Alpha-sitosterol: a new antiviral agent produced by Streptomyces misakiensis and its potential activity against Newcastle disease virus.

BACKGROUND: Newcastle Disease Virus (NDV) causes severe economic losses in the poultry industry worldwide. Hence, this study aimed to discover a novel bioactive antiviral agent for controlling NDV. Streptomyces misakiensis was isolated from Egyptian soil and its secondary metabolites were identified using infrared spectroscopy (IR), gas chromatography-mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy. The inhibitory activity of bioactive metabolite against NDV were examined. Three experimental groups of 10-day-old specific pathogen-free embryonated chicken eggs (SPF-ECEs), including the bioactive metabolite control group, NDV control positive group, and α-sitosterol and NDV mixture-treated group were inoculated. RESULTS: α-sitosterol (Ethyl-6-methylheptan-2-yl]-10,13-dimethyl-dodecahydro-1H-cyclopenta[a]phenanthren-3-ol), a secondary metabolite of S. misakiensis, completely inhibited hemagglutination (HA) activity of the NDV strain. The HA activity of the NDV strain was 8 log2 and 9 log2 for 0.5 and 0.75% RBCs, respectively. The NDV HA activity for the two concentrations of RBCs was significantly (P < 0.0001) inhibited after α-sitosterol treatment. There was a significant (P < 0.0001) decrease in the log 2 of HA activity, with values of - 0.500 (75%, chicken RBCs) before inoculation in SPF-ECEs and - 1.161 (50%, RBCs) and - 1.403 (75%, RBCs) following SPF-ECE inoculation. Compared to ECEs inoculated with NDV alone, the α-sitosterol-treated group showed improvement in histological lesion ratings for chorioallantoic membranes (CAM) and hepatic tissues. The CAM of the α-sitosterol- inoculated SPF-ECEs was preserved. The epithelial and stromal layers were noticeably thicker with extensive hemorrhages, clogged vasculatures, and certain inflammatory cells in the stroma layer in the NDV group. However, mild edema and inflammatory cell infiltration were observed in the CAM of the treated group. ECEs inoculated with α-sitosterol alone showed normal histology of the hepatic acini, central veins, and portal triads. Severe degenerative alterations, including steatosis, clogged sinusoids, and central veins, were observed in ECEs inoculated with NDV. Mild hepatic degenerative alterations, with perivascular round cell infiltration, were observed in the treated group. CONCLUSION: To the best of our knowledge, this is the first study to highlight that the potentially bioactive secondary metabolite, α-sitosterol, belonging to the terpene family, has the potential to be a biological weapon against virulent NDV. It could be used for the development of innovative antiviral drugs to control NDV after further clinical investigation.

Bioactive metabolites of Streptomyces misakiensis display broad-spectrum antimicrobial activity against multidrug-resistant bacteria and fungi.

Background: Antimicrobial resistance is a serious threat to public health globally. It is a slower-moving pandemic than COVID-19, so we are fast running out of treatment options. Purpose: Thus, this study was designed to search for an alternative biomaterial with broad-spectrum activity for the treatment of multidrug-resistant (MDR) bacterial and fungal pathogen-related infections. Methods: We isolated Streptomyces species from soil samples and identified the most active strains with antimicrobial activity. The culture filtrates of active species were purified, and the bioactive metabolite extracts were identified by thin-layer chromatography (TLC), preparative high-performance liquid chromatography (HPLC), nuclear magnetic resonance (NMR) spectroscopy, and gas chromatography-mass spectrometry (GC-MS). The minimum inhibitory concentrations (MICs) of the bioactive metabolites against MDR bacteria and fungi were determined using the broth microdilution method. Results: Preliminary screening revealed that Streptomyces misakiensis and S. coeruleorubidus exhibited antimicrobial potential. The MIC50 and MIC90 of S. misakiensis antibacterial bioactive metabolite (ursolic acid methyl ester) and antifungal metabolite (tetradecamethylcycloheptasiloxane) against all tested bacteria and fungi were 0.5 µg/ml and 1 µg/mL, respectively, versus S. coeruleorubidus metabolites: thiocarbamic acid, N,N-dimethyl, S-1,3-diphenyl-2-butenyl ester against bacteria (MIC50: 2 µg/ml and MIC90: 4 µg/mL) and fungi (MIC50: 4 µg/ml and MIC90: 8 µg/mL). Ursolic acid methyl ester was active against ciprofloxacin-resistant strains of Streptococcus pyogenes, S. agalactiae, Escherichia coli, Klebsiella pneumoniae, and Salmonella enterica serovars, colistin-resistant Aeromonas hydrophila and K. pneumoniae, and vancomycin-resistant Staphylococcus aureus. Tetradecamethylcycloheptasiloxane was active against azole- and amphotericin B-resistant Candida albicans, Cryptococcus neoformans, C. gattii, Aspergillus flavus, A. niger, and A. fumigatus. Ursolic acid methyl ester was applied in vivo for treating S. aureus septicemia and K. pneumoniae pneumonia models in mice. In the septicemia model, the ursolic acid methyl ester-treated group had a significant 4.00 and 3.98 log CFU/g decrease (P < 0.05) in liver and spleen tissue compared to the infected, untreated control group. Lung tissue in the pneumonia model showed a 2.20 log CFU/g significant decrease in the ursolic acid methyl ester-treated group in comparison to the control group. The haematological and biochemical markers in the ursolic acid methyl ester-treated group did not change in a statistically significant way. Moreover, no abnormalities were found in the histopathology of the liver, kidneys, lungs, and spleen of ursolic acid methyl ester-treated mice in comparison with the control group. Conclusion: S. misakiensis metabolite extracts are broad-spectrum antimicrobial biomaterials that can be further investigated for the potential against MDR pathogen infections. Hence, it opens up new horizons for exploring alternative drugs for current and reemerging diseases.

Biocontrol of multi-drug resistant pathogenic bacteria in drainage water by locally isolated bacteriophage.

In areas with limited water resources, the reuse of treated drainage water for non-potable purposes is increasingly recognised as a valuable and sustainable water resource. Numerous pathogenic bacteria found in drainage water have a detrimental impact on public health. The emergence of antibiotic-resistant bacteria and the current worldwide delay in the production of new antibiotics may make the issue of this microbial water pollution even more challenging. This challenge aided the resumption of phage treatment to address this alarming issue. In this study, strains of Escherichia coli and Pseudomonas aeruginosa and their phages were isolated from drainage and surface water from Bahr El-Baqar and El-Manzala Lake in Damietta governorate, Egypt. Bacterial strains were identified by microscopical and biochemical examinations which were confirmed by 16 S rDNA sequencing. The susceptibility of these bacteria to several antibiotics revealed that most of the isolates had multiple antibiotic resistances (MAR). The calculated MAR index values (> 0.25) categorized study sites as potentially hazardous to health. Lytic bacteriophages against these multidrug-resistant strains of E. coli and P. aeruginosa were isolated and characterized. The isolated phages were found to be pH and heat stable and were all members of the Caudovirales order as recognized by the electron microscope. They infect 88.9% of E. coli strains and 100% of P. aeruginosa strains examined. Under laboratory conditions, the use of a phage cocktail resulted in a considerable reduction in bacterial growth. The removal efficiency (%) for E. coli and P. aeruginosa colonies increased with time and maximized at 24 h revealing a nearly 100% reduction after incubation with the phage mixture. The study candidates new phages for detecting and controlling other bacterial pathogens of public health concern to limit water pollution and maintain adequate hygiene.

Streptomyces coeruleorubidus as a potential biocontrol agent for Newcastle disease virus.

BACKGROUND: Newcastle disease virus (NDV) is a severe disease that affects domestic and wild birds. Controlled antibiotics derived from probiotics have been examined as prospective solutions for preserving seroconversion in NDV-vaccinated fowl. In this study, the secondary metabolite "telomycin" was extracted from Streptomyces coeruleorubidus (S. coeruleorubidus) isolated from Egypt's cultivated soil. The structure of telomycin was determined by the elucidation of spectroscopic analysis, including nuclear magnetic resonance (NMR) and mass spectrometry (MS) spectra, and comparison with the literature. The antiviral activity of the secondary metabolite was tested by checking its effect on NDV hemagglutination activity (HA). Moreover, HA of NDV was tested after inoculation of NDV (control) and a combination of telomycin and NDV in 10- days- specific pathogen-free embryonated chicken eggs (SPF-ECE) daily candling. Histopathological examination was performed for chorioallantoic membranes and liver of SPF-ECE. RESULTS: S. coeruleorubidus secondary metabolite "telomycin" showed complete hemagglutination inhibition (HI) activity of NDV strain (MN635617) with log106 infectivity titers (EID50/mL). The HA of NDV strain was 8 log2 and 9 log2 with 0.5% and 0.75% of chicken RBCs, respectively. Preserved structures of chorioallantoic-membranes (CAM) with dilated capillary networks were observed in the treated group inoculated with telomycin and NDV. Histological changes in SPF-ECE liver were examined after inoculation in ova to further characterize the telomycin effect. Telomycin and NDV mixture inoculated group showed preserved cytoarchitecture of hepatocytes with the presence of perivascular foci of lymphocytes. The group that was inoculated with telomycin alone showed normal histology of hepatic acini, central veins, and portal triads. CONCLUSION: S. coeruleorubidus telomycin is a promising bioactive agent that might be a biological weapon against a deadly chicken NDV that costs farmers a lot of money.

Assessment of the Diagnostic Potential of miR-29a-3p and miR-92a-3p as Circulatory Biomarkers in Acute Myeloid Leukemia.

BACKGROUND: Acute myeloid leukemia (AML) is a set of Myeloproliferative neoplasms that are identified by excessive growth of myeloid blasts and production of abnormal blood cells. AML is the most common type of acute leukemia that occurs in adults. In addition, AML progresses rapidly and is considered a fatal disease. Thus, there is an urgent need to find new targets for molecularly designed therapies. In This study, we evaluated the circulatory levels of microRNA-29a-3p (miR-29a-3p) and miR-92a-3p beside exploring the expression pattern of their target gene myeloid cell leukemia sequence1 (MCL1) to investigate the role of these molecules in AML pathophysiology and to assess their ability to diagnose AML patients. METHODS: 40 adult AML patients along with 20 healthy subjects were enrolled in this study. Plasma were separated from venous blood samples, collected on EDTA, of all individuals were used to assess circulating miRNAs' levels. In the meantime, total RNA was extracted from isolated leukocytes and was used to quantify target mRNA transcript levels. RESULTS: Our data revealed that the circulating levels of miR-29a-3p and miR-92a-3p exhibited significant reduction in 90% and 100% of AML patients, respectively, when compared to the control group (p<0.001). On the other hand, the transcript level of the target gene of these miRNAs, MCL1, showed a sharp increase in 77.5% (p<0.001) of AML patients, along with a negative correlation with its regulatory miRNAs, miR-29a-3p and miR-92a-3p. CONCLUSION: Our data validates the negative regulatory role of miR-29a-3p and miR-92a-3p to the expression levels of MCL1 in peripheral blood and indicates that these miRNAs can be used as non-invasive diagnostic markers. Furthermore, our study highlights the therapeutic potential of miR-29a-3p and miR-92a-3p to target and downregulate a very important gene (MCL1), which is highly implicated in the pathogenesis of AML.