Antimicrobial Resistance Trends of Escherichia coli Isolates from Outpatient and Inpatient Urinary Infections over a 20-Year Period.

Antimicrobial resistance is a worldwide problem, and resistance in Enterobacteriaceae, particularly Escherichia coli and Klebsiella pneumoniae, is a critical threat to human health. Inappropriate and unnecessary use of antibiotics in human health care is the most common cause for the development and spread of antimicrobial resistance. In this work, we retrospectively analyzed the antimicrobial data of E. coli strains isolated from midstream urinary samples over a 20-year period (2000-2019). The aim was to provide useful information to clinicians to prescribe a more appropriate empirical antibiotic therapy. A total of 30,955 unique E. coli isolates from positive midstream urine samples of inpatients (1,198) and outpatients (29,757) were collected. Except for carbapenems, over time all the antibiotics tested showed increasing resistance rates in both groups (p < 0.0001). On the other hand, fosfomycin and nitrofurantoin presented significant decreasing trends in resistance rate (p < 0.05). There were significant increases in extended-spectrum ß-lactamases- and multidrug resistance positive isolates starting in 2000 (p < 0.0001), with similar results in both groups. Ciprofloxacin, gentamicin, trimethoprim/sulfamethoxazole, and third-generation cephalosporin resistances significantly increased with increasing age (p < 0.0001). Collectively, E. coli resistance rates severely increased during the study period, except for fosfomycin and nitrofurantoin. The need of monitoring studies about antibiotic nonsusceptibilities at local and regional levels are necessary to enhance the focus on antimicrobial stewardship, to reduce antimicrobial consumption and to detect alarming resistance mechanisms.

Baseline characteristics of COVID-19 Italian patients admitted to Desio Hospital, Lombardy: a retrospective study.

The correlation of clinical, radiological and laboratory findings of patients at admission in the Emergency Department (ED) with clinical severity and risk of mortality was investigated. Adult coronavirus disease 2019 (COVID-19) patients hospitalized in March 2020 in Desio Hospital, Lombardy, were retrospectively included in the study, and categorized in terms of disease severity and adverse outcome. Out of the 175 patients enrolled, 79% presented one or more comorbidities, with cardiovascular disease being the most frequent (62%). More than half of the patients showed lymphocytopenia and 20% thrombocytopenia. The patients in the severe group presented higher absolute neutrophil count (ANC), C-reactive protein (CRP), AST, LDH, procalcitonin (PCT) and BUN values compared to the non-severe group (p < .05). Increased odds of mortality associated with older age (OR = 22.43; 95% CI 5.22-96.27), partial pressure of oxygen (PaO2)/fraction of inspired oxygen (FIO2) ratio < 200 (OR = 4.97; 95% CI 1.55-15.84), clinical severity (OR = 21.32; 95% CI 2.27-200.13), creatinine > 106.08 µmol/L (OR = 2.87; 95% CI 1.04-7.92) and creatine kinase > 2.90 µkat/L (OR = 3.80; 95% CI 1.31-10.9) were observed on admission (p < .05). The above findings may contribute to identify early risk factors of poor prognosis, and to select the most appropriate management for patients.

Next Generation Sequencing in Newborn Screening in the United Kingdom National Health Service.

Next generation DNA sequencing (NGS) has the potential to improve the diagnostic and prognostic utility of newborn screening programmes. This study assesses the feasibility of automating NGS on dried blood spot (DBS) DNA in a United Kingdom National Health Service (UK NHS) laboratory. An NGS panel targeting the entire coding sequence of five genes relevant to disorders currently screened for in newborns in the UK was validated on DBS DNA. An automated process for DNA extraction, NGS and bioinformatics analysis was developed. The process was tested on DBS to determine feasibility, turnaround time and cost. The analytical sensitivity of the assay was 100% and analytical specificity was 99.96%, with a mean 99.5% concordance of variant calls between DBS and venous blood samples in regions with ≥30× coverage (96.8% across all regions; all variant calls were single nucleotide variants (SNVs), with indel performance not assessed). The pipeline enabled processing of up to 1000 samples a week with a turnaround time of four days from receipt of sample to reporting. This study concluded that it is feasible to automate targeted NGS on routine DBS samples in a UK NHS laboratory setting, but it may not currently be cost effective as a first line test.

A comparison of methods for EGFR mutation testing in non-small cell lung cancer.

EGFR mutation testing of tumor samples is routinely performed to predict sensitivity to treatment with tyrosine kinase inhibitors for patients with non-small cell lung cancer. At least 9 different methodologies are employed in UK laboratories, and the aim of this study was to compare the sensitivity of different methods for the detection of EGFR mutations. Participating laboratories were sent coded samples with varying mutation loads (from 0% to 15%) to be tested for the p.Leu858Arg (p.L858R) missense mutation and c.2235_2249del exon 19 deletion. The p.L858R mutation and deletions within exon 19 of the EGFR gene account for ∼90% of mutation-positive cases. The 11 laboratories used their standard testing method(s) and submitted 15 sets of results for the p.L858R samples and 10 for the exon 19 deletion. The p.Leu858Arg (p.L858R) mutation was detected at levels between 1% and 7.5% by Sanger sequencing, pyrosequencing, real-time polymerase chain reaction (PCR), amplification refractory mutation system, and capillary electrophoresis single-strand conformation analysis. The c.2235_2249del mutation was detected at 1% to 5% by fragment size analysis, Sanger sequencing or real-time PCR. A mutation was detected in 24/25 (96%) of the samples tested which contained 5% mutated DNA. The 1% sensitivity claimed for commercial real-time PCR-targeted EGFR tests was achieved and our results show greater sensitivity for the Sanger sequencing and pyrosequencing screening methods compared to the 10% to 20% detection levels cited on clinical diagnostic reports. We conclude that multiple methodologies are suitable for the detection of acquired EGFR mutations.

C9ORF72 expansions, parkinsonism, and Parkinson disease: a clinicopathologic study.

OBJECTIVE: To determine the histopathologic bases for the observed incidence of parkinsonism in families with C9ORF72 expansions, which typically cause amyotrophic lateral sclerosis (ALS) and/or frontotemporal dementia. METHODS: DNA was extracted from 377 brains with the histopathologic diagnosis of idiopathic Parkinson disease or related disorders and analyzed for C9ORF72 expansions. α-Synuclein and p62 immunohistochemistry of the substantia nigra (SN) was undertaken in brains of 17 ALS cases with (C9ORF72+) and 51 without (C9ORF72-) the C9ORF72 expansion. RESULTS: Only 1 of 338 cases with pathologically confirmed idiopathic Parkinson disease had a C9ORF72 expansion. Similarly, only 1 of 17 C9ORF72+ brains displayed features suggestive of α-synucleinopathy. In contrast, p62-positive, TDP-43-negative neuronal cytoplasmic inclusions within the SN were considerably more frequent in C9ORF72+ brain tissue than in the C9ORF72- brains (p = 0.005). Furthermore, there was a more marked loss of dopaminergic neurons in the SN of C9ORF72+ ALS brains than C9ORF72- ALS brains (p = 0.029). CONCLUSIONS: SN involvement is common in C9ORF72+ ALS but can be clearly distinguished from Parkinson disease-related mechanisms by the presence of p62-positive inclusions and the absence of α-synuclein-positive Lewy bodies or Lewy neurites.

Concurrence of multiple sclerosis and amyotrophic lateral sclerosis in patients with hexanucleotide repeat expansions of C9ORF72.

BACKGROUND: Crossover in the pathogenic mechanisms of amyotrophic lateral sclerosis (ALS) and multiple sclerosis (MS) has been described but is poorly understood. A GGGGCC hexanucleotide repeat expansion of C9ORF72 has recently been identified in a significant proportion of patients with ALS. METHODS: In approximately 650 patients diagnosed with ALS from the North of England we identified seven patients who initially presented with MS. DNA obtained from five patients with MS-ALS and 215 patients with MS alone was screened for the C9ORF72 expansion. Post-mortem material was examined from two patients with MS-ALS. Gene expression profiling was performed on lymphoblastoid cells and levels of CXCL10 were measured in cerebrospinal fluid (CSF) from patients with ALS with and without the C9ORF72 expansion and controls. RESULTS: Concurrence of MS and ALS is higher than expected in our population. The C9ORF72 expansion was identified in 80% of patients with MS-ALS but not in those with MS alone. In the presence of preceding MS, C9ORF72-ALS was more rapidly progressive. MetaCore analysis identified alteration of the NF-кB pathway in C9ORF72-ALS and non-C9ORF72-ALS. NF-кB activation is associated with increased expression of the neuroprotective cytokine CXCL10 but, in C9ORF72-ALS, CXCL10 is downregulated and CSF levels are reduced. CONCLUSIONS: We propose that MS-associated neuroinflammation may affect penetrance and progression of the C9ORF72 expansion. In particular, the NF-кB pathway is activated in MS and appears to be dysfunctional in C9ORF72-ALS. Aberrant downregulation of CXCL10 may explain the predisposition of C9ORF72 expansion carriers to develop ALS in the context of MS and NF-кB activation, and offers a potential therapeutic target.

Clinico-pathological features in amyotrophic lateral sclerosis with expansions in C9ORF72.

Intronic expansion of the GGGGCC hexanucleotide repeat within the C9ORF72 gene causes frontotemporal dementia and amyotrophic lateral sclerosis/motor neuron disease in both familial and sporadic cases. Initial reports indicate that this variant within the frontotemporal dementia/amyotrophic lateral sclerosis spectrum is associated with transactive response DNA binding protein (TDP-43) proteinopathy. The amyotrophic lateral sclerosis/motor neuron disease phenotype is not yet well characterized. We report the clinical and pathological phenotypes associated with pathogenic C9ORF72 mutations in a cohort of 563 cases from Northern England, including 63 with a family history of amyotrophic lateral sclerosis. One hundred and fifty-eight cases from the cohort (21 familial, 137 sporadic) were post-mortem brain and spinal cord donors. We screened DNA for the C9ORF72 mutation, reviewed clinical case histories and undertook pathological evaluation of brain and spinal cord. Control DNA samples (n = 361) from the same population were also screened. The C9ORF72 intronic expansion was present in 62 cases [11% of the cohort; 27/63 (43%) familial, 35/500 (7%) cases with sporadic amyotrophic lateral sclerosis/motor neuron disease]. Disease duration was significantly shorter in cases with C9ORF72-related amyotrophic lateral sclerosis (30.5 months) compared with non-C9ORF72 amyotrophic lateral sclerosis/motor neuron disease (36.3 months, P < 0.05). C9ORF72 cases included both limb and bulbar onset disease and all cases showed combined upper and lower motor neuron degeneration (amyotrophic lateral sclerosis). Thus, clinically, C9ORF72 cases show the features of a relatively rapidly progressive, but otherwise typical, variant of amyotrophic lateral sclerosis associated with both familial and sporadic presentations. Dementia was present in the patient or a close family member in 22/62 cases with C9ORF72 mutation (35%) based on diagnoses established from retrospective clinical case note review that may underestimate significant cognitive changes in late disease. All the C9ORF72 mutation cases showed classical amyotrophic lateral sclerosis pathology with TDP-43 inclusions in spinal motor neurons. Neuronal cytoplasmic inclusions and glial inclusions positive for p62 immunostaining in non-motor regions were strongly over-represented in the C9ORF72 cases. Extra-motor pathology in the frontal cortex (P < 0.0005) and the hippocampal CA4 subfield neurons (P < 0.0005) discriminated C9ORF72 cases strongly from the rest of the cohort. Inclusions in CA4 neurons were not present in non-C9ORF72 cases, indicating that this pathology predicts mutation status.

Contribuiçäo ao plano de vacinaçäo anti-sarampo / Vaccination plan against measles

Este trabalho mostra os resultados positivos da intensificaçäo da campanha de vacinaçäo contra o sarampo, denominada "Plano Impacto", realizada nos dias 30 e 31 de julho de 1984 com o apoio da Secretaria da Saúde do Estado de Säo Paulo. Pelo "Plano Impacto", vacinou-se aproximadamente 3,6 vezes mais que nos dias normais da campanha de vacinaçäo. A cobertura vacinal após a campanha foi de 89,7%, oferecendo uma segurança à imunidade populacional local. Avaliou-se o lucro imediato de cada criança vacinada, representado pela näo hospitalizaçäo por sarampo ou suas complicaçöes (Cr$ 102,94 - Cr$ 3.356,71), confrontando-o com o custo de uma dose da vacina (Cr$608,00). Discutiu-se também a idade ideal para vacinaçäo contra o sarampo, que depende da epidemiologia local da doença. Questionou-se o risco dado pela mudança desta idade (de 7 a 9 meses) adotada desde o mês de abril de 1984, no Brasil