BIOTROPICA 39(5): 628–636 2007 10.1111/j.1744-7429.2007.00314.

Low Interannual Variation of Mating System and Gene Flow of Symphonia globulifera
in the Brazilian Amazon

Francimary da Silva Carneiro1 , Alexandre Magno Sebbenn2,3,4 , Milton Kanashiro1 , and Bernd Degen3
1 Embrapa Amazônia Oriental, C.P. 48, 66095-100, Belém-PA, Brazil
2 Instituto Florestal de São Paulo, Caixa Postal 1322, 01059-970, São Paulo, Brazil
3 Bundesforschungsanstalt für Forst- und Holzwirtschaft (BFH), Institut für Forstgenetik Forstpflanzenzüchtung, Sieker Landstrasse 2, D-22927,
Grosshansdorf, Germany

ABSTRACT
Six microsatellite loci were used to compare the mating system and gene flow in two consecutive years of a natural, unlogged population of Symphonia globulifera in a
500 ha experimental plot in the Brazilian Amazon (Flona Tapajós). The species had a low density of reproductive trees per hectare (d = 0.46 trees/ha). We analyzed 205
trees and 261 and 487 open-pollinated seeds from 26 and 30 mother-trees in the years 2002 and 2003, respectively. A significant spatial genetic structure was detected
for the adult trees for distances up to 100 m. We observed only small interannual differences in multilocus outcrossing rate (tˆm(2002) = 1.000, tˆm(2003) = 0.998),
biparental inbreeding (tˆm − tˆs (2002) = 0.093, tˆm − tˆs (2003) = 0.063), and paternity correlation (r̂ p(2002) = 0.414, r̂ p(2003) = 0.386). The number of pollen donors
contributing to mating of each tree in both years was estimated to be low ( N̂ e p = 2.4 − 4.0). Using TWOGENER analysis to calculate the density of reproductive trees
and the distance of pollen dispersal for normal and exponential models, the lowest error was detected for exponential model. For this model, the estimated density
of reproductive trees was lower in 2002 (dˆe (2002) = 0.03 trees/ha) than 2003 (dˆe (2003) = 0.19 trees/ha), resulting in a higher distance of pollen dispersal in 2002
(δ̂(2002) = 444 m) than 2003 (δ̂(2003) = 154 m), although these changes did not affect the outcrossing and correlated mating rates.

Abstract in Portuguese is available at http://www.blackwell-synergy.com/loi/btp

Key words: biparental inbreeding; Neotropical tree species; pollen dispersal; spatial genetic structure; TWOGENER.

THE MATING SYSTEM AND GENE DISPERSAL OF TROPICAL TREES have
been proven to be highly dynamic and sensitive to ecological factors.
Many studies describe variation of outcrossing rate among different
populations of the same species (Murawski et al. 1994, Aldrich &
Hamrick 1998, Lee 2000, Sebbenn et al. 2000, Collevatti et al.
2001, Rocha & Aguilar 2001, Obayashi et al. 2002, Dick et al.
2003a, Funch et al. 2003, Moraes et al. 2004) and a limited num-
ber of studies describe year-to-year variation in outcrossing rate
in the same population (Murawski et al. 1990, Hall et al. 1996,
Caron et al. 1998, Rocha & Aguilar 2001, Hufford & Hamrick
2003, Kenta et al. 2004). For perennial plants, single-season studies
may provide a limited characterization of generational patterns of
mating (Irwing et al. 2003). The level of outcrossing and the distri-
bution of effective pollen might change from one flowering event
to another simply as a function of demographic structure, phe-
nology, and pollinator community composition and abundance.
Whereas some studies with insect-pollinated tropical tree species
showed little variation in the mating system among years (Hall
et al. 1996, Kenta et al. 2004), others demonstrated substantial
variation in outcrossing rates (Caron et al. 1998, Rocha & Aguilar
2001, Hufford & Hamrick 2003,), paternity correlation (Rocha &
Aguilar 2001), and in pollen composition among years (Irwing et al.
2003).
Received 1 June 2006; revision accepted 17 December 2006.
4 Corresponding author; e-mail: alexandresebbenn@yahoo.com.br
Symphonia globulifera L. F. (Clusiaceae) is a hermaphroditic
species with an exceptionally large geographic distribution from
Mexico to Rio de Janeiro, Brazil and it is also present in tropical
West Africa. Range wide phylogeographic investigation (Dick et al.
2003b) has indicated the colonization of Central and South Amer-
ica through marine dispersal from Africa during the mid-Miocene
(ca 15 mya). There are contrasting reports on the community of
pollinators of S. globulifera at different places. In Costa Rica, Pas-
carella (1992) observed Lepidoptera as the most important pollina-
tors whereas, in Central French Guiana, Gill et al. (1998) identified
perching birds, and, in the Central Amazon, Bittrich and Amaral
(1996) and Maues (2001) observed hummingbirds as the principal
pollinators. In South America, the density of S. globulifera is also
extremely variable among populations, with 122 trees/ha (diameter
at breast height [dbh] > 10 cm) in Guiana (Andel 2003), over 10.5
trees/ha in French Guiana (Degen et al. 2004), less than 1 tree/ha in
Panama (Center for Tropical Forest Sciences, 2000 forest census),
and 0.46 tree/ha in Tapajós in Brazil. This geographic variation in
the type of pollinator and tree density probably affects the mating
system of different S. globulifera populations. Variation in mating
system due to changes in type and abundance of pollinators with lat-
itude gradient has been observed in some studies (Molina-Freaner
et al. 2003, Silva-Montellano & Eguiarte 2003, Valiente-Banuet
et al. 2004). For example, in Agave lechuguilla, populations in the
South had an excess of heterozygous individuals, whereas popula-
tions in the Central section had genotypic frequencies near Hardy–
Weinberg equilibrium and those in the North had an excess of

628

C 2007 The Author(s)

Journal compilation

C 2007 by The Association for Tropical Biology and Conservation

homozygous individuals (inbreeding). This pattern was consistent
with the higher rates of pollinator visiting and greater fruit set in the
South (Silva-Montellano & Eguiarte 2003). Changes in population
density can also affect the mating system of a species, altering levels
of self-fertilization and correlated mating, as has been detected in
studies comparing mating system in trees localized in continuous
forest and isolated in fragmented landscape or in logging popu-
lations (Murawski et al. 1990, 1994; Aldrich & Hamrick 1998;
Sebbenn et al. 2001; Cascante et al. 2002; Obayashi et al. 2002;
Fuchs et al. 2003; Quesada et al. 2004; Lobo et al. 2005). Alteration
in population density can also affect the species self-incompatibility
system, changing the mating system from predominantly outcross-
ing to a mixed mating system as has been suggested for Paquira
quinata (Fuchs et al. 2003) and Ceiba pentandra (Lobo et al. 2005).
The present study aimed to quantify interannual variation of
mating system and gene flow in a population of S. globulifera with
low density of reproductive individuals in continuous forest in the
Brazilian Amazon. The results were compared to published data
from Northern populations (Aldrich & Hamrick 1998, Degen et al.
2004) where the type of pollinators and population density are dif-
ferent. We expected that the low density of reproductive individuals
in the Amazon Forest would correspond to high fertilization rate,
long pollen flow distance, low biparental mating (absence of spatial
FIGURE 1. Spatial position of S. globulifera trees in the 500-ha experimental plot at Tapajós (Brazilian Amazon). Squares show the position of the sampled seed
trees.
Mating System and Gene Flow in Symphonia globulifera 629
genetic structure), and a low number of pollen donors contributing
to the offspring.
METHODS
SITE AND THE SAMPLE SIZE.—The study population is situated in
a 500-ha experimental plot in the Tapajós National Forest (TNF)
South of Santarém, Pará State, Brazil at 2◦ 51 S and 54◦ 57 W and
175 m above sea level (Fig. 1). The area is a flat plateau covered
by a dense terra firme forest. The climate is tropical with a mean
annual rainfall of 2000 mm (peaks from February to May) and a
dry season of 2–3 mo (August to October). Annual temperature
averages 27◦ C in the humid lowland areas. The 500-ha experimen-
tal plot was established as an intensive study plot for the ecological
and genetic studies of the Dendrogene Project (Embrapa Amazônia
Oriental/DFID, Kanashiro et al. 2002). The 500-ha plot was sub-
divided into six subplots with four 100-ha subplots (subplots 3–6)
and two 50-ha subplots each (subplots 1 and 2) (Fig. 1). All S.
globulifera trees with dbh > 20 cm were identified and mapped in
subplots 1–6, resulting in a density of 0.46 reproductive trees per
hectare. Additionally, in subplot 5 all trees with dbh > 10 cm were
also identified and mapped, as we aimed to use this data set for
630 Carneiro, Sebbenn, Kanashiro, and Degen
simulation studies of forest logging (Fig. 1), resulting in a density
of 0.83 trees/ha. The distance among trees ranged from 14 m to
2962 m, with an average of 1234 m. We sampled cambium from
205 adult trees representing more than 90 percent of all mapped
trees. In 2002, 261 seeds were collected direct from the crown of
26 mother-trees (9–15 seeds per tree) and in 2003, 487 seeds were
collected from the crowns of 30 mother-trees (12–30 seeds per tree)
(Fig. 1). In 2002, the number of fruiting trees was higher than
2003: in 19 cases seeds could be collected from the same trees in
both years. The distance between sampled seed trees varied between
26 m and 2951 m with a mean of 1282 m in 2002 and 1295 m in
2003.
MICROSATELLITE ANALYSIS.—DNA was extracted from the cam-
bium of adult trees and from the seeds according to the protocol
of Colpaert et al. (2005). Six primer pairs Sg03, Sg18, Sg06, Sg08,
Sg09, and Sg10 were used for microsatellite analysis according to
Degen et al. (2004) and Vinson et al. (2005). Amplification prod-
ucts were visualized by electrophoresis with a LIZ-500 size Standard
(Applied Biosystems Incorporated, ABI; San Mateo, CA, U.S.A.) on
an ABI 310 automated sequencer. Allele sizes were scored using the
program Genotyper from Applied Biosystems. The observed segre-
gation of alleles within families showed co-dominant inheritance of
the microsatellite fragments.
DATA ANALYSIS.—The fixation index was estimated for adults and
offspring as Fˆ = 1 − ( Ĥo / Ĥe ), where Ho is the observed heterozy-
gosity, and He is the expected heterozygosity at Hardy–Weinberg
equilibrium. The significance of F values was tested using Monte
Carlo permutation methods (Degen et al. 1999). The total co-
efficient of inbreeding in seeds (Fo ) was portioned into its com-
ponents resulting from self-fertilization ( F̂ s = 0.5sˆ (1 + Fˆ p ),
where Fp is the coefficient of inbreeding of seed-trees) and mating
among relatives ( F̂ tm −ts = θ̂ p , where θ p is the coancestry coeffi-
cient between outcrossed relatives). Thus, it can be estimated as
Fˆ o = Fˆ s + F̂ tm −ts = 0.5sˆ (1 + F̂ p ) + tˆm θ̂ p , without assuming
inbreeding equilibrium.
The spatial distribution of genotypes was analyzed for the adult
trees using the coancestry coefficient (θ x y ) between pairs of trees
within ten distance classes, at 50-m intervals. We used the 50-m in-
tervals to get comparable results to the study of Degen et al. (2004)
in French Guiana. We also used smaller (25 m) and larger (100 m)
intervals to estimate the impact of the scale of intervals, but the
correlograms showed the same general pattern. The coancestry co-
efficient measures the probability of identity by descent of two
randomly sampled alleles at the same locus in two different individ-
uals. The coancestry coefficient was estimated according to Loiselle
et al. (1995) and Hardy and Vekemans (2002). In theoretical terms,
the θ x y parameter ranges from zero to 1, assuming a value of zero
in the absence of relatedness between individuals, 0.0625 for first-
cousins, 0.125 for half-sibs, 0.25 for full-sibs and parent-offspring,
and 0.5 for self-sibs and clones. The 95% CI of the multilocus
coancestry coefficient was estimated from the standard error, calcu-
lated by Jackknifing over loci.
The mating system was analyzed using the mixed-mating
model (Ritland & Jain 1981) and the correlated mating model
(Ritland 1989). Mating system parameters, including single-locus
(ts ) and multilocus outcrossing rate (tm ), the probability of mating
among relatives (tm − ts ), selfing correlation (r s ) or the probability
of randomly sampling two sibs and both are self-sibs, multilocus
paternity correlations (the likelihood that two randomly selected
offspring from a single mother share the same father, r p(m) ), and
pollen and ovule allele frequencies were estimated using MLTR pro-
gram (Ritland 2004). Standard errors for parameter estimates were
based on 500 bootstraps. The effective number of pollen donors,
as N̂ e p = 1/r̂ p , was computed from paternity correlation. Av-
erage coancestry coefficient within families ( x y ) was estimated
as  ˆ x y = r̂ x y /2, from relatedness correlation (r xy ) within fami-
lies, r̂ x y = 0.25(1 + Fˆ p )[4sˆ + (tˆ2m + r̂ s tˆm sˆ )(1 + r̂ p )] (Ritland
1989), where Fp is the inbreeding coefficient in the parental gener-
ation and s is the selfing rate (sˆ = 1 − tˆm ). The variance effective
size (N e (v) ) was estimated using the Cockerham (1969) estimator,
N̂ e (v) = 0.5/ ˆ x y . The 95% CI for the parameters was estimated
√
from standard deviation (SD) as C I95% = me a n ±1.96(S D/ m),
where ‘mean’ is the average estimated parameter, and m is the num-
ber of sample families. The confidence interval of the estimates was
used to verify if there were statistical differences between years in
the estimated parameters.
Following Smouse et al. (2001) we made a TWOGENER analysis
using the progeny arrays of two reproductive events. The principle
of this method is to estimate the differentiation of allele frequencies
among the pollen pools ( f t ) sampled by several mother-trees in
the population. The relation between  f t and dispersal distance
has been shown for given dispersal curves (Austerlitz & Smouse
2001a), allowing the development of several estimates of pollen dis-
persal (Austerlitz & Smouse 2002). A general estimate is based on
the global  f t measured across all the seed-trees of the population.
This provides an estimate of the pollen dispersal distance (δ) assum-
ing a given dispersal curve and a density of reproducing adults (d)
in the landscape. Pairwise  f t estimates between all mother-trees
in the population were calculated to derive pollen dispersal distance
(δ) and density (d) of reproductive adults (joint estimation). As
noted by Austerlitz and Smouse (2002), the pairwise analysis yields
more accurate estimates of  f t but requires larger sample sizes.
The analysis also included information on the spatial position and
genotypes of all sampled reproductive trees so the estimates could
be corrected according to the observed spatial genetic structure. We
tested the normal and exponential dispersal function, and used a
proportion of the known adult density to estimate density of repro-
ducing adults. In the Tapajós National Forest, S. globulifera had a
density of 0.83 trees (dbh > 10 cm), but observations showed that
only trees with dbh > 20 cm flowered. Moreover, we found a vari-
ation in flowering periods among trees. Thus, as only reproductive
trees are relevant for studying gene flow by pollen, we estimated
the pollen dispersal assuming an upper bound of the density of
reproductive trees of 0.46 (d max ) and a lower bound of 0.1 (d min ).
We computed the 99% CI of  f t by bootstrapping among loci
with 1000 replicates (Weir 1996). Following Austerlitz and Smouse
(2001b) the parameter  f t was corrected for the coefficient of in-
breeding in the parental generation (Fp ) as  ˆ = ˆ ft (1 + Fˆ p ). We
ft

used the parameter  f t to estimate the effective number of pollen
donors: N̂ e p = 1/2 ˆ  (Smouse et al. 2001). Again we used the
ft
upper bound of 0.46 trees/ha (d max ), and the lower bound of 0.1

trees/ha (d min ). Assuming a homogeneous pollen dispersal around
a tree, we estimated the effective pollination neighborhood area as,
 e p = π(δ̂ 2 ) (Nason et al. 1998).
RESULTS
SPATIAL GENETIC STRUCTURE.—There was a strong spatial genetic
structure indicated by positive coancestry coefficients (θ x y ) between
pairs of trees up to a distance of 100 m (Fig. 2). The highest
coancestry coefficient was obtained for the distance class from zero
to 50 m (θ̂x y = 0.119;P < 0.05), which is comparable to the value
for half-sibs (θ x y = 0.125).
MATING SYSTEM.—We did not detect great differences in mating
system parameters between the two reproductive years (Table 1).
Estimates of the multilocus outcrossing rate were not statistically
different from one in both reproductive years (tˆm(2002) = 1.000,
tˆm(2002) = 0.998; P > 0.05). The single-locus outcrossing rate
(ts ) was significantly lower than the multilocus rate in both years
(tˆs (2002) = 0.907,P < 0.05, tˆs (2003) = 0.936, P < 0.05), and
significantly different between reproductive years (Table 1). Dif-
ferences between multilocus and single-locus estimates were sig-
nificantly different from zero (tˆm − tˆs (2002) = 0.093, P < 0.05;
tˆm − tˆs (2003) = 0.063, P < 0.05), suggesting the occurrence of
mating among relatives. But, at 95% confidence limits, these es-
timates are not different between years. The self-correlation was
low and statistically different from zero for the two reproductive
events, (r̂ s (2002) = 0.110,P < 0.05; r̂ s (2003) = 0.109, P < 0.05),
but not significantly different between years, (Table 1), showing low
variation in individual outcrossing rate among seed-trees. Paternity
correlation estimates were high and statistically different from zero
(r̂ p(2002) = 0.414,P < 0.05; r̂ p(2003) = 0.386, P < 0.05), but
FIGURE 2. Correlogram with the coancestry coefficient in ten distance classes, for all pairwise comparisons among trees of S. globulifera at Tapajós, Pará, Brazil.
Dotted lines indicate the limits of the 95% CI obtained by Jackknifing among loci. (∗ P < 0.05).
Mating System and Gene Flow in Symphonia globulifera 631
not statistically different between years (Table 1). From paternity
correlation, the number of pollen donors was estimated to be about
three.
FIXATION INDEX, COANCESTRY AND VARIANCE EFFECTIVE SIZE
WITHIN PROGENY.—The fixation index was positive and statisti-
cally different from zero in adult trees (Fp ) and sampled seeds (Fo )
from both years (Table 1), indicating probable inbreeding. The par-
tition of the fixation index in the seeds, due to selfing and biparental
inbreeding indicates that all inbreeding came from mating among
relatives, F̂ o = Fˆ tm −ts = θ̂ p . The estimated average coancestry co-
efficient and the variance effective size within progeny were higher
than expected for half-sibs (0.125 and 4, respectively) and not sta-
tistically different between years.
GENE FLOW BY TWOGENER ANALYSIS.—The differentiation of
pollen pools among seed-trees corrected by inbreeding coefficient
( f t ) was statistically different from zero, but not different between
years (Table 1). From TWOGENER analysis we estimated that 3–4
trees were the fathers (Nep ) of the sampled seeds of each mother-
tree, but these values were not statistically different between years.
Both the 2002 and 2003 analyses, using the fixed density of 0.46
and 0.10 reproductive trees/ha, converged to the same estimated
density of reproductive trees, pollen dispersal distance, and error
(Table 2). The joint estimation of pollen dispersion distance (δ)
and density of reproductive trees decreased the estimated tree den-
sity in all analysis. In 2002, the estimated tree density decreased
to 0.05 trees/ha (normal model) and 0.03 trees/ha (exponential
model), and in 2003 down to 0.23 trees/ha (normal model) and
0.19 trees/ha (exponential model). Differences in the error, calcu-
lated as a quadratic criterion by fitting the observed and expected
pairwise  f t values (Austerlitz & Smouse 2002), were quite small
among the models. In general, the smallest error was obtained for
632 Carneiro, Sebbenn, Kanashiro, and Degen

TABLE 1. Estimates for mating system and gene flow in two reproductive years in a population of S. globulifera.

Seeds

Parameter 2002 2003

Multilocus outcrossing rate: tm 1.000 (0.982–1.018)a 0.998 (0.982–1.013)

Single-locus outcrossing rate: ts 0.907 (0.902–0.912) 0.936 (0.931–0.941)
Outcrossing rate among relatives: tm − ts 0.093 (0.076–0.110) 0.063 (0.049–0.077)
Self-correlation: r s 0.110 (0.098–0.122) 0.109 (0.101–0.117)
Multilocus correlation of paternity : r p 0.414 (0.401–0.427) 0.386 (0.378–0.394)
Effective number of pollen donors: N̂ e p = 1/r̂ p 2.4 (2.3–2.5) 2.6 (2.5–2.7)
Fixation index in adult trees Fp 0.086 (0.040–0.245)∗∗ –
Fixation index in families Fo 0.078 (0.054–0.127)∗∗ 0.101 (0.038–0.174)∗∗
Coancestry coefficient within progeny:  x y 0.192 (0.183b –0.204c ) 0.189 (0.176–0.211)
ˆ xy
Variance effective size: N̂ e (v) = 0.5/ 2.60 (2.45–2.74) 2.65 (2.36–2.83)
Genetic divergence among pollen pools:  f t 0.135 (0.072–0.190)d 0.173 (0.132–0.221)
Corrected genetic divergence among pollen pools:  ˆ t =
ˆ f t/(1 + Fˆ p )e 0.125 (0.070–0.180) 0.160 (0.120–0.200)
f
ˆ 
Effective number of pollen donors: N̂ e p = 1/2 t 4.0 (2.8–7.1) 3.1 (2.5–4.2)
f

95% CI, mean ± 1.96EP; b  ˆ x y (95%−l o wer ) = 0.125(1 + Fˆ p(min) [4ˆs min + (tˆ2 ˆ x y (95 −up per ) =
m(max) + r̂ s (min) t m(max) s (min) )(1 + r̂ p(min) )];
a ˆ ˆ c 

ˆ
0.125(1 + F p(max) [4ˆs max + (tˆm(min) + r̂ s (max) tˆm(min) sˆ(max) )(1 + r̂ p(max) )], where max and min are the maximum and minimum values estimated for the
2

95% CI; d 99% SE; e Corrected for inbreeding in the parental generation (Austerlitz & Smouse 2001b); ∗∗ , P < 0.01.

the joint estimation in the exponential model. Thus, we considered DISCUSSION

the joint estimation of pollen dispersal distance and density of repro-
ductive trees and the exponential model as the best. The estimates
of the average pollen dispersal distance (δ) from the normal model
were lower than the exponential model in both years (Table 2). In
2002, the pollen dispersal distance was higher than in 2003 for
both normal and exponential models. For the exponential model,
in 2002, the observed pollen dispersal was 444 m and in 2003 it
was 154 m. Due to the fact that the distance of pollen dispersal was
higher in 2002, the effective pollination neighborhood area, esti-
mated as a circular area around seed-trees, was also higher in this year
(Table 2).
VARIATION AMONG YEARS IN MATING SYSTEM PARAMETERS.—In gen-
eral, we did not find great differences in mating system parameters
between years. The outcrossing rates, mating among relatives, cor-
related mating, and differentiation in pollen pool among seed-trees
were similar and not statistically different among years (Table 1).
Low variation of outcrossing rates between two different years has
also been reported for the tropical trees Pithecellobium elegans (Hall
et al. 1996) and Dipterocarpus tempehes (Kenta et al. 2004) (P. el-
egans: tˆ(1991) = 0.97, tˆ(1992) = 0.92; D. tempehes: tˆ(1996) = 0.93,
tˆ(1998) = 0.96). However, other studies have detected significant

TABLE 2. Density of reproductive trees (d), mean pollen dispersal distance (δ), and pollination neighborhood area (Aep ) estimated for the normal and exponential dispersal
model (TWOGENER). The error is a quadratic criterion for the fit between expected and observed values for pairwise  i j estimates (Austerlitz & Smouse 2002).

Dispersal Density Year 2002 Year 2003

Function contrasting d (trees/ha) δ̂ (m) error  e p d (trees/ha) δ̂ (m) error  e p

Normal fixed 0.46 103 6.64 3.3 0.46 88 0.87 2.4

fixed 0.10 216 6.60 14.6 0.10 186 1.15 10.8
estimated 0.05 296 6.60 27.4 0.23 101 1.19 3.2
Exponential fixed 0.46 115 6.62 4.1 0.46 99 0.87 3.1
fixed 0.10 243 6.59 18.5 0.10 210 0.87 13.8
estimated 0.03 444 6.55 61.7 0.19 154 0.86 7.4

differences among years (Murawski et al. 1990, Hall et al. 1996,
Caron et al. 1998, Rocha & Aguilar 2001, Hufford & Hamrick
2003). For example, Murawski et al. (1990) found for the tropical
tree Cavanillesia platanifolia, multilocus outcrossing rates of 0.569
in 1987 and 0.347 in 1988. Rocha and Aguilar (2001) studied two
consecutive reproductive events in a population of Enterolobium cy-
clocarpum in Costa Rica and observed multilocus outcrossing rates
of 0.812 and 0.924, and paternity correlations of 0.462 and 0.174,
in 1994 and 1995, respectively. Hufford and Hamrick (2003) de-
tected variation in individual outcrossing rates among two repro-
ductive events of Platypodium elegans at Barro Colorado Island (for
example, tree 6: tˆ(1998) = 0.88, tˆ(1999) = 0.96). Thus, S. globu-
lifera did not show high interannual variation in mating system
parameters. One explanation for low differences between years for
outcrossing rates is that S. globulifera might have a ‘cryptic self-
incompatibility’ mechanism, ensuring consistent high outcrossing
levels.
VARIATION AMONG REGIONS.—Our results are in contrast to the
initial hypothesis that the low density of reproductive individu-
als in Amazonian Forest would increase the self-fertilization rate.
The multilocus outcrossing rate estimates in this study (minimum
0.982) were higher than the reported values in previous studies for
the same species in other populations. Aldrich and Hamrick (1998)
estimated an outcrossing rate of 0.920 in a continuous forest, 0.886
in fragmented forests, and 0.731 for pasture trees, all occurring
in low densities of reproductive individuals in Costa Rica. Degen
et al. (2004) estimated an outcrossing rate of 0.920 for a high den-
sity, partially logged population of S. globulifera in French Guiana.
The results obtained for the population at Tapajós represent mat-
ing under undisturbed, low-density conditions for S. globulifera.
Independent from the tree densities we see a tendency of increasing
self-fertilization with increasing level of disturbance (undisturbed
population in Brazil > partly logged population in French Guiana
> fragmented population in Costa Rica). These results also suggest
that the S. globulifera may have a ‘cryptic self-incompatibility sys-
tem,’ a phenomenon that has been observed in other tropical tree
species. For example, P. quinata, C. pentandra, and Samanea saman
occurring in continuous forest are self-incompatible, but in frag-
mented populations the species showed a partially self-incompatible
system, and the mating system changed to mixed mating (Cascante
et al. 2002, Fuchs et al. 2003, Quesada et al. 2004, Lobo et al. 2005).
This mating system variation has been attributed to the absence of
pollinators and alteration in population density by fragmentation.
Possibly, a flexible mating system may occur in S. globulifera and un-
der a logging regime, if pollinator behavior has changed, the species
may be able to change mating system and spatially isolated trees
produce seeds by selfing. Moreover, it is possible that the number of
pollen donors involved in seed fertilization was reduced, increasing
the relatedness within families, as detected by Aldrich and Hamrick
(1998) comparing the mating system of S. globulifera in trees in
continuous forest and isolated in forest fragments. They observed
that the majority of the seeds of S. globulifera trees in fragmented
forest were fertilized by a few trees located in pasture.
Mating System and Gene Flow in Symphonia globulifera 633
BIPARENTAL INBREEDING.—The results indicated a significant rate
of mating among relatives in both reproductive events, but low
variation between years (Table 1). The mating among relatives can
be explained by observed spatial genetic structure in the popula-
tion (Fig. 2) and limited pollen dispersal distance. The scale and
strength of the observed spatial genetic structure were not expected
because its known seed dispersers (bats and monkeys) are capable
of transporting seeds for long distances (Dunphy et al. 2004). Still,
the significant fixation index detected in sampled seeds from both
years can be explained by mating among relatives. A mixture of mat-
ings among first cousins (θ p = 0.0625) and half-sibs (θ p = 0.125)
could explain the observed levels of inbreeding. Similar results were
observed by Degen et al. (2004) in French Guiana. The authors
detected high rates of biparental inbreeding (tˆm − tˆs = 0.156) and
significant spatial genetic structure up to 150 m.
For many tree species, higher estimates of fixation index have
been detected in progeny compared to adult trees (see Murawski
1995). The general explanation is that selection against inbred
progeny occurring in early ontogenetic stages reduces the excess
of homozygotes in adult population (Hufford & Hamrick 2003).
But our fixation index for adult trees was positive and bigger than
the fixation index of the seeds from 2002. However, the high fix-
ation index in adult trees could also be due to a Wahlund effect,
if the population is structured in different, reproductively isolated
subpopulations.
CORRELATED MATING.—A similar high proportion of correlated
matings between years in the present population was observed, in-
dicating that part of seeds are full-sibs (Table 1). Correlated mating
can be caused by the behavior of the pollinator, insufficient flow-
ering synchrony, low numbers of reproductive trees, and limited
population size (Lee 2000). However, the study population is large
and there are many reproductive trees outside the plot, so small
population size can be discarded as a potential explanation. All
other factors could have contributed to the high levels of correlated
mating. However, correlated mating in tropical tree species seems
to be a common phenomenon and has been detected in many
species (see Table 3). Thus, open-pollinated seeds usually comprise
mixtures of progeny with different degrees of relatedness. In this
case, for S. globulifera, we found a high proportion of half-sibs
( P̂ H S(2002) = (1 − tˆm )r̂ p = 0.586, P̂ H S(2003) = 0.614), but also
a significant proportion of full-sibs (minimum 38.6%). The mean
coancestry coefficient within progeny was higher than the theo-
retical value of 0.125 expected in half-sib families. This result has
important implications for seeds collected for ex situ conservation,
tree breeding, and reforestation. The high coancestry coefficient
within families requires higher sample size in terms of the number
of families in order to avoid genetic drift due to low effective popu-
lation size. The same conclusion can be drawn from the low values
of variance effective population size (N e (v) ).
DIFFERENTIATION IN POLLEN POOL.—The levels of pollen pool dif-
ferentiation in both reproductive years (Table 2) was lower than
detected in a high-density population of the same species in French
Guiana ( ˆ ft = 0.205, Table 3). But the values of pollen pool
634 Carneiro, Sebbenn, Kanashiro, and Degen

TABLE 3. Density of reproductive trees (d), analysis method ( TWOGENER and MLTR), paternity correlation (r p ), differentiation in pollen pool ( f t ), number of pollen
donors (Nep ), and effective pollination neighborhood area (Aep ) for some tropical tree species

Species Forest d (tree/ha) Method r̂ p ˆ ft

 N̂ e p  e p b (ha) Reference

Carinana legalis Fragmented 1.57 MLTR 0.324 – 3.1 2.0 Sebbenn et al. (2000)
Carinana legalis Fragmented 0.25 MLTR 0.295 – 3.4 13.6 Sebbenn et al. (2000)
Carinana legalis Fragmented 1.80 MLTR 0.212 – 4.7 2.6 Sebbenn et al. (2000)
Ceiba pentandra Fragmented 0.01 MLTR 0.370 – 2.7 270.3 Lobo et al. (2005)
Ceiba pentandra Fragmented 0.03 MLTR 0.600 – 1.7 55.6 Lobo et al. (2005)
Dinizia excelsa Continuous 0.17 TWOGENER – 0.104 4.8 28.3 Dick et al. (2003a)
Dinizia excelsa Fragmented 0.10 TWOGENER – 0.002 299.4 2,994.0 Dick et al. (2003a)
Hevea brasiliensis Continuous 1.00 MLTR 0.351 – 2.8 2.8 Furlani et al. (2005)
Myracrodruon urundeuva Fragmented 0.50 MLTR 0.371 – 2.7 5.4 Moraes et al. (2004)
Myracrodruon urundeuva Fragmented 0.60 MLTR 0.671 – 1.5 2.5 Moraes et al. 2004)
Pachira quinata Continuous > 20.0 MLTR 0.470 – 2.13 0.1 Fuchs et al. (2003)
Pachira quinata Isolated trees 0.01a MLTR 0.740 – 1.35 13.5 Fuchs et al. (2003)
Samanea saman Continuous < 10.0 MLTR 0.027 – 37.0 3.7 Cascante et al. (2002)
Samanea samana Isolated trees 0.01a MLTR 0.120 – 8.3 833.2 Cascante et al. (2002)
Symphonia globulifera Continuous 4.00 TWOGENER – 0.205 2.4 0.6 Degen et al. (2004)
Sextonia rubra Continuous 1.50 TWOGENER – 0.061 8.2 5.5 Veron et al. (2005)

a Isolated tree at least 500 m, dˆ = 1/0.0001.π (500)2 = 0.01. b  e p = N̂ e p /d .

differentiation detected in S. globulifera were generally higher than
estimates in other animal-pollinated tropical trees species (Dizinia
excelsa, ˆ ft = 0.002 − 0.104, Dick et al. 2003a, Sextonia rubra,
ˆ
ft = 0.061, Veron et al. 2005), and wind-pollinated tree species
(Quercus alba,  ˆ ft = 0.036, Smouse et al. 2001; Pinus echinata,
ˆ
ft = 0.095, Dyer and Sork 2001; Quercus lobata,  ˆ ft = 0.136,
ˆ
Sork et al. 2002; Pinus sylvestris, ft = −0.006 − 0.007, Robledo-
Arnuncio et al. 2004). A lower synchronization among flowering
trees or the behavior of pollinators may explain the higher values of
S. globulifera. The high differentiation in pollen pool, as with high
paternity correlation ( ˆ ft = r̂ p /2, Hardy et al. 2004), reduces the
genetic variation within families and increases it among families. As
the pollen pool differentiation and the paternity correlation were
high in both reproductive years, it suggests that a low number of
effective pollen donors mating with each seed-tree (Table 1). These
results are similar to the values reported for another population of
S. globulifera in French Guiana and many other tropical tree species
(see Table 3).
The number of pollen donors estimated from paternity correla-
tion (Ritland 1998) was lower than estimated from differentiation
in pollen, although only the estimates obtained from 2002 were
statistically different. This difference can be attributed to the sta-
tistical approaches used to calculate the Nep parameter. Ritlands
correlated mating model uses a maximum-likelihood method to es-
timate paternity correlation and therefore better exploits multilocus
information (Hardy et al. 2004). The differentiation of pollen pools
is a measure of paternal genetic resemblance within families relative
to that among families (intraclass correlation coefficient). Hence,
we can expect differences between the estimates of the number of
pollen donors by different methods.
EFFECTIVE POLLEN DISPERSAL.—The reduction in the estimated re-
productive tree density in all analyses in relation to fixed density,
suggested that only part of the adult reproductive population par-
ticipated in the reproductive event. In fact, we observed that some
trees produced no seeds in either of the 2 yr, and others only in
1 yr. The results also showed that the density was more reduced
in 2002 than 2003, indicating an annual variation in the number
of reproductive individuals. However, although this alteration did
not change mating system parameters, as already demonstrated, the
pollen dispersal distance was affected (Table 2). The estimated den-
sity of reproductive trees per hectare was lower in 2002 than 2003
and distance of pollen dispersal was higher in 2002 than 2003 for
both normal and exponential models. Pollen dispersal appears to
be negatively correlated with the tree density (Dyers & Sork 2001,
Degen et al. 2004, Veron et al. 2005, Ward et al. 2005). When the
density of reproductive trees is reduced, both the distance among
reproductive trees and the distance of pollen dispersal distance in-
crease. For example, in comparison to fixed density of adult trees
(0.46), the estimated density of reproductive trees in 2002 was
smaller (0.03) than in 2003 (0.19) and, consequently, the distance
of pollen dispersal was 2.9 times bigger in 2002 (444/154). The in-
crease in the distance among reproductive trees forces the pollinator
to travel longer distances between trees to obtain nectar, increasing
the distance of pollen dispersal. However, this apparently does not
affect the number of pollen donors between years (Table 1).
We found a longer pollen dispersal distance in the plot
(Table 2) than had previously been reported. The estimated values
for the effective pollen dispersal distance (δ) in Tapajós were higher
than those reported by Degen et al. (2004) in French Guiana, who
estimated a mean pollen dispersal of 27–53 m. The higher density

of reproductive individuals per hectare of S. globulifera in French
Guiana (10.9 trees/ha) compared to Tapajós National Forest (0.46
trees/ha) is the most likely cause. The pollen dispersal distance of
S. globulifera in Tapajós National Forest is also higher than values
observed in other tree species with low tree density. For example,
Veron et al. (2005) detected a mean pollen dispersal between 65 m
and 89 m, for S. rubra (an insect-pollinated tropical tree species,
with a density of 1.5 trees/ha). Sork et al. (2002) found a mean
pollen dispersal of 64.8 m for wind-pollinated Q. lobata with a
density of 1.19 trees/ha. Higher values were observed by Dick et al.
(2003a) in the Amazon Forest for the insect-pollinated tropical tree
Dinizia excelsa. They found a mean pollen dispersal of 1509 m in
a fragmented landscape and 212 m in an undisturbed forest with
a density of 0.3 trees/ha. Our results support the hypothesis that
pollen dispersal is negatively correlated with the tree density.
CONCLUSION FOR FOREST MANAGEMENT.—Symphonia globulifera is
logged in the Brazilian Amazon Forest for timber production. Con-
sidering that the present analyses of mating system and gene flow
showed interannual variation in only the density of reproductive
trees and pollen dispersal distance, and these changes did not affect
the outcrossing rate and level of correlated mating, we believe that
if the intensity of forest exploitation was not stronger than, for ex-
ample, 40 percent of the reproductive tree, the mating system will
not be negatively affected. A study comparing the mating system
before and after the logging may elucidate this supposition.
Finally, information about the effective pollination neighbor-
hood area is important for the development of sustainable forest
logging plans. This value gives an idea about the size of reproduc-
tive population and thus an indication on critical limits to forest
fragmentation. The effective pollination neighborhood area was es-
timated to reach 61.7 ha. This value is higher than that calculated
for other tropical tree species with similar tree densities and repre-
sents an intermediate value in general (Table 3). Interpreting this
result in terms of forest management, the observed reproductive
neighborhood area suggests that logging will not isolate trees, if the
intensity of the S. globulifera logging is not higher than 50 percent.
Considering that the estimated density of reproductive trees per
hectare ranges from 0.03 (one tree per each 33 ha) to 0.23 (one tree
per each 4 ha), and in the study plot, there are two trees in every 4
ha, we can expect that the logging of one of these two trees will not
isolate the remaining trees.
ACKNOWLEDGMENTS
The study was financially supported by grants from DFID UK
and EMBRAPA Brazil “Dendrogene Project,” by a grant for re-
gional co-operation between French Guiana and Brazil “Forma-
tion supérieure et diffusion des résultats de la recherche pour la
gestion durable des forêts de Guyane Française et d’Amazonie
Brésilienne” as well as by the EU-INCO project “Distribution
of genetic diversity in tree species from the Neotropics based on
DNA fingerprinting assays: Implications for conservation, sustain-
able utilization and management” (contract number: ICA4-CT-
Mating System and Gene Flow in Symphonia globulifera 635
2001-10101, http://thoth.nbu.ac.uk/geneo). The author A.M.S
would like to thank the Conselho National of Desenvolvimento
Cientı́fico e Tecnológico (CNPq) for granting him a scholarship
for postdoctoral studies at the Bundesforschungsanstalt für Forst-
und Holzwirtschaft (BHF) in Germany. We thank Eric Bandou for
his excellent technical assistance in the lab. We are very grateful
to Mauricio Quesada, Stephen Cavers and three anonymous refer-
ees for helpful comments and constructive suggestions on a former
version of the manuscript.
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