bluuebunny/crossref_metadata_2025 huggingface dataset:
Genomic epidemiology of the first London outbreak of antimicrobial resistant sexually transmitted shigellosis
Authors: Mason Lewis , Dallman Timothy , Jenkins Claire , Baker Kate
Published: May 2022

Shigellosis is an intestinal infection caused by Shigella bacteria. Shigella cause an estimated ~200,000 global deaths annually. Antimicrobial resistant (AMR) shigellosis is a significant cause of morbidity in high-income nations, with both multidrug resistant (MDR) and extensively drug resistant (XDR) cases being increasingly reported in Australia, England, and the USA. Sexually transmissible shigellosis was first described in San Francisco, 1974, but it would be a further 30 years before its first description in England. In 2004, London experienced an outbreak of Shigella sonnei (S. sonnei) mediated sexually transmitted shigellosis, associated with men-who-have-sex-with-men (MSM). Since then, sexually transmissible shigellosis has become endemic in England, with a greater than two-fold increase in Shigella diagnoses within sexual health services from 2015 to 2019. Through genomic exploration of samples from the original 2004 outbreak (provided by Public Health England (PHE)), we identified that the 2004 London outbreak isolates clustered within the base of genotype 3.1, lineage III, a lineage which has since gone on to dominate the global epidemiology of S. sonnei. The isolates displayed early evidence of varying degrees of antimicrobial resistance to several drug classes: macrolides, tetracyclines, beta-lactams and sulphonamides. Reconstructing the chronological process of how shigellosis has arrived at its current position in AMR and transmissibility is critical. Further investigation is underway to link this outbreak with MSM-associated shigellosis outbreaks occurring in the early 2000s in other countries to establish whether this lineage globally disseminated; determine the timeframe for global connectivity of shigellosis; and examine the outbreak isolates for virulence determinants.
Engineered anti-viral shRNAs are more effective than lhRNAs in transgenic Aedes aegypti
Authors: Tng Priscilla , Paladino Leonela Carabajal , Anderson Michelle , Dhokiya Vishaal , Keen Elizabeth , Adelman Zach , Fragkoudis Rennos , Noad Rob , Alphey Luke
Published: May 2022

The Aedes aegypti mosquito is a major vector of chikungunya virus (CHIKV), which has no licensed vaccine. Engineered mosquitoes expressing long RNA hairpins (lhRNA) or small RNAs against selected arboviruses have been developed to limit virus replication, but their silencing efficiency has not been compared.

We developed lhRNA and short hairpin RNA (shRNA) arrays against CHIKV non-structural protein nsP2. We used a Tet-response element (TRE) and a tTA transactivator to control expression of lhRNAs (TRE-lhRNA) and shRNAs (TRE-shRNA). Constitutive expression in Aag2 cells was assessed with a PUb-tTA driverand midgut specific expression in transgenic mosquitoes was assessed using Carboxypeptidase A (AeCPA)-tTA as driver. In vitrointerference ability was determined with a CHIKV split replication system, and a synthetic luciferase reporter with mRNA containing the targeted CHIKV sequence (CHIK-FF). In vivo interference was tested by inserting the TRE-lhRNA and TRE-shRNA constructs into a AeCPA-tTA line so both constructs expressed from the same locus, and a TRE reporter line that expressed an AmCyan reporter with an N terminal CHIKV fusion (CHIK-AmC).

In Aag2 cells, shRNAs were more effective than lhRNA in silencing both the CHIKV split replication system (99.7%, S. D. ± 0.47 and 72.8%, S. D. ± 9.50, respectively, P<0.001) and CHIK-FF (98.8%, S. D. ± 0.71 and 50.9%, S. D. ± 7.92, respectively, P<0.05). Similar results were observed in transgenic mosquitoes when comparing AmC expression in the midgut.

This study demonstrates that, in mosquitoes, effectively chosen shRNAs can induce greater interference of the desired viral target than the corresponding lhRNA.
Longevity and neutralisation activity of secretory IgA following SARS-CoV-2 infection
Authors: Ellis Samuel , Burleigh Alice , Way Rosie , Kembou-Ringert Japhette , Masonou Tereza , Gilmour Kimberly , Grandjean Louis , Smith Claire
Published: May 2022

The mucosal barrier is a primary defence against inhaled pathogens, comprising secretory antibodies which have the potential to block viral entry and neutralise infection. There is an ongoing need for greater understanding of the mucosal immunity to SARS-CoV-2 infection. In this study, we investigated mucosal IgA through non-invasive saliva sampling of healthcare workers.

A total of 551 saliva samples were collected from staff at Great Ormond Street Children’s Hospital who previously tested positive for COVID-19. Participant metadata included age, gender, ethnicity and symptoms. IgA titres were measured by ELISA against viral antigens spike protein, nucleocapsid protein, and spike receptor-binding domain. SARS-CoV-2 neutralisation was measured using a VERO E6 cell culture infection assay.

We found that approximately 30% of saliva samples contained detectable IgA specific for at least one of the SARS-CoV-2 antigens. IgA levels in saliva decreased with the time post-infection, and were largely undetectable after six months. IgA titres specific to SARS-CoV-2 were lowest in participants over 60 years old. Specific saliva samples were identified which effectively neutralised SARS-CoV-2 virus infection of epithelial cells.

Our results suggest secretory IgA specific to SARS-CoV-2 can be detected in saliva following infection, an accessible sample type for testing, although titres decreased over time. Some saliva samples were able to neutralise SARS-CoV-2 infectivity against cultured epithelial cells. This data could be used to assess the risk of re-infection with SARS-CoV-2, as well as accelerate efforts to develop effective mucosal vaccination with longer lasting protection.
Exploring foot-and-mouth disease virus antibody interactions using biolayer interferometry
Authors: Shaw Andrew , Burman Alison , Asfor Amin , Ludi Anna , Brocchi Emiliana , Grazioli Santina , King Donald
Published: May 2022

Foot-and-mouth disease virus (FMDV) vaccines protect animals from infection by inducing antibodies. The level of neutralising antibody induced in response to vaccination (or infection), as measured by a virus neutralisation test, is an important parameter with regards to the level of protection afforded against subsequent challenge. However, in addition to overall titre, antibody avidity also represents a crucial metric when assessing the protective efficacy of antibodies. In this project we investigated the use of biolayer interferometry (BLI) to measure the avidity of FMDV antibodies to FMDV antigens. Antibodies targeting site I of the FMDV particle were detected using a commercially synthesised biotinylated peptide. In contrast, the entire antigenic landscape of the FMDV particle was represented by biotinylated FMDV capsids. The antigens were loaded onto Octet streptavidin biosensors at an optimal concentration prior to dipping into antibodies. The sera from different animals varied in avidity, reflecting the quantitative differences in avidity that exist between individual animals in response to FMDV vaccines. Interestingly, the Kdis values obtained for site I vs the entire capsid were different, supporting the importance of other sites beyond site I. Similarly, monoclonal antibodies targeting distinct, known antigenic sites on the capsid surface also resulted in different avidities. The BLI methodology reported here offers a useful approach by which to investigate the strength of antibody interactions at specific sites. In conjunction with recombinant technology, BLI will help aid in investigations into the relative importance of the different antigenic sites with regards to inducing a protective response.
Automated optimization of deep brain stimulation parameters for modulating neuroimaging-based targets
Authors: Malekmohammadi Mahsa , Mustakos Richard , Sheth Sameer , Pouratian Nader , McIntyre Cameron C. , Bijanki Kelly R. , Tsolaki Evangelia , Chiu Kevin , Robinson Meghan E. , Adkinson Joshua A. , Oswalt Denise , Carcieri Stephen
Published: May 2022

Objective

Therapeutic efficacy of deep brain stimulation (DBS) in both established and emerging indications, is highly dependent on accurate lead placement and optimized clinical programming. The latter relies on clinicians’ experience to search among available sets of stimulation parameters and can be limited by the time constraints of clinical practice. Recent innovations in device technology have expanded the number of possible electrode configurations and parameter sets available to clinicians, amplifying the challenge of time constraints. We hypothesize that patient specific neuroimaging data which can effectively assist the clinical programming using automated algorithms.

Approach

This paper introduces the DBS Illumina 3D algorithm as a tool which uses patient-specific imaging to find stimulation settings that optimizes activating a target area while minimizing the stimulation of areas outside the target that could result in unknown or undesired side effects. This approach utilizes preoperative neuroimaging data paired with the postoperative reconstruction of lead trajectory to search the available stimulation space and identify optimized stimulation parameters. We describe the application of this algorithm in three patients with treatment-resistant depression who underwent bilateral implantation of DBS in subcallosal cingulate cortex (SCC) and ventral capsule/ventral striatum (VC/VS) using tractography optimized targeting with an imaging defined target previously described.

Main results

Compared to the stimulation settings selected by the clinicians (informed by anatomy), stimulation settings produced by the algorithm achieved similar or greater target coverage, while producing a significantly smaller stimulation area that spills outside the target (P=0.002).

Significance

The DBS Illumina 3D algorithm is seamlessly integrated with the clinician programmer software and effectively and rapidly assists clinicians with the analysis of image based anatomy, and provides a starting point for the clinicians to search the highly complex stimulation parameter space and arrive at the stimulation settings that optimize activating a target area.

Clinical trial registration number: NCT 03437928
Relationship between finger movement characteristics and voxel-based specific regional analysis systems for Alzheimer’s disease
Authors: Sugioka Junpei , Suzumura Shota , Kuno Katsumi , Kizuka Shiori , Sakurai Hiroaki , Kanada Yoshikiyo , Mizuguchi Tomohiko , Kondo Izumi
Published: May 2022

Background

Aging is the most significant risk factor of dementia. Alzheimer’s disease accounts for approximately 60%–80% of all dementia cases in elderly individuals. This study aimed to examine the relationship between finger movements and brain function in patients with Alzheimer’s disease, using VSRAD, and explore the relationship between VSRAD and cognitive function (MoCA-J).

Methods

Patients diagnosed with AD, at the Center for Comprehensive Care and Research on Memory Disorders, were included in the study. The diagnostic criteria were based on the National Institute on Aging-Alzheimer’s Association. Patients were excluded based on set criteria. A finger-tapping device was used for all measurements. Participants performed the following task, in order: non-dominant hand, dominant hand, simultaneously, and alternate hands. Movements were measured for 15 s each. The relationship between distance and output was measured. MRI measurements were performed, after which VSRAD was performed using sagittal section 3D T1-weighted images. The Z-score was used to calculate the degree of atrophy. Pearson’s product-moment correlation coefficient was used to analyze the relationship between the Z-score and mean values of the parameters in the finger-tapping movements. Statistical significance was set at p < 0.05.

Results

Sixty-two patients were included in the study. Z-scores for MoCA-J analysis, percentage of total brain atrophy in the white matter, and other VSRAD parameters showed varying negative correlations (r = -0. 28, p = 0. 035, r = -0. 31, p = 0. 020, and r = -0. 52, p < 0. 001), respectively. Varying positive correlations between Z-score and the SD of distance rate of velocity peak in extending movements in the non-dominant hand were observed.

Conclusion

The SD of distance rate of velocity peak in extending movements and the Z-score showed a significant relationship. An association between neuropsychological tests and overall degree of brain atrophy was also observed.
Effects of virtual reality-based aerobic endurance training on the functional fitness of healthy older adults: A systematic review
Authors: Restrepo Daniela Ramírez , Ruiz Julialba Castellanos , Mesa Lina María Montealegre , Narváez Carolina Márquez , Rendón Santiago Murillo
Published: May 2022

Objetive

to analytically and systematically review and integrate the available evidence of the effects of virtual reality-based aerobic endurance training on functional fitness of healthy older adults.

Materials and Methods

This systematic review of randomized controlled trials was conducted through searches in Pubmed, Science Direct, Scopus, PEDro, Web of Science, Lilacs, Scielo, Dialnet, and IEEE Xplore between February and May 2021. The evaluation of bias and methodological quality was performed following the parameters proposed by the Cochrane Manual of Systematic Reviews of Interventions and the PEDro Scale. Review Manager Software (Version 5.4.1) served for a summary of bias.

Results

Three clinical trials were selected. None of the selected studies employed any exclusively aerobic intervention with virtual reality; however, statistically significant findings for this type of intervention were found in functional fitness such as aerobic endurance, upper and lower limb strength, agility, and dynamic balance.

Conclusions

aerobic endurance training through the use of virtual reality may provide positive effects on the functional fitness of the healthy elderly. This has implications for clinical practice, as it fosters the development of more beneficial, attractive, motivational, and quality interventions, the prevention of common conditions, and the use of technological tools by the elderly population. However, as for research implications, an increase of knowledge on the effects of technologies in the elderly functional capacity, and an implementation of scientific models or theories on physical activity and exercise are recommended with the purpose of explaining the phenomena due to the use of technologies within the processes of functional rehabilitation.
Health workers’ Perspective on the Feasibility and Acceptability of the Introduction of AgRDT for COVID-19 in Kisumu County, Western Kenya
Authors: Omollo M. , Odero I. A. , Barsosio H.C. , Kariuki S. , Kuile F. Ter , Okello S.O. , Oyoo K. , K’Oloo A. , Otieno K. , Duijn S. van , Houben N. , Milimo E. , Aroka R. , Odhiambo A. , Onsongo S. N. , de Wit T.F. Rinke
Published: May 2022

COVID-19 pandemic remains a major global public health challenge also in Low- and Middle-Income Countries (LMIC), due to fragile health systems, limited resources and personnel, low testing and counseling capacity, community perceptions, among others. In Kisumu County of Western Kenya, a unique Public Private Partnership (PPP) was rolled-out to increase testing and capacity building by linking private facilities to the ongoing public sector efforts in combating COVID-19. It became increasingly clear that centralized PCR testing for COVID-19 was too labor-intensive, expensive, prone to machine breakdowns and stock-outs of essential reagents, resulting in long turn-around times and sometimes even adaptations of patient selection criteria. A clear need was identified for rapid point-of-care COVID-19 testing (AgRDT). After successful field evaluation, RDT for COVID-19 was offered through the PPP. This paper aimed to understand the health workers perspective on the feasibility and acceptability of the introduction of the AgRDT in Kisumu County.

In-Depth Interviews were conducted with selected health workers (n=23) from the participating facilities and analyzed using Nvivo 11 The health workers accepted the use of AgRDT as it enabled the strengthening of the existing health system, increased testing capacity and provided capacity building opportunities. Challenges included poor management of results discrepant with PCR gold standard.

The health workers applauded the introduction of AgRDT with the Kisumu County Department of Health as a more realistic and user-friendly approach, leading to fast turn-around times and increased personal safety experience.
Antibiogram of Clinical Isolates from Primary and Secondary Healthcare Facilities: A Step Towards Antimicrobial Stewardship
Authors: Aika Isabel Naomi , Enato Ehijie
Published: May 2022

Antibiogram development and use is a core element of antimicrobial stewardship practice, such data is scarce in healthcare settings in developing countries. The study aims to determine the epidemiology of clinical isolates and their antibiograms in secondary healthcare (SHC) and primary healthcare (PHC) facilities in Benin City, Nigeria. This was a retrospective study in three laboratories in SHC and PHC facilities. Microbial culture and susceptibility report over the past 4 years was collated. Ethical Clearance was obtained from the Edo State Ministry of Health and Benin City. Data were entered and analyzed using SPSS version 22. Of the 819 isolates, urine, semen and vagina swab were most represented (50.7%; 16.1%; 13.2%). S.aureus (60.6%) and coliform organism (31.9%) were commonly isolated. High resistance of 75%->90% was seen with penicillin, cephalosporin, macrolide, aminoglycoside and floroquinolone against S. aureus, Ps aeruginosa, coliform and E. coli. Resistance to all antimicrobials was observed in 11.7% of the isolates, multidrug resistance (MDR) was found to be 61.4%. MDR for Ps aeruginosa, coliform, E coli, and staph aureus were 95.5%, 67.3% 25.6% and 82.8% respectively. In the SHC, 15.4% of isolates were resistant to all antibacterial compared to none in the PHC. There was consistent yearly increase in resistance to more than six agents in both centers. Gender difference in antimicrobial resistance was observed. High MDR observed in this study emphasizes the need for routine antibiogram and its use in updating treatment guidelines to reflect the current resistance pattern to available antimicrobials.
Brain injury accelerates the onset of a reversible age-related microglial phenotype associated with hyperphagocytosis and inflammatory neurodegeneration
Authors: Ritzel Rodney M. , Li Yun , Jiao Yun , Lei Zhuofan , Doran Sarah J. , He Junyun , Shahror Rami A. , Henry Rebecca J. , Liu Shaolin , Stoica Bogdan A. , Faden Alan I. , Szeto Gregory , Loane David J. , Wu Junfang
Published: May 2022

Lipofuscin is an autofluorescent (AF) pigment formed by lipids and misfolded proteins that accumulates in post-mitotic cells with advanced age. Here we immunophenotyped microglia in the brain of old C57BL/6 mice (>18 months-old) and demonstrate that in comparison to young mice, one third of old microglia are AF, characterized by profound changes in lipid and iron content, phagocytic activity, and oxidative stress. Pharmacological depletion of microglia in old mice eliminated the AF microglia following repopulation and reversed microglial dysfunction. Age-related neurological deficits and neurodegeneration after traumatic brain injury (TBI) were attenuated in old mice lacking AF microglia. Furthermore, hyperphagocytic activity and lipid accumulation in microglia persisted for up to one year after TBI, were modified by Apoe4 genotype, and chronically driven by phagocyte-mediated oxidative stress. Thus, AF may reflect a pathological state in aging microglia associated with hyperphagocytosis and inflammatory neurodegeneration that can be further accelerated by TBI.

Teaser

Traumatic brain injury accelerates age-related pathological phagocytosis and lipofuscin formation in microglia.
The Effects of Propofol Anaesthesia on Molecular-enriched Networks During Resting-state and Naturalistic Stimulation
Authors: Lawn Timothy , Martins Daniel , O’Daly Owen , Williams Steve , Howard Matthew , Dipasquale Ottavia
Published: May 2022

Placing a patient in a state of anaesthesia is crucial for modern surgical practice. However, the mechanisms by which anaesthetic drugs, such as propofol, impart their effects on consciousness remain poorly understood. Propofol potentiates GABAergic transmission, which purportedly has direct actions on cortex as well as indirect actions via ascending neuromodulatory systems. Functional imaging studies to date have been limited in their ability to unravel how these effects on neurotransmission impact system-level dynamics of the brain. Here, we leveraged advances in multi-modal imaging, Receptor-Enriched Analysis of functional Connectivity by Targets (REACT), to investigate how different levels of propofol-induced sedation alters neurotransmission-related functional connectivity (FC), both at rest and when individuals are exposed to naturalistic auditory stimulation. Propofol increased GABA-A- and noradrenaline transporter-enriched FC within occipital and somatosensory regions respectively. Additionally, during auditory stimulation, the network related to the vesicular acetylcholine transporter showed reduced FC within the right superior temporal gyrus, regardless of level of anaesthesia, and a spatial configuration correlating with a broad range of meta-analytic measures of audition- and emotion-related cognition. In bringing together these micro- and macro-scale systems, we provide support for both direct GABAergic and indirect noradrenergic-related network changes under anaesthesia and describe a cognition-related reconfiguration of the cholinergic network, highlighting the utility of REACT to explore the molecular substrates consciousness and cognition.
Influenza Virus-Induced Autophagy Is Not Always Associated with MUC1 Expressions
Authors: Ozkan Hasan Selcuk , Ozarslan Mustafa Alper , Vatansever Gokhan , Cicek Candan
Published: May 2022

Influenza virus-induced autophagy frequently accompanies apoptosis and results in cell death in cells infected with the virus. Autophagy has been well-known to be modulated by the mTOR/PI3K/Akt pathway, which plays an important role in response to the presence of energy sources and external stimulants. This pathway can also be modulated by MUC1, which has extracellular and intracellular components, and playing an important role in metastasis and chemotherapeutic resistance. In this study, the aim is to observe the changes in MUC1 expressions, which is known to have sialic acid residues within MUC1, therefore serving as a receptor for influenza viruses, and consequent changes in autophagy markers such as mTOR and LC3b, after inoculation of cancer cells with influenza virus.

Fluorescence was detected for LC3b, mTOR and MUC1 in all influenza-positive cell lines (MCF-7, He-La, A-549) except for RD in immunofluorescence studies. In influenza-negative MCF-7, HeLa, RD and A-549 cells no fluorescence was detected. In the supernatant of all influenza-positive cells, except for RD cells, positive results were obtained for MUC1, MAP1LC3B and MTOR genes (Ct 20,01-32,04) in RT-qPCR, respectively encoding for MUC1, LC3b, and mTOR. In RT-qPCR, in cell lines without influenza inoculation, only in the A-549 cells, the gene expressions were found to be negative. In other cell lines, positive results were obtained (Ct 21,98-23,78). In RD cells, which have not been inoculated with the virus, all pathways were positive.

In cells with adenocarcinoma structure, the alterations and presence of autophagy pathways and MUC1 expressions activated via influenza viruses were confirmed. However, this has not been proved using nucleic acid assays. The reason why might be the detection of only gene expressions and mRNA presence via the tests used in the study. In future studies, expressions of these genes in protein level should be detected using more advanced tests.
High-throughput single-molecule quantification of individual base stacking energies in nucleic acids
Authors: Punnoose Jibin Abraham , Thomas Kevin J. , Chandrasekaran Arun Richard , Vilcapoma Javier , Hayden Andrew , Kilpatrick Kacey , Vangaveti Sweta , Chen Alan , Banco Thomas , Halvorsen Ken
Published: May 2022

Base stacking interactions between adjacent bases in DNA and RNA are known to be important for many biological processes, for drug development, and in other biotechnology applications. While previous work has estimated base stacking energies between pairs of bases, the individual contributions of each base to the stacking interaction has remained unknown. Here, we developed a novel methodology using a custom Centrifuge Force Microscope to perform high-throughput single molecule experiments to measure base stacking energies between individual adjacent bases. We found stacking energies strongest between purines (G|A at −2.3 ± 0.2 kcal/mol) and weakest between pyrimidines (C|T at −0.4 ± 0.1 kcal/mol). Hybrid stacking with phosphorylated, methylated, and RNA bases had no measurable effect, but a fluorophore modification reduced stacking energy. The implications of the work are demonstrated with three applications. We experimentally show that base stacking design can influence assembly and stability of a DNA nanostructure, modulate kinetics of enzymatic ligation, and determine accuracy of force fields in molecular dynamics (MD) simulations. Our results provide new insights into fundamental DNA interactions that are critical in biology and can inform rational design in diverse biotechnology applications.
Targeted hydrolysis of native potato protein: A novel route for obtaining hydrolysates with improved interfacial properties
Authors: Echers Simon Gregersen , Jafarpour Ali , Yesiltas Betül , García-Moreno Pedro J. , Greve-Poulsen Mathias , Hansen Dennis , Jacobsen Charlotte , Overgaard Michael Toft , Hansen Egon Bech
Published: May 2022

Peptides and protein hydrolysates are promising alternatives to substitute chemical additives as functional food ingredients. In this study, we present a novel approach for producing a potato protein hydrolysate with improved emulsifying and foaming properties by data-driven, targeted hydrolysis. Based on previous studies, we selected 15 emulsifier peptides derived from abundant potato proteins, which were clustered based on sequence identity. Through in silico analysis, we determined that from a range of industrial proteases (Neutrase (Neut), Alcalase (Alc), Flavorzyme (Flav) and Trypsin (Tryp)), Tryp was found more likely to release peptides resembling the target peptides. After applying all proteases individually, hydrolysates were assayed for in vitro emulsifying and foaming properties. No direct correlation between degree of hydrolysis and interfacial properties was found. Tryp produced a hydrolysate (DH=5.4%) with the highest (P<0.05) emulsifying and foaming abilities, good stabilities, and high aqueous solubility. Using LC-MS/MS, we identified >10,000 peptides in each hydrolysate. Through peptide mapping, we show that random overlapping with known peptide emulsifiers is not sufficient to quantitatively describe hydrolysate functionality. While Neut hydrolysates had the highest proportion of peptides with target overlap, they showed inferior interfacial activity. In contrast, Tryp was able to release specifically targeted peptides, explaining the high surface activity observed. While modest yields and residual unhydrolyzed protein indicate room for process improvement, this work shows that data-driven, targeted hydrolysis is a viable, interdisciplinary approach to facilitate hydrolysis design for production of functional hydrolysates from alternative protein sources.
Cardiac fibroblasts regulate cardiomyocyte hypertrophy through dynamic regulation of type I collagen
Authors: Meng Qinghang , Bowers Stephanie L. K. , Kuwabara Yasuhide , Huo Jiuzhou , Minerath Rachel , York Allen J. , Sargent Michelle A. , Prasad Vikram , Saviola Anthony J. , Galindo David Ceja , Hansen Kirk C. , Vagnozzi Ronald J. , Yutzey Katherine E. , Molkentin Jeffery D.
Published: May 2022

Rationale

Cardiomyocytes and fibroblasts in the heart communicate through both secreted growth factors as well as through sensing the structural properties of the extracellular matrix that each helps generate. Previous studies have shown that defects in fibroblast activity during disease stimulation result in altered cardiomyocyte hypertrophy, although the role that collagen might play in this communication is unknown.

Objective

Here we investigated how type I collagen maturation and disease-responsive matrix expansion in the heart by cardiac fibroblasts impacts cardiac fibrosis and cardiomyocyte hypertrophy.

Methods and Results

We generated and characterized Col1a2-/- mice using standard gene-targeting. Col1a2-/- mice were viable, although by young adulthood their hearts showed alterations in extracellular matrix mechanical properties, as well as an unanticipated activation of cardiac fibroblasts and induction of a progressive fibrotic response. This included increases in fibroblast number and a progressive cardiac hypertrophy, with reduced functional performance by 9 months. Col1a2-loxP targeted mice were also generated and crossed with the tamoxifen-inducible Postn-MerCreMer knock-in mice to delete the Col1a2 gene in myofibroblasts post-pressure overload injury, to more specifically implicate fibroblasts as effectors of cardiomyocyte hypertrophy in vivo. Opposite to the gradual induction of cardiac hypertrophy observed in germline Col1a2-/- mice as they matured developmentally, adult fibroblast-specific deletion of Col1a2 during pressure overload protected these mice from cardiac hypertrophy in the first week with a delayed fibrotic response. However, this reduction in hypertrophy due to myofibroblast-specific Col1a2 deletion was gradually lost over 2 and 6 weeks of pressure overload as augmented fibrosis returned.

Conclusions

Defective type I collagen in the developing heart alters the structural integrity of the extracellular matrix that leads to fibroblast expansion, activation, fibrosis and hypertrophy with progressive cardiomyopathy in adulthood. However, acute deletion of type I collagen production for the first time in the adult heart during pressure overload prevents ECM expansion and inhibits cardiomyocyte hypertrophy, while gradual restoration of fibrosis again permitted hypertrophy comparable to controls.
De novo mapping of the apicomplexan Ca2+-responsive proteome
Authors: Herneisen Alice L. , Li Zhu-Hong , Chan Alex W. , Moreno Silvia NJ , Lourido Sebastian
Published: May 2022

Apicomplexan parasites cause persistent mortality and morbidity worldwide through diseases including malaria, toxoplasmosis, and cryptosporidiosis. Ca2+ signaling pathways have been repurposed in these eukaryotic pathogens to regulate parasite-specific cellular processes governing the transition between the replicative and lytic phases of the infectious cycle. Despite the presence of conserved Ca2+-responsive proteins, little is known about how specific signaling elements interact to impact pathogenesis. We mapped the Ca2+-responsive proteome of the model apicomplexan T. gondii via time-resolved phosphoproteomics and thermal proteome profiling. The waves of phosphoregulation following PKG activation and stimulated Ca2+ release corroborate known physiological changes but identify specific proteins operating in these pathways. Thermal profiling of parasite extracts identified many expected Ca2+-responsive proteins, such as parasite Ca2+-dependent protein kinases. Our approach also identified numerous Ca2+-responsive proteins that are not predicted to bind Ca2+, yet are critical components of the parasite signaling network. We characterized protein phosphatase 1 (PP1) as a Ca2+-responsive enzyme that relocalized to the parasite apex upon Ca2+ store release. Conditional depletion of PP1 revealed that the phosphatase regulates Ca2+ uptake to promote parasite motility. PP1 may thus be partly responsible for Ca2+-regulated serine/threonine phosphatase activity in apicomplexan parasites.
Large RNP granules in C. elegans oocytes have distinct phases of RNA binding proteins
Authors: Elaswad Mohamed T. , Watkins Brooklynne M. , Sharp Katherine G. , Munderloh Chloe , Schisa Jennifer A.
Published: May 2022

The germ line provides an excellent in vivo system to study the regulation and function of RNP granules. Germ granules are conserved germ line-specific RNP granules that are positioned in the C. elegans adult gonad to function in RNA maintenance, regulation, and surveillance. In C. elegans, when oogenesis undergoes an extended meiotic arrest, germ granule proteins and other RNA binding proteins assemble into much larger RNP granules whose hypothesized function is to regulate RNA metabolism and maintain oocyte quality. To gain insight into the function of oocyte RNP granules, in this report we characterize distinct phases for four protein components of RNP granules in arrested oocytes. We find the RNA binding protein PGL-1 is dynamic and has liquid-like properties, while the intrinsically disordered protein MEG-3 has gel-like properties, similar to the properties of the two proteins in small germ granules of embryos. We find that MEX-3 exhibits several gel-like properties but is more dynamic than MEG-3, while CGH-1 is dynamic but does not consistently exhibit liquid-like characteristics and may be an intermediate phase within RNP granules. These distinct phases of RNA binding proteins correspond to, and may underlie, differential responses to stress. Interestingly, in oocyte RNP granules MEG-3 is not required for the condensation of PGL-1 or other RNA binding proteins, which differs from the role of MEG-3 in small, embryonic germ granules. Lastly, we show the PUF-5 translational repressor appears to promote MEX-3 and MEG-3 condensation into large RNP granules; however, this role may be associated with regulation of oogenesis.
PH Domain-Mediated Autoinhibition and Oncogenic Activation of Akt
Authors: Bae Hwan , Viennet Thibault , Park Eunyoung , Chu Nam , Salguero Antonieta L. , Eck Michael J. , Arthanari Haribabu , Cole Philip A.
Published: May 2022

Previously, we reported that the conformation of autoinhibited Akt, a Ser/Thr protein kinase that plays a central role in metabolism and cancer, may be shifted by small molecule allosteric inhibitors limiting the mechanistic insights from existing X-ray structures that have relied on such compounds (Chu, Viennet, et al, 2020). Here we discover unexpectedly that a single mutation, R86A, in Akt’s PH domain exhibits intensified autoinhibitory features with enhanced PH domain-kinase domain affinity. Structural and biochemical analysis uncovers the importance of a key interaction network involving Arg86, Glu17, and Tyr18 that controls Akt conformation and activity. Our studies also shed light on the molecular basis for E17K Akt activation as an oncogenic driver.
The leucine-rich repeat (LRR) domain of NLRP3 is required for NLRP3 inflammasome activation in macrophages
Authors: Duan Yanhui , Wang Jihong , Cai Juan , Kelley Nathan , He Yuan
Published: May 2022

The NLRP3 inflammasome is a critical component of innate immunity that defends the host from microbial infections. However, its aberrant activation contributes to the pathogenesis of several inflammatory diseases. Activation of the NLRP3 inflammasome induces the secretion of proinflammatory cytokines IL-1β and IL-18, and pyroptotic cell death. NLRP3 contains a leucine-rich repeat (LRR) domain at its C-terminus. Although posttranslational modifications in this LRR domain have been shown to regulate NLRP3 inflammasome activation, the role of the entire LRR domain in NLRP3 inflammasome activation remains controversial. Here, we generated mouse macrophages that express an endogenous NLRP3 mutant lacking the LRR domain. Deletion of the LRR domain destabilized endogenous NLRP3 protein and abolished NLRP3 inflammasome activation in macrophages. Furthermore, using NLRP3-deficient macrophages that are reconstituted with NLRP3 mutants lacking the LRR domain, we found that deletion of the LRR domain inhibited NLRP3 inflammasome activation. Mechanistically, deletion of the LRR domain abolished NLRP3 self-association, oligomerization, and interaction with the essential regulator NEK7. Our results demonstrate a critical role for the LRR domain in NLRP3 inflammasome activation.
Coordination of CcpA and CodY regulators in Staphylococcus aureus USA300 strains
Authors: Poudel Saugat , Hefner Ying , Szubin Richard , Sastry Anand , Gao Ye , Nizet Victor , Palsson Bernhard O.
Published: May 2022

The complex crosstalk between metabolism and gene regulatory networks makes it difficult to untangle individual constituents and study their precise roles and interactions. To address this issue, we modularized the transcriptional regulatory network (TRN) of the StaphylococcusaureusUSA300 strain by applying Independent Component Analysis (ICA) to 385 RNA sequencing samples. We then combined the modular TRN model with a metabolic model to study the regulation of carbon and amino acid metabolism. Our analysis showed that regulation of central carbon metabolism by CcpA and amino acid biosynthesis by CodY are closely coordinated. In general,S. aureusincreases the expression of CodY-regulated genes in the presence of preferred carbons sources such as glucose. This transcriptional coordination was corroborated by metabolic model simulations that also showed increased amino acid biosynthesis in the presence of glucose. Further, we found that CodY and CcpA cooperatively regulate the expression of ribosome hibernation promoting factor, thus linking metabolic cues with translation. In line with this hypothesis, expression of CodY-regulated genes is tightly correlated with expression of genes encoding ribosomal proteins. Together, we propose a coarse-grained model where expression ofS. aureusgenes encoding enzymes that control carbon flux and nitrogen flux through the system is coregulated with expression of translation machinery to modularly control protein synthesis. While this work focuses on three key regulators, the full TRN model we present contains 76 total independently modulated sets of genes, each with the potential to uncover other complex regulatory structures and interactions.

Importance

Staphylococcus aureusis a versatile pathogen with an expanding antibiotic resistance profile. The biology underlying its clinical success emerges from an interplay of many systems such as metabolism and gene regulatory networks. This work brings together models for these two systems to establish fundamental principles governing the regulation of S. aureus central metabolism and protein synthesis. Studies of these fundamental biological principles are often confined to model organisms such asEscherichia coli. However, expanding these models to pathogens can provide a framework from which complex and clinically important phenotypes such as virulence and antibiotic resistance can be better understood. Additionally, the expanded gene regulatory network model presented herein can deconvolute the biology underlying other important phenotypes in this pathogen.
Bridging the gap between bioinformatics and the clinical and public health microbiology laboratory: An ISO-accredited genomics workflow for antimicrobial resistance
Authors: Sherry Norelle L. , Horan Kristy , Ballard Susan A. , da Silva Anders Gonçalves , Gorrie Claire L. , Schultz Mark B. , Stevens Kerrie , Valcanis Mary , Sait Michelle , Stinear Timothy P. , Howden Benjamin P. , Seemann Torsten
Published: May 2022

Realising the promise of genomics to revolutionise routine AMR diagnosis and surveillance has been a long-standing challenge in clinical and public health microbiology. We have directly addressed this issue by creating and validating abritAMR, an ISO-accredited bioinformatics platform for genomics-based bacterial AMR gene detection. abritAMR utilises the NCBI AMRFinderPlus for detection of AMR genes and mutations, with additional features to classify AMR determinants into an antibiotic class. We validated abritAMR by comparing with multiplex PCR or gold-standard reference genomes, together representing 1500 different bacteria across 29 genera and covering 415 antibiotic resistance alleles. We also assessed inference of phenotypic resistance by comparing genomic predictions with agar dilution results for 864 Salmonella spp. Performance of abritAMR was excellent, detecting AMR genes with 99.9% accuracy (95% CI 99.9-99.9%), 97.9% sensitivity (95% CI 97.5-98.4%) and 100% specificity (100-100%). Phenotypic inference of resistance for Salmonella spp. was equally impressive, with 98.9% accuracy (98.7-99.1%). Validation data were submitted to the governing authority and ISO15189 accreditation was achieved. Implementation of abritAMR resulted in streamlined bioinformatics and reporting pathways, and it was readily updated and re-verified with database revisions or changes in reporting requirements. abritAMR is publicly and freely available to assist clinical and public health microbiology laboratories everywhere harness the power of AMR genomics in their professional practice.
peaks2utr: a robust Python tool for the annotation of 3’ UTRs
Authors: Haese-Hill William , Crouch Kathryn , Otto Thomas D.
Published: May 2022

Annotation of non-model organisms is an open problem, especially the detection of untranslated regions (UTRs). Correct annotation of UTRs is crucial in transcriptomic analysis to accurately capture the expression of each gene yet is mostly overlooked in annotation pipelines. Here we present peaks2utr, an easy-to-use Python command line tool that uses the UTR enrichment of single-cell technologies, such as 10x Chromium, to accurately annotate 3’ UTRs for a given canonical annotation.

Availability and Implementation

peaks2utr is implemented in Python 3 (≥ 3.8). It is available via PyPI at https://pypi.org/project/peaks2utr and GitHub at https://github.com/haessar/peaks2utr. It is licensed under GNU GPLv3.
Optimum time for hand pollination in yam (Dioscorea spp.)
Authors: Mondo Jean M. , Agre Paterne A. , Asiedu Robert , Akoroda Malachy O. , Asfaw Asrat
Published: May 2022

Hand pollination success rate is low in yam (Dioscorea spp.), due partly to suboptimal weather conditions. Thus, determining the most suitable time for pollination could improve the pollination success in yam breeding programs. We performed continuous hand pollination within flowering windows of D. rotundata and D. alata for two consecutive years to determine the most appropriate month, week, and hours of the day allowing maximum pollination success. In D. alata crossing block, we observed significant differences among crossing hours for pollination success (p = 0.003); morning hours (8–12 a.m.) being more conducive than afternoons (12–5 p.m.). No significant differences existed between crossing hours in D. rotundata, though the mid-day seemed optimal. For both species, the time interval 11–12 a.m. was more appropriate for crossing while 4–5 p.m. was the poorest. However, in vitro pollen germination tests showed that mid-day pollen collection (12 noon – 2 p.m.) had better results than both extremes, though there were strong genotypic effects on outcomes. Pollination success rates differed significantly among months for D. alata (p < 0.001) but not for D. rotundata (p > 0.05). Differences in pollination success existed across weeks within flowering windows of both D. alata (p < 0.001) and D. rotundata (p = 0.004). The seed production efficiency (SPE) had a similar trend as the pollination success rate. No clear pattern existed between the pollination time and the seed setting rate (SSR) or seed viability (SV), though their dynamics varied with weeks and months. This study provided an insight on the dynamics of pollination outcomes under the influence of pollination times and allows detecting months, weeks, and hours of the day when hybridization activities should be focused for better results.
Reverse-engineering method for XPCS studies of non-equilibrium dynamics
Authors: Ragulskaya Anastasia , Starostin Vladimir , Begam Nafisa , Girelli Anita , Rahmann Hendrik , Reiser Mario , Westermeier Fabian , Sprung Michael , Zhang Fajun , Gutt Christian , Schreiber Frank
Published: May 2022

X-ray photon correlation spectroscopy (XPCS) is a powerful tool in the investigation of dynamics covering a broad time and length scale. It has been widely used to probe dynamics for systems in both equilibrium and non-equilibrium states; in particular, for systems undergoing a phase transition where the structural growth kinetics and the microscopic dynamics are strongly intertwined. The resulting time-dependent dynamic behavior can be described using the two-time correlation function (TTC), which, however, often contains more interesting features than the component along the diagonal, and cannot be easily interpreted via the classical simulation methods. Here, a reverse engineering (RE) approach is proposed based on particle-based heuristic simulations. This approach is applied to an XPCS measurement on a protein solution undergoing a liquid–liquid phase separation. It is demonstrated that the rich features of experimental TTCs can be well connected with the key control parameters including size distribution, concentration, viscosity and mobility of domains. The dynamic information obtained from this RE analysis goes beyond the existing theory. The RE approach established in this work is applicable for other processes such as film growth, coarsening or evolving systems.
Vineyard or olive garden: an investment decision
Authors: Riaz Yasir , Abdullah Iqra
Published: May 2022

Learning outcomes

The learning outcomes are as follows:• demonstrate an understanding of the fruit farming process;• compare the pros and cons of farming different fruits;• understand the general characteristics of entrepreneurs and identify the issues they face;• compare the risk and returns from two types of investments;• apply capital budgeting techniques to ascertain the best available investment option; and• perform sensitivity analysis based on different anticipated situations for a new business.

Case overview/synopsis

Since his birth, Mr Bashir Khan, a 45-year-old father of four, lived as a farmer in Kallar Kahar, Pakistan. He owned 15 acres of land which he used to cultivate wheat and millet. He decided to start fruit farming after harvesting wheat in April 2021 to satisfy his long-standing desire to own a garden. He recently met a friend who was earning well from grape farming, who suggested that Khan set up a vineyard farm which could become a profitable venture for him. At the same time, Khan learned that the government had declared the Potohar region of Pakistan an olive valley, and was giving massive subsidies on olive cultivation. Khan now had a choice of fruits to plant on his land. One of his relatives, Omar Khayam, was an accountant at a firm. Khayam offered to conduct a feasibility analysis for Khan, to provide the relevant data, and help Khan select a high profit-yielding fruit farm.

Complexity academic level

Introductory finance courses at the undergraduate and postgraduate levels as well as executive training courses focused on the agri-finance discipline.

Supplementary materials

Teaching notes are available for educators only.

Subject code

CSS 1: Accounting and Finance.