bluuebunny/crossref_metadata_2025 huggingface dataset:
Origin and dynamics of Mycobacterium tuberculosis subpopulations that predictably generate drug tolerance and resistance
Authors: Sebastian Jees , Thomas Anooja , Levine Carly , Shrestha Riju , Levy Shawn , Safi Hassan , Pentakota Sri Ram , Kumar Pradeep , Alland David
Published: October 2022

Initial responses to tuberculosis treatment are poor predictors of final therapeutic outcomes in drug-susceptible disease suggesting that treatment success depends on features that are hidden within a small minority of the overall infecting Mycobacterium tuberculosis (Mtb) population. We developed a multi-transwell robotic system to perform numerous parallel cultures of genetically barcoded Mtb exposed to steady-state concentrations of rifampicin to uncover these difficult to eliminate minority populations. We found that tolerance repeatedly emerged from at least two subpopulations of barcoded cells, one that could not grow on solid agar media and a second that could form colonies, but whose kill curves diverged from the general bacterial population within 4 and 16 days of drug exposure, respectively. These tolerant subpopulations reproducibly passed through a phase characterized by multiple unfixed resistance mutations followed by emergent drug resistance in some cultures. Barcodes associated with drug resistance identified an especially privileged subpopulation that was rarely eliminated despite 20 days of drug treatment even in cultures that did not contain any drug resistant mutants. The association of this evolutionary scenario with a defined subset of barcodes across multiple independent cultures suggested a transiently heritable phenotype, and indeed glpK phase variation mutants were associated with up to 16 % of the resistant cultures. Drug tolerance and resistance were eliminated in ΔruvA mutant consistent with the importance of bacterial stress responses. This work provides a window into the origin and dynamics of bacterial drug tolerant subpopulations whose elimination may be critical to developing rapid and resistance free cures.

Importance

Tuberculosis is unusual among bacterial diseases in that treatments which can rapidly resolve symptoms do not predictably lead to a durable cure unless treatment is continued for months after all clinical and microbiological signs of disease have been eradicated. Using a novel steady-state antibiotic exposure system combined with chromosomal barcoding, we identified small hidden Mycobacterium tuberculosis subpopulations that repeatedly enter into a state of drug tolerance with a predisposition to develop fixed drug resistance after first developing a cloud of unfixed resistance mutations. The existence of these difficult to eradicate subpopulations may explain the need for extended treatment regimen for tuberculosis. Their identification provides opportunities to test genetic and therapeutic approaches that may result in shorter and more effective TB treatments.
Circularization of rv0678 for genotypic bedaquiline resistance testing of Mycobacterium tuberculosis
Authors: Limberis Jason D , Nalyvayko Alina , Ernst Joel D , Metcalfe John Z
Published: October 2022

Circular DNA offers benefits over linear DNA in diagnostic and field assays, but currently, circular DNA generation is lengthy, inefficient, highly dependent on the length and sequence of DNA, and can result in unwanted chimeras. We present streamlined methods for generating PCR-targeted circular DNA from a 700bp amplicon of rv0678, the high GC content (65%) gene implicated in Mycobacterium tuberculosis bedaquiline resistance, and demonstrate that these methods work as desired. We employ self-circularization with and without splints, a Gibson cloning-based approach, and novel two novel methods for generating pseudocircular DNA. The circular DNA can be used as a template for rolling circle PCR followed by long-read sequencing, allowing for the error correction of sequence data, and improving the confidence in the drug resistance determination and strain identification; and ultimately improving patient treatment.
The natural history of homonymous hemianopia revisited
Authors: Saionz Elizabeth L. , Cavanaugh Matthew R. , Johnson Brent A. , Harrington Donald , Aguirre Geoffrey K. , Huxlin Krystel R.
Published: October 2022

Objective

To re-evaluate the longitudinal progression of stroke-induced homonymous visual field defects using strictly automated perimetry (Zeiss Humphrey Systems), rigorous inclusion/exclusion criteria, and quantitative analyses.

Methods

A retrospective chart review of stroke patients diagnosed with “homonymous hemianopia”, who underwent monocular Humphrey visual field (HVF) perimetry using the 24-2 SITA standard pattern from 2011-2019, was conducted at a large US academic medical center. Reliable tests (<20% fixation losses, false positives, and false negatives) were identified and analyzed with generalized estimating equations to extract temporal trends in perimetric mean deviation (PMD) and deficit area.

Results

Of 532 patients with “homonymous hemianopia”, sequential, reliable HVFs were only available for 36 patients in the right eye, and 30 patients in the left eye, ranging from 7 days to 58 months post-stroke. Both PMD and deficit area improved early, within the first 3 months post-stroke; however, this was followed by a subsequent decline in performance >1 year post-stroke. Changes were similar between eyes.

Conclusion

We discovered that a large portion of occipital stroke patients do not receive comprehensive ophthalmologic follow-up and, even then, only a fraction of HVFs performed are reliable enough for rigorous analysis. Nonetheless, reliable HVFs in such patients confirmed early visual improvement after stroke, consistent with prior reports. However, in contrast with prior, qualitative reports, there was no stability of the deficit beyond 6 months post-stroke; instead, gradual worsening erased the initial spontaneous improvement, especially >1 year post-stroke.
A NURSING PHILOSOPHY OF FAMILY EMPOWERMENT IN CARING UNDER-FIVE CHILDREN ILLNESS
Authors: Rachmawati Praba Diyan , Arief Yuni Sufyanti , Pandin Moses Glorino Rumambo
Published: October 2022

Introduction

Sick children will be at risk of experiencing growth and development disorders and experiencing severe conditions to decreased quality of life. Quality of care through proper management of sick children under five is a priority. However, in the field of pediatric nursing, there are still obstacles to the application of family empowerment in caring for sick children, so it is important to examine a literature review with a philosophical approach the application of family empowerment in caring for sick children, as an effort to optimize the care of sick children under-five.

Method

This study was based on the results of the Literature Review. Articles were obtained from 3 databases, namely Scopus, Science Direct and PubMed. The keywords used in searching the literature in this study were ((parent) OR (mother) AND (parental AND empowerment) OR (engagement) AND (children) OR (sick AND children)). Articles searched from 2018-2022, which were open access and in English, from this literature review search, found 12 articles.

Result

Based on a philosophical approach of family empowerment in the care of sick children under-five, family empowerment interventions with the principle of involving families in care, and increasing family knowledge and skills in caring for sick children can be implemented as an effort to optimize care for sick toddlers.

Conclusion

The results of this literature review can be used as a basis for nursing interventions that require parental involvement in caring for sick children. Family empowerment programs that are planned and structured can be applied in the care of sick children at home or in the hospital.
Autonomy-Supportive Sexual Health Communication and Sexual Health Behaviors for Black and Latino MSM in the House Ball Community: Protocol for a Social Network Analysis and Exploratory Structural Equation Model
Authors: Smith Martez D. R. , Leblanc Natalie M. , Nelson LaRon E. , McMahon James M.
Published: October 2022

Background

Black and Latino men who have sex with men (MSM) have the highest risk of HIV of any group in the U.S. Prevalence could be even higher among Black and Latino MSM belonging to the House Ball Community (HBC), an understudied community comprised primarily of Black and Latino sexual and gender minorities, with HIV prevalence from non-probability samples ranging from 4% to 38%.

Objectives

Basic Psychological Needs Theory will be utilized to understand how sexual health communication (SHC) influences sexual health behaviors of HBC-MSM. The proposed study will advance this goal by describing characteristics of SHC embedded within social and sexual networks, and identifying the effects of SHC on sexual health behaviors among a sample of Black and Latino HBC-MSM.

Methods

This study entails cross-sectional quantitative survey design with internet-based data collection to test a theory-driven model of the effects of autonomy supportive communication on sexual health behaviors. Respondent-driven sampling (RDS) and internet driven sampling (ID) will be used to recruit a sample of 200 HBC-MSM. We will utilize egocentric network analysis to describe (a) the HBC-specific social and/or sexual network members who provide SHC; (b) the degree of autonomy support provided by network members, and (c) the sexual health behaviors characterizing the sample of HBC-MSM. Structural equation modeling (SEM) will be performed to test associations between autonomy supportive sexual health communication (independent variable) and sexual health behaviors (outcome), with needs satisfaction and intrinsic motivation as intervening mediators.

Discussion

Such knowledge is necessary to expand understanding of how SHC impacts sexual health behaviors for HBC-MSM. The study provides an critical perspective on sexual health behaviors and motivations as participants operate in HBC. Knowledge generated from this study will help improve current HIV prevention interventions, as well as inform the development of future interventions, tailored to HBC-MSM.

Trial registration

N/A
Considerations for ensuring safety during telerehabilitation of people with stroke. A protocol for a scoping review
Authors: Gutierrez-Arias Ruvistay , González-Mondaca Camila , Marinkovic-Riffo Vinka , Ortiz-Puebla Marietta , Paillán-Reyes Fernanda , Seron Pamela
Published: October 2022

Introduction

Exercise interventions have a positive impact on people with stroke. However, access to exercise interventions is variable, and there may be a delay in the start of rehabilitation. Telerehabilitation has enabled the delivery of exercise interventions replacing the traditional face-to-face approach. Aspects related to the safety of people with stroke should be considered to avoid adverse events during the delivery of exercise interventions remotely. However, such information is scattered in the literature, and the detail with which measures taken during the implementation of exercise interventions for people with stroke are reported is unknown.

Objective

To summarise measures or aspects targeted at reducing the incidence of adverse events during the delivery of exercise interventions through telerehabilitation in patients after stroke.

Materials and methods

A scoping review will be conducted. A systematic search in MEDLINE-Ovid, Embase-Ovid CENTRAL, CINAHL Complete (EBSCOhost), and other resources will be carried out. We will include primary studies, published in full text in any language, involving people with stroke who undergo telerehabilitation where exercise is the main component. Two reviewers will independently select studies and extract data, and disagreements will be resolved by consensus or a third reviewer. The results will be reported in a narrative form, using tables and figures to support them.

Discussion

To implement this strategy within rehabilitation services, one of the first aspects to be solved is to ensure the safety of people. The results of this scoping review could contribute an information base for clinicians and decision-makers when designing remotely delivered exercise intervention programs.

Registration number

INPLASY202290104.
Treating intrusive memories after trauma in healthcare workers: a Bayesian adaptive randomised trial developing an imagery-competing task intervention
Authors: Ramineni Varsha , Millroth Philip , Iyadurai Lalitha , Jaki Thomas , Kingslake Jonathan , Highfield Julie , Summers Charlotte , Bonsall Michael B. , Holmes Emily A.
Published: October 2022

Intensive care unit (ICU) staff continue to face recurrent work-related traumatic events throughout the COVID-19 pandemic. Intrusive memories (IMs) of such traumatic events comprise sensory image-based memories. Harnessing research on preventing IMs with a novel behavioural intervention on the day of trauma, here we take critical next steps in developing this approach as a treatment for ICU staff who are already experiencing IMs days, weeks, or months post-trauma. To address the urgent need to develop novel mental health interventions, we used Bayesian statistical approaches to optimise a brief imagery-competing task intervention to reduce the number of IMs. We evaluated a digitised version of the intervention for remote, scalable delivery. We conducted a two-arm, parallel-group, randomised, adaptive Bayesian optimisation trial. Eligible participants worked clinically in a UK NHS ICU during the pandemic, experienced at least one work-related traumatic event, and at least three IMs in the week prior to recruitment. Participants were randomised to receive immediate or delayed (after four weeks) access to the intervention.

Primary outcome was the number of IMs of trauma during week 4, controlling for baseline week. Analyses were conducted on an intention-to-treat basis as a between-group comparison. Prior to final analysis, sequential Bayesian analyses were conducted (n=20,23,29,37,41,45) to inform early stopping of the trial prior to the planned maximum recruitment (n=150). Final analysis (n=75) showed strong evidence for a positive treatment effect (Bayes factor, BF=1.25 × 106): the immediate arm reported fewer IMs (median=1, IQR=0-3) than the delayed arm (median=10, IQR=6-16.5). With further digital enhancements, the intervention (n=28) also showed a positive treatment effect (BF=7.31). Sequential Bayesian analyses provided evidence for reducing IMs of work-related trauma for healthcare workers. This methodology also allowed us to rule out negative effects early, reduced the planned maximum sample size, and allowed evaluation of enhancements. Trial RegistrationNCT04992390(www.clinicaltrials.gov).
Factors associated with persistent hypertension one year postpartum in persons with gestational hypertension or preeclampsia
Authors: Livergood M Christine , Mahlum Leigh , Hauck Jordan , Tallmadge Maggie , Hoppe Kara , Palatnik Anna
Published: October 2022

Objective

To identify factors associated with persistent hypertension one-year postpartum after pregnancy complicated by gestational hypertension or preeclampsia.

Study Design

A retrospective case-control study of postpartum patients who had a diagnosis of gestation hypertension or preeclampsia during a recent pregnancy and attended one-year postpartum annual exam and blood pressure check between 2014 and 2019 in a single academic medical center. Cases were defined as persons with persistent hypertension one year postpartum, using the 2017 American College of Cardiology/American Heart Association (ACC/AHA) guidelines, defining stage I hypertension as systolic blood pressure ≥130 mmHg or diastolic blood pressure ≥80 mmHg. Controls were defined as non-pregnant persons with normal blood pressure (BP) at one year postpartum. Using bivariate and multivariate analyses, demographic, clinical and labor characteristics were compared between persons who had persistent hypertension one-year postpartum and controls.

Results

Of the 595 persons included in this analysis, 268 (45.0%) had persistent hypertension one year postpartum. Bivariate analyses demonstrated that older maternal age, higher body mass index (BMI) at first prenatal visit, at delivery, and one year postpartum, mild-range BP (compared to normal BP) prior to discharge, and patients with elevated BP at 6-week postpartum visit, were more likely to have persistent hypertension one-year postpartum. In contrast, nulliparity was associated with lower risk of having persistent hypertension at one-year postpartum. Multivariate logistic regression demonstrated that mild range BP prior to discharge (aOR 1.78, 95%CI 1.16-2.72), elevated BPs at 6 weeks postpartum (aOR 2.01, 95% CI 1.36-3.00), and higher BMI at one-year postpartum (aOR 1.07, 95%CI 1.00-1.14), remained to be significantly associated with higher odds of persistent hypertension one-year postpartum, while nulliparity remained to be associated with lower odds of persistent hypertension one-year postpartum (aOR 0.55, 95%CI 0.36-0.84)

Conclusion

In this cohort, 45% of patients with gestational hypertension or preeclampsia had persistent hypertension one-year postpartum by the 2017 ACC/AHA hypertension definition. Patients that had mildly elevated BPs in the immediate postpartum period as well as at 6 weeks postpartum, and higher BMI one year postpartum, had higher risk of having at least stage I HTN one year following pregnancy complicated by gestational hypertension or preeclampsia.
Efficacy and safety of endocrine therapy after mastectomy in patients with hormone receptor positive breast ductal carcinoma in situ:retrospective cohort study
Authors: Niu Nan , Zhang Yinan , Bai Yang , Wang Xin , Yan Shunchao , Song Dong , Xu Hong , Liu Tong , Hua Bin , Zhang Yingchao , Liu Jinchi , Qiao Xinbo , Liu Jiaxiang , Zheng Xinyu , Cao Hongyi , Liu Caigang
Published: October 2022

Background

More than half of Chinese patients with hormone receptor positive (HR+) ductal carcinoma in situ (DCIS) are treated with mastectomy, and usually followed by endocrine therapy (ET). Given that long-term ET can cause severe adverse effects it is important to determine the beneficial effect and safety of adjuvant ET after mastectomy on the disease-free survival (DFS) and adverse events in patients with HR+ DCIS.

Methods

To explore beneficial effect and safety of ET after mastectomy in patients with HR+ DCIS, we performed a multicenter, population-based study. This retrospective study analyzed the DFS and adverse events in 1037 HR+ DCIS Chinese patients with or without post-mastectomy ET from eight Breast Centers between 2006-2016. The median follow-up time was 86 months.

Results

There were 791 DCIS patients receiving ET (ET group). Those patients were followed-up for a median of 86 months (range, 60-177 months). There were 23 cases with tumor recurrence or distant metastasis. There were similar 5-year DFS rates and DFS between the ET and non-ET groups, even for those with high risk factors. Conversely, 37.04% of patients suffered from adverse events after ET, which were significantly higher than those in the non-ET group.

Conclusion

ET after mastectomy did not benefit patients with HR+ DCIS for their DFS, rather increased adverse events in those patients. Therefore, ET after mastectomy may not be recommended for patients with HR+ DCIS, even for those with high-risk factors, such as multifocal, microinvasive and higher T stage.

Funding

This study was supported by grants from Outstanding Scientific Fund of Shengjing Hospital (201803) and Outstanding Young Scholars of Liaoning Province (2019-YQ-10).
Long COVID Risk and Pre-COVID Vaccination: An EHR-Based Cohort Study from the RECOVER Program
Authors: Brannock M Daniel , Chew Robert F , Preiss Alexander J , Hadley Emily C , McMurry Julie A , Leese Peter J , Girvin Andrew T , Crosskey Miles , Zhou Andrea G , Moffitt Richard A , Funk Michele Jonsson , Pfaff Emily R , Haendel Melissa A , Chute Christopher G ,   the N3C and RECOVER Consortia
Published: October 2022

Importance

Characterizing the effect of vaccination on long COVID allows for better healthcare recommendations.

Objective

To determine if, and to what degree, vaccination prior to COVID-19 is associated with eventual long COVID onset, among those a documented COVID-19 infection.

Design, Settings, and Participants

Retrospective cohort study of adults with evidence of COVID-19 between August 1, 2021 and January 31, 2022 based on electronic health records from eleven healthcare institutions taking part in the NIH Researching COVID to Enhance Recovery (RECOVER) Initiative, a project of the National Covid Cohort Collaborative (N3C).

Exposures

Pre-COVID-19 receipt of a complete vaccine series versus no pre-COVID-19 vaccination.

Main Outcomes and Measures

Two approaches to the identification of long COVID were used. In the clinical diagnosis cohort (n=47,752), ICD-10 diagnosis codes or evidence of a healthcare encounter at a long COVID clinic were used. In the model-based cohort (n=199,498), a computable phenotype was used. The association between pre-COVID vaccination and long COVID was estimated using IPTW-adjusted logistic regression and Cox proportional hazards.

Results

In both cohorts, when adjusting for demographics and medical history, pre-COVID vaccination was associated with a reduced risk of long COVID (clinic-based cohort: HR, 0.66; 95% CI, 0.55-0.80; OR, 0.69; 95% CI, 0.59-0.82; model-based cohort: HR, 0.62; 95% CI, 0.56-0.69; OR, 0.70; 95% CI, 0.65-0.75).

Conclusions and Relevance

Long COVID has become a central concern for public health experts. Prior studies have considered the effect of vaccination on the prevalence of future long COVID symptoms, but ours is the first to thoroughly characterize the association between vaccination and clinically diagnosed or computationally derived long COVID. Our results bolster the growing consensus that vaccines retain protective effects against long COVID even in breakthrough infections.

Key Points

Question

Does vaccination prior to COVID-19 onset change the risk of long COVID diagnosis?

Findings

Four observational analyses of EHRs showed a statistically significant reduction in long COVID risk associated with pre-COVID vaccination (first cohort: HR, 0.66; 95% CI, 0.55-0.80; OR, 0.69; 95% CI, 0.59-0.82; second cohort: HR, 0.62; 95% CI, 0.56-0.69; OR, 0.70; 95% CI, 0.65-0.75).

Meaning

Vaccination prior to COVID onset has a protective association with long COVID even in the case of breakthrough infections.
IL-13 associated epithelial remodeling correlates with clinical severity in nasal polyposis
Authors: Kotas Maya E. , Patel Neil N. , Cope Emily K. , Gurrola Jose G. , Goldberg Andrew N. , Pletcher Steven D. , Seibold Max A. , Moore Camille M. , Gordon Erin D.
Published: October 2022

Background

Epithelial remodeling is a histopathologic feature of chronic inflammatory airway diseases including chronic rhinosinusitis (CRS). Cell type shifts and their relationship to CRS endotypes and severity are incompletely described.

Objective

The purpose of this study was to understand the relationship of epithelial cell remodeling to inflammatory endotypes and disease outcomes in CRS.

Methods

Using cell type transcriptional signatures derived from epithelial single cell sequencing, we analyzed bulk RNA sequencing data from sinus epithelial brushes obtained from patients with CRS with and without nasal polyps in comparison to healthy controls.

Results

The airway epithelium in nasal polyposis displayed increased tuft cell transcripts and decreased ciliated cell transcripts along with an IL-13 activation signature. In contrast, chronic rhinosinusitis without polyps showed an IL-17 activation signature. IL-13 activation scores were associated with increased tuft cell, goblet cell and mast cell scores and decreased ciliated cell scores. Furthermore, the IL-13 score was strongly associated with a previously reported activated (“polyp”) tuft cell score and a prostaglandin E2 (PGE2) activation signature. The Lund-McKay score, a computed tomographic metric of sinus opacification, correlated positively with activated tuft cell, mast cell, PGE2, and IL-13 and negatively with ciliated cell transcriptional signatures.

Conclusions

These results demonstrate that cell type alterations and PGE2 stimulation are key components of IL-13 induced epithelial remodeling in nasal polyposis, while IL-17 signaling is more prominent in CRS without polyps, and that clinical severity correlates with the degree of IL-13 induced epithelial remodeling.

Key Messages

Cell type signatures from single cell RNA sequencing, applied to bulk sequenced RNA sinus brushes, suggest increased tuft cells and mast cells and decreased ciliated cells in nasal polyp epithelium.

IL-17 signaling rather than IL-13 signaling is observed in epithelium from CRSsNP.

IL-13-drives epithelial remodeling and prostaglandin E2 signatures correlated with clinical measures of sinus opacification in CRS.

Capsule Summary

Measures of epithelial remodeling, including both IL-13 and PGE2 induced epithelial activation and cell type specific transcript alterations, correlate with a radiographic metric of disease severity in CRSwNP.
Changes in Fentanyl Distribution in California
Authors: Dugan Miah V. , Shah Ali H. , Anthony Trinidy R. , Famosa Rafiat , Piper Brian J.
Published: October 2022

Background

Fentanyl is a synthetic opioid that is commonly given as a medication to alleviate pain. This drug can be administered through multiple routes, hence making it easy to exploit at high rates. Due to the flexibility in which it can be taken, it increases the ease of both access and use. The purpose of this study was to analyze trends in the distribution of fentanyl and its formulations across Medicaid enrollees in California and among the 3-digit registrant zip codes in California over the period of pre-pandemic (2018–2019) to the early stages of the COVID-19 pandemic (2020).

Methods

Using the Automated Reports and Consolidated Ordering System (ARCOS), the distribution of fentanyl across California was compiled from 2018 to 2020. Utilizing ARCOS, the number of individuals within the source population who lived in one of California’s many zip codes was observed. To analyze the fentanyl distribution trend, we used Google Sheets, GraphPad Prism (Version 9.3.0 [463]), and Microsoft 365 Excel. These were helpful to organize the Medicaid, ARCOS data, and as well as to create graphs. The Medicaid database was used to compile the number of fentanyl formulations prescribed from 2018 to 2020 across California.

Results

The analyses from both databases provided insight into the difference in fentanyl distribution in California from the years 2018 to 2020. After looking further into the many 3-digit registrant zip codes as well as Medicaid enrollees, it was found that there was a decrease in the distribution of fentanyl and its formulations. Additionally, it was found that the distribution of fentanyl as a medication by business activities also decreased from 2018 to 2020.

Conclusion

The results indicate that there was more fentanyl being distributed and prescribed before the pandemic (2018– 2019). On the other hand, when we considered the effects of the pandemic, during 2020, there was quite a drastic decrease in the amount of fentanyl being prescribed and distributed to those living in California and those enrolled in Medicaid.
Copy number variation onABCC2-DNMBP lociimpacts the diversity and composition of the gut microbiota in pigs
Authors: Ramayo-Caldas Yuliaxis , Crespo-Piazuelo Daniel , Morata Jordi , González-Rodríguez Olga , Sebastià Cristina , Castello Anna , Dalmau Antoni , Ramos-Onsins Sebastian , Alexiou Konstantinos G. , Folch Josep M. , Quintanilla Raquel , Ballester Maria
Published: October 2022

Background

Genetic variation in the pig genome partially modulates the composition of porcine gut microbial communities. Previous studies have been focused on the association between single nucleotide polymorphisms (SNPs) and the gut microbiota, but little is known about the relationship between structural variants and gut microbial traits.

Results

The main goal of this study was to assess the effect of porcine genome copy number variants (CNVs) on the diversity and composition of pig gut microbiota. For this purpose, we used whole-genome sequencing data to undertake a comprehensive identification of CNVs followed by a genome-wide association analysis between the estimated CNV status and the gut bacterial diversity in a commercial Duroc pig population. A CNV predicted as gain (DUP) partially harboringABCC2-DNMBP lociwas associated with richness (p-value=5.41×10−5) and Shannon α-diversity (p-value=1.42×10−4). Thein-silicopredicted gain of copies was validated by real-time quantitative PCR (qPCR), and its segregation, and positive association with the richness and Shannon α-diversity of the porcine gut bacterial ecosystem was confirmed in an unrelated F1 (Duroc×Iberian) cross. Furthermore, despite genetic and environmental differences between both populations, the gut microbiota of DUP samples showed a significant over-abundance of theDesulfovibrio, Blautia, Phascolarctobacterium, Faecalibacterium, SuccinivibrioandAnaerovibriogenera.

Conclusions

In summary, this is the first study that evaluate the putative modulatory role of CNVs on pig gut microbiota. Our results advice the relevance of considering the role of host-genome structural variants as modulators of microbial ecosystems, and suggest theABCC2-DNMBPCNV as a host-genetic factor for the modulation of the diversity and composition of the gut microbiota in pigs.
A thermostable phospholipase C obtained by consensus design
Authors: Val Diego S. , Nardo Luisina Di , Marchisio Fiorela , Peiru Salvador , Castelli María Eugenia , Abriata Luciano , Menzella Hugo G. , Rasia Rodolfo M.
Published: October 2022

Proteins’ extraordinary performance in recognition and catalysis have led their use in a range of applications. But proteins obtained from natural sources are oftentimes not suitable for direct use in industrial or diagnostic setups. Natural proteins, evolved to optimally perform a task in physiological conditions, usually lack the stability required to be used in harsher conditions. Therefore, the alteration of the stability of proteins is commonly pursued in protein engineering studies.

Here we achieved a substantial thermal stabilization of a bacterial Zn(II) dependent phospholipase C by consensus sequence design. We retrieved and analyzed sequenced homologs from different sources, selecting a subset of examples for expression and characterization. A non-natural consensus sequence showed the highest stability and activity among those tested. Comparison of activity and stability parameters of this stabilized mutant and other natural variants bearing similar mutations allow us to pinpoint the sites most likely to be responsible for the enhancement. Point mutations in these sites alter the unfolding process of the consensus sequence. We show that the stabilized version of the protein retains full activity even in the harsh oil degumming conditions, making it suitable for industrial applications.
Simultaneous Membrane and RNA Binding by Tick-Borne Encephalitis Virus Capsid Protein
Authors: Pulkkinen L. I. A. , Barrass S. V. , Lindgren M. , Pace H. , Överby A. K. , Anastasina M. , Bally M. , Lundmark R. , Butcher S. J.
Published: October 2022

Tick-borne encephalitis virus is an enveloped, pathogenic, RNA virus in the family Flaviviridae, genus Flavivirus. Viral particles are formed when the nucleocapsid, consisting of an RNA genome and multiple copies of the capsid protein, buds through the endoplasmic reticulum membrane and acquires the viral envelope and the associated proteins. The coordination of the nucleocapsid components to the sites of assembly and budding are poorly understood. Here, we investigate nucleocapsid assembly by characterizing the interactions of the wild-type and truncated capsid proteins with membranes by using biophysical methods and model membrane systems. We show that capsid protein initially binds membranes via electrostatic interactions with negatively-charged lipids which is followed by membrane insertion. Additionally, we show that membrane-bound capsid protein can recruit viral genomic RNA. We confirm the biological relevance of the biophysical findings by using mass spectrometry to show that purified virions contain negatively-charged lipids. Our results suggest that nucleocapsid assembly is coordinated by negatively-charged membrane patches on the endoplasmic reticulum and that the capsid protein mediates direct contacts between the nucleocapsid and the membrane.
Introducing anrbcL andtrnL reference library to aid in the metabarcoding analysis of foraged plants from semi-arid eastern South African savannas
Authors: Botha Danielle , du Plessis Mornè , Siebert Frances , Barnard Sandra
Published: October 2022

The success of a metabarcoding study is determined by the extent of taxonomic coverage and the quality of records available in the DNA barcode reference database used. This study aimed to create anrbcLa andtrnL (UAA) DNA barcode sequence reference database of plant species that are potential herbivore foraging targets and commonly found in semi-arid savannas of eastern South Africa. A study-area-specific species list of 755 species was compiled. Thereafter, reference libraries forrbcLa andtrnL (UAA) sequences were created mined from sequence databases according to specific quality criteria to ensure accurate taxonomic coverage and resolution. The taxonomic reliability of these reference libraries was evaluated by testing for the presence of a barcode gap, identifying a data-appropriate identification threshold, and determining the identification accuracy of reference sequences via primary distance-based criteria. The finalrbcLa reference dataset consisted of 1238 sequences representing 318 genera and 562 species. The finaltrnL dataset consisted of 921 sequences representing 270 genera and 461 species. Barcode gaps were found for 76% of the taxa in therbcL barcode reference dataset and 68% of the taxa in thetrnL barcode reference dataset. The identification success rate, calculated with thek-nn criterion was 85.86% for therbcL dataset and 73.72% for thetrnL dataset. The datasets forrbcL andtrnL combined during this study are not presented as a complete DNA reference library, but rather as two datasets that should be used in unison to identify plants present in the semi-arid eastern savannas of South Africa.
Electron microscopy reveals toroidal shape of master neuronal cell differentiator REST – RE1-Silencing Transcription factor
Authors: Veverka Pavel , Brom Tomáš , Janovič Tomáš , Stojaspal Martin , Pinkas Matyáš , Nováček Jiří , Hofr Ctirad
Published: October 2022

The RE1-Silencing Transcription factor (REST) is essential for neuronal differentiation. Here, we report the first 18.5-angstrom electron microscopy structure of human REST. The refined electron map suggests that REST forms a torus that can accommodate DNA double-helix in the central hole. Additionally, we quantitatively described REST binding to the canonical DNA sequence of the neuron-restrictive silencer element. We developed protocols for the expression and purification of full-length REST and the shortened variant REST-N62 produced by alternative splicing. We tested the mutual interaction of full-length REST and the splicing variant REST-N62. Revealed structure-function relationships of master neuronal repressor REST will allow finding new biological ways of prevention and treatment of neurodegenerative disorders and diseases.
Live Cell Multicolour Lifetime Imaging Using Genetically Encodable Fluorophores
Authors: Starling Tobias , Carlon-Andres Irene , Iliopoulou Maro , Williamson David J. , Padilla-Parra Sergi
Published: October 2022

Nine fluorescent proteins (FPs) simultaneous imaging has been demonstrated in a single acquisition using fluorescence lifetime imaging microscopy (FLIM) combined with pulsed interleaved excitation (PIE) for three laser lines. We also show how to unmix spectrally similar FPs in a pixel-by-pixel manner with an analytical non-fitting solution.
Setd2 inactivation sensitizes lung adenocarcinoma to inhibitors of oxidative respiration and mTORC1 signaling
Authors: Walter David M. , Gladstein Amy C. , Doerig Katherine R. , Natesan Ramakrishnan , Baskaran Saravana G. , Gudiel A. Andrea , Adler Keren M. , Acosta Jonuelle O. , Wallace Douglas C. , Asangani Irfan A. , Feldser David M.
Published: October 2022

SETD2 is a tumor suppressor that is frequently inactivated in several cancer types. The mechanisms through which SETD2 inactivation promotes cancer are unclear, and whether targetable vulnerabilities exist in these tumors is unknown. Here we identify heightened mTORC1-associated gene expression programs and functionally higher levels of oxidative metabolism and protein synthesis as prominent consequences of Setd2 inactivation in KRAS-driven mouse models of lung adenocarcinoma. Blocking oxidative respiration and mTORC1 signaling abrogates the high rates of tumor cell proliferation and tumor growth specifically in SETD2-deficient tumors. Our data nominate SETD2 deficiency as a functional marker of sensitivity to clinically actionable therapeutics targeting oxidative respiration and mTORC1 signaling.
Inversion polymorphism in a complete human genome assembly
Authors: Porubsky David , Harvey William T. , Rozanski Allison N. , Ebler Jana , Höps Wolfram , Ashraf Hufsah , Hasenfeld Patrick ,   Human Pangenome Reference Consortium (HPRC),   Human Genome Structural Variation Consortium (HGSVC), Paten Benedict , Sanders Ashley D. , Marschall Tobias , Korbel Jan O. , Eichler Evan E.
Published: October 2022

The completion of the human genome significantly improved our ability to discover and interpret genome copy number variation. In order to understand its impact on the characterization of inversion polymorphisms, we remapped data from 41 human genomes and 10 new samples against the telomere-to-telomere (T2T) reference genome as compared to the standard GRCh38 reference. Our analysis shows a ~21% increase in sensitivity identifying and improving mapping of 63 inversions. We further identify 26 misorientations within GRCh38, and show that the T2T reference is three times more likely to represent the correct orientation of the major human allele. As a result, we report a significant bias for inversions accumulating within the pericentromeric regions of specific chromosomes and show that functional annotations around inverted regions, such as topological-associated domains, can be better interpreted.
A novel nuclease is the executing part of a bacterial plasmid defense system
Authors: Weiß Manuela , Giacomelli Giacomo , Ben Assaya Mathilde , Grundt Finja , Haouz Ahmed , Peng Feng , Petrella Stéphanie , Wehenkel Anne Marie , Bramkamp Marc
Published: October 2022

Cells are continuously facing the risk of taking up foreign DNA that can compromise genomic and cellular integrity. Therefore, bacteria are in a constant arms race with mobile genetic elements such as phages, transposons and plasmids. They have developed several active strategies against invading DNA molecules that can be seen as a bacterial ‘innate immune system’. Anti-phage systems are usually organized in ‘defense islands’ and can consist of restriction-modification (R-M) systems, CRISPR-Cas, and abortive infection (Abi) systems. Despite recent advances in the field, much less is known about plasmid defense systems. We have recently identified the MksBEFG system inCorynebacterium glutamicumas a novel plasmid defense system which comprises homologues of the condensin system MukFEB. Here, we investigated the molecular arrangement of the MksBEFG complex. Importantly, we identified MksG as a novel nuclease that degrades plasmid DNA and is, thus, the executing part of the system. The crystal structure of MksG revealed a dimeric assembly through its DUF2220 C-terminal domains. This domain is homologous to the TOPRIM domain of the topoisomerase II family of enzymes and contains the corresponding divalent ion binding site that is essential for DNA cleavage in topoisomerases, explaining thein vitronuclease activity of MksG. We further show that the MksBEF subunits exhibit an ATPase cycle similar to MukBEFin vitroand we reason that this reaction cycle, in combination with the nuclease activity provided by MksG, allows for processive degradation of invading plasmids. Super-resolution localization microscopy revealed that the Mks system is spatially regulated via to the polar scaffold protein DivIVA. Introduction of plasmids increases the diffusion rate and alters the localization of MksG, indicating an activation of the systemin vivo.
Phosphorylation chemistry of the Bordetella PlrSR TCS and its contribution to bacterial persistence in the lower respiratory tract
Authors: Barr Sarah A. , Kennedy Emily N. , McKay Liliana S. , Johnson Richard M. , Ohr Ryan J. , Cotter Peggy A. , Bourret Robert B.
Published: October 2022

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Bordetella species cause lower respiratory tract infections in mammals. B. pertussis and B. bronchiseptica are the causative agents of whooping cough and kennel cough, respectively. The current acellular vaccine for B. pertussis protects against the pertussis toxin but does not prevent transmission or colonization. Cases of B. pertussis infections are on the rise even in areas of high vaccination. The PlrSR two-component system, is required for persistence in the mouse lung. A partial plrS deletion strain and a plrS H521Q strain cannot survive past three days in the lung, suggesting PlrSR works in a phosphorylation dependent mechanism. We characterized the biochemistry of B. bronchiseptica PlrSR and found that both proteins function as a canonical two-component system. His521 and Glu522 were essential for PlrS autophosphorylation. Asn525 was essential for phosphatase activity. The PAS domain was critical for both PlrS autophosphorylation and phosphatase activities. PlrS can both phosphotransfer to and exert phosphatase activity towards PlrR. Unexpectedly, PlrR forms a tetramer when unphosphorylated and a dimer upon phosphorylation. Finally, we demonstrated the importance of PlrS phosphatase activity for persistence within the murine lung. By characterizing PlrSR we hope to guide future in vivo investigation for development of new vaccines and therapeutics.

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GRAPHICAL ABSTRACT
Growth performance and nutritional composition of Clarias gariepinus in flood-and-drain aquaponics coupled with three species of leafy vegetable
Authors: Olanrewaju Gbolaga O. , Sarpong David D. , Aremu Abiola O. , Ade-Ademilua Elizabeth O.
Published: October 2022

Recirculating aquaponics integrate aquaculture and hydroponics with the aid of microorganisms to ensure a sustainable supply of fish and vegetables. In this study, we designed and constructed a flood-and-drain aquaponic system with Clarias gariepinus as the aquaculture and Celosia argentea, Corchorus olitorius, and Ocimum gratissimum as plant components. Nitrogenous waste from the aquaculture unit was circulated to the plant growth beds, which served as bio-oxidizers of toxic ammonia to nitrate and returned less toxic water back to the aquaculture unit. An evaluation of the growth parameters of C. gariepinus in aquaponics and control fish tanks revealed that aquaponics-raised C. gariepinus gained an additional 205.6% of their initial mean weight, whereas those grown in the control fish tank gained an additional 182.2% of their initial mean weight. The majority (37.5%) of aquaponics-raised C. gariepinus weighed 750 g -1 kg, whereas the highest percentage of C. gariepinus raised in the control fish tank (23.4%) weighed 500 g - 700 g. C. gariepinus raised in aquaponics had significantly higher gross feed conversion efficiency and protein efficiency ratio than those raised in the control tank. The mortality rate in the aquaponic fish tank was 0% compared to the 11.43% mortality rate in the control fish tank. There was no significant difference in the nutritional composition of C. gariepinus raised in either tank; however, the aquaponic fish tank had a higher nitrogen retention rate. This study showed that C. gariepinus raised in aquaponics had better biomass accumulation than those raised in conventional fishponds.
Incipient parallel evolution of SARS-CoV-2 Deltacron variant in South Brazil
Authors: Sant’Anna Fernando Hayashi , Andreis Tiago Finger , Salvato Richard Steiner , Muterle Ana Paula , Comerlato Juliana , Gregianini Tatiana Schaffer , Barcellos Regina Bones , Godinho Fernanda Marques , Resende Paola Cristina , da Luz Wallau Gabriel , Castro Thaís Regina y , Casarin Bruna Campestrini , de Almeida Vieira Andressa , Schwarzbold Alexandre Vargas , de Arruda Trindade Priscila , Giannini Gabriela Luchiari Tumioto , Freese Luana , Bristot Giovana , Brasil Carolina Serpa , de Oliveira Rocha Bruna , Martins Paloma Bortolini , de Oliveira Francine Hehn , van Oosterhout Cock , Wendland Eliana
Published: October 2022

With the coexistence of multiple lineages and increased international travel, recombination and gene flow are likely to become increasingly important in the adaptive evolution of SARS-CoV-2. This could result in the incipient parallel evolution of multiple recombinant lineages. However, identifying recombinant lineages is challenging, and the true extent of recombinant evolution in SARS-CoV-2 may be underestimated. This study describes the first SARS-CoV-2 Deltacron recombinant case identified in Brazil. We demonstrate that the recombination breakpoint is at the beginning of Spike gene (S). The 5′ genome portion (circa 22 kb) resembles the AY.101 lineage (VOC Delta), and the 3′ genome portion (circa 8 kb nucleotides) is most similar to the BA.1.1 lineage (VOC Omicron). Furthermore, evolutionary genomic analyses indicate that the new strain emerged after a single recombination event between lineages of diverse geographical locations in December 2021 in South Brazil. This Deltacron, named AYBA-RS, is one out of almost 30 recombinants described this year. The submission of only four sequences in the GISAID database suggests that this Brazilian lineage had a minor epidemiological impact. On the other hand, the recent emergence of this and various other Deltacron recombinant lineages (i.e., XD, XF, and XS) suggests that gene flow and recombination may play an increasingly important role in the COVID-19 pandemic. We explain the evolutionary and population genetic theory that support this assertion, and we conclude that this stresses the need for continued genomic and epidemiological surveillance. This is particularly important for countries where multiple variants are present, as well as for countries that receive significant inbound international travel.