Validation of an HPLC-MS/MS method with QuEChERS extraction using isotopic dilution to analyze chlordecone in human serum.

Until 1993, chlordecone (CLD) was extensively used in banana fields in French West Indies. In a previous study, CLD was detected in 90 % of Martinican and Guadeloupean adult's serum. In order to simplify the analyses of CLD in the serum, a new QuEChERS-HPLC-MS/MS method was implemented and validated by the Pasteur Institute of Guadeloupe (IPG). This method was validated with accuracy profiles according to the French Standard NF V03-110 plus the ISO 15189 and European guidelines. Linearity, repeatability, accuracy, intermediate precision, specificity, limit of detection (LOD), limit of quantification (LOQ) and uncertainty were determined. The accuracy profile allowed the method to be validated between 0.06  µg L-1 and 1.00  µg L-1 of serum. The LOD was 0.02  µg L-1, the LOQ was 0.06  µg L-1 and the uncertainty of the method was 21 %. A comparison of 49 serum samples between the IPG (LC-MS/MS) and the LEAE-CART (GC-HRMS) laboratories demonstrated that this new method can reliably determine CLD in human serum. Stability tests were performed and duration of the storage of raw samples and extracts before analysis by HPLC-MS/MS. Raw samples were stable after collection for at least one week at 5 °C or 25 °C and for at least 3 months at -20 °C. Extracts in acetonitrile were stable for at least 1 month at -20 °C. These stability results facilitate the daily use of the method. This method should help the entire population of Guadeloupe and Martinique by allowing a routinely analyzed for CLD and will be useful for future projects aimed at improving population health monitoring.

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.

Thromboelastographic study of the snakebite-related coagulopathy in Djibouti.

: Hemostasis disorders are one of the major clinical conditions of snakebites and are because of mechanisms which may disrupt vessels, platelets, clotting factors and fibrinolysis. Thromboelastography (TEG) could help to understand these effects in the clinical practice. A retrospective study reports a series of patients presenting a snakebite-related coagulopathy, treated with antivenom and monitored with conventional tests and TEG in a French military treatment facility (Republic of Djibouti, East Africa) between August 2011 and September 2013. Conventional coagulation assays (platelets, prothrombin time, activated partial thromboplastin time, fibrinogen) and TEG measurements were taken on arrival and at various times during the first 72 h of hospitalization, at the discretion of the physician. The study included 14 patients (median age 28 years). Bleedings were present in five patients. All patients received antivenom. A coagulopathy was present in all patients and was detected by both conventional assays and TEG. None exhibited thrombocytopenia. Prothrombin time and fibrinogen remained abnormal for most of patients during the first 72 h. The TEG profiles of 11 patients (79%) showed incoagulability at admission (R-time > 60 min). TEG distinguished 10 patients with a generalized clotting factor deficiency and 4 patients with an isolated fibrinogen deficiency after an initial profile of incoagulability. Hyperfibrinolysis was evident for 12 patients (86%) after Hour 6. Snake envenomations in Djibouti involve a consumption coagulopathy in conjunction with delayed hyperfibrinolysis. TEG could improve medical management of the condition and assessment of additional therapeutics associated with the antivenom.

Failure of dihydroartemisinin plus piperaquine treatment of falciparum malaria by under-dosing in an overweight patient.

BACKGROUND: Artemisinin-based combination therapy (ACT) introduced in the mid-1990s has been recommended since 2005 by the World Health Organization as first-line treatment against Plasmodium falciparum in all endemic countries. In 2010, the combination dihydroartemisinin-piperaquine (DP) was recommended for the treatment of uncomplicated P. falciparum malaria. DP is one of the first-line treatments used by the French army since 2013. CASE PRESENTATION: A case of P. falciparum clinical failure with DP at day 20 was described in a 104 kg French soldier deployed in Djibouti. He was admitted to hospital for supervision of oral treatment with DP [40 mg dihydroartemisinin (DHA) plus 320 mg piperaquine tetraphosphate (PPQ)]. This corresponded to a cumulative dose of 4.6 mg/kg DHA and 37 mg/kg PPQ in the present patient, which is far below the WHO recommended ranges. No mutation was found in the propeller domain of the Kelch 13 (k13) gene, which is associated with artemisinin resistance in Southeast Asia. Pfmdr1 N86, 184F, S1034 and N1042 polymorphisms and haplotype 72-76 CVIET for the pfcrt gene were found in the present case. There was no evidence of resistance to DP. CONCLUSION: This case confirms the risk of therapeutic failure with dihydroartemisinin-piperaquine by under-dosing in patients weighing more than 100 kg. This therapeutic failure with DP by under-dosing highlighted the importance of appropriate dosing guidelines and the need of research data (efficacy, pharmacokinetics and pharmacodynamics) in over-weight patient group.

Clinical Survey of Dengue Virus Circulation in the Republic of Djibouti between 2011 and 2014 Identifies Serotype 3 Epidemic and Recommends Clinical Diagnosis Guidelines for Resource Limited Settings.

Dengue virus is endemic globally, throughout tropical and sub-tropical regions. While the number of epidemics due to the four DENV serotypes is pronounced in East Africa, the total number of cases reported in Africa (16 million infections) remained at low levels compared to Asia (70 million infections). The French Armed forces Health Service provides epidemiological surveillance support in the Republic of Djibouti through the Bouffard Military hospital. Between 2011 and 2014, clinical and biological data of suspected dengue syndromes were collected at the Bouffard Military hospital and analyzed to improve Dengue clinical diagnosis and evaluate its circulation in East Africa. Examining samples from patients that presented one or more Dengue-like symptoms the study evidenced 128 Dengue cases among 354 suspected cases (36.2% of the non-malarial Dengue-like syndromes). It also demonstrated the circulation of serotypes 1 and 2 and reports the first epidemic of serotype 3 infections in Djibouti which was found in all of the hospitalized patients in this study. Based on these results we have determined that screening for Malaria and the presence of the arthralgia, gastro-intestinal symptoms and lymphopenia < 1,000cell/ mm3 allows for negative predictive value and specificity of diagnosis in isolated areas superior to 80% up to day 6. This study also provides evidence for an epidemic of Dengue virus serotype 3 previously not detected in Djibouti.

Concomitant human infections with 2 cowpox virus strains in related cases, France, 2011.

We investigated 4 related human cases of cowpox virus infection reported in France during 2011. Three patients were infected by the same strain, probably transmitted by imported pet rats, and the fourth patient was infected by another strain. The 2 strains were genetically related to viruses previously isolated from humans with cowpox infection in Europe.

The French Military influenza surveillance system (MISS): overview of epidemiological and virological results during four influenza seasons--2008-2012.

QUESTION UNDER STUDY: Influenza is a viral infection caused by a pathogen with considerable ability for genetic mutation, which is responsible for seasonal outbreaks as well as pandemics. This article presents the results of epidemiological and virological monitoring of four successive influenza outbreaks in the French armed forces, for the period 2008 to 2012. METHODS: The main events monitored were acute respiratory infection (ARI). Weekly incidence rates were calculated by relating cases to the number of servicepersons monitored. RESULTS: In continental France, the incidence rates for ARI and for medical consultation attributable to influenza were highest during the pandemic and decreased to reach their lowest values in 2010­2011 and 2011­2012. In terms of virological results, the 2008­2009 outbreak was mainly due to the A(H3N2) virus, while the 2009­2010 pandemic and the following season saw the emergence of the A(H1N1) pdm09 strain. The last season 2011­2012 was characterised by a predominant circulation of A(H3N2) viruses. CONCLUSIONS: Despite some limitations, the MISS represents a good source of information about influenza in young people. Virological results are compatible with those reported by most other influenza surveillance networks, but could be improved by a better knowledge of the other respiratory viruses in circulation in the military community.

[Microbiology method validation: illustration with Bact'Alert 3D]. / Vérification sur site en microbiologie: exemple du Bact'Alert 3D (Biomérieux(©)).

As far as laboratories accreditation according to ISO 15189 is concerned, validation of each method used has to be achieved. Manufacturer's recommendations are not always helpful in this context. That's why each laboratory must proceed systematically to a logical risk analysis. Few publications are available on this subject concerning microbiology. We propose to illustrate it with the example of a blood culture automate: the Bact'Alert 3D commercialised by Biomérieux(®).

[In-house PCR practical knowledge: a sufficient validation according to NF EN ISO 15189?]. / Bilan d'utilisation d'une PCR « maison ¼ qualitative : une validation acceptable en regard de la norme NF EN ISO 15189 ?

As far as molecular biology is concerned, numerous are laboratories using in-house method. Used since many years, medical biologists have often a good practical knowledge about them. According to ISO 15189 accreditation, is this knowledge sufficient for the validation of these methods? We have asked this question ourselves about our in-house Leishmaniasis PCR.

[Internal quality control for antimicrobial susceptibility test: an experience feedback]. / Controle de qualite interne en antibiologie: retour d'experience.

Internal quality control (IQC) position in antimicrobial susceptibility testing must be evaluated attentively before using. Ours S. aureus ATCC 25923 use during year 2011 has given precious information that can be useful for other laboratories. First, IQC should never be used before checking that all the realisation process steps are controlled. It will then appear that reagents are the most susceptible to give false results. That's what happens in 2.74% of antimicrobial susceptibility test. IQC is then useful to limit their clinical's impact. However, IQC use also shows that quality improvement will be difficult without industrial producer's involvement.

[Automatic platelets numbering with citrate as anticoagulant: is the result valid?]. / Numération automatique des plaquettes sur citrate de sodium : le résultat est-il exact ?

Platelets count is usually realised on EDTA anticoagulant. This one is sometimes able to generate platelets agregats. That is the reason why the first thing to do encountering thrombopenia is to check for agregats on blood thin smear. In case of positive result, a control can be asked using another anticoagulant. The most used is sodium citrate. A correction has to be applied to the automat result because blood is diluted in anticoagulant. But no one says those haematological automats are exact on citrate as they are on EDTA. That's what we wanted to check.

[Apport de l'immunocapture-Elisa au diagnostic de l'infection à virus grippal A(H1N1)pdm09 : une alternative à la RT-PCR ?] / Apport de l'immunocapture-Elisa au diagnostic de l'infection à virus grippal A(H1N1)pdm09 : une alternative à la RT-PCR ?

AIM OF THE STUDY: To evaluate immunocapture-Elisa (ICE) sensitivity versus RT-PCR in diagnosis of influenza A(H1N1)pdm09 virus infection. METHODS: Sixty-seven RT-PCR-negative and 282 RT-PCR-positive nasopharyngeal swabs collected during winter 2009-2010 have been analyzed using ICE. RESULTS: Among all the samples tested, a sensitivity of 31.3% was found for ICE. The sensitivity of ICE was directly correlated to the virus load determined through the number of cycling reactions necessary to reach detection by RT-PCR. CONCLUSION: ICE can be a suitable method compared to RT-PCR when RT-PCR cannot be used for economical or epidemiological reasons. Its sensitivity is largely dependent of the nasopharyngeal sampling quality.

Splenic infarction during Plasmodium ovale acute malaria: first case reported.

The splenic complications of acute malaria include two different prognostic and treatment entities: splenic infarction and splenic rupture. This is the first case of splenic infarction during an acute malaria due to Plasmodium ovale in a 34-year-old man. As in the majority other described cases of splenic infarction, the course was spontaneously favourable, suggesting that this complication was relatively benign compared to splenic rupture, which is life-threatening and usually necessitating surgery.