Search

Outlines the introduction of audio-tutorial (A-T) methods of teaching a general biology course, including facility planning, costs, and problems encountered. Presents data indicating that good students achieved higher grades with A-T than with conventional courses, but lower achievers performed at about the same level with either method. (AL)

Biomedical science is rapidly developing in terms of more transparency, openness and reproducibility of scientific publications. This is even more important for all studies that are based on results from basic semen examination. Recently two concordant documents have been published: the 6th edition of the WHO Laboratory Manual for the Examination and Processing of Human Semen, and the International Standard ISO 23162:2021. With these tools, we propose that authors should be instructed to follow these laboratory methods in order to publish studies in peer-reviewed journals, preferable by using a checklist as suggested in an Appendix to this article.

Osteolytic lesions (OL) characterize symptomatic multiple myeloma. The mechanisms of how malignant plasma cells (PC) cause OL in one region while others show no signs of bone destruction despite subtotal infiltration remain unknown. We report on a single-cell RNA sequencing (scRNA-seq) study of PC obtained prospectively from random bone marrow aspirates (BM) and paired imaging-guided biopsies of OL. We analyze 148,630 PC from 24 different locations in 10 patients and observe vast inter- and intra-patient heterogeneity based on scRNA-seq analyses. Beyond the limited evidence for spatial heterogeneity from whole-exome sequencing, we find an additional layer of complexity by integrated analysis of anchored scRNA-seq datasets from the BM and OL. PC from OL are characterized by differentially expressed genes compared to PC from BM, including upregulation of genes associated with myeloma bone disease like DKK1, HGF and TIMP-1 as well as recurrent downregulation of JUN/FOS, DUSP1 and HBB. Assessment of PC from longitudinally collected samples reveals transcriptional changes after induction therapy. Our study contributes to the understanding of destructive myeloma bone disease.

The treatment landscape for advanced urothelial cancer has changed dramatically owing to the US Food and Drug Administration approval and introduction of antibody-drug conjugates (ADCs), including enfortumab vedotin and sacituzumab govitecan. Efforts have begun to use these therapies in earlier disease states, specifically bacillus Calmette-Guérin (BCG)-unresponsive non-muscle-invasive bladder cancer (NMIBC). We assessed gene expression associated with these newly approved therapies in a novel cohort of treatment-naïve NMIBC tumors before and after BCG therapy. Multiple genes, including Nectin-4, Trop-2, and Her-2, exhibited increased expression after BCG therapy compared to baseline. However, few of the tumors with increased expression of ADC targets also exhibited increased PD-L1/PD-1 expression. Taken together, these data demonstrate the heterogeneous genomic landscape of BCG-exposed NMIBC, and provide evidence supporting the evaluation of ADCs in NMIBC. PATIENT SUMMARY: We evaluated the potential role of targeted therapies that have been approved in the USA for advanced non-muscle-invasive bladder cancer (NMIBC) that has recurred after treatment with bacillus Calmette-Guérin (BCG). By assessing levels of specific genes and proteins linked to the targeted therapies, we demonstrate that there is rationale for further evaluation of these therapies in NMIBC.

Burkitt lymphoma (BL) is an aggressive lymphoma type that is currently treated by intensive chemoimmunotherapy. Despite the favorable clinical outcome for most patients with BL, chemotherapy-related toxicity and disease relapse remain major clinical challenges, emphasizing the need for innovative therapies. Using genome-scale CRISPR-Cas9 screens, we identified B-cell receptor (BCR) signaling, specific transcriptional regulators, and one-carbon metabolism as vulnerabilities in BL. We focused on serine hydroxymethyltransferase 2 (SHMT2), a key enzyme in one-carbon metabolism. Inhibition of SHMT2 by either knockdown or pharmacological compounds induced anti-BL effects in vitro and in vivo. Mechanistically, SHMT2 inhibition led to a significant reduction of intracellular glycine and formate levels, which inhibited the mTOR pathway and thereby triggered autophagic degradation of the oncogenic transcription factor TCF3. Consequently, this led to a collapse of tonic BCR signaling, which is controlled by TCF3 and is essential for BL cell survival. In terms of clinical translation, we also identified drugs such as methotrexate that synergized with SHMT inhibitors. Overall, our study has uncovered the dependency landscape in BL, identified and validated SHMT2 as a drug target, and revealed a mechanistic link between SHMT2 and the transcriptional master regulator TCF3, opening up new perspectives for innovative therapies.

Potyvirids (the family Potyviridae) are the largest family of plant RNA viruses. Two novel potyvirid viruses, Striga-associated poty-like virus 1 (SaPlV1) and Striga-associated poty-like virus 2 (SaPlV2), were identified from the transcriptome data of purple witchweed (Striga hermonthica). SaPlV1 was most closely related to bellflower veinal mottle virus (BVMoV), the only member of the genus Bevemovirus, and then to macluraviruses (the genus Macluravirus). The SaPlV1 genome encodes a 2462-amino acid (aa) polyprotein that may be cleaved into nine mature peptides. The cleavage sites of SaPlV1, BVMoV, and macluravirus polyproteins shared strong sequence similarities. SaPlV2 was most closely related to celery latent virus, the sole species of the genus Celavirus, which is the most divergent potyvirid genus. The SaPlV2 polyprotein contained 3329 aa and it may be cleaved into at least seven or eight mature peptides. Phylogenetic analysis suggested that SaPlV1 and SaPlV2 may be novel species of the genera Bevemovirus and Celavirus, respectively. The genome sequences of SaPlV1 and SaPlV2 are useful resources for studying the genome evolution of potyvirids. Keywords: Striga-associated poty-like virus 1; Striga-associated poty-like virus 2; Potyviridae; Beve- movirus; Celavirus; purple witchweed; Striga hermonthica.

The BRCA1 tumor suppressor gene encodes a multidomain protein for which several functions have been described. These include a key role in homologous recombination repair (HRR) of DNA double-strand breaks, which is shared with two other high-risk hereditary breast cancer suppressors, BRCA2 and PALB2. Although both BRCA1 and BRCA2 interact with PALB2, BRCA1 missense variants affecting its PALB2-interacting coiled-coil domain are considered variants of uncertain clinical significance (VUS). Using genetically engineered mice, we show here that a BRCA1 coiled-coil domain VUS, Brca1 p.L1363P, disrupts the interaction with PALB2 and leads to embryonic lethality. Brca1 p.L1363P led to a similar acceleration in the development of Trp53-deficient mammary tumors as Brca1 loss, but the tumors showed distinct histopathologic features, with more stable DNA copy number profiles in Brca1 p.L1363P tumors. Nevertheless, Brca1 p.L1363P mammary tumors were HRR incompetent and responsive to cisplatin and PARP inhibition. Overall, these results provide the first direct evidence that a BRCA1 missense variant outside of the RING and BRCT domains increases the risk of breast cancer. Significance: These findings reveal the importance of a patient-derived BRCA1 coiled-coil domain sequence variant in embryonic development, mammary tumor suppression, and therapy response. See related commentary by Mishra et al., p. 6080

In tuberous sclerosis, most cardiac rhabdomyomas regress spontaneously. In some cases, the tumors can cause life-threatening hemodynamic compromise requiring subsequent surgical resection. The mechanistic target of rapamycin inhibitors everolimus and sirolimus have shown to be effective treatments for multiple conditions. There are four reports of off-label treatment with transplacental sirolimus for fetal rhabdomyomas due to tuberous sclerosis complex. The optimal dosing regimen is unknown.We reviewed the medical records of all patients treated prenatally with sirolimus for rhabdomyomas. All fetuses had a clinical and molecular diagnosis of tuberous sclerosis complex (2012 Consensus Diagnostic Criteria, including a positive genetic test). Clinical history, mechanistic target of rapamycin inhibitor dosing and levels, outcome, and adverse events were reviewed after initiation of sirolimus treatment.Three fetuses were treated with maternal sirolimus. Dosing regimens and subsequent trough levels differed from 1 mg/day to 6 mg/day and1.0 ng/mL to 12.2 ng/mL. Cardiac rhabdomyomas gradually shrank in all patients. Growth restriction was noted in one patient. No severe adverse events occurred during the treatment period.Maternal sirolimus appears to be a safe treatment option in prenatally detected rhabdomyomas with possible need for intervention. Follow-up visits with fetal ultrasound, echocardiography, and laboratory work should be performed weekly during the treatment period. The optimal dosing and trough level timepoints remain unclear. Based on our results, we recommend a sirolimus starting dose of at least 2 mg/m

The ectoparasitic nematode Xiphinema index transmits grapevine fanleaf virus (GFLV) during feeding on grapevine roots, causing fanleaf degeneration in the plant. Hence, resistance breeding is a key to develop novel rootstocks to overcome such threats. In past years, various grapevine species were screened, and a few candidates with partial resistance were identified. However, they were hardly sufficient for viticulture because of their many agronomical defects. To develop reliably resistant rootstocks applicable in viticulture, multiple Vitis spp. genotypes were analyzed using root inoculation with nematodes in glass vials as an early and easy evaluation test. Resistance levels were evaluated 35 days after inoculation based on nematode reproduction factors, focusing on juveniles and eggs. Infection of grapevines with GFLV was analyzed after inoculation with viruliferous X. index. With this fast screening system, putative candidates with resistances against X. index have been identified for future breeding programs. Particularly, genotypes with the genetic background of Vitis aestivalis and Vitis labrusca were found to be nematode-resistant.

Understanding the relationship between migratory performance and fitness is crucial for predicting population dynamics of migratory species. In this study, we used geolocators to explore migration performance (speed and duration of migratory movements, migratory timings) and its association with breeding phenology and productivity in an Afro-Palearctic insectivore, the European bee-eater (Merops apiaster), breeding in Iberian Peninsula. Bee-eaters migrated at higher travel speeds and had shorter travel duration in spring compared to autumn. Individuals that departed earlier or spent fewer days in-flight arrived earlier to the breeding areas. Our results show overall positive, but year-specific, linkages between arrival and laying dates. In one year, laying was earlier and productivity was higher, remaining constant throughout the season, while in the subsequent year productivity was lower and, importantly, declined with laying date. These results suggest that arriving earlier can be advantageous for bee-eaters, as in years when breeding conditions are favourable, early and late breeders produce high and similar number of fledglings, but when conditions are unfavourable only early breeders experience high productivity levels.

Viral tropism within the brain and the role(s) of vertebrate immune response to neurotropic flaviviruses infection is largely understudied. We combine multimodal imaging (cm-nm scale) with single nuclei RNA-sequencing to study Langat virus in wildtype and interferon alpha/beta receptor knockout (Ifnar–/–) mice to visualize viral pathogenesis and define molecular mechanisms. Whole brain viral infection is imaged by Optical Projection Tomography coregistered to ex vivo MRI. Infection is limited to grey matter of sensory systems in wildtype mice, but extends into white matter, meninges and choroid plexus in Ifnar–/– mice. Cells in wildtype display strong type I and II IFN responses, likely due to Ifnb expressing astrocytes, infiltration of macrophages and Ifng-expressing CD8+ NK cells, whereas in Ifnar–/–, the absence of this response contributes to a shift in cellular tropism towards non-activated resident microglia. Multimodal imaging-transcriptomics exemplifies a powerful way to characterize mechanisms of viral pathogenesis and tropism.

Plasmodium species are frequently host-specific, but little is currently known about the molecular factors restricting host switching. This is particularly relevant for P. falciparum, the only known human-infective species of the Laverania sub-genus, all other members of which infect African apes. Here we show that all tested P. falciparum isolates contain an inactivating mutation in an erythrocyte invasion associated gene, PfEBA165, the homologues of which are intact in all ape-infective Laverania species. Recombinant EBA165 proteins only bind ape, not human, erythrocytes, and this specificity is due to differences in erythrocyte surface sialic acids. Correction of PfEBA165 inactivating mutations by genome editing yields viable parasites, but is associated with down regulation of both PfEBA165 and an adjacent invasion ligand, which suggests that PfEBA165 expression is incompatible with parasite growth in human erythrocytes. Pseudogenization of PfEBA165 may represent a key step in the emergence and evolution of P. falciparum., Here, Proto et al. show that human infective Plasmodium falciparum isolates contain an inactivating mutation in the erythrocyte invasion associated gene PfEBA165, while homologues of ape-infective Laverania species are intact, and that expression of intact PfEBA165 is incompatible with parasite growth in human erythrocytes.

Key message The phenomic predictive ability depends on the genetic architecture of the target trait, being high for complex traits and low for traits with major QTL. Abstract Genomic selection is a powerful tool to assist breeding of complex traits, but a limitation is the costs required for genotyping. Recently, phenomic selection has been suggested, which uses spectral data instead of molecular markers as predictors. It was shown to be competitive with genomic prediction, as it achieved predictive abilities as high or even higher than its genomic counterpart. The objective of this study was to evaluate the performance of phenomic prediction for triticale and the dependency of the predictive ability on the genetic architecture of the target trait. We found that for traits with a complex genetic architecture, like grain yield, phenomic prediction with NIRS data as predictors achieved high predictive abilities and performed better than genomic prediction. By contrast, for mono- or oligogenic traits, for example, yellow rust, marker-based approaches achieved high predictive abilities, while those of phenomic prediction were very low. Compared with molecular markers, the predictive ability obtained using NIRS data was more robust to varying degrees of genetic relatedness between the training and prediction set. Moreover, for grain yield, smaller training sets were required to achieve a similar predictive ability for phenomic prediction than for genomic prediction. In addition, our results illustrate the potential of using field-based spectral data for phenomic prediction. Overall, our result confirmed phenomic prediction as an efficient approach to improve the selection gain for complex traits in plant breeding.

Background: A significant subset of patients with Alzheimer’s disease (AD) exhibit low bone mineral density and are therefore more fracture-prone, relative to their similarly aged neurotypical counterparts. In addition to chronic immune hyperactivity, behavioral dysregulation of effector peripheral sympathetic neurons—which densely innervate bone and potently modulate bone remodeling—is implicated in this pathological bone reformation. Objective: Thus, there exists a pressing need for a robust in vitro model which allows interrogation of the paracrine interactions between the putative mediators of AD-related osteopenia: sympathetic neurons (SNs) and mesenchymal stem cells (MSCs). Methods: Toward this end, activated SN-like PC12 cells and bone marrow derived MSCs were cultured in poly(ethylene glycol) diacrylate (PEGDA) hydrogels in the presence or absence of the AD-relevant inflammatory cytokine tumor necrosis factor alpha (TNF-α) under mono- and co-culture conditions. Results: PC12s and MSCs exposed separately to TNF-α displayed increased expression of pro-inflammatory mediators and decreased osteopontin (OPN), respectively. These data indicate that TNF-α was capable of inducing a dysregulated state in both cell types consistent with AD. Co-culture of TNF-α-activated PC12s and MSCs further exacerbated pathological behaviors in both cell types. Specifically, PC12s displayed increased secretion of interleukin 6 relative to TNF-α stimulated monoculture controls. Similarly, MSCs demonstrated a further reduction in osteogenic capacity relative to TNF-α stimulated monoculture controls, as illustrated by a significant decrease in OPN and collagen type I alpha I chain. Conclusion: Taken together, these data may indicate that dysregulated sympathetic activity may contribute to AD-related bone loss.

It remains largely unclear how thymocytes translate relative differences in T cell receptor (TCR) signal strength into distinct developmental programs that drive the cell fate decisions towards conventional (Tconv) or regulatory T cells (Treg). Following TCR activation, intracellular calcium (Ca2+) is the most important second messenger, for which the potassium channel K2P18.1 is a relevant regulator. Here, we identify K2P18.1 as a central translator of the TCR signal into the thymus-derived Treg (tTreg) selection process. TCR signal was coupled to NF-κB-mediated K2P18.1 upregulation in tTreg progenitors. K2P18.1 provided the driving force for sustained Ca2+ influx that facilitated NF-κB- and NFAT-dependent expression of FoxP3, the master transcription factor for Treg development and function. Loss of K2P18.1 ion-current function induced a mild lymphoproliferative phenotype in mice, with reduced Treg numbers that led to aggravated experimental autoimmune encephalomyelitis, while a gain-of-function mutation in K2P18.1 resulted in increased Treg numbers in mice. Our findings in human thymus, recent thymic emigrants and multiple sclerosis patients with a dominant-negative missense K2P18.1 variant that is associated with poor clinical outcomes indicate that K2P18.1 also plays a role in human Treg development. Pharmacological modulation of K2P18.1 specifically modulated Treg numbers in vitro and in vivo. Finally, we identified nitroxoline as a K2P18.1 activator that led to rapid and reversible Treg increase in patients with urinary tract infections. Conclusively, our findings reveal how K2P18.1 translates TCR signals into thymic T cell fate decisions and Treg development, and provide a basis for the therapeutic utilization of Treg in several human disorders.

The ability to spread attention over items or locations is as important for everyday functioning as the ability to focus narrowly. Little is known about neuronal processes involved in broad monitoring, but indirect evidence suggests a role of nicotinic acetylcholine receptors (nAChRs). The present study tested whether the prototypical nAChR agonist nicotine enhances the ability of humans and rodents to maintain a broad attentional window. Fifty-three never-smokers wearing a nicotine (7 mg/24 h) or placebo patch performed an attention task requiring detection of stimuli presented randomly in one of four peripheral locations, with a central cue predicting the target location or indicating the need to spread attention over all locations. Nineteen rats performed the 5-choice serial reaction time task requiring detection of stimuli presented randomly in a horizontal array of five locations. Performance after nicotine (0.1 and 0.2 mg/kg) or vehicle administration was analyzed as a function of target location eccentricity. In human subjects, nicotine caused greater reaction time reduction when all locations were monitored than when a single location was cued. In rats, nicotine attenuated the decline in stimulus detections and the increase in omission errors with greater target location eccentricity. The findings represent cross-species evidence that nAChR agonism facilitates the ability to spread attention broadly. This suggests that nAChR hypofunction may be central to broad monitoring deficits as seen, for example, in schizophrenia. The homology of findings between the rodent and the human paradigm contributes to validating a translational strategy for treatment development.

Under environmental stress, plants and algae employ a variety of strategies to protect the photosynthetic apparatus and maintain photostasis. To date, most studies on stress acclimation have focused on model organisms which possess limited to no tolerance to stressful extremes. We studied the ability of the Antarctic alga Chlamydomonas sp. UWO 241 (UWO 241) to acclimate to low temperature, high salinity or high light. UWO 241 maintained robust growth and photosynthetic activity at levels of temperature (2 °C) and salinity (700 mM NaCl) which were nonpermissive for a mesophilic sister species, Chlamydomonas raudensis SAG 49.72 (SAG 49.72). Acclimation in the mesophile involved classic mechanisms, including downregulation of light harvesting and shifts in excitation energy between photosystem I and II. In contrast, UWO 241 exhibited high rates of PSI-driven cyclic electron flow (CEF) and a larger capacity for nonphotochemical quenching (NPQ). Furthermore, UWO 241 exhibited constitutively high activity of two key ascorbate cycle enzymes, ascorbate peroxidase and glutathione reductase and maintained a large ascorbate pool. These results matched the ability of the psychrophile to maintain low ROS under short-term photoinhibition conditions. We conclude that tight control over photostasis and ROS levels are essential for photosynthetic life to flourish in a native habitat of permanent photooxidative stress. We propose to rename this organism Chlamydomonas priscuii.

The cortico–basal ganglia–thalamo–cortical loop is one of the fundamental network motifs in the brain. Revealing its structural and functional organization is critical to understanding cognition, sensorimotor behaviour, and the natural history of many neurological and neuropsychiatric disorders. Classically, this network is conceptualized to contain three information channels: motor, limbic and associative1–4. Yet this three-channel view cannot explain the myriad functions of the basal ganglia. We previously subdivided the dorsal striatum into 29 functional domains on the basis of the topography of inputs from the entire cortex5. Here we map the multi-synaptic output pathways of these striatal domains through the globus pallidus external part (GPe), substantia nigra reticular part (SNr), thalamic nuclei and cortex. Accordingly, we identify 14 SNr and 36 GPe domains and a direct cortico-SNr projection. The striatonigral direct pathway displays a greater convergence of striatal inputs than the more parallel striatopallidal indirect pathway, although direct and indirect pathways originating from the same striatal domain ultimately converge onto the same postsynaptic SNr neurons. Following the SNr outputs, we delineate six domains in the parafascicular and ventromedial thalamic nuclei. Subsequently, we identify six parallel cortico–basal ganglia–thalamic subnetworks that sequentially transduce specific subsets of cortical information through every elemental node of the cortico– basal ganglia–thalamic loop. Thalamic domains relay this output back to the originating corticostriatal neurons of each subnetwork in a bona fide closed loop., Mesoscale connectomic mapping of the cortico–basal ganglia–thalamic network reveals key architectural and information processing features.

Chysodeixis includens (Walker) is a polyphagous economic pest in agricultural landscapes. To detect the occurrence of this pest in the field, trapping using sex pheromone lures is often implemented. However, other plusiine species are cross-attracted to these lures and may be misidentified as C. includens due to their morphological similarities. The objectives of this study were to provide region-specific information on the abundance of C. includens throughout the year as well as document the occurrence of related plusiines crossattracted to C. includens sex pheromone traps in the Florida Panhandle. Twelve commercial fields of peanut and twelve commercial fields of cotton located across Escambia, Santa Rosa, and Jackson counties were monitored with Trécé delta traps baited with C. includens sex pheromone lures (Alpha Scents, Inc.; West Linn, OR) from June 2017 to June 2019. There was no difference in C. includens flight across dryland or irrigated fields. Identifications revealed that in addition to C. includens, the following species of the subfamily Plusiinae were crossed-attracted: Argyrogramma verruca (Fabricius) (Lepidoptera: Noctuidae), Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae), Ctenoplusia oxygramma (Geyer) (Lepidoptera: Noctuidae), and Rachiplusia ou (Guenée) (Lepidoptera: Noctuidae). The occurrence of each species in the region and their flight phenology are documented. Chysodeixis includens abundance was greatest in September and decreased through December. Due to the high abundance of C. oxygramma and similar flight phenology to C. includens, this is the likeliest species to skew estimations and influence management decisions of C. includens, especially early in the crop season, when C. includens abundance is low. Made available in DSpace on 2022-05-24T05:01:37Z (GMT). No. of bitstreams: 1 Alexandre-Seasonal-flight-patterns.pdf: 30387592 bytes, checksum: 403e5b227747fa7fc1a3e4cad2be3161 (MD5) Previous issue date: 2021

Purpose Growth differentiation factor 11 (GDF11) is a key signaling protein required for proper development of many organ systems. Only one prior study has associated an inherited GDF11 variant with a dominant human disease in a family with variable craniofacial and vertebral abnormalities. Here, we expand the phenotypic spectrum associated with GDF11 variants and document the nature of the variants. Methods We present a cohort of six probands with de novo and inherited nonsense/frameshift (4/6 patients) and missense (2/6) variants in GDF11. We generated gdf11 mutant zebrafish to model loss of gdf11 phenotypes and used an overexpression screen in Drosophila to test variant functionality. Results Patients with variants in GDF11 presented with craniofacial (5/6) , vertebral (5/6), neurological (6/6), visual (4/6), cardiac (3/6), auditory (3/6) and connective tissue abnormalities (3/6). gdf11 mutant zebrafish show craniofacial abnormalities and body segmentation defects that match some patient phenotypes. Expression of the patients’ variants in the fly showed that one nonsense variant in GDF11 is a severe loss-of-function (LOF) alleles whereas the missense variants in our cohort are partial LOF variants. Conclusion GDF11 is needed for human development, particularly neuronal development, and LOF GDF11 alleles can affect the development of numerous organs and tissues.

OBJECTIVE Smoking has been associated with increased systemic lupus erythematosus (SLE) risk, but the biologic basis for this association is unknown. Our objective was to investigate whether women's smoking was positively associated with SLE-associated proinflammatory chemokines/cytokines (stem cell factor [SCF], B lymphocyte stimulator [BLyS], interferon-γ-inducible 10-kd protein [IP-10], and interferon-α); or negatively associated with antiinflammatory cytokine interleukin-10 (IL-10); and whether associations were modified by SLE-related autoantibody status. METHODS The Nurses' Health Study (NHS, n = 121,700) and NHSII (n = 116,429) cohorts were begun in 1976 and 1989. In 1988-1990 (NHS) and 1996-1999 (NHSII), ~25% of participants donated blood samples. We identified 1,177 women without SLE with banked samples, and we tested by enzyme-linked immunoassay (ELISA) for chemokines/cytokines as well as anti-Sm, anti-Ro/SSA, anti-La/SSB, and anti-RNP. Antinuclear antibodies (ANAs) were detected by HEp-2 cell indirect immunofluorescence, and anti-double-stranded DNA antibodies and were assayed by ELISA. Smoking was assessed until blood draw. Separate tobit and linear regression analyses, adjusted for potential confounders, modeled associations between smoking and log-transformed chemokine/cytokine concentrations. Analyses were stratified by autoantibody status. Effect estimates were calculated as ratios of geometric means expressed as percentage differences. RESULTS Among the 15% of current/recent versus 85% of past/never smokers, BLyS levels were 8.7% higher (P < 0.01) and were 24% higher (P < 0.0001) among those who were ANA positive. Current/recent smokers had IL-10 concentrations 46% lower (P < 0.01) than past/never smokers; each 10 pack-years of smoking was associated with a 17% decrease in IL-10 level (P < 0.001). Smoking was not associated with IP-10 or SCF. CONCLUSION Elevated BLyS and lower IL-10 levels among current smokers, particularly among ANA-positive women, may be involved in SLE pathogenesis.

Objectives Iloperidone is an atypical antipsychotic drug that is widely used for the treatment of schizophrenia. hERG 3.1, alternatively spliced form of hERG 1A, is considered a potential target for an antipsychotic drug. The present study was designed to study the effects of iloperidone on hERG 1A/3.1 heterotetrameric channels. Methods The interactions of iloperidone with hERG 1A/3.1 heterotetrameric channels stably expressed in HEK cells were investigated using the whole-cell patch-clamp technique and western blot analysis. Results Iloperidone inhibited the hERG 1A/3.1 tail currents at -50 mV in a concentration-dependent manner with an IC50 value of 0.44 μM. The block of hERG 1A/3.1 currents by iloperidone was voltage-dependent and increased over a range of voltage for channel activation. However, the block by iloperidone was voltage-independent at more depolarized potentials where the channels were fully activated. A fast application of iloperidone inhibited the hERG 1A/3.1 current elicited by a 5-s depolarizing pulse to +60 mV to fully inactivate the hERG 1A/3.1 currents. Iloperidone also induced a rapid and reversible inhibition of hERG 1A/3.1 tail currents during repolarization. However, iloperidone had no effect on either hERG 1A or hERG 1A/3.1 channel trafficking to the cell membrane. Conclusions Our results indicated that iloperidone concentration-dependently inhibited hERG 1A/3.1 currents by preferentially interacting with the open states of channels, but not by the disruption of membrane trafficking or surface membrane expression of hERG 1A and hERG 1A/3.1 channel proteins.

Copper nitrite reductase (CuNiR) is a copper enzyme that converts nitrite to nitric oxide and is an important part of the global nitrogen cycle in bacteria. The relatively simple CuHis3 binding site of the CuNiR active site has made it an enticing target for small molecule modeling and de novo protein design studies. We have previously reported symmetric CuNiR models within parallel three stranded coiled coil systems, with activities that span a range of three orders of magnitude. In this report, we investigate the same CuHis3 binding site within an antiparallel three helical bundle scaffold, which allows the design of asymmetric constructs. We determine that a simple CuHis3 binding site can be designed within this scaffold with enhanced activity relative to the comparable construct in parallel coiled coils. Incorporating more complex designs or repositioning this binding site can decrease this activity as much as 15 times. Comparing these constructs, we reaffirm a previous result in which a blue shift in the 1s to 4p transition energy determined by Cu(I) X-ray absorption spectroscopy is correlated with an enhanced activity within imidazole-based constructs. With this step and recent successful electron transfer site designs within this scaffold, we are one step closer to a fully functional de novo designed nitrite reductase.

STK11 encodes for the protein liver kinase B1, a serine/threonine kinase which is involved in a number of physiological processes including regulation of cellular metabolism, cell polarity and the DNA damage response. It acts as a tumour suppressor via multiple mechanisms, most classically through AMP-activated protein kinase-mediated inhibition of the mammalian target of rapamycin signalling pathway. Germline loss-of-function mutations in STK11 give rise to Peutz-Jeghers syndrome, which is associated with hamartomatous polyps of the gastrointestinal tract, mucocutaneous pigmentation and a substantially increased lifetime risk of many cancers. In the sporadic setting, STK11 mutations are commonly seen in a subset of adenocarcinomas of the lung in addition to a number of other tumours occurring at various sites. Mutations in STK11 have been associated with worse prognoses across a range of malignancies and may be a predictor of poor response to immunotherapy in a subset of lung cancers, though further studies are needed before the presence of STK11 mutations can be implemented as a routine clinical biomarker.

The lack of bacteriophages capable of infecting the Listeria species, Listeria grayi, is academically intriguing and presents an obstacle to the development of bacteriophage-based technologies for Listeria. We describe the isolation and engineering of a novel L. grayi bacteriophage, LPJP1, isolated from farm silage. With a genome over 200,000 base pairs, LPJP1 is the first and only reported jumbo bacteriophage infecting the Listeria genus. Similar to other Gram-positive jumbo phages, LPJP1 appeared to contain modified base pairs, which complicated initial attempts to obtain genomic sequence using standard methods. Following successful sequencing with a modified approach, a recombinant of LPJP1 encoding the NanoLuc luciferase was engineered using homologous recombination. This luciferase reporter bacteriophage successfully detected 100 stationary phase colony forming units of both subspecies of L. grayi in four hours. A single log phase colony forming unit was also sufficient for positive detection in the same time period. The recombinant demonstrated complete specificity for this particular Listeria species and did not infect 150 non-L. grayi Listeria strains nor any other bacterial genus. LPJP1 is believed to be the first reported lytic bacteriophage of L. grayi as well as the only jumbo bacteriophage to be successfully engineered into a luciferase reporter.

Human cytomegalovirus congenital infection is the leading non-genetic cause of fetal malformation in developed countries. There are currently no safe antivirals for use during pregnancy. Placental trophoblast cells specifically secrete exosomes containing miRNA from the chromosome 19 miRNA cluster (C19MC) which confer viral resistance to recipient cells. We show the highly expressed C19MC miRNA miR-517a-3p inhibits HCMV replication and viral protein expression in both fibroblast and trophoblast cell cultures (71.6% and 50.4% inhibition of HCMV DNA at 7 days post infection respectively; p 0.05). This naturally occurring molecule has potential for opening-up antiviral therapeutic strategies for pregnancy.

Carbon monoxide (CO) is a therapeutic gas with therapeutic potential in intestinal bowel disease. Therapeutic efficacy in the gastrointestinal tract (GIT) must be paired with safe and convenient use. Therefore, we designed an oral CO releasing system (OCORS) pairing tunable CO release into the GIT while preventing the release of any other molecule from within the device, causing safety concerns. The dimensions of the device, which is manufactured from 3D printed components, are within compendial limits. This is achieved by controlling CO decarbonylation from a molybdenum complex with a FeCl

Mammary tumors in dogs hold great potential as naturally occurring breast cancer models in translational oncology, as they share the same environmental risk factors, key histological features, hormone receptor expression patterns, prognostic factors, and genetic characteristics as their human counterparts. We aimed to develop in vitro tools that allow functional analysis of canine mammary tumors (CMT), as we have a poor understanding of the underlying biology that drives the growth of these heterogeneous tumors. We established the long-term culture of 24 organoid lines from 16 dogs, including organoids derived from normal mammary epithelium or benign lesions. CMT organoids recapitulated key morphological and immunohistological features of the primary tissue from which they were derived, including hormone receptor status. Furthermore, genetic characteristics (driver gene mutations, DNA copy number variations, and single-nucleotide variants) were conserved within tumor-organoid pairs. We show how CMT organoids are a suitable model for in vitro drug assays and can be used to investigate whether specific mutations predict therapy outcomes. Specifically, certain CMT subtypes, such as PIK3CA mutated, estrogen receptor-positive simple carcinomas, can be valuable in setting up a preclinical model highly relevant to human breast cancer research. In addition, we could genetically modify the CMT organoids and use them to perform pooled CRISPR/Cas9 screening, where library representation was accurately maintained. In summary, we present a robust 3D in vitro preclinical model that can be used in translational research, where organoids from normal, benign as well as malignant mammary tissues can be propagated from the same animal to study tumorigenesis.

BACKGROUND AND AIMS HCV infection is a major risk factor that can lead to chronic liver disease, including fibrosis, cirrhosis, and HCC. Progression of chronic liver disease by HCV infection is caused by a complex intercellular reaction. Especially, exosomes and microRNAs (miRNAs) from HCV-infected hepatocytes play a role in the pathogenesis of liver disease by facilitating cellular communication between parenchymal and nonparenchymal cells. However, the underlying mechanism of secretions of exosome and miRNAs during HCV infection is still open for study. APPROACH AND RESULTS In this study, we demonstrated a pathway for the release of exosome and exosomal miRNAs through caspase-3/pannexin 1 (Panx1)/P2X4 activation during HCV infection in hepatocytes. We found that HCV infection induced the stimulation of exosome release and activation of the caspase-3/Panx1/P2X4 pathway in Huh7.5.1 cells. In addition, miR-122 and miR-146a levels in extracellular exosomes from HCV-infected cells were dramatically increased whereas intracellular miR122 and miR-146a expression had no large changes. Notably, secretions of exosomes and exosomal miRNAs were decreased by inhibition of caspase 3, Panx1, and P2X4 whereas inhibition of ROCK-1 cleavage did not affect these during HCV infection in Huh7.5.1 cells. CONCLUSIONS These results suggested that HCV infection caused secretions of exosomes and exosomal miRNAs dependent on the caspase 3/Panx1/P2X4 pathway. Our study provides a possible therapeutic intervention using Panx1 suppression for liver disease development mediated by exosomes from HCV-infected hepatocytes.

Self-propelling micro- and nano-motors (MNMs) have been extensively investigated as an emerging oral drug delivery carrier for gastrointestinal (GI) tract diseases. However, the propulsion of current MNMs reported so far is mostly based on the redox reaction of metals (such as Zn and Mg) with severe propulsion gas generation, remaining non-degradable residue in the GI tract. Here, we develop a bioinspired enzyme-powered biopolymer micromotor mimicking the mucin penetrating behavior of Helicobacter pylori in the stomach. It converts urea to ammonia and the subsequent increase of pH induces local gel-sol transition of the mucin layer facilitating the penetration into the stomach tissue layer. The successful fabrication of micromotors is confirmed by high-resolution transmission electron microscopy, electron energy loss spectroscopy, dynamic light scattering analysis, zeta-potential analysis. In acidic condition, the immobilized urease can efficiently converted urea to ammonia, comparable with that of neutral condition because of the increase of surrounding pH during propulsion. After administration into the stomach, the micromotors show enhanced penetration and prolonged retention in the stomach for 24 h. Furthermore, histological analysis shows that the micromotors are cleared within 3 days without causing any toxicity in the GI tract. The enhanced penetration and retention of the micromotors as an active oral delivery carrier in the stomach would be successfully harnessed for the treatment of various GI tract diseases., Graphical abstract Image 1, Highlights • Polydopamine (PDA) hollow microcapsules are biocompatible, biodegradable, and muco-adhesive. • Urease-powered PDA micromotors are propelled by the decomposition of urea to ammonia and carbon dioxide. • Micromotors increase the local pH to induce the gel-sol transition of the mucus layer. • Micromotors result in enhanced penetration and prolonged retention in the stomach. • Micromotors can be effectively used for oral drug delivery with complete clearance from the GI tract.

Purpose We characterize the clinical and molecular phenotypes of six unrelated individuals with intellectual disability and autism spectrum disorder who carry heterozygous missense variants of the PRKAR1B gene, which encodes the R1β subunit of the cyclic AMP-dependent protein kinase A (PKA). Methods Variants of PRKAR1B were identified by single- or trio-exome analysis. We contacted the families and physicians of the six individuals to collect phenotypic information, performed in vitro analyses of the identified PRKAR1B-variants, and investigated PRKAR1B expression during embryonic development. Results Recent studies of large patient cohorts with neurodevelopmental disorders found significant enrichment of de novo missense variants in PRKAR1B. In our cohort, de novo origin of the PRKAR1B variants could be confirmed in five of six individuals, and four carried the same heterozygous de novo variant c.1003C>T (p.Arg335Trp; NM_001164760). Global developmental delay, autism spectrum disorder, and apraxia/dyspraxia have been reported in all six, and reduced pain sensitivity was found in three individuals carrying the c.1003C>T variant. PRKAR1B expression in the brain was demonstrated during human embryonal development. Additionally, in vitro analyses revealed altered basal PKA activity in cells transfected with variant-harboring PRKAR1B expression constructs. Conclusion Our study provides strong evidence for a PRKAR1B-related neurodevelopmental disorder.

Animals create diverse structures, both individually and cooperatively, using materials from their environment. One striking example are the nests birds build for reproduction, which protect the offspring from external stressors such as predators and temperature, promoting reproductive success. To construct a nest successfully, birds need to make various decisions, for example regarding the nest material and their time budgets. To date, research has focused mainly on species where one sex is primarily responsible for building the nest. In contrast, the cooperative strategies of monogamous species in which both sexes contribute to nest building are poorly understood. Here we investigated the role of both sexes in nest building and fitness correlates of behaviour in wild, monogamous jackdaw pairs (Corvus monedula). We show that both partners contributed to nest building and behaved similarly, with females and males present in the nest box for a comparable duration and transporting material to the nest equally often. However, while females spent more time constructing the nest, males tended to invest more time in vigilance, potentially as a means of coping with competition for nest cavities. These findings suggest a moderate degree of division of labour, which may facilitate cooperation. Moreover, some aspects of behaviour were related to proxies of reproductive success (lay date and egg volume). Females that contributed relatively more to bringing material laid earlier clutches and pairs that spent less time together in the nest box had larger eggs. Thus, selection pressures may act on how nest building pairs spend their time and cooperatively divide the labour. We conclude that cooperative nest building in birds could be associated with monogamy and obligate biparental care, and provides a vital but relatively untapped context through which to study the evolution of cooperation.HighlightsIn wild monogamous jackdaws, mates behaved similarly and cooperated to build their nest.Females built more and called more frequently; males tended to be more vigilant.Females that contributed relatively more to transporting nest material laid earlier clutches.Pairs that spent more time together in the nest box had smaller eggs.Cooperation may be crucial in light of obligate biparental care and nest site competition.

In [In SPECT/CT phantom measurements (NaI cameras, N = 2; CZT, N = 1), peak contrast recovery (CRpeak) was derived from sphere inserts or myocardial insert (cardiac phantom; signal-to-background ratio [SBR], 10:1 or 5:1). In 25 positive and 38 negative patients (reference: endomyocardial biopsy or clinical diagnosis), Perugini scores and heart-to-contralateral (H/CL) count ratios were derived from planar images 1 hour and 3 hour p.i.In phantom measurements, accuracy of myocardial CRpeak at SBR 10:1 (H-mode, 0.95-0.99) and reproducibility at 5:1 (H-mode, 1.02-1.14) was comparable for H-mode and L-mode. However, L-mode showed higher variability of background counts and sphere CRpeak throughout the field of view than H-mode. In patients, sensitivity/specificity were ≥ 95% for H/CL ratios at both time points and visual scoring 3 hour. At 1 hour, visual scores showed specificity of 89% and reduced reader's confidence.Early DPD images provided no additional value for visual scoring or H/CL ratios. In SPECT/CT, H-mode is preferred over L-mode, especially if quantification is applied apart from the myocardium.ANTECEDENTES: En la gammagrafía con [ 99mTc]-DPD para amiloidosis cardiaca ATTR, se recomienda la adquisición de imágenes planares a las 3h p.i y SPECT/CT en modo L. Este estudio investigó si las imágenes planares más tempranas (1h p.i.) tienen beneficios y si la adquisición del SPECT/CT debería preferirse en modo H (ángulo de detección de 180 °) o en modo L (90 °). MéTODOS: En las mediciones con el fantoma del SPECT/CT (cámaras de NaI, n= 2; CZT, n=1), la recuperación pico de contraste (CRpico) se derivó de inserciones de esfera o de inserciones miocárdicas (fantoma cardiaco; relación señal-fondo [SBR], 10:1 o 5: 1). En 25 pacientes positivos y 38 negativos (referencia: biopsia endomiocárdica o diagnóstico clínico), el score de Perugini y la relación de cuentas corazón-contralateral (H/CL) se derivaron de imágenes planares a 1h y 3h p.i.En mediciones con el fantoma, la precisión del CRpico miocárdico en SBR 10:1 (modo H, 0.95-0.99) y la reproducibilidad a 5:1 (modo H, 1.02-1.14) fue comparable para el modo H y el modo L. Sin embargo, el modo L mostró una mayor variabilidad de las cuentas de fondo y del CRpico de la esfera en todo el campo de visión que el modo H. En pacientes, la sensibilidad/especificidad fue ≥ 95% para las relaciones H/CL en ambos puntos de tiempo y con un score visual a las 3 h. A 1h, los scores visuales mostraron una especificidad de (89%) y disminución de la confianza del lector.Las imágenes tempranas de DPD no proporcionaron ningún valor adicional para el score visual o las relaciones H/CL. En SPECT/CT, se prefiere el modo H sobre el modo L, especialmente si la cuantificación se aplica aparte del miocardio.摘要: 背景: [99mTc]Tc-DPD闪烁显像检查心肌ATTR淀粉样变性时, 建议使用3h p.i.平面图像和L模式SPECT/CT扫描。本研究探讨了早期平面图像(1h p.i.)是否有益, 以及SPECT/CT采集是否应首选H模式(探测器角度180°)或L模式(90°)。 方法: 在SPECT/CT模体测量中(NaI相机, n=2;CZT, n=1), 峰值造影剂恢复(CRpeak)来自球体插入物或心肌插入物(心脏模体; 信号与背景比[SBR], 10:1 或5:1)。在25个阳性和38个阴性病人中(参考标准: 心内膜活检或临床诊断), Perugini分数和心脏/对侧 (H / CL)计数比率来自平面图像1 h和3 h p.i.。 结果: 在模体测量中, 心肌CRpeak的SBR准确性 10:1 (H-mode, 0.95 - 0.99)和可重复性5:1 (H-mode, 1.02 - 1.14), H模式和L 模式相当。然而, 与H模式相比, L模式在整个视野中表现出更高的背景计数和球形CRpeak变异性。在患者中, 两个时间点的H/CL比值以及3h视觉评分的敏感性/特异性均≥95%。1h时, 视觉评分特异性为89%, 阅读者的信心降低。 结论: DPD早期图像对视觉评分和H/CL比值没有额外的价值。在SPECT/CT中, H模式优于L模式, 特别是心肌之外定量时。.

Vertebrates harbour gut microbial communities containing hundreds of bacterial species, most of which have never been cultivated or isolated in the laboratory. The lack of cultured representatives from vertebrate gut microbiotas limits the description and experimental interrogation of these communities. Here, we show that representatives from >50% of the bacterial genera detected by culture-independent sequencing in the gut microbiotas of fence lizards, house mice, chimpanzees, and humans were recovered in mixed cultures from frozen faecal samples plated on a panel of nine media under a single growth condition. In addition, culturing captured >100 rare bacterial genera overlooked by culture-independent sequencing, more than doubling the total number of bacterial sequence variants detected. Our approach recovered representatives from 23 previously uncultured candidate bacterial genera, 12 of which were not detected by culture-independent sequencing. Results identified strategies for both indiscriminate and selective culturing of the gut microbiota that were reproducible across vertebrate species. Isolation followed by whole-genome sequencing of 161 bacterial colonies from wild chimpanzees enabled the discovery of candidate novel species closely related to the opportunistic pathogens of humans Clostridium difficile and Hungatella hathewayi. This study establishes culturing methods that improve inventories and facilitate isolation of gut microbiota constituents from a wide diversity of vertebrate species.

SARS‐CoV‐2 mortality has been extensively studied in relation to host susceptibility. How sequence variations in the SARS‐CoV‐2 genome affect pathogenicity is poorly understood. Starting in October 2020, using the methodology of genome‐wide association studies (GWAS), we looked at the association between whole‐genome sequencing (WGS) data of the virus and COVID‐19 mortality as a potential method of early identification of highly pathogenic strains to target for containment. Although continuously updating our analysis, in December 2020, we analyzed 7548 single‐stranded SARS‐CoV‐2 genomes of COVID‐19 patients in the GISAID database and associated variants with mortality using a logistic regression. In total, evaluating 29,891 sequenced loci of the viral genome for association with patient/host mortality, two loci, at 12,053 and 25,088 bp, achieved genome‐wide significance (p values of 4.09e−09 and 4.41e−23, respectively), though only 25,088 bp remained significant in follow‐up analyses. Our association findings were exclusively driven by the samples that were submitted from Brazil (p value of 4.90e−13 for 25,088 bp). The mutation frequency of 25,088 bp in the Brazilian samples on GISAID has rapidly increased from about 0.4 in October/December 2020 to 0.77 in March 2021. Although GWAS methodology is suitable for samples in which mutation frequencies varies between geographical regions, it cannot account for mutation frequencies that change rapidly overtime, rendering a GWAS follow‐up analysis of the GISAID samples that have been submitted after December 2020 as invalid. The locus at 25,088 bp is located in the P.1 strain, which later (April 2021) became one of the distinguishing loci (precisely, substitution V1176F) of the Brazilian strain as defined by the Centers for Disease Control. Specifically, the mutations at 25,088 bp occur in the S2 subunit of the SARS‐CoV‐2 spike protein, which plays a key role in viral entry of target host cells. Since the mutations alter amino acid coding sequences, they potentially imposing structural changes that could enhance viral infectivity and symptom severity. Our analysis suggests that GWAS methodology can provide suitable analysis tools for the real‐time detection of new more transmissible and pathogenic viral strains in databases such as GISAID, though new approaches are needed to accommodate rapidly changing mutation frequencies over time, in the presence of simultaneously changing case/control ratios. Improvements of the associated metadata/patient information in terms of quality and availability will also be important to fully utilize the potential of GWAS methodology in this field.

Current knowledge on environmental distribution and taxon richness of free-living bacteria is mainly based on cultivation-independent investigations employing 16S rRNA gene sequencing methods. Yet, 16S rRNA genes are evolutionarily rather conserved, resulting in limited taxonomic and ecological resolutions provided by this marker. The faster evolving protein-encoding gene priB was used to reveal ecological patterns hidden within a single operational taxonomic unit (OTU) defined by >99% 16S rRNA sequence similarity. The studied subcluster PnecC of the genus Polynucleobacter represents a ubiquitous group of abundant freshwater bacteria with cosmopolitan distribution, which is very frequently detected by diversity surveys of freshwater systems. Based on genome taxonomy and a large set of genome sequences, a sequence similarity threshold for delineation of species-like taxa could be established. In total, 600 species-like taxa were detected in 99 freshwater habitats scattered across three regions representing a latitudinal range of 3,400 km (42 degrees N to 71 degrees N) and a pH gradient of 4.2 to 8.6. In addition to the unexpectedly high richness, the increased taxonomic resolution revealed structuring of Polynucleobacter communities by a couple of macroecological trends, which was previously only demonstrated for phylogenetically much broader groups of bacteria. An unexpected pattern was the almost complete compositional separation of Polynucleobacter communities of Ca2+-rich and Ca2+-poor habitats. This compositional pattern strongly resembled the vicariance of plant species on silicate and limestone soils. The new cultivation-independent approach presented opened a window to an incredible, previously unseen diversity, and enables investigations aiming on deeper understanding of how environmental conditions shape bacterial communities and drive evolution of free-living bacteria.

Background and aims Animal models of human disease are a key component of translational hepatology research, yet there is no consensus on which model is optimal for NAFLD. Approach and results We generated a database of 3,920 rodent models of NAFLD. Study designs were highly heterogeneous, and therefore, few models had been cited more than once. Analysis of genetic models supported the current evidence for the role of adipose dysfunction and suggested a role for innate immunity in the progression of NAFLD. We identified that high-fat, high-fructose diets most closely recapitulate the human phenotype of NAFLD. There was substantial variability in the nomenclature of animal models: a consensus on terminology of specialist diets is needed. More broadly, this analysis demonstrates the variability in preclinical study design, which has wider implications for the reproducibility of in vivo experiments both in the field of hepatology and beyond. Conclusions This systematic analysis provides a framework for phenotypic assessment of NAFLD models and highlights the need for increased standardization and replication.

The objective of the study was to describe the complexity of diagnosis and evaluation of Zika-exposed pregnant women/fetuses and infants in a U.S. Congenital Zika Program. Pregnant women/fetuses and/or infants referred for clinical evaluation to the Congenital Zika Program at Children’s National (Washington, DC) from January 2016 to June 2018 were included. We recorded the timing of maternal Zika-virus (ZIKV) exposure and ZIKV laboratory testing results. Based on laboratory testing, cases were either confirmed, possible, or unlikely ZIKV infection. Prenatal and postnatal imaging by ultrasound and/or magnetic resonance imaging (MRI) were categorized as normal, nonspecific, or as findings of congenital Zika syndrome (CZS). Of 81 women–fetus/infant pairs evaluated, 72 (89%) had confirmed ZIKV exposure; 18% of women were symptomatic; only a minority presented for evaluation within the time frame for laboratory detection. Zika virus could only be confirmed in 29 (40%) cases, was possible in 26 (36%) cases, and was excluded in 17 (24%) cases. Five cases (7%) had prenatal ultrasound and MRI findings of CZS, but in only three was ZIKV confirmed by laboratory testing. Because of timing of exposure to presentation, ZIKV infection could not be excluded in many cases. Neuroimaging found CZS in 7% of cases, and in many patients, there were nonspecific imaging findings that warrant long-term follow-up. Overall, adherence to postnatal recommended follow-up evaluations was modest, representing a barrier to care. These challenges may be instructive to future pediatric multidisciplinary clinics for congenital infectious/noninfectious threats to pregnant women and their infants.

Targeting the androgen receptor by depriving testosterone with gonadotropin-releasing hormone agonists or antagonists, or surgical castration, has been the backbone of metastatic prostate cancer treatment. Although most prostate cancers initially respond to androgen deprivation, metastatic castration-resistant prostate cancer evolves into a heterogeneous disease with diverse drivers of progression and mechanisms of therapeutic resistance. Development of castrate resistance phenotype is associated with lethality despite the recent noteworthy strides gained via increase in therapeutic options. Identification of novel therapeutics to further improve survival and achieve durable responses in metastatic castration-resistant prostate cancer is a clinical necessity. In this review, we outline the existing avengers for treatment of metastatic castration-resistant prostate cancer by clinical presentation, placing into context the clinical state of the patient, such as burden of disease and symptoms. Doing so might aid in the ability to optimize the sequence of agents and allow for maximal exposure to life-prolonging therapeutics. Realizing the limitations of the androgen signaling inhibition, we explore the androgen-indifferent prostate cancer: the mutants. Classically, these subtypes have been associated with variant histology, but androgen-indifferent prostate cancer features are now frequently observed in association with heterogeneous morphologies, including double-negative prostate cancers, lacking both androgen receptor and neuroendocrine features, or clinicopathologic criteria, such as the aggressive variant prostate cancer criteria. The framework of new avengers against metastatic castration-resistant prostate cancer based on mechanism, including DNA repair, immune checkpoint inhibition, PTEN/PI3K/AKT pathway, prostate-specific membrane antigen targets, bispecific T-cell engagers, and radionuclide therapies, is summarized in this review.

This dissertation lays the foundation for a research program that highlights the important contribution that poorly-dispersing short-range endemic (SRE) organisms can have on advancing empirical and theoretical evolutionary biology. Here, SREs are represented by members of the harvestmen (Opiliones) superfamily Ischyropsalidoidea. In Chapter 1, phylogenetic analyses of an ancient rapid diversification at the base of Ischyropsalidoidea uncovers considerable incongruence across gene trees. Incongruent gene trees are not recovered in frequencies expected from a simple multispecies coalescent model, and incomplete lineage sorting is rejected as the sole contributor to gene tree conflict. In Chapter 2, an integrative taxonomy tests existing, and puts forward novel species-level taxonomic hypotheses within the Acuclavella species endemic to the inland temperate rainforest of the U.S. northern Rocky Mountains. Incongruence among taxonomic disciplines suggests rapid morphological evolution across multiple recent diversifications. Results suggest at least two undescribed species exist within inland populations of Acuclavella. Alternative combinations of incongruence across taxonomic disciplines possibly represent different stages of the speciation continuum, and suggest that different evolutionary mechanisms underlie speciation in thorn harvestmen, thus highlighting the importance of integrative species delimitation for evolutionary research. The disjunct inland temperate rainforest has become a model for recent biogeography, with a complex and recent evolutionary history suggesting that this inland forest was a refuge from Pleistocene glaciation that was compartmentalized into variously isolated pockets during the last glacial maximum (LGM). In Chapter 3, genomic-scale data are used to evaluate the evolutionary consequences of the LGM. This research finds that rivers were a major cause of refugia compartmentalization, with populations and species structured by rivers well before the LGM. Additionally, a negative correlation between elevation and heterozygosity suggests that populations were confined to lower valleys during the LGM with post-glacial colonization of higher elevations. The combined effects of rivers and glaciers paints a picture of a complex Pleistocene glacial refugia system during the LGM. This relatively high-resolution inference of past evolutionary events within an established biogeographic model system points towards the undervalued importance of using poorly-dispersing short-range endemic taxa for biogeography and phylogeography.

The dynamics and phenotypes of intratumoral myeloid cells during tumor progression are poorly understood. Here we define myeloid cellular states in gliomas by longitudinal single-cell profiling and demonstrate their strict control by the tumor genotype: in isocitrate dehydrogenase (IDH)-mutant tumors, differentiation of infiltrating myeloid cells is blocked, resulting in an immature phenotype. In late-stage gliomas, monocyte-derived macrophages drive tolerogenic alignment of the microenvironment, thus preventing T cell response. We define the IDH-dependent tumor education of infiltrating macrophages to be causally related to a complex re-orchestration of tryptophan metabolism, resulting in activation of the aryl hydrocarbon receptor. We further show that the altered metabolism of IDH-mutant gliomas maintains this axis in bystander cells and that pharmacological inhibition of tryptophan metabolism can reverse immunosuppression. In conclusion, we provide evidence of a glioma genotype-dependent intratumoral network of resident and recruited myeloid cells and identify tryptophan metabolism as a target for immunotherapy of IDH-mutant tumors. Platten and colleagues find that tryptophan metabolism by myeloid cells contributes to immunosuppressive microenvironment uniquely in IDH-mutant gliomas, which can be overcome by inhibiting this pathway in murine tumor models.