![]() Genes revealed by whole genome sequencing and screening of phage collections will potentially yield new generations of antimicrobials. In our experience, some prophage regions are recalcitrant to cloning, most likely due to toxicity of the gene products to the bacterium. 21 Thus, the phage gene pool is larger and more diverse than the rest of the chromosome. Even closely related genomes appear to possess different sets of prophages. The majority of the sequenced bacterial genomes reveal the presence of one or more partial or complete prophage genomes. Today, technologies exist that allow cost-effective sequencing of hundreds of viral or bacterial genomes per year, and we can anticipate in the not-too-distant future further advances that might allow routine whole-genome screening of every pathogen encountered in a clinic 20 or on contaminated foodstuff. 19) or detection by receptor binding, to list just a few. 18), reporter bacteriophages (reviewed in ref. Newer developments include phage-amplification assays coupled with MALDI-MS, 16, 17 detection by lysis products (reviewed in ref. Phage-based detection methods confer a faster and more sensitive detection. Phage particles or their components have also been used successfully as detection agents for pathogens. Phage lytic enzyme application in food production has received intensive research interest, as they present highly effective and practical means of decontamination (reviewed in refs. 8 - 13 First phage preparations, such as Listex TM (Micreos) or ListShield TM (Intralytix), have received approval from regulatory agencies and are being used in food production. Numerous studies attest to the efficacy of selected phages or phage cocktails against foodborne pathogens, such as Listeria, Salmonella or E. 1 - 3, 7īesides human therapy approaches, whole-phage preparations have also been widely evaluated as biocontrol agents for food production. 7 Despite the apparent attractiveness of phages as antimicrobials, history is replete with false starts that have suppressed the field for decades at a time. Since their discovery around 1915–1917, phages have served as excellent research tools, 6 although the promise of their antibacterial potential has not been fully realized. This activity, combined with a decline in the discovery of new classes of antibiotics that are effective against these resistant bacteria in the past several decades, has brought about a renewed interest in alternatives to antibiotics, such as phages or phage-encoded lytic enzymes. Since the introduction of antibiotics in the 1940s to treat bacterial infections in humans and livestock, the widespread use, and in many instances misuse, has resulted in the current crisis with multi-drug resistant bacteria. Despite the fact that they are non-toxic to animals and plants, 1 phages are not as widely used for biocontrol and therapeutics as one would imagine. Isolation of new phages is rapid, facile and inexpensive, and there is an abundant supply of phages in nature, making them ideal weapons to combat bacterial infections. They are in a constant evolutionary arms race with host bacteria the survival of phages over millions of years is a testament to their ability to overcome bacterial resistance mechanisms by constantly evolving in parallel with their hosts. Based on our experience we propose several general considerations regarding sample quality, the choice of technology and a “blended approach” for generating reliable whole genome sequences of phages.īacteriophages (phages) are natural viral predators of bacteria. Here we describe conclusions drawn from our efforts in sequencing hundreds of bacteriophage genomes from a variety of Gram-positive and Gram-negative bacteria using Sanger, 454, Illumina and PacBio technologies. Sequencing phage genomes poses several challenges (1) obtaining pure phage genomic material, (2) PCR amplification biases and (3) complex nature of their genetic material due to features such as methylated bases and repeats that are inherently difficult to sequence and assemble. However, whole genome sequencing of bacterial viruses has not kept pace with this revolution, despite the fact that their genomes are orders of magnitude smaller in size compared with bacteria and other organisms. The 2nd and 3rd generation sequencing technologies, based on cloning-free, massively parallel sequencing, have enabled the generation of a deluge of genomic sequences of both prokaryotic and eukaryotic origin in the last seven years. The dawn of next generation sequencing technologies has opened up exciting possibilities for whole genome sequencing of a plethora of organisms.
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