Exploring the Fitness Landscape of TEM-1 Beta-Lactamase: A Survey of Intragenic Epistasis & the Fitness Effects of InDels

The distribution of fitness effects, or fitness landscape, of a protein offers a picture of the relationship between mutations and their effects on a broad scale. Mutations in a protein can have positive, negative, or neutral effects on its function. Mapping these effects on a large scale allows us to better understand evolutionary dynamics in nature and informs our ability to better engineer proteins in the laboratory. I used deep-mutational scanning techniques, including saturation mutagenesis, high-throughput selection, and DNA deep-sequencing, to explore two important and understudied aspects of the fitness landscape of TEM-1 beta-lactamase. First, I examined pairwise intragenic epistasis among sequential amino acid substitutions in TEM-1. Epistasis, or interactions between mutations, plays a central role in shaping the fitness landscape, but a clear picture of the prevalence and patterns of epistasis has yet to emerge. This study is the first to systematically examine pairwise epistasis throughout an entire protein performing its native function in its native host. I explored the relationship between epistasis and secondary structure, solvent accessibility, distance from the active site, amino acid identity, and individual mutant effect. I found pervasive negative epistasis, particularly in highly structured regions of the protein and among buried residues, and a high frequency of negative sign epistasis among individually beneficial mutations. Second, I present a near-comprehensive analysis of the fitness effects of single amino acid insertions and deletions (InDels) in TEM-1. Short InDels are a common type of mutation in nature, often having important consequences, such as opening new pathways for adaptation. InDels also represent a useful source of variation in the protein engineering toolbox. Yet despite their importance and utility, the distribution of fitness effects of InDels is vastly understudied compared to substitutions. I found InDels to be largely deleterious, but notable regions of tolerance were observed throughout the protein. I found secondary structure, weighted contact number, and evolutionary variation in class A beta-lactamases to be the most predictive of their fitness effects

Intradermal Administration of Fractional Doses of Inactivated Poliovirus Vaccine: A Dose-Sparing Option for Polio Immunization.

A fractional dose of inactivated poliovirus vaccine (fIPV) administered by the intradermal route delivers one fifth of the full vaccine dose administered by the intramuscular route and offers a potential dose-sparing strategy to stretch the limited global IPV supply while further improving population immunity. Multiple studies have assessed immunogenicity of intradermal fIPV compared with the full intramuscular dose and demonstrated encouraging results. Novel intradermal devices, including intradermal adapters and disposable-syringe jet injectors, have also been developed and evaluated as alternatives to traditional Bacillus Calmette-Guérin needles and syringes for the administration of fIPV. Initial experience in India, Pakistan, and Sri Lanka suggests that it is operationally feasible to implement fIPV vaccination on a large scale. Given the available scientific data and operational feasibility shown in early-adopter countries, countries are encouraged to consider introducing a fIPV strategy into their routine immunization and supplementary immunization activities

Prefrontal photobiomodulation produces beneficial mitochondrial and oxygenation effects in older adults with bipolar disorder

There is growing evidence of mitochondrial dysfunction and prefrontal cortex (PFC) hypometabolism in bipolar disorder (BD). Older adults with BD exhibit greater decline in PFC-related neurocognitive functions than is expected for age-matched controls, and clinical interventions intended for mood stabilization are not targeted to prevent or ameliorate mitochondrial deficits and neurocognitive decline in this population. Transcranial infrared laser stimulation (TILS) is a non-invasive form of photobiomodulation, in which photons delivered to the PFC photo-oxidize the mitochondrial respiratory enzyme, cytochrome-c-oxidase (CCO), a major intracellular photon acceptor in photobiomodulation. TILS at 1064-nm can significantly upregulate oxidized CCO concentrations to promote differential levels of oxygenated vs. deoxygenated hemoglobin (HbD), an index of cerebral oxygenation. The objective of this controlled study was to use non-invasive broadband near-infrared spectroscopy to assess if TILS to bilateral PFC (Brodmann area 10) produces beneficial effects on mitochondrial oxidative energy metabolism (oxidized CCO) and cerebral oxygenation (HbD) in older (≥50 years old) euthymic adults with BD (N = 15). As compared to sham, TILS to the PFC in adults with BD increased oxidized CCO both during and after TILS, and increased HbD concentrations after TILS. By significantly increasing oxidized CCO and HbD concentrations above sham levels, TILS has the potential ability to stabilize mitochondrial oxidative energy production and prevent oxidative damage in the PFC of adults with BD. In conclusion, TILS was both safe and effective in enhancing metabolic function and subsequent hemodynamic responses in the PFC, which might help alleviate the accelerated neurocognitive decline and dysfunctional mitochondria present in BD

Physiological Stress Responses to a Live-Fire Training Evolution in Career Firefighters

Firefighters have a physically demanding job that exposes them to many specific and unique stressors, which contribute to cardiovascular disease (CVD) risk (i.e., cardiovascular strain, inflammation, and oxidative stress) or even mortality. At present, the literature lacks data evaluating these physiological stress responses amongst firefighters in any realistic or simulated scenarios, such as a live-fire training evolution (LFTE). Given the elevated risk of premature mortality, there is a critical need to better understand the physiological stress responses to an LFTE. This information could aid in developing nutritional, training, and other various interventions to mitigate stress load and reduce the incidence of CVD among this population. PURPOSE: To assess the physiological stress response to an LFTE among firefighters. METHODS: Seventy-six (n = 76) career firefighters completed an LFTE. Salivary samples were collected pre, immediately post, and 30-min post the LFTE, and analyzed the following stress markers: α-amylase (AA), secretory immunoglobulin-A (SIgA), and cortisol. One-way repeated measures analysis of variance was used to assess changes over time. Fisher’s LSD and Cohen’s d effect size calculations were used for Post hoc analysis. RESULTS: Significant main effects for time were found for AA, SIgA, and cortisol (p\u3c0.001). Fisher’s LSD post hoc analysis found AA, SIgA, and cortisol concentrations were all significantly elevated immediately post LFTE compared to pre (p\u3c0.0001) and 30-min post (p\u3c0.0001). Medium to large effect sizes were noted for AA, SIgA, and cortisol with respect to changes pre to immediately post-LFTE (d = 0.84, 0.61, and 0.78, respectively). CONCLUSION: While many studies have shown increased inflammation and oxidative stress, as well as adverse cardiovascular and metabolic responses to firefighting activities, these data provide insight into the physiological stress placed upon a firefighter engaging in fire-suppressive evolutions

Understanding Sow Sexual Behavior and the Application of the Boar Pheromone to Stimulate Sow Reproduction

In this chapter, we review the sexual behavior of domestic pigs, and the visible or measurable anatomical features of the pig that will contribute to detecting sows in estrus. We also summarize olfactory organs, and the effects of a sexual pheromone on pig’s biology and sow reproductive performance. We discuss the role of a live boar in the heat detection where the female is in breeding crates. However, there is an increasing interest in being able to breed sows without a boar present. Farm workers must be trained on the fine points of estrus detection so that they can work in a safe and productive setting. After a review of olfactory biology of the pig, the chapter explains how new pheromonal technology, such as BOARBETTER®, aids in the process of heat detection with or without a live boar. To achieve reproductive success, the persons breeding must assimilate all fine points of pig sexual behavior and possess a clear understanding of what they should be looking for in each sow they expect to breed

Rapid changes in public perception toward a conservation initiative

Rapid, widespread changes in public perceptions and behaviors have the potential to influence conservation outcomes. However, few studies have documented whether and how such shifts occur throughout the span of a conservation initiative. We examined the 2020 ballot initiative to reintroduce wolves into Colorado, which passed with less support than prior surveys had estimated. We conducted a postelection survey of Colorado residents using the same methods as our preelection survey to compare responses between surveys and to official election results. Reported voting in favor of wolf reintroduction in the postelection survey decreased in comparison to voting intentions shared in the preelection survey, but not enough to reflect the actual vote. While bias from survey methods and/or sampling contributed to differences, we also found evidence that public perception changed. Specifically, beliefs about the potential for negative impacts of wolves increased, while beliefs about the potential for positive impacts of wolves decreased. Our findings highlight the need to conduct longitudinal monitoring of public perception given perceptions may be highly fluid as different entities attempt to sway voters. In addition, to better understand evolving perceptions, survey methods and sampling need to be improved

Mechanism of KMT5B haploinsufficiency in neurodevelopment in humans and mice

KMT5B gene; Neurodevelopment; MiceGen KMT5B; Neurodesenvolupament; RatolinsGen KMT5B; Neurodesarrollo; RatonesPathogenic variants in KMT5B, a lysine methyltransferase, are associated with global developmental delay, macrocephaly, autism, and congenital anomalies (OMIM# 617788). Given the relatively recent discovery of this disorder, it has not been fully characterized. Deep phenotyping of the largest (n = 43) patient cohort to date identified that hypotonia and congenital heart defects are prominent features that were previously not associated with this syndrome. Both missense variants and putative loss-of-function variants resulted in slow growth in patient-derived cell lines. KMT5B homozygous knockout mice were smaller in size than their wild-type littermates but did not have significantly smaller brains, suggesting relative macrocephaly, also noted as a prominent clinical feature. RNA sequencing of patient lymphoblasts and Kmt5b haploinsufficient mouse brains identified differentially expressed pathways associated with nervous system development and function including axon guidance signaling. Overall, we identified additional pathogenic variants and clinical features in KMT5B-related neurodevelopmental disorder and provide insights into the molecular mechanisms of the disorder using multiple model systems.This work was supported by LB692 Nebraska Tobacco Settlement Biomedical Research Development Program (to H.A.F.S.); The Simons Foundation Autism Research Initiative–Bridge to Independence Award SFARI 381192 (to H.A.F.S.); The A*STAR, Singapore, IAF-PP Program H17/01/a0/004 (to C.Y.L.); The Wong Boon Hock Society research program Yong Loo Lin School of Medicine (to Z.X.C.); NIH training grant 2T32GM008638-25 (L.B.); The Intramural Research Program of the National Human Genome Research Institute (to W.G.); The National Center for Advancing Translational Sciences of the NIH award number TL1TR001880 (to S.E.S.); The Eunice Kennedy Shriver National Institute of Child Health and Human Development award number HD009003-01 (to S.E.S.); Institute for Translational Medicine and Therapeutics of the Perelman School of Medicine at the University of Pennsylvania (to S.E.S.); and Swiss National Science Foundation (SNSF) grant 320020_179547 and funds from the University of Zurich Research Priority Programs (URPP) AdaBD: Adaptive Brain Circuits in Developments (to A.Rau.). F.J.K. was funded by the Deutsche Forschungsgemeinschaft grant number FOR 2488. In silico modeling was supported by the Spanish Ministerio de Ciencia e Innovación grant number PID2019-111217RB-I00 (to X.d.l.C.). This study used data from the DDD study. The DDD study presents independent research commissioned by the Health Innovation Challenge Fund (grant number HICF-1009-003). This study makes use of DECIPHER (www.deciphergenomics.org), which is funded by Wellcome (grant number 223718/Z/21/Z). See Nature PMID: 25533962 or www.ddduk.org/access.html for full acknowledgement

Neuropsychological deficits in Posterior Cortical Atrophy and typical Alzheimer's disease:A meta-analytic review

Aims: To identify cognitive tests that best differentiate between Posterior Cortical Atrophy (PCA) and typical Alzheimer's Disease (tAD), as well as PCA and healthy control (HC) participants. Method: Medline, PsycInfo and Web of Science were systematically searched using terms related to PCA, tAD, and cognitive testing. Seventeen studies were identified, including 441 PCA, 391 tAD, and 284 HC participants. Standardised effect sizes of mean scores were calculated to measure performance differences on cognitive tests for PCA versus tAD and PCA versus HC groups. Meta-analyses used a random effects model. Results: The most discriminating cognitive tests for PCA and tAD presentations were measures of visuospatial function and verbal memory. Large, significant effect sizes were produced for all measures of visuospatial function, most notably for Rey-Osterrieth Copy (Hedges' g =-2.79), VOSP Fragmented letters (Hedges' g =-1.73), VOSP Dot Counting (Hedges' g =-1.74), and VOSP Cube Analysis (Hedges' g =-1.98). For measures of verbal memory, the RAVLT delay and Digit Span Backwards produced significant medium effects (Hedges' g = .62 and-.56, respectively). Conclusion: Establishing a common framework for testing individuals with PCA has important implications for diagnosis and treatment, and forms a practical objective for future research. Findings from this meta-analysis suggest that measures of visuospatial function and verbal memory would form an important part of this framework. Crown Copyright (c) 2021 Published by Elsevier Ltd. All rights reserved