Detection of mecA Genes in Hospital-Acquired MRSA and SOSA Strains Associated with Biofilm Formation.

Methicillin-resistant (MR) Staphylococcus aureus (SA) and others, except for Staphylococcus aureus (SOSA), are common in healthcare-associated infections. SOSA encompass largely coagulase-negative staphylococci, including coagulase-positive staphylococcal species. Biofilm formation is encoded by the icaADBC operon and is involved in virulence. mecA encodes an additional penicillin-binding protein (PBP), PBP2a, that avoids the arrival of ß-lactams at the target, found in the staphylococcal cassette chromosome mec (SCCmec). This work aims to detect mecA, the bap gene, the icaADBC operon, and types of SCCmec associated to biofilm in MRSA and SOSA strains. A total of 46% (37/80) of the strains were S. aureus, 44% (35/80) S. epidermidis, 5% (4/80) S. haemolyticus, 2.5% (2/80) S. hominis, 1.25% (1/80) S. intermedius, and 1.25% (1/80) S. saprophyticus. A total of 85% were MR, of which 95.5% showed mecA and 86.7% ß-lactamase producers; thus, Staphylococcus may have more than one resistance mechanism. Healthcare-associated infection strains codified type I-III genes of SCCmec; types IV and V were associated to community-acquired strains (CA). Type II prevailed in MRSA mecA strains and type II and III in MRSOSA (methicillin-resistant staphylococci other than Staphylococcus aureus). The operon icaADBC was found in 24% of SA and 14% of SOSA; probably the arrangement of the operon, fork formation, and mutations influenced the variation. Methicillin resistance was mainly mediated by the mecA gene; however, there may be other mechanisms that also participate, since biofilm production is related to genes of the icaADBC operon and methicillin resistance was not associated with biofilm production. Therefore, it is necessary to strengthen surveillance to prevent the spread of these outbreaks both in the nosocomial environment and in the community.

Aptamers: A Cutting-Edge Approach for Gram-Negative Bacterial Pathogen Identification.

Early and accurate diagnoses of pathogenic microorganisms is essential to correctly identify diseases, treating infections, and tracking disease outbreaks associated with microbial infections, to develop precautionary measures that allow a fast and effective response in epidemics and pandemics, thus improving public health. Aptamers are a class of synthetic nucleic acid molecules with the potential to be used for medical purposes, since they can be directed towards any target molecule. Currently, the use of aptamers has increased because they are a useful tool in the detection of specific targets. We present a brief review of the use of aptamers to detect and identify bacteria or even some toxins with clinical importance. This work describes the advances in the technology of aptamers, with the purpose of providing knowledge to develop new aptamers for diagnoses and treatment of different diseases caused by infectious microorganisms.

Isolation and Identification of Multidrug-Resistant Klebsiella pneumoniae Clones from the Hospital Environment.

Global dispersion, hospital outbreaks, and lineage relationships between emerging antibiotic-resistant strains such as Klebsiella pneumoniae are of public health interest. This study aimed to isolate and identify K. pneumoniae clones from third-level healthcare hospitals in Mexico to establish their multidrug-resistant phenotype, phylogeny, and prevalence. Biological and abiotic surface samples were used to isolate K. pneumoniae strains and to test their antibiotic susceptibility to classify them. The housekeeping genes: gapA, InfB, mdh, pgi, phoE, ropB, and tonB were used for multilocus sequence typing (MLST). Phylogenetic networks were constructed with 48 strains. Isolated strains (93) were mainly from urine and blood, 96% were resistant to ampicillin as expected, 60% were extended-spectrum ß-lactamases (ESBL), 98% were susceptible to ertapenem and meropenem and 99% were susceptible to imipenem, 46% were multi-drug resistant (MDR), 17% were extensively-drug resistant (XDR), 1% were pan-drug resistant (PDR), and 36% were not classified. The tonB, mdh, and phoE genes were the most variable, and the InfB gene showed positive selection. The most prevalent sequence types (STs) were ST551 (six clones), ST405 (six clones), ST1088 (four clones), ST25 (four clones), ST392 (three clones), and ST36 (two clones). ST706 was PDR, and ST1088 clones were MDR; neither of these STs has been reported in Mexico. The strains analyzed were from different hospitals and locations; thus, it is important to maintain antibiotic surveillance and avoid clone dissemination to prevent outbreaks, adaptation to antibiotics, and the transmission of antibiotic resistance.

The SEB1741 Aptamer Is an Efficient Tool for Blocking CD4+ T Cell Activation Induced by Staphylococcal Enterotoxin B.

Staphylococcal enterotoxin B (SEB) is a protein produced by Staphylococcus aureus, which is toxic to humans. It is well known for its ability to stimulate the exacerbated activation of proinflammatory CD4+ T cells (Th1 profile), and in vitro studies have been conducted to understand its mechanism of action and its potential use as an immune therapy. However, the efficiency of the SEB1741 aptamer in blocking SEB has not been experimentally demonstrated. METHODS: Enrichment CD4+ T cells were stimulated with SEB, and as a blocker, we used the SEB1741 aptamer, which was previously synthesised by an "in silico" analysis, showing high affinity and specificity to SEB. The efficiency of the SEB1741 aptamer in blocking CD4+ T cell activation was compared with that of an anti-SEB monoclonal antibody. Flow cytometry and Bio-Plex were used to evaluate the T-cell function. RESULTS: In vitro, SEB induced the activation of CD4+ T cells and favoured a Th1 profile; however, the SEB1741 aptamer was highly efficient in decreasing the frequency of CD4+ T cells positive to ki-67 and CD69 cells, this means that proliferation and activation of CD4+ T cells was decreased. Moreover, the production of interleukin 2 (IL-2) and interferon-gamma (IFN-γ) was affected, suggesting that the Th1 profile is not present when the SEB1441 aptamer is used. Thus, the SEB1741 function was similar to that of anti-SEB. CONCLUSIONS: The SEB1741 aptamer is a valuable tool for blocking CD4+ T cell activation and the subsequent release of proinflammatory cytokines by SEB stimulation.

In silico strategies for modeling RNA aptamers and predicting binding sites of their molecular targets.

RNA aptamers are single-stranded nucleic acids of 20-100 nucleotides, with high sensitivity and specificity against particular molecular targets. In vitro production and selection of aptamers can be performed using the SELEX method. However, this procedure requires considerable time and cost. In this sense, bioinformatics tools play an important role in reducing the time and cost associated with development and production of aptamers. In this article, we propose bioinformatics strategies for modeling and analysis of the interaction with molecular targets for two RNA aptamers: ATP binding RNA aptamer and iSpinach aptamer. For this purpose, molecular modeling of the tertiary structure of the aptamers was performed with two servers (SimRNA and RNAComposer); and AutoDock Vina and rDock programs were used to dock their respective ligands. The predictions developed with these methods could be used for in silico design of RNA aptamers, through a simple and accessible methodology.Supplemental data for this article is available online at https://doi.org/10.1080/15257770.2021.1951754 .

Differential Gene Expression Profile Induced by Valproic Acid (VPA) in Pediatric Epileptic Patients.

Epilepsy is a neuronal disease that affects up to 70 million people worldwide. The development of effective therapies to combat childhood epilepsy requires early biomarkers. Here, we performed a whole-genome microarray analysis in blood cells to identify genes differentially expressed between epileptic and epileptic valproic acid (VPA)-treated children versus normal children to obtain information about the gene expression to help us to understand genetic aspects of this disease. We found that the most significant differentially expressed genes were related to the transcriptional factor cAMP-response element binding protein (CREB) that is overexpressed in children with epilepsy compared with normal children, and 6 and 12 months of VPA treatment reversed several of these changes. Interestingly, leukocyte-associated immunoglobulin-like receptor 1 (LAIR1), a type I transmembrane glycoprotein that binds collagen proteins and contains CREB binding sites, was one of the more up-regulated genes in epileptic patients, and treatment with VPA strongly reversed its up-regulation. CREB up-regulates genes related to epilepsy; here, we suggest that LAIR1 could activate CREB, and together, they trigger epilepsy. After VPA treatment, LAIR1 repressed genes by disrupting the functional LAIR1⁻CREB complex, resulting in successful treatment. A functional microarray analysis offers new information that could open novel avenues of research in biomarker discovery, which may be useful for the early identification of children with a predisposition to epilepsy.

Helicobacter pylori: detection of iceA1 and iceA2 genes in the same strain in Mexican isolates.

BACKGROUND AND AIMS: Helicobacter pylori iceA1 and iceA2 gene amplification is usually performed to identify mixed populations as both genes are apparently reportedly exclusive. However, some strains isolated from Mexico show both iceA genes. The aim of this study was to establish the frequency of these genes in Mexican isolates and genomic diversity of the H. pylori strains. METHODS: One hundred thirty six biopsies were obtained from 68 patients (39 children and 29 adults). The presence of H. pylori was confirmed in 3/18 children and 6/19 adults by culture. There were 93 clinical strains isolated from nine patients. Additionally, we studied 37 strains from a strain collection isolated from 10 patients. The strains were genotyped and dual iceA genes were identified by polymerase chain reaction (PCR) and amplicons were sequenced. In addition, an enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) assay was performed as fingerprinting method. RESULTS: The genotypification of the H. pylori isolates indicated that all strains were vacA+; 86% babA2+, 86% cagA+, 82% vacA s1m1+, 19% iceA1+, 9% iceA2+, and 72% of them carried both iceA1 and iceA2 genes. The ERIC-PCR profiling revealed that the strains clustered in eight genetic groups depending on the presence of iceA1, iceA2 or both. A basic local multiple alignment analysis of the nucleotide sequences revealed that the iceA1 and iceA2 genes exhibited no relevant similarity. CONCLUSION: The results here showed the presence of triple-positive strains (babA, cagA, vacA) of H. pylori and strains carrying simultaneously both iceA1 and iceA2 genes.