Supporting play exploration and early developmental intervention versus usual care to enhance development outcomes during the transition from the neonatal intensive care unit to home: a pilot randomized controlled trial

Background While therapy services may start in the Neonatal Intensive Care Unit (NICU) there is often a gap in therapy after discharge. Supporting Play Exploration and Early Development Intervention (SPEEDI) supports parents, helping them build capacity to provide developmentally supportive opportunities starting in the NICU and continuing at home. The purpose of this single blinded randomized pilot clinical trial was to evaluate the initial efficacy of SPEEDI to improve early reaching and exploratory problem solving behaviors. Methods Fourteen infants born very preterm or with neonatal brain injury were randomly assigned to SPEEDI or Usual Care. The SPEEDI group participated in 5 collaborative parent, therapist, and infant interventions sessions in the NICU (Phase 1) and 5 at home (Phase 2). Parents provided daily opportunities designed to support the infants emerging motor control and exploratory behaviors. Primary outcome measures were assessed at the end of the intervention, 1 and 3 months after the intervention ended. Reaching was assessed with the infant supported in an infant chair using four 30 s trials. The Early Problem Solving Indicator was used to evaluate the frequency of behaviors during standardized play based assessment. Effect sizes are including for secondary outcomes including the Test of Infant Motor Performance and Bayley Scales of Infant and Toddler Development. Results No group differences were found in the duration of toy contact. There was a significant group effect on (F1,8 = 4.04, p = 0.08) early exploratory problem-solving behaviors with infants in the SPEEDI group demonstrating greater exploration with effect sizes of 1.3, 0.6, and 0.9 at the end of the intervention, 1 and 3 months post-intervention. Conclusions While further research is needed, this initial efficacy study showed promising results for the ability of SPEEDI to impact early problem solving behaviors at the end of intervention and at least 3 months after the intervention is over. While reaching did not show group differences, a ceiling effect may have contributed to this finding. This single blinded pilot RCT was registered prior to subject enrollment on 5/27/14 at ClinicalTrials.Gov with number NCT02153736

Insecticidal genes of Yersinia spp.: taxonomical distribution, contribution to toxicity towards Manduca sexta and Galleria mellonella, and evolution

<p>Abstract</p> <p>Background</p> <p>Toxin complex (Tc) proteins termed TcaABC, TcdAB, and TccABC with insecticidal activity are present in a variety of bacteria including the yersiniae.</p> <p>Results</p> <p>The <it>tc </it>gene sequences of thirteen <it>Yersinia </it>strains were compared, revealing a high degree of gene order conservation, but also remarkable differences with respect to pseudogenes, sequence variability and gene duplications. Outside the <it>tc </it>pathogenicity island (<it>tc</it>-PAI^<it>Ye</it>) of <it>Y. enterocolitica </it>strain W22703, a pseudogene (<it>tccC2'</it>/<it>3'</it>) encoding proteins with homology to TccC and similarity to tyrosine phosphatases at its C-terminus was identified. PCR analysis revealed the presence of the <it>tc</it>-PAI^{<it>Ye </it>}and of <it>tccC2'</it>/<it>3'</it>-homologues in all biotype 2–5 strains tested, and their absence in most representatives of biotypes 1A and 1B. Phylogenetic analysis of 39 TccC sequences indicates the presence of the <it>tc</it>-PAI^{<it>Ye </it>}in an ancestor of <it>Yersinia</it>. Oral uptake experiments with <it>Manduca sexta </it>revealed a higher larvae lethality of <it>Yersinia </it>strains harbouring the <it>tc</it>-PAI^{<it>Ye </it>}in comparison to strains lacking this island. Following subcutaneous infection of <it>Galleria mellonella </it>larvae with five non-human pathogenic <it>Yersinia </it>spp. and four <it>Y. enterocolitica </it>strains, we observed a remarkable variability of their insecticidal activity ranging from 20% (<it>Y. kristensenii</it>) to 90% (<it>Y. enterocolitica </it>strain 2594) dead larvae after five days. Strain W22703 and its <it>tcaA </it>deletion mutant did not exhibit a significantly different toxicity towards <it>G. mellonella</it>. These data confirm a role of TcaA upon oral uptake only, and suggest the presence of further insecticidal determinants in <it>Yersinia </it>strains formerly unknown to kill insects.</p> <p>Conclusion</p> <p>This study investigated the <it>tc </it>gene distribution among yersiniae and the phylogenetic relationship between TccC proteins, thus contributing novel aspects to the current discussion about the evolution of insecticidal toxins in the genus <it>Yersinia</it>. The toxic potential of several <it>Yersinia </it>spp. towards <it>M. sexta </it>and <it>G. mellonella </it>demonstrated here for the first time points to insects as a natural reservoir for yersiniae.</p

A beta-herpesvirus with fluorescent capsids to study transport in living cells.

Fluorescent tagging of viral particles by genetic means enables the study of virus dynamics in living cells. However, the study of beta-herpesvirus entry and morphogenesis by this method is currently limited. This is due to the lack of replication competent, capsid-tagged fluorescent viruses. Here, we report on viable recombinant MCMVs carrying ectopic insertions of the small capsid protein (SCP) fused to fluorescent proteins (FPs). The FPs were inserted into an internal position which allowed the production of viable, fluorescently labeled cytomegaloviruses, which replicated with wild type kinetics in cell culture. Fluorescent particles were readily detectable by several methods. Moreover, in a spread assay, labeled capsids accumulated around the nucleus of the newly infected cells without any detectable viral gene expression suggesting normal entry and particle trafficking. These recombinants were used to record particle dynamics by live-cell microscopy during MCMV egress with high spatial as well as temporal resolution. From the resulting tracks we obtained not only mean track velocities but also their mean square displacements and diffusion coefficients. With this key information, we were able to describe particle behavior at high detail and discriminate between particle tracks exhibiting directed movement and tracks in which particles exhibited free or anomalous diffusion

Degradation of Cellular miR-27 by a Novel, Highly Abundant Viral Transcript Is Important for Efficient Virus Replication In Vivo

Cytomegaloviruses express large amounts of viral miRNAs during lytic infection, yet, they only modestly alter the cellular miRNA profile. The most prominent alteration upon lytic murine cytomegalovirus (MCMV) infection is the rapid degradation of the cellular miR-27a and miR-27b. Here, we report that this regulation is mediated by the ∼1.7 kb spliced and highly abundant MCMV m169 transcript. Specificity to miR-27a/b is mediated by a single, apparently optimized, miRNA binding site located in its 3'-UTR. This site is easily and efficiently retargeted to other cellular and viral miRNAs by target site replacement. Expression of the 3'-UTR of m169 by an adenoviral vector was sufficient to mediate its function, indicating that no other viral factors are essential in this process. Degradation of miR-27a/b was found to be accompanied by 3'-tailing and -trimming. Despite its dramatic effect on miRNA stability, we found this interaction to be mutual, indicating potential regulation of m169 by miR-27a/b. Most interestingly, three mutant viruses no longer able to target miR-27a/b, either due to miRNA target site disruption or target site replacement, showed significant attenuation in multiple organs as early as 4 days post infection, indicating that degradation of miR-27a/b is important for efficient MCMV replication in vivo

Cytomegalovirus microRNAs Facilitate Persistent Virus Infection in Salivary Glands

Micro (mi)RNAs are small non-coding RNAs that regulate the expression of their targets' messenger RNAs through both translational inhibition and regulation of target RNA stability. Recently, a number of viruses, particularly of the herpesvirus family, have been shown to express their own miRNAs to control both viral and cellular transcripts. Although some targets of viral miRNAs are known, their function in a physiologically relevant infection remains to be elucidated. As such, no in vivo phenotype of a viral miRNA knock-out mutant has been described so far. Here, we report on the first functional phenotype of a miRNA knock-out virus in vivo. During subacute infection of a mutant mouse cytomegalovirus lacking two viral miRNAs, virus production is selectively reduced in salivary glands, an organ essential for virus persistence and horizontal transmission. This phenotype depends on several parameters including viral load and mouse genetic background, and is abolished by combined but not single depletion of natural killer (NK) and CD4+ T cells. Together, our results point towards a miRNA-based immunoevasion mechanism important for long-term virus persistence

Transcriptional Activation of the Adenoviral Genome Is Mediated by Capsid Protein VI

Gene expression of DNA viruses requires nuclear import of the viral genome. Human Adenoviruses (Ads), like most DNA viruses, encode factors within early transcription units promoting their own gene expression and counteracting cellular antiviral defense mechanisms. The cellular transcriptional repressor Daxx prevents viral gene expression through the assembly of repressive chromatin remodeling complexes targeting incoming viral genomes. However, it has remained unclear how initial transcriptional activation of the adenoviral genome is achieved. Here we show that Daxx mediated repression of the immediate early Ad E1A promoter is efficiently counteracted by the capsid protein VI. This requires a conserved PPxY motif in protein VI. Capsid proteins from other DNA viruses were also shown to activate the Ad E1A promoter independent of Ad gene expression and support virus replication. Our results show how Ad entry is connected to transcriptional activation of their genome in the nucleus. Our data further suggest a common principle for genome activation of DNA viruses by counteracting Daxx related repressive mechanisms through virion proteins

Supporting play exploration and early developmental intervention versus usual care to enhance development outcomes during the transition from the neonatal intensive care unit to home: a pilot randomized controlled trial

Abstract Background While therapy services may start in the Neonatal Intensive Care Unit (NICU) there is often a gap in therapy after discharge. Supporting Play Exploration and Early Development Intervention (SPEEDI) supports parents, helping them build capacity to provide developmentally supportive opportunities starting in the NICU and continuing at home. The purpose of this single blinded randomized pilot clinical trial was to evaluate the initial efficacy of SPEEDI to improve early reaching and exploratory problem solving behaviors. Methods Fourteen infants born very preterm or with neonatal brain injury were randomly assigned to SPEEDI or Usual Care. The SPEEDI group participated in 5 collaborative parent, therapist, and infant interventions sessions in the NICU (Phase 1) and 5 at home (Phase 2). Parents provided daily opportunities designed to support the infants emerging motor control and exploratory behaviors. Primary outcome measures were assessed at the end of the intervention, 1 and 3 months after the intervention ended. Reaching was assessed with the infant supported in an infant chair using four 30 s trials. The Early Problem Solving Indicator was used to evaluate the frequency of behaviors during standardized play based assessment. Effect sizes are including for secondary outcomes including the Test of Infant Motor Performance and Bayley Scales of Infant and Toddler Development. Results No group differences were found in the duration of toy contact. There was a significant group effect on (F1,8 = 4.04, p = 0.08) early exploratory problem-solving behaviors with infants in the SPEEDI group demonstrating greater exploration with effect sizes of 1.3, 0.6, and 0.9 at the end of the intervention, 1 and 3 months post-intervention. Conclusions While further research is needed, this initial efficacy study showed promising results for the ability of SPEEDI to impact early problem solving behaviors at the end of intervention and at least 3 months after the intervention is over. While reaching did not show group differences, a ceiling effect may have contributed to this finding. This single blinded pilot RCT was registered prior to subject enrollment on 5/27/14 at ClinicalTrials.Gov with number NCT02153736

Additional file 3: of Supporting play exploration and early developmental intervention versus usual care to enhance development outcomes during the transition from the neonatal intensive care unit to home: a pilot randomized controlled trial

SPEEDI Activity Booklet. Includes the text from the SPEEDI activity booklet provided to parents toward the end of phase 1, for implementation in phase 2. Parents used the activity log in this appendix to document which activities were completed each day during Phase 2 of the SPEEDI intervention. (DOCX 19 kb

Post-transcriptional regulation of miR-27 in murine cytomegalovirus infection

In mammals, microRNAs (miRNAs) can play diverse roles in viral infection through their capacity to regulate both host and viral genes. Recent reports have demonstrated that specific miRNAs change in expression level upon infection and can impact viral production and infectivity. It is clear that miRNAs are an integral component of viral–host interactions, and it is likely that both host and virus contain mechanisms to regulate miRNA expression and/or activity. To date, little is known about the mechanisms by which miRNAs are regulated in viral infection. Here we report the rapid down-regulation of miR-27a in multiple mouse cell lines as well as primary macrophages upon infection with the murine cytomegalovirus. Down-regulation of miR-27a occurs independently from two other miRNAs, miR-23a and miR-24, located within the same genomic cluster, and analysis of pri-miRNA levels suggest that regulation occurs post-transcriptionally. miR-27b, a close homolog of miR-27a (20/21 nucleotide identity), also decreases upon infection, and we demonstrate that both miR-27a and miR-27b exert an antiviral function upon over-expression. Drug sensitivity experiments suggest that virus entry is not sufficient to induce the down-regulation of miR-27 and that the mechanism requires synthesis of RNA. Altogether, our findings indicate that miR-27a and miR-27b have antiviral activity against MCMV, and that either the virus or the host encodes molecule(s) for regulating miR-27 accumulation, most likely by inducing the rapid decay of the mature species