Anticancer, antibacterial, and antifungal activities of arum palaestinum plant extracts

Traditional natural remedies have long played an important role in the treatment of cancer and infectious diseases. A part from the high cost and undesirable side effects associated with synthetic drugs, increased interest has intensified to determine the biological effects of plant extracts on malignant cells as alternative for conventional drugs used in the markets. The medicinal properties of Arum Palaestinum Boiss were in-vitro investigated in this research project. Arum palaestinum is chosen based on its use in traditional palestinian herbal medicine. The leaves of this plant were air dried in the shade and then three types of extract were obtained and their antimicrobial and for anticancer activity testing. Investigations on three different cancer cell lines (C2Cl2, 3T3-L1, Hela) revealed direct inhibitory effect of the extracts. This effect differs according to concentrations used. Aqueous boiled extract was more effective at lower extract concentrations as compared to other two extracts. Arum palaestinum plant extracts showed no inhibitory effect on bacterial species (Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa) as well as yeasts (Candida albicans). In conclusion, Arum palaestinum has potentially anticancer effect. Further investigations are required to confirm this conclusion and to elucidate the mechanisms of actions and toxicity of this herb

Occurrence and molecular characterization of metallo-β-lactamases (MBLs) among Acinetobacter baumannii isolates from cancer patients: Acinetobacter baumannii cancer patients

Background: During the last decade, the prevalence of carbapenem-resistant infection associated with multidrug resistant (MDR) Acinetobacter baumanniiin patients has been continuously increasing.  This prospective study aimed to determine the occurrence and molecular characterization of metallo-β-lactamases (MBLs) and carbapenem hydrolyzing oxacillinases among A. baumannii isolates from cancer patients over a period of 6-month. Methods:Antimicrobial susceptibility profile of 70 randomly collected A. baumannii isolates was first determined using disc diffusion test, and second, the MICs of 45 representative multidrug resistant (MDR) isolates were tested to useful drugs in treatment of their infections using E-test.  PCR assays were used to detect the common four types of class D carbapenem hydrolyzing oxacillinases, two types of class A carbapenemases, four types of class B metallo-β –lactamases, and prevalence of Class 1 Integron among MDR isolates. Results: All 70 isolates were MDR, including 100% resistance to meropenom, aztreonem, piperacillin/tazobactum and 99% to carbapenem.  All isolates carried blaOXA-23 and blaOXA-51, but none carried a blaOXA-24 like or blaOXA-58. The isolates also were positive for NDM-1, NDM, VIM, GES, KPC and SPM at the rates of 29%, 20%, 29%,19%,7% and 2%, respectively.  Class 1 Integron was positive in 82% of A. baumanniiisolates. The clonal relationship of 42 MDR A. baumanniiisolates using ERIC-PCR and constructed dendrogram showed 3 major genotype clusters of genetically related isolates. These include 4 genotype groups, each composed of 2 isolates with 100 % similarity of DNA bands.  Conclusion:This study demonstrates that A. baumannii colonize frequently cancer patients in association with antibiotic treatment. The organism is mostly carrying wide spectrum of antibiotic resistance genetic factors, especially many types of ESBLs and MBLs and Class 1 Integron. This fact should be considered when therapy is selected for treatment of patients infected with MDR A. baumannii. Key words. Acinetobacter baumannii, ESBLs, MBLs, Class 1 Integron, Jordanian cancer patients. &nbsp

Value of phenotyic and genotypic identification of Acinetobacter baumannii isolates from two hospitals in Jordan

Acinetobacter baumannii isolates have been recovered from hospitalized patients over the past few years from two hospitals in Jordan. Phenotypic and biochemical characterization in addition to disc susceptibility testing of all clinical isolates indicated that all of them were belonging to A. baumannii. A high degree of conservation of both the ITS length and the ITS sequence was observed, and their identities were further confirmed by amplified ribosomal DNA gene restriction analysis (ARDRA). The application of ARDRA for the identification of Acinetobacter species has several advantages over phenotypic identification. ARDRA considered rapid and reliable and universally applicable method for identification of most of the Acinetobacter genomic species, thus contribute to better understanding of the clinical importance and epidemiology of isolates

Molecular characterization and antibiotic susceptibility profiles of Helicobacter pylori isolated from patients with Gastrodeudenal diseases in Jordan

Introduction:  Helicobacter pylori is a major cause of more than 80% of chronic active gastritis and other gastrodeudonal diseases worldwide. Successful treatment of H. pylori routinely requires the use of multiple agents with different mechanisms including compounds inhibiting acid secretion in conjunction with antibiotics. However, recent data showed the emergence of resistant clinical strains particularly against metronidazole and clarithromycin. The aim of this study is to determine the prevalence of and the susceptibility of H. pylori isolates recovered from patients with gastrodeudonal diseases to several antimicrobial agents. Materials and Methods: A prospective study has been conducting on Jordanian patients attended the gastrointestinal unit of the Jordan university hospital starting from 2014-2015 with gastroduodenal diseases. Antral and corpus mucosal biopsies from the stomach of each patient were used for the isolation of H. pylori on selective culture media. Presumptive H. pylori colonies were subsequently confirmed by biochemical tests and standard 16S rDNA PCR assay. The antimicrobial susceptibility testing was performed by standard agar diffusion methods according to CLSI. Subsequently, MICs were determined by E test and standard agar dilution method. Molecular typing of the clinical strains was performed using multiplex PCR for the detection of vacA and cagA genotypes. Metronidazole resistance was characterized by molecular methods for the detection of rdxA gene mutations. Results: Among 72 symptomatic patients, 13 (23%) patients showed positive H. pylori infection by both rapid urease test and culture. The antibiotic susceptibility profile showed that all of the isolates were sensitive to amoxicillin.  Resistance to, clarithromycin, ciprofloxacin and levofloxacin were observed in 15%, 23% and 8% of the isolates respectively while 92% of the strains were resistant to metronidazole (MIC ≥ 32 ug/ml). Metronidazole resistance due to mutations in rdxA gene was only observed in one strain (8%) suggesting other resistance mechanisms. Correlation between antibiotic resistance and virulence factors was statistically not significant (p > 0.05). Conclusion: The present study showed that the prevalence of metronidazole resistance among clinical isolates of H. pylori is very high. Lower resistance to other antibiotics are reported. Concern should be taken into consideration when triple therapy is used for the treatment of H. pylori in our region

: a cis antisense RNA operates in trans in S. aureus

International audienceAntisense RNAs (asRNAs) pair to RNAs expressed from the complementary strand, and their functions are thought to depend on nucleotide overlap with genes on the opposite strand. There is little information on the roles and mechanisms of asRNAs. We show that a cis asRNA acts in trans, using a domain outside its target complementary sequence. SprA1 small regulatory RNA (sRNA) and SprA1(AS) asRNA are concomitantly expressed in S. aureus. SprA1(AS) forms a complex with SprA1, preventing translation of the SprA1-encoded open reading frame by occluding translation initiation signals through pairing interactions. The SprA1 peptide sequence is within two RNA pseudoknots. SprA1(AS) represses production of the SprA1-encoded cytolytic peptide in trans, as its overlapping region is dispensable for regulation. These findings demonstrate that sometimes asRNA functional domains are not their gene-target complementary sequences, suggesting there is a need for mechanistic re-evaluation of asRNAs expressed in prokaryotes and eukaryotes

Effect of Low Temperature on Growth and Ultra-Structure of Staphylococcus spp

The effect of temperature fluctuation is an important factor in bacterial growth especially for pathogens such as the staphylococci that have to remain viable during potentially harsh and prolonged transfer conditions between hosts. The aim of this study was to investigate the response of S. aureus, S. epidermidis, and S. lugdunensis when exposed to low temperature (4°C) for prolonged periods, and how this factor affected their subsequent growth, colony morphology, cellular ultra-structure, and amino acid composition in the non-cytoplasmic hydrolysate fraction. Clinical isolates were grown under optimal conditions and then subjected to 4°C conditions for a period of 8 wks. Cold-stressed and reference control samples were assessed under transmission electron microscopy (TEM) to identify potential ultra-structural changes. To determine changes in amino acid composition, cells were fractured to remove the lipid and cytoplasmic components and the remaining structural components were hydrolysed. Amino acid profiles for the hydrolysis fraction were then analysed for changes by using principal component analysis (PCA). Exposure of the three staphylococci to prolonged low temperature stress resulted in the formation of increasing proportions of small colony variant (SCV) phenotypes. TEM revealed that SCV cells had significantly thicker and more diffuse cell-walls than their corresponding WT samples for both S. aureus and S. epidermidis, but the changes were not significant for S. lugdunensis. Substantial species-specific alterations in the amino acid composition of the structural hydrolysate fraction were also observed in the cold-treated cells. The data indicated that the staphylococci responded over prolonged periods of cold-stress treatment by transforming into SCV populations. The observed ultra-structural and amino acid changes were proposed to represent response mechanisms for staphylococcal survival amidst hostile conditions, thus maintaining the viability of the species until favourable conditions arise again

nocoRNAc: Characterization of non-coding RNAs in prokaryotes

<p>Abstract</p> <p>Background</p> <p>The interest in non-coding RNAs (ncRNAs) constantly rose during the past few years because of the wide spectrum of biological processes in which they are involved. This led to the discovery of numerous ncRNA genes across many species. However, for most organisms the non-coding transcriptome still remains unexplored to a great extent. Various experimental techniques for the identification of ncRNA transcripts are available, but as these methods are costly and time-consuming, there is a need for computational methods that allow the detection of functional RNAs in complete genomes in order to suggest elements for further experiments. Several programs for the genome-wide prediction of functional RNAs have been developed but most of them predict a genomic locus with no indication whether the element is transcribed or not.</p> <p>Results</p> <p>We present <smcaps>NOCO</smcaps>RNAc, a program for the genome-wide prediction of ncRNA transcripts in bacteria. <smcaps>NOCO</smcaps>RNAc incorporates various procedures for the detection of transcriptional features which are then integrated with functional ncRNA loci to determine the transcript coordinates. We applied RNAz and <smcaps>NOCO</smcaps>RNAc to the genome of <it>Streptomyces coelicolor </it>and detected more than 800 putative ncRNA transcripts most of them located antisense to protein-coding regions. Using a custom design microarray we profiled the expression of about 400 of these elements and found more than 300 to be transcribed, 38 of them are predicted novel ncRNA genes in intergenic regions. The expression patterns of many ncRNAs are similarly complex as those of the protein-coding genes, in particular many antisense ncRNAs show a high expression correlation with their protein-coding partner.</p> <p>Conclusions</p> <p>We have developed <smcaps>NOCO</smcaps>RNAc, a framework that facilitates the automated characterization of functional ncRNAs. <smcaps>NOCO</smcaps>RNAc increases the confidence of predicted ncRNA loci, especially if they contain transcribed ncRNAs. <smcaps>NOCO</smcaps>RNAc is not restricted to intergenic regions, but it is applicable to the prediction of ncRNA transcripts in whole microbial genomes. The software as well as a user guide and example data is available at <url>http://www.zbit.uni-tuebingen.de/pas/nocornac.htm</url>.</p

Experimental discovery of small RNAs in Staphylococcus aureus reveals a riboregulator of central metabolism

Using an experimental approach, we investigated the RNome of the pathogen Staphylococcus aureus to identify 30 small RNAs (sRNAs) including 14 that are newly confirmed. Among the latter, 10 are encoded in intergenic regions, three are generated by premature transcription termination associated with riboswitch activities, and one is expressed from the complementary strand of a transposase gene. The expression of four sRNAs increases during the transition from exponential to stationary phase. We focused our study on RsaE, an sRNA that is highly conserved in the bacillales order and is deleterious when over-expressed. We show that RsaE interacts in vitro with the 5′ region of opp3A mRNA, encoding an ABC transporter component, to prevent formation of the ribosomal initiation complex. A previous report showed that RsaE targets opp3B which is co-transcribed with opp3A. Thus, our results identify an unusual case of riboregulation where the same sRNA controls an operon mRNA by targeting two of its cistrons. A combination of biocomputational and transcriptional analyses revealed a remarkably coordinated RsaE-dependent downregulation of numerous metabolic enzymes involved in the citrate cycle and the folate-dependent one-carbon metabolism. As we observed that RsaE accumulates transiently in late exponential growth, we propose that RsaE functions to ensure a coordinate downregulation of the central metabolism when carbon sources become scarce

Global Regulatory Functions of the Staphylococcus aureus Endoribonuclease III in Gene Expression

RNA turnover plays an important role in both virulence and adaptation to stress in the Gram-positive human pathogen Staphylococcus aureus. However, the molecular players and mechanisms involved in these processes are poorly understood. Here, we explored the functions of S. aureus endoribonuclease III (RNase III), a member of the ubiquitous family of double-strand-specific endoribonucleases. To define genomic transcripts that are bound and processed by RNase III, we performed deep sequencing on cDNA libraries generated from RNAs that were co-immunoprecipitated with wild-type RNase III or two different cleavage-defective mutant variants in vivo. Several newly identified RNase III targets were validated by independent experimental methods. We identified various classes of structured RNAs as RNase III substrates and demonstrated that this enzyme is involved in the maturation of rRNAs and tRNAs, regulates the turnover of mRNAs and non-coding RNAs, and autoregulates its synthesis by cleaving within the coding region of its own mRNA. Moreover, we identified a positive effect of RNase III on protein synthesis based on novel mechanisms. RNase III–mediated cleavage in the 5′ untranslated region (5′UTR) enhanced the stability and translation of cspA mRNA, which encodes the major cold-shock protein. Furthermore, RNase III cleaved overlapping 5′UTRs of divergently transcribed genes to generate leaderless mRNAs, which constitutes a novel way to co-regulate neighboring genes. In agreement with recent findings, low abundance antisense RNAs covering 44% of the annotated genes were captured by co-immunoprecipitation with RNase III mutant proteins. Thus, in addition to gene regulation, RNase III is associated with RNA quality control of pervasive transcription. Overall, this study illustrates the complexity of post-transcriptional regulation mediated by RNase III

Genetic Pathway in Acquisition and Loss of Vancomycin Resistance in a Methicillin Resistant Staphylococcus aureus (MRSA) Strain of Clonal Type USA300

An isolate of the methicillin-resistant Staphylococcus aureus (MRSA) clone USA300 with reduced susceptibility to vancomycin (SG-R) (i.e, vancomycin-intermediate S. aureus, VISA) and its susceptible “parental” strain (SG-S) were recovered from a patient at the end and at the beginning of an unsuccessful vancomycin therapy. The VISA phenotype was unstable in vitro generating a susceptible revertant strain (SG-rev). The availability of these 3 isogenic strains allowed us to explore genetic correlates of antibiotic resistance as it emerged in vivo. Compared to the susceptible isolate, both the VISA and revertant strains carried the same point mutations in yycH, vraG, yvqF and lspA genes and a substantial deletion within an intergenic region. The revertant strain carried a single additional frameshift mutation in vraS which is part of two component regulatory system VraSR. VISA isolate SG-R showed complex alterations in phenotype: decreased susceptibility to other antibiotics, slow autolysis, abnormal cell division and increased thickness of cell wall. There was also altered expression of 239 genes including down-regulation of major virulence determinants. All phenotypic properties and gene expression profile returned to parental levels in the revertant strain. Introduction of wild type yvqF on a multicopy plasmid into the VISA strain caused loss of resistance along with loss of all the associated phenotypic changes. Introduction of the wild type vraSR into the revertant strain caused recovery of VISA type resistance. The yvqF/vraSR operon seems to function as an on/off switch: mutation in yvqF in strain SG-R turns on the vraSR system, which leads to increase in vancomycin resistance and down-regulation of virulence determinants. Mutation in vraS in the revertant strain turns off this regulatory system accompanied by loss of resistance and normal expression of virulence genes. Down-regulation of virulence genes may provide VISA strains with a “stealth” strategy to evade detection by the host immune system