EFLM Task Force Preparation of Labs for Emergencies (TF-PLE) recommendations for reinforcing cyber-security and managing cyber-attacks in medical laboratories.

The healthcare systems are a prime target for cyber-attacks due to the sensitive nature of the information combined with the essential need for continuity of care. Medical laboratories are particularly vulnerable to cyber-attacks for a number of reasons, including the high level of information technology (IT), computerization and digitization. Based on reliable and widespread evidence that medical laboratories may be inadequately prepared for cyber-terrorism, a panel of experts of the Task Force Preparation of Labs for Emergencies (TF-PLE) of the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) has recognized the need to provide some general guidance that could help medical laboratories to be less vulnerable and better prepared for the dramatic circumstance of a disruptive cyber-attack, issuing a number of consensus recommendations, which are summarized and described in this opinion paper.

Intravenous Artesunate for the Treatment of Severe Imported Malaria: Implementation, Efficacy, and Safety in 1391 Patients.

BACKGROUND: Intravenous artesunate is the World Health Organization-recommended first-line treatment for severe malaria worldwide, but it is still not fully licensed in Europe. Observational studies documenting its safety and efficacy in imported malaria are thus essential. METHODS: We prospectively collected clinical and epidemiological features of 1391 artesunate-treated patients among 110 participant centers during the first 7 years (2011-2017) of a national program implemented by the French Drug Agency. RESULTS: Artesunate became the most frequent treatment for severe malaria in France, rising from 9.9% in 2011 to 71.4% in 2017. Mortality was estimated at 4.1%. Treatment failure was recorded in 27 patients, but mutations in the Kelch-13 gene were not observed. Main reported adverse events (AEs) were anemia (136 cases), cardiac events (24, including 20 episodes of conduction disorders and/or arrhythmia), and liver enzyme elevation (23). Mortality and AEs were similar in the general population and in people with human immunodeficiency virus, who were overweight, or were pregnant, but the only pregnant woman treated in the first trimester experimented a hemorrhagic miscarriage. The incidence of post-artesunate-delayed hemolysis (PADH) was 42.8% when specifically assessed in a 98-patient subgroup, but was not associated with fatal outcomes or sequelae. PADH was twice as frequent in patients of European compared with African origin. CONCLUSIONS: Artesunate was rapidly deployed and displayed a robust clinical benefit in patients with severe imported malaria, despite a high frequency of mild to moderate PADH. Further explorations in the context of importation should assess outcomes during the first trimester of pregnancy and collect rare but potentially severe cardiac AEs.

Characterization of four BCHE mutations associated with prolonged effect of suxamethonium.

Butyrylcholinesterase (BChE) deficiency is characterized by prolonged apnea after the use of muscle relaxants (suxamethonium or mivacurium) in patients who have mutations in the BCHE gene. Here, we report the characterization of four BCHE mutations associated with prolonged effect of suxamethonium (amino acid numbering based on the matured enzyme): p.20delValPheGlyGlyThrValThr, p.Leu88His, p.Ile140del and p.Arg386Cys. Expression of recombinant BCHE mutants, kinetic analysis and molecular dynamics were undertaken to understand how these mutations induce BChE deficiency. Three of the mutations studied (p.20delValPheGlyGlyThrValThr, p.Ile140del and p.Arg386Cys) lead to a "silent" BChE phenotype. Recombinant BCHE expression studies for these mutants revealed BChE activity levels comparable to untransfected cells. Only the last one (hBChE-L88H) presented BChE activity in the transfected cell culture medium. This BChE mutant (p.Leu88His) is associated with a lower kcat value compare to the wild-type enzyme. Molecular dynamics simulations analyses suggest that a destabilization of a structure implicated in enzyme activity (Ω-loop) can explain the modification of the kinetic parameter of the mutated protein.

Performances, feasibility and acceptability of nasopharyngeal swab, saliva and oral-self sampling swab for the detection of severe acute respiratory syndrome coronavirus 2.

Molecular diagnosis on nasopharyngeal swabs (NPS) is the current standard for COVID-19 diagnosis, but saliva may be an alternative specimen to facilitate access to diagnosis. We compared analytic performances, feasibility and acceptability of NPS, saliva, and oral-self sampling swab for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A prospective, multicenter study was conducted in military hospitals in France among adult outpatients attending COVID-19 diagnosis centers or hospitalized patients. For each patient, all samples were obtained and analyzed simultaneously with RT-PCR or transcription-mediated amplification method. Clinical signs, feasibility, and acceptability for each type of sample were collected. A total of 1220 patients were included, corresponding to 1205 NPS and saliva and 771 OS. Compared to NPS, the sensitivity, specificity, and kappa coefficient for tests performed on saliva were 87.8% (95% CI 83.3-92.3), 97.1% (95% CI 96.1-98.1), and 0.84 (95% CI 0.80-0.88). Analytical performances were better in symptomatic patients. Ct values were significantly lower in NPS than saliva. For OS, sensitivity was estimated to be 61.1% (95% CI 52.7-69.4) and Kappa coefficient to be 0.69 (95% CI 0.62-0.76). OS was the technique preferred by the patients (44.3%) before saliva (42.4%) and NPS (13.4%). Instructions were perceived as simple by patients (> 90%) for saliva and OS. Finally, the painful nature was estimated to be 0.9 for OS, on a scale from 0 to 10, and to be 5.3 for NPS. Performances of OS are not sufficient. Saliva is an acceptable alternative to NPS for symptomatic patient but the process required additional steps to fluidize the sample.

The C5 Variant of the Butyrylcholinesterase Tetramer Includes a Noncovalently Bound 60 kDa Lamellipodin Fragment.

Humans with the C5 genetic variant of butyrylcholinesterase (BChE) have 30-200% higher plasma BChE activity, low body weight, and shorter duration of action of the muscle relaxant succinylcholine. The C5 variant has an extra, slow-moving band of BChE activity on native polyacrylamide gel electrophoresis. This band is about 60 kDa larger than wild-type BChE. Umbilical cord BChE in 100% of newborn babies has a C5-like band. Our goal was to identify the unknown, 60 kDa protein in C5. Both wild-type and C5 BChE are under the genetic control of two independent loci, the BCHE gene on Chr 3q26.1 and the RAPH1 (lamellipodin) gene on Chr 2q33. Wild-type BChE tetramers are assembled around a 3 kDa polyproline peptide from lamellipodin. Western blot of boiled C5 and cord BChE showed a positive response with an antibody to the C-terminus of lamellipodin. The C-terminal exon of lamellipodin is about 60 kDa including an N-terminal polyproline. We propose that the unknown protein in C5 and cord BChE is encoded by the last exon of the RAPH1 gene. In 90% of the population, the 60 kDa fragment is shortened to 3 kDa during maturation to adulthood, leaving only 10% of adults with C5 BChE.

Molecular characterization of blaNDM-1 in a sequence type 235 Pseudomonas aeruginosa isolate from France.

An NDM-1 carbapenemase-producing Pseudomonas aeruginosa isolate was recovered from a patient hospitalized in France after a previous hospitalization in Serbia. Genetic studies revealed that the blaNDM-1 gene was surrounded by insertion sequence ISAba125 and a truncated bleomycin resistance gene. This blaNDM-1 region was a part of the variable region of a new complex class 1 integron bearing IS common region 1 (ISCR1). The presence of ISPa7 upstream of this integron suggests insertion in a chromosomally located Tn402-like structure.

Pharmacogenetic Guidelines for Psychotropic Drugs: Optimizing Prescriptions in Clinical Practice.

The modalities for prescribing a psychotropic (dose and choice of molecule) are currently unsatisfactory, which can lead to a lack of efficacy of the treatment associated with prolonged exposure of the patient to the symptoms of his or her illness and the side effects of the molecule. In order to improve the quality of treatment prescription, a part of the current biomedical research is dedicated to the development of pharmacogenetic tools for individualized prescription. In this guideline, we will present the genes of interest with level 1 clinical recommendations according to PharmGKB for the two major families of psychotropics: antipsychotics and antidepressants. For antipsychotics, there are CYP2D6 and CYP3A4, and for antidepressants, CYP2B6, CYP2D6, and CYP2C19. The study will focus on describing the role of each gene, presenting the variants that cause functional changes, and discussing the implications for prescriptions in clinical practice.

Phenotype and genotype characteristics of 58 patients showing a prolonged effect of succinylcholine: A four-year experience.

INTRODUCTION: This study sought to describe the phenotype and genotype characteristics of patients referred to our laboratory to undergo further assessment due to a suspicion of a prolonged effect of suxamethonium attributed to BChE deficiency. METHODS: All patients referred to our laboratory from January 2016 to December 2019 due to the suspicion of a prolonged effect of suxamethonium were included in this study. The determination of BChE activity and genotyping using complete nucleotide sequencing of the entire complementary DNA-coding region with flanking intron-exon boundaries were completed. RESULTS: During this four-year period, 58 patients were referred to our laboratory for the investigation of prolonged neuromuscular block due to BChE deficiency. Among them, 52 showed a BChE deficiency related to BCHE gene mutations. The most commonly detected genotype was compound homozygous atypical variant (p.Asp98Gly)/homozygous Kalow variant (p.Ala569Thr) (p.[Asp98Gly;Ala567Thr];[p.Asp98Gly;Ala567Thr]). Further, we recorded four new BCHE variants, which seem to be associated with prolonged post suxamethonium apnoea: p.(Trp205Cys), p.(Leu222His), p.(Glu469Gln), and p.(Lys276Ter). CONCLUSION: During a four-year period, among the 58 patients referred to our laboratory, we have found four new BCHE variants, which seem to be associated with prolonged post suxamethonium apnoea (p.(Trp205Cys), p.(Leu22His), p.(Glu469Gln), and p.(Lys276Ter)).

[Tuberculous meningitis: diagnosis and therapeutic difficulties]. / Méningites tuberculeuses: difficultés diagnostiques et thérapeutiques.

Tuberculosis remains a public-health problem in 2010 with 9 millions cases and 1,7 million deaths worldwide each year. Tuberculosis meningitis is rare (0.5 to 1%) but is associated with high mortality and disability among survivors. An early starting of treatment is crucial. Despite molecular biology methods, microbiological diagnosis remains a challenge for the biologist. We report here 2 cases of tuberculous meningitis with different clinical and biological presentations, which underline diagnosis and therapeutic difficulties encountered in the management of this disease. The first one occurred in an HIV infected patient and the second one was caused by a multidrug-resistant strain. Clinical issues were severe with important neurological residual disability and death. Biological methods available for tuberculous meningitis diagnosis are exposed.

[SFBC working group "Biochemical markers of COVID-19"]. / Groupe de travail SFBC « Marqueurs biochimiques de COVID-19 ¼.

The SARS-CoV-2 virus is responsible for an epidemic disease called COVID-19, which was initially evidenced in Wuhan, China, and spread very rapidly in China and around the world. In France, the first isolated case seems now to be reported in December 2019, stage 3 of the COVID-19 epidemic was triggered on March 14th, the start of the planned containment exit from May 11th. Healthcare services have faced a large influx of patients who may be beyond their capacity to receive and care, particularly in the Large-East and Ile-de-France regions. Some patients show an evolution of the disease never observed before with other coronaviruses and develop in a few days a very important inflammatory reaction, which can lead to death of patients. A working group of the French Society of Clinical Biology (SFBC) was set up with the objective of providing updated information on the current status of the biological prescriptions (focusing on biochemistry ones) and their evolution during the epidemic, and of analyzing the biological parameters associated with comorbidities and patient evolution in order to link biological results with medical events. The expanded working group covers all sectors of medical biology in France and extends to the French-speaking world: hospital sectors (CHU and CH, Army Training Hospitals) and the private sector opening a field of view on the biological situation in establishments for dependent elderly, social establishments and clinical medical institutions. The purpose of this article is the presentation of this working group and its immediate and future actions.

Rare Dihydropyrimidine Dehydrogenase Variants and Toxicity by Floropyrimidines: A Case Report.

Variations in the activity, up to absolute deficiency, of the enzyme dihydropyrimidine dehydrogenase (DPD), result in the occurrence of adverse reactions to chemotherapy, and have been included among the pharmacogenetic factors underlying inter-individual variability in response to fluoropyrimidines. The study of single-nucleotide polymorphisms of the DPYD gene, which encodes the DPD enzyme, is one of the main parameters capable of predicting reduced enzymatic activity and the consequent influence on fluoropyrimidine treatment, in terms of reduction of both adverse reactions and therapeutic efficacy in disease control. In this paper, we describe a patient with metastatic breast cancer showing signs of increased toxicity following capecitabine therapy. The DPD enzyme activity analysis revealed a partial deficiency. The study of the most frequent polymorphisms of the DPYD gene suggested a wild-type genotype but indicated a novel variant c.1903A>G (p.Asn635Asp), not previously described, proximal to the splice donor site of exon 14. After excluding the potential pathogenic feature of the newly-identified variant, we performed cDNA sequencing of the entire DPYD coding sequence. This analysis identified the variants c.85T>C and c.496A>G, which were previously described as pivotal components of the haplotype associated with decreased enzyme activity and suggested that both variant alleles are related to DPD deficiency. The clinical case findings described in this study emphasize the importance of performing complete genetic analysis of the DPYD gene in order to identify rare and low frequency variants potentially responsible for toxic reactions to fluoropyrimidine treatment.

Molecular epidemiological of extended-spectrum ß-lactamase producing Escherichia coli isolated in Djibouti.

INTRODUCTION: While the molecular epidemiology of extended-spectrum-b-lactamase (ESBL)-producing E. coli is well known in Europe due to effective surveillance networks and substantial literature, data for Africa are less available, especially in Djibouti. METHODOLOGY: We studied 31 isolates of ESBL-producing E. coli from Djibouti and compared these molecular results with data available in Africa. RESULTS: Susceptibility rates were 3.2% for ceftazidim, 48.4% for piperacillin-tazobactam, 90.3% for amikacine and 16.1% for ofloxacin. No isolate showed resistance to carbapenems or colistin. 30 E. coli (96.8%) were positive to blaCTX-M-15, 1 (3.2%) to blaCTX-M-14  and 10 (32.3%) to narrow-broad-spectrum blaTEM. No blaSHV were detected. Fluoroquinolone resistance analysis showed that 30 ofloxacin-resistant E. coli had the mutation Ser-83->Leu on the gyrA gene. 24 E. coli (77.4%) harboured the plasmid-borne aac(6 ')-Ib-cr gene. No E. coli carried the genes qnrA, qnrB and qepA. 10 isolates (32.3%) belonging to the ST131 clone. The plasmid incompatibility group most widely represented in our collection was IncFIA/IB/II. CONCLUSIONS: There is no major difference with African epidemiology. In particular, we notice the international diffusion of specific clonal group ST131.