Validation of plasma amino acid profile using UHPLC-mass spectrometer (QDa) as a screening method in a metabolic disorders reference centre: Performance and accreditation concerns.

INTRODUCTION: Amino acid (AA) analysis in plasma is essential for diagnosis and monitoring of inborn errors of metabolism (IEM). The efficacy of patient management is governed by the rapidity of AA profile availability, along with the robustness of the method. French quality guidelines and progress made in analytical techniques have led biologists to develop AA profile exploration via mass spectrometry (MS). OBJECTIVES: The aim of this study was to validate an analytical method with a single quadrupole mass spectrometer (MS) and to suggest reference values in regard to French quality and IEM society recommendations. DESIGN AND METHODS: Plasma samples from patients were deproteinised and derivatised with AccqTag™ reagent. Analysis was performed by reverse-phase chromatography coupled to QDA detector. We evaluated accuracy, intra-days and inter-days precision and limit of quantification by the ß-expectation tolerance interval method for 27 AA. Method comparison was performed with the standard method (ion exchange chromatography, IEC) on Jeol Aminotac® and to tandem MS. Reference values were established on AA concentrations of the cohort of patients who had no IEM. RESULTS: Our method allowed the separations of almost all amino acids with a total run time of 12 min. Separation of isoleucine and alloisoleucine was incomplete (R = 0.55) but without impact on biological interpretation. Precision, accuracy and quantification were satisfactory (intra-days coefficient of variation (CV) was <5%, inter-days precision CV <10% and accuracy <15%). The limits of quantification were validated between 1 and 900 µmol/L. Results were comparable between the new method and IEC. CONCLUSION: Ultimately, we validated a rapid method on plasma for quantifying 27 amino acids that can be used in routine practice, according to French quality laboratories and SFEIM (French Society of Inborn Error of Metabolism) recommendations. Furthermore, estimated reference values were similar to those found in published studies focusing on other methods. Despite a lower specificity compared to tandem MS, the simplicity and rapidity of our method are the main advantage of this technique and place it as a major tool in IEM diagnosis and management.

External validation of Gout-calculator performance on a cohort of acute arthritis (SYNOLACTATE) sparing distal joints such as hallux and midfoot. A cross-sectional study of 170 patients.

OBJECTIVE: To evaluate the performance of the Gout-calculator in a cohort of consecutive acute arthritis affecting large and intermediate joints (without an attack on hallux or midfoot joints). METHODS: A retrospective study. Gout-calculator data were collected in medical records of patients included in the prospective consecutive cohort of acute arthritis called SYNOLACTATE. The diagnosis of gout was defined by the presence of sodium urate crystals in synovial fluid. The diagnostic performance of the Gout-calculator was studied by performing an ROC curve with the calculation of its AUC (95% CI) as well as the calculation of Sensitivity (Se), Specificity (Sp), and positive likelihood ratio (LR+). RESULTS: 170 patients with acute arthritis were included. Variables associated with the diagnosis of gout were as follows: serum uric acid > 350 µmol/L (OR 5.52 (2.52-12.1), p < 0.001), joint redness (OR 5.08 (1.85-14.0), p = 0.001), previous patient-reported arthritis attack (OR 4.04 (1.92-8.49), p < 0.001), male (OR 3.00 (1.17-7.69), p = 0.02), hypertension or cardiovascular disease (OR 2.33 (1.07-5.06), p = 0.03). The median (IQR) of Gout-calculator was significantly higher in gouty arthritis (7.0 [5.5-8.1]) than in associated-CPP acute arthritis (4.0 [2.0-5.8]), septic arthritis (3.0 [2.0-5.1]), or others arthritis (3.5 [2.0-5.5]). The AUC was 0.833 (0.768-0.897) with for the threshold ≥ 8, a Se at 27.5% (0.161-0.428), Sp 97.7% (0.934-0.992), and LR+ 11.9 (3.5-40). CONCLUSION: Despite diagnostic performances close to those published, the use of the Gout-calculator is not sufficient for the diagnosis of gout or to exclude the differential diagnosis of septic arthritis in the SYNOLACTATE cohort. KEY POINTS: • For a Gout-calculator threshold of ≤ 4, Sensitivity is 92.5%, Specificity 50.8% and LR- 0.15 to the gout diagnosis. • For a Gout-calculator threshold of > = 8, Sensitivity is 27.5%, Specificity 97.7% and LR+ 11.9 to the gout diagnosis. • In a population of acute arthritis affecting large joints, Gout-calculator is not sufficient to discriminate between gouty arthritis and septic arthritis.

Maternal and neonatal outcomes and prognostic factors in acute fatty liver of pregnancy.

OBJECTIVE: To report complications of Acute Fatty Liver of pregnancy (AFLP), a rare liver disease of pregnancy, and identify prognostic factors for mothers and children. STUDY DESIGN: We conducted a retrospective descriptive study over 18 years in three French maternities. Demographic, clinical, biological data, and outcomes of patients and their infants were reviewed. RESULTS: 142,450 pregnancies from centers were studied. Eighteen patients with AFLP were identified The prevalence of AFLP was estimated as 1/7,914 pregnancies. Prolonged prothrombin time was identified as a risk factor of maternal complications (OR = 0.86, p = 0.0493). Gestational age at delivery was the only risk factor associated with fetal or neonate complications (OR = 0.37, p = 0.0417). One boy died of previously undiagnosed ß-oxidation deficiency at eight months. CONCLUSION: In AFLP, prothrombin time must be carefully monitored to anticipate major maternal complications. Infants born to mothers with ALFP should be screened as early as possible for mitochondrial fatty acid oxidation deficiency.

Doubling immunochemistry laboratory testing efficiency with the cobas e 801 module while maintaining consistency in analytical performance.

OBJECTIVES: The new immunochemistry cobas e 801 module (Roche Diagnostics) was developed to meet increasing demands on routine laboratories to further improve testing efficiency, while maintaining high quality and reliable data. DESIGN AND METHODS: During a non-interventional multicenter evaluation study, the overall performance, functionality and reliability of the new module was investigated under routine-like conditions. It was tested as a dedicated immunochemistry system at four sites and as a consolidator combined with clinical chemistry at three sites. RESULTS: We report on testing efficiency and analytical performance of the new module. Evaluation of sample workloads with site-specific routine request patterns demonstrated increased speed and almost doubled throughput (maximal 300 tests per h), thus revealing that one cobas e 801 module can replace two cobas e 602 modules while saving up to 44% floor space. Result stability was demonstrated by QC analysis per assay throughout the study. Precision testing over 21 days yielded excellent results within and between labs, and, method comparison performed versus the cobas e 602 module routine results showed high consistency of results for all assays under study. In a practicability assessment related to performance and handling, 99% of graded features met (44%) or even exceeded (55%) laboratory expectations, with enhanced reagent management and loading during operation being highlighted. CONCLUSION: By nearly doubling immunochemistry testing efficiency on the same footprint as a cobas e 602 module, the new module has a great potential to further consolidate and enhance laboratory testing while maintaining high quality analytical performance with Roche platforms.

Iron excess upregulates SPNS2 mRNA levels but reduces sphingosine-1-phosphate export in human osteoblastic MG-63 cells.

We aimed to study the mechanisms involved in bone-related iron impairment by using the osteoblast-like MG-63 cell line. Our results indicate that iron impact the S1P/S1PR signalizing axis and suggest that iron can affect the S1P process and favor the occurrence of osteoporosis during chronic iron overload. INTRODUCTION: Systemic iron excess favors the development of osteoporosis, especially during genetic hemochromatosis. The cellular mechanisms involved are still unclear despite numerous data supporting a direct effect of iron on bone biology. Therefore, the aim of this study was to characterize mechanisms involved in the iron-related osteoblast impairment. METHODS: We studied, by using the MG-63 cell lines, the effect of iron excess on SPNS2 gene expression which was previously identified by us as potentially iron-regulated. Cell-type specificity was investigated with hepatoma HepG2 and enterocyte-like Caco-2 cell lines as well as in iron-overloaded mouse liver. The SPNS2-associated function was also investigated in MG-63 cells by fluxomic strategy which led us to determinate the S1P efflux in iron excess condition. RESULTS: We showed in MG-63 cells that iron exposure strongly increased the mRNA level of the SPNS2 gene. This was not observed in HepG2, in Caco-2 cells, and in mouse livers. Fluxomic study performed concomitantly on MG-63 cells revealed an unexpected decrease in the cellular capacity to export S1P. Iron excess did not modulate SPHK1, SPHK2, SGPL1, or SGPP1 gene expression, but decreased COL1A1 and S1PR1 mRNA levels, suggesting a functional implication of low extracellular S1P concentration on the S1P/S1PR signalizing axis. CONCLUSIONS: Our results indicate that iron impacts the S1P/S1PR signalizing axis in the MG-63 cell line and suggest that iron can affect the bone-associated S1P pathway and favor the occurrence of osteoporosis during chronic iron overload.

Prospective evaluation of serum anti-Müllerian hormone dynamics in 250 women of reproductive age treated with chemotherapy for breast cancer.

AIM: Women of reproductive age with breast cancer generally receive gonadotoxic chemotherapy. Fertility issues are of great concern for them. However, little is known on ovarian damage during chemotherapy and its evolution during long-term follow-up. The aim of this study was to provide a detailed description of serum anti-Müllerian hormone (AMH) evolution during chemotherapy and 24-month follow-up. METHODS: This prospective cohort study was conducted in 250 patients, aged 18-39 years, diagnosed with breast cancer and treated with adjuvant/neoadjuvant chemotherapy. Each patient underwent blood AMH measurement at each chemotherapy cycle, and at 6, 12 and 24 months after chemotherapy. Menses occurrence was also recorded. RESULTS: Mean basal AMH level was 4.19 ± 4.84 ng/mL, and was negatively correlated with age. Serum AMH level rapidly decreased in all patients after each chemotherapy cycle to undetectable levels in most of them, and slowly increased in 45% of the patients during the 24-month follow-up. AMH decrease was significantly associated with age and basal AMH level, but not with cyclophosphamide dose and tamoxifen use. The prevalence of chemotherapy-related amenorrhoea was 92.4% at the end of chemotherapy; women with amenorrhoea being significantly older and having lower basal AMH than women who resumed menses. CONCLUSIONS: Our study confirms rapid and deep ovarian reserve alteration in young women receiving chemotherapy for breast cancer, and shows moderate AMH recovery in some patients. Although AMH cannot alone predict fertility potential, these new data emphasise the need for post-treatment ovarian insufficiency follow-up, strongly support the use of fertility preservation strategies and may provide new tools for improved counselling.

[Screening for carriage of Staphylococcus aureus prior to mammary reconstruction in the prevention of surgical site infection]. / En reconstruction mammaire: intérêt du dépistage du portage de Staphylococcus aureus dans la prévention de l'infection du site opératoire.

The incidence of prosthesis infections after breast reconstruction is of the order of 4% to 13% according to the literature. In surgical patients, Staphylococcus aureus (S. aureus) is the bacterial species most often responsible for surgical site infections. In cardiac surgery, screening for carriage of S. aureus and preoperative decontamination are carried out routinely before prosthetic surgery. We retrospectively reviewed data from patients at our institution between January 2011 and December 2013. Our series showed that the prosthesis infection rates were in the range of 5.92% in 2008 with an ISO rate of S. aureus 3.61%. Routine screening for prosthetic reconstructions was performed to assess the impact of preoperative decontamination patients in carriers of S. aureus. This screening was done in 381 patients: 17.8% of patients were carriers of S. aureus ; 11 patients have an ISO (or an incidence rate of 2.88%) ; 5 patients have an ISO S. aureus (an incidence of S. aureus ISO 1.3%). The introduction of the screening process, allowed a drop of 5.92% ISO rate at 1.46% with a passage of S. aureus SSI rates of 3, 60% to 0.72%. In the near future, studies are needed to confirm these encouraging results, to demonstrate the efficacy of preoperative decontamination in carriers of S. aureus patients before laying prosthesis.

Heavy/light chain specific immunoglobulin ratios provides no additional information than serum proteins electrophoresis and immunofixation for the diagnosis and the follow-up of intact immunoglobulin multiple myeloma patients.

BACKGROUND: Serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE) are used for diagnosis and follow-up of patients with intact immunoglobulin multiple myeloma. However, the numerous limitations of these methods led to the development of a nephelometric immunoassay (Hevylite™) for the specific measurement of serum IgGκ, IgGλ, IgAκ and IgAλ concentrations. METHODS: In this study, we evaluated the correlation between this assay and SPE and IFE in 114 sera of 15 patients (12 IgG and 3 IgA patients) and its impact on the clinical care of patients, especially for diagnosis, for the evaluation of residual disease and for early detection of relapse. RESULTS: At inclusion and during follow-up, we found a good correlation between monoclonal immunoglobulin concentrations and SPE (R(2)=0.902 for IgA and R(2)=0.915 for IgG) and nephelometric quantification (R(2)=0.948 for IgA and R(2)=0.920 for IgG) for the evaluation of monoclonal and polyclonal immunoglobulins. Our results illustrate that the Hevylite™ test is less sensitive than the IFE for detection of residual disease: 5 patients who obtained very good partial response or complete response had normalization of the Hevylite™ ratio while IFE was still positive. A relapse had been detectable with the Hevylite™ ratio 1 to 2 months earlier than with SPE and IFE in 3 patients out of 15, but no recommendations for treating patients with only slight biological relapse are available. CONCLUSION: Our results demonstrate that heavy/light chain specific immunoglobulin ratios provides no additional information than serum proteins electrophoresis and immunofixation for the diagnosis and the follow-up of intact immunoglobulin multiple myeloma patients. We also studied the correlation between the concentration of total immunoglobulin measured by Hevylite™ (sum of Ig'κ + Ig'λ) and nephelometric measurement of total IgG or IgA. For this correlation analysis, all 114 sera were analyzed. The correlation coefficient was R(2) = 0.948 for IgA and R(2) = 0.920 for IgG.

Occult cancer in specimens of reduction mammaplasty aimed at symmetrization. A multicentric study of 2718 patients.

Women who have undergone surgical treatment for breast cancer often benefit from a contralateral reduction mammaplasty (CRM) aimed at symmetrization of the contralateral breast unaffected by the initial cancer. In our 7-year multicentric study (12 centers) of 2718 patients, incidence of CRM cancers (CRMc) was 1.47% (n = 40) [95% CI 1.05%-2.00%]. The CRMc group had significantly more initial mammary cancers of invasive lobular carcinoma (ILC, 22.5% vs 12.0%) and ductal carcinoma in situ (DCIS, 35.0% vs 21.6%) types than the healthy CRM group (p = 0.017). 35.0% (n = 14) of patients had en bloc resection; 25.0% (n = 10) of surgical specimens were correctly oriented. En bloc resection and orientation of surgical specimens enable precise pinpointing of the CRMc. A salvage lumpectomy may be proposed as an option when margins are invaded. The histological distribution of the 40 CRMc (mean size 12.7 mm) was carcinoma in situ (CIS) 70%, ILC 12.5%, invasive ductal carcinoma (IDC) 12.5% and tubular carcinoma (TC) 5.0%.

[The role of urinary markers in the assessment and follow-up of lower urinary tract disorders: a literature review]. / Place des biomarqueurs urinaires dans le diagnostic et le suivi des troubles du bas appareil urinaire: une revue de la littérature.

OBJECTIVES: To conduct a literature review on the role of urinary biomarkers in the initial assessment and follow-up of lower urinary tract symptoms. METHODS: A literature review was conducted in August 2014 using the Medline/Pubmed database limiting the search to work in English or French. RESULTS: Most studies were of level of evidence 2 or 3 (prospective cohort, controlled or not) and mainly about overactive bladder and bladder pain syndrome. Nerve Growth Factor (NGF) was the most studied and apparently the most promising in the evaluation of overactive bladder (OAB) and neurogenic detrusor overactivity (NDO). Urinary levels of ATP, prostaglandin E2 (PGE2), Brain-Derived Neurotrophic Factor (BDNF) and some cytokines were also significantly higher in most studies in patients with NDO or OAB. Epidermal Growth Factor (EGF), Heparin-Binding EGF (HBEGF) and Antiproliferative Factor (APF) were the most studied urinary markers in bladder pain syndrome, with a significant increase (EGF APF) or decrease (HBEGF) in cases of interstitial cystitis (compared to healthy controls). The urinary N-terminal-telopeptide (NTx) could be predictive of a failed mid-urethral sling. However, few studies reported the diagnostic values of the markers, their association with urodynamic parameters were rarely evaluated and the existence of a publication bias is likely. No randomized controlled study has so far compared the urinary markers to urodynamic evaluation. CONCLUSION: In the future, urinary markers could complete or replace urodynamic examination. However, to date, there is no high level of evidence study comparing these markers to urodynamics and their use can therefore not be recommended in daily practice.

Identification of Nine New RAI1-Truncating Mutations in Smith-Magenis Syndrome Patients without 17p11.2 Deletions.

Smith-Magenis syndrome (SMS) is an intellectual disability syndrome with sleep disturbance, self-injurious behaviors and dysmorphic features. It is estimated to occur in 1/25,000 births, and in 90% of cases it is associated with interstitial deletions of chromosome 17p11.2. RAI1 (retinoic acid induced 1; OMIM 607642) mutations are the second most frequent molecular etiology, with this gene being located in the SMS locus at 17p11.2. Here, we report 9 new RAI1-truncating mutations in nonrelated individuals referred for molecular analysis due to a possible SMS diagnosis. None of these patients carried a 17p11.2 deletion. The 9 mutations include 2 nonsense mutations and 7 heterozygous frameshift mutations leading to protein truncation. All mutations map in exon 3 of RAI1 which codes for more than 98% of the protein. RAI1 regulates gene transcription, and its targets are themselves involved in transcriptional regulation, cell growth and cell cycle regulation, bone and skeletal development, lipid and glucide metabolisms, neurological development, behavioral functions, and circadian activity. We report the clinical features of the patients carrying these deleterious mutations in comparison with those of patients carrying 17p11.2 deletions.

Array-CGH Analysis Suggests Genetic Heterogeneity in Rhombencephalosynapsis.

Rhombencephalosynapsis is an uncommon, but increasingly recognized, cerebellar malformation defined as vermian agenesis with fusion of the hemispheres. The embryologic and genetic mechanisms involved are still unknown, and to date, no animal models are available. In the present study, we used Agilent oligonucleotide arrays in a large series of 57 affected patients to detect candidate genes. Four different unbalanced rearrangements were detected: a 16p11.2 deletion, a 14q12q21.2 deletion, an unbalanced translocation t(2p;10q), and a 16p13.11 microdeletion containing 2 candidate genes. These genes were further investigated by sequencing and in situ hybridization. This first microarray screening of a rhombencephalosynapsis series suggests that there may be heterogeneous genetic causes.

Pure tubular carcinoma of the breast and sentinel lymph node biopsy: a retrospective multi-institutional study of 234 cases.

BACKGROUND: Pure Tubular Carcinoma (PTC) of the breast is a rare histological subtype of invasive breast cancer characterized by a low rate of lymph node involvement. Currently there is no consensus on less surgical axillary node staging according to this histological subtype. METHODS: We performed a retrospective multi-institutional study. Inclusion criteria were PTC, sentinel lymph node detection (SLND) and conservative breast surgery. RESULTS: From January 1999 to December 2006, 234 patients were included in the study from 9 institutions. The median pathological tumor size was 9.59 (1-22) mm. SLN were successfully detected in 98% (229/234) of patients. Among the 234 patients, a macrometastasis was found in 6 cases (2.5%), micrometastasis in 15 cases (6.4%), and isolated cells in 2 cases (0.8%). In the case of patients with SLND macrometastasis, half of them had macrometastasis in the complementary axillary lymphadenectomy, and none in the case of SLN only micrometastasis or isolated cells. Of the 122 patients with a pathological tumor size <10 mm, none had sentinel node macrometastasis. According to a multivariate analysis, pathological tumor size (>10 mm) was the only parameter significatively linked to the risk of lymph node involvement (p = 0.007). CONCLUSION: In a large multi-institutional series with SLND, we have shown that the risk of axillary lymph node involvement in PTC is very low. In the case of PTC <10 mm, we suggest that surgical axillary evaluation, even with SLND, may not be warranted.

[MRI evaluation of residual breast carcinoma after neoadjuvant chemotherapy]. / Cancer du sein traité par chimiothérapie néoadjuvante: évaluation du reliquat tumoral par l'IRM mammaire.

PURPOSE: This study aims to evaluate the sensibility and specificity of MRI in the detection and size measuring of residual breast cancer in patients treated with neoadjuvant chemotherapy before surgery. PATIENTS AND METHODS: This is a retrospective study of 32 women, who underwent breast MRI before and after neoadjuvant treatment. MRI has been confronted to surgical pathology results. RESULTS: The sensibility of MRI to assess pathologic Complete Response (no invasive residual tumor) was excellent (100%) but the specificity was low (55,5%). There was no false negative case and four false positive cases (Two ductal carcinomas in situ and two scars-like fibrosis). When MRI outcomes were compared with the presence or absence of invasive or in situ residual carcinoma, only one false negative case was noticed (one "in situ" residual tumor). The correlation between tumor size measured by MRI and histopathology was low (r=0,32). Underestimations of tumor size were due to non-continuous tumor regression or invasive lobular carcinoma or association of invasive carcinoma and intra ductal breast cancer. Over estimations of tumor size were due to chemotherapy-induced changes. CONCLUSION: MRI is a sensitive but poorly specific method to assess the pathological complete response after neoadjuvant chemotherapy. Estimation of tumor size and detection of isolated residual in situ carcinoma are fare. Therefore, surgical intervention remains necessary whatever the MRI outcomes.

[Atypical epithelial hyperplasia of the breast: current state of knowledge and clinical practice]. / Hyperplasie épithéliale atypique du sein : bilan des connaissances et pratique clinique.

INTRODUCTION: The diagnosis of atypical epithelial hyperplasia (AEH) increases with breast cancer screening. AEH is divided in three groups: atypical ductal hyperplasia, columnar cell lesions with atypia, lobular neoplasia. The management of women with AEH is not consensual because of uncertainty about their diagnosis related to the type of the biopsy sampling (core needle biopsy or surgical excision) and their controversial clinical signification between risk marker and true precursor of breast cancer. MATERIAL AND METHODS: A systematic review of published studies was performed. Medline baseline interrogation was performed with the following keywords: atypical ductal hyperplasia, columnar cell lesions with atypia, lobular neoplasia, core needle biopsy, breast cancer, precursor lesion, hormonal replacement therapy. For each breast lesion, identified publications (English or French) were assessed for clinical practise in epidemiology, diagnosis and patient management. RESULTS: With immunohistochemistry and molecular studies, AEH seems to be precursor of breast cancer. But, epidemiological studies show low rate of breast cancer in women with AEH. AEH were still classified as risk factor of breast cancer. CONCLUSION: Because of high rate of breast cancer underestimation, surgical excision is necessary after the diagnosis of AEH at core needle biopsy. Surgical oncology rules and collaboration with radiologist are required for this surgery. A second operation was not required due to involved margins by AEH (except with pleiomorphic lobular neoplasia) because local control of breast cancer seems to be unchanged. Besides, hormonal replacement therapy for patient with AEH is not recommended because of lack of studies about this subject.

[Breast cancer prevention: from chemoprevention to prophylactic surgery]. / Prévention des cancers du sein : de la chimioprévention à la chirurgie prophylactique.

INTRODUCTION: The breast cancer prevention is based on mastectomy hormonal deprivation (surgical or chemical) and the use of drugs acting on cell signalization pathways, which provoke the cancerization (these drugs are not officially authorized in France). MATERIAL AND METHODS: Analysis of the literature selected from the Medline base on the keywords: breast cancer; chemoprevention; prophylactic surgery; tamoxifene; raloxifene; BRCA. RESULTS: Four trials on the chemoprevention by tamoxifene show a reduction in the breast cancer incidence from 22% up to 33% in the treated patients, limited to oestrogen-dependant cancers, especially in the populations at risk high (histological or genetic) even in the event of concomitant hormonal replacement therapy. The benefit seems continue in time. Raloxifene and tamoxifene effects are comparable with bone benefits and a less risk of endometrial cancer for raloxifene, but the risk of venous thrombosis is still persisting. The breast prophylactic surgery is effective mainly in case of genetic elevated risk when it is practiced in the young age, and requires a patient agreement (the decision needs to follow the patient advice after complete information). The prophylactic ovariectomy has a positive impact on the mammal risk even in the high genetic risk women. CONCLUSION: The breast cancer prevention requires a better selection of the patients, an adaptation of the type of prevention taking account of the balance risks and benefits (mammals and extramammals) before a clinical use in routine.

Multicolour FISH and quantitative PCR can detect submicroscopic deletions in holoprosencephaly patients with a normal karyotype.

Holoprosencephaly (HPE) is the most common structural malformation of the developing forebrain. At birth, nearly 50% of children with HPE have cytogenetic anomalies. Approximately 20% of infants with normal chromosomes have sequence mutations in one of the four main HPE genes (SHH, ZIC2, SIX3, and TGIF). The other non-syndromic forms of HPE may be due to environmental factors or mutations in other genes, or potentially due to submicroscopic deletions of HPE genes. We used two complementary assays to test for HPE associated submicroscopic deletions. Firstly, we developed a multicolour fluorescent in situ hybridisation (FISH) assay using probes for the four major HPE genes and for two candidate genes (DISP1 and FOXA2). We analysed lymphoblastoid cell lines (LCL) from 103 patients who had CNS findings of HPE, normal karyotypes, and no point mutations, and found seven microdeletions. We subsequently applied quantitative PCR to 424 HPE DNA samples, including the 103 samples studied by FISH: 339 with CNS findings of HPE, and 85 with normal CNS and characteristic HPE facial findings. Microdeletions for either SHH, ZIC2, SIX3, or TGIF were found in 16 of the 339 severe HPE cases (that is, with CNS findings; 4.7%). In contrast, no microdeletion was found in the 85 patients at the mildest end of the HPE spectrum. Based on our data, microdeletion testing should be considered as part of an evaluation of holoprosencephaly, especially in severe HPE cases.

[Chloroquine sensitivity of Plasmodium falciparum at the Gamkalley Clinic and the Nigerian armed forces PMI (Niamey, Niger)]. / Chloroquinosensibilité de Plasmodium falciparum à la clinique Gamkalley et à la PMI des Forces armées nigériennes (Niamey, Niger).

In vivo tests for Plasmodium falciparum were carried out in 1998 during the rainy season among children in Niamey, in the Republic of Niger. Chloroquine was prescribed at 25 mg/kg for 3 days in febrile patients with uncomplicated P. falciparum malaria. Forty-five 1-5 year-olds and thirty-three 6-15 year-olds were included in the study. A group of 53 adult patients was also surveyed to evaluate the efficacy of chloroquine in semi-immune persons. Body temperature and blood smears including parasitemia were recorded on days 0, 3, 7 and 14. Less than 10% of the patients were delinquent. Around 75% of the patients were successfully treated in the 1-5 year-olds and 6-15 year old age groups. Relapses were observed in 20% of the 1-5 year-olds (early relapses 8.9%, late relapses 11.1%) and in 16.7% in the 1-15 year-olds (early relapses 6.4%, late relapses 10.3%). Among adults, successful treatment was obtained in 86.8% of the cases and early and late relapses were respectively observed in 3.8% and 1.9% of the cases. All the patients with malaria relapses were cured with second-line treatments (pyrimethamine-sulfadoxine or quinine). According to these results, chloroquine resistance appears to be moderate in Niamey. Therefore chloroquine should remain the first line treatment of uncomplicated P. falciparum malaria in this population.