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2.
Emerg Infect Dis ; 27(1)2020 Nov 30.
Artículo en Inglés | MEDLINE | ID: mdl-33256890

RESUMEN

We investigated the dynamics of seroconversion in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. During March 29-May 22, 2020, we collected serum samples and associated clinical data from 177 persons in London, UK, who had SARS-CoV-2 infection. We measured IgG against SARS-CoV-2 and compared antibody levels with patient outcomes, demographic information, and laboratory characteristics. We found that 2.0%-8.5% of persons did not seroconvert 3-6 weeks after infection. Persons who seroconverted were older, were more likely to have concurrent conditions, and had higher levels of inflammatory markers. Non-White persons had higher antibody concentrations than those who identified as White; these concentrations did not decline during follow-up. Serologic assay results correlated with disease outcome, race, and other risk factors for severe SARS-CoV-2 infection. Serologic assays can be used in surveillance to clarify the duration and protective nature of humoral responses to SARS-CoV-2 infection.

3.
Lancet Oncol ; 20(12): 1760-1772, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31668592

RESUMEN

BACKGROUND: Myeloma causes profound immunodeficiency and recurrent, serious infections. Around 5500 new cases of myeloma are diagnosed per year in the UK, and a quarter of patients will have a serious infection within 3 months of diagnosis. We aimed to assess whether patients newly diagnosed with myeloma benefit from antibiotic prophylaxis to prevent infection, and to investigate the effect on antibiotic-resistant organism carriage and health care-associated infections in patients with newly diagnosed myeloma. METHODS: TEAMM was a prospective, multicentre, double-blind, placebo-controlled randomised trial in patients aged 21 years and older with newly diagnosed myeloma in 93 UK hospitals. All enrolled patients were within 14 days of starting active myeloma treatment. We randomly assigned patients (1:1) to levofloxacin or placebo with a computerised minimisation algorithm. Allocation was stratified by centre, estimated glomerular filtration rate, and intention to proceed to high-dose chemotherapy with autologous stem cell transplantation. All investigators, patients, laboratory, and trial co-ordination staff were masked to the treatment allocation. Patients were given 500 mg of levofloxacin (two 250 mg tablets), orally once daily for 12 weeks, or placebo tablets (two tablets, orally once daily for 12 weeks), with dose reduction according to estimated glomerular filtration rate every 4 weeks. Follow-up visits occurred every 4 weeks up to week 16, and at 1 year. The primary outcome was time to first febrile episode or death from all causes within the first 12 weeks of trial treatment. All randomised patients were included in an intention-to-treat analysis of the primary endpoint. This study is registered with the ISRCTN registry, number ISRCTN51731976, and the EU Clinical Trials Register, number 2011-000366-35. FINDINGS: Between Aug 15, 2012, and April 29, 2016, we enrolled and randomly assigned 977 patients to receive levofloxacin prophylaxis (489 patients) or placebo (488 patients). Median follow-up was 12 months (IQR 8-13). 95 (19%) first febrile episodes or deaths occurred in 489 patients in the levofloxacin group versus 134 (27%) in 488 patients in the placebo group (hazard ratio 0·66, 95% CI 0·51-0·86; p=0·0018. 597 serious adverse events were reported up to 16 weeks from the start of trial treatment (308 [52%] of which were in the levofloxacin group and 289 [48%] of which were in the placebo group). Serious adverse events were similar between the two groups except for five episodes (1%) of mostly reversible tendonitis in the levofloxacin group. INTERPRETATION: Addition of prophylactic levofloxacin to active myeloma treatment during the first 12 weeks of therapy significantly reduced febrile episodes and deaths compared with placebo without increasing health care-associated infections. These results suggest that prophylactic levofloxacin could be used for patients with newly diagnosed myeloma undergoing anti-myeloma therapy. FUNDING: UK National Institute for Health Research.


Asunto(s)
Antibacterianos/uso terapéutico , Profilaxis Antibiótica/métodos , Neutropenia Febril/prevención & control , Infecciones/tratamiento farmacológico , Levofloxacino/uso terapéutico , Mieloma Múltiple/tratamiento farmacológico , Anciano , Método Doble Ciego , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Mieloma Múltiple/patología , Pronóstico , Estudios Prospectivos
5.
Arch Dis Child Fetal Neonatal Ed ; 101(6): F507-F512, 2016 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26951742

RESUMEN

BACKGROUND: Neonatal gram-negative (GN) infections are associated with high mortality and morbidity. Early appropriate antibiotic treatment is vital and gentamicin is the most frequently used antibiotic on neonatal units (NNUs). Antimicrobial breakpoints are predominantly based on adult data and the relationship between minimum inhibitory concentrations (MICs) and outcome in neonates is unclear. We aimed to determine the MIC of GN pathogens causing neonatal infections and relate this to clinical outcomes. METHODS: MICs for eight antibiotics plus extended spectrum ß-lactamase (ESBL) production were determined for invasive GN bacterial isolates from eight UK NNUs. European Committee on Antimicrobial Susceptibility Testing breakpoints were applied. MIC was correlated with clinical outcome using multivariable regression analysis. RESULTS: 118 isolates from 116 patients were analysed. The median birth gestation and postnatal age was 27 weeks (IQR 24.6-32.3) and 20 days (IQR 5-44), respectively. Pathogens included Escherichia coli (51%), Klebsiella spp (23%) and Enterobacter spp (22%). 10-day attributable mortality was 18.1% (21 patients) with the highest mortality from Pseudomonas aeruginosa infections. ESBL producers accounted for 13.8% of the isolates. In regression analysis, increasing gentamicin MIC was associated with increased mortality in gentamicin treated patients across the full MIC range (OR per loge increase in MIC: 2.29; 95% CI 1.23 to 4.26, p=0.009), including susceptible isolates only (MIC ≤4) (OR 3.05; 95% CI 1.10 to 8.46, p=0.032). CONCLUSIONS: Neonatal mortality from GN infections remains high and is associated with increasing gentamicin MIC, even for isolates deemed susceptible. A better understanding of population-specific MICs and aminoglycoside dosing is required to guide empiric antibiotic treatment.

6.
J Antimicrob Chemother ; 71(4): 897-902, 2016 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-26817487

RESUMEN

INTRODUCTION: Increasing use of nucleic acid amplification tests (NAATs) as the primary means of diagnosing gonococcal infection has resulted in diminished availability of Neisseria gonorrhoeae antimicrobial susceptibility data. We conducted a prospective diagnostic assessment of a real-time PCR assay (NGSNP) enabling direct detection of gonococcal ciprofloxacin susceptibility from a range of clinical sample types. METHODS: NGSNP, designed to discriminate an SNP associated with ciprofloxacin resistance within the N. gonorrhoeae genome, was validated using a characterized panel of geographically diverse isolates (n = 90) and evaluated to predict ciprofloxacin susceptibility directly on N. gonorrhoeae-positive NAAT lysates derived from genital (n = 174) and non-genital (n = 116) samples (n = 290), from 222 culture-confirmed clinical episodes of gonococcal infection. RESULTS: NGSNP correctly genotyped all phenotypically susceptible (n = 49) and resistant (n = 41) panel isolates. Ciprofloxacin-resistant N. gonorrhoeae was responsible for infection in 29.7% (n = 66) of clinical episodes evaluated. Compared with phenotypic susceptibility testing, NGSNP demonstrated sensitivity and specificity of 95.8% (95% CI 91.5%-98.3%) and 100% (95% CI 94.7%-100%), respectively, for detecting ciprofloxacin-susceptible N. gonorrhoeae, with a positive predictive value of 100% (95% CI 97.7%-100%). Applied to urogenital (n = 164), rectal (n = 40) and pharyngeal samples alone (n = 30), positive predictive values were 100% (95% CI 96.8%-100%), 100% (95% CI 87.2%-100%) and 100% (95% CI 82.4%-100%), respectively. CONCLUSIONS: Genotypic prediction of N. gonorrhoeae ciprofloxacin susceptibility directly from clinical samples was highly accurate and, in the absence of culture, will facilitate use of tailored therapy for gonococcal infection, sparing use of current empirical treatment regimens and enhancing acquisition of susceptibility data for surveillance.


Asunto(s)
Antibacterianos/uso terapéutico , Ciprofloxacino/farmacología , Genitales/microbiología , Gonorrea/tratamiento farmacológico , Gonorrea/microbiología , Pruebas de Sensibilidad Microbiana/métodos , Neisseria gonorrhoeae/efectos de los fármacos , Farmacorresistencia Bacteriana/efectos de los fármacos , Femenino , Humanos , Masculino , Medicina de Precisión , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los Resultados
7.
Lancet Infect Dis ; 13(11): 936-45, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24007915

RESUMEN

BACKGROUND: Diagnosis of Clostridium difficile infection is controversial because of many laboratory methods, compounded by two reference methods. Cytotoxigenic culture detects toxigenic C difficile and gives a positive result more frequently (eg, because of colonisation, which means that individuals can have the bacterium but no free toxin) than does the cytotoxin assay, which detects preformed toxin in faeces. We aimed to validate the reference methods according to clinical outcomes and to derive an optimum laboratory diagnostic algorithm for C difficile infection. METHODS: In this prospective, multicentre study, we did cytotoxigenic culture and cytotoxin assays on 12,420 faecal samples in four UK laboratories. We also performed tests that represent the three main targets for C difficile detection: bacterium (glutamate dehydrogenase), toxins, or toxin genes. We used routine blood test results, length of hospital stay, and 30-day mortality to clinically validate the reference methods. Data were categorised by reference method result: group 1, cytotoxin assay positive; group 2, cytotoxigenic culture positive and cytotoxin assay negative; and group 3, both reference methods negative. FINDINGS: Clinical and reference assay data were available for 6522 inpatient episodes. On univariate analysis, mortality was significantly higher in group 1 than in group 2 (72/435 [16·6%] vs 20/207 [9·7%], p=0·044) and in group 3 (503/5880 [8·6%], p<0·001), but not in group 2 compared with group 3 (p=0·4). A multivariate analysis accounting for potential confounders confirmed the mortality differences between groups 1 and 3 (OR 1·61, 95% CI 1·12-2·31). Multistage algorithms performed better than did standalone assays. INTERPRETATION: We noted no increase in mortality when toxigenic C difficile alone was present. Toxin (cytotoxin assay) positivity correlated with clinical outcome, and so this reference method best defines true cases of C difficile infection. A new diagnostic category of potential C difficile excretor (cytotoxigenic culture positive but cytotoxin assay negative) could be used to characterise patients with diarrhoea that is probably not due to C difficile infection, but who can cause cross-infection.


Asunto(s)
/aislamiento & purificación , Diarrea/diagnóstico , Diarrea/microbiología , Enterocolitis Seudomembranosa/diagnóstico , Enterocolitis Seudomembranosa/microbiología , Adolescente , Adulto , Anciano , Área Bajo la Curva , Niño , Preescolar , ADN Bacteriano/química , ADN Bacteriano/genética , Enterotoxinas/análisis , Heces/microbiología , Femenino , Glutamato Deshidrogenasa/análisis , Humanos , Modelos Logísticos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Valor Predictivo de las Pruebas , Estudios Prospectivos , Curva ROC , Sensibilidad y Especificidad , Adulto Joven
8.
FEMS Microbiol Lett ; 233(2): 333-9, 2004 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-15063504

RESUMEN

Alternative sigma factors are key global regulators that coordinate bacterial responses to environmental changes necessary for adaptation and survival. In turn these sigma factors are controlled by regulators such as anti-sigma and anti-anti-sigma factors. In this report, using a cDNA-total RNA subtractive hybridisation strategy that we have developed previously, we identified increased transcription of a potential sigma factor regulatory gene, Rv1364c, in Mycobacterium bovis BCG upon phagocytosis by macrophages and this was confirmed by Northern blot analysis. Primer extension analysis revealed the use of alternative promotors, P1 and P2, and that the increased expression inside macrophages coincided with promoter switching from P2 to P1. Rv1364c (653 amino acids), originally annotated as RsbU, contains structural domains homologous to the PAS redox sensor, the protein phosphatases anti-anti-sigma factor RsbU/SpoIIE, the protein kinase anti-sigma factor RsbW/SpoIIAB and the anti-anti-sigma factor RsbV/SpoIIAA found in other bacteria. These findings have important implications for understanding coordination of the expression of sigma factors under intra-macrophage conditions. Other potentially differentially expressed genes, including genes for fatty acid metabolism, membrane transportors, heat shock proteins, potential sigma factors and energy metabolic pathways are also listed and their biological significance discussed.


Asunto(s)
Macrófagos/microbiología , Infecciones por Mycobacterium/microbiología , Mycobacterium bovis/genética , Mycobacterium bovis/metabolismo , Factor sigma/genética , Secuencia de Bases , Regulación Bacteriana de la Expresión Génica , Humanos , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Regiones Promotoras Genéticas/fisiología , Transcripción Genética/fisiología
9.
J Exp Med ; 198(5): 693-704, 2003 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-12953091

RESUMEN

Little is known about the biochemical environment in phagosomes harboring an infectious agent. To assess the state of this organelle we captured the transcriptional responses of Mycobacterium tuberculosis (MTB) in macrophages from wild-type and nitric oxide (NO) synthase 2-deficient mice before and after immunologic activation. The intraphagosomal transcriptome was compared with the transcriptome of MTB in standard broth culture and during growth in diverse conditions designed to simulate features of the phagosomal environment. Genes expressed differentially as a consequence of intraphagosomal residence included an interferon gamma- and NO-induced response that intensifies an iron-scavenging program, converts the microbe from aerobic to anaerobic respiration, and induces a dormancy regulon. Induction of genes involved in the activation and beta-oxidation of fatty acids indicated that fatty acids furnish carbon and energy. Induction of sigmaE-dependent, sodium dodecyl sulfate-regulated genes and genes involved in mycolic acid modification pointed to damage and repair of the cell envelope. Sentinel genes within the intraphagosomal transcriptome were induced similarly by MTB in the lungs of mice. The microbial transcriptome thus served as a bioprobe of the MTB phagosomal environment, showing it to be nitrosative, oxidative, functionally hypoxic, carbohydrate poor, and capable of perturbing the pathogen's cell envelope.


Asunto(s)
Regulación Bacteriana de la Expresión Génica , Macrófagos/microbiología , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Fagosomas/microbiología , Transcripción Genética , Animales , Ratones , ARN Bacteriano/genética
10.
Microbiology (Reading) ; 147(Pt 8): 2293-2305, 2001 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-11496006

RESUMEN

Identifying genes that are differentially expressed by Mycobacterium bovis BCG after phagocytosis by macrophages will facilitate the understanding of the molecular mechanisms of host cell-intracellular pathogen interactions. To identify such genes a cDNA-total RNA subtractive hybridization strategy has been used that circumvents the problems both of limited availability of bacterial RNA from models of infection and the high rRNA backgrounds in total bacterial RNA. The subtraction products were used to screen a high-density gridded Mycobacterium tuberculosis genomic library. Sequence data were obtained from 19 differential clones, five of which contained overlapping sequences for the gene encoding mycocerosic acid synthase (mas). Mas is an enzyme involved in the synthesis of multi-methylated long-chain fatty acids that are part of phthiocerol dimycocerosate, a major component of the complex mycobacterial cell wall. Northern blotting and primer extension data confirmed up-regulation of mas in intracellular mycobacteria and also revealed a putative extended -10 promoter structure and a long untranslated upstream region 5' of the mas transcripts, containing predicted double-stranded structures. Furthermore, clones containing overlapping sequences for furB, groEL-2, rplE and fadD28 were identified and the up-regulation of these genes was confirmed by Northern blot analysis. The cDNA-RNA subtractive hybridization enrichment and high density gridded library screening, combined with selective extraction of bacterial mRNA represents a valuable approach to the identification of genes expressed during intra-macrophage residence for bacteria such as M. bovis BCG and the pathogenic mycobacterium, M. tuberculosis.


Asunto(s)
Aciltransferasas/metabolismo , Regulación Bacteriana de la Expresión Génica , Macrófagos/microbiología , Mycobacterium bovis/enzimología , Mycobacterium bovis/crecimiento & desarrollo , Hibridación de Ácido Nucleico/métodos , Aciltransferasas/química , Aciltransferasas/genética , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Cartilla de ADN , ADN Complementario , Biblioteca Genómica , Humanos , Datos de Secuencia Molecular , Mycobacterium bovis/genética , Mycobacterium tuberculosis/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Regiones Promotoras Genéticas , ARN Bacteriano/genética , ARN Bacteriano/metabolismo , ARN Mensajero/metabolismo , Transcripción Genética , Células Tumorales Cultivadas
11.
Microbiology (Reading) ; 147(Pt 2): 459-471, 2001 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-11158363

RESUMEN

Mycobacterium tuberculosis resides within the macrophages of the host, but the molecular and cellular mechanisms of survival are poorly understood. Recent evidence suggests that the attenuated vaccine strain Mycobacterium bovis BCG is both a deletion and regulatory mutant, yet retains both its immunoprotective and intra-macrophage survival potential. In an attempt to define M. bovis BCG genes expressed during interaction with macrophages, the patterns of protein synthesis were examined by both one- and two-dimensional gel electrophoresis of BCG while inside the human leukaemic macrophage cell line THP-1. This study demonstrated that BCG expresses proteins while resident inside macrophages that are not expressed during in vitro growth in culture media or under conditions of heat shock. Western blotting analysis revealed that some of the differentially expressed proteins are specifically recognized by human M. tuberculosis-infected sera. Proteome analysis by two-dimensional electrophoresis and MS identified six abundant proteins that showed increased expression inside macrophages: 16 kDa alpha-crystallin (HspX), GroEL-1 and GroEL-2, a 31.7 kDa hypothetical protein (Rv2623), InhA and elongation factor Tu (Tuf). Identification of proteins by such a strategy will help elucidate the molecular basis of the attenuation and the vaccine potential of BCG, and may provide antigens that distinguish infection with M. tuberculosis from vaccination with BCG.


Asunto(s)
Proteínas Bacterianas/metabolismo , Macrófagos/microbiología , Mycobacterium bovis/metabolismo , Fagocitosis , Proteoma , Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/genética , Western Blotting , Electroforesis en Gel Bidimensional , Humanos , Mycobacterium bovis/crecimiento & desarrollo , Mycobacterium tuberculosis/inmunología , Tuberculosis/inmunología , Tuberculosis/microbiología
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