First Molecular Identification and Genetic Characterization of Theileria lestoquardi in Sheep of the Maghreb Region

Theileria lestoquardi is the most prominent Theileria species in small ruminants that causes malignant theileriosis of sheep in Africa and Asia. In the present survey, blood samples and ticks were collected in Kebili (southern Tunisia) from 166 Queue Fine de l'Ouest sheep. Giemsa‐stained blood smears, immunofluorescent antibody test (IFAT) and PCR were performed. The DNA was extracted from blood and analysed by PCR targeting 18S rRNA gene of Theileria spp. and then sequenced. A total number of 140 ticks were collected from a total number of 166 sheep during the four seasons. The ticks belonged to two genera and 4 species; the most frequent tick was Hyalomma excavatum 84.3% (118/140) and then Rhipicephalus spp. 15.7% (22/140). Only two animals had positive Giemsa‐stained blood smears, and they were also positive by IFAT. The amplicons had 99.3 and 99.6% homology with the BLAST published T. lestoquardi amplicons. To our knowledge, this is the first report of T. lestoquardi in small ruminants within the Maghreb region.     

Prévalence d’infection des ovins par Toxoplasma gondii en Tunisie

The aim of this study is to estimate the prevalence of Toxoplasma gondii infection in 527 sheep from 4 governorates of Tunisia by Elisa (350 animals) and PCR (177 animals). The seroprevalence in sheep was estimated to be 1.8% (N = 166) in the governorate of Siliana (North Tunisia) and 19% (N = 184) in the governorate of Kasserine (Central Tunisia) with a commercial Elisa kit. T. gondii DNA was extracted from the apex of the heart in 25.5% (N = 106) of sheep from the Sidi-Bouzid governorate (Central Tunisia) and 12.7% (N = 71) from the Ben-Arous governorate (North Tunisia). There was no statistically significant difference between different age categories’ prevalence within each locality. Our results indicate that T. gondii infection is frequent in Tunisian sheep. The implementation of a national control programme against toxoplasmosis should not neglect sheep as a frequently infected intermediate host.     

Long-term follow-up of children with risk organ-negative Langerhans cell histiocytosis after 2-chlorodeoxyadenosine treatment

The nucleoside analogue, 2-chlorodeoxyadenosine (2CDA), was reported to be an active treatment for childhood Langerhans cell histiocytosis (LCH) without risk organ (RO−) involvement. However, we lack data on long-term effects of 2CDA treatment, including the disease reactivation rate, permanent sequelae and long-term tolerance. This study included 44 children from the French LCH registry, treated for a RO− LCH with 2CDA monotherapy (median number of six courses). The median age at the beginning of 2CDA was 3·6 years (range, 0·3–19·7 years) and the median follow-up after was 5·4 years (range, 0·6–15·1 years). Objective response to 2CDA was observed in 25 patients (56·8%), while six patients (13·6%) had stable disease and 13 patients (29·5%) exhibited progressive disease. Among patients without progression, only two experienced disease reactivation after 2CDA discontinuation. The five-year cumulative incidence of disease progression or reactivation after 2CDA therapy initiation was 34·3%. The lymphopenia reported in all cases [72% below absolute lymphocyte count (ALC) of 0·5 G/l], was addressed with appropriate prophylactic measures. Other toxicities above grade 2 were uncommon, and no second malignant neoplasm or neuropathy was reported. The five-year overall survival was 97·7%. In conclusion, we could confirm that 2CDA monotherapy was a beneficial long-term therapy for treating patients with RO− LCH. Appropriate management of induced immune deficiency is mandatory.     

Concurrent occurrence of a gastric cancer with a hepatocellular carcinoma: report of a case: incidental or causal association?

Little information regarding synchronous gastric cancer associated with hepatocellular carcinoma is available and has been sporadically reported. We report a new case of 60 years old patient operated for gastric carcinoma. The radiological investigations revealed a hepatic nodule which correspond to a hepatocellular carcinoma on histological examination. The aim of this study is to clarify the clinicopathologic and therapeutic features of this association.     

Compound heterozygosity of novel missense mutations in the gamma-glutamyl-carboxylase gene causes hereditary combined vitamin K-dependent coagulation factor deficiency

Hereditary combined vitamin K-dependent (VKD) coagulation factor deficiency is an autosomal recessive bleeding disorder associated with defects in either the gamma-carboxylase, which carboxylates VKD proteins to render them active, or the vitamin K epoxide reductase (VKORC1), which supplies the reduced vitamin K cofactor required for carboxylation. Such deficiencies are rare, and we report the fourth case resulting from mutations in the carboxylase gene, identified in a Tunisian girl who exhibited impaired function in hemostatic VKD factors that was not restored by vitamin K administration. Sequence analysis of the proposita did not identify any mutations in the VKORC1 gene but, remarkably, revealed 3 heterozygous mutations in the carboxylase gene that caused the substitutions Asp31Asn, Trp157Arg, and Thr591Lys. None of these mutations have previously been reported. Family analysis showed that Asp31Asn and Thr591Lys were coallelic and maternally transmitted while Trp157Arg was transmitted by the father, and a genomic screen of 100 healthy individuals ruled out frequent polymorphisms. Mutational analysis indicated wild-type activity for the Asp31Asn carboxylase. In contrast, the respective Trp157Arg and Thr591Lys activities were 8% and 0% that of wild-type carboxylase, and their compound heterozygosity can therefore account for functional VKD factor deficiency. The implications for carboxylase mechanism are discussed.     

Simultaneous LC determination of paracetamol and related compounds in pharmaceutical formulations using a carbon-based column

A simple, rapid and convenient high performance liquid chromatographic method, which permits the simultaneous determination of paracetamol, 4-aminophenol and 4-chloracetanilide in pharmaceutical preparation has been developed. The chromatographic separation was achieved on porous graphitized carbon (PGC) column using an isocratic mixture of 80/20 (v/v) acetonitrile/0.05 M potassium phosphate buffer (pH 5.5) and ultraviolet detection at 244 nm. Correlation coefficient for calibration curves in the ranges 1–50 μg ml⁻¹ for paracetamol and 5–40 μg ml⁻¹ for 4-aminophenol and 4-chloroacetanilide were >0.99. The sensitivity of detection is 0.1 μg ml⁻¹ for paracetamol and 0.5 μg ml⁻¹ for 4-aminophenol and 4-chloroacetanilide. The proposed liquid chromatographic method was successfully applied to the analysis of commercially available paracetamol dosage forms with recoveries of 98–103%. It is suggested that the proposed method should be used for routine quality control and dosage form assay of paracetamol in pharmaceutical preparations. The chromatographic behaviour of the three compounds was examined under variable mobile phase compositions and pH, the results revealed that selectivity was dependent on the organic solvent and pH used. The retention selectivity of these compounds on PGC was compared with those of octadecylsilica (ODS) packing materials in reversed phase liquid chromatography. The ODS column gave little separation for the degradation product (4-aminophenol) from paracetamol, whereas PGC column provides better separation in much shorter time.     

Biological validation of bio-engineered red blood cell productions

The generation in vitro of cultured red blood cells (cRBC) could become an alternative to classical transfusion products. However, even when derived from healthy donors, the cRBC generated in vitro from hematopoietic stem cells may display alterations resulting from a poor controlled production process. In this context, we attempted to monitor the quality of the transfusion products arising from new biotechnologies.For that purpose, we developed an in vitro erythrophagocytosis (EP) test with the murine fibroblast cell line MS-5 and human macrophages (reference method). We evaluated 38 batches of cRBC, at the stage of reticulocyte, generated from CD34⁺ cells isolated from placental blood or by leukapheresis. We showed that (i) the EP test performed with the MS-5 cell line was sensitive and can replace human macrophages for the evaluation of cultured cells. (ii) The EP tests revealed disparities among the batches of cRBC. (iii) The viability of the cells (determined by calcein-AM test), the expression of CD47 (antiphagocytosis receptor) and the externalization of phosphatidylserine (PS, marker of phagocytosis) were not critical parameters for the validation of the cRBC. (iv) Conversely, the cell deformability determined by ektacytometry was inversely correlated with the intensity of the phagocytic index.Assuming that the culture conditions directly influence the quality of the cell products generated, optimization of the production mode could benefit from the erythrophagocytosis test.     

Congenital coronary aneurysm. Three case reports

Congenital coronary aneurysms are an unusual anatomical entity. Their prognosis appears to be particularly dependent on the presence or absence of aneurysm thrombosis. We report three cases of congenital coronary aneurysms, diagnosed in one case after myocardial infarction. Two patients were treated successfully by an exclusion of the aneurysm and coronary bypass and the third patient was treated medically. The aim of this study is to discuss the clinical features, prognosis and management of this disease.     

Simultaneous determination of naproxen and related compounds by HPLC using porous graphitic carbon column

A simple, selective and sensitive high performance liquid chromatographic (HPLC) method has been developed for the simultaneous determination of naproxen and its main degradation products such as 1-(6-methoxy-2-naphthyl) ethanol (MNE), 2-methoxy-6-ethyl naphthalene (MEN) and 2-acetyl-6-methoxy naphthalene (AMN). The separation of these compounds was achieved on porous graphitic carbon (PGC) column using tetrahydrofuran-methanol as the mobile phase, and the effluent from the column was monitored at 272 nm. At a flow rate of 1 ml min(-1), the retention time of the last eluting compound was less than 10 min. Correlation coefficient for calibration curves in the ranges 2-25 microg ml(-1) for all compounds studied were greater than 0.999. The sensitivity of detection is 0.05 microg l(-1) for naproxen, MNE and MEN and 0.20 microg ml(-1) for AMN. The reproducibility of the peak area of these compounds using isocratic elution were quite high, and the standard deviations (S.D.) were below 2% (n=5). The reproducibility of retention times of these compounds was within 1% (n=5). The proposed liquid chromatographic method was successfully applied to the analysis of commercially available naproxen sodium (NS) dosage forms with recoveries of 98.8-102%. A comparative study shows that the selectivity of these compounds on PGC column was different to that obtained with octadecyl silica (ODS) columns.