Tatiana de Campos

Structure of genetic diversity among common bean ( Phaseolus vulgaris L.) varieties of Mesoamerican and Andean origins using new developed microsatellite markers

A common bean genomic library was constructed using the ‘IAC-UNA’ variety enriched for (CT) and (GT) for microsatellite motifs. From 1,209 sequenced clones, 714 showed microsatellites distributed over 471 simple and 243 compound motifs. GA/CT and GT/CA were the most frequent motifs found among these sequences. A total of 123 microsatellites has been characterized. Out of these, 87 were polymorphic (73.7%), 33 monomorphic (26.8%), and 3 (2.4%) did not amplify at all. In a sample of 20 common bean materials selected from the Agronomic Institute Germplasm Bank, the number of alleles per locus varied 2–9, with an average of 2.82. The polymorphic information content (PIC) of each marker varied from 0.05 to 0.83, with a 0.45 average value. Cluster and principal coordinate analysis of the microsatellite data were consistent with the original assignment of the germplasm accessions into the Andean and Mesoamerican gene pools of common bean. Low polymorphism levels detected could be associated with the domestication process. These microsatellites could be a valuable resource for the bean community because of their use as new markers for genetic studies.

Subpathotypes of Avian Pathogenic Escherichia coli (APEC) Exist as Defined by their Syndromes and Virulence Traits

Avian pathogenic Escherichia coli (APEC) strains cause different types of systemic extraintestinal infections in poultry, collectively termed colibacillosis, which can cause significant economic losses in the poultry industry. To date, there have been no descriptions of genes or characteristics that allow for the classification of avian strains pathotypes responsible for causing specific diseases in their hosts. In this study we aimed to characterize avian E. coli strains representing 4 groups, including one of commensal strains (AFEC – Avian Fecal Escherichia coli) and 3 groups of APEC strains, where each group is responsible for causing a different disease syndrome in their respective hosts (septicemia, omphalitis and swollen head syndrome). We chose to examine several biological characteristics of these strains including: adhesion to eukaryotic cells, pathogenicity levels according to the lethal dose (50%) assay, phylogenetic group and virulence gene profiles. The comparison of strains based on these genotypic and phenotypic traits, using multivariate statisticals tools and complex networks, allowed us to infer information about the population structure of the studied groups. Our results indicate that APEC strains do not constitute a unique homogeneous group, but rather a structured set of subgroups, where each one is associated with a specific infectious syndrome which can possibly be used to define pathotypes or subpathotypes within APEC strains. These results offer new possibilities with which to study the genes responsible for various pathogenetic processes within APEC strains, and for vaccine development. It may be important to consider these subgroups when developing a vaccine in an effort for obtain cross protection, which has not yet been successfully accomplished when working with APEC strains.

Genetic diversity in cultivated carioca common beans based on molecular marker analysis

A wide array of molecular markers has been used to investigate the genetic diversity among common bean species. However, the best combination of markers for studying such diversity among common bean cultivars has yet to be determined. Few reports have examined the genetic diversity of the carioca bean, commercially one of the most important common beans in Brazil. In this study, we examined the usefulness of two molecular marker systems (simple sequence repeats – SSRs and amplified fragment length polymorphisms – AFLPs) for assessing the genetic diversity of carioca beans. The amount of information provided by Roger’s modified genetic distance was used to analyze SSR data and Jaccards similarity coefficient was used for AFLP data. Seventy SSRs were polymorphic and 20 AFLP primer combinations produced 635 polymorphic bands. Molecular analysis showed that carioca genotypes were quite diverse. AFLPs revealed greater genetic differentiation and variation within the carioca genotypes (Gst = 98% and Fst = 0.83, respectively) than SSRs and provided better resolution for clustering the carioca genotypes. SSRs and AFLPs were both suitable for assessing the genetic diversity of Brazilian carioca genotypes since the number of markers used in each system provided a low coefficient of variation. However, fingerprint profiles were generated faster with AFLPs, making them a better choice for assessing genetic diversity in the carioca germplasm.

Species boundaries inferred from microsatellite markers in the Kielmeyera coriacea complex (Calophyllaceae) and evidence of asymmetric hybridization

Kielmeyera coriacea is one of the most characteristic entities of the Cerrado, a savanna vegetation of Central Brazil and it is a morphologically highly variable polyploid species. In the revision of the genus, an earlier variety was elevated to a related species. However, this proposal has not been accepted by the scientific community and the problematic taxonomic circumscription of the group raises doubts about various scientific studies on plants identified as K. coriacea. In the present study, we investigated the subject by a population genetics approach using eight microsatellite markers in an attempt to understand better the relationships between the proposed species. Like morphological and anatomical observations, molecular studies of highly polymorphic markers showed a great similarity between K. coriacea and K. grandiflora. However, this tool could effectively screen the boundaries of the species complex and, in addition, detect an introgressive hybridization process.

Isolation and characterization of microsatellite loci in the stingless bee Melipona interrupta manaosensis (Apidae: Meliponini)

Melipona interrupta manaosensis is an important pollinator in the Neotropics. This stingless bee is reared by locals for honey production and also for conservation purposes. Eleven microsatellite markers were isolated from M. i. manaosensis and characterized in 32 individuals from the Meliponary of the Instituto Nacional de Pesquisas da Amazônia, Manaus, AM, Brazil. The number of alleles observed for each locus ranged from 2 to 5 (average 2.77). The polymorphism information content (PIC) varied between 0.08 to 0.62 (average 0.34) and the discriminating power (D) ranged from 0.61 to 0.99 (average 0.85). The observed heterozygosity (HO) and the expected heterozygosity (HE) ranged from 0.04 to 1.00 and 0.04 to 0.67, respectively. These microsatellites will be useful for future genetic studies of M. i. manaosensis populations and correlated species contributing with Amazon biodiversity preservation.

Occurrence of virulence-related sequences and phylogenetic analysis of commensal and pathogenic avian Escherichia coli strains (APEC)

ABSTRACT.- Campos T.A., Lago J.C., Nakazato G., Stehling E.G., Brocchi M., CastroA.F.P. & Silveira W.D. 2008. Occurrence of virulence-related sequences andphylogenetic analysis of commensal and pathogenic avian Escherichia coli strains(APEC). Pesquisa Veterinaria Brasileira 28(10):533-540. Departamento de Microbiologiae Immunologia, Instituto de Biologia, Unicamp, Cidade Universitraria Zeferino Vaz s/n,Campinas, SP 13081-862, Brazil. E-mail: wds@unicamp.brThe presence of iron uptake ( irp -2, fyu A, sit A, fep C, iuc A), adhesion ( iha, lpf A O157/O141 , lpf A O157/O154 , efa, tox B) and invasion ( inv, ial- related DNA sequences and assignment tothe four main Escherichia coli phylogenetic groups (A, B1, B2 e D) were determined in 30commensal E. coli strains isolated from healthy chickens and in 49 APEC strains isolatedfrom chickens presenting clinical signs of septicemia (n=24) swollen head syndrome(n=14) and omphalitis (n=11) by PCR. None of the strains presented DNA sequencesrelated to the

Molecular Diversity and Genetic Structure of Guineagrass (Panicum maximum Jacq.), a Tropical Pasture Grass

Guineagrass (Panicum maximum Jacq.) is a forage grass found in tropical and subtropical regions. It is an apomictic and tetraploid species from Africa. The objective of this study was to evaluate the genetic diversity of guineagrass accessions sampled from its regions of origin, which is in Tanzania and Kenya. In this study, a total of 396 accessions were analyzed, and a collection of reproducible and informative microsatellites was developed. Thirty microsatellites were employed to characterize these accessions. A total of 576 clones were sequenced from microsatellite-enriched libraries. Flanking primers were designed for 116 microsatellite loci and screened using a sample of 25 guineagrass accessions. The thirty selected polymorphic microsatellites employed in this study produced a total of 192 bands when evaluated in the 396 P. maximum accessions, with an average of 6.4 bands per microsatellite. Four genetic clusters were identified in the collection using STRUCTURE analysis, and these results were confirmed using AMOVA. The largest genetic variation was found within clusters (65.38%). This study revealed that the collection of accessions from the P. maximum region of origin was a rich source of genetic variability. The geographical distances and genetic similarities among accessions did not indicate a significant association between genetic and geographical variation, supporting the natural interspecific crossing between P. maximum, P. infestum and P. trichocladum as the origin of the high genetic variability and the existence of an agamic complex formed by these three species.

Isolation and characterization of microsatellite markers from the stingless bee Nannotrigona testaceicornis

Conservation of natural populations and handling of breeding programs would benefit from the availability of molecular markers. Stingless bees are one of the most important pollinators in several ecosystems. Thus, seventeen microsatellite markers were developed from an enriched genomic library of Nannotrigona testaceicornis. They were characterized using 50 samples. The expected and observed heterozygosities ranged from 0.59 to 0.89 and from 0.39 to 0.79, respectively. These markers will contribute to advance researches on the genetic conservation, characterization and preservation of the Brazilian native bees.

Potato cultivar identification using molecular markers

O objetivo deste trabalho foi avaliar um conjunto de marcadores microssatelites para identificacao e caracterizacao varietal das cultivares de batata mais amplamente utilizadas no Brasil. O DNA das 14 variedades de batata foi genotipado com marcadores microssatelites, e os alelos foram visualizados em gel de poliacrilamida corado com prata. Vinte e quatro marcadores foram avaliados e apenas um loco foi monomorfico. Com base no padrao de bandas, foi obtido um conjunto com dois microssatelites capazes de identificar e diferenciar todas as cultivares analisadas.

Microsatellite markers isolated from polyploid Kielmeyera coriacea Mart. & Zucc. (Clusiaceae) from an enriched genomic library

Kielmeyera coriacea is a morphologically highly variable polyploid species, characteristic of the Cerrado, the savanna-like vegetation of the Central-Brazil. It was subject of various biological studies and, like others cerrado plants, this species suffers strong anthropogenic pressure. To access the genetic diversity of populations of this species, its taxonomic circumscriptions and to study supposed hybridization events, we have developed 11 polymorphic microsatellite loci from a CT/GT-enriched genomic library. The band pattern showed 12–32 bands per locus, and 2–8 bands per locus per individual. All microsatellites successfully amplified PCR products in K. grandiflora, the putative sister species, and all revealed similar multibanded pattern.