Effects of habitat suitability for vectors, environmental factors and host characteristics on the spatial distribution of the diversity and prevalence of haemosporidians in waterbirds from three Brazilian wetlands.

BACKGROUND: Wetlands are ecosystems in which vectors of avian haemosporidians live and reproduce and where waterbirds join to breed in colonies. Brazil has wetlands at different latitudes, which enables testing the influence of the ecological factors on the prevalence and diversity of haemosporidians. We identified avian haemosporidians in waterbird species in three wetlands and investigated the effects of vector habitat suitability, landscape and host characteristics on the diversity and prevalence of these parasites. METHODS: We created a map with the probability of occurrence of avian haemosporidian vectors using maximum-entropy modelling based on references addressing species known to be vectors of haemosporidians in birds in Brazil. We determined the prevalence and diversity index of haemosporidians in the great egret (Ardea alba) (n = 129) and roseate spoonbill (Platalea ajaja) (n = 180) and compared the findings to data for the wood stork (Mycteria americana) (n = 199). RESULTS: We report the first record of Plasmodium in the family Threskiornithidae: four lineages in the roseate spoonbill, which also presented one lineage of Haemoproteus. In the family Ardeidae, we found three Plasmodium lineages in the great egret. The similar habitat suitability for vectors found in three wetlands explains the pattern of haemosporidian diversity determined for great egret and wood stork populations. Comparisons of haemosporidian diversity within each waterbird species and between regions showed a higher level in the central-western roseate spoonbill population than in the northern population (P = 0.021). Removing the host effect, we discussed the results obtained in terms of characteristics of the Pantanal region. Comparisons of Plasmodium spp. prevalence among waterbird species within the same wetland showed higher level in roseate spoonbill (74%) than those found in the great egret (21%) and wood stork (11%). Excluding the environmental effect, we interpreted result focusing host characteristics that favour infection: time required for nestlings to be covered by feathers and migratory behaviour. CONCLUSIONS: The map of habitat suitability showed that wetlands located in a 30° latitudinal range offer similar conditions for avian vectors species and diversity of haemosporidians. The lineages described in waterbirds were previously identified in birds of prey as Plasmodium paranucleophilum.

Use of noninvasive 'bug-eggs' to enable comparative inferences on genetic mating system with and without parental information: A study in a cattle egret colony.

Colonial waterbirds such as herons, egrets and spoonbills exhibit ecological characteristics that could have promoted the evolution of conspecific brood parasitism and extra-pair copulation. However, an adequate characterization of the genetic mating systems of this avian group has been hindered by the lack of samples of elusive candidate parents which precluded conducting conventional parentage allocation tests. Here, we investigate the genetic mating system of the invasive cattle egret using hematophagous insects contained in fake eggs to collect blood from incubating adults in a wild breeding colony. We tested a protocol with a previously unused Neotropical Triatominae, Panstrongylus megistus, obtained blood samples from males and females in 31 nests built on trees, drew blood from 89 nestlings at those nests, and genotyped all samples at 14 microsatellite loci, including six new species-specific loci. We comparatively addressed the performance of parentage allocation versus kinship classification of nestlings to infer the genetic mating system of cattle egrets. In line with previous behavioral observations, we found evidence in support of a non-monogamous genetic mating system, including extra-pair paternity (EPP) and conspecific brood parasitism (CBP). Parentage allocation tests detected a higher percentage of nests with alternative reproductive tactics (EPP: 61.7%; CBP: 64.5%) than the kinship classification method (EPP: 50.0%; CBP: 43.3%). Overall, these results indicate that rates of alternative reproductive tactics inferred in the absence of parental genetic information could be underestimated and should be interpreted with caution. This study highlights the importance of incorporating samples from candidate parents to adequately determine the genetic mating system of a species. We expand knowledge on the reproductive tactics of colonial waterbirds, contributing novel data on the genetic mating system of the cattle egret, valuable for the design of management strategies for this invasive bird.

Genetic differentiation among populations of the Roseate Spoonbill (Platalea ajaja; Aves: Pelecaniformes) in three Brazilian Wetlands.

Effective population size, levels of genetic diversity, gene flow, and genetic structuring were assessed in 205 colonial Roseate spoonbills from 11 breeding colonies from north, central west, and south Brazil. Colonies and regions exhibited similar moderate levels of diversity at five microsatellite loci (mean expected heterozygosity range 0.50-0.62; allelic richness range 3.17-3.21). The central west region had the highest Ne (59). F ST values revealed low but significant genetic structuring among colonies within the north and within the south regions. Significant global genetic structuring was found between the northern and central western populations as well as between the northern and southern populations. An individual-based Bayesian clustering method inferred three population clusters. Assignment tests correctly allocated up to 64% of individuals to their source regions. Collectively, results revealed complex demographic dynamics, with ongoing gene flow on a local scale, but genetic differentiation on a broader scale. Populations in the three regions may all be conserved, but special concern should be given to central western ones, which can significantly contribute to the species' gene pool in Brazil.

Conspecific brood parasitism in the white-faced ibis Plegadis chihi (Aves: Pelecaniformes) revealed by microsatellites' based kinship-reconstruction.

The white-faced ibis Plegadis chihi Vieillot, 1817 (Pelecaniformes: Threskiornithidae) is a socially monogamous colonially breeding bird in which behavioral and ecological observations suggest the occurrence of conspecific brood parasitism (CBP). We inferred aspects of the genetic mating system of P. chihi in nature, using a genetic approach in the absence of parental information. We used five heterologous microsatellite loci and a multiple-step methodological approach to infer kinship patterns among 104 pairs of nestlings sampled inside 80 nests in a breeding colony from southern Brazil. The estimated effective population size was 69 white-faced ibises (95% CI: 50-98), enough to ensure long-term population survival. Kinship patterns were identified for 38% of the analyzed pairs: 60% of the diagnosed pairs were identified as full-siblings, 2.5% as half-siblings and 37.5% as unrelated individuals. CBP could explain the presence of unrelated nestlings within broods, in agreement with available non-genetic evidence. The presence of half-siblings within broods could indicate extra-pair paternity. Results suggest that a non-strictly monogamous genetic mating system may be present in the white-faced ibis. This study is the first molecular approach to better characterize the reproductive behavior of P. chihi in the wild. Our findings set the stage for further research to investigate the possible causes and consequences of alternative reproductive strategies in this species.

Blood parasites in nestlings of wood stork populations from three regions of the American continent.

This study documents the prevalences and lineages of hemoparasites in wood stork nestlings from 3 regions of the American continent: southeastern United States (n = 90), northern Brazil (n = 74), and central-western Brazil (n = 125). Identification was based on PCR amplification of a mitochondrial small subunit ribosomal RNA gene. A fragment of the hemoparasite cytochrome B gene in infected individuals was utilized for Bayesian phylogenetic analysis. Four wood stork nestlings were infected by Haemoproteus, 1 from northern Brazil and 3 from the United States, and all shared the same haplotype. Morphological analysis confirmed the infection of the U.S. birds by Haemoproteus. Infection by Plasmodium was found in wood stork nestlings from northern (6) and central-western Brazil (14). Five Plasmodium lineages (MYCAMP1-2, and MYCAMP4-6) were found in the Brazilian central-western region and 3 Plasmodium lineages (MYCAMP2-3, and MYCAMP7) were found in the northern region. The most prevalent haplotype (MYCAMP2) differs from the others by 1 mutation, and the less prevalent haplotypes are derived from MYCAMP2. We did not find Plasmodium or Haemoproteus in nestlings younger than 15 and 30 days old, respectively. This is the first documentation of Plasmodium and Haemoproteus infection in wood storks in Brazilian breeding populations. Potential connectivity among wood stork populations was indirectly supported by the presence of identical Haemoproteus lineages in U.S. and northern Brazilian populations, and by the presence of identical Plasmodium haplotypes in the northern and central-western Brazilian populations.

Polymorphic heterologous microsatellite loci for population genetics studies of the white-faced ibis Plegadis chihi (Vieillot, 1817) (Pelecaniformes, Threskiornithidae).

We screened 44 heterologous microsatellites isolated in species of the families Threskiornithidae, Ciconiidae and Ardeidae for their use in a migratory waterbird, the white-faced ibis Plegadis chihi (Vieillot, 1817) (Threskiornithidae). Of the screened loci, 57% amplified successfully and 24% were polymorphic. In two breeding colonies from southern Brazil (N = 131) we detected 32 alleles (2-10 alleles/locus). Average He over all loci and colonies was 0.55, and the combined probability of excluding false parents, 98%. There was no departure from HWE in any loci or population. Eru6 and Eru4 loci were in non-random association in the Alvorada colony, and NnNF5 and Eru5 in both populations. AMOVA analysis indicated that most of the genetic diversity was contained within populations. Structure analysis suggested a single population, and F(ST) value showed weak genetic structuring (F(ST) = 0.009, p = 0.05). The two populations are apparently connected through gene-flow. The panel of six microsatellites optimized here was sufficiently informative for characterizing the genetic diversity and structure in these natural populations of the white-faced ibis. The information generated could be useful in future studies of genetic diversity, relatedness and the mating system in Plegadis chihi and related species.

Sedentary nestlings of Wood Stork as monitors of mercury contamination in the gold mining region of the Brazilian Pantanal.

Sedentary organisms that are at top trophic levels allow inference about the level of local mercury contamination. We evaluated mercury contamination in feather tissue of nestling Wood Storks (Mycteria americana), sampled in different parts of the Brazilian Pantanal that were variably polluted by mercury releases from gold mining activities. Levels of mercury in feathers sampled in seven breeding colonies were determined by atomic absorption spectroscopy, and the mean value of mercury concentration was 0.557 µg/g, dry weight (n=124), range 0.024-4.423 µg/g. From this total sample, 21 feathers that represent 30% of nestlings collected in Porto da Fazenda and Tucum colonies, in the northern region, ranged from 1.0 to 4.43 µg/g, dry weight (median value=1.87 µg/g). We found significant differences among regions (H=57.342; p=0<0.05). Results suggest that permanently flooded areas, or along mainstream rivers are more contaminated by mercury than dry areas, regardless of the distance from the gold mining center, which is located in the northern Pantanal. Highest values found in nestlings feathers were similar to those found in feathers of adult birds and in tissues of adult mammals that are less sedentary and were captured in the same region of Pantanal. These findings indicate that mercury released has been biomagnified and it is present in high concentrations in tissues of top consumers. We suggest a program to monitor mercury availability in this ecosystem using sedentary life forms of top predators like Wood Storks or other piscivorous birds.

Parentage and relatedness in captive and natural populations of the Roseate Spoonbill (Aves: Ciconiiformes) based on microsatellite data.

This study constitutes a first approach to evaluate the use of genetic information and relatedness estimators in the investigation of questions related to mating system and parentage in ex situ and in situ populations of the Roseate Spoonbill. We assessed the parentage assignments in 17 supposed families from US captive populations and investigated the genetic relationships among 67 nestlings sampled within 28 nests in Brazilian natural breeding colonies. Estimations of genetic relatedness values, hypothesis testing methods, simulations and maximum likelihood approaches were performed on data from four microsatellite loci. Parentage was confirmed in 61.5% of the registered parent-offspring relationships at zoo parks. Inconsistencies in assignments were investigated and the likely parents were identified for most of the hatchlings. Matings among relatives, not previously noticed based on behavioral observations, were identified by the use of genetic analyses. In natural populations, 33% of the sampled dyads were confidently classified as full-sibs. Above 25% of the analyzed dyads were unrelated, indicating that more than one parent-pair may have been responsible for the progeny. Our results demonstrate that genetic information can augment the precision in parentage assignment in captive Roseate Spoonbill populations, and this approach can contribute to their management and conservation. Results obtained using three different methodologies are concordant and point to the existence of a mating system other than monogamy for this species in the wild. The approaches implemented in this study can be applied to other waterbird species in which capture of adults is difficult.

Population genetic study of mitochondrial DNA in Roseate spoonbill (Aves; Platalea ajaja) breeding colonies from the Pantanal wetlands, Brazil.

Five breeding colonies of the Roseate spoonbill (Aves: Platalea ajaja) from two Brazilian wetland areas (Pantanal and Taim marshes) were sampled, and domain I of the mitochondrial DNA control region (483 bp) was sequenced in 50 birds. The average haplotype diversity (h = 0.75, s = 0.071) and average nucleotide diversity (pi = 0.004, s = 0.003) were evaluated, and nonsignificant differences were found among the colonies studied. The lack of differentiation among breeding colonies revealed by AMOVA analysis was explained either as a consequence of high gene flow or recent expansion. The significantly negative results of the neutrality tests (Fu's F ( s ) = -23.271, P < 0.01; Tajima's D = -1.941, P < 0.01) associated with the star shape of the haplotype tree and mismatch distribution data are evidence supporting the idea that these populations underwent a recent demographic expansion in the Pantanal region. The average time since the expansion is estimated to be 25,773 years, and this agrees with a period of increased moisture that occurred during the last glacial period.

Demographic history of wood stork (Mycteria americana) Brazilian Pantanal colonies revealed by mitochondrial DNA analysis

We used mitochondrial DNA (mtDNA) sequences to investigate the demographic history of the wood stork (Mycteria americana) populations in the Brazilian Pantanal. Sequences of 390/460 bp fragment of the mtDNA control region were analyzed in 62 wood stork specimens from 8 colonies using neutrality tests, phylogeographic, and coalescent analyses. Population expansion was supported by the significantly negative values of Tajima's (D = -2.071) and Fu's (Fs = -14.544) statistics and the unimodal pattern of mismatch distribution. Nested clade analyses indicated a historic range expansion event and recurrent gene flow that was restricted by isolation by distance as explanations for the haplotype distribution among the sampled colonies. High genetic diversity and the strictly unidirectional gene flow pattern emphasized the conservation importance of preserving the southern Pantanal colonies. Coalescence analyses suggested that northern and southern colonies diverged approximately 6,250 years before the present (YBP), and that their most recent common ancestor was approximately 18,900 YBP. Our results suggest that the contemporary wood stork Pantanal population originated from a more geographically limited founder population. Potential source populations may have occurred in the southern Pantanal or ancestry may reside in populations inhabiting the Brazilian central plateau or areas closer to the equatorial region.