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Numerous affinity purification-mass spectrometry (AP-MS) and yeast two-hybrid screens have each defined thousands of pairwise protein-protein interactions (PPIs), most of which are between functionally unrelated proteins. The accuracy of these networks, however, is under debate. Here, we present an AP-MS survey of the bacterium Desulfovibrio vulgaris together with a critical reanalysis of nine published bacterial yeast two-hybrid and AP-MS screens. We have identified 459 high confidence PPIs from D. vulgaris and 391 from Escherichia coli Compared with the nine published interactomes, our two networks are smaller, are much less highly connected, and have significantly lower false discovery rates. In addition, our interactomes are much more enriched in protein pairs that are encoded in the same operon, have similar functions, and are reproducibly detected in other physical interaction assays than the pairs reported in prior studies. Our work establishes more stringent benchmarks for the properties of protein interactomes and suggests that bona fide PPIs much more frequently involve protein partners that are annotated with similar functions or that can be validated in independent assays than earlier studies suggested.

UnlabelledTransposon mutagenesis with next-generation sequencing (TnSeq) is a powerful approach to annotate gene function in bacteria, but existing protocols for TnSeq require laborious preparation of every sample before sequencing. Thus, the existing protocols are not amenable to the throughput necessary to identify phenotypes and functions for the majority of genes in diverse bacteria. Here, we present a method, random bar code transposon-site sequencing (RB-TnSeq), which increases the throughput of mutant fitness profiling by incorporating random DNA bar codes into Tn5 and mariner transposons and by using bar code sequencing (BarSeq) to assay mutant fitness. RB-TnSeq can be used with any transposon, and TnSeq is performed once per organism instead of once per sample. Each BarSeq assay requires only a simple PCR, and 48 to 96 samples can be sequenced on one lane of an Illumina HiSeq system. We demonstrate the reproducibility and biological significance of RB-TnSeq with Escherichia coli, Phaeobacter inhibens, Pseudomonas stutzeri, Shewanella amazonensis, and Shewanella oneidensis. To demonstrate the increased throughput of RB-TnSeq, we performed 387 successful genome-wide mutant fitness assays representing 130 different bacterium-carbon source combinations and identified 5,196 genes with significant phenotypes across the five bacteria. In P. inhibens, we used our mutant fitness data to identify genes important for the utilization of diverse carbon substrates, including a putative d-mannose isomerase that is required for mannitol catabolism. RB-TnSeq will enable the cost-effective functional annotation of diverse bacteria using mutant fitness profiling.ImportanceA large challenge in microbiology is the functional assessment of the millions of uncharacterized genes identified by genome sequencing. Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach to assign phenotypes and functions to genes. However, the current strategies for TnSeq are too laborious to be applied to hundreds of experimental conditions across multiple bacteria. Here, we describe an approach, random bar code transposon-site sequencing (RB-TnSeq), which greatly simplifies the measurement of gene fitness by using bar code sequencing (BarSeq) to monitor the abundance of mutants. We performed 387 genome-wide fitness assays across five bacteria and identified phenotypes for over 5,000 genes. RB-TnSeq can be applied to diverse bacteria and is a powerful tool to annotate uncharacterized genes using phenotype data.

Background Weight status and weight perception have a significant impact on life satisfaction. As overweight prevalence increases in Canada, it is important to understand how accuracy of weight perception (AWP) is associated with life satisfaction. This study explored the association between AWP and life satisfaction among Canadian adults with and without anxiety and/or mood disorders. Methods Using data from the 2015–2018 cycles of the Canadian Community Health Survey, an indicator of AWP was created to capture concordance between perceived weight and actual weight status. Univariate and multivariate Gaussian generalized linear models were assessed while stratifying by sex and presence of anxiety and/or mood disorders. Results Our sample included 88 814 males and 106 717 females. For both sexes, perceiving oneself as overweight or underweight, regardless of actual weight status, was associated with lower life satisfaction (β = −0.93 to −0.30), compared to those who accurately perceived their weight as 'just about right'. Perceiving oneself as overweight or underweight was associated with more pronounced differences in life satisfaction scores in those with anxiety and/or mood disorders (β = −1.49 to −0.26) than in those without these disorders (β = −0.76 to −0.25). Conclusion Weight perception is more indicative of life satisfaction than actual weight status, especially in those with anxiety and/or mood disorders. [ABSTRACT FROM AUTHOR]

Context: On March 14, 2020, the Ontario, Canada health insurance plan approved COVID-19 physician virtual billing codes; family physicians (FPs) rapidly adopted a new model of care. Virtual care may remain post-pandemic; however, its future should be informed by evidence that considers access and continuity. Objective: 1) to determine FP virtual visit volumes and patient characteristics and 2) to explore FPs' perspectives on virtual visit adoption and implementation. Study Design: Mixed methods: Secondary analysis of health administrative (HA) data and semi-structured qualitative interviews with FPs. Setting or Dataset: London and Middlesex County, Ontario, Canada. HA data through ICES, Ontario entity holding data. Population studied: FPs and their patients. Outcome Measures: Volumes of FP in-person and virtual visits during early pandemic; characteristics of patients receiving care; FPs' perspectives on adopting and delivering virtual care. Results: Overall visit volume dropped by 36% during first wave, recovered to pre-pandemic levels by October 2020. Sharp in-person visit drop of 73% and virtual visit uptake from 0.08% of total visits to 57% within two weeks of March 2020. FPs described this initial drop in volume as patients not seeking care and practices lacking PPE. The move to virtual care was largely to telephone visits. Patient characteristics compared to pre-pandemic, the proportion seeking care were older (46 vs 50 years), more vulnerable (38% vs 41%), and more multimorbidity (33% vs 41%). This was consistent with FP reports that healthier patients stayed away, routine care deferred, sicker patients needed to be seen. FPs believed most vulnerable patients had access to care but cautioned highly vulnerable such as those homeless did not have cell phone access or a safe place to receive calls. Rural FPs reported access issues because of lack of high-speed internet. FPs attributed success of virtual care to the continuity in relationships they had with patients that were established in person pre-pandemic. Conclusions: FPs moved rapidly to virtual care. FP offices remained open despite PPE concerns but overall volumes dropped initially. Vulnerable and sicker patients received care but FPs expressed concern for highly vulnerable and rural residents. FPs believed they could offer patient-centred care over the phone but indicated the importance of maintaining in-person care to build relationships., Competing Interests: Authors report none., (2021 Annals of Family Medicine, Inc.)