Anand Prakash Maurya

The on-going coronavirus disease 19 (COVID-19) pandemic has caused a very high number of infections and deaths around the globe. The absence of vaccine/drugs to counter COVID-19 has scrambled scientific communities to repurpose available medicines/vaccines. As the virus is known to mutate, using the whole genome sequences, the transmission dynamics and molecular evolutionary models were evaluated for South Asian countries to determine the evolutionary rate of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Phylogenetic analyses were done using the data available on National Center for Biotechnology Information (NCBI). Different nucleotide substitution models and molecular evolutionary models were analyzed to see how SARS-CoV-2 was transmitted in the populations. Models for the viral ‘S’ and ‘N’ protein from selected strains were constructed, validated, and analyzed to determine the mutations and discover the potential therapeutics against this deadly viral disease. We found that the Hasegawa-Kishino-Yano (HKY) nucleotide substitution model was the best model with the lowest Bayesian information criterion (BIC) scores. Molecular clock RelTime analysis showed the evolutionary rate of SARS-CoV-2 substitutions in the genome was at 95% confidence interval, and heterogeneity was observed. Several mutations in the viral S-protein were found with one in the receptor-binding domain concerning SARS-CoV-2/Wuhan-1/S-Protein. Nucleocapsid protein also showed mutations in the strains from India and Sri Lanka. Our analysis suggests that SARS-CoV-2 is evolving at a diverse rate. The mutation leading to substitution in the nucleotide sequence occurred in the genome during the transmission of COVID-19 among individuals in the South Asian countries.

Bacterial pathogens resistant to multiple antibiotics are emergent threat to the public health which may evolve in the environment due to the co-selection of antibiotic resistance, driven by poly aromatic hydrocarbons (PAHs) and/or heavy metal contaminations. The co-selection of antibiotic resistance (AMR) evolves through the co-resistance or cross-resistance, or co-regulatory mechanisms, present in bacteria. The persistent toxic contaminants impose widespread pressure in both clinical and environmental setting, and may potentially cause the maintenance and spread of antibiotic resistance genes (ARGs). In the past few years, due to exponential increase of AMR, numerous drugs are now no longer effective to treat infectious diseases, especially in cases of bacterial infections. In this mini-review, we have described the role of co-resistance and cross-resistance as main sources for co-selection of ARGs; while other co-regulatory mechanisms are also involved with cross-resistance that regulates multiple ARGs. However, co-factors also support selections, which results in development and evolution of ARGs in absence of antibiotic pressure. Efflux pumps present on the same mobile genetic elements, possibly due to the function of Class 1 integrons (Int1), may increase the presence of ARGs into the environment, which further is promptly changed as per environmental conditions. This review also signifies that mutation plays important role in the expansion of ARGs due to presence of diverse types of anthropogenic pollutants, which results in overexpression of efflux pump with higher bacterial fitness cost; and these situations result in acquisition of resistant genes. The future aspects of co-selection with involvement of systems biology, synthetic biology and gene network approaches have also been discussed.

Plasmids are potent vehicles for spread of antibiotic resistance genes in bacterial populations and often persist in the absence of selection due to efficient maintenance mechanisms. We previously constructed non-conjugative high copy number plasmid vectors that efficiently displace stable plasmids from enteric bacteria in a laboratory context by blocking their replication and neutralising their addiction systems. Here we assess a low copy number broad-host-range self-transmissible IncP-1 plasmid as a vector for such curing cassettes to displace IncF and IncK plasmids. The wild type plasmid carrying the curing cassette displaces target plasmids poorly but derivatives with deletions near the IncP-1 replication origin that elevate copy number about two-fold are efficient. Verification of this in mini IncP-1 plasmids showed that elevated copy number was not sufficient and that the parB gene, korB, that is central to its partitioning and gene control system, also needs to be included. The resulting vector can displace target plasmids from a laboratory population without selection and demonstrated activity in a mouse model although spread is less efficient and requires additional selection pressure.

Background & objectives: Pseudomonas aeruginosa possessing chromosomally inducible bla _PDC along with other intrinsic mechanism causes infection with high mortality rate. It is difficult to detect inducible AmpC enzymes in this organism and is usually overlooked by routine testing that may lead to therapeutic failure. Therefore, three different inducers were evaluated in the present study to assess their ability of induction of bla _PDC in P. aeruginosa. Methods: A total of 189 consecutive Pseudomonas isolates recovered from different clinical specimens (November 2011-April 2013) were selected for the study. Isolates were screened with cefoxitin for AmpC β-lactamases and confirmed by modified three-dimensional extract test (M3DET). Inductions were checked using three inducers, namely, clavulanic acid, cefoxitin and imipenem along with ceftazidime. Molecular screening of AmpC β-lactamase genes was performed by PCR assay. Antimicrobial susceptibility and minimum inhibitory concentrations (MICs) were determined, and repetitive extragenic palindromic-PCR of all bla _PDC harbouring isolates was performed. Results: Inducible phenotype was observed in 42 (24.3%) of 97 (56%) isolates confirmed by M3DET. Among these, 22 isolates harboured chromosomal bla _PDC gene, and cocarriage of both chromosomal and plasmid-mediated bla _AmpC genes was observed in seven isolates. Cefoxitin-ceftazidime-based test gave good sensitivity and specificity for detecting inducible AmpC enzymes. Isolates harbouring bla _PDC showed high MIC against all tested cephalosporins and monobactam. DNA fingerprinting of these isolates showed 22 different clones of P. aeruginosa. Interpretation & conclusions: P. aeruginosa harbouring inducible (chromosomal) and plasmid-mediated AmpC β-lactamase is a matter of concern as it may limit therapeutic option. Using cefoxitin-ceftazidime-based test is simple and may be used for detecting inducible AmpC β-lactamase amongst P. aeruginosa.

The current study reports dissemination of highly stable bla OXA-10 family of beta lactamases among diverse group of nosocomial isolates of Gram-negative bacilli within a tertiary referral hospital of the northern part of India. In the current study, a total number of 590 Gram negative isolates were selected for a period of 1 year (i.e. 1st November 2011–31st October 2012). Members of Enterobacteriaceae and non fermenting Gram negative rods were obtained from Silchar Medical College and Hospital, Silchar, India. Screening and molecular characterization of β-lactamase genes was done. Integrase gene PCR was performed for detection and characterization of integrons and cassette PCR was performed for study of the variable regions of integron gene cassettes carrying bla OXA-10. Gene transferability, stability and replicon typing was also carried out. Isolates were typed by ERIC as well as REP PCR. Twenty-four isolates of Gram-negative bacilli that were harboring bla OXA-10 family (OXA-14, and OXA16) with fact that resistance was to the extended cephalosporins. The resistance determinant was located within class I integron in five diverse genetic contexts and horizontally transferable in Enterobacteriaceae, was carried through IncY type plasmid. MIC values were above break point for all the tested cephalosporins. Furthermore, co-carriage of bla CMY-2 was also observed. Multiple genetic environment of bla OXA-10 in this geographical region must be investigated to prevent dissemination of these gene cassettes within bacterial population within hospital settings.