Low-Toxicity and High-Efficiency Streptomyces Genome Editing Tool Based on the Miniature Type V-F CRISPR/Cas Nuclease AsCas12f1.

Genome editing tools based on SpCas9 and FnCpf1 have facilitated strain improvements for natural product production and novel drug discovery in Streptomyces. However, due to high toxicity, their editing requires high DNA transformation efficiency, which is unavailable in most streptomycetes. The transformation efficiency of an all-in-one editing tool based on miniature Cas nuclease AsCas12f1 was significantly higher than those of SpCas9 and FnCpf1 in tested streptomycetes, which is due to its small size and weak DNA cleavage activity. Using this tool, in Streptomyces coelicolor, we achieved 100% efficiency for single gene or gene cluster deletion and 46.7 and 40% efficiency for simultaneous deletion of two genes and two gene clusters, respectively. AsCas12f1 was successfully extended to Streptomyces hygroscopicus SIPI-054 for efficient genome editing, in which SpCas9/FnCpf1 does not work well. Collectively, this work offers a low-toxicity, high-efficiency genome editing tool for streptomycetes, particularly those with low DNA transformation efficiency.

Antibacterial activity, computational analysis and host toxicity study of thymol-sulfonamide conjugates.

Methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin resistant Enterococcus faecalis (VRE) are notorious pathogenic multidrug resistant (MDR) bacteria in both hospital and community sectors, and today the first antibacterial drug sulfamethoxazole is ineffective. The monoterpene phenol, thymol was conjugated with seven sulfa drug derivatives individually, adopting the dye-azo synthesis protocol, and conjugates were characterized using spectral analysis techniques such as, UV, FTIR, MS, HPLC, 1H NMR, 13C NMR and SEM. Conjugates were assessed for antibacterial activity in vitro and in silico; the zone of inhibition, MIC and MBC values of each conjugate were determined against isolated MRSA and VRE strains from clinical samples. As 3-dimentional structures of dihydropteroate synthases (DHPSs) of targeted bacteria are not available in protein database, homology models of DHPS enzymes of both bacteria were generated and validated by Ramachandran plots. Seven conjugates were used as ligands in molecular docking against MRSA-DHPS and VRE-DHPS. Additionally bioinformatics tools, PASS prediction, Lipinski rules of five, computational LD50 value, toxicity class, HOMO, LUMO and EPS plots were carried out to assess standard drug-likeliness properties of conjugates. Zone size inhibition of the conjugate, 4b (thymol+sulfadiazine) against MRSA and VRE strains on agar plates were 20 and 40µg/mL as the lowest MIC and MBC values, respectively; while the reference antibiotic ampicillin had the lowest MIC and MBC values at 80 to 180µg/mL. In vitro host-toxicity testing was carried out with cultured human-lymphocytes from umbilical cord blood, and 4b was broadly non-toxic to human cells at 15,000mg/L. Thus, 4b could be promoted a newer antibacterial, against gruesome MDR bacteria.

Antiproliferative and hepatoprotective activity of metabolites from Corynebacterium xerosis against Ehrlich Ascites Carcinoma cells.

OBJECTIVE: To find out the effective anticancer drugs from bacterial products, petroleum ether extract of Corynebacterium xerosis. METHODS: Antiproliferative activity of the metabolite has been measured by monitoring the parameters like tumor weight measurement, tumor cell growth inhibition in mice and survival time of tumor bearing mice, etc. Hepatoprotective effect of the metabolites was determined by observing biochemical, hematological parameters. RESULTS: It has been found that the petroleum ether extract bacterial metabolite significantly decrease cell growth (78.58%; P<0.01), tumor weight (36.04 %; P<0.01) and increase the life span of tumor bearing mice (69.23%; P<0.01) at dose 100 mg/kg (i.p.) in comparison to those of untreated Ehrlich ascites carcinoma (EAC) bearing mice. The metabolite also alters the depleted hematological parameters like red blood cell, white blood cell, hemoglobin (Hb%), etc. towards normal in tumor bearing mice. Metabolite show no adverse effect on liver functions regarding blood glucose, serum alkaline phosphatases, glutamic pyruvic transaminase, glutamic oxaloacetic transaminase activity and serum billirubin, etc. in normal mice. Histopathological observation of these mice organ does not show any toxic effect on cellular structure. But in the case of EAC bearing untreated mice these hematological and biochemical parameters deteriorate extremely with time whereas petroleum ether extract bacterial metabolite receiving EAC bearing mice nullified the toxicity induced by EAC cells. CONCLUSION: Study results reveal that metabolite possesses significant antiproliferative and hepatoprotective effect against EAC cells.

Antiproliferative and hepatoprotective activity of metabolites from Corynebacterium xerosis against Ehrlich Ascites Carcinoma cells

Objective: To find out the effective anticancer drugs from bacterial products, petroleum ether extract of Corynebacterium xerosis.Methods:parameters like tumor weight measurement, tumor cell growth inhibition in mice and survival time of tumor bearing mice, etc. Hepatoprotective effect of the metabolites was determined by observing biochemical, hematological parameters.Results:It has been found that the petroleum ether extract bacterial metabolite significantly Antiproliferative activity of the metabolite has been measured by monitoring the decrease cell growth (78.58%; P<0.01), tumor weight (36.04 %; P<0.01) and increase the life span of tumor bearing mice (69.23%; P<0.01) at dose 100 mg/kg (i.p.) in comparison to those of untreated Ehrlich ascites carcinoma (EAC) bearing mice. The metabolite also alters the depleted hematological parameters like red blood cell, white blood cell, hemoglobin (Hb%), etc. towards normal in tumor bearing mice. Metabolite show no adverse effect on liver functions regarding blood glucose, serum alkaline phosphatases, glutamic pyruvic transaminase, glutamic oxaloacetic transaminase activity and serum billirubin, etc. in normal mice. Histopathological observation of these mice organ does not show any toxic effect on cellular structure. But in the case of EAC bearing untreated mice these hematological and biochemical parameters deteriorate extremely with time whereas petroleum ether extract bacterial metabolite receiving EAC bearing mice nullified the toxicity induced by EAC cells.Conclusion:Study results reveal that metabolite possesses significant antiproliferative and hepatoprotective effect against EAC cells.

Antiproliferative and hepatoprotective activity of metabolites from Corynebacterium xerosis against Ehrlich Ascites Carcinoma cells

<p><b>OBJECTIVE</b>To find out the effective anticancer drugs from bacterial products, petroleum ether extract of Corynebacterium xerosis.</p><p><b>METHODS</b>Antiproliferative activity of the metabolite has been measured by monitoring the parameters like tumor weight measurement, tumor cell growth inhibition in mice and survival time of tumor bearing mice, etc. Hepatoprotective effect of the metabolites was determined by observing biochemical, hematological parameters.</p><p><b>RESULTS</b>It has been found that the petroleum ether extract bacterial metabolite significantly decrease cell growth (78.58%; P<0.01), tumor weight (36.04 %; P<0.01) and increase the life span of tumor bearing mice (69.23%; P<0.01) at dose 100 mg/kg (i.p.) in comparison to those of untreated Ehrlich ascites carcinoma (EAC) bearing mice. The metabolite also alters the depleted hematological parameters like red blood cell, white blood cell, hemoglobin (Hb%), etc. towards normal in tumor bearing mice. Metabolite show no adverse effect on liver functions regarding blood glucose, serum alkaline phosphatases, glutamic pyruvic transaminase, glutamic oxaloacetic transaminase activity and serum billirubin, etc. in normal mice. Histopathological observation of these mice organ does not show any toxic effect on cellular structure. But in the case of EAC bearing untreated mice these hematological and biochemical parameters deteriorate extremely with time whereas petroleum ether extract bacterial metabolite receiving EAC bearing mice nullified the toxicity induced by EAC cells.</p><p><b>CONCLUSION</b>Study results reveal that metabolite possesses significant antiproliferative and hepatoprotective effect against EAC cells.</p>

Hepatoprotective effect of acetone semicarbazone on Ehrlich ascites carcinoma induced carcinogenesis in experimental mice.

OBJECTIVE: To determine the hepatoprotective effect of acetone semicarbazone (ASC) in vivo in normal and Ehrlich ascites carcinoma (EAC) bearing male Swiss albino mice. METHODS: Drug-induced changes in biochemical and behavioral parameters at dose of 2.0 mg/kg body weight for 14 d and nullifying the toxicity induced by EAC cells were studied. The histopathology studies of the protective effects of ASC on vital organs were also assessed. RESULTS: The administration of ASC made insignificant changes in body weight and behavioral (salivation, diarrhea, muscular numbness) changes during treatment period due to minor toxicity were minimized after the treatment in normal mice. The biochemical parameters, including serum glutamate pyruvate transaminase, glutamate oxaloactate transaminase, alkaline phosphatase, serum glucose, cholesterol, urea, triglyceride and billirubin changed modestly in normal mice receiving ASC. Though the treatment continued, these values gradually decreased to normal level after the treatment. In EAC bearing mice, the toxic effects due to EAC cells in all cases were nullified by treatment with the ASC. Significant abnormalities were not detected in histology of the various organs of the normal mice treated with ASC. CONCLUSIONS: ASC can, therefore, be considered safe in formulating novel anticancer drug, as it exhibits strong protective effect against EAC cell bearing mice.

Hepatoprotective effect of acetone semicarbazone on Ehrlich ascites carcinoma induced carcinogenesis in experimental mice

Objective:To determine the hepatoprotective effect of acetone semicarbazone (ASC) in vivo in normal and Ehrlich ascites carcinoma (EAC) bearing male Swiss albino mice. Methods:Drug-induced changes in biochemical and behavioral parameters at dose of 2.0 mg/kg body weight for 14 d and nullifying the toxicity induced by EAC cells were studied. The histopathology studies of the protective effects of ASC on vital organs were also assessed. Results:The administration of ASC made insignificant changes in body weight and behavioral (salivation, diarrhea, muscular numbness) changes during treatment period due to minor toxicity were minimized after the treatment in normal mice. The biochemical parameters, including serum glutamate pyruvate transaminase, glutamate oxaloactate transaminase, alkaline phosphatase, serum glucose, cholesterol, urea, triglyceride and billirubin changed modestly in normal mice receiving ASC. Though the treatment continued, these values gradually decreased to normal level after the treatment. In EAC bearing mice, the toxic effects due to EAC cells in all cases were nullified by treatment with the ASC. Significant abnormalities were not detected in histology of the various organs of the normal mice treated with ASC. Conclusions: ASC can, therefore, be considered safe in formulating novel anticancer drug, as it exhibits strong protective effect against EAC cell bearing mice.

Hepatoprotective effect of acetone semicarbazone on Ehrlich ascites carcinoma induced carcinogenesis in experimental mice

<p><b>OBJECTIVE</b>To determine the hepatoprotective effect of acetone semicarbazone (ASC) in vivo in normal and Ehrlich ascites carcinoma (EAC) bearing male Swiss albino mice.</p><p><b>METHODS</b>Drug-induced changes in biochemical and behavioral parameters at dose of 2.0 mg/kg body weight for 14 d and nullifying the toxicity induced by EAC cells were studied. The histopathology studies of the protective effects of ASC on vital organs were also assessed.</p><p><b>RESULTS</b>The administration of ASC made insignificant changes in body weight and behavioral (salivation, diarrhea, muscular numbness) changes during treatment period due to minor toxicity were minimized after the treatment in normal mice. The biochemical parameters, including serum glutamate pyruvate transaminase, glutamate oxaloactate transaminase, alkaline phosphatase, serum glucose, cholesterol, urea, triglyceride and billirubin changed modestly in normal mice receiving ASC. Though the treatment continued, these values gradually decreased to normal level after the treatment. In EAC bearing mice, the toxic effects due to EAC cells in all cases were nullified by treatment with the ASC. Significant abnormalities were not detected in histology of the various organs of the normal mice treated with ASC.</p><p><b>CONCLUSIONS</b>ASC can, therefore, be considered safe in formulating novel anticancer drug, as it exhibits strong protective effect against EAC cell bearing mice.</p>