Screening of biological activities of fungi associated with pistachio bark beetle, Chaetoptelius vestitus (Coleoptera, Curculionidae), infesting pistachio cultivations in Tunisia.

AIMS: In this study, we aimed to explore the enzymatic diversity, the entomopathogenic and the antimicrobial potentialities of fungi associated with the pistachio bark beetle, Chaetoptelius vestitus. METHODS AND RESULTS: A total of 40 isolates were screened for enzymatic diversity. Most of them, 92·5%, were able to produce at least two of the screened enzymes. Pathogenic assays performed on C. vestitus showed a high entomopathogenic activity of the isolates Ata_io_1 (A. tamarii), Fve_io_1 (F. verticillioides), Tpi_io_1 (T. pinophilus), Pal_io_1 (P. album), Pbi_io_2 (Penicillium bilaiae) and Pch_io_1 (P. chrysogenum), as based on mean mortality of C. vestitus. A screening of antimicrobial activity using well diffusion method showed that the isolates Tro_io_1 (T. pinophilus), Tat_io_1 (T. atroroseus) and Pch_io_1 (P. chrysogenum) presented the highest antibacterial activity. Furthermore, Mgu_io_1 (M. guilliermondii), Asc_io_1 (A. sclerotiorum), Ata_io_1 (A. tamarii), G. lavendula (Gla_io_1), Pva_io_1 (P. variotii), Pul_io_1 (P. ulaiense), Tat_io_1 (T. atroroseus) and Tro_io_1 (T. roseum) were active against at least two of the three tested fungal phytopathogens. CONCLUSIONS: Fungal isolates representing entomopathogenic activity and a broad spectrum of antimicrobial activities can be considered as promising resources for biological pistachio trees protection. SIGNIFICANCE AND IMPACT OF THE STUDY: Fungi associated with C. vestitus were investigated for detecting their potential biotechnological applications.

Phytopathogenic and antagonistic potentialities of fungi associated with pistachio bark beetle, Chaetoptelius vestitus (Coleoptera, Curculionidae), infesting pistachio (Pistacia vera) in Tunisia.

AIM: This study investigated the biodiversity of fungi associated with the pistachio bark beetle, Chaetoptelius vestitus, in Tunisia. We evaluated the phytopathogenic activities and tested antagonistic potentialities with respect to phytopathogens. METHODS AND RESULTS: A total of 41 fungal isolates were randomly isolated from C. vestitus adults and galleries. We identified 28 species belonging to 13 genera using ITS sequences of the ribosomal RNA operons. Pathogenicity assays performed using the excised shoot method revealed that isolates Aal_io_1 (Alternaria alternata), Feq_io_1 (Fusarium equiseti), Fgra_io_1 (Fusarium graminearum), Fve_io_1 (Fusarium verticilloides), Tro_io_1 (Trichothecium roseum) and Nqu_io_1 (Nothophoma quercina) displayed a high pathogenic activity on pistachio stems. Estimation of the antagonistic potentialities of isolated fungi against several phytopathogenic isolates as tested using a dual-culture method showed that isolates Tpi_io_1 (Talaromyces pinophilus), Pbi_io_2 (Penicillium bilaiae), Asc_io_1 (Aspergillus sclerotiorum) and Gla_io_1 (Geosmithia lavendula) displayed a broad range of antagonistic activities. CONCLUSION: Fungi associated with C. vestitus had a variable range of pathogenic activity on pistachio stem. Phytopathogenic fungi were antagonized by different fungal isolates which could be promising in pistachio protection against phytopathogenic fungi. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first that investigated the diversity of fungi associated with C. vestitus and evaluated both their phytopathogenic activity and antagonistic potential against fungal phytopathogens.

Screening of bacterial isolates related to olive orchard pests in Tunisia using 16S ribosomal RNA and evaluation of their biotechnological potential.

AIM: This study investigated the bacterial diversity of dead insects collected from olive tree orchards widely cultivated in Tunisia. We aimed to explore metabolic diversity, screen enzymatic activities for biotechnological applications and carry out preliminary bioassays for bio-insecticide development. METHODS AND RESULTS: A total of 50 bacteria strains were randomly isolated from four different biotopes situated in Sfax (Tunisia). 16S rRNA gene sequencing was used to identify 24 species, and isolates were evaluated for enzymatic activity and antimicrobial potential. Nineteen of the total number of Bacillus strains have enzymatic activity compared to strains from the other genera (14/50). Our bacterial collection was evaluated for antimicrobial potential against bacterial and fungal isolates. Isolates B8-2 (Bacillus subtilis) showed strong antibacterial activities and isolates B2-3 (Bacillus licheniformis) and B10-1 (Serratia marcescens) showed the highest antifungal activity. Seventeen of the total number of isolates caused greater than 50% mortality rate of second and fourth instar larvae of Ephestia kuehniella. CONCLUSION: The selected species from olive orchards represent a broad spectrum of antimicrobial and insecticidal activities and can be considered promising resources in biological control. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteria isolated from olive pests in olive orchards were investigated for detecting their potential biotechnological applications.

Septic elbow arthritis in children: Epidemiology and outcome.

INTRODUCTION: Septic elbow arthritis is a rare disease, especially in children. The aim of this study was to report the functional outcome of our pediatric population with both demographic and microbiological features. METHODS: We retrospectively reviewed all our cases of pediatric septic elbow arthritis between 2007 and 2017. Demographic, microbiological, biological, and radiological features were analyzed. Functional outcome was evaluated according to the Morrey Elbow Performance Score (MEPS). RESULTS: In total, 14 cases were reviewed. The mean age was 9 years. Microbiological findings revealed Staphylococcus aureus in 10 cases. The mean white blood cells count in synovial fluid was 101,306 cells per mm 3. The skin was the most frequently reported entry portal. The mean MEPS score was 86.13 points (excellent in eight cases, good in three cases, fair in two cases, and poor in two cases). Stiffness was observed in three cases. No growth disturbance was reported. DISCUSSION: Septic elbow arthritis in children is rare. Biological inflammatory markers are sensitive but not specific. White blood cell count in the synovial fluid is specific. S. aureus is the most frequently cultured agent. Treatment is based on surgical debridement, antibiotics, and elbow immobilization. Concomitant osteomyelitis is frequent and should be diagnosed and treated simultaneously. Functional outcomes are usually satisfactory in contrast to osteomyelitis. CONCLUSION: Pediatric septic elbow arthritis is rare. It may be primary or secondary to trauma and surgery of the elbow. Demographic, biological, and microbial features are similar to those reported for osteomyelitis; however, functional outcomes are better with a negligible rate of growth disturbance.

Partial purification and characterization of chiIO8, a novel antifungal chitinase produced by Bacillus cereus IO8.

AIMS: To investigate the distribution of chitinase IO8 in Bacillus cereus strains, the enhancing effects of the chitinase-producing B. cereus strains on biocontrol potential by dual culture assay and in vivo assay against Botrytis cinerea and also the enhancing effects of the chiIO8 on disinfectant properties against seed-borne diseases. Moreover, the application of chiIO8 treatment was also observed to improve the germinative energy. METHODS AND RESULTS: The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose anion-exchange chromatography and Sephacryl S-400 high-resolution gel chromatography. The method gave a 5.8-fold increase in the specific activity and had a yield of 17%. The molecular weight of the partially purified chitinase chiIO8 was found to be around 30 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The optimal pH and optimal temperature of the partially purified chitinase were pH 6.5 and 65°C, respectively. The thermostable chitinase still retained the activity after incubation for 100 min at 65°C, and it was increased about 1.25 times than that of the control (before heating) when the enzyme solution heated at 65°C for 60 min. The partially purified chitinase chiIO8 displays a wide inhibitory spectrum towards all phytopathogenic fungi tested. chiIO8 also exhibited effective disinfectant properties against seed-borne diseases. CONCLUSION: The present investigation emphasizes the potential of chitinase-producing micro-organism as promising biocontrol agents of fungal plant pathogens with chitinous cell wall. The novel chitinase chiIO8 proved an efficient, environmentally safe and user-friendly solution. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first investigation devoted exclusively to analyse the distribution of chitinase in B. cereus. It infers that the chitinase produced by B. cereus might play a role in the activity of the biopesticide.

First Report of a Branch Dieback of Olive Trees in Tunisia Caused by a Phoma sp.

From 2007 to 2008, a new dieback of branches of olive trees was observed in several orchards in central and southern Tunisia. The appearance of this new syndrome coincided with warm temperatures and frequent rainfall from February to April 2007. Affected trees were observed in seven commercial orchards; disease incidence ranged from 1 to 9% and affected trees were randomly distributed in each orchard. Symptoms included abundant dead branches and wilted leaves remained attached. Distinct brown areas appeared on the bark of current-year shoots as well as on larger branches. Cankers on branches that were >2 years old were difficult to detect but were conspicuous in current-year shoots. To determine the etiology of this new syndrome, a study was carried out on samples of affected branches collected from 2007 to 2008 from different areas of the country. Unidentified species of Chaetomium and Phoma were isolated from the margins of the cankers. Koch's postulates were performed with one isolate each of a Chaetomium sp. and a Phoma sp on 2-year-old olive trees, cv. Chemlali, grown in 13-cm-diameter pots containing a sand/lime/peat mixture. Stems were inoculated by placing 10 µl of conidial suspension (106 conidia/ml) on 1-cm-long longitudinal stem wounds that had been made with a sterile scalpel. Control plants were wounded, but inoculum was replaced with sterile distilled water. Three sets of 10 plants each were wound inoculated with each of the fungi on the same day. Inoculated plants were covered with a polyethylene plastic bag to retain moisture and incubated for 2 months at 30°C with a 12-h photoperiod. After 45 days, only branches inoculated with the Phoma isolate showed brown discoloration areas at the inoculation sites. A Phoma sp. was recovered from necrotic bark from each of the 10 inoculated plants. Conidia were hyaline, unicellular, slightly ellipsoidal, and 4.8 to 6.3 × 1.8 to 2.2 µm. To confirm the identification, DNA extraction was done with hyphae collected from a 7-day-old culture on PDA after incubation at 25°C (1). Fungal primers ITS1 and ITS4 (3) were used for amplification. Purified amplicons were directly sequenced using the ITS1 and ITS4 primers for the internal transcribed spacer region of rDNA. A BLAST search of the GenBank database revealed 96% homology with Phoma sp. isolate (AJ972865.1) and 98% homology with Phoma medicaginis isolate (DQ026014.1). P. incompta has been reported as responsible for branch dieback of olive tree in Italy (2). To our knowledge, this is the first report of a canker disease of olive caused by a Phoma sp. in Tunisia. References: (1) S. R. Tendulkar et al. Biotechnol. Lett. 22:1941, 2003. (2) L. Tosi and A. Zazzerini. Petria 4:161, 1994. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.