Morphological variability in rupicolous species of the Acianthera prolifera complex (Orchidaceae) occurring in southeastern Brazil

We carried out multivariate morphometric analysis of 23 floral characters in seven populations of a complex of four species of Acianthera (Orchidaceae) occurring in Brazilian campo rupestre (rocky field) vegetation (A.?hamosa, A.?limae, A.?modestissima, and A.?prolifera) that flower synchronously and are partially intercompatible, and one putative hybrid population between A.?limae and A.?prolifera. We also carried out cluster analysis involving these eight populations plus 21 populations of a previously published study belonging to another species complex of Acianthera occurring in campo rupestre, including 12 floral characters in the analysis. Allopatric species pollinated by the same group of Diptera showed higher floral similarity among themselves than to a sympatric species pollinated by another group of Diptera. Such patterns indicate the existence of floral convergence in allopatric species and/or radiation in sympatric species. The analysis also indicated that there is more floral similarity between species of different complexes but that share the same group of pollinators. Large overlap was observed between A.?limae and the putative sympatric hybrids, indicating the occurrence of later generations of hybrids and/or individuals of A.?limae with introgression. The results do not support A.?hamosa and A.?modestissima as distinct species. These taxa are geographically isolated, occurring in different environments, are recognized only by vegetative characters that show high phenotypic plasticity, and share the same pollinators, being interfertile.     

Reproductive isolation in the Aegean Ophrys omegaifera complex (Orchidaceae)

The orchid genus Ophrys operates a system of sexual deception by which high specificity of pollination is attained. Reproductive isolation in Ophrys mainly rests upon prezygotic isolation mechanisms. The level of genetic separateness of Ophrys taxa with different pollinators is therefore likely determined by the fidelity of pollinators. The present study employs genetic fingerprinting to investigate this in the east Aegean Ophrys omegaifera s.l. complex, also including O. dryis, a west Mediterranean species of this complex. Ophrys fleischmannii, O. basilissa, and the west Mediterranean O. dyris, are found to be well-separated genetic entities whereas O. omegaifera s.str. and the putative hybrid taxon, O. sitiaca, are found to be genetically inseparable across their entire range of co-occurrence. This suggests that specific pollinators have high enough fidelity to act as effective isolating factors in east Aegean O. omegaifera s.l. as a whole, but that the situation in the species pair of O. sitiaca and O. omegaifera is likely to be more complex.     

Morphometric analysis of Epidendrum secundum (Orchidaceae) in southeastern Brazil

One of the largest genera of Orchidaceae with about 1125 species, Epidendrum L. presents several taxonomic problems, mainly due to the great variability of some of its species. The present study aims at evaluating the influence of different environments on the morphology of individuals from Epidendrum secundum , using morphometry to compare plants under cultivation and plants collected directly from the field. Eighty specimens maintained in cultivation at the Institute of Botany of São Paulo (Brazil) and 146 individuals originating from three natural populations, totaling 226 specimens, were analyzed with multivariate methods (PCA, CVA, DA). The fact that plants growing in rocky outcrops and plants from the Atlantic rainforest showed the largest morphological differentiation among themselves, indicates that phenotypes are strongly influenced by the habitat. Because plants collected from both habitats, maintained under cultivation for at least 20 years maintained their differentiation, such differentiation may have a genetic component. Because individuals collected in such environments present the largest morphological dimensions, new habitats generated by human activity, such as highway margins, seem to be quite favorable for these plants.     

Reproductive biology of Astyanax fasciatus (Pisces: Characiformes) in a reservoir in southeastern Brazil

To study the reproductive biology of Astyanax fasciatus (Cuvier, 1819) in Furnas Reservoir, Minas Gerais, Brazil, a total of 1307 specimens were captured in bimonthly sampling from August 1993 to July 1994. The sex ratio showed a predominance of females, representing 65.2% of the total sample. The stages of gonadal maturation and spawning type were determined through macro- and microscopic analysis of the gonads as well as by variation in the gonadosomatic indices. A . fasciatus showed fractionated spawning and reproduction throughout the year, with spawning peaks influenced by water temperature and rainfall. Males and females initiated gonadal maturation at a mean standard length of 6.4 and 6.5 cm, respectively. During gonadal maturation, females showed a reduction in hepatosomatic index (from 1.06 to 0.83), suggesting that hepatic substances are transferred from the liver to the ovaries. In the reproductive period, A. fasciatus had lower stomach repletion and coelomic fat indices, indicating that this species ate less and consumed more fat reserves when reproduction had the maximum GSI (11.3 for females and 2.7 for males). Weight of the ovaries had a positive influence on the condition factor (K1), as the lowest K2 value (weight without gonads) were obtained during maturation, confirming the influence of the ovaries upon the physiological condition of the females. In weight–length relationship, parameter b was 3.02 for females and 2.23 for males, and the correlation coefficient ( r ²) was 0.84 and 0.66, respectively.     

Reproductive biology of two tree species of Leguminosae in a Montane Rain Forest in southeastern Brazil

Studies on sympatric species that belong to the same taxonomic group and have the same pattern of pollination and seed dispersion, but different patterns of geographical distribution and structural parameters may be worthwhile to understand the evolution of their compatibility system. We studied the floral biology, pollinators and reproductive systems of two tree species in Itatiaia, an area covered with montane rain forest. Pseudopiptadenia contorta is widespread in Brazil but is present at a low relative density at the study site, while Pseudopiptadenia leptostachya is restrictively distributed but presents one of its highest relative density values in Itatiaia. Both species have small flowers arranged in compact spicate raceme inflorescences that constitute the pollination unit. Pollinators are native stingless bees and European honeybees that collect pollen from their nectarless flowers. Although both species have similar floral characteristics and pollination system, their compatibility system differs. Fruit set after manual pollinations and observation of the pollen tube growth showed that P. leptostachya is self-compatible while P. contorta only set fruits after cross-pollinations, and is thus a self-incompatible species. However, fruit set did not differ between the species, indicating that structural parameters are not affected by the compatibility system. Thus, the difference between these two species in terms of reproductive system may be related to other factors such as patterns of geographical distribution.     

Reproductive versus floral isolation among morphologically similar Serapias L. species (Orchidaceae)

Flowers of the Mediterranean orchid genus Serapias L. form small, dark tubes that vary among taxa in diameter and depth. Visiting insects use the floral tube as shelter and act as pollinators if they touch the sticky viscidium at the rear of the tube and remove the pollinarium. It has been assumed that floral tube size and shape limit access to the flowers and thus may act as a barrier to gene flow between different Serapias species. Here we investigated floral characters and nuclear microsatellite markers in populations belonging to three morphologically similar Serapias species to test whether these species show evidence for floral or reproductive isolation. We found strong overlap of floral traits between two species, suggesting that floral isolation is nonexistent between them. Microsatellite markers applied to the same populations were highly polymorphic and revealed clear genetic differentiation among all three species. These results suggest that reproductive isolation exists, despite the lack of floral isolation between two of the species. In contrast to morphological characters, diagnostic microsatellite alleles were found for all Serapias species. The microsatellite markers could thus provide a useful tool to identify Serapias species and further investigate evolutionary relationships in this fascinating orchid lineage.     

Reproductive biology and conservation genetics ofSerapias vomerace (Orchidaceae)

Serapias vomeracea is considered to be a particular example of the deceptive strategy of terrestrial orchids. Information on its reproductive biology and population genetic variation is needed to develop conservation strategies. The pollination biology and breeding system were investigated in four populations ofS. vomeracea. Hand-pollination experiments showed thatS. vomeracea is self-compatible. Bagged inflorescence produced no fruits; artificial pollination resulted in 89.7–100% fruit set through induced autogamy, 91.1–100% with geitonogamous, and 89.3–94.4% following xenogamous pollination. Fruit set in the open-pollinated control ranged from 13.4 to 20.9%. We evaluated the spatial genetic structure of 13 populations using five nuclear microsatellite loci. Italian and isle of Corfu populations ofS. vomeracea are characterized by a low mean allele number, ranging from 2.4 to 3.8, by intermediate to high heterozygosity levels, ranging from 0.50 to 0.71, and by frequent deviations from the Hardy-Weinberg equilibrium (9 out 13 populations). Most of theS. vomeracea populations exhibited significant heterozygote excess. The population size reduction is most probably the cause of the low genetic variation observed in the populations ofS. vomeracea. For the genetic conservation and management ofS. vomeracea suitable strategies should be developed based on these data.     

Reproductive biology and conservation genetics of Goodyera procera (Orchidaceae)

Goodyera procera is an endangered terrestrial orchid in Hong Kong. Information on its reproductive biology and pattern of genetic variation is needed to develop efficient conservation strategies. Pollination experiments showed that the species is self-compatible, but dependent on pollinators for fruit set. Bagged plants produced no fruits. Artificial pollinations resulted in 92% fruit set through selfing, 94% with geitonogamous pollination, and 95% following xenogamous pollination. Fruit set in the open-pollinated control was 75% at the same sites. Allozyme electrophoresis and random amplified polymorphic DNA (RAPD) were used to evaluate genetic variation and structure of 15 populations of Goodyera procera. Despite its outbreeding system, allozyme data revealed low variation both at the population (P = 21.78%, A = 1.22, and H = 0.073) and species (P = 33%, A = 1.33, and H = 0.15) levels, in comparison with other animal-pollinated outbreeding plant species. However, RAPD variation was relatively high (P = 55.13% and H = 0.18 at the population level, and P = 97.03% and H = 0.29 at the species level). G_ST estimates indicated high levels of genetic differentiation among populations (G_ST = 0.52 and I = 0.909 ± 0.049 based on allozyme data, and G_ST = 0.39 and I = 0.859 ± 0.038 based on RAPD data), much above the average for outcrossing species, suggesting that gene flow was limited in this species. Based on these data, suitable strategies were developed for the genetic conservation and management of the species.     

Reproductive isolation mechanisms among four closely-related species of Conospermum (Proteaceae)

The effectiveness of geographical isolation, ecological isolation, temporal isolation, mechanical isolation, ethological isolation, cross-incompatibility, hybrid inviability, hybrid sterility and hybrid breakdown as practical barriers to gene flow in the field between Conospermum taxifolium, C. ericifolium, C. ellipticum and C. longifolium has been quantified. The barriers to gene flow between C. ericifolium and C. ellipticum are completely effective, owing to their allopatric distributions. The barriers to gene flow between C. taxifolium and these two species are only partially effective, as their ecological separation breaks down in intermediate habitats, and partially-fertile F₁ plants can grow in the areas of overlap. The barriers to gene flow between C. longifolium and the other three species are almost completely effective, as cross-incompatibility is very high and the F₁ plants are female-sterile.     

Reproductive biology and pollination of southeastern Brazilian Stanhopea Frost ex Hook. (Orchidaceae)

Reproductive biology and pollination of Stanhopea lietzei and Stanhopea insignis were studied in a semi-deciduous mesophytic forest in the Serra do Japi (SJ), and in the coastal plain of Picinguaba, both in the State of São Paulo, Brazil. Floral morphology, pollination, breeding system and fruit set of both species were investigated. S. lietzei and S. insignis are pollinator-specific, being pollinated by male bees of Eufriesea (Apidae, Euglossini), which collect the fragrance produced by pluricellular osmophores at the base of the saccate hypochile. S. lietzei and S. insignis were pollinated by Eufriesea pulchra and Eufriesea purpurata, respectively. Observations using substances present in the floral fragrance of both studied species as chemical baits were also performed. E. purpurata was attracted by benzyl alcohol, the major compound of the perfume of S. insignis, while E. pulchra was attracted by none of the compounds used. Both studied Stanhopea are self-compatible but pollinator dependent. Self-pollination, however, tends to be avoided by floral mechanisms. In experimental self- and cross-pollinations the proportion of fruit abortion was high and related to resource limitation. The reproductive success of S. lietzei and S. insignis was low as a consequence of deficient pollen transference while pollinator scarcity was the main factor.     

Isolation mechanisms between two sympatric Sophronitis (Orchidaceae) species endemic to Northeastern Brazil

Sophronitis sincorana and S. pfisteri (Orchidaceae) are endemic to the campo rupestre vegetation, northeastern Brazil, occurring sympatrically and flowering synchronously. In this work we studied the reproductive biology of both species and investigated reproductive barriers between them. Both species are partially self-compatible, pollinator dependent, and are pollinated by males and worker bees of Bombus brevivillus. Pollinaria are deposited on the anterior portion of the mesonotum of these insects in S. pfisteri, and on the scutellum in S. sincorana. The flowers of both species are deceptive, as they have a dry cuniculus. Morphological differences in the labellum and the gynostemium impede inter-specific pollination of S. pfisteri with pollen from S. sincorana. Very low fruiting results from inter-specific crosses of S. sincorana with pollen from S. pfisteri. Although viable seeds can be produced from experimental inter-specific crosses, no natural hybrids were found. Species integrity appears to be maintained by additive unidirectional reproductive barriers between them.     

稻蝗属三近缘种间的生殖隔离机制及其进化意义

通过对日本稻蝗、中华稻蝗台湾亚种和小翅稻蝗的种间交配、交配后精子传送等的研究,探讨了其生殖隔离机制及其进化意义。结果表明： 有共同分布区域的日本稻蝗与中华稻蝗台湾亚种及日本稻蝗与小翅稻蝗的交配率显著低于种内交配,即使交配也无精子的传送,存在强烈的行为隔离和完全的机械隔离。分布不重叠的中华稻蝗台湾亚种与小翅稻蝗之间,小翅稻蝗的雌虫与中华稻蝗台湾亚种雄虫的交配率显著低于种内交配,反向交配时则和种内交配率无显著差异;正反交配都能完成精子传送,显示出不对称、不完全的行为隔离,而无机械隔离的存在。分布不重叠的中华稻蝗台湾亚种与小翅稻蝗生殖隔离的进化速度慢于分布重叠的日本稻蝗与中华稻蝗台湾亚种及日本稻蝗与小翅稻蝗。     

Phenology and pollination biology of eight Peperomia species (Piperaceae) in semideciduous forests in southeastern Brazil

Data on flowering phenology and pollination of Peperomia species are virtually non-existent. This study presents data on the pollination biology of eight Peperomia species from south-eastern Brazil, including the flowering phenology, pollination system, and reproductive success. Data on flowering phenology were recorded weekly and exclusion experiments on inflorescences provided data on autonomous self- and wind pollination. Direct visual observations were made and insect visits were recorded. Four Peperomia species showed continuous flowering, while the others were seasonal and flowered in the wet season. Pollination by wind and Syrphidae was confirmed for two self-incompatible Peperomia species. The remaining species are self-compatible and their high fruit set may be accounted for by autonomous self-pollination and perhaps agamospermy. Although the floral morphology of Peperomia species suggests wind- and/or insect pollination, most of the species studied exhibit autogamy and perhaps agamospermy as the main method of reproduction.     

Pollination biology of four sympatric species of Habenaria (Orchidaceae: Orchidinae) from southern Brazil

The pollination process and breeding system of the sympatric Habenaria johannensis, H. macronectar, H. megapotamensis and H. montevidensis was documented for native populations from Rio Grande do Sul, Brazil. All species investigated offer a nectar reward (mean values of total sugars ranging from 18 to 26%) concealed in a spur. Habenaria montevidensis is pollinated by butterflies (Hesperiidae, especially of the genus Urbanus) that carry pollinaria on their eyes; the other three species are pollinated by Sphingidae. Habenaria johannensis is pollinated by the moths Manduca rustica and M. sexta that carry the pollinaria at the base of the proboscis. Habenaria macronectar is pollinated by the moths Eumorpha labrusca and M. cf. lucetius, and these bear pollinaria between the palpi. Habenaria megapotamensis is pollinated by moths of M. cf. lucetius that bear the pollinaria on the proboscis. All species studied are self‐compatible, but pollinator dependent. They also displayed high reproductive success (ranging from 69.48 to 97.40%) and male efficiency factors slightly higher than 1, suggesting that at least one flower was pollinated for each flower acting as pollen donor. At the study sites, the investigated Habenaria spp. are isolated (in terms of pollination) by a set of factors that includes differing floral morphologies, different pollinators and/or different pollinarium placement on the pollinator. © 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2012, ??, ??–??.     

Reproductive isolation and hybridization in sympatric populations of three Dactylorhiza species (Orchidaceae) with different ploidy levels

Background and Aims

The potential for gene exchange between species with different ploidy levels has long been recognized, but only a few studies have tested this hypothesis in situ and most of them focused on not more than two co-occurring species. In this study, we examined hybridization patterns in two sites containing three species of the genus Dactylorhiza (diploid D. incarnata and D. fuchsii and their allotetraploid derivative D. praetermissa).

Methods

To compare the strength of reproductive barriers between diploid species, and between diploid and tetraploid species, crossing experiments were combined with morphometric and molecular analyses using amplified fragment length polymorphism markers, whereas flow cytometric analyses were used to verify the hybrid origin of putative hybrids.

Key Results

In both sites, extensive hybridization was observed, indicating that gene flow between species is possible within the investigated populations. Bayesian assignment analyses indicated that the majority of hybrids were F₁ hybrids, but in some cases triple hybrids (hybrids with three species as parents) were observed, suggesting secondary gene flow. Crossing experiments showed that only crosses between pure species yielded a high percentage of viable seeds. When hybrids were involved as either pollen-receptor or pollen-donor, almost no viable seeds were formed, indicating strong post-zygotic reproductive isolation and high sterility.

Conclusions

Strong post-mating reproductive barriers prevent local breakdown of species boundaries in Dactylorhiza despite frequent hybridization between parental species. However, the presence of triple hybrids indicates that in some cases hybridization may extend the F₁ generation.     

Reproductive biology of the armoured catfish Loricariichthys castaneus (Castelnau, 1855) in Lajes reservoir,southeastern Brazil

Sex ratio, gonadal development, breeding season and fecundity of the armoured catfish Loricariichthys castaneus were described to assess its reproductive strategy in a Brazilian tropical reservoir. In total, 226 specimens (199 females and 27 males) were captured from September 2005 to August 2006 and examined in the laboratory. Females outnumbered males and achieved sizes larger than 330 mm TL. Oocyte development, determined by histological analysis, was asynchronous with oocyte size, ranging from pre‐spawning (27–270 μm) to spawning (243–3460 μm), followed by a sharp decrease in the mean oocyte diameter postspawning (590–730 μm) as the spawning proceeded. Spawning occurred throughout most of the year, peaking in August–September and reaching a low in April–May, according to variations in GSI and frequencies of stages of gonadal development. Batch fecundity ranged from 242 to 833 vitellogenic oocytes (relative fecundity = 2.27 oocytes g^?1), averaging 483 oocytes, and was positively related to gonad weight (P = 0.00003). Oocyte diameters ranged from 0.027 to 5.59 mm, with vitellogenic diameters ranging from 2.08 to 5.59 mm. Continuous development of oocytes throughout the year suggests that L. castaneus presents indeterminate fecundity and is a batch‐spawner. These attributes, associated with parental care and a wide reproductive period, correspond to an equilibrium strategy that has proved to be effective in the Lajes reservoir.     

Reproductive biology of the rock shrimp Sicyonia dorsalis (Decapoda: Penaeoidea) from the southeastern coast of Brazil

Summary

Sicyonia dorsalis is not commercially exploited in Brazil, but it forms a critical link in marine food chains by converting detritus food sources into microorganism biomass that is available for higher trophic levels. We examined the reproductive biology of this species, sampled for five and a half years in a tropical locality, the southeastern coast of São Paulo (23° S), Brazil. Monthly samples were taken from January 1998 to June 2003 at depths of 5 to 45 m. Degree of ovarian development was used to examine breeding in adult females. Recruitment was estimated by changes in proportion of individuals in juvenile size classes of carapace length (CL #8.2 mm) in relationship to the total number of individuals. The largest immature female measured 9.2 mm CL, but 75% of females were smaller than 7.3 mm CL. The largest immature male measured 6.8 mm CL. The relative frequency of reproductive females was negatively correlated with the relative frequency of recruitment (p <0.001). Reproductive females were found in every season, with peaks in December-February following the recruitment peak in March-June, and a second peak of spawning in May-July and of recruitment in September-November. The continuous reproductive pattern observed for S. dorsalis, combined with the hypothesis that during spring and summer the phytoplankton production is higher, when the South Atlantic central water mass intrudes into the region during upwelling events, suggests that food availability for protozoeal and mysis larvae may be an important selective factor shaping the seasonal breeding pattern for S. dorsalis. The classical paradigm of continuous reproduction at lower latitudes, with increased seasonality of the breeding period at higher latitudes, seems to be valid for this species.     

Reproductive biology of Epidendrum tridactylum (Orchidaceae: Epidendroideae): a reward-producing species and its deceptive flowers

Epidendrum L. (Epidendroideae, Laeliinae) is one of the largest orchid genera distributed throughout tropical America. Based on data on reproductive phenology, floral morphology, labellar histochemistry, pollinators, pollination mechanisms and breeding system, this study explored the reproductive biology of Epidendrum tridactylum Lindl. growing in the state of São Paulo, southeastern Brazil. Epidendrum tridactylum produces an average of 87 rewardless flowers with a longitudinal protuberance on the median portion of its labellum, whose epidermal cells produce a pleasant citric fragrance. This fragrance attracts both male and female flies of families Calliphoridae, Sarcophagidae and Syrphidae that end up drinking the extrafloral nectar produced at the base of the bracts. Pollinaria are removed when the fly proboscis contacts the viscidium in search of nectar. After removing the pollinarium, the fly gets trapped in the flower, which frightens it, thus inhibiting any possible intent to immediately visit another flower and, at least presumably, reducing the possibility of geitonogamy. Since E. tridactylum is self-incompatible, the positive consequence of such behavior is a reduction of pollen loss. The fruits obtained through experimental cross-pollinations present highly variable percentages of potentially viable seeds. This is the first comprehensive study on the reproductive biology of a myophilous Epidendrum and a pioneer report of effective pollinators picking extrafloral nectar from Orchidaceae.