Conservation and genetic diversity of populations of Oryza glumaepatula Steud. in ferruginous mountaintop lakes of the Brazilian Amazonia

Highlights

• Wild rice populations are more heterozygous on mountaintops than in lowlands.

• Mountaintop populations are genetically very distinct from lowland populations.

• In contrast to the rest of the species, mountaintop plants are outcrossing.

Abstract

Natural populations of Oryza glumaepatula, a wild relative of cultivated rice, were found in lakes at mountaintops of the Carajás Ridge, in Carajás National Forest. This species is generally found in aquatic environments of lowlands surrounded by tropical forests, and this is a rare case in which the species colonized high altitude lakes, situated amidst campo rupestre (rupicolous vegetation). This peculiarity instigated an interest on the natural history of these populations, especially on which lowland populations were the sources of the propagules that colonized the Carajás lakes. This study aimed at describing the genetic structure and divergence among four mountaintop populations in order to lay grounds for their long-term conservation. As an external comparison, four O. glumaepatula populations were used, three from Amazon Basin rivers (Solimões, Negro and Xingu), and another from the Paraguay Basin, in the Pantanal biome, in a total of 257 individuals. Eight microsatellite loci were evaluated in the eight populations. The Carajás populations showed less genetic diversity than the others, which suggests that they went through a founder effect as of their initial settling on the ridge. The fixation index was negative for the Carajás lakes populations, and this excess of heterozygotes is unexpected for a traditionally selfing species. This may be the result of selection for the ferruginous environment and/or due to the vegetative propagation of highly heterotic individuals. Two groups were detected, showing a lower divergence among the populations found in the ferruginous lakes at the mountaintops of the Carajás Ridge, when compared to the populations from riverine environments in the lowlands. For genetic conservation purposes, a large number of individuals was transplanted, based on the information generated in this work, to lakes devoid of natural populations of O. glumaepatula in a nearby ridge possessing the same rupicolous vegetation.

Introduction

Ferruginous rupestrian grasslands or “campos rupestres” are a rare and old-grown ecosystem, characterized by herbaceous and/or shrub vegetation with high species richness and high endemism (Conceição et al., 2016). These fields originate in ironstone (“canga”) on open, dry areas at mountaintops and their plants show extraordinary adaptations to extremely restrictive edaphic conditions, both physically and chemically (Schaefer et al., 2016). The ferruginous “campo rupestre” of the Serra dos Carajás mountain ridge, at the Carajás National Forest (Carajás Flona, for short), in the Brazilian Amazonia (Pará State), still has an additional peculiarity: the occurrence of many rupestrian lakes.

These lakes vary widely in their seasonality, some retaining water in a volume enough to hold their vegetation, others drying up and keeping part of their vegetation in a latent form. The predominant plant species are grasses.

The species collected in the mountaintops of Serra dos Carajás is one of four species of native wild rice found in the Neotropics: O. latifolia Desv., O. grandiglumis (Döll) Prod., O. alta Swallen e O. glumaepatula Steud. (Morishima, 1994). “Native” means that they were not introduced by humans and “wild” means that they were never domesticated. These species are, however, occasionally consumed by native people and other riverside human communities, after having been harvested directly from natural, non-planted, wild-rice fields, always serving as a diet supplement, never a staple food (Oliveira, 1993). Oryza glumaepatula is the only diploid of the four Neotropical species, the others being tetraploid, and can be crossed with O. sativa L. with only mild technical difficulty, thus being utilized in rice breeding programs (Brondani et al., 2001, Yoshimura et al., 2010) for the introgression of important traits from the wild species, such as disease resistance and self-incompatibility, enlarging the genetic basis of the crop (Karasawa et al., 2012).

Populations of O. glumaepatula are widely distributed geographically, and occur from Mexico to Paraguay, always associated to river basins. In Brazil, populations of O. glumaepatula can be found in the Amazonas-Solimões basins, Paraguay (including the wetlands of Pantanal Matogrossense) and Tocantins-Araguaia basins, besides ‘Baixada Maranhense’ (an extensive wetland in the state of Maranhão formed by several confluent tributaries) (Oliveira, 1993, Oliveira, 1994, Brondani et al., 2002, Brondani et al., 2005, Vaughan et al., 2003). O. glumaepatula presents biannual, annual or perennial populations, depending on their geographical location (Oliveira, 1993, Akimoto et al., 1998), growing along the seasonally flooded river banks and both permanent and temporary lakes, normally in deeper waters than do the tetraploid species, but seldom deeper than about 10 m. Vegetative propagation by spontaneous abscission of the culms, followed by their dispersion by the stream, as well as the dispersal by seeds, favors the formation of new populations or groupings both down- and upstream due to the action of the river cycle, the wind, fish and birds (Black, 1950, Akimoto et al., 1998). When vegetative propagation occurs, the new colonies are clones of individuals of the populations of origin. The dispersal of culms and seeds promotes the reduction of genetic diversity among populations and regions of the same river basin, so that the river basin becomes a unity in the hierarchical genetic structuring of the species.

Rock-embedded high-altitude lakes are an extremely uncommon habitat for Oryza in the Neotropics, and this called our attention to the natural history of the populations growing in them. The following questions naturally arose: (1) Where did the propagules that colonized the Serra dos Carajás mountaintop lakes come from? Has there been solely one or rather several sources of origin? (2) What effect did colonization have on the genetic variation of these populations at both molecular and phenotypic levels? (3) What the relative roles did phenotypic plasticity and local adaptation have in the successful colonization of an environment with a very high content of oxidized iron by plants coming from soils and waters with very low iron content? (4) If adaptation has indeed played a role, which vegetative or reproductive traits were involved? (5) Why have some lakes been colonized and others have not? All these questions are relevant and, in all, represent a quite complex problem, with a varied palette of methods required for its study. In this work, only question (2) was dealt with, the others being left for future studies.

To address the molecular part of question (2) we adopted microsatellite markers. Microsatellite markers (or SSR) have been used for measuring genetic diversity and structure for the conservation of wild rice species in several regions of the world. For instance, to investigate the Asian O. rufipogon Griff. (Gao et al., 2000a, Gao et al., 2002a, Gao et al., 2002b, Gao, 2004), considered the ancestor of the cultivated species O. sativa L. (Khush, 1997); O. officinalis Wall ex Watt., O. granulata Nees et Arn. ex Watt., O. minuta J.S. Presl. ex C.B. Presl., and O. ridleyi Hook. (Ishii and McCouch, 2000, Gao et al., 2000b, Gao et al., 2005, Zhou et al., 2003, Gao, 2005); the Australian O. australiensis Domin. and the Neotropical O. latifolia Desv., and particularly, O. glumaepatula, for which primers were selected and tested (Brondani et al., 2001, Brondani et al., 2002, Karasawa et al., 2007a), and used later in the estimation of the genetic structure and the reproduction system of Brazilian populations (Brondani et al., 2005, Karasawa et al., 2007b).

The objective of this study was to investigate, with microsatellite markers, the genetic diversity and structure of four populations of O. glumaepatula from the ferruginous, rock-embedded lakes of Carajás, a type of environment in which genus Oryza has never been the object of any scientific study whatsoever, comparing with four O. glumaepatula populations originated in aquatic environments (rivers) in the lowlands surrounded by tropical forests. In the long run, this study aims to provide practical information for the genetic conservation of these Carajás populations and for the understanding of their adaptation to that extremely iron-oxide-rich environment.