Molecular based assessment of genetic diversity within Barbary fig (Opuntia ficus indica (L.) Mill.) in Tunisia

In this work, we report for the first time on the analysis of genetic diversity within a set of 36 Tunisian Opuntia ficus indica (L.) Mill. ecotypes using RAPD markers.     

Origin and genetic diversity of Tunisian grapes as revealed by microsatellite markers

Ten SSR loci, previously developed for grapevine, were analyzed to evaluate the genetic variability, cultivar relatedness, and parentage in a collection of 61 autochthonous Vitis vinifera cultivars from Tunisia.The number of alleles per locus ranged from 6 to 11, while the number of genotype patterns varied between 10 and 21. The expected heterozygosity varied between 0.621 and 0.855 and the observed heterozygosity was higher than 0.9 at 4 loci (VVMD28, VVMD5, VVIP31 and VVS2) indicating that the SSRs were highly informative.Cluster analysis using unweighted pair group method with arithmetic averaging (UPGMA) suggested 14 groups among studied cultivars and 53 grapevine denominations out of 61 were unequivocally distinguished, with all accessions showing at least one-specific combination of alleles.On the other hand, in order to overcome the existing confusion in Tunisian grapevine nomenclature, of the analyzed homonymous pairs of cultivars, only ‘Balta 2’ and ‘Balta 3’ have shown identical allelic profiles, consistent with their being the same genotype. Hence, nomenclature distinction is meaningless and only one denomination should be retained.Due to the high overall power of exclusion (Q) (greater than 99.99%) and to the absence of null alleles, the set of microsatellite loci used is appropriate to determine parentage in Tunisian grapevines beyond any reasonable doubt. The analysis of fingerprints indicated that the Tunisian grape vines have evolved through out crossing between five possible parents: Balta 1, Beldi Baddar, Beldi Rafraf, Beldi Local Rafraf and Khedhiri 3.     

Morphological variation in Astragalus hamosus L. and Coronilla scorpioides L. populations of Tunisia

Morphological variation between Astragalus hamosus and Coronillascorpioides populations was studied using local germoplasm collected in northern and central Tunisia. Twenty-one morphological traits were recorded and data were analysed using complementary statistical analysis. Considerable variation based on morphological and agronomical traits was found between populations and for both species. Pod and peduncle lengths as well as flowers number were the most discriminant variables between populations. Differences were also found for variables like seed number per pod, seed yield and dry matter production. Significant correlations were found between plant traits and the environmental parameters of the origin sites. This study revealed enough variation among populations to initiate a selection programme for pasture improvement in arid and semi-arid areas of Tunisia, where the two species are well adapted. This revised version was published online in July 2006 with corrections to the Cover Date.     

Trichomonas gallinae, in comparison to Tetratrichomonas gallinarum, induces distinctive cytopathogenic effects in tissue cultures

In the present study the interaction of three genetically different clonal cultures of Trichomonas gallinae and Tetratrichomonas gallinarum with a permanent chicken liver (LMH) and a permanent quail fibroblast (QT35) cell culture was studied. Proliferation of T. gallinae cells was associated with a disintegration of the cell monolayer. The initial lesions on the LMH monolayer consisted of a progressive accumulation of the flagellate, forming clumps attached to the monolayer. A prolonged incubation time was characterized by appearance of holes in the cell monolayer with accumulation of trichomonads at their periphery. According to the severeness of the monolayer disruption differences among three tested T. gallinae clones were noticed. Furthermore, filtrates obtained either from axenic cultures of T. gallinae or from infected cell cultures produced a cytopathogenic effect similar to the protozoal cells, on both types of cell cultures. However, the destructive effect of the flagellates and their cell-free filtrates was much more pronounced on the LMH monolayer in comparison with the QT35 cells. Furthermore, freshly seeded LMH and QT35 cells suspended in cell-free filtrates of T. gallinae were unable to form a confluent monolayer. In comparison to T. gallinae, clonal cultures of T. gallinarum or their cell-free filtrates produced no effect on both types of monolayers. Interestingly, the cell-free filtrates obtained from both trichomonad species had an effect on the viability of both cell cultures. However, the cytotoxic effect of T. gallinarum filtrates was less severe than that recorded by T. gallinae. Consequently, for the first time a destruction of specified monolayers induced by T. gallinae-free filtrates could be demonstrated.     

Genetic structure and differentiation among grapevines (<Emphasis Type="Italic">Vitis vinifera</Emphasis>) accessions from Maghreb region

Three gene pools representative of Vitis vinifera L. subsp. vinifera (=subsp. sativa Beger) growing in the Maghreb regions (North Africa) from Tunisia (44), Algeria (31) and Morocco (18) and 16 wild grape accessions (Vitis vinifera L. subsp. sylvestris (Gmelin) Beger) from Tunisia were analysed for genetic diversity and differentiation at twenty nuclear microsatellites markers distributed throughout the 19 grape chromosomes. 203 alleles with a mean number of 10.15 alleles per locus were observed in a total of 109 accessions. Genetic diversities were high in all populations with values ranging from 0.6775 (Moroccan cultivars) to 0.7254 (Tunisian cultivars). F _st pairwise values between cultivated grapevine populations were low but found to be significantly different from zero. High F _st pairwise values were shown between wild and cultivated compartments. Two parent offspring relationships, two synonyms and two clones of the same cultivar were detected. The rate of gene flow caused by vegetative dissemination of cultivated grapevine plants was not sufficient to genetically homogenise the pools of cultivars grown in different regions. The Neighbour Joining cluster analysis showed a clear separation according to geographical origins for the cultivated grapevines gene pools and revealed a high dissimilarity between cultivated and wild grapevine. However, three cultivars (Plant d’Ouchtata 1, Plant de Tabarka 3 and Plant d’Ouchtata 3) are very close to wild accessions and may result from a hybridisation between cultivated and wild accessions. The high level of differentiation between cultivated and wild accessions indicates that the cultivated accessions do not derive directly from local wild populations but could mostly correspond to imported materials introduced from others regions during historical times or derived from crossing between them.     

Biochemical and molecular investigation of oxidative stress associated with urolithiasis induced by increased dietary calcium or protein in chickens

This study was carried out to evaluate the effects of induced urolithiasis by high dietary calcium (Ca) or protein levels on biochemical analyte levels, redox status, selected inflammatory cytokines and histopathology in chickens. A total of 90 one-day-old white Hy-Line chicks were fed basal control diets containing 20% crude protein (CP) and 1% Ca until they reached 44 days of age. After that, the birds were divided into three groups (30 birds per group). All management factors (light, temperature, ventilation, stock density and diet) were identical among the three groups throughout the study except for the dietary Ca and protein percentages. Group I was fed a control diet containing 20% CP and 1% Ca, group II was fed a high-Ca diet containing 5% Ca, and group III was fed a high-protein diet containing 25% CP. Our findings clearly demonstrated that dietary imbalance (caused by high-Ca or high-CP levels) per se in chickens was physiologically harmful, as it was accompanied by post-mortem lesions; biochemical, redox status and histopathological alterations; and upregulation of inflammatory cytokines (interleukin (IL)-1β and IL-6). In particular, the birds fed the high-Ca diet clearly exhibited the most obvious alterations in most of the endpoints. In conclusion, this study constitutes the first extensive investigation of the effects of high-Ca or high-protein diets induced urolithiasis on growth performance, redox status, inflammatory cytokine levels and pathological characterization in chickens.     

Characterization of single nucleotide polymorphism in Tunisian grapevine genome and their potential for population genetics and evolutionary studies

In this study, two gene fragments corresponding to the VvMYBA1 and VvMYBA2 loci were sequenced on a sample of grapes including cultivated and wild accessions originating from Tunisia, Germany and France. A total of 42 SNPs were detected in the sequenced fragments giving an average of 1 SNP every 33 bp. High level of polymorphism was observed in the samples either in cultivated or wild accessions. Pattern of nucleotide diversity indicates a non departure from neutrality expectations for wild grapevine sample for gene VvMYBA1 and VvMYBA2 and for cultivated sample for gene VvMYBA1. However, a linkage to a selective sweep was revealed for cultivated grapevine gene pool in gene VvMYBA2. A genetic structure of the studied sample according to accession taxonomic status was revealed by the UPGMA clustering with a considerable overlap. This result was confirmed by significant but low genetic differentiation values between cultivated and wild sample. The number of migrants Nm based on sequence data information between Tunisian cultivars and Tunisian wild accessions showed a low level of gene flow between those germplasms. This finding indicates that Tunisian cultivars do not derive directly from local wild populations but could mostly correspond to imported materials introduced during historical times. However, the possibility that some cultivars derived from ancestral events of local domestication or cross hybridization with native wild plants was not completely excluded for Tunisian grapevine accessions.     

Genetic stability of long-term micropropagated Opuntia ficus-indica (L.) Mill. plantlets as assessed by molecular tools: Perspectives for in vitro conservation

Randomly amplified polymorphic DNA markers (RAPD) were employed to assess the level of genetic stability of long term micropropagated prickly pear (Opuntia ficus-indica) plantlets.Thirteen micropropagated plantlets were chosen from a clonal collection of shoots that originated from a single mother shoot. This clonal collection had been maintained under in vitro culture conditions for at least 5 years, as achieved for the time by axillary branch multiplication in Opuntia ficus-indica.Twenty arbitrary primers were used to compare RAPD patterns between in vitro raised material and the mother plant. Only 11 primers were found to yield distinct and reproducible amplification products resulting in a total of 87 amplified products, out of which 82 bands were monomorphic across all the plantlets and 5 showed polymorphisms.Cluster analysis performed on the basis of similarity indices indicated that all micropropagated plantlets and their mother plant grouped together in one major cluster with a 91% level of similarity.Low level of genetic variation has been detected, as polymorphic bands accounted for just 2.79% of the total genetic variation. This very low level of genetic variation, despite more than 5 years of in vitro culture, demonstrates the genetic stability of Opuntia ficus-indica and indicates that the axillary branch multiplication method is highly reliable for the multiplication of genetically true-to-type plant material.The high degree of clonal fidelity detected here, recommend the use of axillary-branching micropropagation technique for the safe in vitro conservation of prickly pear interesting genetic resources.     

Some Pharmacokinetic Data for Danofloxacin in Healthy Goats

The pharmacokinetics of danofloxacin was determined in five clinically normal adult female goats after intravenous (IV) or intramuscular (IM) doses of 1.25 mg/kg body weight. Blood and urine samples were collected from each animal at precise time intervals. Serum and urine concentrations were determined using microbiological assay methods and the data were subjected to kinetic analysis. After intravenous injection, the serum concentration–time curves of danofloxacin were characteristic of a two-compartment open model. The drug was rapidly distributed and eliminated with half-lives of 17.71±1.38 min and 81.18±3.70 min, respectively. The drug persisted in the central, highly perfused organs with a K ₁₂/K ₂₁ ratio of 0.67±0.25. The mean volume of distribution at a steady state (V _dss) was 1.42±0.15 L/kg. After intramuscular administration, the serum concentration peaked after 0.58±0.04 h at approximately 0.33±0.01 g/ml. While danofloxacin could be detected in serum for 4 and 6 h, it was recovered in urine for up to 24 and 72 h after IV and IM administration, respectively. The systemic bioavailability after IM injection was 65.70%±10.28% and the serum protein-bound fraction was 13.55±1.78%.