Juvenile ecthyma gangrenosum caused by Pseudomonas aeruginosa revealing an underlying neutropenia: case report and review of the literature.

BACKGROUND: Ecthyma gangrenosum (EG) is characterized by the occurrence of erythematous, violaceous or haemorrhagic macules and/or vesicles, often evolving into necrotic ulcers, with a central grey-black eschar. It is a rare skin condition, usually occurring in immunocompromised patients suffering from bacterial sepsis caused by Pseudomonas aeruginosa. However, seemingly healthy children have been diagnosed with this skin disease as well. OBJECTIVES: We report the workup of a case of vulvar EG caused by P. aeruginosa in a toddler, which led to a diagnosis of an underlying neutropenia. Moreover, we provide a brief literature review on those cases of EG where an underlying primary immunodeficiency, neutropenia in particular, was eventually diagnosed. METHODS: A one-and-a-half-year-old girl presented with a history of recurrent (respiratory) infections and the sporadic occurrence of purpuric, vulvar ulcers. Workup consisted of microbiological and haematological investigations, including repeated blood analyses. RESULTS: Bacterial swabs from the vulvar ulcers showed the growth of P. aeruginosa. No concomitant sepsis was present, but laboratory investigations pointed towards a cyclic neutropenia, coinciding with the occurrence of the EG lesions. Topical gentamicin ointment allowed the skin lesions to heal faster. Following the administration of granulocyte colony-stimulating factor (G-CSF), the girl experienced less infections in general and had no recurrence of EG lesions in particular. Treatment with G-CSF could eventually be stopped, and the neutropenia, ultimately transient in nature, completely resolved. CONCLUSION: Children presenting with (anogenital) EG should always alert a physician to consider a potentially underlying immunodeficiency, neutropenia in particular.

On the systematics of Phlebotomus betisi and two new related species from Laos with proposal of the new subgenus Lewisius.

Phlebotomus betisi was described from Malaysia and classified after its description in the subgenus Larroussius. It was the only species to have a pharyngeal armature composed of dot-like teeth and an annealed spermatheca whose head is carried by a neck in females. Males were characterized by having a style bearing five spines and a simple paramere. The study of sandflies originating from a cave in Laos enabled us to discover and describe two sympatric species close to Ph. betisi Lewis & Wharton, 1963 and new for Science: Ph. breyi Vongphayloth & Depaquit n. sp., and Ph. sinxayarami Vongphayloth & Depaquit n. sp. They were characterized morphologically, morphometrically, geomorphometrically, molecularly, and proteomically (MALDI-TOF). All approaches converged to validate the individualization of these species whose morphological differential characters lay in the two genders by the observation of the interocular suture and by the length of the last two segments of the maxillary palps. In males, the length of the genital filaments discriminates these species. Females are distinguished by the length of the ducts of the spermathecae as well as by the narrow or enlarged shape of the neck bearing their head. Lastly, the particular position of the spines of the gonostyle coupled with molecular phylogeny led us to remove these three species from the subgenus Larroussius Nizulescu, 1931 and to classify them in a new subgenus: Lewisius Depaquit & Vongphayloth n. subg.

Title: Sur la systématique de Phlebotomus betisi et de deux nouvelles espèces apparentées du Laos avec proposition du nouveau sous-genre Lewisius. Abstract: Phlebotomus betisi a été décrit de Malaisie et fut classé après sa description dans le sous-genre Larroussius. C'était la seule espèce à posséder chez la femelle une armature pharyngienne composée de dents en forme de points et à avoir une spermathèque annelée dont la tête est portée par un cou. Les mâles se caractérisaient par un style porteur de cinq épines et par un paramère simple. L'étude de Phlébotomes originaires d'une grotte du Laos nous a permis de découvrir et de décrire deux espèces sympatriques proches de Ph. betisi Lewis & Wharton, 1963 et nouvelles pour la Science : Ph. breyi Vongphayloth & Depaquit n. sp., et Ph. sinxayarami Vongphayloth & Depaquit n. sp. Elles ont été caractérisées morphologiquement, morphométriquement, géomorphométriquement, moléculairement et protéomiquement (MALDI-TOF). Toutes ces approches convergent pour valider l'individualisation de chacune de ces espèces dont les caractères morphologiques différentiels reposent dans les deux sexes par l'observation de la suture interoculaire et par la longueur des deux derniers segments des palpes maxillaires. Chez les mâles, la longueur des filaments génitaux discrimine ces espèces. Les femelles sont distinguées par la longueur des conduits des spermathèques ainsi que par la forme étroite ou élargie du cou portant la tête de ces spermathèques. Enfin, la position particulière des épines sur le gonostyle couplée à une phylogénie moléculaire nous amène à extraire ces trois espèces du sous genre Larroussius Nitzulescu, 1931 pour les classer dans un nouveau sous-genre : Lewisius Depaquit & Vongphayloth n. subg.

MALDI-TOF MS Limits for the Identification of Mediterranean Sandflies of the Subgenus Larroussius, with a Special Focus on the Phlebotomus perniciosus Complex.

Leishmania infantum is the agent of visceral leishmaniasis in the Mediterranean basin. It is transmitted by sandflies of the subgenus Larroussius. Although Phlebotomus perniciosus is the most important vector in this area, an atypical Ph. perniciosus easily confused with Ph. longicuspis has been observed in North Africa. MALDI-TOF MS, an important tool for vector identification, has recently been applied for the identification of sandflies. Spectral databases presented in the literature, however, include only a limited number of Larroussius species. Our objective was to create an in-house database to identify Mediterranean sandflies and to evaluate the ability of MALDI-TOF MS to discriminate close species or atypical forms within the Larroussius subgenus. Field-caught specimens (n = 94) were identified morphologically as typical Ph. perniciosus (PN; n = 55), atypical Ph. perniciosus (PNA; n = 9), Ph. longicuspis (n = 9), Ph. ariasi (n = 9), Ph. mascittii (n = 3), Ph. neglectus (n = 5), Ph. perfiliewi (n = 1), Ph. similis (n = 9) and Ph. papatasi (n = 2). Identifications were confirmed by sequencing of the mtDNA CytB region and sixteen specimens were included in the in-house database. Blind assessment on 73 specimens (representing 1073 good quality spectra) showed a good agreement (98.5%) between MALDI-TOF MS and molecular identification. Discrepancies concerned confusions between Ph. perfiliewi and Ph. perniciosus. Hierarchical clustering did not allow classification of PN and PNA. The use of machine learning, however, allowed discernment between PN and PNA and between the lcus and lcx haplotypes of Ph. longicuspis (accuracy: 0.8938 with partial-least-square regression and random forest models). MALDI-TOF MS is a promising tool for the rapid and accurate identification of field-caught sandflies. The use of machine learning could allow to discriminate similar species.

Thiodicarb and methomyl tissue distribution in a fatal multiple compounds poisoning.

Thiodicarb is a nonsystemic carbamate insecticide whose acetylcholinesterase activity is related to its main methomyl degradation product. A 40-year-old woman was found dead in her car. Empty packages of medicines and an open bottle of Larvin containing thiodicarb were found near her body. No signs of violence nor traumatic injuries were noticed upon autopsy, and police investigations strongly suggested a suicide. Systematic toxicological analysis performed on postmortem specimens revealed the presence of various sedatives, hypnotics, and antipsychotic drugs in blood, urine, and gastric content. Some of the compounds identified were determined at blood concentrations well above the known therapeutic concentrations: zolpidem (2.87 mg/L), bromazepam (2.39 mg/L), nordazepam (4.21 mg/L), and levopremazine (0.64 mg/L). Specific analysis of thiodicarb and of its methomyl metabolite was then performed on all fluids and tissues collected during autopsy by liquid chromatography ion trap tandem mass spectrometry (LC-MS-MS). The anticholinesterase capacity of blood, urine, and gastric content collected at autopsy was 83%, 82%, and 32%, respectively (normal value: 0%). The presence of thiodicarb in the bottle found near the body corroborates the hypothesis of an intake of that compound. Although thiodicarb was only detected in gastric content (24.3 mg/L), its methomyl metabolite was quantified in most postmortem tissues and fluids: gastric content (19.9 mg/L), peripheral blood (0.7 mg/L), urine (8.5 mg/L), bile (2.7 mg/L), liver (0.7 mg/kg), kidney (1.7 mg/kg), lung (1.5 mg/kg), brain (9.3 mg/kg), and heart (3.6 mg/kg).

Specific and sensitive analysis of nefopam and its main metabolite desmethyl-nefopam in human plasma by liquid chromatography-ion trap tandem mass spectrometry.

A specific and sensitive liquid chromatography-tandem mass spectrometric (LC-MS-MS) method using an ion trap spectrometer was developed for quantitation of nefopam and desmethyl-nefopam in human plasma. Nefopam, desmethyl-nefopam and the internal standard (ethyl loflazepate) were extracted in a single step with diethyl ether from 1 mL of alkalinized plasma. The mobile phase consisted of acetonitrile with 0.1% formic acid (50:50, v:v). It was delivered at a flow-rate of 0.3 mL/min. The effluent was monitored by MS-MS in positive-ion mode. Ionisation was performed using an electrospray ion source operating at 200 degrees C. Nefopam and desmethyl-nefopam were identified and quantified in full scan MS-MS mode using a homemade MS-MS library. Calibration curves were linear over the concentration range of 0.78-100 ng/mL with determination coefficients >0.996. This method was fast (total run time<6 min), accurate (bias<12.5%), and reproducible (intra- and inter-assay precision<17.5%) with a quantitation limit of 0.78 ng/mL. The high specificity and sensitivity achieved by this method allowed the determination of nefopam and desmethyl-nefopam plasma levels in patients following either intermittent or continuous intravenous administration of nefopam.

Sensitive bioassay of bupivacaine in human plasma by liquid-chromatography-ion trap mass spectrometry.

A sensitive high-performance liquid chromatography-tandem mass spectrometric (HPLC-MS-MS) method, using an ion trap spectrometer, was developed for quantitation of bupivacaine in human plasma. Bupivacaine and an internal standard (ropivacaine) were extracted in a single step from 100 microL of alkalinized plasma with diethyl-ether. The mobile phase consisted of acetonitrile with 0.1% formic acid (50:50, v/v), and was delivered at a flow rate of 0.3 mL/min. The effluent was detected by MS-MS in positive ion mode. Ionisation was performed, using an electrospray ion source, operating at 200 degrees C. The selected reaction monitoring transitions m/z 289-->m/z 140 and m/z 275-->m/z 126 were chosen for bupivacaine and ropivacaine, respectively. Calibration curves were linear over the concentration range of 3.90-500 microg/L with determination coefficients >0.996. The method is accurate (bias <10%) and reproducible (intra-assay and inter-assay precision <15%), with a quantitation limit of 3.90 microg/L, using only 100 microL of plasma. The high specificity and sensitivity, achieved by this fast method (total run-time <3 min), allowed the determination of bupivacaine plasma levels in pediatric patients, following epidural administration of bupivacaine.

Body condition and habitat use by Hermann's tortoises in burnt and intact habitats.

UNLABELLED: In Mediterranean regions, fires threaten terrestrial tortoises. Nevertheless, varying proportions of adults survive fire; these surviving individuals can play a central role for population recovery. The regions devastated by fire often include important habitat of Hermann's tortoises (Testudo hermanni hermanni), so assessing the ability of survivors to persist is essential for conserving the species. Body-condition indices provide an integrative estimate of how well individuals cope with environmental variations and impacts, including fires. Between 2002 and 2009, we monitored Hermann's tortoises in intact and burnt habitats in southeastern France. In summer 2003, a strong fire ravaged half of the surveyed zone, providing an opportunity to compare body condition of tortoises between intact and burnt areas over time. Six years later, the impact of fire on vegetation was still marked; large trees were abundant in the intact area, whereas open shrub vegetation prevailed in the burnt area. In both areas, the mean body condition of tortoises fluctuated over time; however, there were no differences between the two areas. A radio-tracking experiment demonstrated that individuals from each area were residents, and not vagrants commuting between areas. We also assessed changes in body condition and microhabitat use in radio-tracked individuals. We found no significant differences between the tortoises living in the burnt and intact areas, despite subtle differences in habitat use. IN CONCLUSION: (i) surviving tortoises in an area ravaged by fire can maintain their body condition like individuals living in an intact area, and thus, individuals from burnt areas should not be translocated to supposedly better areas; and (ii) depopulated burnt areas are likely to be appropriate for population-augmentation programmes.

Trichobilharzia spp. in natural conditions in Annecy Lake, France.

Annecy Lake is a well-known focus of human cercarial dermatitis in France. Identification of the parasites, however, was not performed in the past. Previous studies suspected two species, Trichobilharzia franki and Trichobilharzia regenti, based on the presence of parasites in mallards and/or morphological identification of snails emitting ocellate furcocercariae. Following a standardized molecular approach, we studied snails and furcocercariae and compared their haplotypes with those deposited in GenBank. The selected markers were the second internal transcribed spacer ITS-2 for the snails and ITS-2 and D2 domain of the ribosomal DNA for the parasites. Our results confirm the presence of T. franki and T. regenti and two probable new species that could be potential agents of cercarial dermatitis. All the snails emitting the ocellate furcocercariae belong to the same species identified as Radix peregra (=Radix ovata = Radix balthica). Parasite-host relationships between species of the genus Trichobilharzia and snails of the genus Radix do not seem to be as specific as supposed previously.

Identification and quantification of 8 sulfonylureas with clinical toxicology interest by liquid chromatography-ion-trap tandem mass spectrometry and library searching.

BACKGROUND: Identification of sulfonylureas in blood may be useful in the evaluation of hypoglycemic crises of unknown origin. The aim of the present study was to develop a highly selective liquid chromatography-electrospray tandem mass spectrometry (MS-MS) method using an ion-trap detector for rapid screening, identification, and quantification of sulfonylureas in human plasma. METHODS: After standard liquid-liquid extraction with glisoxepide as an internal standard, 8 sulfonylureas (glibenclamide, glipizide, gliclazide, glibornuride, glimepiride, carbutamide, chlorpropamide, and tolbutamide) were eluted from a C18 column within 10 min with an isocratic mobile phase. Drugs were identified and quantified in full-scan MS-MS mode by use of a homemade MS-MS library. We used the assay in 134 cases of hypoglycemic crises of unknown origin. RESULTS: No ion suppression effect was noted for the analytes at their specific retention-time windows. For all drugs, assay validation showed good linearity (r2>0.990) and acceptable imprecision and recovery based on commonly used criteria of acceptance. The mean extraction recoveries were 63%-87% for 5 sulfonylureas but <45% for 3 (carbutamide, chlorpropamide, and tolbutamide). Nevertheless, the high sensitivity of the MS instrument made possible detection and quantification of all 8 drugs at subtherapeutic to toxic concentrations with good precision. Sulfonylureas were found in 9 hypoglycemic patients. CONCLUSION: The described assay method allows accurate, rapid identification and quantification of 8 sulfonylureas in human plasma and can be used for specific diagnosis of factitious hypoglycemia caused by ingestion of these drugs.

Discrepancy in patch test results with wool wax alcohols and Amerchol L-101.

Wondering why Amerchol L-101 (containing wool wax alcohols obtained from the hydrolysis of wool fat) appears as a test material (100%) in a cosmetics series (Chemotechnique Diagnostics AB), as wool wax alcohols were already tested in the European standard series, we added Amerchol L-101 (100%) systematically to the standard series. From 8 April 1991 to 28 February 1992, a total of 393 patients were tested: 3.05% (n = 12) showed positive test results (+, +2, +3) for wool wax alcohols as well as Amerchol L-101 (100%), 0.3% (n = 1) for wool wax alcohols only, and 11.1% (n = 44) for Amerchol L-101 (100%) only. Thus, the positive tests for Amerchol L-101 (100%) had a higher frequency of 3.4 compared to those for wool wax alcohols. In the meantime, we also added Amerchol L-101 50% (Trolab) to the standard series. Of all patients tested (n = 223) 2.7% (n = 6) had positive reactions to wool wax alcohols and 12.1% (n = 27) to Amerchol L-101 100% and/or 50%. Thus, positive reactions to Amerchol L-101 (100% and 50%) were 4.5 higher, whereas the rate of concordancy decreased. It is remarkable that 8 patients only had a positive reaction to Amerchol L-101 50% and 9 only to Amerchol L-101 100%. It seems that we miss many diagnoses in contact allergy by using only the standard wool wax alcohols test (30%).

Contact allergy to resorcinol monobenzoate.

Resorcinol monobenzoate is an ultraviolet light absorber used in cellulose plastics and is known to cause allergic contact dermatitis reactions. It is the most important allergen in spectacle frames but can be a potential allergen in shoes as well. We report 7 cases of allergy to resorcinol monobenzoate with different clinical presentations. Our series further illustrates several possible cross-sensitizers.

Diagnosis of cutaneous T-cell lymphoma detecting T-cell receptor gamma chain gene monoclonality by denaturing gradient gel electrophoresis.

Cutaneous T-cell lymphomas represent a group of malignant lymphoproliferative disorders characterised by the occurrence of a monoclonal population of T-lymphocytes. Diagnosis of early stages of this disease is a difficult challenge for both the dermatologist and the dermatopathologist. With the aid of the polymerase chain reaction it is possible to amplify specific regions of the T-cell receptor gamma gene. The amplification products can then be separated by denaturing gradient gel electrophoresis in order to detect a monoclonal population of T-lymphocytes in the infiltrate. We studied 4 patients with the clinicopathologic diagnosis of mycosis fungoides and 2 patients diagnosed as large plaque parapsoriasis. A monoclonal population was detected in 3 of the 4 mycosis fungoides cases and in 1 of the patients with large plaque parapsoriasis. This indicates that our analysis can help us establishing a diagnosis, and it can also help us to identify patients with a possible early stage of the disease, which clinically or histologically is not yet recognised as such.

Toxic epidermal necrolysis associated with treatment for preterm labor.

We report a 29-year-old pregnant woman who developed toxic epidermal necrolysis at 29 weeks of gestation after administration of ritodrine, indomethacin and betamethasone. Toxic epidermal necrolysis is an unreported side effect of this widely used combination of medications. Since toxic epidermal necrolysis is a potentially fatal disease, awareness of a possible association is warranted.

Diagnosis of Cutaneous T-Cell Lymphoma Detecting T-Cell Receptor γ Chain Gene Monoclonality By Denaturing Gradient Gel Electrophoresis.

Cutaneous T-cell lymphomas represent a group of malignant lymphoproliferative disorders characterised by the occurrence of a monoclonal population of T-lymphocytes. Diagnosis of early stages of this disease is a difficult challenge for both the dermatologist and the dermatopathologist. With the aid of the polymerase chain reaction it is possible to amplify specific regions of the T-cell receptor gamma gene. The amplification products can then be separated by denaturing gradient gel electrophoresis in order to detect a monoclonal population of T-lymphocytes in the infiltrate. We studied 4 patients with the clinicopathologic diagnosis of mycosis fungoides and 2 patients diagnosed as large plaque parapsoriasis. A monoclonal population was detected in 3 of the 4 mycosis fungoides cases and in 1 of the patients with large plaque parapsoriasis. This indicates that our analysis can help us establishing a diagnosis, and it can also help us to identify patients with a possible early stage of the disease, which clinically or histologically is not yet recognised as such.

Contact and photocontact allergy to ketoprofen. The Belgian experience.

Topical ketoprofen (KP) is widely used because of its anti-inflammatory effect. However, photocontact dermatitis is a side-effect. Between May 2001 and June 2002, the Belgian Contact & Environmental Dermatitis Group conducted a prospective, open patch and photopatch test study in 20 patients suspected of KP dermatitis. Severe skin symptoms requiring systemic corticotherapy occurred in 47%. 5 patients were hospitalized. 1 patient showed prolonged photosensitivity. All patients were tested with KP and the other constituents of KP gel. Attribution to KP was demonstrated in all cases. Patch and photopatch tests with KP 2% in petrolatum showed contact photoallergy in 17 patients, contact allergy in 1 patient and photoaggravated contact allergy in 2 patients. 5 patients also reacted to the fragrance components lavender (Lavandula augustifolia) oil and/or neroli (Citrus aurantium dulcis) oil 5% in alcohol. However, in 4 of these, irritant reactions to the ethanolic dilutions could not be ruled out. Additional tests with 3 non-steroidal anti-inflammatory drugs without benzophenone structure ibuprofen, naproxen and diclofenac identified only 1 contact allergic reaction to diclofenac. Cross-reactivity to the substituted benzophenones, oxybenzone and sulisobenzone occurred only to the first in less than 30% of the patients. A high frequency (69%) of contact allergy to fragrance mix was found. Dermatologists should be aware of the severity of photoallergic reactions to KP and the risk of cross-sensitization.

Itraconazole penetrates the nail via the nail matrix and the nail bed--an investigation in onychomycosis.

Nail-matrix kinetics were studied in 21 patients (19 with onychomycosis, two with tinea corporis) as soon as taking itraconazole (Sporanox) 100 mg daily for up to 7 months. Itraconazole was detected in the distal nail as soon as 1 month after the start of therapy (42 ng/g in fingernails and 16 ng/g in toenails). During the course of treatment, this concentration rose and reached a mean of 160 ng/g in fingernail clippings and 197 ng/g in toenail clippings. Moreover, in fingernails of 12 out of 21 patients and in toenails of six out of 20 patients, itraconazole was detected in the distal nail clippings before full outgrowth of the fastest-growing nail. In most patients, itraconazole was detected in the distal nail clippings earlier than would be expected if the drug were incorporated only via the nail matrix, indicating that in addition to the nail matrix, a second route of penetration into the nail exists, i.e. the nail bed.

Influence of C6 and CNS1 brain tumors on methotrexate pharmacokinetics in plasma and brain tissue.

PURPOSE: Comparison of the influence of two different brain tumors (C6 and CNS1 glioma) on methotrexate (MTX) disposition in plasma, brain, and tumor tissue extracellular fluid (ECF). METHODS: Serial collection of plasma samples and brain ECF dialysates after i.v. bolus administration of MTX (50 mg kg(-1)) for 4 h. Quantitation of MTX concentrations by HPLC-UV. RESULTS: Histological studies revealed a 3-fold higher number of blood vessels in CNS1 than in C6 tumor tissue. In vivo recoveries (reverse dialysis) were significantly different in tumor tissue (C6: 8.0 +/- 3.8%; CNS1: 4.9 +/- 2.5%), and in the contralateral hemisphere (C6: 6.0 +/- 4.0%; CNS1: 3.9 +/- 2.5%) between the two tumors. Area under the concentration-time curve (AUC) in plasma was 30% higher in CNS1 than in C6 due to a lower systemic clearance. Maximum MTX levels in brain tumor ECF were significantly higher in CNS1 than in C6, and decreased faster in CNS1 than in C6 tumor-bearing rats. Penetration in tumor ECF (AUC(ECF)/AUC(Plasma) ratio) was similar in CNS1 and C6. MTX concentrations in contralateral hemisphere were significantly lower than in tumor tissue and dependent on tumor model. CONCLUSION: C6 and CNS1 brain tumors have a distinct yet highly variable impact on MTX penetration in brain and brain tumor ECF.