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Aspergillus flavus is the second leading cause of invasive and non-invasive aspergillosis. Secretion of hydrolytic enzymes is considered as a virulence factor in this species. Our work aimed to study in vitro production of some virulence factors, to evaluate the biofilm production against human and avian A. flavus isolates and to investigate the antifungal susceptibility agents. Hydrolytic enzymes, biofilm production and molecular typing were studied for 62 human and 36 avian A. flavus isolates by specific solid media and six microsatellite markers. The susceptibility to antifungal agents was evaluated for 37 human isolates. All human and avian A. flavus isolates showed positive activities of extracellular hydrolase: phospholipase, protease and hemolysin. A positive elastase activity was seen in 64.51% of human A. flavus isolates and 86.1% of avian A. flavus isolates. All A. flavus in these two populations formed biofilms. Statistical significant difference was observed for the mean phospholipase activities (P = 0.025) and biofilm quantification (P = 0.0001) between human and avian A. flavus isolates. The in vitro susceptibility results showed a resistance in 83.7%, 81.08% and 16.21% of A. flavus isolates respectively to amphotericin B, itraconazole and posaconazole. No association was noted between all virulence factors and the genotypes of human and avian isolates. Our study allowed us to show that human strains have a higher production of extracellular hydrolases and biofilm then avian strains. These virulence factors appear to act synergistically to contribute to the virulence of A. flavus strains. Moreover, significant correlation between virulence patterns and antifungal susceptibility profiles was observed.

Introduction. Trichosporon asahii has been recognized as an opportunistic agent having a limited sensitivity to antifungal treatment. Hypothesis/Gap Statement. Molecular mechanisms of azole resistance have been rarely reported for Trichosproron asahii. Similar to other fungi, we hypothesized that both ERG11 gene mutation and efflux pumps genes hyper-expression were implicated. Aim. The current work aimed to study the sensitivity of clinical T. asahii isolates to different antifungal agents and to explore their resistance mechanisms by molecular methods including real-time PCR and gene sequencing. Methods. The sensitivity of T. asahii isolates to fluconazole, amphotericin B and voriconazole was estimated by the Etest method. Real-time PCR was used to measure the relative expression of Pdr11, Mdr and ERG11 genes via the ACT1 housekeeping gene. Three pairs of primers were also chosen to sequence the ERG11 gene. This exploration was followed by statistical study including the receiver operating characteristic (ROC) curve analysis to identify a relationship between gene mean expression and the sensitivity of isolates. Results. In 31 clinical isolates, the resistance frequencies were 87, 16.1 and 3.2 %, respectively, for amphotericin B, fluconazole and voriconazole. Quantitative real-time PCR demonstrated that only Mdr over-expression was significantly associated with FCZ resistance confirmed by univariate statistical study and the ROC curve analysis (P ERG11 sequencing revealed two mutations H380G and S381A in TN325U11 (MIC FCZ=8 µg ml−1) and H437R in TN114U09 (MIC FCZ=256 µg ml−1) in highly conserved regions (close to the haem-binding domain) but their involvement in the resistance mechanism has not yet been assigned. Conclusion. T. asahii FCZ resistance mechanisms are proven to be much more complex and gene alteration sequence and/or expression can be involved. Only Mdr gene over-expression was significantly associated with FCZ resistance and no good correlation was observed between FCZ and VCZ MIC values and relative gene expression. ERG11 sequence alteration seems to play a major role in T. asahii FCZ resistance mechanism but their involvement needs further confirmation.

We report a case of a young woman with acquired immune deficiency syndrome admitted with fever and abdominal pain. Saccharomyces kluyveri was isolated in blood culture. She was successfully treated with fluconazole. This case demonstrates the pathogenicity of Saccharomyces Kluyveri in immunocompromised host.

Purpose. The objectives of our study were species identification and genotyping of Trichosporon isolates collected at the Parasitology and Mycology Laboratory in Sfax, Tunisia. Methodology. Molecular identification was carried out by analysing the IGS1 regions of the rDNA of 30 Trichosporon isolates. Results. Trichosporon asahii was the most frequent species detected. Furthermore, four genotypes were identified in Tunisia: 1 (46.4 %), 4 (35.7 %), 7 (14.3 %) and 3 (3.6 %). In vitro antifungal susceptibility testing of the isolates showed that voriconazole exhibited the highest activity. Conclusion. This is the first reported study of genotype identification of T. asahii in Tunisia and even in the African continent.

OBJECTIVES Rhizopus arrhizus is recognized as an emergent agent of superficial and invasive mucormycosis. Despite an increasing number of these infections, the molecular epidemiology of Rhizopus species has not been well studied. MATERIALS AND METHODS In this study, 43 R. arrhizus strains (25 environmental and 18 clinical isolates) were genotyped using six novel panels of microsatellite markers. RESULTS Upon the analysis of 43 isolates, 4-8 distinct alleles were detected for each marker. The discriminatory power for the individual markers ranged from 0·522 to 0·830. The combination of all six markers yielded 33 different haplotypes with a high degree of discrimination (0·989 D value). A four-marker combination were selected as the most parsimonious panel achieving D > 0·95. One clinical isolate and one environmental isolate shared the same genotype suggesting the possible nosocomial outbreak of mucormycosis in hospitalized patients. We have noted that the strains isolated from cutaneous mucormycosis were different from the strains isolated from rhino-orbito-cerebral mucormycosis. Then, the hypothesis of particular tropism of infectious strains for a given site is not excluded. The standardized indices of association IA and rBarD were significantly different from zero (P

Free-living amoebae (FLA) are opportunistic and ubiquitous protozoa that are widely found in various environmental sources. They are known to cause serious human infections. The aim of our study was to detect FLA and Acanthamoeba spp. in hospital water circuits. Eighty-four water samples were collected over a period of 4 months (September-December 2011) from different wards of the Sfax University Hospital (surgical services, intensive care unit, operating theater, and water storage tanks). FLA were detected in 53.5 % of samples as follows: surgical services (80 %), operating theater and surgical intensive care unit (13.3 %), medical intensive care unit (0 %), water storage tanks (6.6 %). The predominant morphotype was the acanthopodial (89 %). The others morphotypes were as follows: monopodial (40 %), dactylopodial (22 %), rugosa (62 %), eruptive (24 %), fan shaped (18 %), and polypodial (18 %). Acanthamoeba was found in 40 samples (47.6 %). 64.2 % of isolates were identified as Acanthamoeba spp. by PCR, using primers to amplify a region of 18S rDNA which showed variation in the product length. Sequence analysis of five PCR products identified Acanthamoeba sp. These isolates belong to T4, T10, and T11 genotypes, and to our knowledge this is the first report of the T10 and T11 genotype in Tunisia.The occurrence of potentially pathogenic FLA in the hospital environment may represent a health risk for patients, since these organisms can cause severe opportunistic illness and also can harbor pathogenic agents. Thus, increased awareness regarding these parasites and recognition of their importance, particularly in immunocompromised patients is crucial.

We report a case of invasive fungal sinusitis caused by Scopulariopsis in 57 year-old man who had recurrence of orbital cellulitis. CT-scan and magnetic resonance imaging found an orbital cellulitis associated to a left frontal sinusitis with bone erosion and calcification. Patient was treated by surgical debridement and voriconazole. Culture of excised tissue was positive for Scopulariopsis.

Yarrowia lipolytica is weakly pathogenic yeast, which is rarely isolated from the blood. We report unusual cases of Y. lipolytica fungemia occurred between October 2012 and June 2014 in the intensive care unit (ICU) of the UH Habib Bourguiba Sfax. During this period, 55 cases of Y. lipolytica septicemia were diagnosed. There were 44 men and 11 women (sex ratio = 4).The median age was 43 years. The broad-spectrum antibiotics (100 %), the catheterization (96 %), and the prolonged hospitalization in ICU (91 %) were the main risk factors. Patients were hospitalized in ICU, mostly, for polytraumatism (45.4 %), pneumopathy (9 %), and post-operative complications (7 %). Fever unresponsive to broad-spectrum antibacterial therapy was the predominant sign of infection (83.6 %). Y. lipolytica was isolated in one or several blood cultures (14.5 %) and in the catheter tip culture of nine patients (16.3 %).Treatment was based on intravenous amphotericin B (58.2 %), fluconazole (45.4 %) and/or removal catheter (69 %). Apyrexia or blood cultures sterilization was obtained for 34 patients (61.8 %). Y. lipolytica candidemia is an opportunistic and emerging human yeast pathogen. It can reach to the bloodstream of immunocompromised or critically ill patients during hospitalization through intravascular catheterization. Further clinical data need to be evaluated for formulating management strategies of seriously ill patients infected with uncommon fungal agents.

Background The aim of this study was to determine the biofilm formation, the extracellular enzymatic activities of 182 clinical isolates of the Candida parapsilosis complex. Methods Molecular identification of the C. parapsilosis species complex was performed using PCR RFLP of SADH gene and PCR sequencing of ITS region. The susceptibility of ours isolates to antifungal agents and molecular mechanisms underlying azole resistance were evaluated. Results 63.5% of C. parapsilosis were phospholipase positive with moderate activity for the majority of strains. None of the C. metapsilosis or C. orthopsilosis isolates was able to produce phospholipase. Higher caseinase activities were detected in C. parapsilosis (Pz = 0.5 ± 0.18) and C. orthopsilosis (Pz = 0.49 ± 0.07) than in C. metapsilosis isolates (Pz = 0.72 ± 0.1). 96.5% of C. parapsilosis strains and all isolates of C. metapsilosis and C. orthopsilosis produced gelatinase. All the strains possessed the ability to show haemolysis on blood agar. C. metapsilosis exhibited the low haemolysin production with statistical significant differences compared to C. parapsilosis and C. orthopsilosis. The biofilm forming ability of C. parapsilosis was highly strain dependent with important heterogeneity, which was less evident with both C. orthopsilosis and C. metapsilosis. Some C. parapsilosis isolates met the criterion for susceptible dose dependent to fluconazole (10.91%), itraconazole (16.36%) and voriconazole (7.27%). Moreover, 5.45% and 1.82% of C. parapsilosis isolates were respectively resistant to fluconazole and voriconazole. All strains of C. metapsilosis and C. orthopsilosis were susceptible to azoles; and isolates of all three species exhibited 100% of susceptibility to caspofungin, amphotericin B and 5-flucytosine. Conclusions A combination of molecular mechanisms, including the overexpression of ERG11, and genes encoding efflux pumps (CDR1, MDR1, and MRR1) were involved in azole resistance in C. parapsilosis.

Candida glabrata has emerged as an opportunistic pathogen of considerable importance in invasive and superficial infections. Aims To analyze the development of fluconazole resistance in patients under treatment through epidemiological survey in our hospital. Patients and methods Twenty two patients (89 clinical strains) were collected. Molecular typing of isolates was performed by polymorphic markers. Analysis of gene expression was realized by reverse transcriptase-real time polymerase chain reactions (RT-qPCR). Results Genetic analysis showed that 63% persists with apparently unchanged strains (n = 14). Among them, four showed fluconazole resistance development. A strain replacement was observed in 6 patients and two patients selected more resistant isolates during the course of treatment. An analysis of Candida glabrata cerebellar degeneration-related protein 1 (CgCDR1), Candida glabrata cerebellar degeneration-related protein 2 (CgCDR2) and Candida glabrata sterol 14 alpha-demetylase Erg 11 (CgERG11) expression revealed an over-expression in 10 resistant isolates. Conclusion This study demonstrated that C. glabrata strain undergo frequent changes in vivo . The increase in CgCDR1 and CgCDR2 expression was the most mechanism associated with fluconazole resistance.

Summary Invasive fungal infections are a major complication and an important cause of morbidity and mortality among solid organ transplant recipients. Their diagnosis is difficult and their prognosis is often pejorative. Objective The aim of this study was to report the cases of invasive fungal infections in renal transplant recipients in Habib Bourguiba Sfax university hospital and to identify the main fungal agents. Materials and methods It is a retrospective study of invasive fungal infections in renal transplant recipient reported in our hospital from January 1995 to February 2013. Results Invasive fungal infections were diagnosed in 11 cases (3.4%) among 321 renal transplant recipients. These infections included four cases of pneumocystosis, two cases of candidiasis, two cases of aspergillosis, two cases of cryptococcosis and one case of mucormycosis. There were six men and five women. The mean age was 37 years. The infection was late in 63% of cases (> 3 months after transplantation). The prolonged corticosteroid and immunosuppressive therapy were the main risk factors (100%) followed by renal failure (45%), graft rejection (45%), broad spectrum antibiotics (45%), CMV infection (36%), neutropenia (36%) and dialysis (18%). The evolution under treatment was favourable only in two cases (18%). Conclusion Invasive fungal infections are not common among kidney transplant recipients. However, they remain an important cause of morbidity and mortality in this group of patients. Prevention, early diagnosis and appropriate management are necessary to improve prognosis and reduce mortality rate.

The genus Fusarium, initially known for its important agro-economic impact, is more and more often implicated in human pathology. In fact, multiples allergic, toxic and infectious manifestations are more reported in immunocompetent and immunocompromised hosts. The objective of our study was to analyse the epidemiological, mycological and clinical features of fusariosis reported in our CHU. Eighty-seven cases of Fusarium infections were collected: 34 cases of onychomycosis (39%), 26 cases of intertrigos (30%), 25 cases of keratomycosis (29%), one case of atypical invasive fusariosis due to Fusarium oxysporum species complex (FOSC) and one case of localized gingivolabial fusariosis due to Fusarium solani species complex (FSSC) in a patient with leukemia in phase of deep bone marrow aplasia, whose outcome was favorable after exiting of aplasia period and a treatment by amphotericine B. The case of pseudotumoral cutaneous fusariosis to F. oxysporum complicated with osteolysis and septic arthritis occurred in a pregnant woman without any immune deficit. The evolution was fatal in spite of prescription of multiple systemic antifungals. Concerning keratomycosis, Fusarium was the first agent responsible for these infections (43%). The corneal traumatism was found in 37.5% of cases and FSSC was the most isolated (72%). For superficial dermatomycosis, Fusarium was the third agent of onychomycosis in molds (25%). The most isolated species were FSSC (68%) and FOSC (20%). The intertrigo frequency was 0.07% and they were mostly caused by FSSC (84%) and FOSC (16%). Fusarium is an important cause of mold infections in our region. So, the species identification is useful because some species are resistant to the most common systemic antifungal agents.

Candida parapsilosis, which was previously considered to be a complex of three genetically distinct groups, has emerged as a significant agent of nosocomial infections. Recently, this complex was separated into three species: C. parapsilosis sensu stricto, C. orthopsilosis, and C. metapsilosis In Tunisia, data pertaining to these fungi are limited. Thus, the purpose of our study was to determine by BanI PCR-RFLP and ITS sequencing, the occurrence of Candida parapsilosis complex among 182 isolates identified as C. parapsilosis by phenotypical methods. C. parapsilosis sensu stricto represented 94.5% of all isolates, while C. metapsilosis and. C. orthopsilosis were identified in 3.3% and 2.2%, respectively. Sequence analysis of internal transcribed spacer region confirmed and revealed only one genotype among the C. parapsilosis sensu stricto strains, three genotypes among six C. metapsilosis strains and two genotypes among four C. orthopsilosis strains.

Trichophyton verrucosum is the most frequent etiologic agent of cattle dermatophytosis. Throughout the world, it was the second most common agent of zoophilic dermatophytes in human. The aim of our study was to evaluate the efficacy of the PCR- RFLP and PCR-sequencing methods for the identification and differentiation of T. verrucosum strains.Thirty-six clinical strains identified by morphological characteristics as T. verrucosum were isolated from patients referred to parasitology-mycology laboratory of Sfax University Hospital. Identification of our strains by conventional methods was confirmed by molecular methods in 94.4% of cases. Two strains were reclassified as T. violaceum PCR products digested with HinfI produced three profiles and two patterns with MvaI. Sequence analysis revealed a polymorphism in the ITS1and 5.8S regions. Analysis and alignment of consensus sequences has distinguished two types of genotypes among our T. verrucosum strains. The ITS type I was the dominant genotype (93.7%). Phylogenetic study showed that one cluster comprised T. verrucosum strains with ITS type I and species of T. mentagrophytes complex. It was related to Arthroderma vanbreuseghemii complex. The other cluster contained the two T. verrucosum strains with ITS type II, and was related to Arthroderma benhamiae complex. In this study, most of T. verrucosum isolates were type I, dissimilar to others rare studies where type II has been the most common. Specie and strain differentiation is relevant because it helps in prescribing the correct treatment and determining the source of the infection.

Free-living amoebae are widely distributed in soil and water. Small number of them was implicated in human disease: Acanthamoeba spp., Naegleria fowleri, Balamuthia mandrillaris and Sappinia diploidea. Some of the infections were opportunistic, occurring mainly in immunocompromised hosts (Acanthamoeba and Balamuthia encephalitis) while others are non opportunistic (Acanthamoeba keratitis, Naegleria meningoencephalitis and some cases of Balamuthia encephalitis). Although, the number of infections caused by these amoebae is low, their diagnosis was still difficult to confirm and so there was a higher mortality, particularly, associated with encephalitis. In this review, we present some information about epidemiology, ecology and the types of diseases caused by these pathogens amoebae.

Candida lipolytica (actuellement connu sous le nom de Yarrowia lipolytica ) est une levure emergente omnipresente dans l’environnement. Elle peut etre responsable d’infections invasives nosocomiales humaines, en particulier les septicemies associees aux catheters, chez les patients a risque. Les objectifs de notre travail ont ete d’analyser les conditions favorisant la survenue des septicemies a Y. lipolytica , de typer les souches de Y. lipolytica par PCR-sequencage et PCR-RFLP et d’etudier la sensibilite in vitro de cette levure aux antifongiques. Patients et methodes Il s’agit d’une etude retrospective rapportant les cas d’infections a Y. lipolytica diagnostiques au laboratoire de parasitologie-mycologie du CHU Habib Bourguiba de Sfax durant une periode de 23 mois (octobre 2012–aout 2014). Nous avons realise une etude moleculaire pour 26 souches : PCR-RFLP et PCR-sequencage. Nous avons amplifie les regions ITS a l’aide de deux amorces universelles ITS1 et ITS4. Les enzymes utilisees etaient AfaI et DraI. Pour l’etude de la sensibilite aux antifongiques, nous avons utilise la methode du Sensititre YeastOne et l’E-test. Resultats Nous avons collige 64 patients ayant eu une infection a Y. lipolytica . Il s’agissait de 63 cas de septicemies a Y. lipolytica dont 98,4 % provenaient du service de reanimation. L’antibiotherapie a large spectre (100 %), le catheterisme vasculaire (93,6 %) et le sejour prolonge en reanimation (92 %) etaient les principaux facteurs de risques associes a ces septicemies. La fievre prolongee resistante a une antibiotherapie a large spectre etait le signe clinique le plus frequemment retrouve (85,7 %). Le traitement des septicemies a Y. lipolytica a repose dans la majorite des cas sur le fluconazole en association avec l’amphotericine B et/ou l’ablation du catheter vasculaire. Apres amplification par PCR, nous avons obtenu une bande de 358 pb pour les 26 souches. La digestion enzymatique de ces souches a genere deux bandes de 142 pb et 215 pb pour l’enzyme AfaI et de 79 pb et 278 pb pour l’enzyme DraI. Le sequencage de 15 souches a donne un seul type de sequence avec un pourcentage de similarite de 99 % avec les sequences de Y. lipolytica deposees dans la banque de donnees. L’etude de la sensibilite aux antifongiques a montre que Y. lipolytica etait sensible au fluconazole dans 47,6 % des cas, a l’amphotericine B dans 95,2 % des cas, a la caspofungine dans 22,2 % des cas et au voriconazole dans 92 % des cas. Par contre, nos souches etaient resistantes au flucytosine dans 77,7 % des cas et a l’anidulafungine dans 100 % des cas. Conclusion Y. lipolytica est un pathogene emergent responsable de septicemies liees aux catheters. L’etude epidemiologique est un outil essentiel qui nous a permis de connaitre le profil epidemiologique de cette levure et les conditions favorisant la survenue des septicemies a Y. lipolytica dans nos hopitaux. L’etude moleculaire a permis de resoudre rapidement les difficultes d’identification morphologique. L’etude de la sensibilite aux antifongiques a permis de connaitre le profil de sensibilite de nos souches.

Resume Trichophyton verrucosum est un dermatophyte zoophile responsable de divers aspects cliniques, notamment des lesions inflammatoires agressives de la peau et des teignes inflammatoires. Le but de cette etude a ete de preciser le profil clinique, epidemiologique des dermatophyties a T. verrucosum . Patients et methodes Etude retrospective portant sur 35 918 patients suspects de dermatomycoses colliges dans notre laboratoire durant une periode de 13 ans allant de 1998 a 2010. Resultats T. verrucosum a ete isole chez 178 patients soit 1,2 % des cas de dermatophyties. La variete ochraceum etait la plus frequente (60 %), suivie par la variete album (40 %). La frequence d’isolement de T. verrucosum a augmente, en passant d’un cas en 1998 a 37 cas en 2010. L’âge moyen etait 22,7 ans (extremes : 1,5–71 ans). Un pourcentage de 74,2 de nos patients ont ete de sexe masculin et 61 % de provenance urbaine. La notion de contact avec les animaux a ete notee dans 32,5 % des cas, essentiellement des bovins et des ovins. Dans 7,3 % des cas, les malades avaient des antecedents familiaux de dermatophytie. Les epidermophyties circinees etaient les plus frequentes (62,2 %), suivies par les teignes du cuir chevelu (31,6 %). Dix cas de sycosis, un cas de mycose de la plante des pieds et un cas d’atteinte pubienne ont ete diagnostiques. Les lesions de la peau glabre etaient erythemato-squameuses (82,5 %) et pustulo-inflammatoires (17,5 %), touchant surtout les membres superieurs (58,3 %) et la face (25,8 %). Les lesions de teigne etaient squameuses (30,8 %) et pustulo-inflammatoires (41 %). Onze cas de kerion de Celse ont ete diagnostiques (28,2 %). T. verrucosum etait isole a partir d’un seul site (79,7 %), deux sites (16,9 %) et trois sites ou plus (3,4 %). Conclusion Les dermatophyties a T. verrucosum sont en recrudescence dans notre region, cela s’expliquerait certainement par une modification du comportement de la population urbaine a niveau socioeconomique bas, qui se livre a une activite complementaire d’elevage.

Resume Objectif Le but de ce travail a ete d’etudier aussi bien les aspects epidemiologiques que mycologiques de la flore fongique a levures au CHU de Sfax, Tunisie. Materiel et methodes Il s’agit d’une etude retrospective portant sur les infections superficielles et profondes a levures diagnostiquees entre 1993 et 2006. Resultats Nous avons collige 9712 cas de candidoses. C. albicans etait l’espece majoritaire (51,9 %), suivie par C. tropicalis (19,8 %) et C. glabrata (17 %). Elle etait l’espece majoritaire au niveau : buccal (78,3 %), vulvovaginal (61,8 %), perionyxis (58,5 %), urines (41,5 %) et auriculaire (31,3 %). Mais elle occupait la deuxieme place dans les candidemies (29,6 %). C. tropicalis etait la premiere espece isolee au niveau des hemocultures (36,7 %) et elle a occupe le deuxieme rang dans les autres candidoses superficielles a part les vulvovaginites ou elle a cede la place a C. glabrata (26,2 %). Cette derniere a ete isolee dans 17,6 % des hemocultures et elle a ete a l’origine d’abces profonds dans trois cas. C. parapsilosis etait isolee essentiellement au niveau des hemocultures (11,5 %), au niveau auriculaire (26,8 %) et ungueal (10,8 %). Les levures du genre Malassezia ont ete diagnostiquees dans 2326 cas. D’autres levures ont ete moins frequemment isolees : Cryptococcus neoformans (sept cas), Saccharomyces cerevisiae (38 cas) et Trichosporon (55 cas). Conclusion Une surveillance continue de la distribution des levures et l’analyse de la signification clinique de ces changements sont justifiees.

L'objectif de notre etude a ete de detecter les amibes libres par des methodes morphologiques et d'identifier Acanthamoeba spp. par PCR dans l'eau des unites dentaires. Materiel et methodes: il s'agit d'une etude prospective portant sur 196 prelevements d'eau a partir des circuits dentaires (94 echantillons realises avant purge et consultation des malades et 102 echantillons apres consultation des malades) et 39 echantillons d'eau de robinet. Resultats: l'etude morphologique a permis de detecter les amibes libres dans 69 % des echantillons. Le morphotype predominant etait le monopodial (39,2 %). 18 souches d'Acanthamoeba ont ete detectees dans l'eau du circuit dentaire (13,3 %) et trois souches dans l'eau de robinet (10 %). L'amplification du gene 18S de l'ADN ribosomal de ces souches d'Acanthamoeba spp. a ete positive dans 100 % des cas. Conclusion: les amibes libres, dont Acanthamoeba, ont ete isolees aussi bien dans l'eau de robinet que dans les unites dentaires. Des cas d'infections amibiennes directement attribuables aux soins dentaires n'ont pas ete demontres, cependant, la presence de bacteries intracellulaires ou sein des amibes libres doit etre consideree comme un risque potentiel surtout chez les patients immunodeprimes. L'assainissement de ces unites s'impose au moins par la mise en place d'un filtre de porosite 0,2 microns avant l'etape d'entree de l'eau dans les circuits dentaires afin de limiter l'eventuel passage des amibes libres.

Rhodotorula is emerging as an important cause of nosocomial and opportunistic infections. We present two cases of Rhodotorula mucilaginosa fungemia diagnosed at our hospital during the last decade. The first case was of a term neonate who presented congenital heart disease (interventricular communication) and body dysmorphic disorder. He was admitted for respiratory failure and sepsis. The second case involved in a 33-year-old woman that had Hodgkinien lymphoma associated to tuberculosis. Identification was performed using commercial systems and confirmed by PCR sequencing of internal transcribed spacer, ITS1 and ITS2 regions of rDNA. Antifungal susceptibility tested by sensititre yeast revealed susceptibility to amphotericin B and resistance to fluconazole for the two strains. These cases emphasize the emerging importance of Rhodotorula sp. as a pathogen and it must be considered a potential pathogen in patients with immunosupression and with central venous catheters. Correct identification is mandatory for appropriate management, as Rhodotorula spp. are resistant to antifungal agents, such as fluconazole.

Les infections fongiques ont connu ces dernieres decennies une incidence croissante. Aspergillus fumigatus est un champignon filamenteux, saprophyte et sporulant. Il est responsable d’infections invasives chez les immunodeprimes. Sa virulence est multifactorielle. L’activite enzymatique et la capacite de former un biofilm des souches d’ A. fumigatus isolees chez 18 patients asthmatiques hospitalises a l’hopital Hedi Chaker de Sfax ont ete testees in vitro sur des milieux specifiques appropries afin de comparer la virulence de 22 souches collectees a partir des prelevements cliniques et 18 souches a partir des prelevements environnementaux. La production des hydrolases extracellulaires a montre que nos souches etaient productrices de phospholipases (67,5 %), de proteases (77,5 %), d’hemolysines (92,5 %) et d’elastases (75 %). Une production significativement plus elevee d’hydrolases extracellulaires a ete notee chez les souches cliniques ( p p A. fumigatus . L’etude de la formation de biofilm sur des plaques en polystyrene a montre que tous les isolats etaient aptes d’adherer a la surface de ce dispositif. Cependant, les souches cliniques ont montre une production plus elevee de biofilm ( p En conclusion, notre etude nous a permis de montrer que les souches cliniques ont une production plus elevees d’hydrolases extracellulaires et de biofilm. Ces facteurs de virulence expliquent bien le pouvoir pathogene d’ A. fumigatus dans le developpement des infections invasives.

Les levures du genre Malassezia sont des levures d’actualite ayant connu, dans cette derniere decennie, une evolution taxonomique importante et un interet accru en pathologie humaine et animale. L’identification de ces levures peut se faire par les techniques phenotypiques, biochimiques et physiologiques qui restent laborieuses et parfois imprecises vu que certaines especes presentent des caracteristiques tres proches. Nous avons mis au point une PCR multiplex en temps reel avec des sondes specifiques pour detecter les six especes de Malassezia : M. globosa, M. furfur, M. restricta, M. sympodialis, M. slooffiae et M. pachydermatis. Notre etude prospective a ete realisee sur 120 echantillons collectes a partir de 60 animaux domestiques (23 chevres, 10 chiens, 15 vaches, 3 chats, 8 lapins et 1 âne) et de 70 echantillons humains (28 patients atteints de pityriasis versicolor, 17 eleveurs et 25 personnes representant un groupe temoin). Quinze M. pachydermatis ont ete identifies chez les animaux. Nous avons identifie 61 isolats a partir des prelevements humains : M. globosa (28), M. furfur (15), M. restricta (6), M. sympodialis (8), M. slooffiae (2) et M. pachydermatis (2). La PCR en temps reel nous a permis aussi de detecter huit cas de co-infection chez 6 patients et 2 eleveurs. L’amplification a ete specifique pour chacune des especes de Malassezia : M. globosa, M. pachydermatis, M. sympodialis, M. furfur, M. slooffiae et M. restricta. En outre, aucun signal positif n’a ete retrouve par la PCR en temps reel pour les souches n’appartenant pas au genre Malassezia. Nos resultats montrent que l’analyse etait tres efficace pour identifier les especes de Malassezia. L’application de PCR multiplex en temps reel fournit un systeme d’identification sensible et rapide pour les especes de Malassezia, qui peut etre applique dans les enquetes epidemiologiques et la pratique courante.

Trichophyton interdigitale is the second most frequent cause of superficial fungal infections of various parts of the human body. Studying the population structure and genotype differentiation of T. interdigitale strains may lead to significant improvements in clinical practice. The present study aimed to develop and select suitable variable-number tandem-repeat (VNTR) markers for 92 clinical strains of T. interdigitale . On the basis of an analysis of four VNTR markers, four to eight distinct alleles were detected for each marker. The marker with the highest discriminatory power had eight alleles and a D value of 0.802. The combination of all four markers yielded a D value of 0.969 with 29 distinct multilocus genotypes. VNTR typing revealed the genetic diversity of the strains, identifying three populations according to their colonization sites. A correlation between phenotypic characteristics and multilocus genotypes was observed. Seven patients harbored T. interdigitale strains with different genotypes. Typing of clinical T. interdigitale samples by VNTR markers displayed excellent discriminatory power and 100% reproducibility.

Geotrichum capitatum is an uncommon cause of invasive infections in immunocompromised patients, particularly those with hematological malignancies and severe neutropenia. The aim of this study was to report the cases of invasive geotrichosis in our hospital. It is a retrospective study of invasive geotrichosis diagnosed in the Laboratory of Parasitology–Mycology of the UH Habib Bourguiba, Sfax, from January 2005 to August 2013. Six cases of invasive Geotrichum infections were diagnosed. There were three men and three women. The mean age was 35 years. Five patients have acute myeloid leukemia with a profound neutropenia, and one patient was hospitalized in the intensive care unit for polytraumatism. Clinically, the prolonged fever associated with pulmonary symptoms was the predominant symptom (n = 5). Geotrichum capitatum was isolated in one or more blood culture. Two patients had urinary tract infections documented by multiple urine cultures positive for G. capitatum. Five patients received conventional amphotericin B alone or associated with voriconazole. The outcome was fatal in four cases. Invasive geotrichosis is rare, but particularly fatal in immunocompromised patients. Approximately, 186 cases have been reported in the literature. The prognostic is poor with mortality over 50 %. So, early diagnosis and appropriate management are necessary to improve prognosis.

Resume Les Trichosporon sont des levures de plus en plus incriminees dans la survenue d’infections profondes, notamment chez les patients d’oncohematologie. Nous rapportons deux observations de septicemie a Trichosporon asahii diagnostiquees au laboratoire de parasitologie et mycologie du CHU Habib-Bourguiba de Sfax, Tunisie. Notre premier patient presentait une leucemie aigue myeloide avec une fievre persistante malgre la mise en place d’une antibiotherapie a large spectre. Le second cas avait une anemie avec une fievre resistante aux antibiotiques. T. asahii a ete isole des hemocultures dans les deux cas. Les patients ont ete mis sous amphotericine B mais l’evolution etait fatale. Les trichosporonoses sont rares mais particulierement graves chez les sujets immunodeprimes avec un pronostic souvent pejoratif malgre le traitement antifongique.

Fungal keratitis is responsible for a significant burden of blinding disease in the developing world. Objective The aim of this study was to determine the etiological agents, predisposing factors and therapy of keratomycosis in our region. Methods Retrospective study of 60 patients with clinically and cultured confirmed fungi keratitis, who were attended at department of mycology in Sfax (1995 to 2012). Results The mean age of patients was 47.2 years (sex ratio: 1.58). At least, one presumed predisposing factor was identified in 83.3% of cases. Corneal traumatism was established as the most common predisposing factor (61.6%) with vegetative matter (42.4%). Patients had corneal ulcer in 40% or abscess in 47.6%. All cases were positive on direct microscopy and 93% of cultures were positive. Filamentous fungi form the major etiologic agents (83%): Fusarium species (49% with F. solani [66%]), Aspergillus sp. (22%), Alternaria (5%), Scedosporium sp. (2%); and non-identified mold in (5%). Yeast were identified in 17% of cases. Topical agents were used in 97% of cases: ketoconazole 2%, amphotericin B (0.5%). Fluconazole per os was administrated for 11% of cases, itraconazole (2 cases) and voriconazole (one case). Keratoplasty was indicated for 27% of cases. The outcome was favorable in 16% of patients. Among the patients, 71% had persistent corneal deposit sequelae. Four patients lost the eyeball. Conclusion Corneal traumatism was the principal risk factor for fungal keratitis in young and middle-aged farmers. Fusarium solani is the predominant cause in Sfax. Early diagnosis, coupled with appropriate treatment, is crucial for increasing the chance of complete recovery.

In recent years, an increase of systemic Candida infections was noted. Thus, identification and susceptibility testing to antifungals became of considerable importance. The technique of dilution in liquid medium developed by « National committee for clinical laboratory standards » NCCLS or more recently named CLSI « Clinical and laboratory standards institute » is the reference method most used. The European committee "European committee on antibiotic susceptibility testing" or EUCAST has made progress by determining the susceptibility of strains within a shorter time. However, the use of these techniques is limited especially in clinical microbiology laboratories. Other techniques for determining antifungal sensitivity have been developed such as those based on agar diffusion (E-test and disk diffusion), on microdilution (Sensititre yeastOne, Vitek 2 AST-YS01), on flow cytometry techniques and the MALDI-TOF.

Although the arsenal of agents with anti-Aspergillus activity has expanded over the last decade, mortality due to invasive aspergillosis remains unacceptably high. Resistance of the Aspergillus spp. species to antifungal drugs increased in the last 20 years with the increase in antifungal drugs use and might partially account for treatment failures. Recent advances in our understanding of mechanisms of antifungal drug action in Aspergillus, along with the standardization of in vitro susceptibility testing methods, have brought resistance testing to the forefront of clinical mycology. Recent modifications in taxonomy and understanding of the acquired resistance mechanisms of Aspergilli to drugs should support a better management of Aspergillus infections. In this paper, we review the current knowledge on epidemiology and underlying mechanisms involved in antifungal resistance in Aspergillus.

Invasive aspergillosis (IA) is a major cause of morbidity and mortality in profoundly neutropenic patients. Delayed diagnosis and therapy may lead to poor outcomes. Aims The objective of this study was to assess the performance characteristics of the galactomannan (GM) assay in serum and bronchoalveolar lavage specimens for the diagnosis of IA in neutropenic patients with hematological malignancies. We also evaluated the prognostic outcome. Patients and methods A total of 1198 serum samples and 42 BAL from 235 neutropenic patients were tested with a GM elisa platelia test. We used Cox modeling of time to 6- and 12-week mortality for GM level at the time of diagnosis (GM0) and GM decay in the week following diagnosis in proven and probable IA patients with more than two GM values. Results There were three proven, 55 probable, and four possible cases of IA. The sensitivity and specificity of the GM test were 96.8% and 82.4% respectively. In BAL samples, sensitivity was 86% and the specificity 93%. BAL GM was more sensitive than microscopy (22.2%) and BAL culture (38.9%). Among patients with proven/probable IA, serum and BAL GM were in agreement for 92.8% of paired samples. The hazard ratio (HR) of GM0 and 1-week GM decay per unit increase in Aspergillus enzyme immunoassay (EIA) was 1.044 (95% CI, 0.738 to 1.476) and 0.709 (95% CI, 0.236 to 2.130) respectively. Conclusion We found good correlation between the GM0 and GM decay combination and outcome of IA patients. The GM is a useful tool for diagnosis and monitoring of IA.