Anti-Infective Agents - Volume 19, Issue 2, 2021

Metformin is a dimethyl biguanide known for its anti-hyperglycemic effects since 1922 and, thus, has been used in the treatment of Type 2 Diabetes Mellitus. Apart from this, its function as an anti-inflammatory, anti-oxidant, anti-thrombotic, and anti-microbial agent has also been studied largely. Its effect via inhibition of respiratory complex 1 and glycerol-3-phosphate dehydrogenase (GPD2) in the mitochondria and inhibition of the mammalian target of rapamycin complex 1 (mTORC1) and activation of Adenosine Mono Phosphate dependent kinase (AMPK) in the cytoplasm are its already known mechanisms of action. With the newer proposed effects on endosome/ lysosome regulation via Na+/H+ exchangers and V-ATPase, thereby affecting autophagy, coupled with its use in ARDS owing to its immunomodulatory effects and anti-viral action, the use of metformin against the novel Coronavirus is hypothesized.

Coronavirus Disease-2019 (COVID-19), emerged in Wuhan (China) during December 2019, has now grown into a global scale pandemic that is influencing the civilization of the whole world. As of 25th March 2020, the World Health Organization (WHO) has released 65 situation reports starting from 23rd January 2020 to 26th March 2020 regarding this new disease. The virus has now been identified and named as Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-- CoV-2). Total confirmed cases documented across the globe have reached up to 462,684 including 20,834 confirmed deaths from this disease as of 26th March 2020. This study focuses on the essential developments, or response procedures opted by the various countries and presents an elaborative account of the various steps taken by the Pakistan government to respond to this dynamic situation. As a third world country, the fall out of these actions can have severe repercussions. This study attempted to highlight some of the peculiar trials Pakistan is facing in these challenging times and discusses the effect of COVID-19 from the perspective of the third world nation. We believe that this study will provide a realistic picture of the current scenario taking place in Pakistan and will assist the government and concerned institutions/agencies in developing better plans for effectively dealing with this COVID-19 virus.

Background: Acanthamoeba belongs to the free-living amoebae genus that causes a widespread infection with different severities in humans and animals. Treatment of these opportunistic protozoan infections is still not safe or effective. The aim of the present study was to evaluate the effect of spring extract of Artemisia aucheri on Acanthamoeba in vitro. Methods: Trophozoites and cysts of Acanthamoeba were cultured on non-nutrient agar plates containing bacteria Escherichia coli. Macrophage cells were cultured in RPMI-1640 medium. Different concentrations of spring extracts of A. aucheri were added to cultivated parasites and macrophages and additionally calculated to determine the 50% inhibitory concentration (IC50) for parasites and 50% cytotoxic concentration (CC50) for macrophages. Then, the viability of parasites were evaluated using counting assay. MTT assay was carried out on macrophage cells based on growth inhibition. Furthermore, in order to determine the possible induction of apoptosis in cysts of Acanthamoeba, flow cytometry method was employed using annexin staining. Results: The effect of A. aucheri on trophozoites and cysts of Acanthamoeba was effective and had a reverse relationship with its concentration. The IC50 value of spring extract on trophozoites and cysts of the parasite was estimated to be 200 and 333 μg/ml at 24 h, respectively. According to the results of flow cytometry, the spring A. aucheri extract caused about 20% apoptosis. Conclusion: Regarding the above results, it can be concluded that spring A. aucheri extract was effective against trophozoite and cyst forms of Acanthamoeba in vitro. Therefore, this plant could be an appropriate herbal drug candidate for the treatment of Acanthamoeba infections.

Background: Decaprenylphosphoryl-β-D-ribose epimerase (DprE1), a flavoprotein enzyme engaged in the biosynthesis of decaprenylphosphoryl-β-D-arabinofuranose (DPA), is the only contributor of arabinose residues which is fundamental for the mycobacterium cell wall constituents. DprE1 is an interesting target for antitubercular agent and has been exploring to develop potential chemical entities as antitubercular agents. Objective: The objective of the study is the development of novel antitubercular agents targeting Mtb Decaprenylphosphoryl-β-D-ribose epimerase (DprE1). Methods: A series of isoxazole encompassed 1, 2, 4-triazoles were designed based on the antitubercular potential of triazoles and structural features of DprE1 inhibitors. Designed 1, 2, 4- triazoles were synthesized and characterized by spectral studies. The in vitro anti-TB activity of the compounds was screened against Mycobacterium tuberculosis H37Rv strain by Microplate Almar Blue Assay and in vitro cytotoxicity against normal cell lines by MTT assay. Molecular docking study was carried out on DprE1 enzyme to understand designed compounds interactions with amino acid residues at the active site. Results: Antitubercular activity data revealed that eight compounds (6d, 6e,7d, 7e, 10d, 10e, 11d and 11e) have shown promising antitubercular activity with minimum inhibitory concentration at 1.6μg/mL. Cytotoxicity data of anti-TB active compounds demonstrate good safety profile on normal cell lines. Conclusion: Eight compounds have shown promising antitubercular activity with good safety profile on normal cell lines. Molecular docking study revealed that the synthesized compounds have shown non-covalent interactions with amino acid residues of DprE1 enzyme.

Background: Fungal infections are opportunistic infections that become a serious problem for human health. Objective: Considering the antifungal potential of the triazole nucleus, the study was carried out with the objective of synthesizing some novel triazole derivatives with antifungal potential. Methods: 1,2,4-triazole derivatives were synthesized via a two-step reaction (reported earlier). The first step involves the reaction of substituted benzoic acid with thiocarbohydrazide to form 4- amino-3-(substituted phenyl)-5-mercapto-1, 2, 4-triazole derivatives (1a-1k) while in the second step, synthesized compounds (1a-1k) were then subsequently treated with substituted acetophenone to yield substituted (4-methoxyphenyl-7H-[1, 2, 4] triazolo [3, 4-b][1,3,4] thiadiazine derivatives (2a-2k). All synthesized compounds were characterized by IR, 1H NMR, and Mass spectral data analysis and were screened for their antifungal properties against different fungal strains i.e. Candida tropicalis (ATCC-13803, ATCC-20913), Candida albicans (ATCC-60193), Candida inconspicua (ATCC-16783) and Candida glabrata (ATCC-90030, ATCC-2001). Results: Compound 2d displayed better percentage inhibition (26.29%, 24.81%) than fluconazole (24.44%, 22.96%) against ATCC-16783, ATCC-2001 fungal strains respectively at 100μg/ml. Compound 2f also displayed better percentage inhibition (28.51%) against ATCC-90030 as compared to fluconazone (27.4%) at 200 μg/ml. Similarly, compounds 2e and 2j also exhibited better antifungal properties than fluconazole at 200μg/ml. Compound 2e was found most potent against ATCC-13803 (30.37%) and ATCC-90030 (30.37%) fungal strains as compared to fluconazole (28.14%, 27.4%) at 200 μg/ml respectively whereas compound 2j exhibited better antifungal activity (28.51%) against ATCC-60193 than fluconazole (27.7%) at 200 μg/ml. Conclusion: The results were in accordance with our assertions for triazole derivatives, as all compounds displayed moderate to good antifungal activity.

Background: To date, there is no consensus on the best combination of direct-acting antivirals to treat hepatitis C virus in kidney transplant recipients. Objective: This study aims to analyze the efficacy of a combination of sofosbuvir and ledipasvir regimen for the treatment of hepatitis C virus infected kidney transplant patients. Methods: A cross-sectional study was conducted in a nephrology clinic and the Nephrology Center in Basrah Teaching Hospital from June 2015 to June 2018. Ledifos (90 mg Ledipasvir and 400 mg Sofosbuvir fixed-dose) was given as a single daily dose to all the participants for 12 weeks. Response for therapy was tested by a follow up hepatitis C virus load at the end of 12 weeks and after 24 weeks. The sustained virological response was defined as a negative viral load of hepatitis C virus (aviremia) at the end of therapy. This study was done according to the Helsinki Congress. Results: A total of 60 (16 females) patients with renal transplantation and hepatitis C virus infection were included. The mean age was 40±6.2 years. A sustained virological response was observed in all of the patients who received Ledifos after 12 and 24 weeks of therapy for all genotypes (1a, 1b and 4); p= 0.0001. Genotype 1a was more prevalent among males, in about 34 (56.6%) of the patients; p= 0.0001, and it was the most common genotype that tested negative serologically, 11 (18.3%). Conclusion: Ledifos therapy is an effective and safe option for the treatment of hepatitis C virus infection in the post-renal transplant setting.

Background: Infectious diseases, whether intracellular or extracellular infections, biofilm- mediated, or medical device-associated, have always been a global public health problem, causing millions of deaths each year. The aim of this study was to evaluate the antibiofilm activity of ZnO/ZeoNC against K. pneumoniae along with the biocompatibility of the nanocomposite in vivo model. Objective: The formation of biofilm by K. pneumoniae in the catheter-associated urinary tract causes a nosocomial infection. In this regard, antimicrobial nanomaterials have emerged as potent effective agents against biofilm formation. Nevertheless, nanoparticles have already been a challenge with possible side effects such as inflammation. The ZnO/ZeoNC may exhibit anti-biofilm property with minimal adverse effects. Methods: The biofilm formation of K. pneumoniae strains was exposed to ZnO/ZeoNC and then SEM imaging was performed for morphological investigation of bacteria in biofilm state. The response to ZnO/ZeoNC embedded polyethylene tube of the tissue of mice was also analyzed during the 30-day experiment. Results: The results of this study showed that ZnO/ZeoNC has significant antibiofilm activity against K. pneumoniae strains in its sublethal doses. The ZnO/ZeoNC also caused deformation in K. pneumoniae biofilm. In addition, ZnO/ZeoNC also reduced inflammatory response in cell tissue of rats subjected to polyethylene tube. Conclusion: ZnO/ZeoNC can be used potentially against the infections caused by K. pneumonia biofilm without any irritability on the biotic surface such as the urinary tract.

Introduction: The biofilm formation by Pseudomonas aeruginosa seems to protect the bacteria from antibiotics since these entities are highly resistant to such antimicrobial agents. The aim of this study was to investigate the role of Lactobacillus salivarus, Lactobacillus plantarum supernatants and Cu^II Schiff base complex in eliminating planktonic cells and biofilm of P. aeruginosa. Methods: One hundred specimens of blood, urine, cerebrospinal fluid, respiratory samples, and wound swabs were collected from patients attending three hospitals in Mashhad. All specimens were identified by biochemical tests. The susceptibility of the isolates to the conventional antibiotics was assessed using disk diffusion method. The biofilm formation ability of P. aeruginosa isolates was evaluated by crystal violet assay and confirmed using PCR. The anti-planktonic and antibiofilm ability of L. salivarus, L. plantarum supernatants and Cu^II Schiff base complex was evaluated separately in P. aeruginosa isolates. Results: The highest and lowest resistance rates were detected in Cefazoline (95%) and cefepime (23%), respectively. The thickest biofilm was produced by 8% of P. aeruginosa isolates, 9% and 83% of the isolates were considered as moderate and weak biofilm producers, respectively. The rhlR and lasR genes were reported in 100% of the isolates, but the algD gene existed in 92% of them. Conclusion: Particularly, the Cu^II Schiff base complex could affect both planktonic and biofilm cells by the lowest concentration in comparison of probiotic supernatants. L. plantarum supernatant inhibited planktonic cells at a lower concentration than L. salivarius. Also, L. salivarius showed better antibiofilm activity than another probiotic in lower doses of supernatant. Unlike that, these compounds have not completely eliminated biofilm cells, but only reduced the biofilm formation. Metal Schiff base complex and Lactobacillus supernatants is a potent antimicrobial agent against Pseudomonas aeruginosa biofilm cells.

Background: Flavonoids are among the most abundant plant secondary metabolites and have been shown to have natural abilities as microbial deterrents and anti-infection agents in plants. Naringenin as one of the flavonoids and its derivatives have been reported to exhibit antimicrobial activity. Aim: The aim of the study was to evaluate synthesized novel naringenin derivatives (ND-1 to ND- 12) substituted at 3-position with bulky substituent by using the grinding technique (Claisen- Schmidt reaction) as antimicrobial agents. Methods: Synthesized naringenin derivatives were evaluated for in-vitro antimicrobial activity by “Agar disc diffusion method”. Results: Novel naringenin derivatives showed mild to moderate antimicrobial activity with respect to standard drugs against two gram-positive, two gram-negative bacteria and two fungal strains. The substitution of naringenin derivatives at position 3 with substituted phenyl rings showed variation in activity as chloro, nitro and hydroxyl-substituted phenyl ring showed potent activity while methoxy substituted phenyl ring impede the activity. Conclusion: In conclusion, on the basis of the above findings, the substituted naringenin scaffolds may be selected as a skeleton for the development of flavonoid structurally-related compounds having antimicrobial activity.

Background: Campylobacter jejuni is the most common cause of enteric infections, particularly among children, resulting in severe diarrhea. Increasing drug resistance of this bacterium against standard antibiotics favors investigations into additional anti-Campylobacter medications that are already used to overcome effects on enteric infections. Methods: Anti-bacterial activity using well diffusion assay of seventeen fungal extracts was tested against C. jejuni NCTC11168. The obtained results of antibacterial screening showed that different tested fungal isolates have different antimicrobial activities, where Hericium erinaceus extract was the highest activity against the tested bacterium. Results: Fractionation pattern has been done by column chromatography. Furthermore, purity was estimated by thin layer chromatography (TLC). Minimal inhibitory concertation (MIC) for the purified compound was 7.81μg/ml. Cytotoxicity for the purified compound was evaluated to be 170μg/ml. ¹HNMR, IR and GC-Mass were performed for illustration of the suggested structure of the bioactive compound purified from H. erinaceus. Conclusion: The data presented here suggested that H. erinaceus could potentially be used in modern applications aimed at the treatment or prevention of Campylobacter jejuni infection.

Background: COVID-19 caused by SARS-CoV-2 virus which originated in Wuhan and quickly spread across various countries has taken the form of a pandemic. It is now a major health concern worldwide and finding a solution to this problem is of utmost importance. Understanding its origin, transmission, and interaction with different compounds is essential to find probable inhibitors. Objective: The objective of our study was to search for potential inhibitors of the main protease of SARS-CoV-2 and to assess their drug-like properties. Methods: In our study, 1909 ligands were filtered through the Lipinski filter and their ADMET properties along with mutagenic nature were analyzed. They were screened for inhibitory activity against the Main Protease of SARS-CoV-2 using BIOVIA Discovery studio. Results: After virtual high throughput screening, two compounds- apigenin and N-(4-bromophenyl)- 7-hydroxy-2-iminochromene-3-carboxamide were found to have promising binding energies as well as –CDOCKER energy scores compared to the reported inhibitor. Conclusion: Apigenin seems to be a potential candidate against the main protease of SARS-CoV-2 and must be considered for further experiments.

Aim: This study was aimed to evaluate the emergence of third generation cephalosporin resistant Escherichia coli isolated from the culture of various biological fluids of infants in a tertiary care hospital. Background: Resistance to commonly used antibiotics is consistently increasing during the last decades and it was found varying with time and geographical location. Objectives: To determine the prevalence of third generation cephalosporin sensitivity pattern of E. coli isolated from urine, stool, and throat swab culture from laboratory records. Methods: The retrospective study included the laboratory report of antibiotic sensitivity conducted on infants specimens during the period of 2 years. Third generation cephalosporin sensitivity patterns of E. coli isolated from urine, stool, and throat swab culture were collected and data were statistically analyzed. Results: Reports of 351 samples with positive E. coli were included in the study. More samples were analyzed from infants above 6 months of age with female dominance (184/351). Among the specimens tested, urine samples were highest 128/351 (35.0%). Cephalosporin resistant strain was found in 243/351 samples (69.2%) (p=0.0463) with 218/243 (89.7%) isolated from infants above 6 months of age and 25/243 (10.2%) were from infants below 6 months of age (p=0.038). The frequency of cephalosporin resistant E. coli in urine specimens of male and female infants was significant (p=0.0001). Conclusion: Third generation cephalosporin resistant E. coli strain was found in 69.2% of isolates and more in urine samples from female infants above 6 months of age. The clinicians should consider the emergence of cephalosporin resistant E. coli while selecting antibiotics as empirical treatment.

Background: The emergence of drug resistance is the biggest threat; consequently, novel antimicrobial agents from natural sources is the need of the hour. This study was designed to isolate and evaluate a bioactive compound from actinomycete against human pathogens. Methods: In search of a new antimicrobial compound with enhanced potency, several actinomycetes were isolated and screened for their anidermatophytic acitivity. The bioactive compound was extracted and purified by chromatographic technique. The structure of the extracted bio active compound was elucidated by mass and NMR data. The synthesized bioactive compound was further tested for its antidermatophytic activity by the micro broth dilution method. Results: Streptomyces ARITM 03 was isolated that exhibited antifungal activity against human pathogenic fungi by primary and secondary screening methods. The MIC of bioactive compound was found to be 1250, 312.5 and 2500 μg/ml against tested fungal pathogens A. niger, C. albicans and M. canis, respectively. Chromatography analysis of active metabolite showed a single spot having Rf value of 0.74 and FT-IR spectrum displaying the presence of OH, CO=NH2 functional amide group, and C=O keto groups in the structure. The mass and NMR spectra revealed the molecular formula C26H44N4O6 of bioactive compound. The azole derivative showed an acceptable docking score and exhibited a greater zone of inhibition in comparison to the other compounds. Conclusion: The results presented in this paper provided an insight into the capability of Streptomyces sp. ARITM03 as a potential source of bioactive secondary metabolites compound a molecule to develop other azoles, to be used clinically to overcome adverse effects like gynecomastia and hepatotoxicity due to extensive use of current azole antifungal agents.