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BACKGROUND: Moraxella catarrhalis is a leading cause of otitis media (OM) and chronic obstructive pulmonary disease (COPD). M. catarrhalis contains a Type III DNA adenine methyltransferase (ModM) that is phase-variably expressed (i.e., its expression is subject to random, reversible ON/OFF switching). ModM has six target recognition domain alleles (modM1-6), and we have previously shown that modM2 is the predominant allele, while modM3 is associated with OM. Phase-variable DNA methyltransferases mediate epigenetic regulation and modulate pathogenesis in several bacteria. ModM2 of M. catarrhalis regulates the expression of a phasevarion containing genes important for colonization and infection. Here we describe the phase-variable expression of modM3, the ModM3 methylation site and the suite of genes regulated within the ModM3 phasevarion. RESULTS: Phase-variable expression of modM3, mediated by variation in length of a 5'-(CAAC)n-3' tetranucleotide repeat tract in the open reading frame was demonstrated in M. catarrhalis strain CCRI-195ME with GeneScan fragment length analysis and western immunoblot. We determined that ModM3 is an active N6-adenine methyltransferase that methylates the sequence 5'-ACm6ATC-3'. Methylation was detected at all 4446 5'-ACATC-3' sites in the genome when ModM3 is expressed. RNASeq analysis identified 31 genes that are differentially expressed between modM3 ON and OFF variants, including five genes that are involved in the response to oxidative and nitrosative stress, with potential roles in biofilm formation and survival in anaerobic environments. An in vivo chinchilla (Chinchilla lanigera) model of otitis media demonstrated that transbullar challenge with the modM3 OFF variant resulted in an increased middle ear bacterial load compared to a modM3 ON variant. In addition, co-infection experiments with NTHi and M. catarrhalis modM3 ON or modM3 OFF variants revealed that phase variation of modM3 altered survival of NTHi in the middle ear during early and late stage infection. CONCLUSIONS: Phase variation of ModM3 epigenetically regulates the expression of a phasevarion containing multiple genes that are potentially important in the progression of otitis media.
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Viabilidad Microbiana/genética , Moraxella catarrhalis/enzimología , Moraxella catarrhalis/genética , Otitis Media/microbiología , Metiltransferasa de ADN de Sitio Específico (Adenina Especifica)/genética , Animales , Proteínas Bacterianas/genética , Chinchilla , Modelos Animales de Enfermedad , Epigénesis Genética , Femenino , Expresión Génica , Regulación Bacteriana de la Expresión Génica , Técnicas de Inactivación de Genes , Humanos , Masculino , Infecciones por Moraxellaceae/microbiologíaRESUMEN
Moraxella catarrhalis is a human-restricted opportunistic bacterial pathogen of the respiratory mucosa. It frequently colonizes the nasopharynx asymptomatically, but is also an important causative agent of otitis media (OM) in children, and plays a significant role in acute exacerbations of chronic obstructive pulmonary disease (COPD) in adults. As the current treatment options for M. catarrhalis infection in OM and exacerbations of COPD are often ineffective, the development of an efficacious vaccine is warranted. However, no vaccine candidates for M. catarrhalis have progressed to clinical trials, and information regarding the distribution of M. catarrhalis virulence factors and vaccine candidates is inconsistent in the literature. It is largely unknown if virulence is associated with particular strains or subpopulations of M. catarrhalis, or if differences in clinical manifestation can be attributed to the heterogeneous expression of specific M. catarrhalis virulence factors in the circulating population. Further investigation of the distribution of M. catarrhalis virulence factors in the context of carriage and disease is required so that vaccine development may be targeted at relevant antigens that are conserved among disease-causing strains. The challenge of determining which of the proposed M. catarrhalis virulence factors are relevant to human disease is amplified by the lack of a standardized M. catarrhalis typing system to facilitate direct comparisons of worldwide isolates. Here we summarize and evaluate proposed relationships between M. catarrhalis subpopulations and specific virulence factors in the context of colonization and disease, as well as the current methods used to infer these associations.
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Moraxella catarrhalis/inmunología , Moraxella catarrhalis/patogenicidad , Infecciones por Moraxellaceae/inmunología , Infecciones por Moraxellaceae/microbiología , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Vacunas Bacterianas/inmunología , Humanos , Moraxella catarrhalis/clasificación , Moraxella catarrhalis/genética , Infecciones por Moraxellaceae/tratamiento farmacológico , Infecciones por Moraxellaceae/prevención & control , Otitis Media/tratamiento farmacológico , Otitis Media/inmunología , Otitis Media/microbiología , Otitis Media/prevención & control , Enfermedad Pulmonar Obstructiva Crónica/tratamiento farmacológico , Enfermedad Pulmonar Obstructiva Crónica/inmunología , Enfermedad Pulmonar Obstructiva Crónica/microbiología , Enfermedad Pulmonar Obstructiva Crónica/prevención & control , Virulencia/genética , Virulencia/inmunología , Factores de Virulencia/genética , Factores de Virulencia/inmunologíaRESUMEN
Moraxella catarrhalis is a significant cause of otitis media and exacerbations of chronic obstructive pulmonary disease. Here, we characterize a phase-variable DNA methyltransferase (ModM), which contains 5'-CAAC-3' repeats in its open reading frame that mediate high-frequency mutation resulting in reversible on/off switching of ModM expression. Three modM alleles have been identified (modM1-3), with modM2 being the most commonly found allele. Using single-molecule, real-time (SMRT) genome sequencing and methylome analysis, we have determined that the ModM2 methylation target is 5'-GAR(m6)AC-3', and 100% of these sites are methylated in the genome of the M. catarrhalis 25239 ModM2 on strain. Proteomic analysis of ModM2 on and off variants revealed that ModM2 regulates expression of multiple genes that have potential roles in colonization, infection, and protection against host defenses. Investigation of the distribution of modM alleles in a panel of M. catarrhalis strains, isolated from the nasopharynx of healthy children or middle ear effusions from patients with otitis media, revealed a statistically significant association of modM3 with otitis media isolates. The modulation of gene expression via the ModM phase-variable regulon (phasevarion), and the significant association of the modM3 allele with otitis media, suggests a key role for ModM phasevarions in the pathogenesis of this organism.
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Metilasas de Modificación del ADN/metabolismo , Moraxella catarrhalis/patogenicidad , Infecciones por Moraxellaceae/microbiología , Otitis Media/microbiología , Secuencia de Aminoácidos , Metilasas de Modificación del ADN/química , Cartilla de ADN , Humanos , Espectrometría de Masas , Datos de Secuencia Molecular , Infecciones por Moraxellaceae/enzimología , Otitis Media/enzimología , Reacción en Cadena de la Polimerasa , Homología de Secuencia de AminoácidoRESUMEN
Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is widely used in clinical microbiology laboratories for bacterial identification but its use for detection of antimicrobial resistance (AMR) remains limited. Here, we used MALDI-TOF MS with artificial intelligence (AI) approaches to successfully predict AMR in Pseudomonas aeruginosa, a priority pathogen with complex AMR mechanisms. The highest performance was achieved for modern ß-lactam/ß-lactamase inhibitor drugs, namely, ceftazidime/avibactam and ceftolozane/tazobactam. For these drugs, the model demonstrated area under the receiver operating characteristic curve (AUROC) of 0.869 and 0.856, specificity of 0.925 and 0.897, and sensitivity of 0.731 and 0.714, respectively. As part of this work, we developed dynamic binning, a feature engineering technique that effectively reduces the high-dimensional feature set and has wide-ranging applicability to MALDI-TOF MS data. Compared to conventional feature engineering approaches, the dynamic binning method yielded highest performance in 7 of 10 antimicrobials. Moreover, we showcased the efficacy of transfer learning in enhancing the AUROC performance for 8 of 11 antimicrobials. By assessing the contribution of features to the model's prediction, we identified proteins that may contribute to AMR mechanisms. Our findings demonstrate the potential of combining AI with MALDI-TOF MS as a rapid AMR diagnostic tool for Pseudomonas aeruginosa.IMPORTANCEPseudomonas aeruginosa is a key bacterial pathogen that causes significant global morbidity and mortality. Antimicrobial resistance (AMR) emerges rapidly in P. aeruginosa and is driven by complex mechanisms. Drug-resistant P. aeruginosa is a major challenge in clinical settings due to limited treatment options. Early detection of AMR can guide antibiotic choices, improve patient outcomes, and avoid unnecessary antibiotic use. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is widely used for rapid species identification in clinical microbiology. In this study, we repurposed mass spectra generated by MALDI-TOF and used them as inputs for artificial intelligence approaches to successfully predict AMR in P. aeruginosa for multiple key antibiotic classes. This work represents an important advance toward using MALDI-TOF as a rapid AMR diagnostic for P. aeruginosa in clinical settings.
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OBJECTIVES: New Delhi metallo-ß-lactamases (NDMs) are major contributors to the spread of carbapenem resistance globally. In Australia, NDMs were previously associated with international travel, but from 2019 we noted increasing incidence of NDM-positive clinical isolates. We investigated the clinical and genomic epidemiology of NDM carriage at a tertiary-care Australian hospital from 2016 to 2021. METHODS: We identified 49 patients with 84 NDM-carrying isolates in an institutional database, and we collected clinical data from electronic medical record. Short- and long-read whole genome sequencing was performed on all isolates. Completed genome assemblies were used to assess the genetic setting of blaNDM genes and to compare NDM plasmids. RESULTS: Of 49 patients, 38 (78%) were identified in 2019-2021 and only 11 (29%) of 38 reported prior travel, compared with 9 (82%) of 11 in 2016-2018 (P = .037). In patients with NDM infection, the crude 7-day mortality rate was 0% and the 30-day mortality rate was 14% (2 of 14 patients). NDMs were noted in 41 bacterial strains (ie, species and sequence type combinations). Across 13 plasmid groups, 4 NDM variants were detected: blaNDM-1, blaNDM-4, blaNDM-5, and blaNDM-7. We noted a change from a diverse NDM plasmid repertoire in 2016-2018 to the emergence of conserved blaNDM-1 IncN and blaNDM-7 IncX3 epidemic plasmids, with interstrain spread in 2019-2021. These plasmids were noted in 19 (50%) of 38 patients and 35 (51%) of 68 genomes in 2019-2021. CONCLUSIONS: Increased NDM case numbers were due to local circulation of 2 epidemic plasmids with extensive interstrain transfer. Our findings underscore the challenges of outbreak detection when horizontal transmission of plasmids is the primary mode of spread.
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Brotes de Enfermedades , Plásmidos , beta-Lactamasas , Humanos , beta-Lactamasas/genética , Plásmidos/genética , Masculino , Femenino , Persona de Mediana Edad , Anciano , Australia/epidemiología , Secuenciación Completa del Genoma , Adulto , Infecciones por Enterobacteriaceae/epidemiología , Infecciones por Enterobacteriaceae/transmisión , Infecciones por Enterobacteriaceae/microbiología , Transferencia de Gen Horizontal , Anciano de 80 o más Años , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Genoma BacterianoRESUMEN
Introduction. One third of people with CF in the UK are co-infected by both Staphylococcus aureus and Pseudomonas aeruginosa. Chronic bacterial infection in CF contributes to the gradual destruction of lung tissue, and eventually respiratory failure in this group.Gap Statement. The contribution of S. aureus to cystic fibrosis (CF) lung decline in the presence or absence of P. aeruginosa is unclear. Defining the molecular and phenotypic characteristics of a range of S. aureus clinical isolates will help further understand its pathogenic capabilities.Aim. Our objective was to use molecular and phenotypic tools to characterise twenty-five clinical S. aureus isolates collected from mono- and coinfection with P. aeruginosa from people with CF at the Royal Victoria Infirmary, Newcastle upon Tyne.Methodology. Genomic DNA was extracted and sequenced. Multilocus sequence typing was used to construct phylogeny from the seven housekeeping genes. A pangenome was calculated using Roary, and cluster of Orthologous groups were assigned using eggNOG-mapper which were used to determine differences within core, accessory, and unique genomes. Characterisation of sequence type, clonal complex, agr and spa types was carried out using PubMLST, eBURST, AgrVATE and spaTyper, respectively. Antibiotic resistance was determined using Kirby-Bauer disc diffusion tests. Phenotypic testing of haemolysis was carried out using ovine red blood cell agar plates and mucoid phenotypes visualised using Congo red agar.Results. Clinical strains clustered closely based on agr type, sequence type and clonal complex. COG analysis revealed statistically significant enrichment of COG families between core, accessory and unique pangenome groups. The unique genome was significantly enriched for replication, recombination and repair, and defence mechanisms. The presence of known virulence genes and toxins were high within this group, and unique genes were identified in 11 strains. Strains which were isolated from the same patient all surpassed average nucleotide identity thresholds, however, differed in phenotypic traits. Antimicrobial resistance to macrolides was significantly higher in the coinfection group.Conclusion. There is huge variation in genetic and phenotypic capabilities of S. aureus strains. Further studies on how these may differ in relation to other species in the CF lung may give insight into inter-species interactions.
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Coinfección , Fibrosis Quística , Infecciones Estafilocócicas , Animales , Ovinos , Staphylococcus aureus , Fibrosis Quística/complicaciones , Fibrosis Quística/microbiología , Coinfección/microbiología , Agar , Fenotipo , Infecciones Estafilocócicas/microbiología , Antibacterianos/farmacologíaRESUMEN
Infections caused by metallo-beta-lactamase-producing organisms (MBLs) are a global health threat. Our understanding of transmission dynamics and how MBLs establish endemicity remains limited. We analysed two decades of blaIMP-4 evolution in a hospital using sequence data from 270 clinical and environmental isolates (including 169 completed genomes) and identified the blaIMP-4 gene across 7 Gram-negative genera, 68 bacterial strains and 7 distinct plasmid types. We showed how an initial multi-species outbreak of conserved IncC plasmids (95 genomes across 37 strains) allowed endemicity to be established through the ability of blaIMP-4 to disseminate in successful strain-genetic setting pairs we termed propagators, in particular Serratia marcescens and Enterobacter hormaechei. From this reservoir, blaIMP-4 persisted through diversification of genetic settings that resulted from transfer of blaIMP-4 plasmids between bacterial hosts and of the integron carrying blaIMP-4 between plasmids. Our findings provide a framework for understanding endemicity and spread of MBLs and may have broader applicability to other carbapenemase-producing organisms.
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Integrones , beta-Lactamasas , Integrones/genética , beta-Lactamasas/genética , beta-Lactamasas/metabolismo , Proteínas Bacterianas/genética , Plásmidos/genética , Serratia marcescens/genética , Serratia marcescens/metabolismo , Carbapenémicos/farmacología , Genómica , Pruebas de Sensibilidad Microbiana , Antibacterianos/farmacologíaRESUMEN
BACKGROUND: Resistance to third-generation cephalosporins, often mediated by extended-spectrum beta-lactamases (ESBLs), is a considerable issue in hospital-associated infections as few drugs remain for treatment. ESBL genes are often located on large plasmids that transfer horizontally between strains and species of Enterobacteriaceae and frequently confer resistance to additional drug classes. Whilst plasmid transmission is recognised to occur in the hospital setting, the frequency and impact of plasmid transmission on infection burden, compared to ESBL + strain transmission, is not well understood. METHODS: We sequenced the genomes of clinical and carriage isolates of Klebsiella pneumoniae species complex from a year-long hospital surveillance study to investigate ESBL burden and plasmid transmission in an Australian hospital. Long-term persistence of a key transmitted ESBL + plasmid was investigated via sequencing of ceftriaxone-resistant isolates during 4 years of follow-up, beginning 3 years after the initial study. RESULTS: We found 25 distinct ESBL plasmids. We identified one plasmid, which we called Plasmid A, that carried blaCTX-M-15 in an IncF backbone similar to pKPN-307. Plasmid A was transmitted at least four times into different Klebsiella species/lineages and was responsible for half of all ESBL episodes during the initial 1-year study period. Three of the Plasmid A-positive strains persisted locally 3-6 years later, and Plasmid A was detected in two additional strain backgrounds. Overall Plasmid A accounted for 21% of ESBL + infections in the follow-up period. CONCLUSIONS: Here, we systematically surveyed ESBL strain and plasmid transmission over 1 year in a single hospital network. Whilst ESBL plasmid transmission events were rare in this setting, they had a significant and sustained impact on the burden of ceftriaxone-resistant and multidrug-resistant infections. If onward transmission of Plasmid A-carrying strains could have been prevented, this may have reduced the number of opportunities for Plasmid A to transmit and create novel ESBL + strains, as well as reducing overall ESBL infection burden.
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Klebsiella pneumoniae , beta-Lactamasas , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Australia/epidemiología , Ceftriaxona , Hospitales , Humanos , Klebsiella pneumoniae/genética , Plásmidos/genética , beta-Lactamasas/genéticaRESUMEN
Nisin, a peptide used as a natural food preservative, is employed in this work for the development of a novel nanocarrier system. Stable and uniform nisin-shelled nanoemulsions (NSNE) with a diameter of 100 ± 20 nm were successfully prepared using 20 kHz flow-through ultrasonication technique. The NSNE showed limited toxicity, high bactericidal activity and high drug loading capacity (EE 65 % w/w). In addition, the nisin shell was exploited for the site-specific attachment of a recombinantly produced cancer targeting ligand (αHER2LPETG IgG). Employing a unique two phases (bio-click) approach which involved both Sortase A mediated Azide Bioconjugation (SMAB) and Strain Promoted Azide Alkyne Cycloaddition (SPAAC) reactions, targeted NSNE (NSNEDOX-αHER2 IgG) were successfully assembled and loaded with the chemotherapeutic drug Doxorubicin (DOX). Finally, NSNEDOX-αHER2 IgG showed cancer-specific binding and augmented cytotoxicity to HER2 expressing tumour cells.
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Neoplasias , Nisina , Humanos , Azidas , Nisina/farmacología , Doxorrubicina/farmacología , Inmunoglobulina GRESUMEN
OBJECTIVES: There are scant primary clinical data on antimicrobial resistance (AMR) burden from low- and middle-income countries (LMICs). We adapted recent World Health Organization methodology to measure the effect of third-generation cephalosporin resistance (3GC-R) on mortality and excess length of hospital stay in Fiji. METHODS: We conducted a prospective cohort study of inpatients with Enterobacterales bloodstream infections (BSIs) at Colonial War Memorial Hospital, Suva. We used cause-specific Cox proportional hazards models to estimate the effect of 3GC-R on the daily risk (hazard) of in-hospital mortality and being discharged alive (competing risks), and we used multistate modelling to estimate the excess length of hospital stay. RESULTS: From July 2020 to February 2021 we identified 162 consecutive Enterobacterales BSIs; 3GC-R was present in 66 (40.7%). Crude mortality for patients with 3GC-susceptible and 3GC-R BSIs was 16.7% (16/96) and 30.3% (20/66), respectively. 3GC-R was not associated with the in-hospital mortality hazard rate (adjusted hazard ratio [aHR] 1.13, 95% confidence interval [CI] 0.51-2.53) or being discharged alive (aHR 0.99, 95% CI 0.65-1.50), whereas Charlson comorbidity index score (aHR 1.62, 95% CI 1.36-1.93) and Pitt bacteraemia score (aHR 3.57, 95% CI 1.31-9.71) were both associated with an increased hazard rate of in-hospital mortality. 3GC-R was associated with an increased length of stay of 2.6 days (95% CI 2.5-2.8). 3GC-R was more common among hospital-associated infections, but genomics did not identify clonal transmission. CONCLUSION: Patients with Enterobacterales BSIs in Fiji had high mortality. There were high rates of 3GC-R, which was associated with increased hospital length of stay but not with in-hospital mortality.
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Bacteriemia , Infección Hospitalaria , Bacteriemia/tratamiento farmacológico , Cefalosporinas , Infección Hospitalaria/tratamiento farmacológico , Fiji/epidemiología , Humanos , Tiempo de Internación , Estudios ProspectivosRESUMEN
Background: Staphylococcus aureus bacteraemia (SAB) is one of the commonest bloodstream infections globally and is associated with a high mortality rate. Most published data comes from temperate, high-income countries. We describe the clinical epidemiology, microbiology, management and outcomes of patients with SAB treated in a tropical, middle-income setting at Fiji's largest hospital. Methods: A prospective, observational study was performed of consecutive SAB cases admitted to Colonial War Memorial Hospital (CWMH) in Suva, between July 2020 and February 2021. Detailed demographic, clinical and microbiological data were collected, including the key outcome of in-patient mortality. To estimate the population incidence, all SAB cases diagnosed at the CWMH laboratory were included - even if not admitted to CWMH - with the population of Fiji's Central Division used as the denominator. Findings: A total of 176 cases of SAB were detected over eight-months, which equated to an incidence of 68.8 cases per 100,000 population per year. Of these, 95 cases were admitted to CWMH within 48 h of index culture. Approximately 8.4% (8/95) of admitted cases were caused by methicillin-resistant Staphylococcus aureus (MRSA). All cause in-patient mortality was 25.3%, increasing to 55% among patients aged 60 or older. Interpretation: This reported incidence of SAB in central Fiji is one of the highest in the world. SAB was associated with significant mortality, especially in those over 60 years of age, despite a relatively low frequency of methicillin resistance. Funding: Supported by the National Health and Medical Research Council (Australia) and the GRAM (Global Research on Antimicrobial Resistance) Project, Oxford University (United Kingdom).
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Background: Well tolerated antivirals administered early in the course of COVID-19 infection when the viremia is highest could prevent progression to severe disease. Favipiravir inhibits SARS-CoV-2 viral replication in vitro with evidence of clinical benefit in open label trials. Placebo controlled studies of people with early symptomatic COVID-19 with regular assessments of SARS-CoV-2 viral load can determine if it has an antiviral effect and improves clinical outcomes. Methods: People with PCR-confirmed COVID-19 and 5 days or less of symptoms were randomised 1:1 to favipiravir 1800 mg on day 1, then 800 mg twice daily or matched placebo for 14 days. SARS-CoV-2 viral load was quantitated from second daily self-collected nose-throat swabs while receiving study drug. The primary endpoint was time to virological cure defined as 2 successive swabs negative for SARS-CoV-2 by PCR and secondary outcomes were progression of disease severity, symptom resolution and safety. Findings: Between 31 July 2020 and 19 September 2021, 200 people were enrolled (199 in the community, 1 in hospital) with 190 receiving one or more doses of drug (modified intention to treat [mITT] population). There was no difference in time to virological cure (Log-rank p=0.6 comparing Kaplan Meier curves), progression to hospitalisation (14 favipiravir, 9 placebo; p=0.38), time to symptom resolution (cough, fever, sore throat) and there were no deaths. 51 people related an adverse event that was possibly drug related, but these were evenly distributed (n=24 favipiravir, n=27 placebo). Sensitivity analyses where the definition of virological cure was changed to: a single negative PCR, exclude datapoints based on the presence or absence of human DNA in the swab, a SARS-CoV-2 viral load < 300 copies/mL being considered negative all demonstrated no difference between arms. Interpretation: Favipiravir does not improve the time to virological cure or clinical outcomes and shows no evidence of an antiviral effect when treating early symptomatic COVID-19 infection. Funding: The study was supported in part by grants from the Commonwealth Bank Australia, the Lord Mayor's Charitable Foundation, Melbourne Australia and the Orloff Family Charitable Trust, Melbourne, Australia. JHM is supported by the Medical Research Future Fund, AYP, JT are supported by the Australian National Health and Medical Research Council.
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Respiratory tract infection with SARS-CoV-2 results in varying immunopathology underlying COVID-19. We examine cellular, humoral and cytokine responses covering 382 immune components in longitudinal blood and respiratory samples from hospitalized COVID-19 patients. SARS-CoV-2-specific IgM, IgG, IgA are detected in respiratory tract and blood, however, receptor-binding domain (RBD)-specific IgM and IgG seroconversion is enhanced in respiratory specimens. SARS-CoV-2 neutralization activity in respiratory samples correlates with RBD-specific IgM and IgG levels. Cytokines/chemokines vary between respiratory samples and plasma, indicating that inflammation should be assessed in respiratory specimens to understand immunopathology. IFN-α2 and IL-12p70 in endotracheal aspirate and neutralization in sputum negatively correlate with duration of hospital stay. Diverse immune subsets are detected in respiratory samples, dominated by neutrophils. Importantly, dexamethasone treatment does not affect humoral responses in blood of COVID-19 patients. Our study unveils differential immune responses between respiratory samples and blood, and shows how drug therapy affects immune responses during COVID-19.
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COVID-19 , Anticuerpos Antivirales , Humanos , Inmunidad , Inmunoglobulina G , Inmunoglobulina M , Sistema Respiratorio , SARS-CoV-2 , Índice de Severidad de la Enfermedad , Glicoproteína de la Espiga del CoronavirusRESUMEN
OBJECTIVES: mcr-9.1 is a newly described mobile colistin resistance gene. We have noted its presence in multiple species of carbapenem-resistant Enterobacterales (CRE) from our institution. We aimed to determine the clinical features, genomic context and phenotypic impact of mcr-9.1 carriage in a series of patients between 2010 and 2019. METHODS: We identified 32 patients with mcr-9.1-carrying CRE isolates (mCRE) and collected demographic, antimicrobial exposure and infection data. Whole-genome sequencing (including short and long reads) was performed on 32 isolates. We assessed sequence similarity of mcr-9.1-harbouring plasmids, then compared our findings with plasmids for which sequence data were publicly available. RESULTS: There was no colistin exposure in patients prior to isolation of mCRE. mcr-9.1 was identified on IncHI2 plasmids across four different bacterial species and was co-located with blaIMP-4 in 23/30 plasmids studied. mCRE isolates did not demonstrate phenotypic colistin resistance, either at baseline or following sublethal colistin exposure, thus showing that mcr-9.1 alone is not sufficient for resistance. Publicly available sequence data indicated the presence of carbapenemase genes in 236/619 mcr-9.1-carrying genomes (38%). IncHI2 plasmids carrying mcr-9.1 and carbapenemase genes were detected in genomes from North America, Europe, North Africa, Asia and Oceania. CONCLUSIONS: Spread of mcr-9.1 in CRE from our institution was driven by IncHI2 'superplasmids', so termed because of their large size and their prolific carriage of resistance determinants. These were also detected in global CRE genomes. Phenotypic colistin resistance was not detected in our isolates but remains to be determined from global mCRE.
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Carbapenémicos , Colistina , Farmacorresistencia Bacteriana/genética , Enterobacteriaceae , Plásmidos , Antibacterianos/farmacología , Carbapenémicos/farmacología , Colistina/farmacología , Enterobacteriaceae/efectos de los fármacos , Enterobacteriaceae/genética , Genes Bacterianos , Pruebas de Sensibilidad Microbiana , Plásmidos/genéticaRESUMEN
Moraxella catarrhalis is a leading cause of otitis media and exacerbations of chronic obstructive pulmonary disease; however, its response to iron starvation during infection is not completely understood. Here, we announce a sequential window acquisition of all theoretical fragment ion spectra mass spectrometry (SWATH-MS) data set describing the differential expression of the M. catarrhalis CCRI-195ME proteome under iron-restricted versus iron-replete conditions.
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Moraxella catarrhalis is a leading bacterial cause of otitis media and exacerbations of chronic obstructive pulmonary disease. Here, we announce a transcriptome RNA sequencing data set detailing global gene expression in two M. catarrhalis CCRI-195ME variants with expression of the DNA methyltransferase ModM3 phase varied either on or off.
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Moraxella catarrhalis is a human-adapted, opportunistic bacterial pathogen of the respiratory mucosa. Although asymptomatic colonization of the nasopharynx is common, M. catarrhalis can ascend into the middle ear, where it is a prevalent causative agent of otitis media in children, or enter the lower respiratory tract, where it is associated with acute exacerbations of chronic obstructive pulmonary disease in adults. Phase variation is the high frequency, random, reversible switching of gene expression that allows bacteria to adapt to different host microenvironments and evade host defences, and is most commonly mediated by simple DNA sequence repeats. Bioinformatic analysis of five closed M. catarrhalis genomes identified 17 unique simple DNA sequence repeat tracts that were variable between strains, indicating the potential to mediate phase variable expression of the associated genes. Assays designed to assess simple sequence repeat variation under conditions mimicking host infection demonstrated that phase variation of uspA1 (ubiquitous surface protein A1) from high to low expression occurs over 72 hours of biofilm passage, while phase variation of uspA2 (ubiquitous surface protein A2) to high expression variants occurs during repeated exposure to human serum, as measured by mRNA levels. We also identify and confirm the variable expression of two novel phase variable genes encoding a Type III DNA methyltransferase (modO), and a conserved hypothetical permease (MC25239_RS00020). These data reveal the repertoire of phase variable genes mediated by simple sequence repeats in M. catarrhalis and demonstrate that modulation of expression under conditions mimicking human infection is attributed to changes in simple sequence repeat length.
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Regulación Bacteriana de la Expresión Génica/genética , Moraxella catarrhalis/genética , Adhesión Bacteriana/genética , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas de la Membrana Bacteriana Externa/metabolismo , Expresión Génica/genética , Humanos , Repeticiones de Microsatélite/genética , Moraxella catarrhalis/patogenicidad , Infecciones por Moraxellaceae , Otitis Media/microbiología , Secuencias Repetitivas de Ácidos Nucleicos/genéticaRESUMEN
Moraxella catarrhalis is a host-adapted bacterial pathogen that causes otitis media and exacerbations of chronic obstructive pulmonary disease. This study characterises the conserved M. catarrhalis extracellular nuclease, a member of the ßßα metal finger family of nucleases, that we have named NucM. NucM shares conserved sequence motifs from the ßßα nuclease family, including the DRGH catalytic core and Mg2+ co-ordination site, but otherwise shares little primary sequence identity with other family members, such as the Serratia Nuc and pneumococcal EndA nucleases. NucM is secreted from the cell and digests linear and circular nucleic acid. However, it appears that a proportion of NucM is also associated with the cell membrane and acts as an entry nuclease, facilitating transformation of M. catarrhalis cells. This is the first example of a ßßα nuclease in a Gram negative bacteria that acts as an entry nuclease. In addition to its role in competence, NucM affects cell aggregation and biofilm formation by M. catarrhalis, with ΔnucM mutants having increased biofilm biomass. NucM is likely to increase the ability of cells to survive and persist in vivo, increasing the virulence of M. catarrhalis and potentially affecting the behaviour of other pathogens that co-colonise the otorhinolaryngological niche.