SummaryEscherichia coli biofilm consists of a bacterial colony embedded in a matrix of extracellular polymeric substances (EPS) which protects the microbes from adverse environmental conditions and results in infection. Besides being the major causative agent for recurrent urinary tract infections, E. coli biofilm is also responsible for indwelling medical device-related infectivity. The cell-tocell communication within the biofilm occurs due to quorum sensors that can modulate the key biochemical players enabling the bacteria to proliferate and intensify the resultant infections. The diversity in structural components of biofilm gets compounded due to the development of antibiotic resistance, hampering its eradication. Conventionally used antimicrobial agents have a restricted range of cellular targets and limited efficacy on biofilms. This emphasizes the need to explore the alternate therapeuticals like anti-adhesion compounds, phytochemicals, nanomaterials for effective drug delivery to restrict the growth of biofilm. The current review focuses on various aspects of E. coli biofilm development and the possible therapeutic approaches for prevention and treatment of biofilm-related infections.

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To effectively modulate the gene expression within an infected mammalian cell, the pathogen Mycobacterium tuberculosis would need to bring about epigenetic modifications at appropriate genomic loci. Working on this hypothesis, we show in this study that the mycobacterial protein Rv2966c is a 5-methylcytosine-specific DNA methyltransferase that is secreted out from the mycobacterium and gets localized to the nucleus in addition to the cytoplasm inside the host cell. Importantly, Rv2966c binds to specific DNA sequences, methylates cytosines predominantly in a non-CpG context and its methylation activity is positively influenced by phosphorylation. Interestingly, like the mammalian DNA methyltransferase, DNMT3L, Rv2966c can also interact with histone proteins. Ours is the first study that identifies a protein from a pathogenic bacteria with potential to influence host DNA methylation in a non-canonical manner providing the pathogen with a novel mechanism to alter the host epigenetic machinery. This contention is supported by repression of host genes upon M. tuberculosis infection correlated with Rv2966c binding and non-CpG methylation.

Summary Piwi-interacting RNAs (piRNAs) are important for genome regulation across metazoans, but their biogenesis evolves rapidly. In Caenorhabditis elegans , piRNA loci are clustered within two 3-Mb regions on chromosome IV. Each piRNA locus possesses an upstream motif that recruits RNA polymerase II to produce an ∼28 nt primary transcript. We used comparative epigenomics across nematodes to gain insight into the origin, evolution, and mechanism of nematode piRNA biogenesis. We show that the piRNA upstream motif is derived from core promoter elements controlling snRNA transcription. We describe two alternative modes of piRNA organization in nematodes: in C. elegans and closely related nematodes, piRNAs are clustered within repressive H3K27me3 chromatin, while in other species, typified by Pristionchus pacificus , piRNAs are found within introns of active genes. Additionally, we discover that piRNA production depends on sequence signals associated with RNA polymerase II pausing. We show that pausing signals synergize with chromatin to control piRNA transcription.

Background: Antimicrobial resistance (AMR) is a major global public health issue. In India, access to medicines is poorly regulated and therefore antibiotics in dairy cattle are commonly used by farmers without consulting with veterinarians. This study was conducted to understand practices and knowledge related to antibiotic use and AMR among dairy farmers and veterinary professionals in selected urban and peri-urban areas of India. Methods: A total of 28 focus group discussions with farmers and 53 interviews with veterinary professionals were carried out. Results: Mastitiswas identified as the main animal health challenge. Antibiotic consultation behavior of farmers depended on the availability of veterinarians. Except in Bangalore, farmers were found to often treat animals on their own. They were found unaware of the concept of AMR, but knew the importance of vaccination. Veterinarians included in the study had a good understanding of antibiotics, AMR, and zoonotic diseases. Conclusion: The knowledge level and practices observed in the study related to the use/ abuse of antibiotics can potentially increase the risk of development of AMR and its transfer in the community. Our findings can help support AMR-mitigation efforts in the country, including the design of better policies on antibiotic use in dairy.

Background: Livestock production, particularly the dairy sector, is important for food and nutritional wellbeing of communities in India, it supports livelihoods of many farmers, and contributes to the economy of the country. India is a high consumer of antibiotics and antimicrobial resistant (AMR) bacteria are a major public health concern. Objectives: Our objectives were to identify animal health and drug use practices that may contribute to emergence and spread of AMR in the country, review previous AMR-mitigation strategies, and discuss "theory of change" as an approach to informing the choice of interventions. Methods: We undertook a desk review of literature to identify practices with potential to contribute to emergence and spread of antimicrobial resistance in India. Searches were done in PubMed, Google scholar, and Google. Data were synthesized and discussed by themes. Results: Animal disease surveillance is less developed and infrastructure to support delivery of services is inadequate. Several groups are known to offer animal health services. The untrained "animal health workers" and para-veterinarians are more popular with farmers as they charge less for consultations (compared to veterinarians who are few and charge more). Over-the-counter access of antibiotics, without prescription, and direct marketing of drugs to farmers are common. Because of this, farmers are able to treat their animals and only consult when cases become non-responsive to treatment. Antibiotics are mostly used in management of mastitis cases. Drug withdrawal periods are rarely observed and occurrence of antibiotic-contaminated milk has been reported. Awareness on AMR is low and antimicrobial stewardship in livestock is yet to be developed. Initiatives such as the National programme for containment of AMR, National Action Plan on AMR, and the National Health policy point to government's commitment in addressing the problem of AMR in the country. Conclusion: Several animal health and drug use practices, with potential to cause AMR, have been described, and their contribution can be discussed further by engaging stakeholders in a "theory of change" exercise. Interventions that address AMR from the animal health perspective should be promoted, and incentives to increase their adoption explored.

Objective: The aim of this study was to purify and characterize bacteriocin from the soil isolate Bacillus subtilis GAS101, and to determine its antimicrobial as well as antibiofilm potential. The purified bacteriocin was further analyzed and evaluated for mammalian cell cytotoxicity and the possible mode of action. Material and Methods: Bacteriocin from B. subtilis GAS101 (an animal husbandry soil isolate) was partially purified and checked for antimicrobial and antibiofilm activity against gram-positive and gram-negative bacteria. The molecular weight of bacteriocin was determined using tricine SDS-PAGE gel. The stability of bacteriocin was investigated at various temperatures and pH levels, and its sensitivity towards 8 enzymes and 6 chemicals was determined. Cytotoxicity analysis was performed on a Vero cell line by a tetrazolium dye-based assay. Scanning electron microscopy (SEM) of bacteriocin-treated bacteria was carried out to determine the possible mode of action. Results: Bacteriocin from B. subtilis GAS101 was a potential inhibitor of both the indicator organisms (Staphylococcus epidermidis and Escherichia coli), and had a molecular weight of approximately 6.5 kDa. An in situ gel assay showed a zone of inhibition corresponding to the estimated protein band size. Bacteriocin was stable and showed antibacterial activity in broad ranges of temperature (30–121°C) and pH (2–12). It was sensitive to 4 proteolytic enzymes, which indicated its proteinaceous nature. Bacteriocin showed > 70% cell viability on the mammalian Vero cell line. SEM depicted that the bacteriocin was able to disrupt the bacterial cell membrane as its probable mode of action. Conclusion: Thermostable and pH-tolerant bacteriocin from B. subtilis GAS101, of about 6.5 kDa, showed broad-spectrum antimicrobial and antibiofilm activity.

A mammalian cell utilizes DNA methylation to modulate gene expression in response to environmental changes during development and differentiation. Aberrant DNA methylation changes as a correlate to diseased states like cancer, neurodegenerative conditions and cardiovascular diseases have been documented. Here we show genome-wide DNA methylation changes in macrophages infected with the pathogen M. tuberculosis. Majority of the affected genomic loci were hypermethylated in M. tuberculosis infected THP1 macrophages. Hotspots of differential DNA methylation were enriched in genes involved in immune response and chromatin reorganization. Importantly, DNA methylation changes were observed predominantly for cytosines present in non-CpG dinucleotide context. This observation was consistent with our previous finding that the mycobacterial DNA methyltransferase, Rv2966c, targets non-CpG dinucleotides in the host DNA during M. tuberculosis infection and reiterates the hypothesis that pathogenic bacteria use non-canonical epigenetic strategies during infection.