The lion's share of bacteria in various environments cannot be cloned in the laboratory and thus cannot be sequenced using existing technologies. A major goal of single-cell genomics is to complement gene-centric metagenomic data with whole-genome assemblies of uncultivated organisms. Assembly of single-cell data is challenging because of highly non-uniform read coverage as well as elevated levels of sequencing errors and chimeric reads. We describe SPAdes, a new assembler for both single-cell and standard (multicell) assembly, and demonstrate that it improves on the recently released E+V-SC assembler (specialized for single-cell data) and on popular assemblers Velvet and SoapDeNovo (for multicell data). SPAdes generates single-cell assemblies, providing information about genomes of uncultivatable bacteria that vastly exceeds what may be obtained via traditional metagenomics studies. SPAdes is available online ( http://bioinf.spbau.ru/spades ). It is distributed as open source software.

SPAdes, a new genome assembly algorithm and its applications to single-cell sequencing ( 7th Annual SFAF Meeting, 2012)

Accurate estimates of virus mutation rates are important to understand the evolution of the viruses and to combat them. However, methods of estimation are varied and often complex. Here, we critically review over 40 original studies and establish criteria to facilitate comparative analyses. The mutation rates of 23 viruses are presented as substitutions per nucleotide per cell infection (s/n/c) and corrected for selection bias where necessary, using a new statistical method. The resulting rates range from 10 8 to10 6 s/n/c for DNA viruses and from 10 6 to 10 4 s/n/c for RNA viruses. Similar to what has been shown previously for DNA viruses, there appears to be a negative correlation between mutation rate and genome size among RNA viruses, but this result requires further experimental testing. Contrary to some suggestions, the mutation rate of retroviruses is not lower than that of other RNA viruses. We also show that nucleotide substitutions are on average four times more common than insertions/deletions (indels). Finally, we provide estimates of the mutation rate per nucleotide per strand copying, which tends to be lower than that per cell infection because some viruses undergo several rounds of copying per cell, particularly double-stranded DNA viruses. A regularly updated virus mutation rate data set will be available at www.uv.es/rsanjuan/virmut.

https://www.uv.es/rasanve2/Sanjuan%20JVirol%202010.pdf

Viral mutation rates.

The acronym ESKAPE includes six nosocomial pathogens that exhibit multidrug resistance and virulence: Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. Persistent use of antibiotics has provoked the emergence of multidrug resistant (MDR) and extensively drug resistant (XDR) bacteria, which render even the most effective drugs ineffective. Extended spectrum β-lactamase (ESBL) and carbapenemase producing Gram negative bacteria have emerged as an important therapeutic challenge. Development of novel therapeutics to treat drug resistant infections, especially those caused by ESKAPE pathogens is the need of the hour. Alternative therapies such as use of antibiotics in combination or with adjuvants, bacteriophages, antimicrobial peptides, nanoparticles, and photodynamic light therapy are widely reported. Many reviews published till date describe these therapies with respect to the various agents used, their dosage details and mechanism of action against MDR pathogens but very few have focused specifically on ESKAPE. The objective of this review is to describe the alternative therapies reported to treat ESKAPE infections, their advantages and limitations, potential application in vivo, and status in clinical trials. The review further highlights the importance of a combinatorial approach, wherein two or more therapies are used in combination in order to overcome their individual limitations, additional studies on which are warranted, before translating them into clinical practice. These advances could possibly give an alternate solution or extend the lifetime of current antimicrobials.

/pdf/emerging-strategies-to-combat-eskape-pathogens-in-the-era-of-mchli3dllg.pdf

Emerging Strategies to Combat ESKAPE Pathogens in the Era of Antimicrobial Resistance: A Review.

/pdf/emergence-of-genomic-diversity-and-recurrent-mutations-in-4ug78d8h14.pdf

Emergence of genomic diversity and recurrent mutations in SARS-CoV-2.

The remarkable capacity of some viruses to adapt to new hosts and environments is highly dependent on their ability to generate de novo diversity in a short period of time. Rates of spontaneous mutation vary amply among viruses. RNA viruses mutate faster than DNA viruses, single-stranded viruses mutate faster than double-strand virus, and genome size appears to correlate negatively with mutation rate. Viral mutation rates are modulated at different levels, including polymerase fidelity, sequence context, template secondary structure, cellular microenvironment, replication mechanisms, proofreading, and access to post-replicative repair. Additionally, massive numbers of mutations can be introduced by some virus-encoded diversity-generating elements, as well as by host-encoded cytidine/adenine deaminases. Our current knowledge of viral mutation rates indicates that viral genetic diversity is determined by multiple virus- and host-dependent processes, and that viral mutation rates can evolve in response to specific selective pressures.

/pdf/mechanisms-of-viral-mutation-2li5ef9ina.pdf

Mechanisms of viral mutation

https://digital.csic.es/bitstream/10261/209677/1/Metropolitan%20Wastewater%20Analysis_Randazzo.pdf

Metropolitan wastewater analysis for COVID-19 epidemiological surveillance.

Mutational fitness effects can be measured with relatively high accuracy in viruses due to their small genome size, which facilitates full-length sequencing and genetic manipulation. Previous work has shown that animal and plant RNA viruses are very sensitive to mutation. Here, we characterize mutational fitness effects in single-stranded (ss) DNA and ssRNA bacterial viruses. First, we performed a mutation-accumulation experiment in which we subjected three ssDNA (ΦX174, G4, F1) and three ssRNA phages (Qβ, MS2, and SP) to plaque-to-plaque transfers and chemical mutagenesis. Genome sequencing and growth assays indicated that the average fitness effect of the accumulated mutations was similar in the two groups. Second, we used site-directed mutagenesis to obtain 45 clones of ΦX174 and 42 clones of Qβ carrying random single-nucleotide substitutions and assayed them for fitness. In ΦX174, 20% of such mutations were lethal, whereas viable ones reduced fitness by 13% on average. In Qβ, these figures were 29% and 10%, respectively. It seems therefore that high mutational sensitivity is a general property of viruses with small genomes, including those infecting animals, plants, and bacteria. Mutational fitness effects are important for understanding processes of fitness decline, but also of neutral evolution and adaptation. As such, these findings can contribute to explain the evolution of ssDNA and ssRNA viruses.

/pdf/the-fitness-effects-of-random-mutations-in-single-stranded-52u32hp9rx.pdf

The fitness effects of random mutations in single-stranded DNA and RNA bacteriophages.

Biofilms are clusters of bacteria that live in association with surfaces. Their main characteristic is that the bacteria inside the biofilms are attached to other bacterial cells and to the surface by an extracellular polymeric matrix. Biofilms are capable of adhering to a wide variety of surfaces, both biotic and abiotic, including human tissues, medical devices, and other materials. On these surfaces, biofilms represent a major threat causing infectious diseases and economic losses. In addition, current antibiotics and common disinfectants have shown limited ability to remove biofilms adequately, and phage-based treatments are proposed as promising alternatives for biofilm eradication. This review analyzes the main advantages and challenges that phages can offer for the elimination of biofilms, as well as the most important factors to be taken into account in order to design effective phage-based treatments.

/pdf/phages-for-biofilm-removal-2m949pcta8.pdf

Phages for Biofilm Removal.

Despite being silent with respect to protein sequence, synonymous nucleotide substitutions can be targeted by natural selection directly at the DNA or RNA level. However, there has been no systematic assessment of how frequent this type of selection is. Here, we have constructed 53 single random synonymous substitution mutants of the bacteriophages Qb and UX174 by site-directed mutagenesis and assayed their fitness. Analysis of this mutant collection and of previous studies undertaken with a variety of single-stranded (ss) viruses demonstrates that selection at synonymous sites is stronger in RNA viruses than in DNA viruses. We estimate that this type of selection contributes approximately 18% of the overall mutational fitness effects in ssRNA viruses under our assay conditions and that random synonymous substitutions have a 5% chance of being lethal to the virus, whereas in ssDNA viruses, these figures drop to 1.4% and 0%, respectively. In contrast, the effects of nonsynonymous substitutions appear to be similar in ssRNA and ssDNA viruses.

Pilar Domingo-Calap

Papers

Mechanisms of viral mutation

Metropolitan wastewater analysis for COVID-19 epidemiological surveillance.

The fitness effects of random mutations in single-stranded DNA and RNA bacteriophages.

Phages for Biofilm Removal.

The Fitness Effects of Synonymous Mutations in DNA and RNA Viruses