Help in the Choice of Automated or Semiautomated Immunoassays for Serological Diagnosis of Toxoplasmosis: Evaluation of Nine Immunoassays by the French National Reference Center for Toxoplasmosis.

Toxoplasmosis, a benign infection, is asymptomatic or paucisymptomatic in over 80% of cases, except in immunocompetent patients suffering from ocular toxoplasmosis or in immunocompromised patients with opportunistic or congenital toxoplasmosis. Diagnosis is based mainly on serology testing. Thus, we compared the performance of the nine most commonly used commercial automated or semiautomated immunoassays for IgG and IgM Toxoplasma gondii antibody detection, that is, the Advia Centaur, Architect, AxSYM, Elecsys, Enzygnost, Liaison, Platelia, VIDAS, and VIDIA assays. The assays were conducted on four panels of serum samples derived during routine testing from patients with an interfering disease and who exhibited a low IgG antibody level in one of two clinical settings, namely, acute or chronic toxoplasmosis. As a result, IgG sensitivities ranged from 97.1% to 100%, and IgG specificities ranged from 99.5% to 100%. For IgG quantification, strong differences in IgG titers (expressed in IU/ml) were noted depending on the assay used. IgM sensitivities ranged from 65% to 97.9%, and IgM specificities ranged from 92.6% to 100%. For defining the best serological strategies to be implemented, it appears crucial to compare the diagnostic performance of the different tests with respect to their specificity and sensitivity in detecting the presence of IgG and IgM antibodies.

Ruling out nosocomial transmission of Cryptosporidium in a renal transplantation unit: case report.

BACKGROUND: Cryptosporidium spp. is a ubiquitous parasite affecting humans as well as domestic and wild vertebrates, causing diarrhea in both immunocompetent and immunocompromised hosts worldwide. Its transmission occurs primarily by the fecal-oral route. In humans, C. parvum and C. hominis are the most prevalent species, whereas immunocompetent and immunocompromised individuals can also be infected by other zoonotic species. Renal transplant patients are prone to develop cryptosporidiosis, which can induce severe and life-threatening diarrhea. CASE PRESENTATION: We report here a series of nearly concomitant cases of acute symptomatic cryptosporidiosis in three renal transplant patients attending the Strasbourg University Hospital Nephrology Unit. The clinical presentation was persistent diarrhea and acute renal failure. The diagnosis was confirmed by microscopic stool examination using a modified Ziehl-Neelsen staining method and species identification by molecular tools. All patients were treated with nitazoxanide and recovered from diarrhea after 14 days of therapy. CONCLUSION: Genotypic species identification was not consistent with an epidemic context, thus underlining the need for genotyping to monitor at risk patients.

Toxoplasma seroconversion with negative or transient immunoglobulin M in pregnant women: myth or reality? A French multicenter retrospective study.

Classically, Toxoplasma infection is associated with high levels of specific IgM antibody and a rise in specific IgG levels 1 to 3 weeks later. Atypical IgG seroconversion, without IgM detection or with transient IgM levels, has been described during serologic follow-up of seronegative pregnant women and raises difficulties in interpreting the results. To evaluate the frequency and the characteristics of these atypical cases of seroconversion, an investigation was conducted within the French National Reference Center for Toxoplasmosis, from which 26 cases collected from 12 laboratories belonging to the network were identified. The aim of this work was to retrospectively analyze the results of serologic testing, the treatments administered, and the results of prenatal and postnatal follow-up for these women. In each case, IgG antibodies were detected using both screening and confirmatory tests. IgM antibodies were not detected in 15 cases, and the levels were equivocal or low-positive in 11 cases. The IgG avidity results were low in 16 cases and high in one case. Most of the pregnant women (22/26) were treated with spiramycin from the time that IgG antibodies appeared until delivery. Amniotic fluid was analyzed for Toxoplasma gondii DNA by PCR in 11/26 cases, and the results were negative in all cases. Congenital toxoplasmosis was ruled out in 12/26 newborns. There was no abnormality observed at birth for 10 newborns and no information available for 4 newborns. In conclusion, when the interpretation of serological results is so difficult, it seems cautious to initiate treatment by spiramycin and to follow the pregnant women and their newborns.

Antifungal activity of sodium bicarbonate against fungal agents causing superficial infections.

Although sodium bicarbonate-NaHCO(3) (SB) has many domestic and medical, traditional and empirical uses, only little scientific documentation of its activity is available. The aims of this study were to investigate the antifungal activity of SB on the three fungal groups (yeasts, dermatophytes and molds) responsible for human skin and nail infections. We first evaluated the in vitro antifungal activity of SB on 70 fungal strains isolated from skin and nail infections: 40 dermatophytes, 18 yeasts and 12 molds. A concentration of 10 g/L SB inhibited the growth of 80% of all the fungal isolates tested on Sabouraud dextrose agar. The minimal inhibitory concentration 90 (MIC90) of SB measured on Sabouraud dextrose agar, Sabouraud dextrose broth and potato dextrose broth was 5 g/L for the yeasts, 20 g/L for the dermatophytes and 40 g/L for the molds. In a second step, we prospectively evaluated the ex vivo antifungal activity of SB on 24 infected (15 dermatophytes, 7 yeasts and 2 molds) clinical specimens (15 nails and 9 skin scrapings). The fungal growth was completely inhibited for 19 (79%) specimens and reduced for 4 (17%) specimens after 7 days of incubation on Sabouraud dextrose-chloramphenicol agar supplemented with 10 g/L of SB as compared to Sabouraud dextrose-chloramphenicol agar without SB. In conclusion, we documented the antifungal activity of SB on the most common agents of cutaneous fungal infection and onychomycosis, and we specified the effective concentrations for the different groups of pathogenic fungi. The mechanism of action of SB has yet to be explored.

Human babesiosis in Alsace.

OBJECTIVES: Human babesiosis is a rare parasitic anthropozoonosis transmitted to humans by tick bites. Fifty-six cases of human babesiosis have been recorded in Europe. Two cases of babesiosis were reported in Alsace, France, in 2009. We performed a retrospective observational descriptive study to assess the epidemiology of the disease in Alsace. METHODS: Patients were included if they had a positive serology result for Babesia and/or a positive blood smear and/or a positive PCR result. The tests were performed in the microbiology laboratories of the university hospitals of Strasbourg, the civil hospitals of Colmar, and the hospital of Mulhouse between January 1, 2005 and December 31, 2015. Included patients were divided into three groups: definite case group (positive PCR or positive blood smear or seroconversion), possible case group (positive serology results without seroconversion with a compatible clinical picture and without other confirmed diagnoses), and incompatible case group (positive serology results without seroconversion, without compatible clinical picture and/or with other confirmed diagnoses). The compatible clinical picture was defined by the presence of flu-like symptoms and fever (≥38°C). RESULTS: Fifty-one patients had at least one positive result. Three cases were excluded (missing files). There were six definite cases, 12 possible cases, and 30 incompatible cases. All patients in the definite case group were immunocompetent. No deaths occurred. CONCLUSIONS: Human babesiosis is probably underdiagnosed due to its non-specific symptoms, lack of awareness about the disease, and the difficulty in making a diagnosis.

Different effect of Toxoplasma gondii infection on adhesion capacity of fibroblasts and monocytes.

This study investigated the effect of infection with the apicomplexan parasite Toxoplasma gondii, in combination with the concomitant cytokine environment (IFN-gamma/TNF-alpha), on adhesion of THP-1 monocytic cells to MRC-5 fibroblasts. Surprisingly, infection of THP-1 cells decreased their adhesion to the MRC-5 cell monolayer. This decrease was compensated by IFN-gamma/TNF-alpha stimulation. In contrast, infection of MRC-5 cells significantly increased adhesion, which was synergistically augmented by cytokine stimulation. Levels of ICAM-1 (CD54) on MRC-5 cells, as well as LFA-1 (CD11a) on THP-1 cells, were not changed by infection, neither in resting, nor in cytokine stimulated cells. These results show that T. gondii infection alters adhesion properties and reactivity to cytokine stimulation in a cell-specific way.

New insights in toxoplasmosis immunology during pregnancy. Perspective for vaccine prevention.

Toxoplasma gondii is one of the few pathogens that can cross the placenta. Frequency and severity of transmission vary with gestational age. While acquired toxoplasmosis is already well explored, the control of maternal-foetal transmission of the parasite remains almost unknown. This is partly due to inherent inadequacies of animal models. This review summarises the studies which have been undertaken and shows that the mouse is a valuable model despite obvious differences to the human case. The paramount role of the cellular immune response during primary infection has been consistently shown. Surprisingly, IFN-g has a dual role in this process. While its beneficial effects in the control of toxoplasmosis are well known, it also seems to have transmission-enhancing effects within the placenta and can also directly harm the developing foetus. This shows the importance of designing vaccines which protects both mother and foetus. Therefore, it is useful to study the mechanisms of natural resistance against transmission during a secondary infection. In this setting, the process is more complicated, involving cellular, but also humoral components of the immune system. In summary, even if the whole process is far from being elucidated, important insights have been gained so far which will help us to undertake rational vaccine research.

[Human subcutaneous dirofilariosis due to Dirofilaria repens. A case diagnosed in Strasbourg, France]. / Dirofilariose sous-cutanée humaine due à dirofilaria repens. A propos d'un cas diagnostiqué à Strasbourg, France.

We report a case of human dirofilariosis in a 35 year-old man living in Alsace (North-East France), who presented a subcutaneous nodule of the left cheek, strongly simulating an epidermic cyst. Surgical excision and histopathological examination unexpectedly established the diagnosis by the presence of nematode worm sections identified as Dirofilaria (Nochtiella) repens. This patient living in a rural area of Strasbourg spent one week in Toulon, Southern France. This new observation of cutaneous dirofilariosis constitutes the third Alsatian case described in literature, and emphasizes the presence of this parasitic disease in Southern France.

Species of Metarhizium anisopliae complex implicated in human infections: retrospective sequencing study.

OBJECTIVES: Fungi belonging to the Metarhizium anisopliae complex comprise ubiquitous arthropod pathogenic moulds used as mycopesticides. Rare cases of human infections due to M. anisopliae have been reported. We hypothesize misidentifications of fungal strains implicated in these cases or used in mycopesticides. METHODS: A review of the literature was conducted to identify previously published cases. We collected some of these previous described strains and reported new cases, and a French mycopesticide containing M. anisopliae. All identifications were performed based on elongation factor-1α gene sequencing. RESULTS: We report eight new cases of Metarhizium infection in humans (three from France and five from Australia). The strains isolated from these cases, and three others from already published cases and reported as M. anisopliae, were molecularly identified based on elongation factor-1α (Ef1-α) gene sequencing as follows: Metarhizium robertsii (six), Metarhizium guizhouense (three), Metarhizium brunneum (one) and Metarhizium pingshaense (one). CONCLUSIONS: In this study, we report new human cases of Metarhizium infections, and, based on Ef-1α gene sequencing, we demonstrate the misidentification of species in case reports. We also correct the species identification of a strain reported as M. anisopliae used in a commercially available mycopesticide. According to our results, none of the strains from the human infection reports reviewed belongs to the species M. anisopliae.

Serological diagnosis of Toxoplasma gondii infection: Recommendations from the French National Reference Center for Toxoplasmosis.

Toxoplasmosis manifests no clinical signs in 80% of cases in immunocompetent patient, causing immunization characterized by the persistence of cysts, particularly in brain, muscles, and retina. Assessing the serological status, based on testing for serum toxoplasma IgG and IgM antibodies, is essential in cases that are increasingly at risk for the more severe disease forms, such as congenital or ocular toxoplasmosis. This disease also exposes immunosuppressed patients to reactivation, which can lead to more widespread forms and increased mortality. By interpreting the serological results, we can estimate the risk of contamination or reactivation and define appropriate prophylactic and preventive measures, such as hygienic and dietetic, therapeutic, biological, and clinical follow-up, according to the clinical context. We hereby propose practical approaches based on serological data, resulting from a consensus of a group of experts from the French National Reference Center Network for Toxoplasmosis, according to both routine and specific clinical situations.

Epidemiological study of invasive pulmonary aspergillosis in a haematology unit by molecular typing of environmental and patient isolates of Aspergillus fumigatus.

In order to determine the possible relationship between environmental contamination by Aspergillus fumigatus and occurrence of invasive aspergillosis, a one-year prospective study was carried out in the haematology ward of Hautepierre Hospital, Strasbourg, France. During the study period, 21 environmental isolates and 26 clinical isolates of A. fumigatus were collected. Each was genotyped using a random amplification of polymorphic DNA (RAPD) technique. Thirty-four distinct profiles were identified by RAPD analysis, indicating the great genetic diversity of A. fumigatus isolated from infected patients and from the environment. For two patients, RAPD analysis demonstrated concurrent infection by at least two different strains. In two cases, a genetic similarity was noted between isolates obtained from a patient and from the environment.

Molecular evidence of ovine (G1) and camel (G6) strains of Echinococcus granulosus in Tunisia and putative role of cattle in human contamination.

Three hundred and seventy-two cysts coming from 50 humans, 166 cattle, 153 sheep and 3 camels were collected in order to establish some epidemiological molecular information in Tunisia for the first time. The analysis by PCR-RFLP of ITS1 sequence showed that all the human, ovine and bovine cysts were due to the common sheep strain of Echinococcus granulosus. The sequencing of the CO1 gene of 37 isolates confirm the G1 genotype of this strain. For seven of these isolates, we found the mutation C56T which is present in the three principal intermediate hosts: human (three cysts), cattle (three cysts) and sheep (one cyst). With regard to the G1 genotype, we identified three other point mutations. The camel strain G6 is uniquely found in the three camels isolates and not in the other intermediate hosts analysed. The fertility of the bovine cyst represents 48% that means that this host is involved in a bovine-dog cycle and consequently represents a reservoir of sheep strain in Tunisia. Our results confirm the importance of the prophylaxis measures in order to disrupt the cycle of transmission sheep-dog in Tunisia. Nevertheless, the supervision of bovine infection should be reinforced because this intermediate host may constitute an important link with the human contamination.

[Transfusion-transmitted malaria, preventive measures]. / Le paludisme transfusionnel, les mesures de prévention.

Malaria is a blood parasitic disease in the first place among the most prevalent communicable infectious diseases over the world, which leads to an increased risk of transfusion transmitted malaria. Preventive measures have been undertaken to screen blood donors such as discarding red cell donations according to the medical history, travel history and detection of malarial antibodies. However, these measures may be not sufficient and reliable to avoid the risk of transmission. Preliminary data indicates that combination of travel history, detection of malarial antibodies and antigens by commercialized kits adapted to blood transfusion centres either in endemic or non endemic areas may improve malaria transfusion risk management.

[Evaluation of in vitro efficacy of combined riboflavin and ultraviolet-A (365 nm) for Acanthamoeba]. / Efficacité anti-amibienne in vitro de l'association riboflavine/UV-A.

INTRODUCTION: Acanthamoeba keratitis is a rare but serious disease and is particularly difficult to treat when the diagnosis is delayed, partly because of the limitations of current therapies. The purpose of our study is to evaluate the anti-amoebic effectiveness of riboflavin and UV-A on Acanthamoeba castellani. MATERIALS AND METHODS: We tested the effect of 0.02% chlorhexidine alone (C), the combination of riboflavin 1% and UV-A (UV-A+R), and the combination of the two treatments (R+C+UV-A) on cultures of vegetative and cystic forms of A. castellani. We conducted a parasite count under optical microscopy for each treated area at day 1, 4 and 8. RESULTS: There was a decrease in the number of cysts for all three treatments (C, UV-A+R, R+C+UV-A). This reduction was greater for the plates treated with R+UV-A (P <0.01 at D8) and those treated with C+R+UV-A (P<0.001 at D8) compared to those exposed to chlorhexidine alone (C). There was no decrease in the number of amoebic trophozoites for the three treatments (C, UV-A+R, R+C+UV-A), but encystment was observed. DISCUSSION: Given the in vitro efficacy of riboflavin combined with UV-A against cystic forms of A. castellani and excellent in vivo tolerance of the procedure, the treatment of acanthamoeba keratitis might be improved by this new therapeutic approach.

Depression of the N-demethylation of erythromycin, azithromycin, clarithromycin and clindamycin in murine Toxoplasma infection.

The N-demethylation of macrolides was studied in a murine model of infection. Mice were infected with a cystogenic strain of Toxoplasma gondii (20 or 40 cysts/mouse) and microsomes were prepared from liver homogenates and jejunum villus tip enterocytes on day 10 post-infection. The rate of N-demethylation of the anti-Toxoplasma macrolides azithromycin, clarithromycin and clindamycin was investigated and compared to that of the macrolide erythromycin, a marker of activity of the cytochrome P-450 3A (CYP3A) mono-oxygenases. In infected mice (20 cysts/mouse), the rate of N-demethylation fell in the liver and jejunum for erythromycin (-25% and -35%, respectively), azithromycin (-12% and -10%, respectively), clarithromycin (-23% and -21%, respectively) and clindamycin (-20% and -28%, respectively). The degree of hepatic depression was more marked in mice receiving a 40-cysts burden: for erythromycin (-54%), azithromycin (-29%), clarithromycin (-49%) and clindamycin (-47%).

Loss of oral infectivity of tissue cysts of Toxoplasma gondii RH strain to outbred Swiss Webster mice.

We have developed a method for obtaining cysts of Toxoplasma gondii RH strain. Outbred Swiss Webster mice were infected subcutaneously (s.c.) or intraperitoneally (i.p) with 10(5) tachyzoites and given sulfadiazine 400 mg l-1 + NaHCO3 10 g l-1 in drinking water from day 1 to day 15 post-infection (p.i.). None of the mice infected i.p. survived, compared with 50% of the mice infected s.c. Cysts were detectable in the brain on day 45 p.i., and had ultrastructural features consistent with those of bradyzoites. However, these cysts were incapable of infecting mice via the oral route. In addition, immunofluorescence studies showed the persistence of P36 protein expression, indicating that the conversion to bradyzoites was incomplete.

[Gel fractionation of immunoglobulins in the detection of specific antitoxoplasma IgM. Evaluation of a 1-year study]. / Apport du fractionnement sur gel des immunoglobulines dans la détection des IgM spécifiques antitoxoplasmiques. Bilan d'une année d'étude.

Toxoplasmosis seroconversion is diagnosed on the presence of fluorescent antibodies. IgG level kinetics and presence of IgM at the Remington test are usually the points taken in consideration. But the Remington test can be false positive, or inadequate. The authors report their experience of gel filtration of sera in toxoplasmosis, and discuss the help that this method can be establishing diagnosis in the case of a positive Remington test.

A parasitologically proven case of Toxoplasma pneumonia in an immunocompetent pregnant woman.

We report a case of primary pulmonary toxoplasmosis in an immunocompetent pregnant woman. Seroconversion did not occur until 2 weeks after detection of the parasite in bronchoalveolar lavage fluid. It is suggested that toxoplasma infection should be considered in any atypical pneumonia in which the X-ray shows diffuse pulmonary infiltrates.