Mutational spectrum of breast cancer by shallow whole-genome sequencing of cfDNA and tumor gene panel analysis.

Breast cancer (BC) has different molecular subgroups related to different risks and treatments. Tumor biopsies for BC detection are invasive and may not reflect tumor heterogeneity. Liquid biopsies have become relevant because they might overcome these limitations. We rationalize that liquid cfDNA biopsies through shallow whole genome sequencing (sWGS) could improve the detection of tumor alterations, complementing the genomic profiling. We evaluated the feasibility to detect somatic copy number alterations (SCNAs) in BC using shallow whole genome sequencing (sWGS) in cfDNA from archived samples from National Cancer Institute of Colombia patients. We sequenced tumor tissues from 38 BC patients with different molecular subtypes using a gene panel of 176 genes significantly mutated in cancer, and by liquid biopsies using sWGS on 20 paired samples to detect SCNAs and compare with the tumor samples. We identified an extensive intertumoral heterogeneity between the molecular subtypes of BC, with a mean tumor load of 602 mutations in the gene panel of tumor tissues. There was a 12.3% of concordance in deletions in the cfDNA-tumor pairs considering only the genes covered by the panel encompassing seven genes: BRCA1, CDK12, NF1, MAP2K4, NCOR1, TP53, and KEAP1 in three patients. This study shows the feasibility to complement the genomic analysis of tumor tissue biopsies to detect SCNA in BC using sWGS in cfDNA, providing a wider identification of potential therapeutic targets.

Genomic landscape of early-stage prostate adenocarcinoma in Mexican patients: an exploratory study.

BACKGROUND: Health disparities have been highlighted among patient with prostate adenocarcinoma (PRAD) due to ethnicity. Mexican men present a more aggressive disease than other patients resulting in less favorable treatment outcome. We aimed to identify the mutational landscape which could help to reduce the health disparities among minority groups and generate the first genomics exploratory study of PRAD in Mexican patients. METHODS: Paraffin-embedded formalin-fixed tumoral tissue from 20 Mexican patients with early-stage PRAD treated at The Instituto Nacional de Cancerología, Mexico City from 2017 to 2019 were analyzed. Tumoral DNA was prepared for whole exome sequencing, the resulting files were mapped against h19 using BWA-MEM. Strelka2 and Lancet packages were used to identify single nucleotide variants (SNV) and insertions or deletions. FACETS was used to determine somatic copy number alterations (SCNA). Cancer Genome Interpreter web interface was used to determine the clinical relevance of variants. RESULTS: Patients were in an early clinical stage and had a mean age of 59.55 years (standard deviation [SD]: 7.1 years) with 90% of them having a Gleason Score of 7. Follow-up time was 48.50 months (SD: 32.77) with recurrences and progression in 30% and 15% of the patients, respectively. NUP98 (20%), CSMD3 (15%) and FAT1 (15%) were the genes most frequently affected by SNV; ARAF (75%) and ZNF419 (70%) were the most frequently affected by losses and gains SNCA's. One quarter of the patients had mutations useful as biomarkers for the use of PARP inhibitors, they comprise mutations in BRCA, RAD54L and ATM. SBS05, DBS03 and ID08 were the most common mutational signatures present in this cohort. No associations with recurrence or progression were identified. CONCLUSIONS: This pilot study reveals the mutational landscape of early-stage prostate adenocarcinoma in Mexican men, providing a first approach to understand the mutational patterns and actionable mutations in early prostate cancer can inform personalized treatment approaches and reduce the underrepresentation in genomic cancer studies.

Analysis of in vitro expression of virulence genes related to antibiotic and disinfectant resistance in Escherichia coli as an emerging periodontal pathogen.

The collective involvement of virulence markers of Escherichia coli as an emerging pathogen associated with periodontitis remains unexplained. This study aimed to implement an in vitro model of infection using a human epithelial cell line to determine the virulome expression related to the antibiotic and disinfectant resistance genotype and pulse field gel electrophoresis (PFGE) type in E. coli strains isolated from patients with periodontal diseases. We studied 100 strains of E. coli isolated from patients with gingivitis (n = 12), moderate periodontitis (n = 59), and chronic periodontitis (n = 29). The identification of E. coli and antibiotic and disinfectant resistance genes was performed through PCR. To promote the expression of virulence genes in the strains, an in vitro infection model was used in the human epithelial cell line A549. RNA was extracted using the QIAcube robotic equipment and reverse transcription to cDNA was performed using the QuantiTect reverse transcription kit (Qiagen). The determination of virulence gene expression was performed through real-time PCR. Overall, the most frequently expressed adhesion genes among the isolated strains of gingivitis, moderate periodontitis, and chronic periodontitis were fimH (48%), iha (37%), and papA (18%); those for toxins were usp (33%); those for iron acquisition were feoB (84%), fyuA (62%), irp-2 (61%), and iroN (35%); those for protectins were traT (50%), KpsMT (35%), and ompT (28%); and those for pathogenicity islands were malX (45%). The most common antibiotic and disinfectant resistance genes among gingivitis, moderate periodontitis, and chronic periodontitis strains were sul-2 (43%), blaSHV (47%), blaTEM (45%), tet(A) (41%), dfrA1 (32%), marR-marO (57%), and qacEA1 (79%). The findings revealed the existence of a wide distribution of virulome expression profiles related to the antibiotic and disinfectant resistance genotype and PFGE type in periodontal strains of E. coli. These findings may contribute toward improving the prevention and treatment measures for periodontal diseases associated with E. coli.

Methylation marks in blood DNA reveal breast cancer risk in patients fulfilling hereditary disease criteria.

Less than 15-20% of patients who meet the criteria for hereditary breast and ovarian cancer (HBOC) carry pathogenic coding genetic mutations, implying that other molecular mechanisms may contribute to the increased risk of this condition. DNA methylation in peripheral blood has been suggested as a potential epigenetic marker for the risk of breast cancer (BC). We aimed to discover methylation marks in peripheral blood associated with BC in 231 pre-treatment BC patients meeting HBOC criteria, testing negative for coding pathogenic variants, and 156 healthy controls, through methylation analysis by targeted bisulfite sequencing on 18 tumor suppressor gene promoters (330 CpG sites). We found i) hypermethylation in EPCAM (17 CpG sites; p = 0.017) and RAD51C (27 CpG sites; p = 0.048); ii) hypermethylation in 36 CpG-specific sites (FDR q < 0.05) in the BC patients; iii) four specific CpG sites were associated with a higher risk of BC (FDR q < 0.01, Bonferroni p < 0.001): cg89786999-FANCI (OR = 1.65; 95% CI:1.2-2.2), cg23652916-PALB2 (OR = 2.83; 95% CI:1.7-4.7), cg47630224-MSH2 (OR = 4.17; 95% CI:2.1-8.5), and cg47596828-EPCAM (OR = 1.84; 95% CI:1.5-2.3). Validation of cg47630224-MSH2 methylation in one Australian cohort showed an association with 3-fold increased BC risk (AUC: 0.929; 95% CI: 0.904-0.955). Our findings suggest that four DNA methylation CpG sites may be associated with a higher risk of BC, potentially serving as biomarkers in patients without detectable coding mutations.

Impact of STAT-signaling pathway on cancer-associated fibroblasts in colorectal cancer and its role in immunosuppression.

Colorectal cancer (CRC) remains one of the most commonly diagnosed and deadliest types of cancer worldwide. CRC displays a desmoplastic reaction (DR) that has been inversely associated with poor prognosis; less DR is associated with a better prognosis. This reaction generates excessive connective tissue, in which cancer-associated fibroblasts (CAFs) are critical cells that form a part of the tumor microenvironment. CAFs are directly involved in tumorigenesis through different mechanisms. However, their role in immunosuppression in CRC is not well understood, and the precise role of signal transducers and activators of transcription (STATs) in mediating CAF activity in CRC remains unclear. Among the myriad chemical and biological factors that affect CAFs, different cytokines mediate their function by activating STAT signaling pathways. Thus, the harmful effects of CAFs in favoring tumor growth and invasion may be modulated using STAT inhibitors. Here, we analyze the impact of different STATs on CAF activity and their immunoregulatory role.

Helminth-derived molecules improve 5-fluorouracil treatment on experimental colon tumorigenesis.

Colorectal cancer (CRC) is one of the most prevalent fatal neoplasias worldwide. Despite efforts to improve the early diagnosis of CRC, the mortality rate of patients is still nearly 50%. The primary treatment strategy for CRC is surgery, which may be accompanied by chemotherapy and radiotherapy. The conventional and first-line chemotherapeutic agent utilized is 5-fluorouracil (5FU). However, it has low efficiency. Combination treatment with leucovorin and oxaliplatin or irinotecan improves the effectiveness of 5FU therapy. Unfortunately, most patients develop drug resistance, leading to disease progression. Here, we evaluated the effect of a potential alternative adjuvant treatment for 5FU, helminth-derived Taenia crassiceps (TcES) molecules, on treating advanced colitis-associated colon cancer. The use of TcES enhanced the effects of 5FU on established colonic tumors by downregulating the expression of the immunoregulatory cytokines, Il-10 and Tgf-ß, and proinflammatory cytokines, Tnf-α and Il-17a, and reducing the levels of molecular markers associated with malignancy, cyclin D1, and Ki67, both involved in apoptosis inhibition and the signaling pathway of ß-catenin. TcES+5FU therapy promoted NK cell recruitment and the release of Granzyme B1 at the tumor site, consequently inducing tumor cell death. Additionally, it restored P53 activity which relates to decreased Mdm2 expression. In vitro assays with human colon cancer cell lines showed that therapy with TcES+5FU significantly reduced cell proliferation and migration by modulating the P53 and P21 signaling pathways. Our findings demonstrate, for the first time in vivo, that helminth-derived excreted/secreted products may potentiate the effect of 5FU on established colon tumors.

Complete mitochondrial genome of Gnathostoma binucleatum.

This report describes the mitochondrial genome of the parasite Gnathostoma binucleatum (G. binucleatum), which was obtained from naturally infected freshwater fish in Sinaloa, Mexico (22°46'00.1″N 105°40'21.8″W). G. binucleatum is responsible for human gnathostomiasis and is endemic to Mexico. It belongs to the Spirurida order of the Secernentea class of Nematoda.

Extensive Expression of the Virulome Related to Antibiotic Genotyping in Nosocomial Strains of Klebsiella pneumoniae.

The emergence of hyper-virulent and multidrug-resistant (MDR) strains of Klebsiella pneumoniae isolated from patients with hospital- and community-acquired infections is a serious health problem that increases mortality. The molecular analysis of virulome expression related to antimicrobial-resistant genotype and infection type in K. pneumoniae strains isolated from patients with hospital- and community-acquired infections has been poorly studied. In this study, we analyzed the overall expression of the virulence genotype associated with the antimicrobial resistance genotype and pulse field gel electrophoresis (PFGE) type (PFtype) in K. pneumoniae. We studied 25 strains of K. pneumoniae isolated from patients who developed bacteremia and pneumonia during their hospital stay and 125 strains from outpatients who acquired community-acquired infections. Susceptibility to 12 antimicrobials was determined by Kirby-Bauer. The identification of K. pneumoniae and antibiotic-resistance genes was performed using polymerase chain reaction (PCR). To promote the expression of the virulence genes of K. pneumoniae, an in vitro infection model was used in human epithelial cell lines A549 and A431. Bacterial RNA was extracted with the QIAcube robotic workstation, and reverse transcription to cDNA was performed with the Reverse Transcription QuantiTect kit (Qiagen). The determination of the expression of the virulence genes was performed by real-time PCR. In addition, 57.3% (n = 86) of the strains isolated from patients with hospital- and community-acquired infections were multidrug-resistant (MDR), mainly to beta-lactam antibiotics (CB, AM, CFX, and CF), aminoglycosides (GE), quinolones (CPF and NOF), nitrofurantoin (NF), and sulfamethoxazole/trimethoprim (SXT). The most frequently expressed genes among strains isolated from hospital- and community-acquired infections were adhesion-type, ycfm (80%), mrkD (51.3%), and fimH (30.7%); iron uptake, irp2 (84%), fyuA (68.7%), entB (64.7%), and irp1 (56.7%); and protectins, rpmA (26%), which were related to antibiotic-resistance genes, blaTEM (96%), blaSHV (64%), blaCITM (52.6%), blaCTXM-1 (44.7%), tetA (74%), sul1 (57.3%), aac(3)-IV (40.7%), and aadA1 (36%). The results showed the existence of different patterns of expression of virulome related to the genotype of resistance to antimicrobials and to the PFtypes in the strains of K. pneumoniae that cause hospital- and community-acquired infections. These findings are important and may contribute to improving medical treatment strategies against infections caused by K. pneumoniae.

Molecular transition of SARS-CoV-2 from critical patients during the first year of the COVID-19 pandemic in Mexico City.

Background: The SARS-CoV-2 virus has caused unprecedented mortality since its emergence in late 2019. The continuous evolution of the viral genome through the concerted action of mutational forces has produced distinct variants that became dominant, challenging human immunity and vaccine development. Aim and methods: In this work, through an integrative genomic approach, we describe the molecular transition of SARS-CoV-2 by analyzing the viral whole genome sequences from 50 critical COVID-19 patients recruited during the first year of the pandemic in Mexico City. Results: Our results revealed differential levels of the evolutionary forces across the genome and specific mutational processes that have shaped the first two epidemiological waves of the pandemic in Mexico. Through phylogenetic analyses, we observed a genomic transition in the circulating SARS-CoV-2 genomes from several lineages prevalent in the first wave to a dominance of the B.1.1.519 variant (defined by T478K, P681H, and T732A mutations in the spike protein) in the second wave. Conclusion: This work contributes to a better understanding of the evolutionary dynamics and selective pressures that act at the genomic level, the prediction of more accurate variants of clinical significance, and a better comprehension of the molecular mechanisms driving the evolution of SARS-CoV-2 to improve vaccine and drug development.

Phylogenomics and Biogeography of the Mammilloid Clade Revealed an Intricate Evolutionary History Arose in the Mexican Plateau.

Mexico harbors ~45% of world's cacti species richness. Their biogeography and phylogenomics were integrated to elucidate the evolutionary history of the genera Coryphantha, Escobaria, Mammillaria, Mammilloydia, Neolloydia, Ortegocactus, and Pelecyphora (Mammilloid Clade). We analyzed 52 orthologous loci from 142 complete genomes of chloroplast (103 taxa) to generate a cladogram and a chronogram; in the latter, the ancestral distribution was reconstructed with the Dispersal-Extinction-Cladogenesis model. The ancestor of these genera arose ~7 Mya on the Mexican Plateau, from which nine evolutionary lineages evolved. This region was the site of 52% of all the biogeographical processes. The lineages 2, 3 and 6 were responsible for the colonization of the arid southern territories. In the last 4 Mya, the Baja California Peninsula has been a region of prolific evolution, particularly for lineages 8 and 9. Dispersal was the most frequent process and vicariance had relevance in the isolation of cacti distributed in the south of Mexico. The 70 taxa sampled as Mammillaria were distributed in six distinct lineages; one of these presumably corresponded to this genus, which likely had its center of origin in the southern part of the Mexican Plateau. We recommend detailed studies to further determine the taxonomic circumscription of the seven genera.

Evaluation of genetic alterations in hereditary cancer susceptibility genes in the Ashkenazi Jewish women community of Mexico.

Background: Individuals of Ashkenazi Jewish ancestry have been identified as having higher prevalence of specific pathogenic variants associated with susceptibility to specific rare and chronic diseases. In Mexico, the prevalence and composition of rare cancer predisposing germline variants in Ashkenazi Jewish individuals has not been evaluated. Aim and methods: We aimed to evaluate the prevalence of pathogenic variants by massive parallel sequencing in a panel of 143 cancer-predisposing genes in 341 women from the Ashkenazi Jewish community of Mexico, who were contacted and invited to participate in the study through the ALMA Foundation for Cancer Reconstruction. Pre- and posttest genetic counseling was given and a questionnaire on personal, gyneco-obstetric, demographic and lifestyle variables was conducted. From peripheral blood DNA, the complete coding region, and splicing sites of a panel of 143 cancer susceptibility genes, including 21 clinically relevant genes, were sequenced. The Mexican founder mutation BRCA1 ex9-12del [NC_000017.10(NM_007294):c. (825+1-826-1)_(4,589+1-4,590-1)del] was also evaluated. Results: Among study participants (mean age ±standard deviation: 47 ± 14) 15% reported a personal history of cancer (50/341). Fourteen percent of participants (48/341) were carriers of pathogenic and likely pathogenic variants distributed among seven high-risk genes (APC, CHEK2, MSH2, BMPR1A, MEN1, MLH1, and MSH6), whereas 18.2% (62/341) had variants of uncertain clinical significance in genes associated with breast and ovarian cancer susceptibility (list of genes with VUS). Pathogenic and likely pathogenic variants in 16 susceptibility genes with ambiguous or non-well-established risk association for cancer were detected in 17.6% (60/341) of participants. Sixty four percent of participants reported current alcohol consumption compared with the 39 percent prevalence of alcohol consumption in Mexican women. None of the participants carried the recurrent Ashkenazi and Mexican founder mutations in BRCA1 or BRCA2, but 2% (7/341) had pathogenic Ashkenazi Jewish founder variants in BLM. Conclusion: Our findings show a diverse pathogenic variant composition among the recruited individuals of Ashkenazi Jewish ancestry in Mexico consistent with being a high-risk population for genetic diseases, which warrants further investigation to adequately assess the burden of hereditary breast cancer in this group and implement appropriate preventative programs.

A Maverick-like cluster in the genome of a pathogenic, moderately virulent strain of Gallibacterium anatis, ESV200, a transient biofilm producer.

Introduction: Gallibacterium anatis causes gallibacteriosis in birds. These bacteria produce biofilms and secrete several fimbrial appendages as tools to cause disease in animals. G. anatis strains contain up to three types of fimbriae. Complete genome sequencing is the strategy currently used to determine variations in the gene content of G. anatis, although today only the completely circularized genome of G. anatis UMN179 is available. Methods: The appearance of growth of various strains of G. anatis in liquid culture medium was studied. Biofilm production and how the amount of biofilm was affected by DNase, Proteinase K, and Pronase E enzymes were analyzed. Fimbrial gene expression was performed by protein analysis and qRT-PCR. In an avian model, the pathogenesis generated by the strains G. anatis ESV200 and 12656-12 was investigated. Using bioinformatic tools, the complete genome of G. anatis ESV200 was comparatively studied to search for virulence factors that would help explain the pathogenic behavior of this strain. Results and Discussion: G. anatis ESV200 strain differs from the 12656-12 strain because it produces a biofilm at 20%. G. anatis ESV200 strain express fimbrial genes and produces biofilm but with a different structure than that observed for strain 12656-12. ESV200 and 12656-12 strains are pathogenic for chickens, although the latter is the most virulent. Here, we show that the complete genome of the ESV200 strain is similar to that of the UNM179 strain. However, these strains have evolved with many structural rearrangements; the most striking chromosomal arrangement is a Maverick-like element present in the ESV200 strain.

Mexican Colorectal Cancer Research Consortium (MEX-CCRC): Etiology, Diagnosis/Prognosis, and Innovative Therapies.

In 2013, recognizing that Colorectal Cancer (CRC) is the second leading cause of death by cancer worldwide and that it was a neglected disease increasing rapidly in Mexico, the community of researchers at the Biomedicine Research Unit of the Facultad de Estudios Superiores Iztacala from the Universidad Nacional Autónoma de México (UNAM) established an intramural consortium that involves a multidisciplinary group of researchers, technicians, and postgraduate students to contribute to the understanding of this pathology in Mexico. This article is about the work developed by the Mexican Colorectal Cancer Research Consortium (MEX-CCRC): how the Consortium was created, its members, and its short- and long-term goals. Moreover, it is a narrative of the accomplishments of this project. Finally, we reflect on possible strategies against CRC in Mexico and contrast all the data presented with another international strategy to prevent and treat CRC. We believe that the Consortium's characteristics must be maintained to initiate a national strategy, and the reported data could be useful to establish future collaborations with other countries in Latin America and the world.

Mutational Landscape of Bladder Cancer in Mexican Patients: KMT2D Mutations and chr11q15.5 Amplifications Are Associated with Muscle Invasion.

Bladder cancer (BC) is the most common neoplasm of the urinary tract, which originates in the epithelium that covers the inner surface of the bladder. The molecular BC profile has led to the development of different classifications of non-muscle invasive bladder cancer (NMIBC) and muscle-invasive bladder cancer (MIBC). However, the genomic BC landscape profile of the Mexican population, including NMIBC and MIBC, is unknown. In this study, we aimed to identify somatic single nucleotide variants (SNVs) and copy number variations (CNVs) in Mexican patients with BC and their associations with clinical and pathological characteristics. We retrospectively evaluated 37 patients treated between 2012 and 2021 at the National Cancer Institute-Mexico (INCan). DNA samples were obtained from paraffin-embedded tumor tissues and exome sequenced. Strelka2 and Lancet packages were used to identify SNVs and insertions or deletions. FACETS was used to determine CNVs. We found a high frequency of mutations in TP53 and KMT2D, gains in 11q15.5 and 19p13.11-q12, and losses in 7q11.23. STAG2 mutations and 1q11.23 deletions were also associated with NMIBC and low histologic grade.

Deep DNA sequencing of MGMT, TP53 and AGT in Mexican astrocytoma patients identifies an excess of genetic variants in women and a predictive biomarker.

PURPOSE: Astrocytomas are a type of malignant brain tumor with an unfavorable clinical course. The impact of AGT and MGMT somatic variants in the prognosis of astrocytoma is unknown, and it is controversial for TP53. Moreover, there is a lack of knowledge regarding the molecular characteristics of astrocytomas in Mexican patients. METHODS: We studied 48 Mexican patients, men and women, with astrocytoma (discovery cohort). We performed DNA deep sequencing in tumor samples, targeting AGT, MGMT and TP53, and we studied MGMT gene promoter methylation status. Then we compared our findings to a cohort which included data from patients with astrocytoma from The Cancer Genome Atlas (validation cohort). RESULTS: In the discovery cohort, we found a higher number of somatic variants in AGT and MGMT than in the validation cohort (10.4% vs < 1%, p < 0.001), and, in both cohorts, we observed only women carried variants AGT variants. We also found that the presence of either MGMT variant or promoter methylation was associated to better survival and response to chemotherapy, and, in conjunction with TP53 variants, to progression-free survival. CONCLUSIONS: The occurrence of AGT variants only in women expands our knowledge about the molecular differences in astrocytoma between men and women. The increased prevalence of AGT and MGMT variants in the discovery cohort also points towards possible distinctions in the molecular landscape of astrocytoma among populations. Our findings warrant further study.

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

Background: Bacteriophage 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. Methods: In 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. Results: Phage 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. Conclusions: We 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.

Dual activation profile of monocytes is associated with protection in Mexican patients during SARS-CoV-2 disease.

The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been one of the most catastrophic diseases observed in recent years. It has reported nearly 550 million cases worldwide, with more than 6.35 million deaths. In Mexico, an increased incidence and mortality of this disease were observed, where the immune response has been involved in the magnitude and severity. A critical version of the disease is accompanied by hyperinflammatory responses, with cytokine and defective cellular responses. A detailed understanding of the role of molecules and cells in the immune response during COVID-19 disease may help to generate effective protection mechanisms, improving those we already have. Here we analyzed blood samples obtained from patients at the Hospital Regional de Alta Especialidad de Ixtapaluca (HRAEI), Mexico, which were classified according to living guidance for clinical management of COVID-19 by the World Health Organization: asymptomatic, mild, severe, and critical disease. We observed increased interleukin (IL)-6 levels and a T-CD8+ and T-CD4+ cell reduction correlated with the critical disease version. Importantly, here, we described a significant reduction of CD11b+CD45highCD14low monocytes during severe disease, which displayed a non-classical profile, expressing IL-10, transforming growth factor (TGF)-ß, and indoleamine 2,3-dioxygenase (IDO)1 molecule. Moreover, CD11b+CD45highCD14low monocytes obtained from infected one-dose vaccinated patients (Pfizer® vaccine) who suffered minimal symptoms showed simultaneously a dual classical and no-classical profile expressing pro- and anti-inflammatory cytokines. These results suggest that blood monocytes expressing a dual pro- and anti-inflammatory profile might be a predictive marker for protection in the Mexican population during COVID-19 disease. KEY POINTS : • Exacerbated immune response is associated with COVID-19 severe disease. • Dual monocyte activation profile is crucial for predicting protection during COVID-19. • Vaccination is crucial to induce the dual activation profile in monocytes.

Hypervirulence and Multiresistance to Antibiotics in Klebsiella pneumoniae Strains Isolated from Patients with Hospital- and Community-Acquired Infections in a Mexican Medical Center.

Klebsiella pneumoniae is a pathogenic bacterium associated with different infectious diseases. This study aimed to establish the different association profiles of virulence genes related to the hypermucoviscous phenotype (HM), capsular serotypes, biofilm formation, and multidrug resistance in K. pneumoniae strains from patients with hospital- and community-acquired infections. K. pneumoniae virulence genes and capsular serotypes were identified by PCR, antibiotic susceptibility by the Kirby-Bauer method, HM by the string test, and biofilm formation by measurement in polystyrene microtiter plates. Of a total of 150 strains from patients with hospital- (n = 25) and community-acquired infections (n = 125), 53.3% (80/150) were HM-positive and 46.7% (70/150) were HM-negative. HM-positive (68/80) and HM-negative (67/70) strains were biofilm-forming. Moreover, 58.7% (47/80) HM-positive and 57.1% (40/70) HM-negative strains were multidrug-resistant. Among HM-positive, HM-negative, and serotypes K1 (25/150), K2 (48/150), and non-K1/K2 strains, (77/150) the frequently detected adhesion genes were fimH, mrkD, ycfM, and kpn; entB, irp2, irp1, and ybtS, for iron acquisition; and rmpA for protectins. The gene association pattern fimH/kpn/mrkD/ycfM/entB/irp1/irp2/ybtS/fyuA (18/150) was frequent among the strains. K. pneumoniae strains from patients with hospital- and community-acquired infections demonstrated a wide diversity of virulence gene profiles related to phenotype (hypermucoviscosity, multidrug resistance, and biofilm formation) and serotypes.

Early detection of colorectal cancer somatic mutations using cfDNA liquid biopsies in a murine carcinogenesis model.

Colorectal cancer (CRC) is one of the top five cancers in incidence and mortality worldwide. The early detection of this neoplasm through analysis of circulating free DNA (cfDNA), which carries tumor genetic alterations, as a liquid biopsy, could have a major impact in enhancing early detection and reducing the mortality rate. The aim of this work was to demonstrate the feasibility of using cfDNA as a liquid biopsy for the early detection of CRC. For this purpose, we implemented an azoxymethane and dextran sodium sulfate-induced murine carcinogenesis model to detect oncogenic somatic mutations in Ctnnb1 and Kras during CRC development. To enhance the sensitivity in the detection, E-ice-COLD-PCR was utilized to selectively enrich for mutant alleles, followed by massively parallel sequencing. Driving somatic mutations were detected in Ctnnb1 and Kras in the liquid biopsies of early stages of tumor development, corresponding to the formation of aberrant crypt foci, the first histological alterations that can be identified throughout the formation of CRC. The concentration of cfDNA was increased along the carcinogenic process. Polyclonality in Ctnnb1 was found in tumor samples and cfDNA in this model. On the other hand, the use of cfDNA as a non-invasive test resulted in superior early detection compared to microPET/CT imaging. As a proof-of-principle, this study shows the great potential use of allelic-specific PCR for the detection and enrichment of pathogenic alleles present in cfDNA samples, as a test for early non-invasive detection of CRC. This work provides scientific evidence to set methodological bases that allow early detection of mutations in cfDNA obtained from plasma of CRC in humans.

Variability in Susceptibility to Type I Interferon Response and Subgenomic RNA Accumulation Between Clinical Isolates of Dengue and Zika Virus From Oaxaca Mexico Correlate With Replication Efficiency in Human Cells and Disease Severity.

Dengue and Zika viruses cocirculate annually in endemic areas of Mexico, causing outbreaks of different magnitude and severity every year, suggesting a continuous selection of Flavivirus variants with variable phenotypes of transmissibility and virulence. To evaluate if Flavivirus variants with different phenotypes cocirculate during outbreaks, we isolated dengue and Zika viruses from blood samples of febrile patients from Oaxaca City during the 2016 and 2019 epidemic years. We compared their replication kinetics in human cells, susceptibility to type I interferon antiviral response, and the accumulation of subgenomic RNA on infected cells. We observed correlations between type I interferon susceptibility and subgenomic RNA accumulation, with high hematocrit percentage and thrombocytopenia. Our results suggest that Flaviviruses that cocirculate in Oaxaca, Mexico, have variable sensitivity to the antiviral activity of type I interferons, and this phenotypic trait correlates with the severity of the disease.