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Introduction: The human body is a planet populated by myriads of microorganisms all over its surface and in cavities connected to the outside. Experimental and clinical research is showing that microbial colonizers are a functional and essential part of the human organism. The microbial ecosystem, which is housed in the gastrointestinal tract, provides a "metagenome": genes and additional functions to the genetic resources of the species, which are involved in multiple physiological processes (somatic development, nutrition, immunity, etc.). The human intestine houses lymphoid structures specialized in the induction and regulation of adaptive immunity, and the interaction of the intestinal microbiota with the immune system of the digestive mucosa plays a key role for the individual's homeostasis with the outside world. Some chronic non-communicable inflammatory diseases in developed society are associated with dysbiosis: loss of species richness in the gut microbiota and deviation from the ancestral microbial environment. Generating and maintaining diversity in the gut microbiota is a new clinical goal for health promotion and disease prevention.

Tuberculosis resistance diagnostics have vastly improved in recent years thanks to the development of standardised phenotypic and molecular testing methods. However, these methods are either slow or limited in the number of resistant genotypes they can detect. With the advent of next-generation sequencing (NGS) we can sidestep all those problems, as we can sequence whole tuberculosis genomes at increasingly smaller costs and requiring less and less DNA. In this review, we explain how accumulated knowledge in the field has allowed us to go from phenotypic testing to molecular methods to Whole Genome Sequencing (WGS) for resistance diagnostics. We compare current diagnostic methods with WGS as to their efficacy in detecting resistant cases, and show how forthcoming advances in NGS technologies will be crucial in widespread implementation of WGS as a diagnostic tool., (Copyright © 2019 SEPAR. Publicado por Elsevier España, S.L.U. All rights reserved.)

Introduction: Attention deficit hyperactivity disorder (ADHD) has a complex aetiology, mainly attributed to a number of susceptibility genes and environmental factors. Genetic association studies, however, have been inconsistent and have identified genetic variants with a moderate effect that explain a small proportion of the estimated inheritability of the disorder (< 10%). Recent studies suggest that the gut microbiota and diet play an important role in the development and symptoms of different mental disorders. Nevertheless, no clear evidence exists on the issue. This project proposes an alternative approach to identify mechanisms by which the intestinal microbial ecosystem and diet could contribute to the presence of ADHD., Aim: To identify biomarkers for ADHD by examining the gut microbiota., Subjects and Methods: We conducted a cross-sectional study of adult patients with ADHD (n = 100) and control subjects (n = 100). Measures of ADHD evaluation and eating habits were performed in both groups. Samples of faecal material were obtained from which to extract bacterial DNA, then used to characterise the participants' gut microbiota. A meta-genomic association study was later performed to attempt to correlate the bacterial composition of the intestine with the clinical subtypes of the disorder., Results and Conclusions: Comparing the gut microbiota profiles of subjects with ADHD and controls is expected to help account for the clinical heterogeneity of the disorder and identify new mechanisms involved in its development.

Mycobacterium tuberculosis, the etiological agent of human tuberculosis, causes annually three million deaths and latently infects about two billion people. Immunodeficiency caused by malnutrition, senescence or co-infection with HIV enhances the risk of developing active tuberculosis, either from a primary infection or by reactivation of a latent infection. The increasing appearance of multidrug-resistant strains to existing drugs is worrisome, since it leaves patients practically without treatment options. The understanding of the mechanisms of transmission, pathogenesis and virulence of M. tuberculosis is important. The analysis of its genome shows the presence of alternative sigma factors, transcriptional repressors and activators, two component signaling systems, metabolic enzymes and cellular secretory systems, that are associated with virulence in a series of pathogenic micro-organisms. Environmental stimuli such as pH, temperature, osmolality, oxygen availability are processed, activating or repressing virulence genes. The molecular mechanisms of action of these genes have been elucidated in in vitro and in vivo models.

Introduction: A growing number of sequenced genomes belonging to the Mycobacterium tuberculosis complex has enabled a comparison of strain traits and genomic constitution. These analyses may reveal mechanisms of evolution and genomic variation relevant to tuberculosis pathogenesis., Objective: Multiple alignments were used to analyze the differences between six genomes of the M. tuberculosis complex and to locate regions of variation that may lead to improvements in species identification or in their treatment., Materials and Methods: The Mauve software package was used to perform a multiple alignment of 6 genomes belonging to the M. tuberculosis complex. Regions exclusive to each genome were annotated using the TB database., Results: Percent similarity among the six genomes ranged between 96.1% and 97.8%. The annotation identified intergenic regions, regions associated with transposable elements of the PE-PGRS and PPE families, and regions associated with resistance against bacteriophage., Conclusions: In spite of the high genetic similarity among the tuberculosis strains, genomic variations were elucidated that may be relevant to differences in behavior and virulence, as well as for improvement of strain diagnosis. Regions encoding membrane-associated proteins, possibly related with antigenic variation and immune response, are particularly interesting for studies aimed at seeking tuberculosis treatments.

The bacteriological methods traditionally used in the diagnosis of acute respiratory infections (ARI) have limited sensitivity (culture, direct antigen detection, etc.) or require long periods to obtain results (appearance of antibodies). In the last few years, nucleic acid amplification techniques (NAAT) have been developed that allow pathogen-specific genetic targets to be detected in clinical samples. These techniques have been proven to be more sensitive than culture or direct detection and, unlike serological tests, are effective in the acute phase of the infection. However, NAAT also have certain limitations, such as the occasional presence of amplification inhibitors in clinical samples, the persistence of Mycoplasma pneumoniae or Chlamydophila pneumoniae in the mucosa of some individuals, and the lack of discrimination between pathogen infection and colonization in bacteria forming part of normal respiratory tract flora (Streptococcus pneumoniae). Recently developed real-time NAAT have raised expectations that some of these obstacles will be resolved, since these techniques allow bacterial load to be quantified. In the etiological diagnosis of ARI due to S. pneumoniae, the use of NAAT is still in an experimental phase. In M. pneumoniae and C. pneumoniae, combining NAAT with serological tests could potentially improve diagnosis. NAAT show good sensitivity and specificity in the detection of Legionella; however, the practical utility of these techniques should be weighed against that of antigenuria. NAAT provide advantages over other techniques in Bordetella pertussis. At present, these techniques are not useful in the diagnosis of Coxiella burnetii acute infections.

Cystic fibrosis is the most frequent lethal genetic disease that affects the caucasian population. The main cause of morbidity is the chronic lung infection, being the infection caused by Pseudomonas aeruginosa the most difficult to eradicate. This bacteria can be acquired in direct form, by person-to-person transfer, or indirectly, by hospital acquired infection. The Centro Provincial de Referencia de Fibrosis Quistica functioning in the Hospital de Niños "Sor María Ludovica", in La Plata, cares almost 220 patients aged two months to 45 years. The life expectancy depends of factors like the early diagnosis of the disease and the later acquisition of the chronic lung infection. The purpose of this work was the molecular typing of P. aeruginosa isolates obtained from cystic fibrosis patients to evaluate the genomic relationship among them. The study was carried out using RAPD-PCR. The analysis showed a great genetic heterogeneity among the isolates. The separation of the patients in groups in accordance with its bacteriology, that implies the attendance in different days and the implementation of isolation (or segregation) measures had demonstrated to be, in addition to other strategies, effective in the reduction of cross infections.

Mycobacteria that cause tuberculosis in animals and humans belong to the Mycobacterium tuberculosis complex. Techniques for conventional diagnosis are time-consuming and do not differentiate between different strains belonging to the M. tuberculosis complex. The aim of this study was to evaluate a multiplex PCR assay applicable to mycobacteria in culture with the capacity to differentiate different strains belonging to the M. tuberculosis complex in a reference laboratory. Primers based on genomics regions of difference (RD) consisting in DNA segments that are present in M. tuberculosis, but differentially deleted in several members of M. tuberculosis complex were used in a PCR assay. The test was applied to 86 clinical isolates of mycobacteria. The pattern of amplification allowed differentiating between M. tuberculosis, M. bovis and M. bovis BCG in a single PCR reaction. This PCR multiplex assay may be used in a Reference Laboratory of Tuberculosis Diagnosis as a complementary test to differentiate mycobacteria strains belonging to the M. tuberculosis complex. This test significantly reduces the time period between culture and strain identification, and thus for could favor the adoption of better strain specific antimycobacterial regimens as well as identification of zoonotic transmission of M. bovis to humans.

Listeria monocytogenes is a foodborne human pathogen responsible for invasive infections presenting overall a high mortality. Despite the ubiquity of the microorganism, the actual disease rate is quite low and the disease is most often associated with an underlying predisposition. Foodborne and environmental isolates were traditionally considered of similar pathogenicity compared to clinical isolates. But the analysis of mutations in the genes encoding specific virulence factors (internalin, hemolysin, phospholipases, surface protein ActA and regulator protein PrfA), quantitative studies with cell cultures and population genetics have raised considerable concerns about virulence differences among L. monocytogenes strains. Despite this great step forward, there is not a single marker available to test the virulence of field isolates of this species. In the future, the combination of different molecular markers will probably allow the screening of food contamination by only the virulent clones of L. monocytogenes, thus improving the prevention of foodborne human listeriosis.

Vaccination with attenuated Mycobacterium bovis BCG has been used as the routine procedure to immunize against tuberculosis. Since the efficacy of BCG vaccination is very controversial, the search for new immunoprophylatic tools against tuberculosis is an area of intense interest. Knowledge of the complete sequence of Mycobacterium tuberculosis (Mtb) H37Rv genome and the application of new immunological, biochemical and genetic technologies has led to a detailed understanding of the transcriptome and proteome of this bacterium. Approximately one-third of the human population is infected with Mtb; however, the bacillus is only detected once the symptoms appear and therefore most of the recent efforts have been devoted to the development of a post-infection vaccine. In theory, this vaccine (1) will give rise to an increase in the long-lasting specific immunity against Mtb, (2) will not have significant adverse effects, and (3) will be affordable for the people in third world countries. The main strategies that have been developed include the subunit vaccines, either as a mixture of relevant immunogenic proteins or DNA constructs, recombinant strains of Mycobacterium bovis BCG and Mtb, designed to secrete immunogenic proteins or with attenuated virulence, respectively, and DNA-based vaccines. The subunit vaccines are delivered either as mixtures of immunogenic proteins and adjuvants, or as naked DNA or by viral vectors in order to induce a potent Th1 response. Most of these vaccines have been tested in several kinds of animal models, but they do not fully reproduce the human pathology. However, the results obtained so far are very encouraging and have led to the development of phase I trials in humans.

Purpose/method: We report the case of a 75-year-old man with endogenous endophthalmitis in the left eye. Culture results were negative. PCR for several Gram-negative bacterias genome was performed., Results/conclusions: Polymerase chain reaction showed amplification of Serratia marcescens genome. Our case indicates that PCR can be a valuable tool in the diagnosis of culture negative endophthalmitis.

The bacterial genome is present in the cell within a complex structure, the nucleoid. The nucleoid contains the genomic DNA, and molecules of RNA and proteins. The main proteins of the nucleoid are: RNA polymerase, topoisomerases and the histone-like proteins: HU, H-NS (H1), H, HLP1, IHF and FIS. The DNA molecule in the nucleoid is under helical tension or supercoiling and is organized into 43 +/- 10 topodomains. DNA supercoiling is generated by the activity of the topoisomerases and by DNA-protein interactions. In this review, we analize current knowledge in Escherichia coli about genome organization and proteins of the nucleoid.