Most freshwater bacterial communities are characterized by a few dominant taxa that are often ubiquitous across freshwater biomes worldwide. Our understanding of the genomic diversity within these taxonomic groups is limited to a subset of taxa. Here, we investigated the genomic diversity that enables Limnohabitans, a freshwater genus key in funneling carbon from primary producers to higher trophic levels, to achieve abundance and ubiquity. We reconstructed eight putative Limnohabitans metagenome-assembled genomes (MAGs) from stations located along broad environmental gradients existing in Lake Michigan, part of Earth’s largest surface freshwater system. De novo strain inference analysis resolved a total of 23 strains from these MAGs, which strongly partitioned into two habitat-specific clusters with cooccurring strains from different lineages. The largest number of strains belonged to the abundant LimB lineage, for which robust in situ strain delineation had not previously been achieved. Our data show that temperature and nutrient levels may be important environmental parameters associated with microdiversification within the Limnohabitans genus. In addition, strains predominant in low- and high-phosphorus conditions had larger genomic divergence than strains abundant under different temperatures. Comparative genomics and gene expression analysis yielded evidence for the ability of LimB populations to exhibit cellular motility and chemotaxis, a phenotype not yet associated with available Limnohabitans isolates. Our findings broaden historical marker gene-based surveys of Limnohabitans microdiversification and provide in situ evidence of genome diversity and its functional implications across freshwater gradients.

IMPORTANCE Limnohabitans is an important bacterial taxonomic group for cycling carbon in freshwater ecosystems worldwide. Here, we examined the genomic diversity of different Limnohabitans lineages. We focused on the LimB lineage of this genus, which is globally distributed and often abundant, and its abundance has shown to be largely invariant to environmental change. Our data show that the LimB lineage is actually comprised of multiple cooccurring populations for which the composition and genomic characteristics are associated with variations in temperature and nutrient levels. The gene expression profiles of this lineage suggest the importance of chemotaxis and motility, traits that had not yet been associated with the Limnohabitans genus, in adapting to environmental conditions.

Background:

Care services have not been sufficiently adapted to meet the comprehensive care needs of women living with HIV. Our study objective was to engage patients and providers in codesigning care recommendations to improve care for this population in the province of Quebec.

Background:

Patients with multimorbidity often require services across different health care settings, yet team processes among settings are rarely implemented. We explored perceptions of specialists and family physicians collaborating in a telemedicine interprofessional consultation for patients with multimorbidity to better understand the value of bringing physicians together across the boundaries of health care settings.

A 25-year-old male is admitted with complaints of dry cough for the past 5 years, and increased thirst, urinary frequency and output for the past 18 months. He also complains of shortness of breath on climbing a flight of stairs, and itchy lesions on the scalp and back for the past 2–3 months. There is no history of bone pain or abdominal pain. He has history of bilateral recurrent pneumothoraxes, twice on the right and once on the left side, in the past month. Pleurodesis with povidone iodine is performed on left side and the patient is transferred to your hospital with persistent right pneumothorax with air leak, with an intercostal drainage tube in situ. The patient is a never-smoker with no family history of pneumothorax. On general examination, he has small papules, 1–2 mm in diameter, with scaling over scalp and back. Onycholysis, onychoschisis and subungual splinter haemorrhages are present (figure 1).

Obtaining blood and cerebrospinal fluid is generally less invasive than standard tumor biopsy, and are increasingly used to develop surrogate biomarkers. Leptomeningeal disease, a devastating complication of cancer, represents a unique opportunity for using liquid biopsies for diagnosis, treatment, and to elucidate underlying mechanisms of resistance to therapy.

See related article by Smalley et al., p. 2163

In this retrospective study, whole-genome sequencing (WGS) data generated on an Ion Torrent platform was used to predict phenotypic drug resistance profiles for first- and second-line drugs among Swedish clinical Mycobacterium tuberculosis isolates from 2016 to 2018. The accuracy was ~99% for all first-line drugs and 100% for four second-line drugs. Our analysis supports the introduction of WGS into routine diagnostics, which might, at least in Sweden, replace phenotypic drug susceptibility testing in the future.

Quinolones, such as the antimalarial atovaquone, are inhibitors of the malarial mitochondrial cytochrome bc₁ complex, a target critical to the survival of both liver- and blood-stage parasites, making these drugs useful as both prophylaxis and treatment. Recently, several derivatives of endochin have been optimized to produce novel quinolones that are active in vitro and in animal models. While these quinolones exhibit potent ex vivo activity against Plasmodium falciparum and Plasmodium vivax, their activity against the zoonotic agent Plasmodium knowlesi is unknown. We screened several of these novel endochin-like quinolones (ELQs) for their activity against P. knowlesi in vitro and compared this with their activity against P. falciparum tested under identical conditions. We demonstrated that ELQs are potent against P. knowlesi (50% effective concentration, <117 nM) and equally effective against P. falciparum. We then screened selected quinolones and partner drugs using a longer exposure (2.5 life cycles) and found that proguanil is 10-fold less potent against P. knowlesi than P. falciparum, while the quinolones demonstrate similar potency. Finally, we used isobologram analysis to compare combinations of the ELQs with either proguanil or atovaquone. We show that all quinolone combinations with proguanil are synergistic against P. falciparum. However, against P. knowlesi, no evidence of synergy between proguanil and the quinolones was found. Importantly, the combination of the novel quinolone ELQ-300 with atovaquone was synergistic against both species. Our data identify potentially important species differences in proguanil susceptibility and in the interaction of proguanil with quinolones and support the ongoing development of novel quinolones as potent antimalarials that target multiple species.

Staphylococcus aureus biofilms are a significant problem in health care settings, partly due to the presence of a nondividing, antibiotic-tolerant subpopulation. Here we evaluated treatment of S. aureus UAMS-1 biofilms with HT61, a quinoline derivative shown to be effective against nondividing Staphylococcus spp. HT61 was effective at reducing biofilm viability and was associated with increased expression of cell wall stress and division proteins, confirming its potential as a treatment for S. aureus biofilm infections.

We characterized 29 bla_CTX-M-27-harboring plasmids of Escherichia coli sequence type 131 (ST131) sublineage C1/H30R isolates from healthy individuals and long-term-care facility (LTCF) residents. Most (27/29) plasmids were of the FIA, FIB, and FII multireplicon type with the same plasmid multilocus sequence typing (pMLST). Several plasmids (7/23) from LTCF residents harbored only bla_CTX-M-27 as the resistance gene; however, their fundamental structures were very similar to those of previously isolated bla_CTX-M-27/F1:A2:B20 plasmids, suggesting their prevalence as a newly arising public health concern.

Fluoroquinolones are reported to possess immunomodulatory activity; hence, a novel benzoquinolizine fluoroquinolone, levonadifloxacin, was evaluated in lipopolysaccharide-stimulated human whole-blood (HWB) and mouse acute lung injury (ALI) models. Levonadifloxacin significantly mitigated the inflammatory responses in an HWB assay through inhibition of proinflammatory cytokines and in the ALI model by lowering lung total white blood cell count, myeloperoxidase, and cytokine levels. The immunomodulatory effect of levonadifloxacin, along with promising antibacterial activity, is expected to provide clinical benefits in the treatment of infections.

Insufficient reactivity against cells with low antigen density has emerged as an important cause of chimeric antigen receptor (CAR) T-cell resistance. Little is known about factors that modulate the threshold for antigen recognition. We demonstrate that CD19 CAR activity is dependent upon antigen density and that the CAR construct in axicabtagene ciloleucel (CD19-CD28) outperforms that in tisagenlecleucel (CD19-4-1BB) against antigen-low tumors. Enhancing signal strength by including additional immunoreceptor tyrosine-based activation motifs (ITAM) in the CAR enables recognition of low-antigen-density cells, whereas ITAM deletions blunt signal and increase the antigen density threshold. Furthermore, replacement of the CD8 hinge-transmembrane (H/T) region of a 4-1BB CAR with a CD28-H/T lowers the threshold for CAR reactivity despite identical signaling molecules. CARs incorporating a CD28-H/T demonstrate a more stable and efficient immunologic synapse. Precise design of CARs can tune the threshold for antigen recognition and endow 4-1BB-CARs with enhanced capacity to recognize antigen-low targets while retaining a superior capacity for persistence.

Unlike replicative senescence, oncogene-induced senescence caused heterochromatin-body formation.

Thermo Fisher Scientific announced plans in March to acquire Qiagen in a $11.5 billion deal that could bring morediagnostic offeringsand sample-preparation technologies to one of the world's leading manufacturers of scientific instruments, research services, and laboratory consumables.

A new method called MicroMapping can identify nanoscale protein–protein interactions on live cells.

Monoubiquitinated FANCI and FANCD2 constitute the ID complex, which forms a sliding clamp on DNA.

Genomic analysis of a patient's tumor is the cornerstone of precision oncology, but it does not address whether metastases should be treated differently. Here we tested whether comparative single-cell RNA sequencing (scRNA-seq) of a primary small intestinal neuroendocrine tumor to a matched liver metastasis could guide the treatment of a patient's metastatic disease. Following surgery, the patient was put on maintenance treatment with a somatostatin analog. However, the scRNA-seq analysis revealed that the neuroendocrine epithelial cells in the liver metastasis were less differentiated and expressed relatively little SSTR2, the predominant somatostatin receptor. There were also differences in the tumor microenvironments. RNA expression of vascular endothelial growth factors was higher in the primary tumor cells, reflected by an increased number of endothelial cells. Interestingly, vascular expression of the major VEGF receptors was considerably higher in the liver metastasis, indicating that the metastatic vasculature may be primed for expansion and susceptible to treatment with angiogenesis inhibitors. The patient eventually progressed on Sandostatin, and although consideration was given to adding an angiogenesis inhibitor to her regimen, her disease progression involved non-liver metastases that had not been characterized. Although in this specific case comparative scRNA-seq did not alter treatment, its potential to help guide therapy of metastatic disease was clearly demonstrated.

Gastric cancer (GC) is the fifth-ranked cancer type by associated mortality. The proportion of early diagnosis is low, and most patients are diagnosed at the advanced stages. First-line therapy standardly includes fluoropyrimidines and platinum compounds with trastuzumab for HER2-positive cases. For recurrent disease, there are several alternative options including ramucirumab, a monoclonal therapeutic antibody that inhibits VEGF-mediated tumor angiogenesis by binding with VEGFR2, alone or in combination with other cancer drugs. However, overall response rate following ramucirumab or its combinations is 30%–80% of the patients, suggesting that personalization of drug prescription is needed to increase efficacy of treatment. We report here original tumor RNA sequencing profiles for 15 advanced GC patients linked with data on clinical response to ramucirumab or its combinations. Three genes showed differential expression in the tumors for responders versus nonresponders: CHRM3, LRFN1, and TEX15. Of them, CHRM3 was up-regulated in the responders. Using the bioinformatic platform Oncobox we simulated ramucirumab efficiency and compared output model results with actual tumor response data. An agreement was observed between predicted and real clinical outcomes (AUC ≥ 0.7). These results suggest that RNA sequencing may be used to personalize the prescription of ramucirumab for GC and indicate potential molecular mechanisms underlying ramucirumab resistance. The RNA sequencing profiles obtained here are fully compatible with the previously published Oncobox Atlas of Normal Tissue Expression (ANTE) data.

An 18-yr-old man with a history of intellectual disability, craniofacial dysmorphism, seizure disorder, and obesity was identified to carry a de novo, pathogenic variant in ASXL1 (c.4198G>T; p.E1400X) associated with the diagnosis of Bohring–Opitz syndrome based on exome sequencing. In addition, he was identified to carry a maternally inherited and likely pathogenic variant in MC4R (c.817C>T; p.Q273X) associated with monogenic obesity. Dual genetic diagnosis occurs in 4%–6% of patients and results in unique clinical phenotypes that are a function of tissue-specific gene expression, involved pathways, clinical expressivity, and penetrance. This case highlights the utility of next-generation sequencing in patients with an unusual combination of clinical presentations for several pillars of precision medicine including (1) diagnosis, (2) prognosis and outcome, (3) management and therapy, and (4) utilization of resources.

The T1D Exchange established a learning platform by evaluating the current state of care and engaging 10 diabetes clinics in collaborative quality improvement (QI) activities. Participating clinics are sharing data and best practices to improve care delivery for people with type 1 diabetes. This article describes the design and initial implementation of this platform, known as the T1D Exchange Quality Improvement Collaborative. This effort has laid a foundation for learning from variation in type 1 diabetes care delivery via QI methodology and has demonstrated success in improving processes through iterative testing cycles and transparent sharing of data.

Background:

Limited treatment options are available for oral mucositis, a common, debilitating complication of cancer therapy. We examined the association between daily delivery time of radiotherapy and the severity of oral mucositis in patients with head and neck cancer.

Adeno-associated virus (AAV) recombinants are currently the vector of choice for many gene therapy applications. As experimental therapies progress to clinical trials, the need to characterize recombinant adeno-associated viruses (rAAVs) accurately and reproducibly increases. Accurate determination of rAAV infectious titer is important for determining the activity of each vector lot and for ensuring lot-to-lot consistency. The following protocol developed in our laboratory uses a 96-well TCID₅₀ format and quantitative polymerase chain reaction (qPCR) detection for the determination of rAAV infectious titer.

This protocol is used to determine the concentration of DNase-resistant vector genomes (i.e., packaged in the capsid) in purified recombinant adeno-associated virus (rAAV) preparations. The protocol begins with treatment of the vector stock with DNase I to eliminate unencapsidated AAV DNA or contaminating plasmid DNA. This is followed by a heat treatment to heat-inactivate DNase I, to disrupt the viral capsid, and to release the packaged vector genomes for quantification by real-time polymerase chain reaction (PCR) using a set of standards (linearized plasmid used for vector production) containing known copy numbers. To accomplish high-throughput titration, the primer and probe sets used in real-time PCR are usually designed to target common elements present in most rAAV genomes, such as promoters and poly(A) signals. This strategy significantly reduces the number of PCRs, controls, and turnaround time. Several important controls should be included in the assay as follows: The first two controls should have a known copy number of the rAAV genome plasmid treated or not treated with DNase I. This control tests the effectiveness of DNase treatment. To control for potential cross-contamination between samples during the preparation process, a blank control containing nuclease-free water only should be processed and tested in parallel. A validation vector sample with a known titer should be included in every assay to monitor interassay variability. Finally, for the PCR run, a no-template control (NTC) is included to indicate cross-contamination during PCR setup.

RING-finger E3 ligases are instrumental in the regulation of inflammatory cascades, apoptosis, and cancer. However, their roles are relatively unknown in TGFβ/SMAD signaling. SMAD3 and its adaptors, such as β2SP, are important mediators of TGFβ signaling and regulate gene expression to suppress stem cell–like phenotypes in diverse cancers, including hepatocellular carcinoma (HCC). Here, PJA1, an E3 ligase, promoted ubiquitination and degradation of phosphorylated SMAD3 and impaired a SMAD3/β2SP-dependent tumor-suppressing pathway in multiple HCC cell lines. In mice deficient for SMAD3 (Smad3+/−), PJA1 overexpression promoted the transformation of liver stem cells. Analysis of genes regulated by PJA1 knockdown and TGFβ1 signaling revealed 1,584 co-upregulated genes and 1,280 co-downregulated genes, including many implicated in cancer. The E3 ligase inhibitor RTA405 enhanced SMAD3-regulated gene expression and reduced growth of HCC cells in culture and xenografts of HCC tumors, suggesting that inhibition of PJA1 may be beneficial in treating HCC or preventing HCC development in at-risk patients.Significance: These findings provide a novel mechanism regulating the tumor suppressor function of TGFβ in liver carcinogenesis.

Hepatitis C virus (HCV) is an important and underreported infectious disease, causing chronic infection in ~71 million people worldwide. The limited host range of HCV, which robustly infects only humans and chimpanzees, has made studying this virus in vivo challenging and hampered the development of a desperately needed vaccine. The restrictions and ethical concerns surrounding biomedical research in chimpanzees has made the search for an animal model all the more important. In this review, we discuss different approaches that are being pursued toward creating small animal models for HCV infection. Although efforts to use a nonhuman primate species besides chimpanzees have proven challenging, important advances have been achieved in a variety of humanized mouse models. However, such models still fall short of the overarching goal to have an immunocompetent, inheritably susceptible in vivo platform in which the immunopathology of HCV could be studied and putative vaccines development. Alternatives to overcome this include virus adaptation, such as murine-tropic HCV strains, or the use of related hepaciviruses, of which many have been recently identified. Of the latter, the rodent/rat hepacivirus from Rattus norvegicus species-1 (RHV-rn1) holds promise as a surrogate virus in fully immunocompetent rats that can inform our understanding of the interaction between the immune response and viral outcomes (i.e., clearance vs. persistence). However, further characterization of these animal models is necessary before their use for gaining new insights into the immunopathogenesis of HCV and for conceptualizing HCV vaccines.

BACKGROUND AND PURPOSE:

Endolymphatic hydrops in patients with Menière disease relies on delayed postcontrast 3D-FLAIR sequences. The purpose of this study was to compare the degree of perilymphatic enhancement and the detection rate of endolymphatic hydrops using constant and variable flip angles sequences.

BACKGROUND AND PURPOSE:

Conventional nonadhesive liquid embolic agents currently are the criterion standard for endovascular embolization of cerebral AVMs. However, inadequate distal penetration into the nidus and unstable proximal plug formation are the major limitations of this approach and of the currently available embolic materials. The aim of this study was to evaluate the hypothetic efficacy of combining liquid embolic agents with different properties and viscosities for use in endovascular embolization of cerebral AVMs.

BACKGROUND AND PURPOSE:

NeuroQuant is an FDA-approved software that performs automated MR imaging quantitative volumetric analysis. This study aimed to compare the accuracy of NeuroQuant analysis with visual MR imaging analysis by neuroradiologists with expertise in epilepsy in identifying hippocampal sclerosis.