New evidence for the potential role of Culex pipiens mosquitoes in the transmission cycle of West Nile virus in Tunisia.

Physiological and molecular characteristics of natural populations of Culex pipiens Linnaeus, 1758 (Diptera: Culicidae) were investigated to elucidate how this species is potentially involved in the transmission of West Nile virus in Tunisia. A total of 215 Cx. pipiens females from 11 breeding habitats were analysed in the laboratory to estimate autogeny and stenogamy rates. They were tested individually for the locus CQ11 to distinguish between the two Cx. pipiens forms, pipiens and molestus. All tested Cx. pipiens populations were stenogamous. Females from underground breeding sites were all autogeneous, whereas females from above-ground habitats were mostly anautogeneous. Of all the females tested, 59.7% were identified as pipiens, 22.4% as molestus, and 17.9% as hybrid pipiens/molestus. Furthermore, both Cx. pipiens forms and their hybrids were found to co-occur in sympatry in all sites. The results of this study represent the first evidence that both Cx. pipiens forms and their hybrids are present in Tunisia. Because hybrids able to act as bridge vectors are present in all studied habitats, Tunisia can be considered to have a high degree of receptivity for the establishment of West Nile virus zoonotic cycles.

Early dental epithelial transcription factors distinguish ameloblastoma from keratocystic odontogenic tumor.

The aim of the study was to characterize the molecular relationship between ameloblastoma and keratocystic odontogenic tumor (KCOT) by means of a genome-wide expression analysis. Total RNA from 27 fresh tumor samples of 15 solid/multicystic intraosseous ameloblastomas and 12 sporadic KCOTs was hybridized on Affymetrix whole genome arrays. Hierarchical clustering separated ameloblastomas and KCOTs into 2 distinct groups. The gene set enrichment analysis based on 303 dental genes showed a similar separation of ameloblastomas and KCOTs. Early dental epithelial markers PITX2, MSX2, DLX2, RUNX1, and ISL1 were differentially overexpressed in ameloblastoma, indicating its dental identity. Also, PTHLH, a hormone involved in tooth eruption and invasive growth, was one of the most differentially upregulated genes in ameloblastoma. The most differentially overexpressed genes in KCOT were squamous epithelial differentiation markers SPRR1A, KRTDAP, and KRT4, as well as DSG1, a component of desmosomal cell-cell junctions. Additonally, the epithelial stem cell marker SOX2 was significantly upregulated in KCOT when compared with ameloblastoma. Taken together, the gene expression profile of ameloblastoma reflects differentiation from dental lamina toward the cap/bell stage of tooth development, as indicated by dental epithelium-specific transcription factors. In contrast, gene expression of KCOT indicates differentiation toward keratinocytes.

[Ameloblastoma of the jaws. A retrospective analysis from 1994 to 2007]. / Améloblastomes des mâchoires. Analyse rétrospective de 1994 à 2007.

INTRODUCTION: Ameloblastomas and keratocysts are the most frequent epithelial odontogenic tumors of the jaws. They have a high recurrence rate. This retrospective study reviews the features of ameloblastomas operated on in our unit from 1994 to 2007. PATIENTS AND METHODS: The studied parameters were sex, ethnic origin, age at diagnosis, clinical signs, radiographic presentation, site distribution, histological type, treatment, and follow-up records. RESULTS: One hundred and sixteen patients were included (with 239 surgical samples). The mean age was 36 years, with a majority of Europeans, 60% of multilocular radiolucent lesions with root resorption, mandibular location (93%). Twenty-one percent of the patients presented with an impacted tooth, the third molar in 79% of cases. Fifty percent of the lesions were from 5 to 13cm in length, 10% longer than 13cm. The most common histological type was follicular ameloblastoma. Patients were treated by enucleation in 82% of cases and radical mandibular resection with reconstruction in 11% of cases. The follow-up was documented for 96% of the patients with a 44% recurrence rate. Seventy-four percent of patients with a double recurrence presented with a "follicular" ameloblastoma. DISCUSSION: We prefer a well-performed enucleation which preserves surrounding bone. The high rate of follicular type recurrence should more systematically lead to a combined treatment: periostectomy and tooth extraction. Our data was compared with previously published large series.

First detection of Babesia occultans in Hyalomma ticks from Tunisia.

Descriptions of Babesia occultans have previously been restricted to sub-Saharan Africa. Here, we report the finding, for the first time, of this low or non-pathogenic bovine Babesia species in Tunisia, northern Africa. B. occultans DNA was detected by molecular methods in Hyalomma marginatum unfed ticks collected in 3 bioclimatic regions of Tunisia. The near-full-length 18S rRNA gene was sequenced and compared with related sequences retrieved from GenBank. Phylogenetic analysis indicated that other sequences deposited as Babesia sp. could also correspond to B. occultans, suggesting that this species may have a wide distribution in Mediterranean and Asiatic regions, and not only in sub-Saharan Africa as previously described. A B. occultans-specific Reverse Line Blot (RLB) oligonucleotide probe was designed for future epidemiological studies that would help to clarify this possibility.

[Role of Toscana virus in meningo-encephalitis in Tunisia]. / Rôle du virus Toscana dans les infections neuroméningées en Tunisie.

OBJECTIVES: To detect the presence of Toscana virus (TOSV) circulation in Tunisia and to study its role in viral meningo-encephalitis. PATIENTS AND METHODS: A total of 315 (167 sera and 178 cerobrospinal fluid [CSF]) samples was investigated. These samples are colleted from Tunisian patients with neurological diseases during the period between January 2003 and December 2009. All samples were tested negative for enterovirus, Herpes Simplex virus and West Nile virus. Detection for IgM and IgG specific to TOSV was done by ELISA tests. RESULTS: Specific IgM for TOSV were detected in 10 % of patients with neurological diseases (31 cases). These recent infections were distributed throughout the study period and predominated during summer and automn. Patients were originated, in the majority from the coastal region. IgG were isolated in 22 cases (7 %) corresponding to previous infection. CONCLUSION: This is the first report of TOSV circulating in Tunisia and its frequent implication in neurological diseases. These results incited to include TOSV as one of the viral etiologies to target in the diagnosis of viral meningitis and encephalitis in the country.

[Assessment of the risk of introduction to Tunisia of the Rift Valley fever virus by the mosquito Culex pipiens]. / Estimation du risque d'introduction du virus de la fièvre de la vallée du Rift en Tunisie par le moustique Culex pipiens.

The mosquito Culex pipiens has been involved as vector of the West Nile virus in Tunisia. Its bio-ecological characteristics in combination with some environmental factors have favoured the emergence of this virus in a West-Nile free zone. This leads to question about the potential risk of introducing another arbovirus, the Rift Valley fever (RVF) virus, in Tunisia from neighbouring countries where RVF circulates. In this study, we have evaluated the vector competence of different populations of Cx. pipiens towards two strains of RVF virus, the virulent ZH548 and the avirulent Clone 13 by experimental infections and the genetic differentiation of these populations of Cx. pipiens using four microsatellite loci. We found disseminated infection rates ranging from 0% to 14.7% and a high genetic differentiation among populations without any geographical pattern (no isolation by distance). Thus, although Cx. pipiens is able to sustain an amplification of RVF virus, viral dissemination through mosquito dispersal would be unlikely. However, as RVF is an emerging disease transmitted by several other potential mosquito species (e.g. Ochlerotatus caspius), attention should be maintained to survey livestock and mosquitoes in Tunisia.

Quatre premiers cas de meningites a Toscana Virus dans la Region de Monastir

Les meningites a Toscana Virus ont ete largement decrites en Italie ou le virus a ete isole pour la premiere fois. Actuellement; elles sont de plus en plus decrites dans le pourtour du bassin mediterraneen devenant ainsi une pathologie emergente dans cette region. Nous rapportons; dans ce travail; 4 cas de meningites a Toscana Virus hospitalises dans le service des Maladies Infectieuses de l'hopital Fattouma Bourguiba de Monastir - Tunisie; en rappelant les caracteristiques epidemiologiques; cliniques; biologiques et evolutives. Tous les malades etaient de sexe masculin; demeurant dans une region cotiere et hospitalises pendant la saison estivale. Ils etaient ages en moyenne de 26 ans (14 - 41). Le diagnostic clinique reposait sur la presence d'un syndrome meninge febrile. La ponction lombaire avait montre dans les 4 cas une meningite lymphocytaire a liquide clair avec une glycorrachie normale. La detection des IgM et des IgG dans le sang et le LCR avait permis de confirmer le diagnostic. L'evolution etait favorable sous traitement symptomatique; dans tous les cas; avec un recul de 9 mois

Theileria and Babesia parasites in ticks in Tunisia.

A total 576 adult ixodid ticks, collected in seven sites in Northern Tunisia, were analysed for piroplasm infection. Reverse line blot analysis identified nine different piroplasms, with an overall prevalence of 22.7%. 17.8% were Theileria spp. positive, 8.5% were positive for Babesia spp. and 3.6% of ticks harboured both genera. Although, most of the positive ticks harboured a single infection (17%), mixed infections with two, three or four different piroplasm species were also detected (5.7%).

Clinical, serological, and molecular evidence of ehrlichiosis and anaplasmosis in dogs in Tunisia.

A seroepidemiological survey was conducted in five bioclimatic areas of Tunisia to determine the prevalence of antibodies to Ehrlichia canis and Anaplasma phagocytophilum antigens, surrogate markers of the agents of canine monocytic ehrlichiosis (CME) and canine granulocytic ehrlichiosis, respectively. Among 286 collected sera, 54.2% and 25.2% were seropositive for E. canis and A. phagocytophilum, respectively, by the indirect immunofluorescence antibody (IFA) test. Clinical and hematological tests were done only for 58 sick dogs from Tunis area. A reverse line blot (RLB) hybridization was then used to identify isolated Ehrlichia and Anaplasma species infecting dogs (n = 228). Among them, only two dogs were infected by A. phagocytophilum; ten sample dogs were demonstrated infected by E. canis and ten infected by Ehrlichia sp., from which one dog showed a mixed infection with A. phagocytophilum and E. canis and one with A. phagocytophilum and Ehrlichia sp. RLB findings were confirmed by sequencing; BLAST search against GenBank revealed high similarity of the sequence of Ehrlichia sp. PCR/RLB amplicons with Anaplasma platys 16S rRNA partial sequence.

A molecular survey of Theileria and Babesia parasites in cattle, with a note on the distribution of ticks in Tunisia.

Between October and November 2006, a total of 278 bovine blood samples were examined, and 104 (37.4%) were positive for piroplasms by microscopy. A reverse line blot hybridisation with polymerase chain reaction detected Theileria annulata, T. buffeli, Babesia bovis and B. bigemina in cattle accounting for 48.6% of positive samples. The most frequently found species was T. buffeli, which was present in 39.2% of the samples. T. annulata was found in 48 samples (17.3%). Babesia infections were less frequently detected: B. bovis was found in 6.8% of the samples and B. bigemina in 4.3%. Mixed infections were detected in 45 samples, accounting for seven different combinations of species. Seven Ixodid tick species (Boophilus annulatus, Ixodes ricinus, Hyalomma marginatum, Hyalomma excavatum, Hyalomma detritum, Haemaphysalis punctata and Haemaphysalis sulcata) were collected from examined cattle in the 23 visited farms. I. ricinus was the dominant species (36%), mainly collected in the humid zone, while it seemed to be very rare in the semi-arid zone (where only 15 specimens were collected), whereas B. annulatus was the most commonly collected species in the sub-humid area (68.5% of ticks collected in this zone).

Detection and molecular characterization of Babesia canis vogeli from naturally infected dogs and Rhipicephalus sanguineus ticks in Tunisia.

Canine babesiosis, caused by intra-erythrocytic Babesia, is a tick-borne disease of worldwide importance. No information on canine babesiosis has been documented in Tunisia. Detection and analysis of Babesia species from naturally infected dogs and ticks recovered from dogs were attempted by reverse line blot hybridization and nucleotide sequence analysis based on 18S rRNA gene. Out of 180 blood samples collected from domestic dogs in 4 villages situated in different bioclimatic zones, 12 were positive for Babesia canis vogeli. In addition, a total of 160 Rhipicephalus sanguineus were analysed; only one male was infected by B. canis vogeli. This is the first report on the detection of DNA belonging to B. canis vogeli in domestic dogs and in R. sanguineus in Tunisia.

Widespread distribution and high prevalence of an alpha-proteobacterial symbiont in the tick Ixodes ricinus.

The tick Ixodes ricinus is responsible for the transmission of a number of bacterial, protozoan and viral diseases to humans and animals in Europe and Northern Africa. Female I. ricinus from England, Switzerland and Italy have been found to harbour an intracellular alpha-proteobacterium, designated IricES1, within the cells of the ovary. IricES1 is the only prokaryote known to exist within the mitochondria of any animal or multicellular organism. To further examine the distribution, prevalence and mode of transmission of IricES1, we performed polymerase chain reaction screening of I. ricinus adults from 12 countries across its geographic distribution, including tick colonies that have been maintained in the laboratory for varying periods of time. IricES1 was detected in 100% of field-collected female ticks from all countries examined (n = 128), while 44% of males were found to be infected (n = 108). Those males that are infected appear to harbour fewer bacteria than females. Sequencing of fragments of the 16S rRNA and gyrB genes revealed very low nucleotide diversity among various populations of IricES1. Transmission of IricES1 from engorged adult females to eggs was found to be 100% (n = 31). In tick colonies that had been maintained in the laboratory for several years, a relatively low prevalence was found in females (32%; n = 25). To our knowledge, IricES1 is the most widespread and highly prevalent of any tick-associated symbiont.

The known distribution and ecological preferences of the tick subgenus Boophilus (Acari: Ixodidae) in Africa and Latin America.

A compilation of the known distribution of Boophilus ticks in Africa and Latin America is presented, together with details on climate preferences. B. annulatus is recorded mainly in the western part of a strip from the equator to parallel 20 degrees N. It associates with woodlands and forests (lowland rain forest and secondary grassland). This species is also present in the Mediterranean region, associated to woodland and open areas. B. decoloratus extends southern to parallel 20 degrees N, in woodland with montane vegetation and Zambezian miombo; some records have been collected in the highveld grassland. B. geigyi is mainly collected in the western range of a stripe extending between parallels 5 degrees N and 18 degrees N, associated with Sudanian woodland, lowland rain forest with secondary grassland and woodland. Confirmed records of microplus in Africa are restricted to Malagasy region and south and eastern Africa, being predominant in the Zambezian miombo, deciduous forest with secondary grassland, and woodland. In Latin America, microplus is abundant in the Mesoamerican corridor to Venezuela and Colombia, and southern in Brazil and Argentina. The tick is mainly associated to the biomes of Chaco and Pampas in Argentina, the North-central moist Andes, the Atlantic forest (southern range) and the moist Meso-American vegetation (northern range). Most collections of B. annulatus and B. geigyi came from areas where winter minimum temperature is above 15 degrees C, maximum temperatures remain between 33 and 36 degrees C and maximum rainfall is recorded between June and September. B. decoloratus and African B. microplus are recorded in sites with low temperatures in May-September. Minimum temperature requirements are similar for both B. decoloratus and African B. microplus, and both are around 4 degrees C less than the value recorded for collections of Latin-American B. microplus. The rainfall pattern observed for decoloratus shows a minimum in May and June. The requirements of total rainfall are highest for B. microplus in Latin America, while records of African B. microplus are concentrated in areas of low rainfall between May and October, and high rainfall between November and March (low rainfall in the same period for B. decoloratus). Statistical analysis revealed the existence of populations (demes) with ecologically different requirements within each tick species. Both B. annulatus and B. decoloratus showed many different demes clearly associated to defined areas. The collections of Latin American B. microplus are very homogeneous according climate preferences and well separated from the African counterpart.

[Lyme borreliosis situation in North Africa]. / Situation de la borreliose de Lyme au Maghreb.

Many epidemiological studies were conducted for studying Lyme borreliosis (LB) which represents a new global public health problem. It is now the most common vector-borne disease in Europe and North America. The causative agent Borrelia burgdorferi sl is a bacterial species complex comprising 12 delineated and named species. In North Africa, few studies based on clinical and serological features, have suggested that LB could occur. Indeed, recent studies conducted in Tunisia, Algeria and Morocco have showm that Ixodes ricinus is present in cooler and humid area of these regions. These studies also revealed that this species is a vector of B. burgdorferi sl with high prevalence of infection. Using IFI and PCR tests, the mean rate of Borrelia-infection ranged from 50 to 60% in I. ricinus adult collected in Tunisia and Morocco and from 30 to 40% in nymphs; in contrast, the prevalence in larvae is less than 2.5%. Several strains of B. burgdorfer were isolated from adult and nymph I ricinus collected in Tunisia and Morocco. The identification of these strains and DNAs directly extracted from Ixodes was done by PCR-RFLP and sequence analysis. The results showed that B. lusitaniae (genotypes Poti B2 and Poti B3) is the predominant species circulating in I. ricinus in Tunisia and Morocco, B. garinii and B. burgdorferi ss and B lusitaniae were also present but very rare. These results provide the evidence for the existence of B. burgdorferi sl in North Africa; however, the impact of LB in the human population seem to be negligible and the seroprevalence of Borrelia in forest workers (considered as population at high risk) in Tunisia is less than 4%.

[Dichotomous identification keys of ticks (Acari: Ixodidae), livestock parasites in North Africa]. / Clé dichotomique et identification des tiques (Acari: Ixodidae) parasites du bétail au Maghreb.

The known fauna of livestock in the Maghreb consists of 5 established species. Five other species recorded seem to be introduced by imported animals. In this paper, dichotomous keys are given for species and subspecies of the genus Hyalomma. Rhipicephalus and Haemaphysalis parasite of livestock in North Africa. For the ticks of the other genera which are represented by only one species, we provide an identification description. Notes on geographical distribution, host and period of activity are provided.

High prevalence of Borrelia lusitaniae in Ixodes ricinus ticks in Tunisia.

To investigate whether ticks of the genus Ixodes are infected by Borrelia burgdorferi complex, 490 unfed Ixodes ricinus ticks were collected by flagging in three different areas of Tunisia in 1998. DNAs extracted from 81 adults, 60 nymphs and 38 larvae were analysed after genic amplification of the non-coding spacer between the two copies of the rrl-rrf genes of B. burgdorferi sl. The prevalence of B. burgdorferi sl. in adults, nymphs and larvae was found to be 34, 33.3 and 2.6%, respectively. All DNAs (n = 61) but one were identified as belonging to different genotypes of B. lusitaniae by analysis of the restriction fragment length polymorphism of amplification products. In addition, 290 adults, 14 nymphs and 7 larvae were used to inoculate BSK-H medium to isolate spirochetes. Fifteen strains were isolated from adult ticks in the humid areas of Tunisia, whereas only one was obtained from larvae. Isolates were identified as B. lusitaniae (15/16) and B. garinii (1/16). These results provide new evidence for the existence of Lyme borreliosis in North Africa.

Infection of Ixodes ricinus (Acari: Ixodidae) by Borrelia burgdorferi sensu lato in North Africa.

Free-living adult Ixodes ricinus L, were collected in Amdoun, situated in the Kroumiry mountains in northwestern Tunisia (North Africa). Using direct fluorescence antibody assay, the infection rate of field-collected I. ricinus by Borrelia burgdorferi sensu lato was 30.5% (n = 72). No difference in infection rate was observed between male and female ticks. Spirochetes that had been isolated from I. ricinus from Ain Drahim (Kroumiry Mountains) in 1988 were identified as Borrelia lusitaniae (formerly genospecies PotiB2). This is the first identification of a genospecies of Borrelia burgdorferi sensu lato from the continent of Africa.

Distribution and ecology of ticks (Acari: Ixodidae) infesting livestock in Tunisia: an overview of eighth years field collections.

Ticks (Ixodidae) play a significant role as vectors of pathogens of domestic animals in Tunisia. The major losses caused by ticks are related to transmission of protozoan parasites. These include agents of tropical theileriosis and babesiosis in ruminants. Since 1991, we conducted research studies on tick population of livestock in Tunisia. This overview reports a synthesis on tick distribution, their biology and their role as vectors of pathogens in domestic animals, particularly cattle. During the whole period of the study about 15,000 tick specimens were collected from different zones of the country. A total of 14 species were identified. Hyalomma detritum detritum was the most abundant and important (vector of Theileria annulata) species infesting cattle. Hyalomma dromedarii and Hyalomma impeltatum were collected on domestic ruminants in the arid and desertic zones. Hyalomma marginatum marginatum and Hyalomma anatolicum excavatum were widespread and found on livestock hosts. Ixodes ricinus, vector of Babesia divergens and Borrelia burgdorferi sl, colonises mainly the humid zone. Boophilus annulatus and Rhipicephalus bursa infesting cattle, sheep and goats were found in the sub-humid and semi-arid zones. Haemaphysalis sulcata and Hae. punctata were collected in humid and sub-humid zones on cattle and sheep. Rhipicephalus turanicus were collected in different regions, on different animal species. Rhipicephalus sanguineus, tick of dogs, were often collected on livestock. Only few specimens of Hyalomma marginatum rufipes and Hyalomma franchinii were collected.