Molecular Determinants Of Resistance Research Articles

Current Status of Malaria Control and Elimination in Africa: Epidemiology, Diagnosis, Treatment, Progress and Challenges.

The African continent carries the greatest malaria burden in the world. Falciparum malaria especially has long been the leading cause of death in Africa. Climate, economic factors, geographical location, human intervention and unstable security are factors influencing malaria transmission. Due to repeated infections and early interventions, the proportion of clinically atypical malaria or asymptomatic plasmodium carriers has increased significantly, which easily lead to misdiagnosis and missed diagnosis. African countries have made certain progress in malaria control and elimination, including rapid diagnosis of malaria, promotion of mosquito nets and insecticides, intermittent prophylactic treatment in high-risk groups, artemisinin based combination therapies, and the development of vaccines. Between 2000 and 2022, there has been a 40% decrease in malaria incidence and a 60% reduction in mortality rate in the WHO African Region. However, many challenges are emerging in the fight against malaria in Africa, such as climate change, poverty, substandard health services and coverage, increased outdoor transmission and the emergence of new vectors, and the growing threat of resistance to antimalarial drugs and insecticides. Joint prevention and treatment, identifying molecular determinants of resistance, new drug development, expanding seasonal malaria chemo-prevention intervention population, and promoting the vaccination of RTS, S/AS01 and R21/Matrix-M may help to solve the dilemma. China's experience in eliminating malaria is conducive to Africa's malaria prevention and control, and China-Africa cooperation needs to be constantly deepened and advanced. Our review aims to help the global public develop a comprehensive understanding of malaria in Africa, thereby contributing to malaria control and elimination.

Molecular Profiling of Response and Resistance to Venetoclax-Decitabine Therapy in Acute Myeloid Leukemia

The hypomethylating agent decitabine and the BCL-2 inhibitor venetoclax combination (DEC-VEN) is now standard of care in AML, yet a subset of patients remains refractory, and factors that reduce efficacy remain to be fully elucidated. This study aimed to systematically identify molecular determinants of resistance via single-cell (sc)RNA-seq and scATAC-seq analysis of bone marrow (BM) samples from patients participating in a prospective clinical trial of DEC-VEN (NCT03404193). For scRNA-seq, 17 BM samples (total 39,607 cells) from 12 AML patients (median age 67; 8 responders; 4 non-responders; 5 pre- and post-treatment pairs) and 3 healthy BM controls were integrated and annotated into 23 transcriptional cell clusters. In the Uniform Manifold Approximation and Projection (UMAP) plot, C1 overlapped with healthy hematopoietic stem cells (HSCs), C2 appeared only in AML patients, and C3 overlapped with progenitor cells. C1 was characteristic of refractory AML and persisted after DEC-VEN treatment. (Figure 1) C1 cells showed significantly higher expression of FLT3, BCL2, MCL1 and transcription factors MYB, ETV6, RUNX1, LMO2, JUND compared to HSCs from healthy donors. DEC-VEN treatment markedly increased ribosomal protein ( RP ) genes and mitochondrial electron transfer COX family genes and increased expression of the leukemia stem cell (LSC) marker CD74 in C1 of non-responders. In C2 cells, high expression of LSC marker genes FLT3, PIM1, MYC and transforming master regulator genes MYB, ETV6, RUNX2, LMO2, STAT3, NOTCH1, BCL11A was observed. DEC-VEN significantly upregulated RP genes and CD74 in C2 in non-responders, as in C1. In responders, hemoglobin family genes were significantly increased in C2. C3 cells were characterized by highly expressed iron metabolism genes, which were further increased by DEC-VEN in responders. GSEA analysis demonstrated that DEC-VEN upregulated ATP metabolic processes and ribosomal biosynthesis and downregulated stem cell differentiation genes in refractory C1 and C2. In responders' AML clusters, DEC-VEN upregulated cellular oxidant detoxification and iron ion homeostasis and downregulated myeloid leukocyte activation. Next, we extended the AML transcriptome analysis by profiling accessible chromatin by scATAC-seq using BM from pre- and post- DEC -VEN treatment of 2 AML patients (1 refractory, 1 relapsed after remission with incomplete count recovery [CRi]), and 2 healthy controls (total 41,062 cells). Leukemia-like cluster (LLC) cells showed significantly higher chromatin accessibility of CD74, RUNX1, RUNX2, NOTCH1, LMO2, ETV6, JUND, STAT3, BCL2, BCL11A and enriched binding motifs of RUNX2, ETV6, MYB, JUN, JUND, STAT3 compared to other clusters, which was consistent with gene expression findings of C1 and C2 by scRNA-seq. DEC-VEN treatment induced further increased motif enrichment of STAT3 in LLC in CRi AML, and STAT3 RUNX2, IKZF1 in refractory AML. To interpret T cell kinetics in response to DEC-VEN in AML, T cells from scRNA-seq profiling were subclassified based on canonical gene expression. The results showed that the CD3+ and CD3+CD8+ cell signatures were significantly lower in non-responders compared to responders post-treatment (p<0.005). Notably, the ratio of exhausted CD8+ cell clusters was significantly higher in non-responders than in responders in DEC-VEN post-treatment (p<0.05), with downregulation of CD8+ effector marker GZMK, and regulator of CD8+ cytotoxic T cell development TOX. In summary, we identified a subpopulation of AML cells that overlap with HSCs in UMAP plots and are associated with DEC-VEN resistance. LSC-related genes were enriched in resistant AML clusters, while hemoglobin-related genes were increased in sensitive clusters. DEC-VEN induced mitochondrial metabolism genes in resistant cells and further enhanced erythrocytic differentiation in sensitive cells. In addition, scATAC-seq revealed LSC-specific chromatin accessibility patterns, and depletion of CD8+ T cells was observed in non-responders. These findings shed light on the molecular mechanisms of DEC-VEN resistance.

Genomic analysis of Streptococcus pneumoniae serogroup 20 isolates in Alberta, Canada from 1993-2019.

In the province of Alberta, Canada, invasive disease caused by Streptococcus pneumoniae serogroup 20 (serotypes 20A/20B) has been increasing in incidence. Here, we characterize provincial invasive serogroup 20 isolates collected from 1993 to 2019 alongside invasive and non-invasive serogroup 20 isolates from the Global Pneumococcal Sequencing (GPS) Project collected from 1998 to 2015. Trends in clinical metadata and geographic location were evaluated, and serogroup 20 isolate genomes were subjected to molecular sequence typing, virulence and antimicrobial resistance factor mining, phylogenetic analysis and pangenome calculation. Two hundred and seventy-four serogroup 20 isolates from Alberta were sequenced, and analysed along with 95 GPS Project genomes. The majority of invasive Alberta serogroup 20 isolates were identified after 2007 in primarily middle-aged adults and typed predominantly as ST235, a sequence type that was rare among GPS Project isolates. Most Alberta isolates carried a full-length whaF capsular gene, suggestive of serotype 20B. All Alberta and GPS Project genomes carried molecular resistance determinants implicated in fluoroquinolone and macrolide resistance, with a few Alberta isolates exhibiting phenotypic resistance to azithromycin, clindamycin, erythromycin, tetracycline and trimethoprim-sulfamethoxazole, as well as non-susceptibility to tigecycline. All isolates carried multiple virulence factors including those involved in adherence, immune modulation and nutrient uptake, as well as exotoxins and exoenzymes. Phylogenetically, Alberta serogroup 20 isolates clustered with predominantly invasive GPS Project isolates from the USA, Israel, Brazil and Nepal. Overall, this study highlights the increasing incidence of invasive S. pneumoniae serogroup 20 disease in Alberta, Canada, and provides insights into the genetic and clinical characteristics of these isolates within a global context.

A Review of Resistance to Polymyxins and Evolving Mobile Colistin Resistance Gene (mcr) among Pathogens of Clinical Significance.

The global rise in antibiotic resistance in bacteria poses a major challenge in treating infectious diseases. Polymyxins (e.g., polymyxin B and colistin) are last-resort antibiotics against resistant Gram-negative bacteria, but the effectiveness of polymyxins is decreasing due to widespread resistance among clinical isolates. The aim of this literature review was to decipher the evolving mechanisms of resistance to polymyxins among pathogens of clinical significance. We deciphered the molecular determinants of polymyxin resistance, including distinct intrinsic molecular pathways of resistance as well as evolutionary characteristics of mobile colistin resistance. Among clinical isolates, Acinetobacter stains represent a diversified evolution of resistance, with distinct molecular mechanisms of intrinsic resistance including naxD, lpxACD, and stkR gene deletion. On the other hand, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa are usually resistant via the PhoP-PhoQ and PmrA-PmrB pathways. Molecular evolutionary analysis of mcr genes was undertaken to show relative relatedness across the ten main lineages. Understanding the molecular determinants of resistance to polymyxins may help develop suitable and effective methods for detecting polymyxin resistance determinants and the development of novel antimicrobial molecules.

Decoupled neoantigen cross-presentation by dendritic cells limits anti-tumor immunity against tumors with heterogeneous neoantigen expression.

Cancer immunotherapies, in particular checkpoint blockade immunotherapy (CBT), can induce control of cancer growth, with a fraction of patients experiencing durable responses. However, the majority of patients currently do not respond to CBT and the molecular determinants of resistance have not been fully elucidated. Mounting clinical evidence suggests that the clonal status of neoantigens (NeoAg) impacts the anti-tumor T cell response. High intratumor heterogeneity (ITH), where the majority of NeoAgs are expressed subclonally, is correlated with poor clinical response to CBT and poor infiltration with tumor-reactive T cells. However, the mechanism by which ITH blunts tumor-reactive T cells is unclear. We developed a transplantable murine lung cancer model to characterize the immune response against a defined set of NeoAgs expressed either clonally or subclonally to model low or high ITH, respectively. Here we show that clonal expression of a weakly immunogenic NeoAg with a relatively strong NeoAg increased the immunogenicity of tumors with low but not high ITH. Mechanistically we determined that clonal NeoAg expression allowed cross-presenting dendritic cells to acquire and present both NeoAgs. Dual NeoAg presentation by dendritic cells was associated with a more mature DC phenotype and a higher stimulatory capacity. These data suggest that clonal NeoAg expression can induce more potent anti-tumor responses due to more stimulatory dendritic cell:T cell interactions. Therapeutic vaccination targeting subclonally expressed NeoAgs could be used to boost anti-tumor T cell responses.

Genomic and Antimicrobial Surveillance of Campylobacter Population in Italian Poultry.

Campylobacter is one of the most common foodborne diseases worldwide with increasing rates of antibiotic resistance. Most cases of campylobacteriosis can be traced back to the consumption of poultry meat. Despite many efforts to reduce contamination in farms and in slaughterhouses, the persistence of this pathogen in poultry products remains a problem. This study aimed to evaluate the genetic diversity and antibiotic resistance of 542 C. jejuni and C. coli in Italian poultry, in the framework of two National Monitoring Programs. Genomes were screened for antibiotic resistance, virulence determinants and contextualized within a global collection of C. jejuni. ST2116, ST2863 and ST 832 were the most prevalent and significantly associated with Italian poultry. A worrying increase in resistance to quinolones, fluoroquinolones and tetracycline was observed in C. jejuni, while an increased occurrence of multidrug resistant (MDR) strains and strains resistant to macrolides was detected in C. coli. Low resistance rates were found for aminoglycosides. Molecular resistance determinants were consistent with the phenotypic resistance for tetracycline and quinolones. In silico analysis revealed 119 genes associated with virulence factors, with a notably higher prevalence of some genes in ST2863 genomes. This study highlights the increased resistance to macrolides and the emergence of MDR strains for C. coli, the genetic basis of AMR and the predominance of two genotypes among Campylobacter strains isolated from the Italian poultry farms.

Perchlozone Resistance in Clinical Isolates of Mycobacterium tuberculosis

The emergence of drug-resistant tuberculosis forced the development of new drugs and the screening of more effective or less toxic analogues. Mycolic acid biosynthesis is targeted by several antituberculosis drugs, isoniazid being one of the most important in tuberculosis therapy. Recently, perchlozone, acting on another step in the FAS-II cycle, was officially approved for tuberculosis treatment in the Russian Federation and was included in the Russian national clinical guidelines. Using the serial dilution method on 7H10 agar plates for perchlozone and a Sensititre MYCOTB microdilution plate, we analyzed the phenotypic properties of primary clinical isolates of M. tuberculosis and analyzed the molecular determinants of resistance to isoniazid, ethionamide, and perchlozone. We found a wide variation in the MIC of perchlozone from 2 to 64 mg/L, correlating with the overall resistance profile: the MIC was higher for MDR and pre-XDR isolates. The cross-resistance between ethionamide and perchlozone was driven by mutations in the ethA gene encoding monooxygenase responsible for the activation of both drugs. The presumably susceptible to perchlozone and wild-type strains had MICs ranging from 2 to 4 mg/L, and the breakpoint was estimated to be 4 or 8 mg/L. In conclusion, susceptibility to perchlozone is retained for a part of the MDR strains, as is susceptibility to ethionamide, providing the possibility of therapy for such cases based on phenotypic or molecular analysis.

A mechanism of salt bridge-mediated resistance to FtsZ inhibitor PC190723 revealed by a cell-based screen.

Bacterial cell division proteins, especially the tubulin homologue FtsZ, have emerged as strong targets for developing new antibiotics. Here, we have utilized the fission yeast heterologous expression system to develop a cell-based assay to screen for small molecules that directly and specifically target the bacterial cell division protein FtsZ. The strategy also allows for simultaneous assessment of the toxicity of the drugs to eukaryotic yeast cells. As a proof-of-concept of the utility of this assay, we demonstrate the effect of the inhibitors sanguinarine, berberine, and PC190723 on FtsZ. Though sanguinarine and berberine affect FtsZ polymerization, they exert a toxic effect on the cells. Further, using this assay system, we show that PC190723 affects Helicobacter pylori FtsZ function and gain new insights into the molecular determinants of resistance to PC190723. On the basis of sequence and structural analysis and site-specific mutations, we demonstrate that the presence of salt bridge interactions between the central H7 helix and β-strands S9 and S10 mediates resistance to PC190723 in FtsZ. The single-step in vivo cell-based assay using fission yeast enabled us to dissect the contribution of sequence-specific features of FtsZ and cell permeability effects associated with bacterial cell envelopes. Thus, our assay serves as a potent tool to rapidly identify novel compounds targeting polymeric bacterial cytoskeletal proteins like FtsZ to understand how they alter polymerization dynamics and address resistance determinants in targets.

Evaluation of in-vitro susceptibility of ß-lactam-resistant Gram-negative bacilli to ceftazidime-avibactam and ceftolozane-tazobactam from clinical samples of a general hospital in southern Brazil.

The spread of carbapenemase- and extended-spectrum β-lactamase (ESBL)-producing gram-negative bacilli (GNB) represent a global public health threat that limits therapeutic options for hospitalized patients. This study aimed to evaluate the in-vitro susceptibility of β-lactam-resistant GNB to ceftazidime-avibactam (C/A) and ceftolozane-tazobactam (C/T), and investigate the molecular determinants of resistance. Overall, 101 clinical isolates of Enterobacterales and Pseudomonas aeruginosa collected from a general hospital in Brazil were analyzed. Susceptibility to the antimicrobial agents was evaluated using an automated method, and the minimum inhibitory concentrations (MIC50/90) of C/A and C/T were determined using Etest®. The β-lactamase-encoding genes were investigated using polymerase chain reaction. High susceptibility to C/A and C/T was observed among ESBL-producing Enterobacterales (100% and 97.3% for CLSI and 83.8% for BRCAST, respectively) and carbapenem-resistant P. aeruginosa (92.3% and 87.2%, respectively). Carbapenemase-producing Klebsiella pneumoniae exhibited high resistance to C/T (80%- CLSI or 100%- BRCAST) but high susceptibility to C/A (93.4%). All carbapenem-resistant K. pneumoniae isolates were susceptible to C/A, whereas only one isolate was susceptible to C/T. Both antimicrobials were inactive against metallo-β-lactamase-producing K. pneumoniae isolates. Resistance genes were concomitantly identified in 44 (44.9%) isolates, with bla CTX-M and bla SHV being the most common. C/A and C/T were active against microorganisms with β-lactam-resistant phenotypes, except when resistance was mediated by metallo-β-lactamases. Most C/A- and C/T-resistant isolates concomitantly carried two or more β-lactamase-encoding genes (62.5% and 77.4%, respectively).

Plasmid-mediated quinolone resistance determinants in clinical bacterial pathogens isolated from the Western Region of Ghana: a cross-sectional study.

quinolones are critically important antibiotics that are reserved for treating very severe infections caused by multidrug-resistant bacterial pathogens. However, their indiscriminate uses have resulted in an increased number of resistant strains in many parts of the world including Ghana. We determined the quinolone resistance profile of Gram-negative bacterial pathogens and characterized the underlying molecular determinants of resistance. Gram-negative pathogens obtained from clinical specimens at three hospital laboratories were tested for resistance to quinolones and other commonly used antibiotics. ESBL production among the Enterobacterial isolates was confirmed using the combined disc diffusion method. We then used PCR to determine seven types of plasmid-mediated quinolone resistance genes present in the isolates resistant to nalidixic acid and ciprofloxacin. in this study, 29.5% of the isolates were resistant to ciprofloxacin, with the highest of 50% among E. coli resistance to the other quinolones was levofloxacin (24.4%), norfloxacin (24.9%), and nalidixic acid (38.9%). Significant proportions of the quinolone-resistant isolates were ESBL producers (P-values < 0.001). The aac(6´)-Ib-cr, qnrS, oqxA, and qepA genes were present in 43 (89.6%), 27 (56.3%), 23 (47.9%), and one (2.1%) of the isolates, respectively. None of the isolates tested positive to qnrA, qnrB, and oqxB genes. The presence of the aac(6´)-Ib-cr gene positively correlated with resistance to ceftriaxone, cefotaxime, and gentamicin (P-values < 0.05). high proportions of Gram-negative bacterial isolates were resistant to quinolones and most of these isolates possessed multiple PMQR genes. There is a need to implement measures to limit the spread of these organisms.

High prevalence of multiple antibiotic resistance in clinical E. coli isolates from Bangladesh and prediction of molecular resistance determinants using WGS of an XDR isolate

Multi-drug-resistance (MDR) is a severe public health concern worldwide, and its containment is more challenging in developing countries due to poor antimicrobial resistance (AMR) surveillance and irrational use of antibiotics. The current study investigated 100 clinical E. coli isolates and revealed that 98% of them were MDR. PCR analysis using 25 selected isolates showed the predominance of metallo-β-lactamase gene blaNDM (80%) and ESBL genes blaOXA (48%) and blaCTX-M-15 (32%). The AmpC gene was detected in 68% of the isolates, while 32% was tetC positive. Notably, 34% of the isolates were resistant to carbapenem. Whole genome sequence (WGS) analysis of an extensively drug-resistant (XDR) isolate (L16) revealed the presence of the notorious sequence type 131 responsible for multi-drug-resistant infections, multiple antibiotic resistance genes (ARGs), virulence genes, and mobile genetic elements that pose risks to environmental transmission. Our results indicate that MDR is alarmingly increasing in Bangladesh that critically limits the treatment option against infections and contributes to further aggravation to the prevailing situation of MDR worldwide. The findings of this study will be valuable in designing sustainable strategies to contain MDR in the region.

Antifungal Drug Susceptibility and Genetic Characterization of Fungi Recovered from COVID-19 Patients.

Fungal infections are common complications of respiratory viral infections and are associated with the increased need for intensive care and elevated mortality. Data regarding microbiological and molecular characteristics of such infections in COVID-19 patients are scarce. Here, we performed a comprehensive analysis, including species identification, antifungal susceptibility testing, molecular resistance determinants analysis, typing, and retrospective clinical data review, of fungal isolates recovered from 19 COVID-19 patients, who were hospitalized at the Hackensack University Medical Center (HUMC) in Hackensack, New Jersey, USA, in the initial phase of the pandemic from April–May 2020. In total, 17 Candida albicans, two C. parapsilosis, and two Aspergillus fumigatus were analyzed. All Candida spp. isolates were susceptible to micafungin and azole drugs (fluconazole, voriconazole, posaconazole, itraconazole, isavuconazole). A. fumigatus isolates were susceptible to micafungin and all triazole drugs except fluconazole (intrinsic resistance). Multilocus sequence typing (MLST) of C. albicans isolates revealed 15 different sequence types (STs), which clustered below the clade-defining limit of p-distance < 0.04. Pulsed-field gel electrophoresis (PFGE) karyotyping revealed no chromosomal rearrangements in these isolates. A. fumigatus isolates were of different, non-related genotypes. We speculate that virus- and drug-induced immunosuppression (94.7% of the patients received corticosteroids), together with prolonged hospital stay (median duration of 29 days) and mechanical ventilation (median duration of 24 days) likely increased the susceptibility to secondary respiratory and bloodstream infections in the studied patient population. The presence of fungi in blood or respiratory tract fluid was a prognosticator for poor clinical outcome, which presented as an 89.5% 30-day mortality in our patient cohort.

136 Evolutionary and molecular determinants of resistance to UV-induced skin cancer in naked mole rats

Background. Naked mole rats (NMR) are unusually long-lived and unusually resistant to aging and spontaneous cancer development. NMR live colonially in underground tunnel systems. They are under no direct selection to develop and maintain adaptations against UV-induced tissue damage. Here we test NMR’s resistance to UV-induced DNA damage and skin cancer, and we probe for potential anti-cancer mechanisms.

Multiomics profiling of longitudinal melanoma specimens unravels molecular mechanisms of resistance to sequential targeted and cancer immunotherapies.

e22015 Background: We evaluated spatially-resolved protein profiling of longitudinal tumor specimens derived from two patients with metastatic melanoma who progressed on sequential therapies including targeted therapy (TT) and immune checkpoint blockade (ICB) therapy targeting T cell-surface antigens (CTLA-4 and PD-1). The purpose of this study was to identify molecular determinants of resistance to sequential TT and ICB therapies. Methods: We performed multiplexed and multidimensional spatial protein profiling using NanoString’s GeoMx Digital Spatial Profiling (DSP) platform and single-cell level imaging analysis with Cyclic Immunofluorescence (CycIF) to simultaneously determine dynamic changes in tumor intrinsic signaling pathways and immune response in the tumor microenvironment (TME). Results: The first patient presented with a BRAFV600E-positive brain metastasis. This patient was sequentially treated with ipilimumab (Ipi), the combination of dabrafenib and trametinib, and pembrolizumab (Pembro) and progressed despite of high expression of CD56 NK cell after all treatments. In addition, CycIF analysis revealed drug resistance in a subpopulation of cells that had continued activation of mTOR (pS6) and EGFR pathways. The second patient presented with dermal metastases on the flank with NRASQ61K mutation. This patient was sequentially treated with Pembo, Talimogene Laherparepvec, the combination of Ipi plus nivolumab, and two different investigational agents combined with Pembro. This patient displayed stable disease (SD) on Pembro but eventually progressed on the subsequent therapies. DSP analysis demonstrated CD68/CD40 myeloid cell infiltrates as well as HLA-DR and CD44 in the TME after the last treatment. CycIF analysis revealed dynamic changes in tumoral characteristics including DNA damage and proliferation during treatment. Furthermore, the analysis suggested that there might be a resistant subpopulation in the last tumor biopsy, which is in line with progression of the disease. Conclusions: In this study, we conducted detailed analyses on serial specimens from two patients to precisely define the spatial distribution of immune responses and cancer signaling pathways. The findings propose that concurrent proteomics analysis and immune monitoring of longitudinal tumor biopsies can be informative in clinical evaluation in order to identify salvage therapies to overcome drug resistance.

Abstract LB-299: A comprehensive platform of patient-derived xenografts and matched cell lines mirrors the genomic landscape of colorectal cancer

Abstract Preclinical models that accurately reflect patients’ tumor with regards to histopathology, genomics, and therapeutic response represent a compelling need to fuel the progress of effective cancer treatments. Patient-derived tumor xenografts (PDXs) have significantly accelerated this process, but, although they closely mirror structural and molecular features of the tumor of origin, they still retain important restrictions related to maintenance costs and large-scale screening. To overcome this issue, we have established a novel platform of 2D-cell lines (xeno-cell lines, XL) derived from PDXs of colorectal cancer (CRC) from which patient’s germline gDNA was available. We have characterized XL-cells at multiple levels to confirm their proximity to the PDXs of origin and to assess their suitability as patient avatars in vitro to interrogate functional networks in colorectal cancer. All XL-cells showed an epithelial-like morphology and phenotype, as also confirmed by EMT biomarker transcriptomic analysis. Whole exome and RNA-seq analyses showed that genomic features were consistently preserved between PDXs and matched cell models. Expression analysis revealed the XL-line collection as a significant representative of all CRC subtypes (CMS and CRIS subgroups). Furthermore, genomic analysis allowed the identification of molecular biomarkers of response and resistance to targeted therapies, including EGFR and HER2 blockade. In particular, molecular determinants of resistance to dual-HER2 blockade previously reported in the Heracles-A trial were validated in XL-cells. In conclusion, the XL-cell line and PDX platform represents a unique and comprehensive preclinical tool to validate gene function e to identify novel pharmacological vulnerabilities in colorectal cancer. Citation Format: Luca Lazzari, Giorgio Corti, Claudio Isella, Monica Montone, Pamela Arcella, Eugenia Zanella, Luca Novara, Fabiane Barbosa, Andrea Cassingena, Carlotta Cancelliere, Enzo Medico, Andrea Sartore-Bianchi, Salvatore Siena, Andrea Bertotti, Livio Trusolino, Federica Di Nicolantonio, Michael Linnebacher, Alberto Bardelli, Sabrina Arena. A comprehensive platform of patient-derived xenografts and matched cell lines mirrors the genomic landscape of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-299.

Identification of Molecular Determinants of Resistance to Pyrethroid Insecticides in Aedes aegypti (Diptera: Culicidae) Populations in California, USA.

The first breeding populations of Aedes aegypti (Linnaeus) were identified in California in 2013, and have since been detected in 13 counties. Recent studies suggest two introductions likely occurred, with genetically distinct populations in the central and southern regions of the state. Given the threat of dengue, chikungunya, and Zika virus transmission, it is imperative to understand if these populations harbor genes that could confer resistance to pyrethrin-based insecticides, known as pyrethroids, the most commonly used class of adulticides in the state. In 2017, the California Department of Public Health initiated a pesticide resistance screening program for Ae. aegypti to assess the presence of specific mutations on the sodium channel gene (V1016I and F1534C) associated with knockdown resistance to pyrethroids. Mosquitoes collected between 2015 and 2017 from 11 counties were screened for mutations using real-time polymerase chain reaction assays. Results revealed distinctly different resistance profiles between the central and southern regions. The central population displayed nearly fixed resistant mutations at both loci, whereas the southern population was more variable. The relative proportion of resistant alleles observed in sampled mosquitoes collected in southern California increased each year from 2015 through 2017, indicating potential increases in resistance across this region. The presence of these mutations indicates that these mosquitoes may be predisposed to surviving pyrethroid treatments. Additional biological and biochemical assays will help better elucidate the mechanisms underlying insecticide resistance in California Ae. aegypti and prompt the use of pesticides that are most effective at controlling these mosquitoes.

A systematic review of antimicrobial resistance of typhoidal Salmonella in India.

Background & objectives:The temporal trends in the development of antimicrobial resistance (AMR) among Salmonella Typhi and Salmonella Paratyphi in India have not been systematically reported. We aimed to systematically review the temporal AMR trends (phenotypic and molecular mechanisms) in bacterial isolates from patients with enteric fever over two decades in India.Methods:To identify trends in AMR in India, resistance patterns among 4611 individual S. Typhi isolates and 800 S. Paratyphi A isolates, reported from 1992 to 2017 in 40 publications, were analysed. Molecular resistance determinants were extracted from 22 publications and also reviewed in accordance with the PRISMA guidelines. Articles were sourced using a predefined search strategy from different databases.Results:The analyses suggested that multidrug-resistant (MDR) enteric fever was declining in India and being replaced by fluoroquinolone (FQ) resistance. Mutations in gyrA and parC were key mechanisms responsible for FQ resistance, whereas MDR was largely driven by resistance determinants encoded on mobile genetic elements (plasmids, transposons).Interpretation & conclusions:The results reflect the effect of antimicrobial pressure which has been driving AMR in typhoidal Salmonella in India. Understanding these trends is important in planning future approaches to therapy, which serve as a baseline for assessment of the impact of new typhoid conjugate vaccines against these resistant organisms.

A systematic review of antimicrobial resistance in Salmonella enterica serovar Typhi, the etiological agent of typhoid.

BackgroundThe temporal and spatial change in trends of antimicrobial resistance (AMR) in typhoid have not been systematically studied, and such information will be critical for defining intervention, as well as planning sustainable prevention strategies.Methodology and findingsTo identify the phenotypic trends in AMR, 13,833 individual S. Typhi isolates, reported from 1973 to 2018 in 62 publications, were analysed to determine the AMR preponderance over time. Separate analyses of molecular resistance determinants present in over 4,000 isolates reported in 61 publications were also conducted. Multi-drug resistant (MDR) typhoid is in decline in Asia in a setting of high fluoroquinolone resistance while it is on the increase in Africa. Mutations in QRDRs in gyrA (S83F, D87N) and parC (S80I) are the most common mechanisms responsible for fluoroquinolone resistance. Cephalosporin resistant S. Typhi, dubbed extensively drug-resistant (XDR) is a real threat and underscores the urgency in deploying the Vi-conjugate vaccines.ConclusionFrom these observations, it appears that AMR in S. Typhi will continue to emerge leading to treatment failure, changes in antimicrobial policy and further resistance developing in S. Typhi isolates and other Gram-negative bacteria in endemic regions. The deployment of typhoid conjugate vaccines to control the disease in endemic regions may be the best defence.

Evolution of drug resistance in Mycobacterium tuberculosis: a review on the molecular determinants of resistance and implications for personalized care.

Drug-resistant TB (DR-TB) remains a significant challenge in TB treatment and control programmes worldwide. Advances in sequencing technology have significantly increased our understanding of the mechanisms of resistance to anti-TB drugs. This review provides an update on advances in our understanding of drug resistance mechanisms to new, existing drugs and repurposed agents. Recent advances in WGS technology hold promise as a tool for rapid diagnosis and clinical management of TB. Although the standard approach to WGS of Mycobacterium tuberculosis is slow due to the requirement for organism culture, recent attempts to sequence directly from clinical specimens have improved the potential to diagnose and detect resistance within days. The introduction of new databases may be helpful, such as the Relational Sequencing TB Data Platform, which contains a collection of whole-genome sequences highlighting key drug resistance mutations and clinical outcomes. Taken together, these advances will help devise better molecular diagnostics for more effective DR-TB management enabling personalized treatment, and will facilitate the development of new drugs aimed at improving outcomes of patients with this disease.

224 - Molecular determinants of resistance to CDK4/6 inhibition in ER+ breast cancer

224 - Molecular determinants of resistance to CDK4/6 inhibition in ER+ breast cancer