The effect of a daily quiz (TOPday) on self-confidence, enthusiasm, and test results for biomechanics

Many students in Biomedical Sciences have difficulty understanding biomechanics. In a second-year course, biomechanics is taught in the first week and examined at the end of the fourth week. Knowledge is retained longer if the subject material is repeated. However, how does one encourage students to repeat the subject matter? For this study, we developed ‘two opportunities to practice per day (TOPday)’, consisting of multiple-choice questions on biomechanics with immediate feedback, which were sent via e-mail. We investigated the effect of TOPday on self-confidence, enthusiasm, and test results for biomechanics. All second-year students (n = 95) received a TOPday of biomechanics on every regular course day with increasing difficulty during the course. At the end of the course, a non-anonymous questionnaire was conducted. The students were asked how many TOPday questions they completed (0–6 questions [group A]; 7–18 questions [group B]; 19–24 questions [group C]). Other questions included the appreciation for TOPday, and increase (no/yes) in self-confidence and enthusiasm for biomechanics. Seventy-eight students participated in the examination and completed the questionnaire. The appreciation for TOPday in group A (n = 14), B (n = 23) and C (n = 41) was 7.0 (95 % CI 6.5–7.5), 7.4 (95 % CI 7.0–7.8), and 7.9 (95 % CI 7.6–8.1), respectively (p < 0.01 between A and C). Of the students who actively participated (B and C), 91 and 80 % reported an increase in their self-confidence and enthusiasm, respectively, for biomechanics due to TOPday. In addition, they had a higher test result for biomechanics (p < 0.01) compared with those who did not actively participate (A). In conclusion, the teaching method ‘TOPday’ seems an effective way to encourage students to repeat the subject material, with the extra advantage that students are stimulated to keep on practising for the examination. The appreciation was high and there was a positive association between active participation, on the one hand, and self-confidence, enthusiasm, and test results for biomechanics on the other

Polyadenylation of genomic RNA and initiation of antigenomic RNA in a positive-strand RNA virus are controlled by the same cis-element

Genomes and antigenomes of many positive-strand RNA viruses contain 3′-poly(A) and 5′-poly(U) tracts, respectively, serving as mutual templates. Mechanism(s) controlling the length of these homopolymeric stretches are not well understood. Here, we show that in coxsackievirus B3 (CVB3) and three other enteroviruses the poly(A) tract is ∼80–90 and the poly(U) tract is ∼20 nt-long. Mutagenesis analysis indicate that the length of the CVB3 3′-poly(A) is determined by the oriR, a cis-element in the 3′-noncoding region of viral RNA. In contrast, while mutations of the oriR inhibit initiation of (−) RNA synthesis, they do not affect the 5′-poly(U) length. Poly(A)-lacking genomes are able to acquire genetically unstable AU-rich poly(A)-terminated 3′-tails, which may be generated by a mechanism distinct from the cognate viral RNA polyadenylation. The aberrant tails ensure only inefficient replication. The possibility of RNA replication independent of oriR and poly(A) demonstrate that highly debilitated viruses are able to survive by utilizing ‘emergence’, perhaps atavistic, mechanisms

Dissecting distinct proteolytic activities of FMDV Lpro implicates cleavage and degradation of RLR signaling proteins, not its deISGylase/DUB activity, in type I interferon suppression

Author summary Outbreaks of the picornavirus foot-and-mouth disease virus (FMDV) have significant consequences for animal health and product safety and place a major economic burden on the global livestock industry. Understanding how this notorious animal pathogen suppresses the antiviral type I interferon (IFN-alpha/beta) response may help to develop countermeasures to control FMDV infections. FMDV suppresses the IFN-alpha/beta response through the activity of its Leader protein (L-pro), a protease that can cleave host cell proteins. L(pro)was also shown to have deubiquitinase and deISGylase activity, raising the possibility that L(pro)suppresses IFN-alpha/beta by removing ubiquitin and/or ISG15, two posttranslational modifications that can regulate the activation, interactions and localization of (signaling) proteins. Here, we show that TBK1 and MAVS, two signaling proteins that are important for activation of IFN-alpha/beta gene transcription, are cleaved by L-pro. By generating L(pro)mutants lacking either of these two activities, we demonstrate that L-pro's ability to cleave signaling proteins, but not its deubiquitination/deISGylase activity, correlates with suppression of IFN-beta gene transcription. The type I interferon response is an important innate antiviral pathway. Recognition of viral RNA by RIG-I-like receptors (RLRs) activates a signaling cascade that leads to type I interferon (IFN-alpha/beta) gene transcription. Multiple proteins in this signaling pathway (e.g. RIG-I, MDA5, MAVS, TBK1, IRF3) are regulated by (de)ubiquitination events. Most viruses have evolved mechanisms to counter this antiviral response. The leader protease (L-pro) of foot-and-mouth-disease virus (FMDV) has been recognized to reduce IFN-alpha/beta gene transcription; however, the exact mechanism is unknown. The proteolytic activity of L(pro)is vital for releasing itself from the viral polyprotein and for cleaving and degrading specific host cell proteins, such as eIF4G and NF-kappa B. In addition, L(pro)has been demonstrated to have deubiquitination/deISGylation activity. L-pro's deubiquitination/deISGylation activity and the cleavage/degradation of signaling proteins have both been postulated to be important for reduced IFN-alpha/beta gene transcription. Here, we demonstrate that TBK1, the kinase that phosphorylates and activates the transcription factor IRF3, is cleaved by L(pro)in FMDV-infected cells as well as in cells infected with a recombinant EMCV expressing L-pro.In vitrocleavage experiments revealed that L(pro)cleaves TBK1 at residues 692-694. We also observed cleavage of MAVS in HeLa cells infected with EMCV-L-pro, but only observed decreasing levels of MAVS in FMDV-infected porcine LFPK alpha V beta 6 cells. We set out to dissect L-pro's ability to cleave RLR signaling proteins from its deubiquitination/deISGylation activity, to determine their relative contributions to the reduction of IFN-alpha/beta gene transcription. The introduction of specific mutations, of which several were based on the recently published structure of L(pro)in complex with ISG15, allowed us to identify specific amino acid substitutions that separate the different proteolytic activities of L-pro. Characterization of the effects of these mutations revealed that L-pro's ability to cleave RLR signaling proteins but not its deubiquitination/deISGylation activity correlates with the reduced IFN-beta gene transcription

Immunometabolism pathways as the basis for innovative anti-viral strategies (INITIATE): a Marie Sklodowska-Curie innovative training network

The past century has witnessed major advances in the control of many infectious diseases, yet outbreaks and epidemics caused by (re-) emerging RNA viruses continue to pose a global threat to human health. As illustrated by the global COVID19 pandemic, high healthcare costs, economic disruption and loss of productivity reinforce the unmet medical need to develop new antiviral strategies to combat not only the current pandemic but also future viral outbreaks. Pivotal for effective anti-viral defense is the innate immune system, a first line host response that senses and responds to virus infection. While molecular details of the innate immune response are well characterized, this research field is now being revolutionized with the recognition that cell metabolism has a major impact on the antiviral and inflammatory responses to virus infections. A detailed understanding of the role of metabolic regulation with respect to antiviral and inflammatory responses, together with knowledge of the strategies used by viruses to exploit immunometabolic pathways, will ultimately change our understanding and treatment of pathogenic viral diseases. INITIATE is a Marie Sklodowska-Curie Actions Innovative Training Network (MSCA-ITN), with the goal to train 15 early stage PhD researchers (ESRs) to become experts in antiviral immunometabolism (https://initiate-itn.eu/). To this end, INITIATE brings together a highly complementary international team of academic and corporate leaders from 7 European countries, with outstanding track records in the historically distinct research fields of virology, immunology and metabolism. The ESRs of INITIATE are trained in these interdisciplinary research fields through individual investigator-driven research projects, specialized scientific training events, workshops on academia-industry interactions, outreach &amp; communication. INITIATE will deliver a new generation of creative and entrepreneurial researchers who will be able to face the inevitable future challenges in combating viral diseases

Genus- and species-specific identification of mycoplasmas by 16S rRNA amplification

Systematic computer alignment of mycoplasmal 16S rRNA sequences allowed the identification of variable regions with both genus- and species-specific sequences. Species-specific sequences of Mycoplasma collis were elucidated by asymmetric amplification and dideoxynucleotide sequencing of variable regions, using primers complementary to conserved regions of 16S rRNA. Primers selected for Mycoplasma pneumoniae, M. hominis, M. fermentans, Ureaplasma urealyticum, M. pulmonis, M. arthritidis, M. neurolyticum, M. muris, and M. collis proved to be species specific in the polymerase chain reaction. The genus-specific primers reacted with all mycoplasmal species investigated as well as with members of the genera Ureaplasma, Spiroplasma, and Acholeplasma. No cross-reaction was observed with members of the closely related genera Streptococcus, Lactobacillus, Bacillus, and Clostridium or with any other microorganism tested. On the basis of the high copy number of rRNA, a highly sensitive polymerase chain reaction assay was developed in which the nucleic acid content equivalent to a single organism could be detected

Massive empyema caused by Mycoplasma pneumoniae in an adult: A case report

BACKGROUND: Mycoplasma pneumoniae is responsible for more than 20% of community acquired pneumonia cases, and capable of causing upper respiratory illness as well. Complications of M.pneumoniae infections include CNS involvement but other as pericarditis were also reported. The lack of feasible culture methods and under appreciation of the pathogens ability to cause invasive disease leads to reduced number of diagnosed M.pneumoniae related complications. In contrast to many other respiratory pathogens causing pneumonia, M. pneumoniae related severe pleural complications were almost never reported. CASE PRESENTATION: We report a previously healthy 57 years old woman presented with indolent massive right pleural effusion, leukocytosis and elevated ESR. Extensive microbiological evaluation didn't reveal any pathogen in the pus even before antibiotic treatment was started. Surprisingly, M.pneumoniae DNA was detected in the pus from the empyema using PCR designed to detect M.pneumoniae. A serological assay (Serodia-Myco II) using convalescent serum was indeterminate with a titer of 1:80. The patient responded well to a treatment that included right thoracotomy with pleural decortication and a combination of antibiotics and anti-inflammatory medications. CONCLUSION: M.pneumoniae related empyema was never reported before in adult patients and was reported in only a few pediatric patients. In our patient there was no evidence to any common pathogens even before initiating antibiotic treatment. The only pathogen detected was M.pneumoniae. In this patient, serology was not helpful in establishing the diagnosis of M.pneumoniae related diseases, as was suggested before for older patients. We suggest that M.pneumoniae related empyema is probably under-diagnosed complication due to insensitivity of serology in older patients and under use of other diagnosis methods

Bithiazole Inhibitors of Phosphatidylinositol 4-Kinase (PI4KIIIβ) as Broad-Spectrum Antivirals Blocking the Replication of SARS-CoV-2, Zika Virus, and Human Rhinoviruses

Over half a century since the description of the first antiviral drug, “old” re-emerging viruses and “new” emerging viruses still represent a serious threat to global health. Their high mutation rate and rapid selection of resistance toward common antiviral drugs, together with the increasing number of co-infections, make the war against viruses quite challenging. Herein we report a host-targeted approach, based on the inhibition of the lipid kinase PI4KIIIβ, as a promising strategy for inhibiting the replication of multiple viruses hijacking this protein. We show that bithiazole inhibitors of PI4KIIIβ block the replication of human rhinoviruses (hRV), Zika virus (ZIKV) and SARS-CoV-2 at low micromolar and sub-micromolar concentrations. However, while the anti-hRV/ZIKV activity can be directly linked to PI4KIIIβ inhibition, the role of PI4KIIIβ in SARS-CoV-2 entry/replication is debated

Lack of Detection of XMRV in Seminal Plasma from HIV-1 Infected Men in The Netherlands

Background: Xenotropic murine leukaemia virus-related virus (XMRV) is a recently discovered human gammaretrovirus with yet unknown prevalence and transmission route(s). Its presence in prostate stromal fibroblasts and prostatic secretions suggests that XMRV might be sexually transmitted. We chose to study a compartment closely connected to the prostate, a location where XMRV was detected in independent studies. Seminal plasma samples from HIV-1 infected men were examined as they have an increased probability of acquiring sexually transmitted pathogens. Methodology/Principal Findings: We studied the prevalence of XMRV in 93 seminal plasma samples of 54 HIV-1 infected men living in The Netherlands with a nested PCR amplification specifically targeting the XMRV gag gene. As a control for the presence and integrity of retrovirus particles, HIV-1 was amplified from the same samples with a PCR amplification targeting the env gene of the virus, or HIV-1 was quantified with a real-time PCR amplifying part of the pol gene. Conclusions/Significance: Although HIV-1 was amplified from 25 % of the seminal plasma samples, no XMRV was detected, suggesting that either the prevalence of XMRV is very low in The Netherlands, or that XMRV is not naturally present in th