bluuebunny/crossref_metadata_2025 huggingface dataset:
A screen of alkaline and oxidative formulations for their inactivation efficacy of metal surface adsorbed prions using a steel-bead seed amplification assay
Authors: Heinzer Daniel , Avar Merve , Pfammatter Manuela , Moos Rita , Schwarz Petra , Buhmann Matthias T. , Kuhn Benjamin , Mauerhofer Stefan , Rosenberg Urs , Aguzzi Adriano , Hornemann Simone
Published: June 2023

Iatrogenic transmission of prions, the infectious agents of fatal Creutzfeldt-Jakob disease, through inefficiently decontaminated medical instruments remains a critical issue. Harsh chemical treatments are effective, but not suited for routine reprocessing of reusable surgical instruments in medical cleaning and disinfection processes due to material incompatibilities. The identification of mild detergents with activity against prions is therefore of high interest but laborious due to the low throughput of traditional assays measuring prion infectivity. Here, we report the development of TESSA (sTainlESs steel-beadSeedAmplification assay), a prion seed amplification assay that explores the propagation activity of prions with stainless steel beads. TESSA was applied for the screening of about 70 different commercially available and novel formulations and conditions for their prion inactivation efficacy. One hypochlorite-based formulation, two commercially available alkaline formulations and a manual alkaline pre-cleaner were found to be highly effective in inactivating prions under conditions simulating automated washer-disinfector cleaning processes. The efficacy of these formulations was confirmedin vivoin a murine prion infectivity bioassay, yielding a reduction of the prion titer for the bead surface adsorbed prions below detectability. Our data suggest that TESSA represents an effective method for a rapid screening of prion-inactivating detergents, and that alkaline and oxidative formulations are promising in reducing the risk of potential iatrogenic prion transmission through insufficiently decontaminated instrument surfaces.
SynBot: An open-source image analysis software for automated quantification of synapses
Authors: Savage Justin Tyler , Ramirez Juan , Risher W. Christopher , Wang Yizhi , Irala Dolores , Eroglu Cagla
Published: June 2023

The formation of precise numbers of neuronal connections, known as synapses, is crucial for brain function. Therefore, synaptogenesis mechanisms have been one of the main focuses of neuroscience. Immunohistochemistry is a common tool for visualizing synapses. Thus, quantifying the numbers of synapses from light microscopy images enables screening the impacts of experimental manipulations on synapse development. Despite its utility, this approach is paired with low throughput analysis methods that are challenging to learn and results are variable between experimenters, especially when analyzing noisy images of brain tissue. We developed an open-source ImageJ-based software, SynBot, to address these technical bottlenecks by automating the analysis. SynBot incorporates the advanced algorithms ilastik and SynQuant for accurate thresholding for synaptic puncta identification, and the code can easily be modified by users. The use of this software will allow for rapid and reproducible screening of synaptic phenotypes in healthy and diseased nervous systems.
Vertical transmission ofBorrelia turicatae(Spirochaetales:Borreliaceae) by autogenously reproducingOrnithodoros turicata(Ixodida: Argasidae) female naturally infected with the spirochetes
Authors: Filatov Serhii , Krishnavajhala Aparna , Lopez Job E.
Published: June 2023

Ornithodoros turicatais a vector of relapsing fever (RF) spirochetes in North America and transmitsBorrelia turicataeto a variety of vertebrate hosts. The remarkably long lifespan ofO. turicataand its ability to maintain spirochetes horizontally (between life stages) and vertically to progeny promotes the perpetuation ofB. turicataein nature. Nevertheless, the reproductive biology ofO. turicatais poorly understood. In this report, we collected ticks from a park within a neighborhood of Austin, Texas. They were reared to adulthood and male ticks were individually housed with females. We observed autogenous reproduction by the ticks and further investigated vertical transmission ofB. turicataeby quantifying filial infection rates in a cohort of progeny ticks. These results indicate thatO. turicatatransovarially transmitsB. turicataeduring autogenous reproduction and further signify the tick as a natural reservoir of the spirochetes.

Importance

Previous research has implicatedOrnithodorosticks, includingOrnithodoros turicata, as long-term reservoirs of relapsing fever (RF) spirochetes. Considering the tick’s long lifespan and their efficiency in maintaining and transferring spirochetes within the population, the infection could persist in a given enzootic focus for decades. However, little is known about the relative importance of horizontal and vertical transmission routes in the persistence and evolution of RFBorrelia. Our observations on the reproductive biology ofO. turicatain the absence of vertebrate hosts indicate an additional mechanism by whichB. turicatacan be maintained in the environment. This work establishes the foundation for studyingO. turicatareproduction and spirochete-vector interactions, which will aid in devising control measures forOrnithodorosticks and RF spirochetes.
Recombinant outer membrane vesicles coupled with cytokines act as high-performance adjuvants againstHelicobacter pyloriinfection in mice
Authors: Liu Qiong , Li Biaoxian , Lu Jiahui , Zhang Yejia , Shang Yinpan , Li Yi , Gong Tian , Zhang Chengsheng
Published: June 2023

The widespread prevalence ofHelicobacter pylori(H. pylori) infection remains a great challenge to the human health. The existing vaccines are not ideal for preventingH. pyloriinfection; thus, exploring highly effective adjuvants may improve the immunoprotective efficacy ofH. pylorivaccines. In a previous study, we found that outer membrane vesicles (OMVs), a type of nanoscale particle spontaneously produced by Gram-negative bacteria, could act as adjuvants to boost the immune response to vaccine antigens. In the present study, we explored the potential application of OMVs as delivery vectors for adjuvant development. We constructed recombinant OMVs containing cytokines interleukin 17A or interferon-γ and evaluated their function as adjuvants in combination with inactivated whole-cell vaccine (WCV) or UreB as vaccine antigens. Our results showed that recombinant OMVs as adjuvants could induce stronger humoral and mucosal immune responses in mice than wild-typeH. pyloriOMVs and the cholera toxin (CT) adjuvant. Additionally, the recombinant OMVs significantly promoted Th1/Th2/Th17-type immune responses. Furthermore, the recombinant OMV adjuvant induced more potent clearance ofH. pylorithan CT and wild-type OMVs. Our data suggest that the recombinant OMVs coupled with cytokines may become potent adjuvants for development of novel and effective vaccines againstH. pyloriinfection.

Importance

Helicobacter pylori(H. pylori) is one of the important risk factors for gastric cancer, and its vaccine is crucial for its prevention and control. However, to date, no effective vaccine has been approved. Exploring novel and effective vaccine adjuvants may provide new perspectives and ideas for the development ofH. pylorivaccines. Outer membrane vesicles (OMVs) deserve more attention as a novel form of vaccine and adjuvant. We have long focused on OMVs as vaccine adjuvants to enhance efficacy by delivering eukaryotic plasmid expressing immune promoting cytokines in wild-type OMVs. Our results are expected to provide a new adjuvant form for the development ofH. pylorivaccine, and this adjuvant design strategy can also be used in the development of vaccines for other types of pathogens, including bacteria and even viruses.
Cooperative control of a DNA origami force sensor
Authors: Robbins Ariel , Hildebolt Hazen , Neuhoff Michael , Beshay Peter , Winter Jessica O. , Castro Carlos E. , Bundschuh Ralf , Poirier Michael G.
Published: June 2023

Most biomolecular systems are dependent on a complex interplay of forces. Modern force spectroscopy techniques provide means of interrogating these forces. These techniques, however, are not optimized for studies in constrained or crowded environments as they typically require micron-scale beads in the case of magnetic or optical tweezers, or direct attachment to a cantilever in the case of atomic force microscopy. We implement a nanoscale force-sensing device using a DNA origami which is highly customizable in geometry, functionalization, and mechanical properties. The device, referred to as the NanoDyn, functions as a binary (open or closed) force sensor that undergoes a structural transition under an external force. The transition force is tuned with minor alterations of 1 to 3 DNA oligonucleotides and spans tens of picoNewtons (pN). This actuation of the NanoDyn is reversible and the design parameters strongly influence the efficiency of resetting the initial state, with higher stability devices (≳10 pN) resetting more reliably during repeated force-loading cycles. Finally, we show that the opening force can be adjusted in real time by the addition of a single DNA oligonucleotide. These results establish the NanoDyn as a versatile force sensor and provide fundamental insights into how design parameters modulate mechanical and dynamic properties.
Text dialogue analysis Based ChatGPT for Primary Screening of Mild Cognitive Impairment
Authors: Wang Changyu , Liu Siru , Li Aiqing , Liu Jialin
Published: June 2023

Background

AI models tailored to diagnose cognitive impairment have shown excellent results. However, it is unclear whether large linguistic models can rival specialized models by text alone.

Objectives

We would explore the effectiveness of ChatGPT for primary screening of mild cognitive impairment (MCI) and standardize the design steps and components of the prompt.

Methods

We obtained 174 participants from the DementiaBank screening and classified 70% of them into the training set and 30% into the test set. Three dimensions of variables were incorporated, including: vocabulary, syntax and grammar, and semantics. These variables were generated from published studies and statistical analyses. We used R 4.3.0. for the analysis of variables and diagnostic indicators.

Results

The final variables included by published studies included: word frequency and word ratio, phrase frequency and phrase ratio, lexical complexity, syntactic complexity, grammatical components, semantic density, and semantic coherence; variables included in the analysis included: tip-of-the-tongue phenomenon (P < 0.001), difficulty with complex ideas (P < 0.001), and memory issues (P < 0.001). The final GPT4 model achieved the sensitivity (SEN) of 0.8636, specificity (SPE) of 0.9487 and area under the curve (AUC) of 0.9062 on the training set; on the test set, the SEN, SPE and AUC reached 0.7727, 0.8333 and 0.8030, respectively. The prompt consisted of five main parts, including character setting, scoring system setting, indicator setting, output setting, and explanatory information setting.

Conclusion

ChatGPT was effective in primary screening of participants with possible MCI. Improved standardization of prompts by professional clinicians would also improve the performance of the model. It is important to note that ChatGPT is not a substitute for a clinician making a diagnosis.
mRNA-1273 vaccine effectiveness against symptomatic SARS-CoV-2 infection and COVID-19-related hospitalization in children aged 6 months to 5 years
Authors: Aglipay Mary , Maguire Jonathon , Swayze Sarah , Tuite Ashleigh , Mamdani Muhammad , Keown-Stoneman Charles , Birken Catherine , Kwong Jeffrey C
Published: June 2023

Importance

Data on mRNA-1273 (Moderna) vaccine effectiveness in children aged 6 months to 5 years are limited.

Objective

To assess mRNA-1273 vaccine effectiveness against symptomatic SARS-CoV-2 infection and COVID-19-related hospitalization among children aged 6 months to 5 years.

Design, Setting, and Participants

A test-negative study using linked health administrative data in Ontario, Canada. Participants included symptomatic children aged 6 months to 5 years who were tested by RT-PCR.

Exposures

mRNA-1273 vaccination.

Main Outcomes and Measures

Symptomatic SARS-CoV-2 infection and COVID-19-related hospitalization.

Results

We included 3467 test-negative controls and 572 test-positive cases. Receipt of mRNA-1273 was associated with reduced symptomatic SARS-CoV-2 infection (VE=90%; 95%CI: 53, 99%) and COVID-19-related hospitalization (VE=82%; 95%CI: 4, 99%) ≥7 days after the second dose.

Conclusions and Relevance

Our findings suggest mRNA-1273 vaccine effectiveness is initially strong against symptomatic SARS-CoV-2 infection and hospitalization in children aged 6 months to 5 years. Further research is needed to understand long-term effectiveness and the need for booster doses.
How does corruption influence health system efficiency? A case study of two counties in Kenya
Authors: Munywoki Joshua , Nyawira Lizah , Musiega Anita , Njuguna Rebecca G , Tsofa Benjamin , Hanson Kara , Molyneux Sassy , Normand Charles , Jemutai Julie , Barasa Edwine
Published: June 2023

Objective

Efficiency gains are one potential pathway to unlocking additional resources for the health sector. Given that corruption has been cited as a key cause of inefficiency in the health sector, the objective of this study was to examine the influence of corruption on the efficiency of county health systems in Kenya.

Design, setting and analysis

We conducted a qualitative case study in two counties in Kenya. We developed a conceptual framework from a literature review to guide the development of tools and analysis. We collected qualitative data through in-depth interviews (n=26) with county, sub-county, and health facility level respondents across the two counties. We analyzed the data collected using a framework approach.

Results

Corrupt practices reported in the case study counties included non-merit-based recruitment and training of health workers, supply of substandard goods, equipment and infrastructure; theft, embezzlement and misuse of public funds and property; and informal payments. These practices were perceived to impact negatively on health system efficiency by leading to a direct loss of health sector resources, increase in operational costs, poor quality of care, reduced staff motivation and productivity, and reduced access to healthcare services.

Conclusion

The efficiency of county health systems could be enhanced by implementing anti-corruption strategies to tackle the identified corrupt practices.

Strengths and limitations of this study

To the best of our knowledge, this paper is the first of its kind to focus on how corruption affects attainment of health sector goals in Kenya

Our reported findings focus on only two out of forty-seven counties in Kenya.

Transferability of our findings ought to be interpreted with consideration to contextual factors that shaped reported corruption practices

We found study respondents to be cautious when reporting their experiences and perceptions on health sector corruption. It is possible that social desirability bias affected response from our study participants

Given the sensitivity and difficulty of corruption as a study topic, this study was not able to evaluate the effectiveness of existing anti-corruption strategies in Kenya
The effect of natural terrestrial gamma radiation emissions and environmental radon levels on the incidence of cutaneous squamous cell carcinoma of the head and neck
Authors: Couto-González Ivan , Arenaz-Búa Jorge , Taboda-Suárez Antonio , García-García Abel
Published: June 2023

The effects of ionising radiation on the development of cutaneous squamous cell carcinoma have been previously studied in environments in which the levels of ionising radiation have been increased by artificial sources. The purpose of this study is to determine the role that environmental radon concentration and natural gamma radiation emissions may play in the development of head and neck cutaneous squamous cell carcinoma in a geographical area which is known to have high levels of radon and natural terrestrial gamma radiation emissions. A total of 284 patients diagnosed with cutaneous squamous cell carcinoma during the 26-month observation period were included in the study. The overall incidence was 37.33 cases/100,000 people-year. The mean of environmental radon concentration according to their council of residence was 116.69 Bq/m3(40.05) and the mean of natural terrestrial gamma radiation emitted according to their council of residence was 14.25 μRad/hour (3.86). The multiple linear regression analysis revealed that only mean natural terrestrial gamma radiation emissions in the council of residence (P < 0.05), carrying out an outdoor profession (P < 0.05) and the mean number of hours of sunlight per year in the council of residence (P = 0.03) were found to have statistical significance on the incidence of head and neck cutaneous squamous cell carcinoma. Emissions of natural terrestrial gamma radiation have never been proposed as a factor having an influence on the development of cutaneous neoplasms. The implications that this could have in areas with naturally high terrestrial gamma radiation emissions should be more exhaustively studied to assess the true weight of this factor.
Chronic Overlapping Pain Conditions and Nociplastic Pain
Authors: Johnston Keira J.A. , Signer Rebecca , Huckins Laura M.
Published: June 2023

Chronic Overlapping Pain Conditions (COPCs) are a subset of chronic pain conditions commonly comorbid with one another and more prevalent in women and assigned female at birth (AFAB) individuals. Pain experience in these conditions may better fit with a new mechanistic pain descriptor, nociplastic pain, and nociplastic type pain may represent a shared underlying factor among COPCs. We applied GenomicSEM common-factor genome wide association study (GWAS) and multivariate transcriptome-wide association (TWAS) analyses to existing GWAS output for six COPCs in order to find genetic variation associated with nociplastic type pain, followed by genetic correlation (linkage-disequilibrium score regression), gene-set and tissue enrichment analyses. We found 24 independent single nucleotide polymorphisms (SNPs), and 127 unique genes significantly associated with nociplastic type pain, and showed nociplastic type pain to be a polygenic trait with significant SNP-heritability. We found significant genetic overlap between multisite chronic pain and nociplastic type pain, and to a smaller extent with rheumatoid arthritis and a neuropathic pain phenotype. Tissue enrichment analyses highlighted cardiac and thyroid tissue, and gene set enrichment analyses emphasized potential shared mechanisms in cognitive, personality, and metabolic traits and nociplastic type pain along with distinct pathology in migraine and headache. We use a well-powered network approach to investigate nociplastic type pain using existing COPC GWAS output, and show nociplastic type pain to be a complex, heritable trait, in addition to contributing to understanding of potential mechanisms in development of nociplastic pain.
Effects of Chronic Pain Diagnoses on the Antidepressant Efficacy of Transcranial Magnetic Stimulation
Authors: Totonchi Afshar H. , Fishbein J. N. , Martinez E. J. , Chu G. M. , Shenasa M. A. , Ramanathan D. , Herbert M. S.
Published: June 2023

Background

Major depressive disorder (MDD) and chronic pain are highly comorbid and bidirectionally related, such that MDD typically interferes with chronic pain treatment and vice versa. Repetitive transcranial magnetic stimulation (rTMS) over the dorsolateral prefrontal cortex (DLPFC) is effective in treating MDD, but additional research is needed to determine if chronic pain interferes with rTMS for MDD.

Method

Participants were 124 veterans (Mage=49.1, SD=13.8) scheduled for 30 sessions of rTMS across six weeks at the Veterans Affairs San Diego Healthcare System. Depression severity was monitored weekly using the Patient Health Questionnaire-9. Having any pain diagnosis, low back pain, or migraine/headache were assessed by chart review. Latent basis models were used to estimate change and change-by-pain diagnosis in depression scores during rTMS treatment.

Results

A total of 92 participants (74%) had a documented pain diagnosis, 58 (47%) had low back pain, and 32 (26%) had migraine/headache. Depression scores initially decreased (linear slope estimate=-2.04, SE=0.26,p<.0001), but the rate of decrease slowed over time (quadratic slope estimate=0.18, SE=0.04,p<.001). Having any pain diagnosis, low back pain, or migraine/headache did not significantly differentiate overall amount of change. However, individuals with headache/migraine showed greater initial improvement but then an even faster slowing in rate of decrease than those without headache/migraine.

Conclusions

Having any pain diagnosis, low back pain, or headache/migraine did not significantly interfere with improvement in depression; however, headache/migraine affected the timing of change. These data contribute to the ongoing support of rTMS as a viable treatment option for comorbid populations.
LSD increases sleep duration the night after microdosing
Authors: Allen Nathan , Jeremiah Aron , Murphy Robin , Sumner Rachael , Forsyth Anna , Hoeh Nicholas , Menkes David B , Evans William , Muthukumaraswamy Suresh , Sundram Frederick , Roop Partha
Published: June 2023

Microdosing psychedelic drugs, at a level below the threshold to induce hallucinations, is an increasingly common lifestyle practise. However, the effects of microdosing on sleep have not been previously reported. Here we report results from a Phase 1 randomised controlled trial in which 80 healthy adult male volunteers received a six week course of either LSD (10 µg) or placebo with doses self-administered every third day. Participants used a commercially available sleep/activity tracker for the duration of the trial. Data from 3231 nights of sleep showed that on the night after microdosing participants in the LSD group slept an extra 24.3 minutes per night (95% Confidence Interval 10.3 - 38.3 minutes) compared to placebo - with no reductions of sleep observed on the dosing day itself. There were no changes in the proportion of time spent in various sleep stages or in participant physical activity. These results show clear modification of the physiological sleep requirements in healthy volunteers who microdose, and may have implications for the proposed therapeutic effects of microdosing in mood disorders such as major depressive disorder where sleep is frequently disturbed. The clear, clinically significant, changes in objective measurements of sleep observed are difficult to explain as a placebo effect.
Application of Deep Learning Models into the Prediction of Interleukin-6 and -8 Cytokines in Sickle Cell Anemia Patients
Authors: Nguyen Dylan , Abraham Lukas , Amatya Ashesh
Published: June 2023

Interleukin-6 (IL6) and Interleukin-8 (IL8) are cytokines related to general immune function, but within Sickle Cell Anemia (SCA) patients, their overproduction tends to cause auto-immune reactions. These vital cytokines engage in the pathophysiology of SCA, but the extent to which they are associated with the genetics of the disease requires further exploration. This research paper seeks to further the study of IL6 and IL8 in SCA patients as well as the possibilities to predict their presence in patients based on Haptoglobin alleles and various other hematological factors using artificial neural networks. This was done through a cross-sectional study of 60 sickle anemia patients and 74 healthy individuals who provided the basis for the data of this study. The deep learning model found a non-linear correlation between the Haptoglobin alleles and the production of IL6 and IL8, predicting their over presence in SCA patients with an accuracy of 90.9% and r-squared value of 0.88 based on the given inputs. The machine learning models built in this paper have the potential to accelerate the development of targeted treatments and diagnoses to those suffering from Sickle Cell Anemia and its specific immune complications.
Population encoding of stimulus features along the visual hierarchy
Authors: Dyballa Luciano , Rudzite Andra M. , Hoseini Mahmood S. , Thapa Mishek , Stryker Michael P. , Field Greg D. , Zucker Steven W.
Published: June 2023

The retina and primary visual cortex (V1) both exhibit diverse neural populations sensitive to diverse visual features. Yet it remains unclear how neural populations in each area partition stimulus space to span these features. One possibility is that neural populations are organized into discrete groups of neurons, with each group signaling a particular constellation of features. Alternatively, neurons could be continuously distributed across feature-encoding space. To distinguish these possibilities, we presented a battery of visual stimuli to mouse retina and V1 while measuring neural responses with multi-electrode arrays. Using machine learning approaches, we developed a manifold embedding technique that captures how neural populations partition feature space and how visual responses correlate with physiological and anatomical properties of individual neurons. We show that retinal populations discretely encode features, while V1 populations provide a more continuous representation. Applying the same analysis approach to convolutional neural networks that model visual processing, we demonstrate that they partition features much more similarly to the retina, indicating they are more like big retinas than little brains.
Label-free analysis of bacterial growth and lysis at the single-cell level using droplet microfluidics and object detection-oriented deep learning
Authors: Tiwari Anuj , Nikolic Nela , Anagnostidis Vasileios , Gielen Fabrice
Published: June 2023

Bacteria identification and counting at the small population scale is important to many applications in the food safety industry, the diagnostics of infectious diseases and the study and discovery of novel antimicrobial compounds. There is still a lack of easy to implement, fast and accurate methods to count populations of motile cells at the single-cell level. Here, we report a label-free method to count and localize bacterial cells freely swimming in microfluidic anchored picolitre droplets. We used the object detection oriented YOLOv4 deep learning framework for cell detection from bright-field images obtained with an automated Z-stack setup. The neural network was trained to recognizeEscherichia colicell morphology with an average precision of approximately 84%. This allowed us to accurately identify individual cell division events, enabling the study of stochastic bacterial growth starting from initial populations as low as one cell. This work also demonstrates the ability to study single cell lysis in the presence of T7 lytic bacterial viruses (phages). The high precision in cell numbers facilitated the visualization of bacteria-phage interactions over timescale of hours, paving the way towards deciphering phage life cycles in confined environments.
Microglia produce the amyloidogenic ABri peptide in familial British dementia
Authors: Arber Charles , Casey Jackie M. , Crawford Samuel , Rambarack Naiomi , Yaman Umran , Wiethoff Sarah , Augustin Emma , Piers Thomas M. , Rostagno Agueda , Ghiso Jorge , Lewis Patrick A. , Revesz Tamas , Hardy John , Pocock Jennifer M. , Houlden Henry , Schott Jonathan M. , Salih Dervis A. , Lashley Tammaryn , Wray Selina
Published: June 2023

Mutations inITM2Bcause familial British, Danish, Chinese and Korean dementias. In familial British dementia (FBD) a mutation in the stop codon of theITM2Bgene (also known asBRI2) causes a C-terminal cleavage fragment of the ITM2B/BRI2 protein to be extended by 11 amino acids. This fragment, termed amyloid-Bri (ABri), is highly insoluble and forms extracellular plaques in the brain. ABri plaques are accompanied by tau pathology, neuronal cell death and progressive dementia, with striking parallels to the aetiology and pathogenesis of Alzheimer’s disease. The molecular mechanisms underpinning FBD are ill-defined. Using patient-derived induced pluripotent stem cells, we show that expression ofITM2B/BRI2is 34-fold higher in microglia than neurons, and 15-fold higher in microglia compared with astrocytes. This cell-specific enrichment is supported by expression data from both mouse and human brain tissue. ITM2B/BRI2 protein levels are higher in iPSC-microglia compared with neurons and astrocytes. Consequently, the ABri peptide was detected in patient iPSC-derived microglial lysates and conditioned media but was undetectable in patient-derived neurons and control microglia. Pathological examination of post-mortem tissue support ABri expression in microglia that are in proximity to pre-amyloid deposits. Finally, gene co-expression analysis supports a role for ITM2B/BRI2 in disease-associated microglial responses. These data demonstrate that microglia are the major contributors to the production of amyloid forming peptides in FBD, potentially acting as instigators of neurodegeneration. Additionally, these data also suggest ITM2B/BRI2 may be part of a microglial response to disease, motivating further investigations of its role in microglial activation. This has implications for our understanding of the role of microglia and the innate immune response in the pathogenesis of FBD and other neurodegenerative dementias including Alzheimer’s disease.
Lymphocytic choriomeningitis arenavirus requires cellular COPI and AP-4 complexes for efficient replication and virion production
Authors: Byford Owen , Shaw Amelia B. , Tse Hiu Nam , Todd Eleanor J. A. A. , Álvarez-Rodríguez Beatriz , Hewson Roger , Fontana Juan , Barr John N.
Published: June 2023

Lymphocytic choriomeningitis virus (LCMV) is a bisegmented negative-sense RNA virus classified within theArenaviridaefamily of theBunyaviralesorder. LCMV is associated with fatal disease in immunocompromised populations, and as the prototypical arenavirus, acts as a model for the many serious human pathogens within theArenaviridaefamily. Here, we examined the dependence of LCMV multiplication on cellular trafficking components using a recombinant LCMV expressing enhanced green fluorescent protein in conjunction with a curated siRNA library. The screen revealed a requirement for subunits of both the coat protein 1 (COPI) coatamer and adapter protein 4 (AP-4) complexes. By rescuing a recombinant LCMV harbouring a FLAG tagged GP-1 envelope spike (rLCMV-GP1-FLAG) we showed infection resulted in marked co-localization of COPI and AP-4 component with both LCMV nucleoprotein (NP) and GP-1. Time-of-addition studies using brefeldin A (BFA), an ARF-I inhibitor that prevents formation of both COPI and AP-4 complexes, suggested these cellular components were involved in late stages of the LCMV multiplication cycle. Consistent with this finding, BFA treatment at similar late time-points resulted in a marked redistribution of NP and GP-1, and subsequent loss of COPI/AP-4 co-localization. Finally, titration of released virus within supernatant of BFA-treated cells revealed a 10-fold decrease in viral titres, greater than the 2-fold BFA-mediated reduction in NP expression. Taken together, these findings suggest COPI and AP-4 complexes are important host cell factors that are required for efficient LCMV assembly and egress.

Importance

Arenaviruses are rodent-borne, segmented, negative-sense RNA viruses, with several members responsible for fatal human disease, with the prototypic member LCMV being under-recognised as a pathogen capable of inflicting neurological infections with fatal outcome. Here, we assessed the impact of siRNA knockdown of host cell trafficking genes on LCMV multiplication. We reveal the requirement of host cellular COPI and AP-4 complexes for efficient LCMV multiplication, acting late in the replication cycle, at the stages of egress and assembly. Collectively, our findings improve the understanding of arenaviruses host-pathogen interactions and reveal novel cellular trafficking pathways required during infection. Moreover, this study may lead to the discovery of novel therapeutic targets for arenaviruses to prevent serious human disease.
Homing in on the rare virosphere reveals the native host of giant viruses
Authors: Fromm Amir , Hevroni Gur , Vincent Flora , Schatz Daniella , Martinez-Gutierrez Carolina A. , Aylward Frank O. , Vardi Assaf
Published: June 2023

Giant viruses (phylum Nucleocytoviricota) are globally distributed in aquatic ecosystems1,2. They play major roles as evolutionary drivers of eukaryotic plankton3and regulators of global biogeochemical cycles4. Recent metagenomic studies have significantly expanded the known diversity of marine giant viruses1,5–7, but we still lack fundamental knowledge about their native hosts, thereby hindering our understanding of their lifecycle and ecological importance. Here, we aim to discover the native hosts of giant viruses using a novel, sensitive single-cell metatranscriptomic approach. By applying this approach to natural plankton communities, we unraveled an active viral infection of several giant viruses, from multiple lineages, and identified their native hosts. We identify a rare lineage of giant virus (Imitervirales-07) infecting a minute population of protists (class Katablepharidaceae) and revealed the prevalence of highly expressed viral-encoded cell-fate regulation genes in infected cells. Further examination of this host-virus dynamics in a temporal resolution suggested this giant virus controls its host population demise. Our results demonstrate how single-cell metatranscriptomics is a sensitive approach for pairing viruses with their authentic hosts and studying their ecological significance in a culture-independent manner in the marine environment.
MORPHOGENIC VERSUS MITOGENIC ROLES OF SHH ARE SEGREGATED ON DISTINCT EXOSOMES REGULATED BY CELLULAR RAB7 LEVELS
Authors: Walvekar Ankita , Pandey Shivangi , Kamat Siddhesh S. , Damodar Sharat , Ladher Raj K. , Vyas Neha
Published: June 2023

The secreted signaling molecule, Sonic hedgehog (Shh) is involved in patterning and growth of various embryonic tissues across species. This lipid-anchored protein is secreted on extracellular vesicles to activate signaling. Shh has two functions during spinal cord development, acting as a morphogen to pattern the ventral neural tube and a mitogen to maintain neural progenitors. Here, we find that these activities are segregated on to two distinct pools of exosomes. Shh secreted on a classical exosomal pool (Shh-P150) is able to activate ventral threshold targets in the neural tube. In contrast, the mitogenic activity, is elicited by a lighter exosomal pool (Shh-P450). We further show that cellular environment plays a major role in regulating the biogenesis of P150 versus P450 by modulating Rab7 levels. We find that active Rab7 is necessary for ventral neural tube patterning through the regulation of Shh-P150 secretion. The cellular mechanisms involved in packaging and secretion of Shh with different partners on P150 or P450 pools may be more general, enabling the separation of functional activities of signaling molecules during development and disease.
Functional loss ofrffGandrfbB,encoding dTDP-glucose 4,6-dehydratase, changes colony morphology, cell shape, motility and virulence inSalmonellaTyphimurium
Authors: Chakravorty Subhashish , Banerjee Pip , Joseph Joel P. , Pathak Sanmoy , Verma Taru , Das Mrinmoy , Nandi Dipankar
Published: June 2023

Lipopolysaccharide (LPS) O-antigen and enterobacterial common antigen (ECA) play crucial roles in maintaining the structural integrity of the outer membrane in Gram-negative bacteria. Previous studies conducted with either LPS or ECA mutants have highlighted the importance of these cell surface polysaccharides in the physiology ofSalmonella entericaserovar Typhimurium (S. Typhimurium). However, the functional consequences resulting from the abrogation of both O-antigen and ECA synthesis inS. Typhimurium are not well studied. In the present study, we generated single and double gene-deleted mutants ofrffGandrfbB, which are paralogs, encoding dTDP-glucose 4,6-dehydratase that catalyze steps in the synthesis of both O-antigen and ECA. The functional loss of bothrffGandrfbB(ΔrffGΔrfbB), but not in single gene-deleted strains, results in a round cell morphology, smaller colony formation and altered LPS profile. In addition, the ΔrffGΔrfbBstrain displays defects in outer membrane permeability, causing hypersensitivity to bile and cell wall targeting antibiotics, e.g., meropenem and polymyxin B. Transcriptomic analysis identified flagellar and SPI-1 pathway to be highly down-regulated in the ΔrffGΔrfbBstrain which leads to impaired swimming and swarming motility and lower adhesion and invasion of HeLa cells. Importantly, the ΔrffGΔrfbBstrain is less proficient in colonizing Peyer’s patches, spleen and liver, is unable to induce pro-inflammatory cytokines and is attenuated in both the oral and intra-peritoneal models ofS. Typhimurium infection in mice. Overall, this study highlights the importance ofrffGandrfbBin maintaining cell wall integrity, colony and cellular morphology, motility and virulence inS. Typhimurium.
A novel role for the peptidyl-prolylcis-transisomerase Cyclophilin A in DNA-repair following replication fork stalling via the MRE11-RAD50-NBS1 complex
Authors: Bedir Marisa , Outwin Emily , Colnaghi Rita , Bassett Lydia , Abramowicz Iga , O’Driscoll Mark
Published: June 2023

We previously reported that non-homologous end-joining (NHEJ)-defective humanLIG4-/-pre-B lymphocytes were unexpectedly sensitive to killing by the cyclic peptide Cyclosporin A (CsA), a common component of bone marrow transplantation conditioning and maintenance regimes. We also found that CsA induced DNA double strand breaks (DSBs) in LIG4 syndrome patient fibroblasts, specifically upon transit through S-phase. The molecular basis underlying these CsA impacts has not been described hitherto. We postulated that CsA-induced genomic instability may reflect a direct role of Cyclophilin A (CYPA) in DNA repair, as CYPA is the primary physiological target interactor of CsA.

CYPA is the founding member of the Cyclophilin family of peptidyl-prolylcis-transisomerases (PPIs). CsA inhibits the PPI activity of CYPA through occupation of the latter’s enzymatic active site. Using an integrated approach involving CRISPR/Cas9-engineering, siRNA, BioID, co-immunoprecipitation, pathway-specific DNA repair investigations as well as protein expression-interaction analysis, we describe novel impacts of CYPA loss and inhibition of its PPI activity on DNA repair. Prompted by findings from our CYPA-BioID proximity interactome, we validate CYPA interactions with different components of the DNA end resection machinery. Moreover, we characterise a novel and direct CYPA interaction with the NBS1 component of the MRE11-RAD50-NBS1 (MRN) complex, providing evidence that the PPI function of CYPA actively influences DNA repair via direct protein-protein interaction at the level of DNA end resection. Consequently, we demonstrate that CYPA loss or inhibition impairs Homologous Recombination Repair (HRR) following DNA replication fork stalling.

Additionally, we define a set of genetic vulnerabilities associated with CYPA loss and inhibition, identifying DNA replication fork protection as an important determinant of viability herein. Leveraging the novel insights into CYPA biology we have uncovered; we explore examples of how CYPA PPI inhibition may be exploited to selectively kill cells from a variety of different cancers with a shared characteristic genomic instability profile. These findings propose a potential new disease application or repurposing strategy for the non-immunosuppressive CsA analogue class of Cyclophilin inhibitors.
Multiple pathways for glucose phosphate transport and utilization support growth ofCryptosporidium parvum
Authors: Xu Rui , Beatty Wandy L. , Greigert Valentin , Witola William H. , Sibley L. David
Published: June 2023

Cryptosporidium parvumis an obligate intracellular parasite with a highly reduced mitochondrion that lacks the TCA cycle and the ability to generate ATP, making the parasite reliant on glycolysis. Genetic ablation experiments demonstrated that neither of the two putative glucose transporters CpGT1 and CpGT2 were essential for growth. Surprisingly, hexokinase was also dispensable for parasite growth while the downstream enzyme aldolase was required, suggesting the parasite has an alternative way of obtaining phosphorylated hexose. Complementation studies inE. colisupport a role for direct transport of glucose-6-phosphate from the host cell by the parasite transporters CpGT1 and CpGT2, thus bypassing a requirement for hexokinase. Additionally, the parasite obtains phosphorylated glucose from amylopectin stores that are released by the action of the essential enzyme glycogen phosphorylase. Collectively, these findings reveal thatC. parvumrelies on multiple pathways to obtain phosphorylated glucose both for glycolysis and to restore carbohydrate reserves.
Glycoprofiling of proteins as prostate cancer biomarkers: a multinational population study
Authors: Pinkeova Andrea , Tomikova Adela , Bertokova Aniko , Fabinyova Eva , Bartova Radka , Jane Eduard , Hroncekova Stefania , Sievert Karl-Dietrich , Sokol Roman , Jirasko Michal , Kucera Radek , Eder Iris E. , Horninger Wolfgang , Klocker Helmut , Ďubjaková Petra , Fillo Juraj , Bertok Tomas , Tkac Jan
Published: June 2023

The glycoprofiling of two proteins, the free form of the prostate-specific antigen (fPSA) and zinc-α-2-glycoprotein (ZA2G), was assessed to determine their suitability as prostate cancer (PCa) biomarkers. The glycoprofiling of proteins was performed by analysing changes in the glycan composition on fPSA and ZA2G using lectins (proteins recognising glycans,i.e. complex carbohydrates). The specific glycoprofiling of the proteins was performed using magnetic beads (MBs) modified with horseradish peroxidase (HRP) and antibodies that selectively enriched fPSA or ZA2G from human serum samples. Subsequently, the antibody-captured glycoproteins were incubated on lectin-coated ELISA plates. In addition, a novel glycoprotein standard (GPS) was used to calibrate the assay. The glycoprofiling of fPSA and ZA2G was performed in human serum samples obtained from men undergoing prostate biopsy after an elevated serum PSA, and prostate cancer patients with or without prior therapy. The results are presented in the form of a ROC (Receiver Operating Curve). A DCA (Decision Curve Analysis) to evaluate the clinical performance and net benefit of fPSA glycan-based biomarkers was also performed. While the glycoprofiling of ZA2G showed little promise as a potential PCa biomarker, the glycoprofiling of fPSA would appear to have significant clinical potential. Hence, the GIA (Glycobiopsy ImmunoAssay) test integrates the glycoprofiling of fPSA (i.e. two glycan forms of fPSA). The GIA test could be used for early diagnoses of PCa (AUC=0.84; n=501 samples) with a potential for use in therapy-monitoring (AUC=0.85; n=168 samples). Moreover, the analysis of a subset of serum samples (n=215) revealed that the GIA test (AUC=0.81) outperformed the PHI (Prostate Health Index) test (AUC=0.69) in discriminating between men with prostate cancer and those with benign serum PSA elevation.
The Sde phosphoribosyl-linked ubiquitin transferases exploit reticulons to protect the integrity of theLegionella-containing vacuole
Authors: Zhang Mengyun , Kim Seongok , Isberg Ralph R.
Published: June 2023

Legionella pneumophilagrows within host cells by forming a specialized membrane-bound compartment via the Icm/Dot type IV secretion system (T4SS). T4SS translocated Sde proteins promote phosphoribosyl-linked ubiquitination (pR-Ub) of several mammalian cell proteins, including Rtn4. In response to modification, Rtn4 forms tubular ER aggregates around theLegionella-containing vacuole (LCV). The loss ofsdetogether withsdhAresults in severe vacuole disruption at early infection timepoints. We tested if Rtn4 modification allowed it to serve as a physical barrier to protect its degradation from host-derived disruptive compartments. The challenge of thertn4-/-macrophages with ΔsdhApartially phenocopied the challenge ofrtn4+/+with ΔsdeΔsdhA, indicating that Rtn4 plays a role in protecting LCV integrity. Depletion ofrtn1inrtn4-/-macrophages potentiated vacuole permeability when compared tortn4-/-macrophages, consistent with Sde targeting multiple reticulon family members to support vacuole integrity. These results indicate thatL. pneumophilaexploits Rtn4 in cooperation with Rtn1 to establish a replication niche by promoting Sde-mediated tubular ER aggregates, arguing that these proteins function as a physical barrier during early steps of LCV biogenesis.
Prevalence of pathogenic bacteria detected by qPCR from cultured Nile tilapia (Oreochromis niloticusLinnaeus, 1758) in southwest Mexico
Authors: Soto-Rodriguez Sonia A. , Lopez Francis I. Marrujo , Aguilar-Rendon Karla G.
Published: June 2023

Nile tilapia (Oreochromis niloticus) is one of the most important aquaculture species in the world. When bacteria are present in cultured tilapia but do not cause a declared disease, it makes them asymptomatic carrier organisms. Once environmental or nutritional conditions change, an outbreak may occur. This is why it is so important to detect pathogens before outbreaks occur. This is the first study that use molecular techniques based on PCR to estimate prevalence of fish pathogens in southwest Mexico. During 2018, 2019, 2020 and 2022 samples of internal organs and lesions of Nile tilapia were taken and analyzed for detection of the main bacterial tilapia pathogens using one-step PCR or qPCR. A total of 2396 samples from the internal organs of Nile tilapia pond and cage cultured come from the Mexican Pacific southwest states of Guerrero, Oaxaca and Chiapas were analyzed. Most of the sampled tilapias were apparently healthy and had no relation between the clinical signs and the pathogens detection was found. NoFrancisellasp. was detected in any sample,Staphylococcussp. was the most prevalent bacterial genus from the three states over time (from 0 to 64 %). Prevalence ofAeromonassp. was from 0 to 4.3 %, although the fish pathogenA. dhakensiswas not detected. Meanwhile,S. iniaewas only detected in Chiapas in 2019 at low prevalence (1.4 %) andS. agalactiaewas detected in the three sites at high prevalence (from 0 to 59 %). BothStreptococcuscan cause streptococcosis, the most dangerous re-emergent disease to cultured tilapia, which means a great risk for tilapia farming in Mexico.
Self-extinguishing relay waves enable homeostatic control of human neutrophil swarming
Authors: Strickland Evelyn , Pan Deng , Godfrey Christian , Kim Julia S. , Hopke Alex , Degrange Maureen , Villavicencio Bryant , Mansour Michael K. , Zerbe Christa S. , Irimia Daniel , Amir Ariel , Weiner Orion D.
Published: June 2023

Neutrophils exhibit self-amplified swarming to sites of injury and infection. How swarming is controlled to ensure the proper level of neutrophil recruitment is unknown. Using anex vivomodel of infection, we find that human neutrophils use active relay to generate multiple pulsatile waves of swarming signals. Unlike classic active relay systems such as action potentials, neutrophil swarming relay waves are self-extinguishing, limiting the spatial range of cell recruitment. We identify an NADPH-oxidase-based negative feedback loop that is needed for this self-extinguishing behavior. Through this circuit, neutrophils adjust the number and size of swarming waves for homeostatic levels of cell recruitment over a wide range of initial cell densities. We link a broken homeostat to neutrophil over-recruitment in the context of human chronic granulomatous disease.