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.

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.

[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.

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.

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.

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.

First report of Babesia divergens in Tunisia.

Babesia divergens of bovine origin was isolated, for the first time in Tunisia, from a cow of a local breed (Bos taurus) from a locality in the north-west of the country. Gerbils (Meriones unguiculatus) were used for the isolation of the parasite which was inoculated into a splenectomised Friesian calf that developed a babesiosis, thus confirming the presence of the parasite.

Cattle infestation by Hyalomma ticks and prevalence of Theileria in H. detritum species in Tunisia.

Seventy-four cattle, from three farms endemic for tropical theileriosis in the north of Tunisia, were studied for tick populations from June 1991 to June 1992. Ticks were removed from cattle twice a month in the summer and every month the rest of the year. They were identified and assessed for Theileria infection. A total of 5083 Hyalomma adult ticks were collected and the major species found was H. detritum (84.3%). The activity of this species is limited between June and August with a peak in numbers observed at the end of June and the beginning of July. Amongst the 2356 Hyalomma ticks dissected, no evidence of salivary gland infection was found in either H. m. marginatum or H. a. excavatum. However, 12.4% (277/2230) of H. d. detritum dissected ticks were infected with Theileria species and amongst these, 62% had one to two sporoblasts in their salivary glands (range 1-91). The prevalence, but not the intensity, of infection was greater in females than in male ticks, and the cases of tropical theileriosis followed the peak of infected females. This suggests that female ticks have a more important role in theileriosis transmission than male ticks. A significantly lower number of adult H. detritum were collected from calves, at their first tick season, than from adult cattle. Finally, this study showed that the infestation level of cattle by H. d. detritum and the prevalence of Theileria-infection in these cattle varied between the three farms studied.

Epidemiology of tropical theileriosis (Theileria annulata infection of cattle) in an endemic region of Tunisia: characterisation of endemicity states.

A serological survey on tropical theileriosis was conducted on a sample of 54 farms in a region within the semi-arid bioclimatic zone of Tunisia. Screening of cattle sera at a dilution of 1/160 using the indirect immunofluorescent antibody test with the schizont antigen of Theileria annulata, revealed the presence of animals with positive sera in 92.15% of the sampled farms. The exposure of calves to infection in the first season was shown to be significantly lower than in older cattle. Three endemic situations were identified based on the serological profiles of herds and the incidence and age distribution of disease cases. Endemic stability was observed in farms showing a sero-prevalence of 100% in cattle of four theileriosis seasons or more and by the incidence of the highest disease levels in cattle at their second and third theileriosis season. High endemic instability was identified on the basis of low sero-prevalence rates and the occurrence of the highest disease incidence in cattle at fourth theileriosis season or more.

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.

Tropical theileriosis in Tunisia: epidemiology and control.

In Tunisia, tropical theileriosis (Theileria annulata infection of cattle) is one of the major diseases affecting cattle in the summer. Each year about 2,500 clinical cases are recorded in the country, mainly in pure-bred animals. An attempt was made in the North of Tunisia to estimate the direct cost related to clinical cases of TT. In the endemic regions, three states of endemicity were described according to cattle age categories at highest disease risk: (i) endemic stability, (ii) low endemic instability, (iii) high endemic instability. The characterisation of these endemic situations, which are conditioned by the levels of the vector tick population and the quantitative aspects of the infection in ticks, are highly relevant for the development of a live attenuated cell line vaccine against TT targeting the cattle population at disease risk in Tunisia. A research programme was set up to develop the attenuation of four local parasite stocks. Two cell lines infected with two distinct T. annulata stocks were tested in the field on pure-bred animals of different age groups from regions with high disease incidence. The vaccination with each of the two cell lines showed a high efficacy. However significant differences in vaccine reactions rates were observed between the two attenuated cell lines emphasising the importance of achieving an optimal balance between protection and vaccine tolerance particularly in pure-bred lactating cows.

Tunis virus: a new Phlebovirus from Argas reflexus hermanni ticks in Tunisia.

A new Phlebovirus provisionally named Tunis virus has been isolated from Argas reflexus hermanni ticks parasitizing domestic pigeons. It is the first isolation of an arbovirus from Tunisia and the fourth tick-borne virus to be isolated from the Moghreb following Soldado, Essaouira and Kala Iris in Morocco. The pathogenic potential of this virus is briefly discussed according to the behaviour of its vector and previous serosurveys in the country.

[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.

[Experimental study of larval efficiency of Gambusia affinis holbrooki (GIRARD, 1859) (fish-Poecilidae)]. / Etude experimentale de l'efficacite larvivore de Gambusia affinis holbrooki (GIRARD, 1859) (Poisson-Poecilidae).

Experimental study of efficiency of Gambusia affinis holbrooki (GIRARD, 1859) (Fish-Poecilidae). Gambusia affinis is a culiciphage fish which was introduced in Tunisia in 1929 to control mosquito larvae. In the present study, we confirm, under laboratory conditions, the good efficacy of Gambusia affinis against mosquito larvae. The consumption of larvae decreases with the evolution of larvae instars and increases, for the same instar, in relation with the fish size. In fact, the size of the prey is related to the predator's size: fish belonging to the length class [20-24 mm] prefer IIIrd instar, fish from class length [25-29 mm] prefer pupae and IVth instar. The trophic test shows that G. affinis has a clear preference for Culicidae larvae, except fish belonging to the 2 length classes [35-39 mm] and [40-44 mm] for which Culicidae and Ephemeroptera have similar preference. In addition, at high temperature (30 degrees C), Gambusia affinis has a higher efficiency.

[Pollen density calender in Tunisia]. / Le calendrier pollinique de Tunis.

The pollen calendar of the region on Tunis atmosphere was established and the weekly variations of the pollen density of the 25 main taxones were studied. This calendar showed an identity reflecting the surrounding vegetation modulated by the pollen production of neighbouring regions and other far-off areas. This should allow the allergist to predict the critical periods for the patients and to advice him the specific treatment. On the other hand, it would demonstrate the usefulness, in using a series of skin tests, certain European pollens which are partially or totally absent in the atmosphere of Tunis. However, it would be desirable controlling the allergic capacity of pollens which are not tested up to now and could be found in non negligible concentrations in the atmosphere.

[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%.

[Preliminary investigation of four anopheles larvae samples susceptibility to chlorpyrifos in Tunisia]. / Investigation préliminaire de la sensibilité au chlorpyrifos de quatre échantillons larvaires d'anopheles de Tunisie.

Larvae susceptibility to chlorpyrifos is studied on four Tunisian Anopheles larvas samples: Anopheles labranchiae from Rades (South of Tunis) and Menchar (region of Beja), in the North of Tunisia, A. sergentii from Meknassy and A. multicolor from Sidi Bouzid, both in the Centre of Tunisia. The test results of larvae susceplibility indicate that the LC50 and the LC95 values are less than 0.002 and 0.02 mg l-1 respectively and their 95% confidence limits overlap. We also notice that the studied samples show the same susceptibility to chlorpyrifos. The results can be used as a base data further studies on the susceptibility of Anopheles to chemical insecticides.

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%).