In Silico Evaluation of Variant Calling Methods for Bacterial Whole-Genome Sequencing Assays.

Identification and analysis of clinically relevant strains of bacteria increasingly relies on whole-genome sequencing. The downstream bioinformatics steps necessary for calling variants from short-read sequences are well-established but seldom validated against haploid genomes. We devised an in silico workflow to introduce single nucleotide polymorphisms (SNP) and indels into bacterial reference genomes, and computationally generate sequencing reads based on the mutated genomes. We then applied the method to Mycobacterium tuberculosis H37Rv, Staphylococcus aureus NCTC 8325, and Klebsiella pneumoniae HS11286, and used the synthetic reads as truth sets for evaluating several popular variant callers. Insertions proved especially challenging for most variant callers to correctly identify, relative to deletions and single nucleotide polymorphisms. With adequate read depth, however, variant callers that use high quality soft-clipped reads and base mismatches to perform local realignment consistently had the highest precision and recall in identifying insertions and deletions ranging from1 to 50 bp. The remaining variant callers had lower recall values associated with identification of insertions greater than 20 bp.

Pandemic-Related Instructor Talk: How New Instructors Supported Students at the Onset of the COVID-19 Pandemic.

At the same time that COVID-19 cases in the United States first began to increase, fellows in a mentored teaching apprenticeship for postdoctoral scientists began to teach undergraduate seminars. The fellows suddenly needed to support students emotionally and switch to online instruction. They were encouraged to acknowledge and address the pandemic during each class and decided to do so. In this case study, we examined the language fellows used in response to this encouragement, hypothesizing that they would engage in a variety of pandemic-related instructor talk, i.e., language that instructors use in the classroom that is not directly tied to educational content. We analyzed transcripts from 17 2-hour undergraduate biology seminar courses and found 167 instances of pandemic-related instructor talk. We used grounded theory to identify categories that emerged from these quotations: Positive coping mechanisms and self-care; Adjusting to online learning; Compassionate instruction; Personal impacts; COVID-19 and society; Dreaming; and Biology of COVID-19. Talk in these categories may help build relationships among instructors and students. The category about quickly Adjusting to online learning is unique, in that it is unlikely that there will be another time that will require simultaneous and rapid national movement to online instruction. In addition, four of the seven categories are direct consequences of COVID-19 specifically, and thus are unique to this time. Analyzing pandemic-related instructor talk has shed light on how new instructors navigated the trials of teaching in 2020.

Synergistic epistasis enhances the co-operativity of mutualistic interspecies interactions.

Early evolution of mutualism is characterized by big and predictable adaptive changes, including the specialization of interacting partners, such as through deleterious mutations in genes not required for metabolic cross-feeding. We sought to investigate whether these early mutations improve cooperativity by manifesting in synergistic epistasis between genomes of the mutually interacting species. Specifically, we have characterized evolutionary trajectories of syntrophic interactions of Desulfovibrio vulgaris (Dv) with Methanococcus maripaludis (Mm) by longitudinally monitoring mutations accumulated over 1000 generations of nine independently evolved communities with analysis of the genotypic structure of one community down to the single-cell level. We discovered extensive parallelism across communities despite considerable variance in their evolutionary trajectories and the perseverance within many evolution lines of a rare lineage of Dv that retained sulfate-respiration (SR+) capability, which is not required for metabolic cross-feeding. An in-depth investigation revealed that synergistic epistasis across pairings of Dv and Mm genotypes had enhanced cooperativity within SR- and SR+ assemblages, enabling their coexistence within the same community. Thus, our findings demonstrate that cooperativity of a mutualism can improve through synergistic epistasis between genomes of the interacting species, enabling the coexistence of mutualistic assemblages of generalists and their specialized variants.

Vector-Enabled Metagenomic (VEM) Surveys Using Whiteflies (Aleyrodidae) Reveal Novel Begomovirus Species in the New and Old Worlds.

Whitefly-transmitted viruses belonging to the genus Begomovirus (family Geminiviridae) represent a substantial threat to agricultural food production. The rapid evolutionary potential of these single-stranded DNA viruses combined with the polyphagous feeding behavior of their whitefly vector (Bemisia tabaci) can lead to the emergence of damaging viral strains. Therefore, it is crucial to characterize begomoviruses circulating in different regions and crops globally. This study utilized vector-enabled metagenomics (VEM) coupled with high-throughput sequencing to survey begomoviruses directly from whiteflies collected in various locations (California (USA), Guatemala, Israel, Puerto Rico, and Spain). Begomoviruses were detected in all locations, with the highest diversity identified in Guatemala where up to seven different species were identified in a single field. Both bipartite and monopartite viruses were detected, including seven new begomovirus species from Guatemala, Puerto Rico, and Spain. This begomovirus survey extends the known diversity of these highly damaging plant viruses. However, the new genomes described here and in the recent literature appear to reflect the outcome of interactions between closely-related species, often resulting from recombination, instead of unique, highly divergent species.

98% identical, 100% wrong: per cent nucleotide identity can lead plant virus epidemiology astray.

Short-form publications such as Plant Disease reports serve essential functions: the rapid dissemination of information on the geography of established plant pathogens, incidence and symptomology of pathogens in new hosts, and the discovery of novel pathogens. Many of these sentinel publications include viral sequence data, but most use that information only to confirm the virus' species. When researchers use the standard technique of per cent nucleotide identity to determine that the new sequence is closely related to another sequence, potentially erroneous conclusions can be drawn from the results. Multiple introductions of the same pathogen into a country are being ignored because researchers know fast-evolving plant viruses can accumulate substantial sequence divergence over time, even from a single introduction. An increased use of phylogenetic methods in short-form publications could speed our understanding of these cryptic second introductions and aid in control of epidemics.