bluuebunny/crossref_metadata_2025 huggingface dataset:
The demand for wildlife not protected by the CITES multilateral treaty
Authors: Watters Freyja , Stringham Oliver , Shepherd Chris , Cassey Phillip
Published: March 2022

The international wildlife trade presents severe conservation and environmental security risks. However, no international regulatory framework exists to monitor the trade of species not listed in the appendices of the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). We explored the composition and dynamics of internationally regulated versus non-regulated trade, focussing on importations of wild-caught terrestrial vertebrates entering the United States of America (US) from 2009-2018. The prominence of the US in global wildlife imports and its detailed data collection conventions allows a unique opportunity to formally assess this substantial but often overlooked and understudied component of the legal wildlife trade. We found 3.6 times the number of unlisted species in US imports compared with CITES-listed species (1,366 versus 378). CITES-listed species were more likely to face reported conservation threats relative to the unlisted species (71.7% vs 27.5%). Yet, we found 376 unlisted species facing conversation threats, 297 species with unknown population trends and 139 species without an evaluation by the IUCN Red List of Threatened Species. Unlisted species appeared novelly in imports at 5.5 times higher rates relative to CITES-listed species, where unlisted reptiles saw the largest rate of entry, averaging 53 unique species appearing in imports for the first time per year. Overall trade volumes were substantially larger for unlisted imports with approximately 11 times the number of animals relative to CITES-listed imports, however, import volumes were similar when compared at a species-by-species level. We found that the countries that were top exporters for CITES-listed shipments were mostly different from exporters of unlisted species. In highlighting the vulnerabilities of the wild-caught unlisted vertebrate trade entering the US and in the face of increasing global demand, we recommend governments adapt policies to monitor the trade of all wildlife.
Intercellular CRISPR Screens Enhance the Discovery of Cancer Immunotherapy Targets
Authors: Yim Soorin , Hwang Woochang , Han Namshik , Lee Doheon
Published: March 2022

Cancer immunotherapy works through the interplay between immune and cancer cells. Particularly, interactions between cytotoxic T lymphocytes (CTLs) and cancer cells, such as PDCD1 (PD-1) and CD274 (PD-L1), are crucial for removing cancer cells. However, immune checkpoint inhibitors targeting these interactions are effective only to a subset of patients, requiring the development of novel immunotherapy drugs with novel targets.

Genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) screening in either cancer or immune cells has been used to discover regulators of immune cell function as immunotherapeutic targets. However, the method has two main limitations. First, performing CRISPR screens in one cell type alone makes it difficult to identify essential intercellular interactions due to the focus on single genes instead of interactions. Second, pooled screening is associated with high noise levels. Therefore, we propose intercellular CRISPR screens, which perform genome-wide CRISPR screening in every interacting cell type to discover intercellular interactions as immunotherapeutic targets.

Intercellular CRISPR screens use two individual genome-wide CRISPR screens one each in immune and cancer cells to evaluate intercellular interactions that are crucial for killing cancer cells. We used two publicly available genome-wide CRISPR screening datasets obtained while triple-negative breast cancer (TNBC) cells and CTLs were interacting. We analyzed 4825 interactions between 1391 ligands and receptors on TNBC cells and CTLs to assess the effects of intercellular interactions on CTL function by incorporating both CRISPR datasets and the expression levels of ligands and receptors.

Our results showed that intercellular CRISPR screens discovered targets of approved drugs, a few of which were not identifiable using single datasets. To quantitatively evaluate the method’s performance, we used data for cytokines and costimulatory molecules because they constitute the majority of immunotherapeutic targets. Combining both CRISPR datasets improved the F1 score of discovering these genes relative to using single CRISPR datasets by more than twice.

Our results indicate that intercellular CRISPR screens can identify novel immune-oncology targets that were not obtained using individual CRISPR screens. The pipeline can be extended to other cancer and immune cell types, such as natural killer cells, to identify important intercellular interactions as potential immunotherapeutic targets.
Polypyrimidine Tract Binding Protein 1 regulates the activation of mouse CD8 T cells
Authors: D’Angeli Vanessa , Monzón-Casanova Elisa , Matheson Louise S. , Gizlenci Özge , Petkau Georg , Gooding Clare , Berrens Rebecca V. , Smith Christopher W. J. , Turner Martin
Published: March 2022

We show that the RNA-binding protein Polypyrimidine Tract Binding Protein 1 (PTBP1) is dispensable for the development of naïve mouse CD8 T cells, but is necessary for the optimal expansion and production of effector molecules by antigen-specific CD8 T cellsin vivo. PTBP1 has an essential role in regulating the early events following activation of the naïve CD8 T cell leading to IL-2 and TNF production. It is also required to protect activated CD8 T cells from apoptosis. PTBP1 controls alternative splicing of over 400 genes in naïve CD8 T cells in addition to regulating the abundance of ∼200 mRNAs. PTBP1 is required for the nuclear accumulation of c-Fos, NFATc2 and NFATc3, but not NFATc1. This selective effect on NFAT proteins correlates with PTBP1-promoted expression of the shorter Aβ1 isoform and exon 13 skipped Aβ2 isoform of the catalytic A-subunit of calcineurin phosphatase. These findings reveal a crucial role for PTBP1 in regulating CD8 T cell activation.
Neonatal Peyer’s patch cDC activation as a pacemaker of postnatal immune maturation
Authors: Torow N. , Li R. , Hitch T. , Mingels C. , Bounny S. al , van Best N. , Stange E.-L. , Benabid A. , Rüttger L. , Gadermayr M. , Runge S. , Treichel N. , Merhof D. , Rosshart S. , Jehmlich N. , von Bergen M. , Heymann F. , Clavel T. , Tacke F. , Lelouard H. , Costa I. , Hornef M. W.
Published: March 2022

Marked differences exist between the mucosal immune system of the neonate and adult host. The pronounced influence of the enteric microbiota in adults suggests a causal relationship between postnatal colonization and immune maturation. However, using metagenomic, metaproteomic, and functional immunological analyses we demonstrate an early presence of bacteria and immunogenic microbial antigens preceding immune maturation in the small intestine, the primary inductive site of intestinal immunity. Instead, transcriptomic, flow cytometric and histological analysis indicated neonatal Peyer’s patch (PP) mononuclear phagocytes (MNP) as rate limiting factor of postnatal immune maturation. Despite the early presence of MNPs, conventional dendritic cells (cDC) of type 1, 2a and 2b exhibited significant age-dependent differences in tissue distribution and cellular composition. Single cell transcriptional profiling and functional assays revealed decreased antimicrobial and antigen processing/presentation capacity, an overall retarded cell maturation and reduced antigen uptake. In cDC2a this resulted in a reduced proportion of CCR7+migratory cells and a consequent defect in CD4 T cell priming. Interestingly, transcriptional profiling of neonatal DC subsets identified reduced expression of type I interferon (IFN)-stimulated genes (ISG). Type I IFN induction by oral administration of the TLR7 agonist R848 accelerated MNP maturation and enhanced cognate antigen CD4 T cell priming. However, humoral responses to oral vaccination in the presence of R848 were significantly reduced. Together, our results identify PP MNP maturation as pacemaker of postnatal mucosal immune priming, indicate the biological role of delayed maturation and demonstrate that targeted interventional strategies allow manipulation of mucosal responses in early life.
Cardio-audio synchronization elicits neural and cardiac surprise responses in human wakefulness and sleep
Authors: Pelentritou Andria , Pfeiffer Christian , Schwartz Sophie , De Lucia Marzia
Published: March 2022

The human brain can infer temporal regularities in auditory sequences with fixed sound-to-sound intervals and in pseudo-regular sequences where sound onsets are locked to cardiac inputs. Here, we investigated auditory and cardio-audio regularity encoding during sleep, when reduced vigilance may result in altered bodily and environmental stimulus processing. Using electroencephalography and electrocardiography in healthy volunteers (N=26) during wakefulness and sleep, we measured the response to unexpected sound omissions within three auditory regularity conditions: synchronous, where sound and heartbeat are temporally locked, isochronous, with fixed sound-to-sound intervals, and a control condition without specific regularity. During wakefulness and all sleep stages, the cardiac signal following sound omissions exhibited a deceleration over time in the synchronous condition only. At the neural level, both the synchronous and isochronous sequences gave rise to a neural omission response in wakefulness and N2 sleep. Regularity encoding in the synchronous and isochronous conditions was characterized by a modulation of the background oscillatory activity in N2 sleep, outlining a possible mechanism through which the brain aligns periods of high neuronal excitability to the expected sound onset. The violation of auditory and cardio-audio regularity elicits cardiac and neural surprise responses across vigilance stages.

Significance Statement

Across vigilance states, the human brain can generate predictions about the future based on past sensory regularities. While this is evident for environmental stimuli, the role of bodily signals in forming sensory prediction remains unknown. Here, we show that the human brain utilizes the temporal relationship between cardiac and auditory inputs in order to anticipate upcoming sounds during wakefulness and sleep. After presenting sounds in synchrony with the ongoing heartbeat, a sound omission elicited both a heartbeat deceleration and a prediction error signal as measured by the electroencephalographic response. Heartbeat signals support auditory regularity encoding during sleep and wakefulness, highlighting one mechanism for optimizing the detection of unexpected stimuli by taking advantage of the continuously monitored cardiac signals.
Dynamic nucleosome organization after fertilization reveals regulatory factors for mouse zygotic genome activation
Authors: Wang Chenfei , Chen Chuan , Liu Xiaoyu , Li Chong , Wu Qiu , Chen Xiaolan , Yang Lingyue , Kou Xiaochen , Zhao Yanhong , Wang Hong , Gao Yawei , Zhang Yong , Gao Shaorong
Published: March 2022

Chromatin remodeling is essential for epigenome reprogramming after fertilization. However, the underlying mechanisms of chromatin remodeling remain to be explored. Here, we investigated the dynamic changes in nucleosome occupancy and positioning in pronucleus-stage zygotes using ultra low-input MNase-seq. We observed distinct features of inheritance and reconstruction of nucleosome position in both paternal and maternal genomes. Genome-wide de novo nucleosome occupancy in the paternal genome was observed as early as 1 hour after the injection of sperm into ooplasm. The nucleosome positioning pattern was continually rebuilt to form nucleosome depletion regions (NDRs) at promoters and transcription factor (TF) binding sites with differential dynamics in paternal and maternal genomes. NDRs formed more quickly on the promoters of genes involved in zygotic genome activation (ZGA), and this formation is closely connected with histone acetylation, but not transcription elongation or DNA replication. Importantly, we found that NDR establishment on the binding motifs of specific TFs might be associated with their potential pioneer functions in ZGA. Further investigations suggested that the predicted factors MLX and RFX1 played important roles in regulating minor and major ZGA, respectively. Our data not only elucidate the nucleosome positioning dynamics in both male and female pronuclei following fertilization, but also provide an efficient method for identifying key transcription regulators during development.
Mouse Genomic Associations withEx VivoSensitivity to Simulated Space Radiation
Authors: Cekanaviciute Egle , Tran Duc , Nguyen Hung , Macha Alejandra Lopez , Pariset Eloise , Langley Sasha , Babbi Giulia , Malkani Sherina , Penninckx Sébastien , Schisler Jonathan C. , Nguyen Tin , Karpen Gary H. , Costes Sylvain. V.
Published: March 2022

Exposure to ionizing radiation is considered by NASA to be a major health hazard for deep space exploration missions. Ionizing radiation sensitivity is modulated by both genomic and environmental factors. Understanding their contributions is crucial for designing experiments in model organisms, evaluating the risk of deep space (i.e. high-linear energy transfer, or LET, particle) radiation exposure in astronauts, and also selecting therapeutic irradiation regimes for cancer patients. We identified single nucleotide polymorphisms in 15 strains of mice, including 10 collaborative cross model strains and 5 founder strains, associated with spontaneous and ionizing radiation-inducedex vivoDNA damage quantified based on immunofluorescent 53BP1+nuclear foci. Statistical analysis suggested an association with pathways primarily related to cellular signaling, metabolism, tumorigenesis and nervous system damage. We observed different genomic associations in early (4 and 8 hour) responses to different LET radiation, while later (24 hour) DNA damage responses showed a stronger overlap across all LETs. Furthermore, a subset of pathways was associated with spontaneous DNA damage, suggesting 53BP1+foci as a potential biomarker for DNA integrity in mouse models. Based on our results, we suggest several mouse strains as new models to further study the impact of ionizing radiation and validate the identified genetic loci. We also highlight the importance of future humanex vivostudies to refine the association of genes and pathways with the DNA damage response to ionizing radiation and identify targets for space travel countermeasures.
The Flexible Linker of Decorin Binding Protein A from Borrelia burgdorferi is a Target of Antibodies in Lyme disease
Authors: Movahed Elaheh , Vance David J , Van Slyke Greta , Ehrbar Dylan , Yates Jennifer , Kullas Karen , Rudolph Michael , Mantis Nicholas J
Published: March 2022

Decorin binding protein A (DbpA) is a surface adhesin expressed by Borrelia burgdorferi, the causative agent of Lyme disease. While DbpA is one of the most immunogenic B. burgdorferi antigens in Lyme disease patients, the B cell epitopes recognized over the course of infection have not been defined. In this report we profiled ∼300 human serum samples from early, mid, and late-stage Lyme disease for IgM and IgG reactivity with DbpA and a tiled DbpA 18-mer peptide array derived from B.b. strains B31 and 297. Using ELISA and multiplex immunoassays (MIA), we identified 12 DbpA-derived peptides whose reactivity was elevated in Lyme disease patients as compared to healthy controls. The most reactive peptide (“A7”) corresponds to the flexible loop between DbpAB31 α-helix 1 and α-helix 2, implicated in influencing DbpA binding to heparin and dermatan sulfate. An A7-like peptide is also reportedly a target of antibodies in Lyme neuroborreliosis patients. The remaining peptides, while highly reactive in serum samples across disease stages, likely represent non-native epitopes, as antibody reactivity to a subset of peptides in competition assays was unaffected by the addition of soluble DbpA. Moreover, peptide reactivity of any given patient sample rarely correlated with overall DbpA antibody levels, suggesting the antibodies were raised against DbpA degradation products. Insights into B cell epitopes on DbpA elicited during Lyme disease have important implications for understanding how B.burgdorferi persists in the face of an overwhelming antibody response.

IMPORTANCE

The bacterium, Borrelia burgdorferi, is the causative agent of Lyme disease, the most reported tick-borne illness in the United States. In humans, clinical manifestations of Lyme disease are complex and can persist for months, even in the face of a robust antibody response directed against numerous B. burgdorferi surface proteins, including decorin binding protein A (DbpA), which is involved in early stages of infection. In this study we employed ∼300 serum samples from Lyme disease patients to better understand antibody reactivity with specific regions (“epitopes”) of DbpA. We identified one epitope on DbpA recognized by antibodies from almost half of the serum samples tested and that, theoretically, should block DbpA’s ability to function in promoting B. burgdorferi colonization of human hosts.
Protective antibodies and T cell responses to Omicron variant three months after the booster dose of BNT162b2 vaccine
Authors: Naaber Paul , Tserel Liina , Kangro Kadri , Sepp Epp , Jürjenson Virge , Kärner Jaanika , Haljasmägi Liis , Haljasorg Uku , Kuusk Marilin , Gerhold Joachim M. , Planken Anu , Ustav Mart , Kisand Kai , Peterson Pärt
Published: March 2022

The high number of mutations in the Omicron variant of SARS-CoV-2 cause its immune escape when compared to the earlier variants of concern (VOC). At least three vaccine doses are required for the induction of Omicron neutralizing antibodies and further reducing the risk for hospitalization. However, most of the studies have focused on the immediate response after the booster vaccination while the duration of immune response is less known. We here studied longitudinal serum samples from the vaccinated individuals up to three months after their third dose of the BNT162b2 vaccine for their capacity to produce protective antibodies and T cell responses to Wuhan and Omicron variants. After the second dose, the antibody levels to the unmutated spike protein were significantly decreased at three months, and only 4% of the individuals were able to inhibit Omicron spike interaction compared to 47%, 38%, and 14% of individuals inhibiting wild-type, delta, and beta variants’ spike protein. Nine months after the second vaccination, the antibody levels were similar to the levels before the first dose and none of the sera inhibited SARS-CoV-2 wild-type or any of the three VOCs. The booster dose remarkably increased antibody levels and their ability to inhibit all variants. Three months after the booster the antibody levels and the inhibition activity were trending lower but still up and not significantly different from their peak values at two weeks after the third dose. Although responsiveness towards mutated spike peptides was lost in less than 20 % of vaccinated individuals, the wild-type spike-specific CD4+ and CD8+ memory T cells were still present at three months after the booster vaccination in the majority of studied individuals. Our data show that two doses of the BNT62b2 vaccine are not sufficient to protect against the Omicron variant, however, the spike-specific antibodies and T cell responses are strongly elicited and well maintained three months after the third vaccination dose.
Neural representations of vicarious rewards are linked to interoception and prosocial behaviour
Authors: Contreras-Huerta Luis Sebastian , Coll Michel-Pierre , Bird Geoffrey , Yu Hongbo , Prosser Annayah , Lockwood Patricia L. , Murphy Jennifer , Crockett Molly J. , Apps Matthew A.J.
Published: March 2022

Every day we constantly observe other people receiving rewards. Theoretical accounts posit that vicarious reward processing might be linked to people’s sensitivity to internal body states (interoception) and facilitates a tendency to act prosocially. However, the neural processes underlying the links between vicarious reward processing, interoception and prosocial behaviour are poorly understood. Previous research has linked vicarious reward processing to the anterior cingulate gyrus (ACCg) and the anterior insula (AI). Can we predict someone’s propensity to be prosocial or to be aware of interoceptive signals from variability in how the ACCg and AI process rewards? Here, participants monitored rewards being delivered to themselves or a stranger during functional magnetic resonance imaging. Later, they performed a task measuring their willingness to exert effort to obtain rewards for others, and a task measuring their propensity to be aware and use interoceptive signals. Using multivariate similarity analysis, we show that people’s willingness to be prosocial is predicted by greater similarity between self and other representations in the ACCg. Moreover, greater dissimilarity in self-other representations in the AI is linked to interoceptive propensity. These findings highlight that vicarious reward is linked to bodily signals in AI, and foster prosocial tendencies through the ACCg.
RNA G-quadruplex structure contributes to cold adaptation in plants
Authors: Yang Xiaofei , Yu Haopeng , Duncan Susan , Zhang Yueying , Cheema Jitender , Miller J. Benjamin , Zhang Jie , Kwok Chun Kit , Zhang Huakun , Ding Yiliang
Published: March 2022

Nucleotide composition is suggested to infer gene functionality and ecological adaptation of species to distinct environments. However, the underlying biological function of nucleotide composition dictating environmental adaptations is largely unknown. Here, we systematically analyzed the nucleotide compositions of transcriptomes across 1000 plants (1KP) and their corresponding habitats. Intriguingly, we found that plants growing in cold climates have guanine (G)-enriched transcriptomes, which are prone to forming RNA G-quadruplex structures. Both immunofluorescence detection and in vivo structure profiling revealed that RNA G-quadruplex formation in plants was globally enhanced in response to cold. Cold-responsive RNA G-quadruplexes strongly enhanced mRNA stability, rather than affecting translation. Disruption of individual RNA G-quadruplex promoted mRNA decay in the cold, leading to impaired plant cold response. Therefore, we propose that plants adopted RNA G-quadruplex structure as a molecular marker to facilitate their adaptation to the cold during evolution.
DNA supercoiling restricts the transcriptional bursting of neighboring eukaryotic genes
Authors: Patel Heta P. , Coppola Stefano , Pomp Wim , Brouwer Ineke , Lenstra Tineke L.
Published: March 2022

DNA supercoiling has emerged as a major contributor to gene regulation in bacteria. The impact of DNA supercoiling on transcription dynamics in eukaryotes is less clear. Here, using single-molecule dual-color RNA imaging in budding yeast, we show that transcriptional bursting of the divergent and tandem GAL genes is coupled. Upon topoisomerase degradation, supercoils that buildup from transcription inhibit subsequent transcription at neighboring genes, thereby reducing their simultaneous bursting. GAL gene transcription is inhibited more by negative than by positive supercoiling accumulation. Unlike bacteria, wildtype yeast has sufficient topoisomerase levels to minimize inhibition from supercoils at adjacent genes. Overall, we discover fundamental differences in supercoiling-mediated gene regulation between bacteria and yeast and show that rapid supercoiling release in eukaryotes ensures proper gene expression of neighboring genes.

One sentence summary

Transcription causes twisting of the DNA double helix, which can inhibit transcription of adjacent genes.
Loss of NF1 causes tactile hypersensitivity and impaired synaptic transmission in a Drosophila model of autism spectrum disorder
Authors: Dyson Alex , Garg Shruti , Evans D. Gareth , Baines Richard A.
Published: March 2022

Autism Spectrum Disorder (ASD) is a neurodevelopmental condition in which the mechanisms underlying its core symptomatology are largely unknown. Studying animal models of monogenic syndromes associated with ASD, such as neurofibromatosis type 1 (NF1), can offer insights into its aetiology. Here, we show that loss of function of the Drosophila NF1 ortholog results in larval tactile hypersensitivity, paralleling the sensory abnormalities observed in individuals with ASD. Mutant larvae also exhibit synaptic transmission deficits at the glutamatergic neuromuscular junction (NMJ), with increased spontaneous but reduced evoked release. Diminished expression of NF1 specifically within central cholinergic neurons induces both excessive neuronal firing and tactile hypersensitivity, suggesting the two may be linked. Furthermore, both impaired synaptic transmission and behavioural deficits are fully rescued via knockdown of Ras proteins. These findings validate NF1-/-Drosophila as a tractable model of ASD with the potential to elucidate important pathophysiological mechanisms.
Lipobiotin-capture magnetic bead assay for isolation, enrichment and detection of Mycobacterium tuberculosis from saliva
Authors: Hansen Julia , Kolbe Katharina , König Inke R. , Scherliess Regina , Hellfritzsch Marie , Malm Sven , Zallet Julia , Hillemann Doris , Wiesmüller Karl-Heinz , Herzmann Christian , Brandenburg Julius , Reiling Norbert
Published: March 2022

Background

Pulmonary Tuberculosis (TB) is diagnosed through sputum samples. As sputum sampling is challenging in children and cachexic patients, the development of diagnostic tests using saliva appears promising but has been discouraged due to low bacterial load and the poor sensitivity. We present a novel and rapid method to enrich Mycobacterium tuberculosis (Mtb) from saliva, which may serve as a basis for a diagnostic saliva test.

Methods

Lipobiotin-functionalized magnetic beads (LMBs) were incubated with Mtb-spiked PBS and saliva from healthy donors as well as with saliva from TB patients. Flow cytometry was used to evaluate the capacity of the beads to bind Mtb, while real-time quantitative polymerase chain reaction (qPCR) was utilized to detect Mtb and estimate the amount of mycobacterial DNA in different sample types.

Results

We found that LMBs bind Mtb efficiently when compared to non-functionalized beads. The development of an qPCR assay based on the use of LMBs (LMB assay) allowed us to enrich mycobacterial DNA in spiked sample types, including PBS and saliva from healthy donors (enrichment of up to 8.7 fold). In Mtb-spiked saliva samples, we found that the LMB assay improved the detection rate of 102 bacteria in a volume of 5 ml from 0 out of 15 (0%) to 6 out of 15 (40%). Consistent with that, the LMB assay increased the rate of correctly identified saliva samples from TB patients in two independent cohorts.

Conclusions

Implementation of the principle of the LMB-based assay may improve the sensitivity of existing diagnostic techniques, e.g. by functionalizing materials that facilitate Mtb sampling from the oral cavity.
Profiling Genetic Diversity Reveals the Molecular Basis for Balancing Function with Misfolding in Alpha-1 Antitrypsin
Authors: Wang Chao , Zhao Pei , Sun Shuhong , Wang Xi , Balch William E.
Published: March 2022

Genetic variation of alpha-1 antitrypsin (AAT) is responsible for alpha-1-antitrypsin deficiency (AATD) leading to gain-of-toxic aggregation in the liver and loss-of-function onneutrophilelastase (NE) inhibitory activity in the lung contributing tochronicobstructivepulmonarydisease (COPD) during aging. To probe the molecular basis for how biology designs the protein fold to achieve balance between sequence, function and structure contributing to AATD in the population, we measured the intracellular monomer and polymer, secreted monomer and polymer and NE inhibitory activity of 75 alpha-1-antitrypsin (AAT) variants. To address the complex folding dynamics affecting the form and function of the protein fold that is differentially impacted by variants in the population, we applied aGaussianprocessregression (GPR) based machine learning approach termedvariationspatialprofiling (VSP). By using a sparse collection of extant variants to link genotype to phenotype, VSP mapsspatialcovariance (SCV) relationships that quantitate the functional value of every residue in the wild-type (WT) AAT sequence with defined uncertainty in the context of its protein fold design. The SCV-based uncertainty allows us to pinpoint critical short- and long-range residue interactions involving 3 regions-the N-terminal (N1), middle (M2) and carboxyl-terminal (C3) of AAT polypeptide sequence that differentially contribute to the balance between function and misfolding of AAT, thus providing an unanticipated platform for precision therapeutic development for liver and lung disease. By understanding mechanistically the complex fold design of the metastable WT AAT fold, we posit that GPR-based SCV provides a foundation for understanding the evolutionary design of the fold from the ensemble of structures found in the population driving biology for precision management of AATD in the individual.
Field evaluation of a novel, rapid diagnostic assay RLDT, and molecular epidemiology of enterotoxigenicE. coliamong Zambian children presenting with diarrhea
Authors: Silwamba Suwilanji , Chilyabanyama Obvious N. , Liswaniso Fraser , Chisenga Caroline C , Chilengi Roma , Dougan Gordon , Kwenda Geoffrey , Chakraborty Subhra , Simuyandi Michelo
Published: March 2022

Background

EnterotoxigenicEscherichia coli(ETEC) is one of the top aetiologic agents of diarrhea in children under the age of 5 in low-middle income countries (LMICs). The lack of point of care diagnostic tools for routine ETEC diagnosis results in limited data regarding the actual burden and epidemiology in the endemic areas. We evaluated performance of the novel Rapid LAMP based Diagnostic Test (RLDT) for detection of ETEC in stool as a point of care diagnostic assay in a resource-limited setting.

Methods

We conducted a cross-sectional study of 324 randomly selected stool samples from children under 5 presenting with moderate to severe diarrhea (MSD). The samples were collected between November 2012 to September 2013 at selected health facilities in Zambia. The RLDT was evaluated by targeting three ETEC toxin genes [heat labile toxin (LT) and heat stable toxins (STh, and STp)]. Quantitative PCR was used as the “gold standard” to evaluate the diagnostic sensitivity and specificity of RLDT for detection of ETEC. We additionally described the prevalence and seasonality of ETEC.

Results

The study included 50.6% of participants that were female. The overall prevalence of ETEC was 19.8% by qPCR and 19.4 % by RLDT. The children between 12 to 59 months had the highest prevalence of 22%. The study determined ETEC toxin distribution was LT (49%), ST (34%) and LT/ST (16%). The sensitivity and specificity of the RLDT compared to qPCR using a Ct 35 as the cutoff, were 90.7% and 97.5% for LT, 85.2% and 99.3% for STh and 100% and 99.7% for STp, respectively.

Conclusion

The results of this study suggest that RLDT is sufficiently sensitive and specific and easy to implement in the endemic countries. Being rapid and simple, the RLDT also presents as an attractive tool for point-of-care testing at the health facilities and laboratories in the resource-limited settings.

Author Summary

ETEC is one of the top causes of diarrheal diseases in low and middle income countries. The advancement of molecular diagnosis has made it possible to accurately detect ETEC in endemic areas. However, the complexity, infrastructure and cost implication of these tests has made it a challenge to routinely incorporate them in health facilities in endemic settings. The ETEC RLDT is a simple and cost-effective molecular tool that can be used to screen for ETEC in resource limited settings. Here, we described the performance of the RLDT against a qPCR as the gold standard. Our findings showed that the ETEC RLDT performs comparable to the qPCR and would be a suitable screening tool in health facilities in recourse limited settings.
EFFECTS OF HIV INFECTION AND FORMER COCAINE DEPENDENCE ON NEUROANATOMICAL MEASURES AND NEUROCOGNITIVE PERFORMANCE
Authors: Wakim Kathryn-Mary , Freedman Edward G. , Tivarus Madalina E. , Christensen Zachary , Molholm Sophie , Foxe John J.
Published: March 2022

Evidence from animal research, postmortem analyses, and MRI investigations indicate substantial morphological alteration in brain structure as a function of HIV or cocaine dependence (CD). Although previous research on HIV+ active cocaine users suggests the presence of deleterious morphological effects in excess of either condition alone, a yet unexplored question is whether there is a similar deleterious interaction in HIV+ individuals with CD who are currently abstinent. To this end, the combinatorial effects of HIV and CD history on regional brain volume, cortical thickness, and neurocognitive performance was examined across four groups of participants: healthy controls, HIV-negative individuals with a history of CD, HIV+ individuals with no history of CD, HIV+ individuals with a history of CD. Our analyses revealed no statistical evidence of an interaction between both conditions on brain morphometry and neurocognitive performance. While descriptively, individuals with comorbid HIV and a history of CD exhibited the lowest neurocognitive performance scores, using Principle Component Analysis of neurocognitive testing data, HIV was identified as a primary driver of neurocognitive impairment. Higher caudate volume was evident in CD+ participants relative to CD-participants. Taken together, these data provide evidence of independent effects of HIV and CD history on brain morphometry and neurocognitive performance in cocaine-abstinent individuals.
Circadian rhythm of temporal reproduction as a function of stimulus eccentricity in the visual field
Authors: Wang Wei , Yin Xuanzi , Bao Yan
Published: March 2022

The aim of this study was to elucidate how temporal reproduction changed in a diurnal time course. The overestimated reproduction showed a smaller difference to the standard in the morning and evening and a larger difference in the afternoon, and the underestimated reproduction showed an inverse change. No eccentricity effect in temporal production was observed, which indicates homogenous temporal processing shared in the perifoveal and peripheral visual fields.
A Systematic Strategy for Identifying Causal Single Nucleotide Polymorphisms and Their Target Genes on Juvenile Arthritis Risk Haplotypes
Authors: Jiang Kaiyu , Liu Tao , Kales Susan , Tewhey Ryan , Kim Dongkyeong , Park Yungki , Jarvis James N.
Published: March 2022

Introduction

Although genome-wide association studies (GWAS) multiple regions conferring genetic risk for juvenile idiopathic arthritis (JIA), we are still faced with the task of identifying the single nucleotide polymorphisms (SNPs) on the disease haplotypes that exert the biological effects that confer risk. Until we identify the risk-driving variants, identifying the genes influenced by these variants, and therefore translating genetic information to improved clinical care, will remain an insurmountable task. We used a function-based approach for identifying causal variant candidates and the target genes on JIA risk haplotypes.

Methods

We used a massively parallel reporter assay (MPRA) in myeloid K562 cells to query the effects of 5,226 SNPs in non-coding regions on JIA risk haplotypes for their ability to alter gene expression when compared to the common allele. The assay relies on 180 bp oligonucleotide reporters (“oligos”) in which the allele of interest is flanked by its cognate genomic sequence. Barcodes were added randomly by PCR to each oligo to achieve >20 barcodes per oligo to provide a quantitative read-out of gene expression for each allele. Assays were performed in both unstimulated K562 cells and cells stimulated overnight with interferon gamma (IFNg). As proof of concept, we then used CRISPRi to demonstrate the feasibility of identifying the genes regulated by enhancers harboring expression-altering SNPs.

Results

We identified 553 expression-altering SNPs in unstimulated K562 cells and an additional 490 in cells stimulated with IFNg. We further filtered the SNPs to identify those plausibly situated within functional chromatin, using open chromatin and H3K27ac ChIPseq peaks in unstimulated cells and open chromatin plus H3K4me1 in stimulated cells. These procedures yielded 42 unique SNPs (total = 84) for each set. Using CRISPRi, we demonstrated that enhancers harboring MPRA-screened variants in the TRAF1 and LNPEP/ERAP2 loci regulated multiple genes, suggesting complex influences of disease-driving variants.

Conclusion

Using MPRA and CRISPRi, JIA risk haplotypes can be queried to identify plausible candidates for disease-driving variants. Once these candidate variants are identified, target genes can be identified using CRISPRi informed by the 3D chromatin structures that encompass the risk haplotypes.
Decreased spliceosome fidelity inhibits mTOR signalling and promotes longevity via an intron retention event
Authors: Huang Wenming , Kew Chun , Fernandes Stephanie A. , Loerhke Anna , Han Lynn , Demetriades Constantinos , Antebi Adam
Published: March 2022

Changes in splicing fidelity are associated with loss of homeostasis and ageing1–3, yet only a handful of splicing factors have been shown to be causally required to promote longevity 1–3, and the underlying mechanisms and downstream targets in these paradigms remain elusive. Surprisingly, we found a hypomorphic mutation within RNP-6/PUF60, a spliceosome component promoting weak 3’ splice site recognition, which causes aberrant splicing, elevated stress responses, and enhances longevity in Caenorhabditis elegans. Through genetic suppressor screens, we identify a gain-of-function mutation within rbm-39, an RNP-6 interacting splicing factor, which increases nuclear speckle formation, alleviates splicing defects and curtails longevity caused by rnp-6 mutation. By leveraging the splicing changes induced by RNP-6/RBM-39 activities, we uncover a single intron retention event in egl-8/phospholipase C B4 as a key splicing target prolonging life. Genetic and biochemical evidence show that neuronal RNP-6/EGL-8 downregulate mTORC1 signaling to control organismal life span. In mammalian cells, PUF60 downregulation also potently and specifically inhibits mTORC1 signaling. Altogether, our results reveal that splicing fidelity modulates mTOR signaling and suggest a potential therapeutic strategy to delay ageing.
Diverse regulatory pathways modulate “bet hedging” of competence induction in epigenetically-differentiated phase variants ofStreptococcus pneumoniae
Authors: Kwun Min Jung , Ion Alexandru V. , Oggioni Marco R. , Bentley Stephen D. , Croucher Nicholas J.
Published: March 2022

Despite enablingStreptococcus pneumoniaeto acquire antibiotic resistance and evade vaccine-induced immunity, transformation occurs at variable rates across pneumococci. Phase variants of isolate RMV7, distinguished by altered methylation patterns driven by the translocating variable restriction-modification (tvr) locus, differed significantly in their transformation efficiencies and biofilm thicknesses. These differences were replicated when the correspondingtvralleles were introduced into an RMV7 derivative lacking the locus. RNA-seq identified differential expression of the type 1 pilus, causing the variation in biofilm formation, and inhibition of competence induction in the less transformable variant, RMV7domi. This was partly attributable to lower expression of ManLMN in RMV7domi, which promoted competence induction through importingN-acetylglucosamine. This effect was potentiated by orthologues of the gram-negative competence regulatory machinery. Furthermore, a phage-related chromosomal island was more active in RMV7domi, which inhibited transformation by increasing expression of the stress response proteins ClpP and HrcA. However, HrcA increased competence induction in the other variant, with its effects depending on Ca2+supplementation or heat shock. Hence the heterogeneity in transformation efficiency likely reflects the diverse signalling pathways by which it is affected. This regulatory complexity will modulate population-wide responses to synchronising quorum sensing signals to produce co-ordinated yet stochastic “bet hedging” behaviour.
Zoonotic Tick Borne Pathogens in Ticks at Human-Wildlife-Livestock interfaces in Kwale County, Kenya
Authors: Godani Salim Kobo , Menza C. Nelson , Muturi W. Margaret
Published: March 2022

Background

Ticks are vectors that can harbor pathogens like viruses, protozoa and bacteria that can cause zoonotic diseases in human. Human gets infected through tick bites where the pathogens are passed to the human blood. One tick bite can lead to transmission of more than one disease. Ticks are obligate hematophagous, infest most vertebrates globally. Lack of surveillance information about the tick-borne pathogens and diseases has made it impossible to assess its impact to the community.

Materials and Methods

We morphologically identified ticks collected from two game reserves in Kenya, locations near the Shimba Hills National Reserve (SHNR) and Tsavo National Reserve (TNR). We identified tick-borne pathogens by HRM analysis and sequencing of specific RT-PCR products of Anaplasma, Ehrlichia, and Rickettsia.

Results

A total of 317 (281 adult ticks and 36 nymphs) was collected at near Tsavo National Reserve in Taita Taveta County, which includes seven species. Amblyomma was the most sampled genus with Amblyomma gemma being the most sampled species (n=135, 42.6%). Other Amblyomma species sampled was Amblyomma Variegatum (n= 40, 12.62%). Greatest species diversity was identified in Rhipicephalus genus with four species identified that includes; Rhipicephalus appendiculatus (n=44, 13.9%), Rhipicephalus Averts (n=1, 0.31%), Rhipicephalus Decoloratus (n=5, 1.6%), Rhipicephalus Pulchellus (n= 91, 28.7%). A single species of Hyalomma sp. was sampled. From near Shimba Hill game reserve (SHNR), a total of 240 adult’s ticks were sampled that comprises of eight species. Amblyomma was the most sampled genus and again Amblyomma gemma being the most sampled species (n=156, 65 %). Other Amblyomma species sampled includes; Amblyomma Lepidum (n= 5, 2.1 %), Amblyomma Variegatum (n= 15, 6.3 %). Greatest species diversity was also identified in Rhipicephalus genus with four species identified that includes; Rhipicephalus appendiculatus (n=18, 7.5 %), Rhipicephalus Averts (n=6, 2.5 %), Rhipicephalus Decoloratus (n=4, 1.7 %), Rhipicephalus Pulchellus (n= 34, 14.2 %). The least sampled species was a single species of Hyalomma Scupense (n=2, 0.83 %). At near Tsavo National Reserve (TNR) in Taita Taveta County, a total of three pools of Rhipicephaline appendiculatus were positive for Theileria parva, two pools of Rhipicephaline evertsi for Anaplasma platys and one pool of Amblyomma variegatum nymphs for Rickettsia africae. From near Shimba Hill game reserve (SHNR), Kwale County, Rickettsia africae pathogen was detected in two pools of Am. variegatum and one pool of Am. Gemma. Rickettsia sp. and Anaplasma sp. were detected in Am. Gemma and Rh. evertsi respective. R. aeschlimannii was detected in a pool of Am. Gemma. These findings highlight the risk of transmission of zoonotic R. africae and unclassified Rickettsia spp. to humans causing African tick-bite fever and other spotted-fever group rickettsioses.
Alterations of the axon initial segment in multiple sclerosis
Authors: Senol Aysegul Dilsizoglu , Pinto Giulia , Beau Maxime , Guillemot Vincent , Dupree Jeff L. , Stadelmann Christine , Ranft Jonas , Lubetzki Catherine , Davenne Marc
Published: March 2022

Grey matter damage has been established as a key contributor to disability progression in multiple sclerosis. Aside from neuronal loss and axonal transections, which predominate in cortical demyelinated lesions, synaptic alterations have been detected in both demyelinated plaques and normal-appearing grey matter, resulting in functional neuronal damage. The axon initial segment is a key element of neuronal function, responsible for action potential initiation and maintenance of neuronal polarity. Despite several reports of profound axon initial segment alterations in different pathological models, among which experimental auto-immune encephalomyelitis, whether the axon initial segment is affected in multiple sclerosis is still unknown. Using immunohistochemistry, we analyzed axon initial segments from control and multiple sclerosis tissue, focusing on layer 5/6 pyramidal neurons in the neocortex and Purkinje cells in the cerebellum and performed analysis on the parameters known to control neuronal excitability, i.e., axon initial segment length and position. We found that the axon initial segment length was unchanged among the different multiple sclerosis samples, and not different from controls. In contrast, in both cell types, the axon initial segment position was altered, with an increased soma-axon initial segment gap, in both active and inactive demyelinated lesions. In addition, using a computational model, we show that this increased gap between soma and axon initial segment might increase neuronal excitability. Taken together, these results show for the first time changes of axon initial segments in multiple sclerosis, in active as well as inactive grey matter lesions in both neocortex and cerebellum, which might alter neuronal function.
CF-Seq, An Accessible Web Application for Rapid Re-Analysis of Cystic Fibrosis Pathogen RNA Sequencing Studies
Authors: Neff Samuel L. , Hampton Thomas H. , Puerner Charles , Cengher Liviu , Doing Georgia , Lee Alexandra J. , Koeppen Katja , Cheung Ambrose L. , Hogan Deborah A. , Cramer Robert A. , Stanton Bruce A.
Published: March 2022

Researchers studying cystic fibrosis (CF) pathogens have produced numerous RNA-seq datasets which are available in the gene expression omnibus (GEO). Although these studies are publicly available, substantial computational expertise and manual effort are required to compare similar studies, visualize gene expression patterns within studies, and use published data to generate new experimental hypotheses. Furthermore, it is difficult to filter available studies by domain-relevant attributes such as strain, treatment, or media, or for a researcher to assess how a specific gene responds to various experimental conditions across studies. To reduce these barriers to data re-analysis, we have developed an R Shiny application called CF-Seq, which works with a compendium of 147 studies and 1,446 individual samples from 13 clinically relevant CF pathogens. The application allows users to filter studies by experimental factors and to view complex differential gene expression analyses at the click of a button. Here we present a series of use cases that demonstrate the application is a useful and efficient tool for new hypothesis generation. (CFSeq: http://scangeo.dartmouth.edu/CFSeq/)