Modeling the limits of detection for antimicrobial resistance genes in agri-food samples: a comparative analysis of bioinformatics tools.

BACKGROUND: Although the spread of antimicrobial resistance (AMR) through food and its production poses a significant concern, there is limited research on the prevalence of AMR bacteria in various agri-food products. Sequencing technologies are increasingly being used to track the spread of AMR genes (ARGs) in bacteria, and metagenomics has the potential to bypass some of the limitations of single isolate characterization by allowing simultaneous analysis of the agri-food product microbiome and associated resistome. However, metagenomics may still be hindered by methodological biases, presence of eukaryotic DNA, and difficulties in detecting low abundance targets within an attainable sequence coverage. The goal of this study was to assess whether limits of detection of ARGs in agri-food metagenomes were influenced by sample type and bioinformatic approaches. RESULTS: We simulated metagenomes containing different proportions of AMR pathogens and analysed them for taxonomic composition and ARGs using several common bioinformatic tools. Kraken2/Bracken estimates of species abundance were closest to expected values. However, analysis by both Kraken2/Bracken indicated presence of organisms not included in the synthetic metagenomes. Metaphlan3/Metaphlan4 analysis of community composition was more specific but with lower sensitivity than the Kraken2/Bracken analysis. Accurate detection of ARGs dropped drastically below 5X isolate genome coverage. However, it was sometimes possible to detect ARGs and closely related alleles at lower coverage levels if using a lower ARG-target coverage cutoff (< 80%). While KMA and CARD-RGI only predicted presence of expected ARG-targets or closely related gene-alleles, SRST2 (which allows read to map to multiple targets) falsely reported presence of distantly related ARGs at all isolate genome coverage levels. The presence of background microbiota in metagenomes influenced the accuracy of ARG detection by KMA, resulting in mcr-1 detection at 0.1X isolate coverage in the lettuce but not in the beef metagenome. CONCLUSIONS: This study demonstrates accurate detection of ARGs in synthetic metagenomes using various bioinformatic methods, provided that reads from the ARG-encoding organism exceed approximately 5X isolate coverage (i.e. 0.4% of a 40 million read metagenome). While lowering thresholds for target gene detection improved sensitivity, this led to the identification of alternative ARG-alleles, potentially confounding the identification of critical ARGs in the resistome. Further advancements in sequencing technologies providing increased coverage depth or extended read lengths may improve ARG detection in agri-food metagenomic samples, enabling use of this approach for tracking clinically important ARGs in agri-food samples.

Cognitive Reserve Modifies the Relationship Between Neural Function, Neural Injury and Upper-Limb Recovery After Stroke.

OBJECTIVE: To investigate whether cognitive reserve modifies the relationship between functional connectivity, lesion volume, stroke severity and upper-limb motor impairment and recovery in stroke survivors. METHODS: Ten patients with first-ever ischemic middle cerebral artery stroke completed the Cognitive Reserve Index Questionnaire at baseline. Upper-limb motor impairment and functional connectivity were assessed using the Fugl-Meyer Assessment and electroencephalography respectively at baseline and 3-months post-stroke. A debiased weighted phase lag index was computed to estimate functional connectivity between electrodes. Partial least squares (PLS) regression identified a connectivity model that maximally predicted variance in the degree of upper-limb impairment. Regression models were generated to determine whether cognitive reserve modified the relationship between neural function (functional connectivity), neural injury (lesion volume), stroke severity (National Institutes of Health Stroke Scale) and upper-limb motor impairment at baseline and recovery at 3-months (Fugl-Meyer Assessment). RESULTS: The addition of cognitive reserve to a regression model with a dependent variable of upper-limb motor recovery and independent variables of functional connectivity between the ipsilesional motor cortex and parietal cortex, stroke severity and lesion volume improved model efficiency (∆BIC=-7.07) despite not reaching statistical significance (R2=0.90, p=0.07). Cognitive reserve did not appear to improve regression models examining motor impairment at baseline. CONCLUSIONS: Preliminary observations suggest cognitive reserve might modify the relationship between neural function, neural injury, stroke severity and upper-limb motor recovery. Further investigation of cognitive reserve in motor recovery post-stroke appears warranted.

COMPASS: the COMPletely Arbitrary Sequence Simulator.

SUMMARY: Simulated sequence alignments are frequently used to test bioinformatics tools, but current sequence simulators are limited to defined state spaces. Here, we present the COMPletely Arbitrary Sequence Simulator (COMPASS), which is able to simulate the evolution of absolutely any discrete state space along a tree, for any form of time-reversible model. AVAILABILITY AND IMPLEMENTATION: COMPASS is implemented in Python 2.7, and is freely available for all platforms with the Supplementary Information, as well as at http://labs.carleton.ca/eme/software-and-data. CONTACT: alex_wong@carleton.ca. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A Comparison of Pneumatic Actuators for Soft Growing Vine Robots.

Soft pneumatic actuators are used to steer soft growing "vine" robots while being flexible enough to undergo the tip eversion required for growth. In this study, we compared the performance of three types of pneumatic actuators in terms of their ability to perform eversion, quasi-static bending, dynamic motion, and force output: the pouch motor, the cylindrical pneumatic artificial muscle (cPAM), and the fabric pneumatic artificial muscle (fPAM). The pouch motor is advantageous for prototyping owing to its simple manufacturing process. The cPAM exhibits superior bending behavior and produces the highest forces, whereas the fPAM actuates fastest and everts at the lowest pressure. We evaluated a range of dimensions for each actuator type. Larger actuators can produce more significant deformations and forces, but smaller actuators inflate faster and can evert at a lower pressure. Because vine robots are lightweight, the effect of gravity on the functionality of different actuators is minimal. We developed a new analytical model that predicts the pressure-to-bending behavior of vine robot actuators. Using the actuator results, we designed and demonstrated a 4.8 m long vine robot equipped with highly maneuverable 60 × 60 mm cPAMs in a three-dimensional obstacle course. The vine robot was able to move around sharp turns, travel through a passage smaller than its diameter, and lift itself against gravity.

TLR regulation of SPSB1 controls inducible nitric oxide synthase induction.

The mammalian innate immune system has evolved to recognize foreign molecules derived from pathogens via the TLRs. TLR3 and TLR4 can signal via the TIR domain-containing adapter inducing IFN-ß (TRIF), which results in the transcription of a small array of genes, including IFN-ß. Inducible NO synthase (iNOS), which catalyzes the production of NO, is induced by a range of stimuli, including cytokines and microbes. NO is a potent source of reactive nitrogen species that play an important role in killing intracellular pathogens and forms a crucial component of host defense. We have recently identified iNOS as a target of the mammalian SPSB2 protein. The SOCS box is a peptide motif, which, in conjunction with elongins B and C, recruits cullin-5 and Rbx-2 to form an active E3 ubiquitin ligase complex. In this study, we show that SPSB1 is the only SPSB family member to be regulated by the same TLR pathways that induce iNOS expression and characterize the interaction between SPSB1 and iNOS. Through the use of SPSB1 transgenic mouse macrophages and short hairpin RNA knockdown of SPSB1, we show that SPSB1 controls both the induction of iNOS and the subsequent production of NO downstream of TLR3 and TLR4. Further, we demonstrate that regulation of iNOS by SPSB1 is dependent on the proteasome. These results suggest that SPSB1 acts through a negative-feedback loop that, together with SPSB2, controls the extent of iNOS induction and NO production.

Parent-Adolescent Conflict: Adolescents' Coping Strategies and Preferred Parenting Styles during the Social Movements in Hong Kong, 2019.

This research investigated parent-adolescent conflict, conflict resolution strategies and perceived parenting styles by adolescents during the social movements in Hong Kong in 2019, a period characterized by considerable social unrest in which many young people participated in demonstrations and protests. The study drew on responses from 866 adolescents aged between 11 and 16 who completed a questionnaire that included a conflict issue checklist and elicited respondents' conflict resolution strategies as well as perceived parenting styles. Correlation analysis was performed to identify the correlation of parent-adolescent conflicts with differences in political stances with their parents and other demographic data. Regression analysis was performed to identify the correlation of perceived parenting styles and conflict resolution strategies adopted by adolescents. Results indicated that early adolescents have a higher intensity of conflicts with their parents than late adolescents in this period. Respondents had more intense conflicts with their parents over political differences and ways of expressing their political views than other issues. Those respondents in conflict or ineffective arguing strategies perceived their parents as more authoritarian than those who adopt positive conflict resolution strategies. However, when asked about their ideal ways of resolving conflicts, adolescents preferred problem-solving rather than conflict strategies.

Primary T-cell Vitreoretinal Non-Hodgkin Lymphoma: A Case Report and Literature Review.

A 72-year-old Chinese male presented with unilateral left eye panuveitis, then diagnosed as bilateral T-cell primary vitreoretinal lymphoma (T-PVRL) through chorioretinal biopsy and immunohistochemistry. No CNS nor systemic involvement was found at diagnosis. Despite initiating intravenous and intrathecal chemotherapy and intravitreal methotrexate, the disease eventually spread to the fellow eye with subsequent recurrence and systemic metastasis. To our knowledge, no cases of T-PVRL treated in a silicone-filled eye were reported in the literature. T- PVRL is exceedingly rare, with most PVRL being the malignant B-cell variant. This case highlights the challenges encountered throughout the treatment course of this aggressive entity, including the administration of intravitreal methotrexate in a silicone oil-filled eye. The poor overall survival rate and grim prognosis of T-PVRL are highlighted. Therefore, we recommend prompt tissue biopsy and immediate initiation of systemic chemotherapy and intravitreal methotrexate.

Find Joy in Sorrows: The Role of Hope in Buffering the Psychological Impact of COVID-19 on Chinese University Students in Hong Kong.

The global coronavirus disease 2019 (COVID-19) crisis has exerted significant psychological impacts on university students who have faced drastic changes in the learning mode and suspension of classes. Despite these challenges, many students maintained subjective well-being. In this study, we examined the role of "hope" as a potential protector to maintain their subjective well-being when facing adversity during this global crisis. Specifically, we explored the mediating role of two hope components (agency thinking and pathways thinking) on the association of positive emotions and life satisfaction among Chinese university students. We conducted an online survey at a local university and recruited a total of 315 undergraduates from the humanities, creative arts, and social sciences programs through convenience sampling. Participants confirmed their informed consent and completed a set of self-administered questionnaires measuring positive emotions, hope, life satisfaction, and demographic variables. The results of mediation testing indicated that, during a global crisis such as COVID-19, positive emotions indirectly influenced life satisfaction through agency thinking rather than pathways thinking. The findings highlight the importance of agency thinking among Chinese university students in adverse situations and provide valuable insights for psychological interventions during a crisis. The article concludes by discussing possible explanations and implications of the findings in a post-pandemic world.

Role of the helical structure of the N-terminal region of Plasmodium falciparum merozoite surface protein 2 in fibril formation and membrane interaction.

Merozoite surface protein 2 (MSP2), an abundant glycosylphosphatidylinositol-anchored protein on the surface of Plasmodium falciparum merozoites, is a promising malaria vaccine candidate. MSP2 is intrinsically disordered and forms amyloid-like fibrils in solution under physiological conditions. The 25 N-terminal residues (MSP2(1-25)) play an important role in both fibril formation and membrane binding of the full-length protein. In this study, the fibril formation and solution structure of MSP2(1-25) in the membrane mimetic solvents sodium dodecyl sulfate (SDS), dodecylphosphocholine (DPC), and trifluoroethanol (TFE) have been investigated by transmission electronic microscopy, turbidity, thioflavin T fluorescence, circular dichroism (CD), and nuclear magnetic resonance (NMR) spectroscopy. Turbidity data showed that the aggregation of MSP2(1-25) was suppressed in the presence of membrane mimetic solvents. CD spectra indicated that helical structure in MSP2(1-25) was stabilized in SDS and DPC micelles and in high concentrations of TFE. The structure of MSP2(1-25) in 50% aqueous TFE, determined using NMR, showed that the peptide formed an amphipathic helix encompassing residues 10-24. Low concentrations of TFE favored partially folded helical conformations, as demonstrated by CD and NMR, and promoted MSP2(1-25) fibril formation. Our data suggest that partially folded helical conformations of the N-terminal region of MSP2 are on the pathway to amyloid fibril formation, while higher degrees of helical structure stabilized by high concentrations of TFE or membrane mimetics suppress self-association and thus inhibit fibril formation. The roles of the induced helical conformations in membrane interactions are also discussed.

The N-terminal domains of SOCS proteins: a conserved region in the disordered N-termini of SOCS4 and 5.

Suppressors of cytokine signaling (SOCS) proteins function as negative regulators of cytokine signaling and are involved in fine tuning the immune response. The structure and role of the SH2 domains and C-terminal SOCS box motifs of the SOCS proteins are well characterized, but the long N-terminal domains of SOCS4-7 remain poorly understood. Here, we present bioinformatic analyses of the N-terminal domains of the mammalian SOCS proteins, which indicate that these domains of SOCS4, 5, 6, and 7 are largely disordered. We have also identified a conserved region of about 70 residues in the N-terminal domains of SOCS4 and 5 that is predicted to be more ordered than the surrounding sequence. The conservation of this region can be traced as far back as lower vertebrates. As conserved regions with increased structural propensity that are located within long disordered regions often contain molecular recognition motifs, we expressed the N-terminal conserved region of mouse SOCS4 for further analysis. This region, mSOCS486₋155, has been characterized by circular dichroism and nuclear magnetic resonance spectroscopy, both of which indicate that it is predominantly unstructured in aqueous solution, although it becomes helical in the presence of trifluoroethanol. The high degree of sequence conservation of this region across different species and between SOCS4 and SOCS5 nonetheless implies that it has an important functional role, and presumably this region adopts a more ordered conformation in complex with its partners. The recombinant protein will be a valuable tool in identifying these partners and defining the structures of these complexes.

Where There's a Will, There's a Way? Social and Mental Forces of Successful Adaptation of Immigrant Children in Young Adulthood.

Although the twenty-first century is deemed as a new era of globalization, waves of immigration continue, due to disparities between politically and economically unstable regions and Western democratized and developed countries. Immigration research has therefore reignited its attention on the successful adaptation of immigrants' offspring, which has profound implications for Western immigrant-receiving countries, as well as worldwide stability. Although immigration research mainly informed by the conventional assimilation theory and/or segmented assimilation perspective accentuates the importance of structural factors, termed as social forces here, in relation to immigrant children's successful adaptation in adolescence, an argument of determinism and tenability keeps on and the contribution of human mental resources and determination, termed as mental forces here, in shaping life trajectories of immigrant children should be not ignored. For this, with a representative sample of 3344 immigrant children from the Children of Immigrants Longitudinal Study (CILS), we examined and compared both the effects of social and mental forces measured in adolescence of immigrant children on their multiple adaptation outcomes in terms of college graduation, engagement in postgraduate study, and first and current job attainments in young adulthood with a Bayesian multilevel modeling framework. The results found that both social forces of segmented assimilation theory and mental forces of immigrant children in adolescence were significantly predictive of immigrant children's successful adaptation in young adulthood (OR = 1.088-2.959 and ß = 0.050-0.639 for social forces; OR = 11.290-18.119 and ß = 0.293-0.297 for mental forces), in which, although the latter showed stronger effects than the former, the effects of mental forces on adaptation of immigrant children were conditionally shaped by the contexts of the social forces informed by segmented assimilation theory. The findings of the current study highlight the significance of the organism-environment interaction perspective on immigration research and provide an insight to consider a context-driven response thesis proposed.

Hazardous alcohol use and HIV indicators in six African countries: results from the Population-based HIV Impact Assessments, 2015-2017.

INTRODUCTION: Hazardous alcohol use (HAU), defined as a pattern of alcohol consumption that increases the risk of harmful consequences for the user or others, is associated with an elevated risk of human immunodeficiency virus (HIV) infection and poor health outcomes. We describe the association between people living with HIV (PLHIV) who report HAU and key HIV indicators. Gaps in current literature in estimating HAU on HIV outcomes at the regional level of Eastern and Southern Africa still exist and our analysis aims to address this issue. METHODS: We used weighted pooled data (2015-2017) from the nationally representative Population-based HIV Impact Assessments among adults who provided written consent aged 18-59 years from Eswatini, Malawi, Namibia, Tanzania, Zambia and Zimbabwe. We estimated differences in the prevalence of HIV infection and The Joint United Nations Programme on HIV and AIDS (UNAIDS) 90-90-90 indicators between PLHIV by HAU status using log-binomial regression, stratified by sex. HAU was determined using the Alcohol Use Identification Test-Consumption. RESULTS: Among the 9755 women and 4444 men who tested HIV positive, 6.6% of women and 21.8% of men engaged in HAU. Women who reported HAU were more likely to be HIV positive (adjusted prevalence ratio [aPR] = 1.31, 95% CI: 1.18-1.46) compared to those who did not report HAU. For the UNAIDS 90-90-90 targets, women who engaged in HAU were more likely to be unaware of their HIV-positive status (aPR = 1.22, 95% CI: 1.01-1.47) and not on antiretroviral therapy (ART) (aPR = 1.73, 95% CI: 1.26-2.37). Men who engaged in HAU were more likely to be unaware of their HIV-positive status (aPR = 1.56, 95% CI 1.39-1.76) and not on ART (aPR = 1.72, 95% CI: 1.30-2.29). No difference in viral load suppression, defined as <1000 copies/ml of HIV RNA, was seen by sex. CONCLUSIONS: PLHIV who engage in HAU were more likely to have suboptimal outcomes along the HIV care continuum when compared to those who did not engage in HAU. Targeted interventions, such as alcohol screening for HAU in HIV testing and treatment settings and HIV prevention efforts in alcohol-based venues, may help countries reach HIV epidemic control by 2030.

Effects of Interfering Cue Separation Distance and Amplitude on the Haptic Detection of Skin Stretch.

Multi-sensory haptic cues, which contain several types of tactile stimuli that are presented concurrently to the user, have been shown to be useful for conveying information-rich cues. One limitation of multi-sensory cues is that user perception of individual cue components can be hindered by more salient components of the composite cue. In this article, we investigate how amplitude and distance between cues affect the perception of multi-sensory haptic cues. Specifically, participants' absolute threshold perception of stretch cues was measured in the presence of interfering squeeze cues using a modular testbed. We evaluated ten conditions of varying interference amplitude and distance between cues. We found that interference cue amplitude and distance between cues both have a statistically significant effect on the absolute perception of stretch cues. As interference cue amplitude increases, and as distance between cues decreases, absolute perception of stretch cues worsens. These results inform design considerations for future wearable multi-sensory haptic devices, so that cue salience can be maximized and interference effects minimized.

Systematic Evaluation of Whole Genome Sequence-Based Predictions of Salmonella Serotype and Antimicrobial Resistance.

Whole-genome sequencing (WGS) is used increasingly in public-health laboratories for typing and characterizing foodborne pathogens. To evaluate the performance of existing bioinformatic tools for in silico prediction of antimicrobial resistance (AMR) and serotypes of Salmonella enterica, WGS-based genotype predictions were compared with the results of traditional phenotyping assays. A total of 111 S. enterica isolates recovered from a Canadian baseline study on broiler chicken conducted in 2012-2013 were selected based on phenotypic resistance to 15 different antibiotics and isolates were subjected to WGS. Both SeqSero2 and SISTR accurately determined S. enterica serotypes, with full matches to laboratory results for 87.4 and 89.2% of isolates, respectively, and partial matches for the remaining isolates. Antimicrobial resistance genes (ARGs) were identified using several bioinformatics tools including the Comprehensive Antibiotic Resistance Database - Resistance Gene Identifier (CARD-RGI), Center for Genomic Epidemiology (CGE) ResFinder web tool, Short Read Sequence Typing for Bacterial Pathogens (SRST2 v 0.2.0), and k-mer alignment method (KMA v 1.17). All ARG identification tools had ≥ 99% accuracy for predicting resistance to all antibiotics tested except streptomycin (accuracy 94.6%). Evaluation of ARG detection in assembled versus raw-read WGS data found minimal observable differences that were gene- and coverage- dependent. Where initial phenotypic results indicated isolates were sensitive, yet ARGs were detected, repeat AMR testing corrected discrepancies. All tools failed to find resistance-determining genes for one gentamicin- and two streptomycin-resistant isolates. Further investigation found a single nucleotide polymorphism (SNP) in the nuoF coding region of one of the isolates which may be responsible for the observed streptomycin-resistant phenotype. Overall, WGS-based predictions of AMR and serotype were highly concordant with phenotype determination regardless of computational approach used.

HIV coreceptor phenotyping in the clinical setting.

The introduction of CCR5 antagonists increases the options available for constructing antiretroviral regimens. However, this option is coupled with the caveat that patients should be tested for HIV coreceptor tropism prior to initiating CCR5 antagonist-based therapy. Failure to screen for CXCR4 usage increases the risk of using an ineffective drug, thus reducing the likelihood of viral suppression and increasing their risk for developing antiretroviral resistance. This review discusses current and future methods of determining HIV tropism, with a focus on their utility in the clinical setting for screening purposes. Some of these methods include recombinant phenotypic tests, such as the Monogram Trofile assay, as well as genotype-based predictors, heteroduplex tracking assays, and flow cytometry based methods. Currently, the best evidence supports the use of phenotypic methods, although other methods of screening for HIV coreceptor usage prior to the administration of CCR5 antagonists may reduce costs and increase turnaround time over phenotypic methods. The presence of low levels of X4 virus is a challenge to all assay methods, resulting in reduced sensitivity in clinical, patient-derived samples when compared to clonally derived samples. Gaining a better understanding of the output of these assays and correlating them with clinical progression and therapy response will provide some indication on how both genotype-based, and phenotypic assays for determining HIV coreceptor usage can be improved. In addition, leveraging new technologies capable of detecting low-level minority species may provide the most significant advances in ensuring that individuals with low levels of dual/mixed tropic virus are not inadvertently prescribed CCR5 antagonists.

Prostate MRI evolution in clinical practice: Audit of tumour detection and staging versus prostatectomy with staged introduction of multiparametric MRI and Prostate Imaging Reporting and Data System v2 reporting.

INTRODUCTION: We conducted a retrospective audit to compare dominant nodule detection and local staging before and after the introduction of functional sequences and PI-RADS v2 reporting to MRI prostate scans in routine private practice. METHODS: A retrospective audit was performed of 245 patients in four separate groups undergoing robotic prostatectomy for prostate cancer by a single urologist between 2009 and 2017. The initial 100 consecutive patients had T2 imaging only. The next 43 patients had T2 and DWI. 52 subsequent patients had T2, DWI and DCE sequences (mpMRI). A final 50 consecutive patients had mpMRI using PI-RADS v2 reporting. Preoperative MRI reports were compared with prostatectomy histopathology to determine the sensitivity of MRI in detecting dominant tumour nodule and T3 extension. RESULTS: The addition of DWI and DCE sequences improved sensitivity for detection of dominant tumour nodule, with a significant further increase using PI-RADS v2 reporting (38% for T2 vs. 62% for T2/DWI vs. 67% for mpMRI vs 91% for PI-RADS v2). The accuracy of detecting T3 disease was initially very low. The use of additional imaging techniques did not significantly influence this, but the use of a three category likelihood of extraprostatic extension in the PI-RADS v2 group had a significant increase in detection of T3 disease (sensitivity 27% vs. 23% vs. 38% vs 63%). CONCLUSION: This audit tracks the significant improvements in MRI detection of prostate cancer dominant tumour nodule and T3 extension in patients undergoing prostatectomy with changing techniques and reporting standards in routine clinical practice.

ConFindr: rapid detection of intraspecies and cross-species contamination in bacterial whole-genome sequence data.

Whole-genome sequencing (WGS) of bacterial pathogens is currently widely used to support public-health investigations. The ability to assess WGS data quality is critical to underpin the reliability of downstream analyses. Sequence contamination is a quality issue that could potentially impact WGS-based findings; however, existing tools do not readily identify contamination from closely-related organisms. To address this gap, we have developed a computational pipeline, ConFindr, for detection of intraspecies contamination. ConFindr determines the presence of contaminating sequences based on the identification of multiple alleles of core, single-copy, ribosomal-protein genes in raw sequencing reads. The performance of this tool was assessed using simulated and lab-generated Illumina short-read WGS data with varying levels of contamination (0-20% of reads) and varying genetic distance between the designated target and contaminant strains. Intraspecies and cross-species contamination was reliably detected in datasets containing 5% or more reads from a second, unrelated strain. ConFindr detected intraspecies contamination with higher sensitivity than existing tools, while also being able to automatically detect cross-species contamination with similar sensitivity. The implementation of ConFindr in quality-control pipelines will help to improve the reliability of WGS databases as well as the accuracy of downstream analyses. ConFindr is written in Python, and is freely available under the MIT License at github.com/OLC-Bioinformatics/ConFindr.