Screening and Characterization of Streptomyces spp. Isolated from Three Moroccan Ecosystems Producing a Potential Inhibitor of the Drug Efflux Pump AcrAB-TolC.

Traditional antimicrobial antibiotics are increasingly suffering from the emergence of multidrug resistance among pathogenic microorganisms. The antibiotic era is threatened by the ruthless rise of resistance in bacterial infections. A significant role in these resistance profiles is attributed to multidrug efflux pumps. Hence, much effort is being directed towards developing new compounds to overcome this problem. During our screening program of efflux pumps inhibitors (EPI) produced by bioactive Moroccan Actinobacteria, 210 isolates were screened for their antibacterial activities against Escherichia coli strains containing a system of efflux pump AcrAB-TolC, fully functional, and its mutant, inactivated due to the insertion of transposon Tn903 in AcrAB operon, using the method of agar disc diffusion. The results showed that 14 isolates were able to produce EPI as they were active against the wild type strain but not against the mutant in comparison with the synthetic inhibitor L-Phe-L-Arg-ß-naphthylamide (PaßN). We focused on the highest EPI activity produced by four strains (Z332, Z35/G, Z385/b and 136). Taxonomic studies and the 16S rDNA sequence indicated that these strains belonged to the Streptomyces species. This work could contribute to the discovery of a new class of antibacterial agents that could expand the therapeutic arsenal.

Endophytic actinobacteria of medicinal plant Aloe vera: Isolation, antimicrobial, antioxidant, cytotoxicity assays and taxonomic study

Objective: To explore the new sources of novel bioactive compounds having pharmaceutical and agricultural interest and to search the endophytic actinobacteria from medicinal plants. Methods: NAF-1 an endophyte actinobacteria was isolated from leaves of medicinal plant Aloe vera collected in Marrakesh, Morocco using Bennett agar as selective medium. NAF-1 was tested for its antimicrobial activity against five pathogenic bacteria such asStaphylococcus aureus PIC 53156,Micrococcus luteus ATCC381,Bacillus subtilis ATCC 14579,Pseudomonas aeruginosa DSM 50090 and Escherichia coli ATCC 8739 and four human clinic fungi belonging to the Candida,Aspergillus and Microsporum genera. Several antioxidant activities were studied such as DPPH free radical scavenging,β-carotene and linoleic acid and reducing power assays. The total of phenol and flavonoid was also calculated. Using Artemia salina shrimp assay, the cytotoxicity of NAF-1 crude extract was determined.Results: The results revealed that the actinobacteria showed a high activity (≥20 mm) against only Gram positive bacteria but it had a moderate activity (between 13 and 15 mm) against Human clinic fungi. The isolate also exhibited a LD50 of 14.20 μg/mL in the cytotoxicity assay. The result showed that the crude extract presented an interesting free radical-scavenging activity with IC50 value of (5.58 ± 0.26) μg/mL and a high value of phenolic and flavonoid compounds with (15.41 ± 0.18) μg GAE/mg extract and (11.41± 0.06) μg QE/mg extract respectively. Moreover, the taxonomic position of our endophyte actinobacteria using the morphological and physiological criteria and using16SrRNA gene sequence (polyphasic approach) showed that the NAF-1 isolate was similar to Streptomyces hydrogenans which was never described as an endophyte actinobacteria. Conclusions: This isolated strain appears promising resources of bioactive agents and can be exploited to produce therapeutic agents active against pathogenic disease.

Frequency of ubiquitin and FUS-positive, TDP-43-negative frontotemporal lobar degeneration.

Frontotemporal lobar degeneration (FTLD) is a clinically, genetically and pathologically heterogeneous disorder. Within FTLD with ubiquitin-positive inclusions (FTLD-U), a new pathological subtype named FTLD-FUS was recently found with fused in sarcoma (FUS) positive, TDP-43-negative inclusions, and striking atrophy of the caudate nucleus. The aim of this study was to determine the frequency of FTLD-FUS in our pathological FTLD series, and to describe the clinical, neuroimaging and neuropathological features of FTLD-FUS, especially caudate atrophy. Demographic and clinical data collected prospectively from 387 patients with frontotemporal dementia (FTD) yielded 74 brain specimens. Immunostaining was carried out using a panel of antibodies, including AT-8, ubiquitin, p62, FUS, and TDP-43. Cortical and caudate atrophy on MRI (n = 136) was rated as normal, mild-moderate or severe. Of the 37 FTLD-U cases, 33 were reclassified as FTLD-TDP and four (0.11, 95%: 0.00-0.21) as FTLD-FUS, with ubiquitin and FUS-positive, p62 and TDP-43-negative neuronal intranuclear inclusions (NII). All four FTLD-FUS cases had a negative family history, behavioural variant FTD (bvFTD), and three had an age at onset

TDP-43 pathology in familial frontotemporal dementia and motor neuron disease without Progranulin mutations.

Frontotemporal dementia is accompanied by motor neuron disease (FTD + MND) in approximately 10% of cases. There is accumulating evidence for a clinicopathological overlap between FTD and MND based on observations of familial aggregation and neuropathological findings of ubiquitin-positive neuronal cytoplasmatic inclusions (NCI) in lower motor neurons, hippocampus and neocortex in both conditions. Several familial forms exist with different genetic loci and defects. We investigated the familial aggregation and clinical presentation of FTD + MND cases in a large cohort of 368 FTD patients in The Netherlands. Immunohistochemistry of available brain tissue of deceased patients was investigated using a panel of antibodies including ubiquitin, p62 and TAR DNA-binding protein of 43 kDa antibodies. A total of eight patients coming from six families had a family history positive for FTD + MND (mean age at onset 53.2 +/- 8.4 years). Five patients presented with behavioural changes and cognitive changes followed by motor neuron disease, whereas symptoms of motor neuron disease were the presenting features in the remaining three patients. Other affected relatives in these families showed dementia/FTD, MND or FTD + MND reflecting the clinical interfamilial variation. No mutations were identified in any of the candidate genes, including Superoxide Dismutase 1, dynactin, angiogenin, Microtubule-Associated Protein Tau, valosin-containing protein and progranulin. Available brain tissue of five patients with familial FTD + MND showed NCI in hippocampus, neocortex and spinal cord in all, and neuronal intranuclear inclusions (NII) in two brains. TDP-43 antibody showed robust staining of neuronal inclusions similar in distribution and morphology to NCI and NII. Additionally, TDP-43 antibody also stained ubiquitin-negative glial inclusions in the basal striatum of one case. In conclusion, there exists considerable clinical variation within families with FTD + MND, which may be determined by other genetic or environmental factors. NII are also found in some cases of familial FTD + MND without Progranulin mutations. The observation of glial TDP-43 positive inclusions in one brain is very interesting, although their pathophysiological significance is yet unknown.

Progranulin mutations in Dutch familial frontotemporal lobar degeneration.

Mutations in the progranulin (PGRN) gene have recently been identified in frontotemporal lobar degeneration with ubiquitin inclusions linked to chromosome 17q21. We report here the finding of two novel frameshift mutations and three possible pathogenic missense mutations in the PGRN gene. Furthermore, we determined the frequency of PGRN mutations in familial cases recruited from a large population-based study of frontotemporal lobar degeneration carried out in The Netherlands.

The DeltaK280 mutation in MAP tau favors exon 10 skipping in vivo.

Tau mutations in frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) are associated with changes in alternative splicing of exon 10. The DeltaK280 mutation in exon 10 is exceptional because in vitro observations suggest a dramatic effect on microtubule binding, enhanced self-aggregation, as well as a decrease of the 4R/3R ratio by the ablation of an exon splicing enhancer element. Using immunohistochemistry, Western blotting, and electron microscopy on brain material with the DeltaK280 mutation, we investigated which of these effects is most dominant in vivo. The brain showed abundant Pick bodies in several brain regions, which stained positive with 3-repeat-specific but not with 4-repeat-specific tau antibodies. Western blots of sarkosyl-insoluble tau showed exclusively three repeat (3R0N and 3R1N) tau in most regions, although some 4R1N could be detected in the frontal cortex. In addition, the sarkosyl-soluble tau fraction showed a significantly higher amount of 3-repeat tau. Because quantitative analysis of 4R and 3R mRNA transcripts showed a 4R/3R ratio of only 0.3, association between increased transcription and protein expression was observed. These observations confirm the postulated hypothesis that the DeltaK280 mutation abolishes a splice enhancer element, which overrules the decreased microtubule binding and enhanced self-aggregation.