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Enterobacter cloacae is an emerging pathogen isolated in healthcare-associated infections. A major virulence factor of this bacterium is the type VI secretion system (T6SS). The genome of E. cloacae harbors two T6SS gene clusters (T6SS-1 and T6SS-2), and the functional characterization of both systems showed that these two T6SSs are not expressed under the same conditions. Here, we report that the major histone-like protein HU positively regulates the expression of both T6SSs and, therefore, the function that each T6SS exerts in E. cloacae. Single deletions of the genes encoding the HU subunits (hupA and hupB) decreased mRNA levels of both T6SS. In contrast, the hupA hupB double mutant dramatically affected the T6SS expression, diminishing its transcription. The direct binding of HU to the promoter regions of T6SS-1 and T6SS-2 was confirmed by electrophoretic mobility shift assay. In addition, single and double mutations in the hup genes affected the ability of inter-bacterial killing, biofilm formation, adherence to epithelial cells, and intestinal colonization, but these phenotypes were restored when such mutants were trans-complemented. Our data broaden our understanding of the regulation of HU-mediated T6SS in these pathogenic bacteria. IMPORTANCE: T6SS is a nanomachine that functions as a weapon of bacterial destruction crucial for successful colonization in a specific niche. Enterobacter cloacae expresses two T6SSs required for bacterial competition, adherence, biofilm formation, and intestinal colonization. Expression of T6SS genes in pathogenic bacteria is controlled by multiple regulatory systems, including two-component systems, global regulators, and nucleoid proteins. Here, we reported that the HU nucleoid protein directly activates both T6SSs in E. cloacae, affecting the T6SS-related phenotypes. Our data describe HU as a new regulator involved in the transcriptional regulation of T6SS and its impact on E. cloacae pathogenesis.

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Here, we conducted a comprehensive analysis of 356 Klebsiella pneumoniae species complex (KpSC) isolates that were classified as classical (cl), presumptive hypervirulent (p-hv) and hypermucoviscous-like (hmv-like). Overall, K. pneumoniae (82.3%), K. variicola (2.5%) and K. quasipneumoniae (2.5%) were identified. These isolates comprised 321 cl-KpSC, 7 p-hv-KpSC and 18 hmv-like-KpSC. A large proportion of cl-KpSC isolates were extended-spectrum-ß-lactamases (ESBLs)-producers (64.4%) and 3.4% of isolates were colistin-resistant carrying carbapenemase and ESBL genes. All p-hv-KpSC showed an antibiotic susceptible phenotype and hmv-like isolates were found to be ESBL-producers (8/18). Assays for capsule production and capsule-dependent virulence phenotypes and whole-genome sequencing (WGS) were performed in a subset of isolates. Capsule amount differed in all p-hv strains and hmv-like produced higher capsule amounts than cl strains; these variations had important implications in phagocytosis and virulence. Murine sepsis model showed that most cl strains were nonlethal and the hmv-like caused 100% mortality with 3 × 108 CFUs. Unexpectedly, 3/7 (42.9%) of p-hv strains required 108 CFUs to cause 100% mortality (atypical hypervirulent), and 4/7 (57.1%) strains were considered truly hypervirulent (hv). Genomic analyses confirmed the diverse population, including isolates belonging to hv clonal groups (CG) CG23, CG86, CG380 and CG25 (this corresponded to the ST3999 a novel hv clone) and MDR clones such as CG258 and CG147 (ST392) among others. We noted that the hmv-like and hv-ST3999 isolates showed a close phylogenetic relationship with cl-MDR K. pneumoniae. The information collected here is important to understand the evolution of clinically important phenotypes such as hypervirulent and ESBL-producing-hypermucoviscous-like amongst the KpSC in Mexican healthcare settings. Likewise, this study shows that mgrB inactivation is the main mechanism of colistin resistance in K. pneumoniae isolates from Mexico.

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Carbon capture, utilization, and storage (CCUS) technology offer promising solution to mitigate the threatening consequences of large-scale anthropogenic greenhouse gas emissions. Within this context, this report investigates the influence of NiO deposition on the Li4SiO4 surface during the CO2 capture process and its catalytic behavior in hydrogen production via dry methane reforming. Results demonstrate that the NiO impregnation method modifies microstructural features of Li4SiO4, which positively impact the CO2 capture properties of the material. In particular, the NiO-Li4SiO4 sample captured twice as much CO2 as the pristine Li4SiO4 material, 6.8 and 3.4 mmol of CO2 per gram of ceramic at 675 and 650 °C, respectively. Additionally, the catalytic results reveal that NiO-Li4SiO4 yields a substantial hydrogen production (up to 55 %) when tested in the dry methane reforming reaction. Importantly, this conversion remains stable after 2.5 h of reaction and is selective for hydrogen production. This study highlights the potential of Li4SiO4 both a support and a captor for a sorption-enhanced dry reforming of methane. To the best of our knowledge, this is the first report showcasing the effectiveness of Li4SiO4 as an active support for Ni-based catalysis in the dry reforming of methane. These findings provide valuable insights into the development of this composite as a dual-functional material for carbon dioxide capture and conversion.

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Enterotoxigenic Escherichia coli (ETEC) strains produce at least one of two types of enterotoxins: the heat-labile (LT) and heat-stable (ST) toxins, which are responsible for the watery secretory diarrhoea that is a hallmark of the human ETEC infection. One regulatory system that controls the transcription of virulence genes in pathogenic bacteria is the CpxRA two-component system (TCS). We reported that the eltAB bicistronic operon, which encodes for the A and B subunits of LT, was repressed for the CpxRA TCS by direct binding of CpxR-P from -12 to +6 bp with respect to the transcription start site of eltAB. Moreover, the Cpx-response activation down-regulated the transcription of eltAB genes, and this negative effect was CpxRA-dependent. Our data show that CpxRA TCS is a negative regulator of the LT, one of the main virulence determinants of ETEC.

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Burkholderia cepacia complex (Bcc) species are opportunistic pathogens widely distributed in the environment and often infect people with cystic fibrosis (CF). This study aims to determine which genomovars of the Bcc can cause infections in non-CF patients from a tertiary care hospital in Mexico and if they carry virulence factors that could increase their pathogenicity. We identified 23 clinical isolates that carry the recA gene. Twenty-two of them belongs to the genomovar V (B. vietnamiensis) and one to the genomovar II (B. multivorans). Thirteen pulsotypes were identified among 22 B. vietnamiensis isolates. All clinical isolates produced biofilm were motile and cytotoxic on murine macrophage-like RAW264.7 and in A549 human lung epithelial cells. In conclusion, B. vietnamiensis causes infections in non-CF patients in a tertiary care hospital in Mexico, rapid identification of this pathogen can help physicians to establish a better antimicrobial treatment.

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In the public health portfolio of disaster tools, rapid needs assessments are essential intelligence data mining resources that can assess immediate needs in almost all hazard scenarios. Following prolonged and unusual seismic activity that caused significant structural damage, mainly in the southwest part of the island of Puerto Rico, thousands of area residents were forced to leave their homes and establish improvised camps. The austere environmental exposure and limited access to safety and hygiene services prompted public health authorities to request assistance with conducting a rapid needs assessment of those encampments. This report summarizes the design, organization, and execution of a rapid needs assessment of improvised camps following a strong sequence of earthquakes in Puerto Rico.

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The acquisition of Salmonella pathogenicity island 2 (SPI-2) conferred on Salmonella the ability to survive and replicate within host cells. The ssrAB bicistronic operon, located in SPI-2, encodes the SsrAB two-component system (TCS), which is the central positive regulator that induces the expression of SPI-2 genes as well as other genes located outside this island. On the other hand, CpxRA is a two-component system that regulates expression of virulence genes in many bacteria in response to different stimuli that perturb the cell envelope. We previously reported that the CpxRA system represses the expression of SPI-1 and SPI-2 genes under SPI-1-inducing conditions by decreasing the stability of the SPI-1 regulator HilD. Here, we show that under SPI-2-inducing conditions, which mimic the intracellular environment, CpxRA represses the expression of SPI-2 genes by the direct action of phosphorylated CpxR (CpxR-P) on the ssrAB regulatory operon. CpxR-P recognized two sites located proximal and distal from the promoter located upstream of ssrA. Consistently, we found that CpxRA reduces the replication of Salmonella enterica serovar Typhimurium inside murine macrophages. Therefore, our results reveal CpxRA as an additional regulator involved in the intracellular lifestyle of Salmonella, which in turn adds a new layer to the intricate regulatory network controlling the expression of Salmonella virulence genes. IMPORTANCE SPI-2 encodes a type III secretion system (T3SS) that is a hallmark for the species Salmonella enterica, which is essential for the survival and replication within macrophages. Expression of SPI-2 genes is positively controlled by the two-component system SsrAB. Here, we determined a regulatory mechanism involved in controlling the overgrowth of Salmonella inside macrophages. In this mechanism, CpxRA, a two-component system that is activated by extracytoplasmic stress, directly represses expression of the ssrAB regulatory operon; as a consequence, expression of SsrAB target genes is decreased. Our findings reveal a novel mechanism involved in the intracellular lifestyle of Salmonella, which is expected to sense perturbations in the bacterial envelope that Salmonella faces inside host cells, as the synthesis of the T3SS-2 itself.

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El síndrome de vena cava superior es una entidad infrecuente. La etiología neoplásica es la de mayor prevalencia, también la relacionada a los procedimientos invasivos con catéter venosos centrales. Masculino, 32 años, consulta al servicio de emergencias por cianosis facial súbita, opresión en rostro, tos seca, odinofagia, disfonía, vértigos; no refiere disnea. Neoplasia de colón desde 2019, con colostomía y catéter Port subclavio izquierdo. Al examen: edema en esclavina, cianosis central, petequias múltiples, sangrado ungueal en manos. Angiotomografia muestra defecto de llenado por trombosis reciente en venas yugular interna y braquiocefálica izquierdas, braquiocefálica derecha, arco de la ácigos y cava superior en toda su luz. La tromboaspiración mecánica quirúrgica permitió la resección del trombo y restitución de la circulación, con relativa seguridad y baja mortalidad.

Superior vena cava syndrome is a rare entity. The neoplastic etiology is the most relevant, as well as that related to invasive procedures with central venous catheter. A 32-year-old man consults the Emergency Department for sudden facial cyanosis, facial tightness, dry cough, odynophagia, dysphonia and vertigo without dyspnea. He presents colon neoplasia since 2019, with colostomy and left subclavian Port-catheter. At examination, facial and upper extremity edema, central cyanosis, multiple petechiae and nail bleeding on the hands. The angiotomography shows filling defect fort recent thrombosis in left internal jugular and brachiocephalic vein, right brachiocephalic vein, arch of the azygos vein and superior vena cava in its entire lumen. The surgical mechanical thromboaspiration allowed resection of the thrombus and restitution of circulation, with relative safety and low mortality.

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One important event for the divergence of Salmonella from Escherichia coli was the acquisition by horizontal transfer of the Salmonella pathogenicity island 1 (SPI-1), containing genes required for the invasion of host cells by Salmonella. HilD is an AraC-like transcriptional regulator in SPI-1 that induces the expression of the SPI-1 and many other acquired virulence genes located in other genomic regions of Salmonella. Additionally, HilD has been shown to positively control the expression of some ancestral genes (also present in E. coli and other bacteria), including phoH. In this study, we determined that both the gain of HilD and cis-regulatory evolution led to the integration of the phoH gene into the HilD regulon. Our results indicate that a HilD-binding sequence was generated in the regulatory region of the S. enterica serovar Typhimurium phoH gene, which mediates the activation of promoter 1 of this gene under SPI-1-inducing conditions. Furthermore, we found that repression by H-NS, a histone-like protein, was also adapted on the S. Typhimurium phoH gene and that HilD activates the expression of this gene in part by antagonizing H-NS. Additionally, our results revealed that the expression of the S. Typhmurium phoH gene is also activated in response to low phosphate but independently of the PhoB/R two-component system, known to regulate the E. coli phoH gene in response to low phosphate. Thus, our results indicate that cis-regulatory evolution has played a role in the expansion of the HilD regulon and illustrate the phenomenon of differential regulation of ortholog genes. IMPORTANCE Two mechanisms mediating differentiation of bacteria are well known: acquisition of genes by horizontal transfer events and mutations in coding DNA sequences. In this study, we found that the phoH ancestral gene is differentially regulated between Salmonella Typhimurium and Escherichia coli, two closely related bacterial species. Our results indicate that this differential regulation was generated by mutations in the regulatory sequence of the S. Typhimurium phoH gene and by the acquisition by S. Typhimurium of foreign DNA encoding the transcriptional regulator HilD. Thus, our results, together with those from an increasing number of studies, indicate that cis-regulatory evolution can lead to the rewiring and reprogramming of transcriptional regulation, which also plays an important role in the divergence of bacteria through time.

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BACKGROUND: Cystic fibrosis (CF) is a genetic disease in which thick, sticky mucus is produced in the lungs (and other organs) that impairs ciliary clearance, leading to respiratory problems, increased chronic bacterial infections, and decreased lung function. Staphylococcus aureus is one of the primary bacterial pathogens colonizing the lungs of CF patients. This study aimed to characterize the genetic relatedness of S. aureus, its presence in children with CF, and its cytotoxic activity in THP1 cell-derived macrophages (THP1m). METHODS: Genetic relatedness of S. aureus isolates from a cohort of 50 children with CF was determined by pulsed-field gel electrophoresis (PFGE). The VITEK 2 automated system was used to determine antimicrobial susceptibility, and methicillin-resistance S. aureus (MRSA) was determined by diffusion testing using cefoxitin disk. The presence of mecA and lukPV genes was determined by the polymerase chain reaction and cytotoxic activity of S.aureus on THP1m by CytoTox 96® assay. RESULTS: From 51 S. aureus isolates from 50 children with CF, we identified 34pulsotypes by PFGE. Of the 50 children, 12 (24%) were colonized by more than one pulsotype, and 5/34 identified pulsotypes(14.7%) were shared between unrelated children. In addition, 3/34 pulsotypes (8.8%) were multidrug-resistant (MDR), and2/34 (5.9%) were MRSA. Notably, 30/34 pulsotypes (88.2%) exhibited cytotoxicity on THP1m cells and 14/34 (41.2%) alteredTHP1m monolayers. No isolate carried the lukPV gene. CONCLUSIONS: Although a low frequency of MRSA and MDR wasfound among clinical isolates, most of the S. aureus pulsotypes identified were cytotoxic on THP1m.

INTRODUCCIÓN: La fibrosis quística (FQ) es una enfermedad genética en la que se produce moco espeso y pegajoso en los pulmones (y otros órganos), lo que conduce a problemas respiratorios, incremento de las infecciones bacterianas crónicas y disminución de la función pulmonar. Staphylococcus aureus es uno de los principales patógenos que colonizan los pul-mones de los pacientes con FQ. El objetivo de este trabajo fue caracterizar la relación genética de S. aureus, su presencia en niños con FQ y su actividad citotóxica en macrófagos derivados de células THP1 (THP1m). MÉTODOS: La relación gené-tica de los aislados de S. aureus provenientes de una cohorte de 50 pacientes con FQ fue determinada por electroforesis en gel de campo pulsado (PFGE). La sensibilidad a los antimicrobianos se determinó mediante el sistema automatizado VITEK 2, y la resistencia a la meticilina (SARM) mediante la prueba de difusión utilizando discos de cefoxitina. La presen-cia de los genes mecA y lukPV se determinó mediante reacción en cadena de la polimerasa, y la actividad citotóxica de S. aureus sobre células THP1m mediante el ensayo CytoTox96®. RESULTADOS: A partir de 51 aislados de S. aureus provenientes de 50 niños con FQ se identificaron 34 pulsotipos por PFGE. De los 50 niños, 12 (24%) estaban colonizados por más de un pulsotipo y 5 de los 34 pulsotipos (14.7%) los compartían niños que no estaban relacionados. De los 34 pulsotipos, 3 (8.8%) presentaron multirresistencia (MDR) y 2 (5.9%) fueron SARM. Además, 30 pulsotipos (88.2%) fueron citotóxicos sobre células THP1m y 14 (41.2%) alteraron su monocapa. Ninguno de los pulsotipos presentó el gen lukPV. CONCLUSIONES: Aunque se encontró una baja frecuencia de SARM y MDR en los aislados, la mayoría de los pulsotipos de S. aureus identificados fueron citotóxicos para células THP1m.

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