicuous venation and small fruit (Psidium myrsinites; photograph Stephen A. Harris); (M) seeds few per fruit (Psidium myrsinites; photograph Maurício Mercadante); (N) bark smooth and thinly peeling [P. sartorianum; Proença et al. 2993 (UB)].

In our study, the nuclear markers ITS and ETS both showed high levels of correct section identification, with good bootstrap values for sections and few misclassified species. The chloroplast intergenic spacer psbA–trnH showed a strongly conserved identity within section Apertiflora but poor internal structuring, while the other sections showed high variability; this region may be evolving too fast to be very informative for the whole of the genus. The chloroplast ndhF gene, that expresses NADH-plastoquinone oxidoreductase, as might be expected, showed low variability and low bootstrap values. One of its most informative regions was a 9 bp region (GTTTATTAA), commonly present in other genera of Myrteae such as Calycolpus, Myrceugenia, Mosiera and Myrtus, but frequently lost in Psidium. This 9 bp region was almost always present in the basal section Psidium (except for P. guajava and one accession of P. guineense), but was lost in all species except one each of sections Obversifolia, Mitranthes and Apertiflora.

Evolution, diversity and distribution

Psidium underwent an accelerated rate of diversification from approx. 25 million years ago (mya; under macrofossil calibration) or from approx. 17 mya (under fossil pollen calibration) up to the present (Vasconcelos et al., 2017b). Eugenia L. and Psidium were the only Myrteae genera to show an abrupt increase in diversification rate at the crown (explosive radiation). However, while in the older Eugenia, diversification rates have gradually decreased over the last 10–20 million years, in the younger Psidium they have remained high (Fig. 5). Analyses of diversification through time using molecular phylogenies have been recently criticized (Louca and Pennell, 2020), so these results must be interpreted with care, and a wider sample of Psidium is desirable.

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Fig. 5.

Distinguishing morpho-anatomic characters in Psidium and their occurrence in Tribe Myrteae. (A) Hemixyle habit with lignotuber [P. bergianum; Wood 26934 (USZ)]; (B) variability in leaf pre-foliation and architecture (P. guajava; photograph CEBP); (C) amphistomatic leaves with arrows showing stomata [P. ratterianum; Proença et al. 3016 (UB)]; (D) fruit size, colour and seediness [P. cauliflorum; Tuler 480 (RB)]; (E) stone cell belt (white) in the ovary [P. firmum; Proença 610 (HEPH)]; (F) fruits with fleshy placentas (P. guajava; photograph CEBP); (G) bony seeds with operculum (black) [P. rotundidiscum; Faria et al. 4160 (UB)]; (H) seeds with pulpy testa of irregular, superimposed cells [P. firmum; Proença 638 (HEPH)]. Red = restricted to Psidium; deep pink = restricted to one to three large genera in tribe Myrteae; pale pink = restricted to a few genera in Tribe Myrteae. Photographs by first collectors in vouchered material except (C) (microphotograph by Suzanne Fank de Carvalho). Phylogeny showing diversification in Myrteae reproduced from Vasconcelos et al. (2017b) in accordance with the STM Permissions Guidelines, 28 April 28 2021).

Eugenia and Psidium are the Neotropical genera of Myrtaceae with the highest rates of polyploidy (Costa et al., 2008). Polyploidy is apparently rare in Myrcia, the second largest genus of Neotropical Myrtaceae (two polyploid populations out of 22 counts of 14 species; Forni-Martins and Martins, 2000; Costa and Forni-Martins, 2007; Amorim et al., 2012; Silveira et al., 2021). Eugenia and Psidium are also the genera with the highest diversity of fruit size and colour. Eugenia does not vary greatly in fruit seediness (1–4 seeds are the norm), while Psidium does. Also, Eugenia has a thin or crustaceous seed coat while Psidium has a thick bony seed coat with an operculum. Opercula are not unique to Psidium; they are also found in other genera in Subtribe Ugniinae such as Mosiera, Myrteola and Ugni (Landrum and Sharp, 1989; Salywon and Landrum, 2014), and in Subtribe Pimentinae, such as Pimenta pseudocaryophyllus (Gomes) Landrum and Curitiba prismatica (Landrum and Sharp, 1989; Salywon and Landrum, 2007) and in Amomyrtella (unplaced; Lucas et al., 2019). Psidium is, however, the only genus to combine operculate seeds with both bird (Snow, 1981) and a wide range of other dispersal agents, including mammals (Pizo, 2002; Gressler et al., 2006).

The combination of a wide range of dispersal agents, a wide range of seediness, operculate seeds that can exhibit long-term mechanical dormancy, stone cells in the ovarian tissue, polyploidy and apomixis is tentatively proposed as the main drivers behind the rapid rates of diversification in Psidium, as well as the ability to adapt to many different habitats. Polyploidy can break self-incompatibility (Richards, 1997; Miller and Venable, 2000), facilitate plant invasions, increase flower, fruit and seed size, and propitiate niche shifts by increasing drought tolerance (ter Beest et al., 2012). Speciation in Psidium seems to have occurred mostly by such niche shifts through vegetative specialization and adaptation to different dispersal agents. This is reflected in the great variation in plant architecture and leaf pre-foliation, shape, textures, venation and anatomy, seed number and seed size found within the genus (Rotman, 1976; Landrum, 2005, 2017; Proença et al., 2010, 2013, and references therein). Apomixis facilitates range expansion by generating clonal populations (Hojsgaard and Hörandl, 2015) and has been recorded in two megadiverse Myrtalean genera: Syzygium (Myrtaceae), the most species rich genus in the family (Govaerts et al., 2008), and Miconia (Melastomataceae), the most species-rich woody genus with an exclusively Neotropical distribution (Goldenberg et al., 2013).

Infra-generic classification

Three of the four clades identified in the phylogeny were found to mirror three of the six sections proposed by Berg (1857) in Flora Brasiliensis (Table 1). The alignment of Berg’s sections with these three major clades in our phylogeny was mostly accurate: a relatively small percentage (9–16 % depending on the section) of his names were misclassified. The exception was the artificial sect. Rigidifolia in which the three species were not found to be closely related. Since Berg indicated no type species, judicious lectotypification that would reflect phylogenetic structure was undertaken. To ensure that the infra-generic structure is congruent with the phylogeny, we have recognized two of Berg’s sections, reduced two of them to sub-sections and synonymized two of them. The genus Mitranthes O.Berg, a synonym of Psidium, was raised to section level as the fourth section.

Taxonomic treatment

Psidium L. Species Plantarum 470. 1753. Type: P. guajava L.

Guajava P. Miller, Gard. Dict. Abr. ed. 4. 28 Jan 1754.

Cuiavus C. J. Trew, Pl. Sel. Pinx. Ehret 4: 12. 1754.

Guaiava Adanson, Fam. 2: 88, 563 (‘Guiava’). Jul–Aug 1763.

1. Psidium L. sect. Psidium, Typus: P. guajava L.

• Calyptropsidium O.Berg, Linnaea 27: 347, 349. Jan 1856 (‘1854’). Typus: C. friedrichsthalianum O.Berg.

• Psidium sect. Costata O.Berg, Fl. Bras. 14(1): 396. Typus: P. guajava L. (Lectotype here designated), nom. illeg. to be substituted for Psidium sect. Psidium

• Psidium sect. Crenatifolia O.Berg, Fl. Bras. 14(1): 407. Typus: Psidium alatum O.Berg (Lectotype here designated) = Psidium australe var. suffruticosum (O.Berg) Landrum, syn. nov.

C.E.B.P. wrote the paper. Species circumscription and descriptions were jointly constructed by C.E.B.P. and A.C.T., and anatomical descriptions were led by the latter author. I.R.C. and E.R.F.-M. were major contributors to discussion on cytogenetics. Sampling in the field was done by J.E.Q.F., I.R.C., C.E.B.P. and V.G.S. Modus operandi in the lab, experimental design and analysis and interpretation of molecular results had significant input from T.N.C.V., E.J.L., P.W.I., L.R.M. and P.S.d-C. Most sequences were generated by I.R.C., with significant contributions from C.E.B.P., V.G.S. and T.N.C.V.. All authors contributed with comments and suggestions that improved the final draft of the manuscript. We thank the photographers that kindly contributed images (cited by name in the legends) to Figs 4 and ​and5,5, Maria Rosa Zanatta for assistance in improving the illustrations, and Stephen A. Harris for reviewing an earlier draft of this manuscript.