bluuebunny/crossref_metadata_2025 huggingface dataset:
Urine CXCL10 to Assess BK Polyomavirus Replication After Kidney Transplantation
Authors: Haller Jana , Diebold Matthias , Leuzinger Karoline , Wehmeier Caroline , Handschin Joelle , Amico Patrizia , Hirt-Minkowski Patricia , Steiger Jürg , Dickenmann Michael , Hirsch Hans H. , Schaub Stefan
Published: July 2023

Background.

Urine CXCL10 is a biomarker for renal allograft inflammation induced by rejection, urinary tract infection, or BK polyomavirus (BKPyV) replication. This study aimed to compare urine CXCL10 levels in different stages of BKPyV reactivation and to investigate urine CXCL10 as a biomarker for BKPyV replication.

Methods.

We included 763 urine samples (235 patients) from an interventional, randomized trial obtained in the context of regular screening for urine CXCL10 levels. All urine samples had a complete urine sediment analysis, no rejection episode noted within 30 d before urine collection, and a urine decoy cell analysis was conducted within ±3 d.

Results.

Urine CXCL10 levels were 2.31 ng/mmol in samples without BKPyV viruria, slightly rose to 4.35 ng/mmol with BKPyV viruria, and then markedly increased to 16.42 ng/mmol when decoy cells were detectable, but still in the absence of BKPyV DNAemia (P < 0.001). The highest urine CXCL10 values were observed in samples with BKPyV DNAemia (median 42.59 ng/mmol). The area under the curve of urine CXCL10 levels to detect ≥3 decoy cells was 0.816. At a CXCL10 cutoff of 3 ng/mmol, the negative predictive value was 97%. The area under the curve of urine CXCL10 levels to detect BKPyV DNAemia was 0.882, with a negative predictive value of 99% at a CXCL10 cutoff of 3 ng/mmol.

Conclusions.

Urine CXCL10 levels are already significantly elevated in BKPyV viruria (especially with decoy cell shedding) and further increase with BKPyV DNAemia. Low urine CXCL10 values can rule out the presence of ≥3 decoy cells and BKPyV DNAemia with high certainty.
Assessment and Evaluation of Alcohol Use Disorder in Liver Transplant
Authors: Branagan William T. , Norvell J. P.
Published: July 2023

Alcohol-related liver disease is the most common indication for liver transplantation. It is essential for providers in transplantation to be informed of the state of the science in evaluation of alcohol use disorder (AUD). This review examines the broad range of approaches to the evaluation of AUD ranging from traditional interview approaches to recent literature on artificial intelligence models. The empirical support for methods of evaluation is examined. The authors discuss the use of each method in the context of patients seeking a liver transplant for alcohol-related liver disease. This review emphasizes the importance of using objective assessments so that transplant centers make evidence-based decisions and reduce cognitive bias. The review concludes with a proposed assessment battery for evaluation and bridges to future directions in the field of AUD assessment in liver transplantation.
Evaluation of kit-based loop-mediated isothermal amplification (TB-LAMP) assay in the diagnosis of tubercular lymphadenitis
Authors: Das Saurav , Nischal Neeraj , Singh Binit Kumar , Ray Animesh , Jorwal Pankaj , Soneja Manish , Khan Maroof Ahmad , Sikka Kapil , Sinha Sanjeev , Wig Naveet
Published: July 2023

Background: The rapid and accurate diagnosis of tubercular lymphadenitis remains a challenging task today.WHO endorsed LoopAMP™ MTBC kit (TB-LAMP) as a replacement for sputum-smear microscopy in the diagnosis of pulmonary tuberculosis (PTB). However, no prospective diagnostic accuracy study of TB-LAMP in tubercular lymphadenitis in adults has been done yet. Current study evaluated the diagnostic performance of TB-LAMP in tubercular lymphadenitis (LNTB).Material & Methods: In a prospective observational study conducted at a tertiary care hospital in India, 90 subjects (age>18years) suspected of LNTB were consecutively recruited and followed up till six months between January 2019 and December 2020. Samples were processed for microscopy, culture, GeneXpert®, histopathology and TB-LAMP. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of TB-LAMP against composite reference standard (CRS) and culture were determined.Results : TB-LAMP showed a sensitivity of 83.78% (95% CI, 73.76-90.47) and specificity of 81.25% (95% CI, 56.99-93.41) respectively against CRS. The PPV and NPV were 95.38% (95% CI, 87.29-98.42) and 52.00% (95% CI, 33.50-69.97) respectively. TB-LAMP showed a sensitivity of 88.89% (95% CI, 71.94-96.15) and specificity of 36.17% (95% CI, 23.97-50.46) respectively against culture. The PPV and NPV were 44.44% (95% CI, 32-57.62) and 85% (95% CI, 63.96-94.76) respectively.Conclusion: TB-LAMP can be used over conventional microscopy for the diagnosis of TB in lymph node specimens at primary health care centres. This is the rapid and cost-effective diagnosis of LNTB in resource-limited settings due to good sensitivity and NPV.
Detection and significance of antibody response in the diagnosis of pulmonary tuberculosis using enzyme-linked immunosorbent assay: a case-control study.
Authors: Chaudhary Pooja , Mohanty Keshar K. , Kumar Santosh , Sikarwar Arun P. , Patil Shripad A
Published: July 2023

Objective

An ELISA test was performed to detect antibody response in the serum of pulmonary tuberculosis (PTB) patients with confirmed acid-fast bacilli (AFB) in sputum along with healthy controls using the in-house prepared antigen of Mycobacterium tuberculosis (MTB) H37Rv strain sonicated soluble extract.

Methods

The present study has recruited participants with AFB sputum smear positivity (N = 652) and healthy controls (N= 251). Serum samples of both groups were subjected to antibody analysis by ELISA. The evaluation of diagnostic specificity was based on a positive score represented by titers above the cut–off value. Statistical analysis was performed.

Results

A significant antibody response was (P value < 0.0001) observed in PTB sputum smear-positive patients as compared to healthy controls. We found that 577 PTB patients show positive antibody responses against the prepared sonicated antigen. Antibody detection by using the ELISA method has revealed 88.50% sensitivity and 97.61% specificity.

Conclusion

This immunological test could be much more efficient to detect antibody response in PTB sputum smear-positive patients. Further, the test may be useful in smear-negative cases of TB and those with latent infection.
Whole genome sequencing and genotyping Klebsiella pneumoniae multi-drug resistant hospital isolates from Western Kenya
Authors: Dinda Victor , Kimang’a Andrew Nyerere , Kariuki Daniel , Sifuna Anthony Wawire , O’Brien Thomas James , Welch Martin , Reva Oleg N
Published: July 2023

Objectives

Klebsiella pneumoniae are frequent causes of nosocomial infections worldwide. Sequence type 147 (ST147) has been reported as a major circulating high-risk lineage in many countries, and appears to be a formidable platform for the dissemination of antimicrobial resistance (AMR) determinants. Distribution of this pathogen in hospital of Western Africa is scarcely studied. An objective of this work was to perform whole genome sequencing of AMR isolates from a referral hospital in Kakamega (Kenya) for genotyping and identification of AMR and virulence determinants.

Methods

In total, 15 K. pneumoniae isolates showing a broad spectrum antimicrobial resistance were selected for whole genome sequencing by Illumina HiSeq 2500 platform.

Results

It was found that ST147 has become the dominant lineage among the K. pneumonia causing both the community and the hospital acquired infections from different infection sites, whereas other STs were predominantly uropathogens. Two broad range plasmids enriched in AMR determinants were found in the sequenced isolates.

Conclusions

Our data suggest that the evolutionary success of ST147 may be linked with the acquisition of these plasmids, and their propensity to accrue AMR and virulence determinants. ST147 was reported to be a dominant lineage in many countries worldwide, but it has been not frequent in Africa before. This may indicate an influx of new nosocomial pathogens with new virulence genes into African hospitals from other continents.
Emergence of linezolid-resistant Enterococcus faecium in a tertiary hospital in Copenhagen
Authors: Misiakou Maria-Anna , Hertz Frederik Boetius , Schønning Kristian , Häussler Susanne , Nielsen Karen Leth
Published: July 2023

Linezolid is used as first-line treatment of infections caused by vancomycin-resistant

Enterococcus faecium

. However, resistance to linezolid is increasingly detected. The aim of the present study was to elucidate the causes and mechanisms for the increase in linezolid-resistant

E. faecium

at Copenhagen University Hospital – Rigshospitalet. We therefore combined patient information on linezolid treatment with whole-genome sequencing data for vancomycin- or linezolid-resistant

E. faecium

isolates that had been systematically collected since 2014 (n=458). Whole-genome sequencing was performed for multilocus sequence typing (MLST), identification of linezolid resistance-conferring genes/mutations and determination of phylogenetically closely related strains. The collection of

E. faecium

isolates belonged to prevalent vancomycin-resistant MLST types. Among these, we identified clusters of closely related linezolid-resistant strains compatible with nosocomial transmission. We also identified linezolid-resistant enterococcus isolates not genetically closely related to other isolates compatible with de novo generation of linezolid resistance. Patients with the latter isolates were significantly more frequently exposed to linezolid treatment than patients with related linezolid-resistant enterococcus isolates. We also identified six patients who initially carried a vancomycin-resistant, linezolid-sensitive enterococcus, but from whom vancomycin-resistant, linezolid-resistant enterococci (LVRE) closely related to their initial isolate were recovered after linezolid treatment. Our data illustrate that linezolid resistance may develop in the individual patient subsequent to linezolid exposure and can be transmitted between patients in a hospital setting.
Origin of Osteoclasts: Osteoclast Precursor Cells
Authors: Tsai Jefferson , Kaneko Kaichi , Suh Andrew J. , Bockman Richard , Park-Min Kyung-Hyun
Published: July 2023

Osteoclasts are multinucleated bone-resorbing cells and a key player in bone remodeling for health and disease. Since the discovery of osteoclasts in 1873, the structure and function of osteoclasts and the molecular and cellular mechanisms of osteoclastogenesis have been extensively studied. Moreover, it has been well established that osteoclasts are differentiated in vitro from myeloid cells such as bone marrow macrophages or monocytes. The concept showing that osteoclasts are derived from a specific population (named osteoclast precursor cells [OCPs]) among myeloid cells has been long hypothesized. However, the specific precursor population of osteoclasts is not clearly defined yet. A growing body of work provides evidence of the developmental origin and lifespan of murine osteoclasts, particularly in vivo. Here, we review the emerging evidence that supports the existence of OCPs and discuss current insights into their identity.
Effect of Vitamin B6 on Osteoporosis Fracture
Authors: Welan Rahmani
Published: July 2023

Osteoporosis is a medical condition that causes bones to become weak and brittle, making them more likely to break or fracture. This condition occurs when the body loses too much bone, makes too little bone, or both. Bones are living tissues that are constantly being broken down and rebuilt. However, as we age, this process slows down, and our bodies may be unable to replace bone as quickly as it is broken down. This can lead to a loss of bone density, making bones weaker and more prone to fracture. Unfortunately, osteoporosis often has no symptoms until a bone is broken, so taking steps to prevent this condition is essential. This may include getting enough calcium and vitamin D, engaging in weight-bearing exercise, quitting smoking, limiting alcohol intake, and taking medication if prescribed by a doctor. Vitamin B6, also known as pyridoxine, is a water-soluble vitamin that plays a vital role in various bodily functions. It is part of the B-vitamin complex and is essential for metabolizing proteins, carbohydrates, and fats. Vitamin B6 produces neurotransmitters such as serotonin, which helps regulate mood, and dopamine, which is involved in motivation and reward. Several previous studies have found that women with higher intakes of vitamin B6 have higher bone mineral density (BMD) of the hips and spine than women with lower intakes. Another study found that men and women with higher blood levels of vitamin B6 had higher BMD in the hips and spine.
The ratio of interacting miRNAs’ expressions is a robust biomarker for disease classification in multi-center data
Authors: Zhang Yonghao , Ma Cuidie , Ding Rui , Chen Han , Xu Lida , Yu Changyuan
Published: July 2023

Background

Many miRNA-based diagnostic models have been constructed to distinguish diseased individuals. However, due to the inherent differences across different platforms or within multi-center data, the models usually fail in the generalization for medical application.

Results

Here, we proposed to use the within-sample expression ratios of related miRNA pairs as markers, by utilizing the internal miRNA: miRNA interactions. The ratio of the expression values between each miRNA pair turned out to be more stable cross multiple data source. Moreover, we adopted the genetic algorithm to solve the curse of dimensions when exploring the features.

Conclusions

The application results on three example datasets demonstrated that the expression ratio of interacting miRNA pair is a promising type of biomarker, which is insensitive to batch effects and has better performance in disease classifications.
Training and testing a CNN-based engine for brain MRI scan classification and segmentation
Authors: Dhawan Kunaal , Agrawal Utkarsh
Published: July 2023

Biomedical imaging data (from X-rays, CT scans, MRIs) is a rich and crucial source of information for diagnosing and treating patients, yet the large volume of visual data generated can be challenging even for the most experienced clinical professionals to handle and use for diagnosis with high accuracy. With the traditional diagnosis approaches, patients usually face high costs too. Sometimes, abnormalities remain unspotted, causing delays in intervention which can be the difference between saving and losing patient lives. Most traditional radiomics studies use hand-crafted feature extraction techniques, such as texture analysis, followed by conventional machine learning classifiers, such as random forests and support vector machines (SVMs). In this paper, we establish that X-ray, MRI or CT-image classification using convolutional neural networks (CNNs) could be an efficient, cost-effective and fast approach for diagnosis and interpretation. The research is focused on training a CNN algorithm to develop diagnostic analysis power using 250 brain MRI images and then testing the accuracy and predictive power of the developed CNN algorithm on 5 images. We have shown the superiority of CNN models in image classification compared to traditional ML Classifiers that used an extensive ablation study. We have then compared a selected set of existing models with the new model we have built based on the EfficientNet v2-S architecture with a high classification accuracy of 98% – showing a high predictive power to provide various differential diagnoses from the brain MRI scans.
Attitudes to phage therapy among Australian infectious diseases physicians
Authors: Plymoth Martin , Lynch Stephanie A. , Khatami Ameneh , Sinclair Holly A. , Sacher Jessica C. , Zheng Jan , Lin Ruby CY. , Iredell Jonathan R.
Published: July 2023

Due to the rise in antimicrobial resistance (AMR), there has been an increased interest in phage therapy to treat multi-drug resistant infections. In Australia, phage therapy is predominantly used in small clinical studies or for compassionate use, however, despite its potential expansion in modern medicine, the perception of phage therapy among medical professionals remains largely unknown. Therefore, we conducted a national survey of Australian infectious diseases and clinical microbiology advanced trainees and specialists to assess their knowledge, areas of interest, and concerns around the use of phage therapy in clinical practice in Australia. Our survey received 92 responses from infectious diseases and clinical microbiology professionals across all states of Australia. The majority of those surveyed believed that the current national plan for controlling AMR is inadequate and that phage therapy may be an effective solution; with 97% of respondents indicating that they would consider using phage therapy meeting established guidelines for purity and safety (United States Food and Drug Administration and/or European Union guidelines). The respondents indicated a preference for bespoke therapy, with Gram-negative pathogens highlighted as priority targets. Alongside the phage therapy delivery protocols, therapeutic phage monitoring (TPM; like therapeutic drug monitoring (TDM)) was considered important. Cystic Fibrosis, lung-infections, prosthetic device related infections, and infections among patients following transplantation and/or immunosuppression were highly ranked in terms of priorities for clinical syndromes. Accessibility was highlighted as a barrier to phage therapy, specifically timely access (72%) and logistics of phage procurement and administration (70%). Altogether, these results suggest the support of phage therapy among infectious diseases and clinical microbiology advanced trainees and specialists in Australia, and highlights areas of focus and priority in order to advance phage therapy in modern medicine.
Genetic Susceptibility to Astrovirus Diarrhea in Bangladeshi Infants
Authors: Chen Laura , Munday Rebecca M. , Haque Rashidul , Duchen Dylan , Nayak Uma , Korpe Poonum , Mentzer Alexander J. , Kirkpatrick Beth D. , Wojcik Genevieve L. , Petri William A. , Duggal Priya
Published: July 2023

Astroviral infections commonly cause acute nonbacterial gastroenteritis in children globally. However, these infections often go undiagnosed outside of research settings. There is no treatment available for astrovirus, and Astroviridae strain diversity presents a challenge to potential vaccine development. To address our hypothesis that host genetic risk factors are associated with astrovirus disease susceptibility, we performed a genome-wide association study (GWAS) of astrovirus infection in the first year of life from children enrolled in two Bangladeshi birth cohorts. We identified a novel region on chromosome 1 near the loricrin gene (LOR) associated with astrovirus diarrheal infection (rs75437404, meta-analysis p-value=8.82×10−9, A allele OR=2.71) and on chromosome 10 near the prolactin releasing hormone receptor gene (PRLHR) (rs75935441, meta-analysis p-value=1.33×10−8, C allele OR=4.17). The prolactin-releasing peptide has been shown to influence feeding patterns and energy balance in mice. Additionally, several SNPs in the chromosome 1 locus have previously been associated with expression of innate immune system genesPGLYRP4, S100A9, andS100A12. This study identified two significant host genetic regions that may influence astrovirus diarrhea susceptibility and should be considered in further studies.
The impact ofERP29on the progression of pharyngeal squamous cell carcinoma
Authors: Carron Juliana , Coser Lilian de Oliveira , Kelemen Andrea , Tolstonog Genrich V , Lima Carmen Silvia Passos , Lourenço Gustavo Jacob
Published: July 2023

Objectives

We investigatedERP29gene role on pharynx squamous cell carcinoma (PSCC) progression in cisplatin (CDDP)-sensitive (FaDu and LAU-2063), CDDP-treated (FaDu-CDDP), and CDDP-resistant (FaDu-R) cells.

Materials and Methods

Cells, modified to induceERP29overexpression or silencing, were mainly submitted to cell proliferation, necrosis, and migration assay. E-cadherin immunoexpression was assessed in three-dimensional spheroids. WNT, MAPK, and PI3K/AKT pathways genes’ expression were identified by PCR array and validated by qPCR. The influence of microRNA miR-4421 inhibitor onERP29expression, and its target genes, were quantified by qPCR.

Results

ERP29silencing especially decreased necrotic cell death and increased migration in CDDP-sensitive, treated, and resistant cells, and decreased E-cadherin immunoexpression in CDDP-sensitive three-dimensional-spheroids. During CDDP treatment,ERP29silencing increased cell proliferation. In CDDP-sensitive cells,ERP29silencing increased the expression of several genes involved in WNT, MAPK, and PI3K/AKT pathways and decreasedCASP9expression. During CDDP treatment,ERP29silencing decreasedMDM2andCASP9expression. In CDDP-resistant cells,ERP29silencing increasedSOS1,MAPK1,AKT1,ITGAV, andCCNE1; and decreasedKRAS,JUN,MDM2, andCASP9expression. In addition, miR-4421 inhibition increasedERP29expression and decreasedMAPK1,AKT1,andJUNexpression in CDDP-sensitive cells; andSOS1,MAPK1,AKT1,andITGAVin CDDP-resistant cells, suggesting a potential therapeutic use for miR-4421 inhibitor.

Conclusions

ERP29silencing seems to decrease necrosis and increase migration of PSCC cells by modulating genes enrolled in WNT, MAPK, and PI3K/AKT pathways. Once validated, our data may enable target therapy development based on ensuringERP29expression that could benefit patients with CDDP-sensitive and resistant tumors.
Learning beyond-pairwise interactions enables the bottom-up prediction of microbial community structure
Authors: Ishizawa Hidehiro , Tashiro Yosuke , Inoue Daisuke , Ike Michihiko , Futamata Hiroyuki
Published: July 2023

The way to deal with higher-order effects (i.e., modification of pairwise interactions by third-party species) has been a major consideration in community ecology. Ignoring these effects is not in line with reality, yet fully considering them make the situation overly complex. Here, we propose a simple framework incorporating higher-order effects into a bottom-up community modeling, and assessed its validity using a seven-member synthetic bacterial community on a host plant, duckweed. Our findings revealed that actual interspecies interactions in community could not be predicted from pairwise co-culturing results; however, using information from trio combinations allowed for acceptable prediction. In addition, inclusion of four-, five-, and six-member combinations did not markedly enhance the prediction accuracy from trio-based prediction, suggesting that trio combinations, the smallest unit of higher-order effects, provide a reasonable baseline to unravel complex interaction networks. Building on this finding, we developed a prediction rule to estimate the structure of 4 – 7 member communities based on information from ≤ 3-member combinations, which yielded significantly better accuracy (relative mean square percentage errors of 22.7% – 61.2%) than pairwise-based model (53.5% – 185.2%). This highlights the possibility of establishing a quantitative link between the interspecies interactions and community structure, by observing beyond-pairwise combinations.
Dearth of smoking-induced mutations in NSRO-driven non-small-cell lung cancer despite smoking exposure
Authors: Huang Chen-Yang , Jiang Nanhai , Shen Meixin , Lai Gillianne , Tan Aaron C. , Jain Amit , Saw Stephanie P. , Ang Mei-Kim , Ng Quan Sing , Lim Darren Wan-Teck , Kanesvaran Ravindran , Tan Eng-Huat , Tan Wan Ling , Ong Boon-Hean , Chua Kevin L. , Anantham Devanand , Takano Angela , Lim Tony K.H. , Tam Wai Leong , Sim Ngak Leng , Skanderup Anders J. , Tan Daniel S.W. , Rozen Steven G.
Published: July 2023

Non-small cell lung cancers (NSCLCs) in non-smokers are mostly driven by mutations in the oncogenesEGFR, ERBB2,andMET, and fusions involvingALKandRET. We term these “non-smoking-related oncogenes” (NSROs). In addition to occurring in non-smokers, NSRO-driven tumors also occur in smokers, and the clonal architecture and genomic landscape of these tumors remain unknown. We investigated genomic and transcriptomic alterations in 173 tumor sectors from 48 patients with NSRO-driven or typical-smoking NSCLCs. NSRO-driven NSCLCs in smokers and non-smokers have similar genomic landscapes. Surprisingly, even in patients with prominent smoking histories, the mutational signature caused by tobacco smoking was essentially absent in NSRO-driven NSCLCs. However, NSRO-driven NSCLCs in smokers had higher transcriptomic activities related to regulation of the cell cycle, suggesting that smoking still affects tumor phenotype independently of genomic alterations.

Statement of significance

This study highlights the lack of genomic scars caused by smoking in NSCLCs driven by non-smoking-related oncogenes regardless of smoking history. The impact of smoking on these tumors is mainly non-genomic. The transcriptomic features of NSCLCs associated with smoking may help in the development of therapeutic approaches.
Malaria resistance-related biological adaptation and complex evolutionary footprints of Tai-Kadai people inferred from 796 genomes
Authors: Duan Shuhan , Wang Mengge , Wang Zhiyong , Liu Yan , Jiang Xiucheng , Su Haoran , Cai Yan , Sun Qiuxia , Sun Yuntao , Li Xiangping , Chen Jing , Zhang Yijiu , Yan Jiangwei , Nie Shengjie , Hu Liping , Tang Renkuan , Yun Libing , Wang Chuan-Chao , Liu Chao , Yang Junbao , He Guanglin
Published: July 2023

Pathogen-host adaptative interaction and complex population demographical processes, including admixture, drift and Darwen selection, have considerably shaped the Neolithic-to-Modern Western Eurasian population structure and genetic susceptibility to modern human diseases. However, the genetic footprints of evolutionary events in East Asia keep unknown as the underrepresentation of genomic diversity and the design of large-scale population studies. We reported one aggregated database of genome-wide-SNP variations from 796 Tai-Kadai (TK) genomes, including Bouyei first reported here, to explore the genetic history, population structure and biological adaptative features of TK-speaking people from Southern China and Southeast Asia. We found geography-related population substructure among TK-speaking people using the state-of-the-art population genetic structure reconstruction techniques based on the allele frequency spectrum and haplotype-resolved phased fragments. We found that the Northern TK-speaking people from Guizhou harboured one TK-dominant ancestry maximised in Bouyei people, and the Southern one from Thailand obtained more influences from Southeast Asians and indigenous people. We reconstructed the fitted admixture models and demographic graphs, which showed that TK-speaking people received gene flow from ancient rice farmer-related lineages related to the Hmong-Mien and Austroasiatic people and Northern millet farmers associated with the Sino-Tibetan people. Biological adaptation focused on our identified unique TK lineages related to Bouyei showed many adaptive signatures conferring Malaria resistance and low-rate lipid metabolism. Further gene enrichment, the allele frequency distribution of derived alleles, and their correlation with the incidence of Malaria further confirmed thatCR1played an essential role in the resistance of Malaria in the ancient “Baiyue” tribes.
Mapping gut bacteria into functional niches reveals the ecological structure of human gut microbiomes
Authors: Anthamatten Laura , Rogalla von Bieberstein Philipp , Thabuis Clémentine , Menzi Carmen , Reichlin Markus , Meola Marco , Rodriguez Bertrand , Cordero Otto X. , Lacroix Christophe , de Wouters Tomas , Leventhal Gabriel E.
Published: July 2023

Microbiomes are an essential contributor to the metabolic activity in the human gastrointestinal tract. The fermentation of otherwise indigestible nutritional components like dietary fibers relies on a complex interplay of metabolic pathways that are distributed across the individual bacteria. Yet, which of the bacteria are responsible for which parts of the distributed metabolism and how they should be grouped together is insufficiently understood. Here, we present the NicheMap™, an approach to map the different bacterial taxa that make up the gut microbiome onto the different functional niches of microbial carbohydrate fermentation. Our approach usesin vitromeasurements of bacterial growth and metabolic activity to identify which bacterial taxa are responsible for which metabolic function in the relevant complex context of whole human fecal microbiomes. We identified ‘characteristic taxa’ selected for by a panel growth substrates representative of dietary components that are resistant to digestion by host enzymes. These characteristic taxa offer predictions of which bacteria are stimulated by the various components of human diet. We validated these predictions using microbiome data from a human nutritional supplementation study. We suggest a template of how bacterial taxonomic diversity is organized along the trophic cascade of intestinal carbohydrate fermentation. We anticipate that our results and our approach will provide a key contribution towards building a structure-function map for gut microbiomes. Having such a map on hand is an important step in moving the microbiome from a descriptive science to an interventional one.
Automated Purification of DNA Origami with SPRI Beads
Authors: Chau Chalmers , Mohanan Gayathri , Macaulay Iain , Actis Paolo , Wälti Christoph
Published: July 2023

DNA origami synthesis is a well-established technique and has been employed in various applications. The synthesised origami must be purified to eliminate the excess materials such as DNA oligos and other molecules. While several purification techniques are routinely used, they all have limitations, and none can be automated to simultaneously handle large numbers and quantities of samples. Here we introduce the use of solid-phase immobilisation (SPRI) beads as an easy-to-adopt, scalable, high-throughput and automation-compatible method to purify DNA origami. Not only can this method remove excess oligos and biomolecules with comparable yield to existing methods while maintaining high structural integrity of the origami, but it also allows an automated workflow to simultaneously purify large numbers of samples within a limited time. We envision that the SPRI beads purification approach will improve the scalability of DNA nanostructures synthesis both for research and commercial applications.
Fecal and skin microbiota of two rescued Mediterranean monk seal pups during rehabilitation
Authors: Dosi Aggeliki , Meziti Alexandra , Tounta Eleni , Koemtzopoulos Kimon , Komnenou Anastasia , Dendrinos Panagiotis , Kormas Konstantinos
Published: July 2023

The role of animal host-associated microbiomes is becoming more apparent and defined for wild animals, especially for the species under conservation strategies. This study investigated the succession of fecal and skin bacterial microbiota of two rescued female Mediterranean monk seal (Monachus monachus) pups for most of their rehabilitation period. Bacterial species richness and diversity was assessed by high-throughput sequencing of nine freshly collected fecal samples and four skin swabs per individual. Both the fecal and skin microbiota highly overlapped in their containing operational taxonomic units (OTUs) and abundance patterns. The fecal microbiota was separated in two distinct periods, and was dominated by OTUs related to theShigella,Streptococcus,Enterococcus,LactobacillusandEscherichiagenera in the first period, while in the second period the dominating genera were theClostridium, Blautia, Fusobacterium, Edwardsiellaand Bacteroides. The skin microbiota was highly similar between the two individuals in each sampling and were dominated byPsychrobacter-, Elizabethkingia-andBergeyella-related OTUs. The provided antibiotic treatment along with the provided probiotics and nutritional supplements, resulted in a major turnover of the bacterial microbiota with the potentially detrimental OTUs being eliminated towards the end of the rehabilitation period, prior to the release of the pups in the wild.
Yeast-based heterologous production of the Colletochlorin family of fungal secondary metabolites
Authors: Geistodt-Kiener Aude , Totozafy Jean Chrisologue , Goff Géraldine Le , Vergne Justine , Sakai Kaori , Ouazzani Jamal , Mouille Grégory , Viaud Muriel , O’Connell Richard J. , Dallery Jean-Félix
Published: July 2023

Transcriptomic studies have revealed that fungal pathogens of plants activate the expression of numerous biosynthetic gene clusters (BGC) exclusively when in presence of a living host plant. The identification and structural elucidation of the corresponding secondary metabolites remain challenging. Here we adapted a polycistronic vector for efficient, seamless and cost-effective cloning of biosynthetic genes using in vivo assembly (also called transformation-assisted recombination) directly in Escherichia coli followed by heterologous expression in Saccharomyces cerevisiae. Two vectors were generated with different auto-inducible yeast promoters and selection markers. The effectiveness of these vectors was validated with fluorescent proteins. As a proof-of-principle, we applied our approach to the Colletochlorin family of molecules. These polyketide secondary metabolites were known from the phytopathogenic fungus Colletotrichum higginsianum but had never been linked to their biosynthetic genes. Considering the requirement for an halogenase, and by applying comparative genomics, we identified a BGC putatively involved in the biosynthesis of Colletochlorins in C. higginsianum. Following the expression of those genes in S. cerevisiae, we could identify the presence of the precursor Orsellinic acid, Colletochlorins and their non-chlorinated counterparts, the Colletorins. In conclusion, the polycistronic vectors described herein were adapted for the host S. cerevisiae and allowed to link the Colletochlorin compound family to their corresponding biosynthetic genes. This system will now enable the production and purification of infection-specific secondary metabolites of fungal phytopathogens. More widely, this system could be applied to any fungal BGC of interest.
Machine-learning analysis of factors that shape cancer aneuploidy landscapes reveals an important role for negative selection
Authors: Jubran Juman , Slutsky Rachel , Rozenblum Nir , Rokach Lior , Ben-David Uri , Yeger-Lotem Esti
Published: July 2023

Aneuploidy, an abnormal number of chromosomes within a cell, is considered a hallmark of cancer. Patterns of aneuploidy differ across cancers, yet are similar in cancers affecting closely-related tissues. The selection pressures underlying aneuploidy patterns are not fully understood, hindering our understanding of cancer development and progression. Here, we applied interpretable machine learning (ML) methods to study tissue-selective aneuploidy patterns. We defined 20 types of features of normal and cancer tissues, and used them to model gains and losses of chromosome-arms in 24 cancer types. In order to reveal the factors that shape the tissue-specific cancer aneuploidy landscapes, we interpreted the ML models by estimating the relative contribution of each feature to the models. While confirming known drivers of positive selection, our quantitative analysis highlighted the importance of negative selection for shaping the aneuploidy landscapes of human cancer. Tumor-suppressor gene density was a better predictor of gain patterns than oncogene density, and vice-versa for loss patterns. We identified the contribution of tissue-selective features and demonstrated them experimentally for chr13q gain in colon cancer. In line with an important role for negative selection in shaping the aneuploidy landscapes, we found compensation by paralogs to be a top predictor of chromosome-arm loss prevalence, and demonstrated this relationship for one such paralog interaction. Similar factors were found to shape aneuploidy patterns in human cancer cell lines, demonstrating their relevance for aneuploidy research. Overall, our quantitative, interpretable ML models improve the understanding of the genomic properties that shape cancer aneuploidy landscapes.
Molecular insights into atypical modes of β-arrestin interaction with seven transmembrane receptors
Authors: Maharana Jagannath , Sano Fumiya K. , Sarma Parishmita , Yadav Manish K. , Duan Longhan , Stepniewski Tomasz M. , Chaturvedi Madhu , Ranjan Ashutosh , Singh Vinay , Saha Sayantan , Mahajan Gargi , Chami Mohamed , Shihoya Wataru , Selent Jana , Chung Ka Young , Banerjee Ramanuj , Nureki Osamu , Shukla Arun K.
Published: July 2023

β-arrestins are multifunctional proteins that are critically involved in regulating spatio-temporal aspects of GPCR signaling. The interaction of β-arrestins with GPCRs is typically conceptualized in terms of receptor activation and phosphorylation primarily in the carboxyl-terminus. Interestingly however, there are several GPCRs that harbor majority of phosphorylation sites in their 3rdintracellular loop (ICL3) instead of carboxyl-terminus but still robustly engage β-arrestins. Moreover, there are several 7TMRs that are now characterized as intrinsically-biased, β-arrestin-coupled receptors (ACRs) due to lack of functional G-protein-coupling but robust β-arrestin binding leading to functional outcomes. The molecular basis of β-arrestin interaction and activation upon binding to these types of 7TMRs is currently elusive, and it represents a major knowledge gap in our current understanding of this signaling system. Here, we present seven cryo-EM structures of β-arrestins in basal state, activated by the muscarinic M2 receptor (M2R) through its ICL3, and a β-arrestin-coupled receptor known as decoy D6 receptor (D6R). These structural snapshots combined with biochemical, cellular, and biophysical experiments including HDX-MS and MD simulation provide novel insights into the ability of β-arrestins to preferentially select specific phosphorylation patterns in the receptors, and also illuminate the structural diversity in 7TMR-β-arrestin interaction. Surprisingly, we also observe that the carboxyl-terminus of β-arrestin2 but not β-arrestin1 undergoes structural transition from a β-strand to α-helix upon activation by D6R, which may preclude the core-interaction with the activated receptor. Taken together, our study elucidates previously unappreciated aspects of 7TMR-β-arrestin interaction, and provides important mechanistic clues about how the two isoforms of β-arrestins can recognize and regulate a large repertoire of GPCRs.
Reverse engineering the anti-MUC1 hybridoma antibody 139H2 by mass spectrometry-basedde novosequencing
Authors: Peng Weiwei , Giesbers Koen C.A.P. , Šiborová Marta , Beugelink J. Wouter , Pronker Matti F. , Schulte Douwe , Hilkens John , Janssen Bert J.C. , Strijbis Karin , Snijder Joost
Published: July 2023

Mucin 1 (MUC1) is a transmembrane mucin expressed at the apical surface of epithelial cells at different mucosal surfaces including breast and intestine. In the gastrointestinal tract, MUC1 has a barrier function against bacterial invasion, but can also serve as an entry receptor for pathogenicSalmonellabacteria. Moreover, MUC1 is well known for its aberrant expression and glycosylation in adenocarcinomas The MUC1 extracellular domain contains a variable number of tandem repeats (VNTR) of 20 amino acids, which are heavilyO-linked glycosylated.. Monoclonal antibodies against the MUC1 VNTR can be powerful tools because of their multiplicity of binding and possible applications in the diagnosis and treatment of MUC1-expressing cancers. One such antibody is the hybridoma mouse monoclonal 139H2, which is also widely used as a research tool to study non-cancer MUC1. Here we report direct mass spectrometry-based sequencing of hybridoma-derived 139H2 IgG, which enabled reverse engineering of a recombinant 139H2. The performance of the reverse engineered 139H2 IgG and its Fab fragment were validated by comparison to the hybridoma-derived product in Western blot and immunofluorescence microscopy. The reverse engineering of 139H2 allowed us to characterize binding to the VNTR peptide epitope by surface plasmon resonance (SPR) and solve the crystal structure of the 139H2 Fab fragment in complex with the MUC1 VNTR peptide. These analyses reveal the molecular basis for 139H2 binding specificity to MUC1 and its tolerance toO-glycosylation of the VNTR. The available sequence of 139H2 will allow further development of MUC1-related diagnostics, targeting and treatment strategies.
Metal-induced energy transfer (MIET) imaging of cell surface engineering with multivalent DNA nanobrushes
Authors: Wang Dong-Xia , Liu Bo , Han Gui-Mei , Li Qing-Nan , Kong De-Ming , Enderlein Jörg , Chen Tao
Published: July 2023

The spacing between cells has a significant impact on cell-cell interactions, which are critical to the fate and function of both individual cells and multicellular organisms. However, accurately measuring the distance between cell membranes and the variations between different membranes has proven to be a challenging task. In this study, we employ metal-induced energy transfer (MIET) imaging/spectroscopy to determine and track the inter-membrane distance and variations with nanometer precision. We have developed a DNA-based molecular adhesive called the DNA nanobrush, which serves as a cellular adhesive for connecting plasma membranes of different cells. By manipulating the number of base pairs within the DNA nanobrush, we can modify various aspects of cell-cell interactions, such as adhesive directionality, distance, and forces. We demonstrate that such nanometer-level changes can be detected with the MIET imaging/spectroscopy. Moreover, we successfully employ MIET to measure distance variations between a cellular plasma membrane and a model membrane. This experiment does not only showcase the effectiveness of MIET as a powerful tool for accurately quantifying cell-cell interactions, but does also validate the potential of DNA nanobrushes as cellular adhesives. This innovative method holds significant implications for advancing the study of multicellular interactions.
Survival and Developmental Progression of Unselected Thymocytes in the Absence of the T Cell Adaptor Gads
Authors: Shalah Rose , Marzouk Manal , Hallumi Enas , Klopstock Naama , Yablonski Deborah
Published: July 2023

Thymic development is tightly regulated by TCR signaling via the immune adaptors SLP-76 and LAT. Gads bridges the TCR-induced recruitment of SLP-76 to LAT; yet is not absolutely required for thymic progression. To better identify Gads-dependent developmental transitions, we performed tamoxifen-induced ablation of Gads (GadsiKO), accompanied by expression of tdTomato, and compared the development of Gads-expressing (Tom−) and - ablated (Tom+) thymocytes within the same mouse. The frequency of GadsiKO(Tom+) thymocytes decreased at β-selection and positive selection, confirming the Gads-dependence of these junctures; nevertheless, we observed small populations of positively-selected GadsiKOSP thymocytes. Consistent with a signaling defect, expression of CD5 was strongly impaired at the β-selection checkpoint and within the DP compartment; moreover, GadsiKODP thymocytes exhibited reduced TCR-induced calcium flux. Surprisingly, MHC-non-responding (CD5−) GadsiKODP thymocytes exhibited reduced death by neglect; instead, aberrant populations of CD5−GadsiKOthymocytes progressed as far as the CD4 SP compartment, while lacking key characteristics of positively selected thymocytes. In an experimentally-induced model of death by neglect triggered by CD8 crosslinking, Gads was required for the preferential apoptosis of CD5loDP thymocytes. Our results suggest that Gads promotes passage through TCR-driven developmental checkpoints while also promoting the death by neglect of unselected thymocytes.
It’s a Trap?! Escape from an ancient, ancestral sex chromosome system and implication ofFoxl2as the putative primary sex determining gene in a lizard (Anguimorpha; Shinisauridae)
Authors: Pinto Brendan J. , Nielsen Stuart V. , Sullivan Kathryn A. , Behere Ashmika , Keating Shannon E. , van Schingen-Khan Mona , Nguyen Truong Quang , Ziegler Thomas , Pramuk Jennifer , Wilson Melissa A. , Gamble Tony
Published: July 2023

Although sex determination is ubiquitous in vertebrates, mechanisms of sex determination vary from environmentally-to genetically-influenced. In vertebrates, genetic sex determination is typically accomplished with sex chromosomes. Groups like mammals maintain conserved sex chromosome systems, while sex chromosomes in most vertebrate clades aren’t conserved across similar evolutionary timescales. One group inferred to have an evolutionarily stable mode of sex determination is Anguimorpha, a clade of charismatic taxa including: monitor lizards, Gila monsters, and crocodile lizards. The common ancestor of extant anguimorphs possessed a ZW system that has been retained across the clade. However, the sex chromosome system in the endangered, monotypic family of crocodile lizards (Shinisauridae) has remained elusive. Here, we analyze genomic data to demonstrate thatShinisaurushas replaced the ancestral anguimorph ZW system on LG7 chromosome with a novel ZW system on LG3. The linkage group LG3 corresponds to chromosome 9 in chicken, and this is the first documented use of this syntenic block as a sex chromosome in amniotes. Additionally, this ∼1Mb region harbors approximately 10 genes, including a duplication of the sex-determining transcription factor,Foxl2—critical for the determination and maintenance of sexual differentiation in vertebrates, and thus a putative primary sex determining gene forShinisaurus.
Selective Inhibition of Interleukin-2 Inducible T Cell Kinase (ITK) Enhances Anti-Tumor Immunity in Association with Th1-skewing, Cytotoxic T cell Activation, and Reduced T Cell Exhaustion
Authors: Hsu Lih-Yun , Rosenbaum James T , Verner Erik , Jones William B , Hill Craig M. , Janc James W. , Buggy Joseph J. , Ding Ning , Reneau John C. , Khodadoust Michael S. , Kim Youn H. , Wilcox Ryan A. , Miller Richard A.
Published: July 2023

ITK is a tyrosine kinase expressed predominantly by T lymphocytes. In mice, selective knock-out of the ITK gene produces Th1 skewing of T helper cell differentiation. We synthesized a covalent ITK inhibitor, soquelitinib, that binds ITK with greater than 100-fold selectivity compared to binding to resting lymphocyte kinase (RLK).In vitrostudies with normal or malignant T cells demonstrated that soquelitinib suppresses Th2 cytokine production preferentially with relative sparing of Th1 cytokines. Soquelitinib inhibits thein vivogrowth of several syngeneic murine tumors including those that do not express ITK. Treatment with soquelitinib leads to increased tumor infiltration of normal CD8+ cells that possess enhanced T effector function. Soquelitinib inhibited expression of T cell exhaustion markers and was able to restore T effector function to exhausted cells. Pharmacologic selective ITK inhibition may represent a novel approach to cancer immunotherapy.