Implementation of a novel in vitro model of infection of reconstituted human epithelium for expression of virulence genes in methicillin-resistant Staphylococcus aureus strains isolated from catheter-related infections in Mexico

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) are clinically relevant pathogens that cause severe catheter-related nosocomial infections driven by several virulence factors. METHODS: We implemented a novel model of infection in vitro of reconstituted human epithelium (RHE) to analyze the expression patterns of virulence genes in 21 MRSA strains isolated from catheter-related infections in Mexican patients undergoing haemodialysis. We also determined the phenotypic and genotypic co-occurrence of antibiotic- and disinfectant-resistance traits in the S. aureus strains, which were also analysed by pulsed-field-gel electrophoresis (PFGE). RESULTS: In this study, MRSA strains isolated from haemodialysis catheter-related infections expressed virulence markers that mediate adhesion to, and invasion of, RHE. The most frequent pattern of expression (present in 47.6% of the strains) was as follows: fnbA, fnbB, spa, clfA, clfB, cna, bbp, ebps, eap, sdrC, sdrD, sdrE, efb, icaA, and agr. Seventy-one percent of the strains harboured the antibiotic- and disinfectant-resistance genes ermA, ermB, tet(M), tet(K), blaZ, qacA, qacB, and qacC. PFGE of the isolated MRSA revealed three identical strains and two pairs of identical strains. The strains with identical PFGE patterns showed the same phenotypes and genotypes, including the same spa type (t895), suggesting hospital personnel manipulating the haemodialysis equipment could be the source of catheter contamination. CONCLUSION: These findings help define the prevalence of MRSA virulence factors in catheter-related infections. Some of the products of the expressed genes that we detected in this work may serve as potential antigens for inclusion in a vaccine for the prevention of MRSA-catheter-related infections

Gene expression profiles induced by E6 from non-European HPV18 variants reveals a differential activation on cellular processes driving to carcinogenesis

AbstractCervical cancer in developed countries remains as a major concern on public health policies due to incidence and mortality rates. Persistent infection with high risk human papillomavirus is a necessary etiological agent in the progression to invasive cervical carcinoma. A proposed hypothesis is the association between more aggressive HPV variants and the risk to develop cervical cancer. In order to have a global perspective in terms of cellular transcripts and molecular pathways affected by HPV18 E6 intratype variants; we conducted a genome wide analysis of gene expression. Our results show that E6 derived from non-European variants are able to up-regulate cellular transcripts associated to the hallmarks of cancer; such as cell cycle, migration, Wnt pathway and mTor signaling. Moreover, we were able to show that HPV18 E6 from African variant had a major effect on cellular processes such as cell cycle and migration as confirmed by functional studies

Effect of organic and conventional systems used to grow pecan trees on diversity of soil microbiota

Agronomic management modifies the soil bacterial communities and may alter the carbon fractions. Here, we identify differences in several chemical and biological soil variables, as well as bacterial composition between organic (Org) and conventional (Conv) agronomic management in pecan (Carya illinoinensis) orchards located in Coahuila, Mexico. The analyzed variables were pH, N, P, K, soil organic matter, organic matter quality, soil organic carbon, C/N ratio, carbon fractions, microbial biomass carbon, easily extractable Glomalin, colony-forming units, CO2 emissions, and the enzyme activity. The DNA of soil bacteria was extracted, amplified (V3-V4 16S rRNA), and sequenced using Illumina. To compare variables between agronomic managements, t tests were used. Sequences were analyzed in QIIME (Quantitative Insights Into Microbial Ecology). A canonical correspondence analysis (CCA) was used to observe associations between the ten most abundant phyla and soil variables in both types of agronomic managements. In Org management, variables related to the capture of recalcitrant carbon compounds were significant, and there was a greater diversity of bacterial communities capable of promoting organic carbon sequestration. In Conv management, variables related to the increase in carbon mineralization, as well as the enzymatic activity related to the metabolism of labile compounds, were significant. The CCA suggested a separation between phyla associated with some variables. Agronomic management impacted soil chemical and biological parameters related to carbon dynamics, including bacterial communities associated with carbon sequestration. Further research is still necessary to understand the plasticity of some bacterial communities, as well as the soil–plant dynamics.Fil: Cabrera Rodríguez, Alejandra. Universidad Autónoma de Chapingo; MéxicoFil: Nava Reyna, Erika. Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias; MéxicoFil: Trejo Calzada, Ricardo. Universidad Autónoma de Chapingo; MéxicoFil: García de la Peña, Cristina. Universidad Juárez del Estado de Durango; MéxicoFil: Arreola Avila, Jesús G.. Universidad Autónoma de Chapingo; MéxicoFil: Collavino, Mónica Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; ArgentinaFil: Vaca Paniagua, Felipe. Universidad Nacional Autónoma de México; MéxicoFil: Díaz Velásquez, Clara. Universidad Nacional Autónoma de México; MéxicoFil: Constante García, Vicenta. Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias; Méxic

General Microbiota of the Soft Tick Ornithodoros turicata Parasitizing the Bolson Tortoise (Gopherus flavomarginatus) in the Mapimi Biosphere Reserve, Mexico

The general bacterial microbiota of the soft tick Ornithodoros turicata found on Bolson tortoises (Gopherus flavomarginatus) were analyzed using next generation sequencing. The main aims of the study were to establish the relative abundance of bacterial taxa in the tick, and to document the presence of potentially pathogenic species for this tortoise, other animals, and humans. The study was carried-out in the Mapimi Biosphere Reserve in the northern-arid part of Mexico. Bolson tortoises (n = 45) were inspected for the presence of soft ticks, from which 11 tortoises (24.4%) had ticks in low loads (1–3 ticks per individual). Tick pools (five adult ticks each) were analyzed through 16S rRNA V3–V4 region amplification in a MiSeq Illumina, using EzBioCloud as a taxonomical reference. The operational taxonomic units (OTUs) revealed 28 phyla, 84 classes, 165 orders, 342 families, 1013 genera, and 1326 species. The high number of taxa registered for O. turicata may be the result of the variety of hosts that this tick parasitizes as they live inside G. flavomarginatus burrows. While the most abundant phyla were Proteobacteria, Actinobacteria, and Firmicutes, the most abundant species were two endosymbionts of ticks (Midichloria-like and Coxiella-like). Two bacteria documented as pathogenic to Gopherus spp. were registered (Mycoplasma spp. and Pasteurella testudinis). The bovine and ovine tick-borne pathogens A. marginale and A. ovis, respectively, were recorded, as well as the zoonotic bacteria A. phagocytophilum,Coxiella burnetii, and Neoehrlichia sp. Tortoises parasitized with O. turicata did not show evident signs of disease, which could indicate a possible ecological role as a reservoir that has yet to be demonstrated. In fact, the defense mechanisms of this tortoise against the microorganisms transmitted by ticks during their feeding process are still unknown. Future studies on soft ticks should expand our knowledge about what components of the microbiota are notable across multiple host–microbe dynamics. Likewise, studies are required to better understand the host competence of this tortoise, considered the largest terrestrial reptile in North America distributed throughout the Chihuahuan Desert since the late Pleistocene

Exploring bovine fecal bacterial microbiota in the Mapimi Biosphere Reserve, Northern Mexico

En México la información sobre la microbiota fecal bovina (Bos taurus) es escasa. El presente estudio describe la diversidad y abundancia de bacterias en muestras fecales de bovinos en pastizales, recolectadas en la Reserva de la Biosfera de Mapimí en la parte central del desierto chihuahuense. Las muestras fecales se analizaron mediante secuenciación masiva de siguiente generación de alto rendimiento utilizando la V3-V4 del ARNr 16S en Miseq de Illumina. Se identificaron un total de 17 filos, 24 clases, 33 órdenes, 50 familias, 281 géneros y 297 especies. Firmicutes y Verrucomicrobia fueron los filos más abundantes. Los géneros más abundantes fueron Sporobacter, PAC000748_g (géneros de la familia Ruminococcaceae) y Eubacterium_g23. Se registraron tres géneros (Clostridium, Corynebacterium y Fusobacterium) y una especie (Campylobacter fetus) de bacterias bovinas potencialmente patógenas. Esta información representa una línea base bacteriológica para monitorear el estado de salud intestinal de bovinos en pastoreo y para rastrear posibles interacciones con la microbiota fecal de la fauna nativa itinerante del área.In Mexico, information on the bovine fecal microbiota (Bos taurus) is scarce. The present study describes the diversity and abundance of bacteria in fecal samples from rangeland bovines, collected in the Mapimi Biosphere Reserve in the central part of the Chihuahuan desert. Fecal samples were analysed using high-throughput next generation massive sequencing using V3-V4 16S rRNA on Illumina Miseq. A total of 17 phyla, 24 classes, 33 orders, 50 families, 281 genera, and 297 species were identified. Firmicutes and Verrucomicrobia were the most abundant phyla. The most abundant genera were Sporobacter, PAC000748_g (genera into the Ruminococcaceae family) and Eubacterium_g23. Three genera (Clostridium, Corynebacterium and Fusobacterium) and one species (Campylobacter fetus) potentially pathogenic bovine bacteria were registered. This information represents a bacteriological baseline for monitoring the grazing bovine intestinal health status, and to trace possible interactions with the fecal microbiota of native roaming wildlife in the area

Resistance against two lytic phage variants attenuates virulence and antibiotic resistance in Pseudomonas aeruginosa

BackgroundBacteriophage therapy is becoming part of mainstream Western medicine since antibiotics of clinical use tend to fail. It involves applying lytic bacteriophages that self-replicate and induce cell lysis, thus killing their hosts. Nevertheless, bacterial killing promotes the selection of resistant clones which sometimes may exhibit a decrease in bacterial virulence or antibiotic resistance.MethodsIn this work, we studied the Pseudomonas aeruginosa lytic phage φDCL-PA6 and its variant φDCL-PA6α. Additionally, we characterized and evaluated the production of virulence factors and the virulence in a Galleria mellonella model of resistant mutants against each phage for PA14 and two clinical strains.ResultsPhage φDCL-PA6α differs from the original by only two amino acids: one in the baseplate wedge subunit and another in the tail fiber protein. According to genomic data and cross-resistance experiments, these changes may promote the change of the phage receptor from the O-antigen to the core lipopolysaccharide. Interestingly, the host range of the two phages differs as determined against the Pseudomonas aeruginosa reference strains PA14 and PAO1 and against nine multidrug-resistant isolates from ventilator associated pneumonia.ConclusionsWe show as well that phage resistance impacts virulence factor production. Specifically, phage resistance led to decreased biofilm formation, swarming, and type III secretion; therefore, the virulence towards Galleria mellonella was dramatically attenuated. Furthermore, antibiotic resistance decreased for one clinical strain. Our study highlights important potential advantages of phage therapy’s evolutionary impact that may be exploited to generate robust therapy schemes

Full-Exon Pyrosequencing Screening of BRCA Germline Mutations in Mexican Women with Inherited Breast and Ovarian Cancer

Hereditary breast cancer comprises 10% of all breast cancers. The most prevalent genes causing this pathology are BRCA1 and BRCA2 (breast cancer early onset 1 and 2), which also predispose to other cancers. Despite the outstanding relevance of genetic screening of BRCA deleterious variants in patients with a history of familial cancer, this practice is not common in Latin American public institutions. In this work we assessed mutations in the entire exonic and splice-site regions of BRCA in 39 patients with breast and ovarian cancer and with familial history of breast cancer or with clinical features suggestive for BRCA mutations by massive parallel pyrosequencing. First we evaluated the method with controls and found 41–485 reads per sequence in BRCA pathogenic mutations. Negative controls did not show deleterious variants, confirming the suitability of the approach. In patients diagnosed with cancer we found 4 novel deleterious mutations (c.2805_2808delAGAT and c.3124_3133delAGCAATATTA in BRCA1; c.2639_2640delTG and c.5114_5117delTAAA in BRCA2). The prevalence of BRCA mutations in these patients was 10.2%. Moreover, we discovered 16 variants with unknown clinical significance (11 in exons and 5 in introns); 4 were predicted as possibly pathogenic by in silico analyses, and 3 have not been described previously. This study illustrates how massive pyrosequencing technology can be applied to screen for BRCA mutations in the whole exonic and splice regions in patients with suspected BRCA-related cancers. This is the first effort to analyse the mutational status of BRCA genes on a Mexican-mestizo population by means of pyrosequencing

Latin American study of hereditary breast and ovarian cancer LACAM : a genomic epidemiology approach

Q2Q1Artículo original1-13Purpose: Hereditary Breast and Ovarian Cancer (HBOC) syndrome is responsible for ~5–10% of all diagnosed breast and ovarian cancers. Breast cancer is the most common malignancy and the leading cause of cancer-related mortality among women in Latin America (LA). The main objective of this study was to develop a comprehensive understanding of the genomic epidemiology of HBOC throughout the establishment of The Latin American consortium for HBOC-LACAM, consisting of specialists from 5 countries in LA and the description of the genomic results from the first phase of the study. Methods: We have recruited 403 individuals that fulfilled the criteria for HBOC from 11 health institutions of Argentina, Colombia, Guatemala, Mexico and Peru. A pilot cohort of 222 individuals was analyzed by NGS gene panels. One hundred forty-three genes were selected on the basis of their putative role in susceptibility to different hereditary cancers. Libraries were sequenced in MiSeq (Illumina, Inc.) and PGM (Ion Torrent-Thermo Fisher Scientific) platforms. Results: The overall prevalence of pathogenic variants was 17% (38/222); the distribution spanned 14 genes and varied by country. The highest relative prevalence of pathogenic variants was found in patients from Argentina (25%, 14/57), followed by Mexico (18%, 12/68), Guatemala (16%, 3/19), and Colombia (13%, 10/78). Pathogenic variants were found in BRCA1 (20%) and BRCA2 (29%) genes. Pathogenic variants were found in other 12 genes, including high and moderate risk genes such as MSH2, MSH6, MUTYH, and PALB2. Additional pathogenic variants were found in HBOC unrelated genes such as DCLRE1C, WRN, PDE11A, and PDGFB. Conclusion: In this first phase of the project, we recruited 403 individuals and evaluated the germline genetic alterations in an initial cohort of 222 patients among 4 countries. Our data show for the first time in LA the distribution of pathogenic variants in a broad set of cancer susceptibility genes in HBOC. Even though we used extended gene panels, there was still a high proportion of patients without any detectable pathogenic variant, which emphasizes the larger, unexplored genetic nature of the disease in these populations