Resumo em Inglês:

Mollicutes are cell wall-less bacteria with a genome characterized by its small size. Chromosomal rearrangements help these organisms evade host immune surveillance and hence cause disease. Our goal was to determine genes shared by Mollicutes genomes using the bidirectional best hit methodology. The twelve studied Mollicutes share 210 genes, most of which (> 60%) fall into the following COG categories: translation, ribosomal structure and biogenesis; DNA replication, recombination and repair; nucleotide transport and metabolism and energy production and conversion. Thirty Mollicute-specific genes were identified, 22 of them previously described as essential genes in Mycoplasma genitalium.

Resumo em Inglês:

Bacterial cell division has been studied mainly in model systems such as Escherichia coli and Bacillus subtilis, where it is described as a complex process with the participation of a group of proteins which assemble into a multiprotein complex called the septal ring. Mycoplasmas are cell wall-less bacteria presenting a reduced genome. Thus, it was important to compare their genomes to analyze putative genes involved in cell division processes. The division and cell wall (dcw) cluster, which in E. coli and B. subtilis is composed of 16 and 17 genes, respectively, is represented by only three to four genes in mycoplasmas. Even the most conserved protein, FtsZ, is not present in all mycoplasma genomes analyzed so far. A model for the FtsZ protein from Mycoplasma hyopneumoniae and Mycoplasma synoviae has been constructed. The conserved residues, essential for GTP/GDP binding, are present in FtsZ from both species. A strong conservation of hydrophobic amino acid patterns is observed, and is probably necessary for the structural stability of the protein when active. M. synoviae FtsZ presents an extended amino acid sequence at the C-terminal portion of the protein, which may participate in interactions with other still unknown proteins crucial for the cell division process.

Resumo em Inglês:

The annotation and comparative analyses of the genomes of Mycoplasma synoviae and Mycoplasma hyopneumonie, as well as of other Mollicutes (a group of bacteria devoid of a rigid cell wall), has set the grounds for a global understanding of their metabolism and infection mechanisms. According to the annotation data, M. synoviae and M. hyopneumoniae are able to perform glycolytic metabolism, but do not possess the enzymatic machinery for citrate and glyoxylate cycles, gluconeogenesis and the pentose phosphate pathway. Both can synthesize ATP by lactic fermentation, but only M. synoviae can convert acetaldehyde to acetate. Also, our genome analysis revealed that M. synoviae and M. hyopneumoniae are not expected to synthesize polysaccharides, but they can take up a variety of carbohydrates via the phosphoenolpyruvate-dependent phosphotransferase system (PEP-PTS). Our data showed that these two organisms are unable to synthesize purine and pyrimidine de novo, since they only possess the sequences which encode salvage pathway enzymes. Comparative analyses of M. synoviae and M. hyopneumoniae with other Mollicutes have revealed differential genes in the former two genomes coding for enzymes that participate in carbohydrate, amino acid and nucleotide metabolism and host-pathogen interaction. The identification of these metabolic pathways will provide a better understanding of the biology and pathogenicity of these organisms.

Resumo em Inglês:

Several mollicute genome projects are underway, offering unique opportunities to study genes and metabolic pathways on a genome-wide scale. Here, we have analyzed the conservation and diversity of purine and pyrimidine metabolism in mycoplasmas. An evaluation of discrepancies between genomic analysis and enzymatic data revealed interesting aspects about these organisms. We found important examples in which enzyme activity was reported without the annotation of a corresponding gene. An interesting example concerns phosphopentomutase. In Mollicutes, we have identified CDSs orthologous to sequences recently identified as new phosphopentomutases in archaeobacteria that are structurally related to phosphomannomutases. It is suggested that these sequences could replace the function of phosphopentomutases in mollicutes lacking the canonical phosphopentomutase gene (deoB). Also, the activity of 5'-nucleotidase was reported in mollicutes that do not possess any CDS related to ushA. Hypothetical proteins exhibiting domains similar to newly characterized 5' nucleotidases in Escherichia coli are proposed as possible CDSs related to this enzymatic activity in Mollicutes. Based on our analysis, the reductive genome evolution of Mollicutes does not appear to result in a minimum set of genes nor a minimum set of metabolic functions shared by all mollicute species.

Resumo em Inglês:

ABC transporters represent one of the largest superfamilies of active membrane transport proteins (MTPs) with a highly conserved ATPase domain that binds and hydrolyzes ATP, supplying energy for the uptake of a variety of nutrients and for the extrusion of drugs and metabolic wastes. The complete genomes of a non-pathogenic (J) and pathogenic (7448) strain of Mycoplasma hyopneumoniae, as well as of a pathogenic (53) strain of Mycoplasma synoviae have been recently sequenced. A detailed study revealed a high percentage of CDSs encoding MTPs in M. hyopneumoniae strains J (13.4%), 7448 (13.8%), and in M. synoviae 53 (11.2%), and the ABC systems represented from 85.0 to 88.6% of those CDSs. Uptake systems are mainly involved in cell nutrition and some might be associated with virulence. Exporter systems include both drug and multidrug resistant systems (MDR), which may represent mechanisms of resistance to toxic molecules. No relation was found between the phylogeny of the ATPase domains and the lifestyle or pathogenicity of Mycoplasma, but several proteins, potentially useful as targets for the control of infections, were identified.

Resumo em Inglês:

Mycoplasmas are the smallest known prokaryotes with self-replication ability. They are obligate parasites, taking up many molecules of their hosts and acting as pathogens in men, animals, birds and plants. Mycoplasma hyopneumoniae is the infective agent of swine mycoplasmosis and Mycoplasma synoviae is responsible for subclinical upper respiratory infections that may result in airsacculitis and synovitis in chickens and turkeys. These highly infectious organisms present a worldwide distribution and are responsible for major economic problems. Proteins of the GTPase superfamily occur in all domains of life, regulating functions such as protein synthesis, cell cycle and differentiation. Despite their functional diversity, all GTPases are believed to have evolved from a single common ancestor. In this work we have identified mycoplasma GTPases by searching the complete genome databases of Mycoplasma synoviae and Mycoplasma hyopneumoniae, J (non-pathogenic) and 7448 (pathogenic) strains. Fifteen ORFs encoding predicted GTPases were found in M. synoviae and in the two strains of M. hyopneumoniae. Searches for conserved G domains in GTPases were performed and the sequences were classified into families. The GTPase phylogenetic analysis showed that the subfamilies were well resolved into clades. The presence of GTPases in the three strains suggests the importance of GTPases in 'minimalist' genomes.

Resumo em Inglês:

Mycoplasma synoviae and Mycoplasma hyopneumoniae are wall-less eubacteria belonging to the class of Mollicutes. These prokaryotes have a reduced genome size and reduced biosynthetic machinery. They cause great losses in animal production. M. synoviae is responsible for an upper respiratory tract disease of chickens and turkeys. M. hyopneumoniae is the causative agent of enzootic pneumonia in pigs. The complete genomes of these organisms showed 17 ORFs encoding kinases in M. synoviae and 15 in each of the M. hyopneumoniae strain. Four kinase genes were restricted to the avian pathogen while three were specific to the pig pathogen when compared to each other. All deduced kinases found in the non pathogenic strain (J[ATCC25934]) were also found in the pathogenic M. hyopneumoniae strain. The enzymes were classified in nine families composing five fold groups.

Resumo em Inglês:

Mycoplasmas are very fastidious in their nutritional requirements for in vitro growth and have limited biosynthetic capacity, a reflection of their reduced genomes. As a result, these bacteria depend upon external metabolites for nutrition and growth and have developed dependence on their hosts for survival and maintenance. Protein degradation and peptide importation play an important role in Mycoplasma spp. nutrition, and proteases can play a role in host adaptation and pathogenicity. Here, we present a general survey on the genes involved in protein degradation, secretion and importation, comparing all available Mollicute genomes.

Resumo em Inglês:

This is a report on the analysis of genes involved in translation of the complete genomes of Mycoplasma hyopneumoniae strain J and 7448 and Mycoplasma synoviae. In both genomes 31 ORFs encoding large ribosomal subunit proteins and 19 ORFs encoding small ribosomal subunit proteins were found. Ten ribosomal protein gene clusters encoding 42 ribosomal proteins were found in M. synoviae, while 8 clusters encoding 39 ribosomal proteins were found in both M. hyopneumoniae strains. The L33 gene of the M. hyopneumoniae strain 7448 presented two copies in different locations. The genes encoding initiation factors (IF-1, IF-2 and IF-3), elongation factors (EF-G, EF-Tu, EF-Ts and EF-P), and the genes encoding the ribosome recycling factor (frr) and one polypeptide release factor (prfA) were present in the genomes of M. hyopneumoniae and M. synoviae. Nineteen aminoacyl-tRNA synthases had been previously identified in both mycoplasmas. In the two strains of M. hyopneumoniae, J and 7448, only one set of 5S, 16S and 23S rRNAs had been identified. Two sets of 16S and 23S rRNA genes and three sets of 5S rRNA genes had been identified in the M. synoviae genome.

Resumo em Inglês:

Restriction and Modification (R-M) systems are present in all Mycoplasma species sequenced so far. The presence of these genes poses barriers to gene transfer and could protect the cell against phage infections. The number and types of R-M genes between different Mycoplasma species are variable, which is characteristic of a polymorphism. The majority of the CDSs code for Type III R-M systems and particularly for methyltransferase enzymes, which suggests that functions other than the protection against the invasion of heterologous DNA may exist. A possible function of these enzymes could be the protection against the invasion of other but similar R-M systems. In Mycoplasma hyopneumoniae strain J, three of the putative methyltransferase genes were clustered in a region forming a genomic island. Many R-M CDSs were mapped in the vicinity of transposable elements suggesting an association between these genes and reinforcing the idea of R-M systems as mobile selfish DNA. Also, many R-M genes present repeats within their coding sequences, indicating that their expression is under the control of phase variation mechanisms. Altogether, these data suggest that R-M systems are a remarkable characteristic of Mycoplasma species and are probably involved in the adaptation of these bacteria to different environmental conditions.

Resumo em Inglês:

Mycoplasma hyopneumoniae is the etiological agent of porcine enzootic pneumonia (PEP), a major problem for the pig industry. The mechanisms of M. hyopneumoniae pathogenicity allow to predict the existence of several classes of virulence factors, whose study has been essentially restricted to the characterization of adhesion-related and major antigenic proteins. The now available complete sequences of the genomes of two pathogenic and one non-pathogenic strain of M. hyopneumoniae allowed to use a comparative genomics approach to putatively identify virulence genes. In this preliminary survey, we were able to identify 118 CDSs encoding putative virulence factors, based on specific criteria ranging from predicted cell surface location or variation between strains to previous functional studies showing antigenicity or involvement in host-pathogen interaction. This survey is expected to serve as a first step towards the functional characterization of new virulence genes/proteins that will be important not only for a better comprehension of M. hyopneumoniae biology, but also for the development of new and improved protocols for PEP vaccination, diagnosis and treatment.

Resumo em Inglês:

Mycoplasma sinoviae is a major pathogen of poultry causing synovitis and respiratory infection. M. synoviae hemagglutinin (VlhA) is a lipoprotein encoded by related multigene families that appear to have arisen by horizontal gene transfer. It is an abundant immunodominant surface protein involved in host-parasite interaction mediating binding to host erythrocytes. Herein, we have performed in silico analysis of the vlhA gene product from the Mycoplasma synoviae 53 strain and compared it to the VlhA protein of M. synoviae WUV1853 strain. The VlhA of the M. synoviae 53 strain possesses 569 amino acids and showed 85% identity with the VlhA protein of the M. synoviae WUV1853 strain. Further, a signal peptide was identified from amino acid M1 to D28 and a cleavage site between D28 and Q29, both located in the N-terminal domain of the molecule. Additionally, an insertion of PAPT amino acids was observed between T30-P35 and a deletion of the amino acids GTPGNP within the PRR region of the VlhA from the M. synoviae 53 strain, which may be related to its reduced virulence. Finally, we have identified 17 B cell epitopes and 22 T cells epitopes within the VlhA from the M. synoviae 53 strain. The B cell epitope S263-D277 and the T cell epitopes N45-N54 and G58-N67 showed 100% and 87-100% identity, respectively, with regions of VlhA protein of tested Mycoplasma synoviae and Mycoplasma galisepticum strains. Thus, these peptides represent new candidate molecules for the development of efficient diagnostic assays and new subunit vaccines.

Resumo em Inglês:

Diversification of bacterial species and pathotypes is largely caused by lateral gene transfer (LGT) of diverse mobile DNA elements such as plasmids, phages, transposons and genomic islands. Thus, acquisition of new phenotypes by LGT is very important for bacterial evolution and relationship with hosts. This paper reports a 23 kb region containing fourteen CDSs with similarity to the previous described Integrative Conjugal Element of Mycoplasma fermentans (ICEF). This element, named ICEH, is present as one copy at distinct integration sites in the chromosome of 7448 and 232 pathogenic strains and is absent in the type strain J (non-pathogenic). Notable differences in the nucleotide composition of the insertion sites were detected, and could be correlated to a lack of specificity of the ICEH integrase. Although present in strains of the same organism, the ICEH elements are more divergent than the typical similarity between other chromosomal locus of Mycoplasma hyopneunomiae, suggesting an accelerated evolution of these constins or an ongoing process of degeneration, while maintaining conservation of the tra genes. An extrachromosomal form of this element had been detected in the 7448 strain, suggesting a possible involvement in its mobilization and transference of CDSs to new hosts.

Resumo em Inglês:

Several genomes of mycoplasmas have been sequenced and here we tried to retrieve the evolutionary relationships of nine species using a phylogenomic approach. Several methods were used to build phylogenetic trees based on protein sequence information, gene-order, and gene-content. We also utilized datasets composed of individual and concatenated sets of orthologous proteins, as well as with reduced unreliable alignment regions. Most of our results converge to a single topology, except for the trees built with both the maximum parsimony method and with the gene-order dataset. The gene-content dataset presented trees consistent with most nodes of the convergent tree, but in the gene-order dataset most internal branches were clearly saturated and unreliable. The topological difference between the trees obtained by the diverse methods could not be explained by regions with unreliable alignments or attributed to horizontal gene transfer among the genomes. It is possible that the incongruence between the methods could be associated with their differential sensibility in relation to certain evolutionary factors. Further analysis using other empirical genomic datasets would be necessary in order to better understand the basis of such conflicts.

Resumo em Inglês:

This report describes the transcription apparatus of Mycoplasma hyopneumoniae (strains J and 7448) and Mycoplasma synoviae, using a comparative genomics approach to summarize the main features related to transcription and control of gene expression in mycoplasmas. Most of the transcription-related genes present in the three strains are well conserved among mycoplasmas. Some unique aspects of transcription in mycoplasmas and the scarcity of regulatory proteins in mycoplasma genomes are discussed.

Resumo em Inglês:

We have analyzed the sequenced genomes of three strains of Mycoplasma hyopneumoniae and one strain of M. synoviae, and have found three and two different transposable element families, respectively in each species. In M. hyopneumoniae, the Insertion Sequences of the IS4 family is represented by ISMHp1, a putatively active element. The IS3 family is represented by several degenerated sequences. A third element called tMH was found, which shows some characteristics reminiscent of retrotransposons. In M. synoviae, three different possibly active IS4 elements are present (ISMHp1-like; ISMs1 and IS1634-like elements). The IS30 family is represented by the degenerated IS1630-like element. The IS1634-like element is shown to be involved in chromosomal rearrangements and horizontal gene transfer (HGT). The ISMHp1-like element is shown to relate to the HGT of a 25-kb region from M. gallisepticum to M. synoviae. The fractions of these genomes that correspond to mobile elements varied from 1.35 to 3.13% in M. hyopneumonia strains and was 2.08% in M. synoviae. Although these species possess reduced genomes, they maintain mobile elements, perhaps as a mechanism for genetic variability production.

Resumo em Inglês:

In this work, we predict a structural model for the RecA protein from M. synoviae (MsRecA) by theoretical homology modeling and evaluate the occurrence of polymorphisms in this protein within several isolates of this species. The structural model suggested for MsRecA conserves the main domains present in MtRecA and EcRecA. The L1 and L2 regions showed six and three amino acid substitutions, respectively, which apparently do not affect the conformation and function of MsRecA. The C-terminal domain is shorter than that found in EcRecA and MtRecA, which may increase its capacity to bind dsDNA and displace SSB, compensating the absence of recombination initiation enzymes. The MS59 isolate RecA sequence showed one polymorphism which does not affect its functions since these belong to the same physical-chemical group.