Revised Mimivirus major capsid protein sequence reveals intron-containing gene structure and extra domain

Background: Acanthamoebae polyphaga Mimivirus (APM) is the largest known dsDNA virus. The viral particle has a nearly icosahedral structure with an internal capsid shell surrounded with a dense layer of fibrils. A Capsid protein sequence, D13L, was deduced from the APM L425 coding gene and was shown to be the most abundant protein found within the viral particle. However this protein remained poorly characterised until now. A revised protein sequence deposited in a database suggested an additional N-terminal stretch of 142 amino acids missing from the original deduced sequence. This result led us to investigate the L425 gene structure and the biochemical properties of the complete APM major Capsid protein. Results: This study describes the full length 3430 bp Capsid coding gene and characterises the 593 amino acids long corresponding Capsid protein 1. The recombinant full length protein allowed the production of a specific monoclonal antibody able to detect the Capsid protein 1 within the viral particle. This protein appeared to be post-translationnally modified by glycosylation and phosphorylation. We proposed a secondary structure prediction of APM Capsid protein 1 compared to the Capsid protein structure of Paramecium Bursaria Chlorella Virus 1, another member of the Nucleo-Cytoplasmic Large DNA virus family. Conclusion: The characterisation of the full length L425 Capsid coding gene of Acanthamoebae polyphaga Mimivirus provides new insights into the structure of the main Capsid protein. The production of a full length recombinant protein will be useful for further structural studies

Nanobacteria Are Mineralo Fetuin Complexes

“Nanobacteria” are nanometer-scale spherical and ovoid particles which have spurred one of the biggest controversies in modern microbiology. Their biological nature has been severely challenged by both geologists and microbiologists, with opinions ranging from considering them crystal structures to new life forms. Although the nature of these autonomously replicating particles is still under debate, their role in several calcification-related diseases has been reported. In order to gain better insights on this calciferous agent, we performed a large-scale project, including the analysis of “nanobacteria” susceptibility to physical and chemical compounds as well as the comprehensive nucleotide, biochemical, proteomic, and antigenic analysis of these particles. Our results definitively ruled out the existence of “nanobacteria” as living organisms and pointed out the paradoxical role of fetuin (an anti-mineralization protein) in the formation of these self-propagating mineral complexes which we propose to call “nanons.” The presence of fetuin within renal calculi was also evidenced, suggesting its role as a hydroxyapatite nucleating factor

Carbapenem Resistance and Acinetobacter baumannii in Senegal: The Paradigm of a Common Phenomenon in Natural Reservoirs

Incidence of carbapenem-resistant Acinetobacter baumannii is rising in several parts of the world. In Africa, data concerning this species and its resistance to carbapenems are limited. The objective of the present study was to identify the presence of A. baumannii carbapenem-resistant encoding genes in natural reservoirs in Senegal, where antibiotic pressure is believed to be low. From October 2010 to January 2011, 354 human head lice, 717 human fecal samples and 118 animal fecal samples were screened for the presence of A. baumannii by real time PCR targeting blaOXA51-like gene. For all samples positive for A. baumannii, the carbapenemase-hydrolysing oxacillinases blaOXA23-like and blaOXA24-like were searched for and sequenced, and the isolates harbouring an oxacillinase were genotyped using PCR amplification and sequencing of recA gene. The presence of A. baumannii was detected in 4.0% of the head lice, in 5.4% of the human stool samples and in 5.1% of the animal stool samples tested. No blaOXA24 gene was detected but six fecal samples and three lice were positive for blaOXA23-like gene. The blaOXA23-like gene isolated in lice was likely a new oxacillinase sequence. Finally, the A. baumannii detected in stools were all of recA genotype 3 and those detected in lice, of recA genotype 4. This study shows for the first time a reservoir of blaOXA23-like-positive gene in human head lice and stool samples in Senegal

Draft genome of Gemmata massiliana sp. nov, a water-borne Planctomycetes species exhibiting two variants

International audienceGemmata massiliana is a new Planctomycetes bacterium isolated from a hospital water network in France, using a new culture medium. It is an aerobic microorganism with optimal growth at pH 8, at 30 °C and salinity ≤ 1.25 % NaCl. G. massiliana is resistant to β-lactam antibiotics, due to lack of peptidoglycan in its cell wall.G. massiliana shares a 97 % 16S rRNA gene sequence similarity with the nearest species, Gemmata obscuriglobus; and 99 % similarity with unnamed soil isolates. Its 9,249,437-bp genome consists in one chromosome and no detectable plasmid and has a 64.07 % G + C content, 32.94 % of genes encoding for hypothetical proteins. The genome contains an incomplete 19.6-kb phage sequence, 26 CRISPRs, 3 CAS and 15 clusters of secondary metabolites. G. massiliana genome increases knowledge of a poorly known world of bacteria

clbP Gene, a Potential New Member of the β-Lactamase Family

The colibactin island (pks) of Escherichia coli formed by 19 genes (55-Kb), encodes non-ribosomal peptide (NRP) and polyketide (PK) synthases, which allow the synthesis of colibactin, a suspected hybrid PK-NRP compound that causes damage to DNA in eukaryotic cells. The clbP, an unusual essential gene, is found in the operon structure with the clbS gene in the pks-encoded machinery. Interestingly, the clbP gene has been annotated as a β-lactamase but no previous study has reported its β-lactamase characteristics. In this study, we (i) investigated the β-lactamase properties of the clbP gene in silico by analysing its phylogenetic relationship with bacterial β-lactamase and peptidase enzymes, (ii) compared its three-dimensional (3D) protein structure with those of bacterial β-lactamase proteins using the Phyr2 database and PyMOL software, and (iii) evaluated in vitro its putative enzymatic activities, including β-lactamase, nuclease, and ribonuclease using protein expression and purification from an E. coli BL21 strain. In this study, we reveal a structural configuration of toxin/antitoxin systems in this island. Thus, similar to the toxin/antitoxin systems, the role of the clbP gene within the pks-island gene group appears as an antitoxin, insofar as it is responsible for the activation of the toxin, which is colibactin. In silico, our analyses revealed that ClbP belonged to the superfamily of β-lactamase, class C. Furthermore, in vitro we were unable to demonstrate its β-lactamase activity, likely due to the fact that the clbP gene requires co-expression with other genes, such as the genes present in the pks-island (19 genes). More research is needed to better understand its actions, particularly with regards to antibiotics, and to discover whether it has any additional functions due to the importance of this gene and its toxicity

Proteome analysis of Rickettsia conorii by two-dimensional gel electrophoresis coupled with mass spectrometry.

The availability of genome sequence offers the opportunity to further expand our knowledge about proteins expressed by Rickettsia conorii, strictly intracellular bacterium responsible for Mediterranean spotted fever. Using two-dimensional polyacrylamide gel electrophoresis combined with MALDI-TOF mass spectrometry, we established the first reference map of R. conorii proteome. This approach also allowed identification of GroEL as the major antigen recognized by rabbit serum and sera of infected patients. Altogether, this work opens the way to characterize the proteome of R. conorii, to compare protein profiles of different isolates or of bacteria maintained under different experimental conditions and to identify immunogenic proteins as potential vaccine targets

Proteome analysis of Rickettsia felis highlights the expression profile of intracellular bacteria.

The proteome of Rickettsia felis, an obligate intracellular bacterium responsible for spotted fever, was analyzed using two complementary proteomic approaches: 2-DE coupled with MALDI-TOF, and SDS-PAGE with nanoLC-MS/MS. This strategy allowed identification of 165 proteins and helped to answer some questions raised by the genome sequence of this bacterium. We successfully identified potential virulence factors including two putative adhesins, four proteins of the type IV secretion system, four Sca autotransporters, four components of ABC transporters, some R. felis-specific proteins, and one antitoxin of the toxin-antitoxin system. Notably, the antitoxin was the first to be identified in intracellular bacteria. Only one protein containing rickettsia palindromic repeats was found, whereas none of the split genes, transposases, or tetratricopeptide/ankyrin repeats were detectably expressed. Comparison of the protein expression profiles of R. felis and 23 other bacterial species according to functional categories showed that intracellular bacteria express more proteins related to translation, especially ribosomal proteins. However, the remaining bacteria express more proteins related to energy production and carbohydrate/amino acid metabolism. In conclusion, this study reveals R. felis virulence factor expression and highlights the unique protein expression profile of intracellular bacteria