Antimicrobial drug resistant features of Mycobacterium tuberculosis associated with treatment failure.

Tuberculosis stands as a prominent cause of mortality in developing countries. The treatment of tuberculosis involves a complex procedure requiring the administration of a panel of at least four antimicrobial drugs for the duration of six months. The occurrence of treatment failure after the completion of a standard treatment course presents a serious medical problem. The purpose of this study was to evaluate antimicrobial drug resistant features of Mycobacterium tuberculosis associated with treatment failure. Additionally, it aimed to evaluate the effectiveness of second line drugs such as amikacin, linezolid, moxifloxacin, and the efflux pump inhibitor verapamil against M. tuberculosis isolates associated with treatment failure. We monitored 1200 tuberculosis patients who visited TB centres in Lahore and found that 64 of them were not cured after six months of treatment. Among the M. tuberculosis isolates recovered from the sputum of these 64 patients, 46 (71.9%) isolates were simultaneously resistant to rifampicin and isoniazid (MDR), and 30 (46.9%) isolates were resistant to pyrazinamide, Resistance to amikacin was detected in 17 (26,5%) isolates whereas resistance to moxifloxacin and linezolid was detected in 1 (1.5%) and 2 (3.1%) isolates respectively. Among MDR isolates, the additional resistance to pyrazinamide, amikacin, and linezolid was detected in 15(23.4%), 4(2.6%) and 1(1.56%) isolates respectively. One isolate simultaneously resistant to rifampicin, isoniazid, amikacin, pyrazinamide, and linezolid was also identified. In our investigations, the most frequently mutated amino acid in the treatment failure group was Serine 315 in katG. Three novel mutations were detected at codons 99, 149 and 154 in pncA which were associated with pyrazinamide resistance. The effect of verapamil on the minimum inhibitory concentration of isoniazid and rifampicin was observed in drug susceptible isolates but not in drug resistant isolates. Rifampicin and isoniazid enhanced the transcription of the efflux pump gene rv1258 in drug susceptible isolates collected from the treatment failure patients. Our findings emphasize a high prevalence of MDR isolates linked primarily to drug exposure. Moreover, the use of amikacin as a second line drug may not be the most suitable choice in such cases.

Investigation of Three Morchella Species for Anticancer Activity Against Colon Cancer Cell Lines by UPLC-MS-Based Chemical Analysis.

In search of new anticancer agents, natural products including fungal compounds had been used as potential anticancer agents. The aim of this study was to investigate the anticancer activity of Morchella extracts against colon cancer cell line and UPLC-DAD-MS/MS analysis for the identification of compounds. The cytotoxic activity of the three Morchella species was examined for their anti-carcinogenic properties against the colon cancer cell lines. Phytochemical analyses were performed to screen Morchella for the presence of anti-cancerous compounds. All the fungal extracts inhibited the viability of colon cancer cells in a dose-dependent manner. Major compounds identified in Morchella included amino acid, fatty acid, sterol, flavonoid, peptide, glutamic acid, alkaloid, terpenoid, cyclopyrrolones, and coumarin. Several new compounds were detected among all the three Morchella extracts. In conclusion, all the fungal extracts showed potential inhibition of colon cancer cells and actively arrested the cell viability. It was concluded that the identified bioactive compounds might be the main constituents contributing to the anticancer activity of Morchella against human colon cancer cell lines. Thus, Morchella extracts are a potential source of bioactive compounds with cytotoxicity and could potentially be used as functional food supplements. Due to the nature of impressive findings, this investigation should be undertaken further to allow the studies to explore and develop a potential cytotoxic agents against colon cancer.

Antibacterial Activity of Different Extracts of Ascomata of Morchella conica and M. esculenta (Ascomycota) against Salmonella Species.

Antibiotic resistance has become a worldwide concern, as it has reduced the clinical efficiency of multiple antibiotics. As a result, screening of novel antibacterial substances for antimicrobial potential has increased. Mushrooms are widely known as a source of antimicrobial agents. The current study was designed to investigate the bacteriostatic and bactericidal effects of Morchella conica and M. esculenta against typhoidal and nontyphoidal Salmonella species. The sensitivity of S. typhi, S. paratyphi-A, and S. typhimurium was determined using an agar diffusion assay. The standard broth microdilution method was used to assess the minimal inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC). Bacterial growth and inhibition kinetics were evaluated spectrophotometrically. All of the mushroom extracts were able to inhibit the growth of typhoidal and nontyphoidal Salmonella species. Notably, all of the extracts possessed bacteriostatic effects (MIC: 3.33 ± 0.6 to 16.0 ± 0 mg/mL) and bactericidal effects (MBC: 8-16 mg/mL). The results showed statistically significant differences of antibacterial and bactericidal potential of mushroom extracts against the tested bacteria (P ≤ 0.05). Thus, extracts of Morchella species can be used as natural antibacterial pharmaceuticals. Further mycopharmacological studies must be performed to characterize their metabolites.

Investigation of Morchella esculenta and Morchella conica for their antibacterial potential against methicillin-susceptible Staphylococcus aureus, methicillin-resistant Staphylococcus aureus and Streptococcus pyogenes.

Antimicrobial resistance is an alarming problem, especially due to emergence of methicillin-resistance Staphylococcus aureus (MRSA). World Health Organization (WHO) has already listed MRSA as a top priority pathogen for the development of novel antibacterial agents. Presently, different therapeutic approaches against bacterial infections are in practice which includes targeting bacterial virulence factors, bacteriophage therapy, and manipulation of the microbiome. Natural products have been efficiently used for centuries to combat bacterial infections. Morchella is a natural fungal product which has been reported to possess broad-spectrum biological activities against bacterial infections. Hence, this study was aimed to evaluate the antibacterial efficacy of two macro-fungi against S. aureus, MRSA, and Streptococcus pyogenes (S. pyogenes). The antibacterial potential of both fungal extracts (Morchella esculenta and Morchella conica) was evaluated using disk diffusion and standard broth microdilution methods. The chemical compounds of both fungi were investigated using ultra-performance liquid chromatography mass spectroscopy (UPLC-MS) analysis. All fungal extracts inhibited growth of tested bacteria with inhibitory zone ranging from 10.66 ± 0.3 to 21.00 ± 1.5 mm. The minimum inhibitory concentration (MIC) of tested bacterial growth ranged from 03.33 to 16.0 mg/ml. It was noteworthy that Morchella extracts prevented S. aureus growth in a bactericidal manner with minimal bactericidal concentration (MBC) of 8-16 mg/ml. The extracts were also more effective against MRSA than currently available antibiotics. In conclusion, the growth inhibition of tested bacteria by fungal extracts revealed their potential as antibacterial agents and their compounds may be used as drug candidates.

From Molecular Pathology of COVID 19 to Nigella Sativum as a Treatment Option: Scientific Based Evidence of Its Myth or Reality.

COVID-19 virus is a causative agent of viral pandemic in human beings which specifically targets respiratory system of humans and causes viral pneumonia. This unusual viral pneumonia is rapidly spreading to all parts of the world, currently affecting about 105 million people with 2.3 million deaths. Current review described history, genomic characteristics, replication, and pathogenesis of COVID-19 with special emphasis on Nigella sativum (N. sativum) as a treatment option. N. sativum seeds are historically and religiously used over the centuries, both for prevention and treatment of different diseases. This review summarizes the potential role of N. sativum seeds against COVID-19 infection at levels of in silico, cell lines and animal models.

Antibacterial activity of Pakistani Beri honey compared with silver sulfadiazine on infected wounds: a clinical trial.

OBJECTIVE: To determine the antibacterial activity of Pakistani Beri honey in patients with infected wounds in comparison with silver sulfadiazine. METHOD: Inpatients with infected wounds at a tertiary care hospital were divided in to three equal-sized treatment groups. In Group A, patients were treated with non-Gamma irradiated Beri honey. In Group B, Gamma irradiated Beri honey was used, and in Group C silver sulfadiazine was used. Treatment was for a period of four weeks. Pus swabs were taken at day zero and weeks one and four of treatment. Bacteria were identified using the analytical profile index system API 20E, 20NE and API Staph and antimicrobial susceptibility was done as per the Clinical and Laboratory Standards Institute 2010 guidelines. RESULTS: A total of 90 patients with wounds (n=90) took part in the trial. Out of 90 wounds, 47% were post-traumatic and 37% were postoperative. Overall, average length, width and depth of Group A patients' wounds were significantly reduced (p<0.0001). Out of 144 pus swabs; 99 and 45 were Gram-negative rods and Gram-positive cocci, respectively. Among these Pseudomonas aeruginosa (n=25) and Staphylococcus aureus (n=38) were the major pathogens. Interestingly, bacterial load gradually decreased from baseline to week four due to non-Gamma irradiated Beri honey. Moreover, both the Gram-negative rods and Gram-positive cocci displayed 100% resistance to commonly used antibiotics; the most effective drugs were carbapenem and vancomycin. CONCLUSION: Pakistani Beri honey could be used as an alternative therapeutic option for the management of infected wounds.

Effect of different doses of Manuka honey in experimentally induced mouse typhoid.

Typhoid fever is a major cause of morbidity and mortality in the developing world. Data from World Health Organization (WHO) shows that 21 million cases of typhoid occur globally every year and over 200,000 die each year; most of them at a very young age. The situation in Pakistan is similar. Typhi and other typhoidal salmonellae have developed resistance to chloramphenicol and other first line anti-typhoid. There is a rapid increase in multi-drug resistance (MDR) throughout the world. There is an urgent need to find out alternative medicine to sort out this problem. This study was conducted to establish preventive as well as therapeutic potential of Manuka honey. A total of eighty pathogen free BALB/C mice between 8 weeks to 12 weeks of age, weighing 25-30 grams were taken and divided into 4 groups. Group A, B and C were infected through oral route with 10(8) colony forming unit (CFU) of Salmonella typhimurium ATCC 14028 to produce typhoid like disease in mice. Group A, which comprised of 20 mice was further divided in A1 and A2 given Manuka honey at a dose of 15ml/kg and 20 ml/kg respectively. Group B, which comprised of 20 mice was further divided in B1 and B2 was given Manuka honey at dose of 20ml/kg and 25ml/kg respectively. Clinical features of mouse typhoid, like body temperature, respiratory rate, number of stools and general behavior were recorded twice daily. Blood cultures of mice in different groups were taken at different days to evaluate the establishment of infection as well as to observe the therapeutic and preventive potential of Manuka honey in mouse typhoid. Fisher's Exact, Chi- Square and t-test were used to analyze the data. Significant association was observed in the ultimate fate of mice in Group A1 and Group A2 (P<0.001), showing that from a total of 20 mice in both groups, 10 mice fall in Group A1 of which 10 (100%) developed infection as it was not prevented by honey at a dose of 15ml/kg body weight (15.00±0.00) in Group A1 and ten mice fall in Group A2 of which 10(100%) did not developed an infection as it was prevented by honey at a dose of 20ml/kg body weight (20.00±0.00) in Group A2. Significant association was observed in the ultimate fate of mice in Group B1 and Group B2 (P<0.001) showing that from a total of 20 mice in both groups, 10 mice fall in Group B1 of which 10 (100%) had an infection, which was not treated by honey at a dose of 20 ml/kg body weight. Ten mice fall in Group B2 of which 10 (100%) had an infection, which was treated by honey at a dose of 25 ml/kg body weight (25.00±0.00). Results of the present study suggest that Manuka honey (UMF25±) has a potent anti-typhoid activity in vivo as well. There is an intense need for a carefully designed clinical trial in which this therapeutic potential of Manuka honey should be further evaluated. There is also need for the search of local honeys comparable to Manuka honey as a therapeutic option for typhoid fever.

Evaluation of the antibacterial activity of selected Pakistani honeys against multi-drug resistant Salmonella typhi.

BACKGROUND: The development of resistance to conventional anti-typhoid drugs and the recent emergence of fluoroquinolone resistance have made it very difficult and expensive to treat typhoid fever. As the therapeutic strategies become even more limited, it is imperative to investigate non-conventional modalities. In this context, honey is a potential candidate for combating antimicrobial resistance because it contains a broad repertoire of antibacterial compounds which act synergistically at multiple sites, thus making it less likely that the bacteria will become resistant. The in vitro antibacterial activity of 100 unifloral honey samples against a blood culture isolate of multi-drug resistant (MDR) Salmonella typhi were investigated. METHODS: All honey samples were evaluated for both total (acidity, osmolarity, hydrogen peroxide and non-peroxide activity) and plant derived non-peroxide antibacterial activity by agar well diffusion assay at 50% and 25% dilution in sterile distilled water and 25% in catalase solution. Manuka (Unique Manuka Factor-21) honey was used for comparison. The phenol equivalence of each honey sample from 2% to 7% (w/v) phenol was obtained from regression analysis. The antibacterial potential of each honey sample was expressed as its equivalent phenol concentration. The honey samples which showed antibacterial activity equivalent to or greater than manuka honey were considered therapeutically active honeys. RESULTS: Nineteen honey samples (19%) displayed higher hydrogen peroxide related antibacterial activity (16-20% phenol), which is more than that of manuka honey (21-UMF). A total of 30% of the honey samples demonstrated antibacterial activity between 11 and 15% phenol similar to that of manuka honey while 51% of the honey samples did not exhibit any zone of inhibition against MDR-S. typhi at 50% (w/v) dilution. None of the indigenous honey samples displayed non-peroxide antibacterial activity. Only manuka honey showed non-peroxide antibacterial activity at 25% dilution (w/v) in catalase solution. CONCLUSIONS: The honey samples which displayed antibacterial activity equal to or greater than manuka honey may be useful in the clinical conditions where higher hydrogen peroxide related antibacterial activity is required. Manuka honey, which is known to possess non-peroxide antibacterial activity, warrants further evaluation in a suitable typhoid animal model.

Anti bacterial activity of Nigella sativa against clinical isolates of methicillin resistant Staphylococcus aureus.

BACKGROUND: Methicillin resistant Staphylococcus aureus (MRSA) continues to be one of the commonest pathogens encountered in clinical as well as laboratory practice. It has become a major health problem worldwide. Newer antimicrobials/agents are urgently needed to combat this problem MRSA resistance to various anti-staphylococcal agents. In the back-drop of this difficult situation Nigella sativa commonly known as black seed (ethanolic extract) was aimed at to evaluate if it had any anti-staphylococcal activity. METHODS: The extract was prepared by reflux extraction method. Disc diffusion and in agar dilution methods were performed to assess the antibacterial activity. Staphylococcus aureus ATCC 25923 was used as the standard reference strain. RESULTS: All tested strains of MRSA were sensitive to N. sativa extract at a concentration of 4 mg/disc while the extract had an MIC range of 0.2-0.5 mg/ml. CONCLUSION: The results indicated that N. sativa has inhibitory effect on MRSA. This finding warrants necessity of further investigation of this product of folk medicine.