Dieback and wilting symptoms caused by complex soilborne fungi are nowadays the most serious threatening disease affecting olive trees (Olea europaea) in Tunisia and presumably in many Mediterranean basin countries. Fusarium is one of the important phytopathogenic genera associated with dieback symptoms of olive trees. The objective of the present study was to confirm the pathogenicity of Fusarium spp. isolated from several olive-growing areas in Tunisia. According to the pathogenic test done on young olive trees (cv. Chemlali), 23 out of 104 isolates of Fusarium spp. were found to be pathogenic and the others were weakly or not pathogenic. The pathogenic Fusarium spp. isolates were characterized using molecular methods based on ITS PCR. Isolation results revealed the predominance of Fusarium solani (56.5%) and F. oxysporum species (21.7%) compared to F. chalmydosporum (8.7%), F. brachygibbosum (8.7%) and F. acuminatum (4.34%). Based on pathogenicity test, disease severity was highly variable among the 23 pathogenic isolates tested (P < 0.05) where F. solani was the most aggressive dieback agent. To the best of our knowledge, this is the first work that shows that Fusarium spp. might be a major agent causing dieback disease of olive trees in Tunisia.
Forty-two isolates of Verticillium dahliae were recovered from stem and root samples of olive trees showing symptoms of verticillium wilt in various olive-growing regions in Tunisia. Each isolate was identified based on microscopic observations of morphological and cultural characteristics, pathogenicity tests, as well as PCR amplification using Vd1/Vd2 primers. Genetic diversity among the isolates was investigated using random amplified microsatellites (RAMS) and PCR-RFLP of intergenic spacer region (IGS) of ribosomal DNA (rDNA). A single fragment of approximately 1.7-2.
Since 2006, verticillium wilt of olive induced by Verticillium dahliae has caused considerable economic losses in olive orchards in Tunisia. Using virulence tests, Vegetative Compatibility Grouping (VCG) and Amplified Fragment Length Polymorphism (AFLP) analyses, we investigated the genetic structure of V. dahliae isolates collected from different olive growing
Accepted ArticleThis article is protected by copyright. All rights reserved.regions. In all, 42 isolates of V. dahliae from diseased olive trees were tested. Cluster analysis and principal coordinate analysis revealed that geographic origin was the main factor determining the genetic structure of V. dahliae populations and both methods indicated a genetic separation between the central and coastal isolates. Isolates were divided into two major groups: the AFLP-I group included all isolates from Sidi Bouzid, Kairouan, Kasserine and Sfax (center of the country) and the AFLP-II group included isolates from Monastir, Zaghouane, Sousse, Mahdia (Coastal region), and two isolates from Sfax. Analysis of the molecular variance (AMOVA) indicated a significant level of genetic differentiation among (76%) and within (23%) the two populations. Analyses of both the defoliating (D) and nondefoliating (ND) pathotypes and VCG markers indicated that most of the isolates belong to VCG 2A and 4B/ND pathotype. The disease severity was highly variable among the isolates tested (P < 0.05) with no evidence of association between aggressiveness and geographical origin of the isolates. Overall, results of this study revealed a clear association between the genetic diversity of the isolates and their geographic origin, but not between genetic diversity and virulence patterns.
A rapid, sensitive, and highly reproducible real-time PCR assay was developed to detect the three major Vibrio spp. pathogenic for humans in Tunisian seafood products and sediments. A conventional culture method found 102 (41.3%) of 247 analyzed samples positive for Vibrio spp.; a conventional PCR method found 126 (51%) of the 247 samples positive. Real-time PCR assay found 126 (51.1%) samples positive; V. alginolyticus toxR was the most common, found in 99 (78.57%) of samples, followed by V. parahaemolyticus in 26 (20.63%) and V. cholerae in 1 (0.7%). All culture-positive samples were PCR positive. However, 24 samples that were positive by conventional PCR and real-time PCR were culture negative. Our findings indicate that retail seafood is commonly contaminated with Vibrio spp. and presents a potential risk to human health in Tunisia. These data also indicate that real-time PCR can provide sensitive species-specific detection of Vibrio spp. in seafood without prior isolation and characterization of the bacteria by traditional microbiological methods.
25CTX-M ESBLs have been increasingly reported from human enterobacteria isolates in 26 Tunisia. NDM-1 carbapenemase was recently reported in isolates from Tunisian Hospitals. 27 During a 2-month period (December 2017 to January 2018), we have collected 23 ESBL- 28 producing Enterobacteriaceae (ESBL-E) from urines of patients hospitalized in three health 29 care facilities and from community, situated in two distinct Tunisian geographical regions. 30 They were divided into 15 Escherichia coli and eight Klebsiella pneumoniae. The aim of this 31 study was to characterize ESBL-E with regard to their β-lactamase content and their 32 epidemiological relationship. The results indicated a high rate (47%) of E. coli producing both 33 CTX-M group-1 and -9. For the first time, we demonstrated the presence of E. coli having 34 concomitantly CTX-M-15 and CTX-M-27 which belong to two sequences types (ST), ie. A-35 ST617 (2 isolates) and B2-ST131 subclade C2 (2 isolates). All four E. coli isolates carried a 36 multireplicon IncF with an identical allelic combination, F31:A4:B1. This study reports also 37 the first description of K. pneumoniae belonging to the clone ST147 carrying the 38 carbapenemase NDM-1 in the Tunisian community. In conclusion, our data confirms the need 39 for monitoring the resistance to extended-spectrum cephalosporins and to carbapenems 40 among enterobacteria in Tunisia. 41 42 43Resistance to extended-spectrum cephalosporins (ESC) is widespread among 44 Enterobacteriaceae species and is mainly due to the production of extended-spectrum ß-45 lactamases (ESBLs). The dissemination of ESBLs is become a growing concern since they 46 are considered as a major cause of morbidity and mortality. Up to the end of the 1990s, TEM-47 and SHV-type ESBLs were mainly produced by the Klebsiella pneumoniae and Enterobacter 48 spp. responsible for nosocomial infections (1, 2). Since 2000s, CTX-M ESBLs have gained 49 prominence mainly in Escherichia coli and K. pneumoniae strains and are now considered as 50 pandemic enzymes. Many bla CTX-M variants exist, but they can be divided into two main 51 clusters (groups -1 and -9); each cluster of CTX-M genotypes has a corresponding progenitor 52 gene sharing homology with different environmental Kluyvera spp. from which bla CTX-M 53 genes originated. Among CTX-M enzymes, CTX-M-15 belonging to group 1, has currently 54 been the most frequent all over the world (1, 3) 55 The majority of CTX-M type ESBL-associated E. coli infections is often concentrated within 56 specific extraintestinal pathogenic E. coli (ExPEC) lineages belonging to highly virulent 57 phylogenetic group B2, and recognizable by their sequence type (ST). The bla CTX-M-15 is the 58 dominant ESBL gene in the virulent E. coli ST131 clone, in particular in the subclade ST131-59 C2 (4). However other genetically divergent CTX-M genes also occur in this ST, such as 60 bla CTX-M-14/14-like variants in Canada, China, and Spain (5, 6). A subclade of E. coli ST131 61 producing the CTX-M-27 (group 9 enzyme, CTX-M-1...
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