Luciana W. Zuccherato

Swine and Poultry Pathogens: the Complete Genome Sequences of Two Strains of Mycoplasma hyopneumoniae and a Strain of Mycoplasma synoviae

This work reports the results of analyses of three complete mycoplasma genomes, a pathogenic (7448) and a nonpathogenic (J) strain of the swine pathogen Mycoplasma hyopneumoniae and a strain of the avian pathogen Mycoplasma synoviae; the genome sizes of the three strains were 920,079 bp, 897,405 bp, and 799,476 bp, respectively. These genomes were compared with other sequenced mycoplasma genomes reported in the literature to examine several aspects of mycoplasma evolution. Strain-specific regions, including integrative and conjugal elements, and genome rearrangements and alterations in adhesin sequences were observed in the M. hyopneumoniae strains, and all of these were potentially related to pathogenicity. Genomic comparisons revealed that reduction in genome size implied loss of redundant metabolic pathways, with maintenance of alternative routes in different species. Horizontal gene transfer was consistently observed between M. synoviae and Mycoplasma gallisepticum. Our analyses indicated a likely transfer event of hemagglutinin-coding DNA sequences from M. gallisepticum to M. synoviae.

Color and genomic ancestry in Brazilians: a study with forensic microsatellites.

The population of Brazil, formed by extensive admixture between Amerindians, Europeans and Africans, is one of the most variable in the world. We have recently published a study that used ancestry-informative markers to conclude that in Brazil, at an individual level, color, as determined by physical evaluation, was a poor predictor of genomic ancestry, estimated by molecular markers. To corroborate these findings we undertook the present investigation based on data from 12 commercially available forensic microsatellites that were utilized to estimate the personal genomic origin for each of 752 individuals from the city of São Paulo, belonging to different Brazilian color categories (275 Whites, 192 Intermediates and 285 Blacks). The genotypes permitted the calculation of a personal likelihood-ratio estimator of African or European ancestry. Although the 12 marker set proved capable of discriminating between European and African individuals, we observed very significant overlaps among the three color categories of Brazilians. This was confirmed quantitatively using a Bayesian analysis of population structure that did not demonstrate significant genetic differentiation between the three color groups. These results corroborate and validate our previous conclusions using ancestry-informative markers that in Brazil at the individual level there is significant dissociation of color and genomic ancestry.

A worldwide analysis of beta-defensin copy number variation suggests recent selection of a high-expressing DEFB103 gene copy in East Asia.

Beta‐defensins are a family of multifunctional genes with roles in defense against pathogens, reproduction, and pigmentation. In humans, six beta‐defensin genes are clustered in a repeated region which is copy‐number variable (CNV) as a block, with a diploid copy number between 1 and 12. The role in host defense makes the evolutionary history of this CNV particularly interesting, because morbidity due to infectious disease is likely to have been an important selective force in human evolution, and to have varied between geographical locations. Here, we show CNV of the beta‐defensin region in chimpanzees, and identify a beta‐defensin block in the human lineage that contains rapidly evolving noncoding regulatory sequences. We also show that variation at one of these rapidly evolving sequences affects expression levels and cytokine responsiveness of DEFB103, a key inhibitor of influenza virus fusion at the cell surface. A worldwide analysis of beta‐defensin CNV in 67 populations shows an unusually high frequency of high‐DEFB103‐expressing copies in East Asia, the geographical origin of historical and modern influenza epidemics, possibly as a result of selection for increased resistance to influenza in this region. Hum Mutat 32:743–750, 2011.

Study of AZFc partial deletion gr/gr in fertile and infertile Japanese males

AbstractA recurrent partial azoospermia factor C (AZFc) deletion, called gr/gr, has been reported to be a male infertility risk factor. A specific type of Y chromosome observed in approximately 30% of Japanese males (haplogroup D derived at YAP+) is believed to have a fixed gr/gr deletion. A recent study claimed that spermatogenic failure is more likely in males with D Y chromosomes, because of the gr/gr deletion, the presence of which is not well characterized among D haplogroup chromosomes. We therefore decided to perform a systematic study of the frequency of the gr/gr deletion in the Japanese. We studied fertile and infertile males to investigate the possibility of different gr/gr frequencies. The deletions were detected by use of single tagged-sequences (STSs) and the D haplogroup sub-lineages typing were done by use of the biallelic markers M174, M64, M116.1, 12f2.2, M15, M151, and M125. Analysis of gr/gr deleted Y chromosomes showed that all are classified as haplogroup D2, suggesting a lineage association. The subtype D2b1 was most frequent among the Japanese, in control and infertile samples. The haplogroups D2b2, D*, and D1 were not found in any population group. Remarkably, we observed no statistical difference between haplogroup D sub-lineages of the infertile and control groups, although the statistical power of this study is low. This study suggests lack of significant evidence of increased infertility risk in haplogroup D Japanese males. We were also able to establish the ancestral chromosome that suffered a gr/gr deletion, and propose a new Y chromosome phylogeny for haplogroup D and its derivatives. In summary, we were able to define the frequency of gr/gr deletion in Japanese males and show that the gr/gr deletion was probably present in the ancestral Y chromosome that entered Japan at least 12,000 years ago.

Population Genetics of GYPB and Association Study between GYPB*S/s Polymorphism and Susceptibility to P. falciparum Infection in the Brazilian Amazon

Background Merozoites of Plasmodium falciparum invade through several pathways using different RBC receptors. Field isolates appear to use a greater variability of these receptors than laboratory isolates. Brazilian field isolates were shown to mostly utilize glycophorin A-independent invasion pathways via glycophorin B (GPB) and/or other receptors. The Brazilian population exhibits extensive polymorphism in blood group antigens, however, no studies have been done to relate the prevalence of the antigens that function as receptors for P. falciparum and the ability of the parasite to invade. Our study aimed to establish whether variation in the GYPB*S/s alleles influences susceptibility to infection with P. falciparum in the admixed population of Brazil. Methods Two groups of Brazilian Amazonians from Porto Velho were studied: P. falciparum infected individuals (cases); and uninfected individuals who were born and/or have lived in the same endemic region for over ten years, were exposed to infection but have not had malaria over the study period (controls). The GPB Ss phenotype and GYPB*S/s alleles were determined by standard methods. Sixty two Ancestry Informative Markers were genotyped on each individual to estimate admixture and control its potential effect on the association between frequency of GYPB*S and malaria infection. Results GYPB*S is associated with host susceptibility to infection with P. falciparum; GYPB*S/GYPB*S and GYPB*S/GYPB*s were significantly more prevalent in the in the P. falciparum infected individuals than in the controls (69.87% vs. 49.75%; P<0.02). Moreover, population genetics tests applied on the GYPB exon sequencing data suggest that natural selection shaped the observed pattern of nucleotide diversity. Conclusion Epidemiological and evolutionary approaches suggest an important role for the GPB receptor in RBC invasion by P. falciparum in Brazilian Amazons. Moreover, an increased susceptibility to infection by this parasite is associated with the GPB S+ variant in this population.

Characterization of molecular and cellular phenotypes associated with a heterozygous CNTNAP2 deletion using patient-derived hiPSC neural cells

Neurodevelopmental disorders, such as autism spectrum disorders and schizophrenia, are complex disorders with a high degree of heritability. Genetic studies have identified several candidate genes associated with these disorders, including contactin-associated protein-like 2 (CNTNAP2). Traditionally, in animal models or in vitro, CNTNAP2 has been studied by genetic deletion or transcriptional knockdown, which reduces the expression of the entire gene; however, it remains unclear whether the mutations identified in clinical settings are sufficient to alter CNTNAP2 expression in human neurons. Here, using human induced pluripotent stem cells (hiPSCs) derived from two individuals with a large (289 kb) heterozygous deletion in CNTNAP2 (affecting exons 14–15) and discordant clinical outcomes, we have characterized CNTNAP2 expression patterns in hiPSC neural progenitor cells, two independent populations of hiPSC-derived neurons and hiPSC-derived oligodendrocyte precursor cells. First, we observed exon-specific changes in CNTNAP2 expression in both carriers; although the expression of exons 14–15 is significantly decreased, the expression of other exons is upregulated. Second, we observed significant differences in patterns of allele-specific expression in CNTNAP2 carriers that were consistent with the clinical outcome. Third, we observed a robust neural migration phenotype that correlated with diagnosis and exon- and allele-specific CNTNAP2 expression patterns, but not with genotype. In all, our data highlight the importance of considering the nature, location, and regulation of mutated alleles when attempting to connect genome wide association studies to gene function.

Development of two multiplex mini-sequencing panels of ancestry informative SNPs for studies in Latin Americans: an application to populations of the State of Minas Gerais (Brazil).

Admixture occurs when individuals from parental populations that have been isolated for hundreds of generations form a new hybrid population. Currently, interest in measuring biogeographic ancestry has spread from anthropology to forensic sciences, direct-to-consumers personal genomics, and civil rights issues of minorities, and it is critical for genetic epidemiology studies of admixed populations. Markers with highly differentiated frequencies among human populations are informative of ancestry and are called ancestry informative markers (AIMs). For tri-hybrid Latin American populations, ancestry information is required for Africans, Europeans and Native Americans. We developed two multiplex panels of AIMs (for 14 SNPs) to be genotyped by two mini-sequencing reactions, suitable for investigators of medium-small laboratories to estimate admixture of Latin American populations. We tested the performance of these AIMs by comparing results obtained with our 14 AIMs with those obtained using 108 AIMs genotyped in the same individuals, for which DNA samples is available for other investigators. We emphasize that this type of comparison should be made when new admixture/population structure panels are developed. At the population level, our 14 AIMs were useful to estimate European admixture, though they overestimated African admixture and underestimated Native American admixture. Combined with more AIMs, our panel could be used to infer individual admixture. We used our panel to infer the pattern of admixture in two urban populations (Montes Claros and Manhuaçu) of the State of Minas Gerais (southeastern Brazil), obtaining a snapshot of their genetic structure in the context of their demographic history.

Extensive admixture in Brazilian sickle cell patients: implications for the mapping of genetic modifiers.

To the editor: Genome-wide association studies (GWASs) of sickle cell disease (SCD) patients are a promising tool for identifying genetic modifiers of clinically relevant traits.[1][1],[2][2] In such a study, Solovieff et al[2][2] have uncovered a set of SNPs associated with fetal hemoglobin (HbF)

Identification of secreted virulence factors of Chromobacterium violaceum

Chromobacterium violaceum, a component of tropical soil microbiota, is an opportunistic pathogenic bacterium that can infect humans and other animals. In addition to identifying a large number of genes that demonstrate the vast biotechnological potential of this bacterium, genome sequencing revealed several virulence factors, including different cytolysins, which can be related to its pathogenicity. Here we confirmed these predictions from genomic analyses by identifying, through mass spectrometry, proteins present in the culture supernatant of C. violaceum that may constitute secreted virulence factors. Among them, we identified a secreted collagenase and the product of a gene with sequence similarity to previously characterized bacterial porins.

Comparative transcriptome profiling of virulent and non-virulent Trypanosoma cruzi underlines the role of surface proteins during infection

Trypanosoma cruzi, the protozoan that causes Chagas disease, has a complex life cycle involving several morphologically and biochemically distinct stages that establish intricate interactions with various insect and mammalian hosts. It has also a heterogeneous population structure comprising strains with distinct properties such as virulence, sensitivity to drugs, antigenic profile and tissue tropism. We present a comparative transcriptome analysis of two cloned T. cruzi strains that display contrasting virulence phenotypes in animal models of infection: CL Brener is a virulent clone and CL-14 is a clone that is neither infective nor pathogenic in in vivo models of infection. Gene expression analysis of trypomastigotes and intracellular amastigotes harvested at 60 and 96 hours post-infection (hpi) of human fibroblasts revealed large differences that reflect the parasite’s adaptation to distinct environments during the infection of mammalian cells, including changes in energy sources, oxidative stress responses, cell cycle control and cell surface components. While extensive transcriptome remodeling was observed when trypomastigotes of both strains were compared to 60 hpi amastigotes, differences in gene expression were much less pronounced when 96 hpi amastigotes and trypomastigotes of CL Brener were compared. In contrast, the differentiation of the avirulent CL-14 from 96 hpi amastigotes to extracellular trypomastigotes was associated with considerable changes in gene expression, particularly in gene families encoding surface proteins such as trans-sialidases, mucins and the mucin associated surface proteins (MASPs). Thus, our comparative transcriptome analysis indicates that the avirulent phenotype of CL-14 may be due, at least in part, to a reduced or delayed expression of genes encoding surface proteins that are associated with the transition of amastigotes to trypomastigotes, an essential step in the establishment of the infection in the mammalian host. Confirming the role of members of the trans-sialidase family of surface proteins for parasite differentiation, transfected CL-14 constitutively expressing a trans-sialidase gene displayed faster kinetics of trypomastigote release in the supernatant of infected cells compared to wild type CL-14.