Conjugative transfer of the Lactococcus lactis chromosomal sex factor promotes dissemination of the Ll.LtrB group II intron

The Ll.LtrB group II intron from the low-G+C gram-positive bacterium Lactococcus lactis was the first bacterial group II intron shown to splice and mobilize in vivo. This retroelement interrupts the relaxase gene (ltrB) of three L. lactis conjugative elements: plasmids pRS01 and pAH90 and the chromosomal sex factor. Conjugative transfer of a plasmid harboring a segment of the pRS01 conjugative plasmid including the Ll.LtrB intron allows dissemination of Ll.LtrB among L. lactis strains and lateral transfer of this retroelement from L. lactis to Enterococcus faecalis. Here we report the dissemination of the Ll.LtrB group II intron among L. lactis strains following conjugative transfer of the native chromosomally embedded L. lactis sex factor. We demonstrated that Ll.LtrB dissemination is highly variable and often more efficient from this integrative and conjugative element than from an engineered conjugative plasmid. Cotransfer among L. lactis strains of both Ll.LtrB-containing elements, the conjugative plasmid and the sex factor, was detected and shown to be synergistic. Moreover, following their concurrent transfer, both mobilizable elements supported the spread of their respective copies of the Ll.LtrB intron. Our findings explain the unusually high efficiency of Ll.LtrB mobility observed following conjugation of intron-containing plasmids.     

Comparative Heterochromatin Profiling Reveals Conserved and Unique Epigenome Signatures Linked to Adaptation and Development of Malaria Parasites

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Postnatal periodontal ligament as a novel adult stem cell source for regenerative corneal cell therapy

Corneal opacities are a leading cause of global blindness. They are conventionally treated by the transplantation of donor corneal tissue, which is, restricted by a worldwide donor material shortage and allograft rejection. Autologous adult stem cells with a potential to differentiate into corneal stromal keratocytes (CSKs) could offer a suitable choice of cells for regenerative cell therapy. Postnatal periodontal ligament (PDL) contains a population of adult stem cells, which has a similar embryological origin as CSK, that is cranial neural crest. We harvested PDL cells from young adult teeth extracted because of non‐functional or orthodontic reason and differentiated them towards CSK phenotype using a two‐step protocol with spheroid formation followed by growth factor and cytokine induction in a stromal environment (human amnion stroma and porcine corneal stroma). Our results showed that the PDL‐differentiated CSK‐like cells expressed CSK markers (CD34, ALDH3A1, keratocan, lumican, CHST6, B3GNT7 and Col8A2) and had minimal expression of genes related to fibrosis and other lineages (vasculogenesis, adipogenesis, myogenesis, epitheliogenesis, neurogenesis and hematogenesis). Introduction of PDL spheroids into the stroma of porcine corneas resulted in extensive migration of cells inside the host stroma after 14‐day organ culture. Their quiescent nature and uniform cell distribution resembled to that of mature CSKs inside the native stroma. Our results demonstrated the potential translation of PDL cells for regenerative corneal cell therapy for corneal opacities.     

Interspecific diversity reduces and functionally substitutes for intraspecific variation in biofilm communities

Diversity has a key role in the dynamics and resilience of communities and both interspecific (species) and intraspecific (genotypic) diversity can have important effects on community structure and function. However, a critical and unresolved question for understanding the ecology of a community is to what extent these two levels of diversity are functionally substitutable? Here we show, for a mixed-species biofilm community composed of Pseudomonas aeruginosa, P. protegens and Klebsiella pneumoniae, that increased interspecific diversity reduces and functionally substitutes for intraspecific diversity in mediating tolerance to stress. Biofilm populations generated high percentages of genotypic variants, which were largely absent in biofilm communities. Biofilms with either high intra- or interspecific diversity were more tolerant to SDS stress than biofilms with no or low diversity. Unexpectedly, genotypic variants decreased the tolerance of biofilm communities when experimentally introduced into the communities. For example, substituting P. protegens wild type with its genotypic variant within biofilm communities decreased SDS tolerance by twofold, apparently due to perturbation of interspecific interactions. A decrease in variant frequency was also observed when biofilm populations were exposed to cell-free effluents from another species, suggesting that extracellular factors have a role in selection against the appearance of intraspecific variants. This work demonstrates the functional substitution of inter- and intraspecific diversity for an emergent property of biofilms. It also provides a potential explanation for a long-standing paradox in microbiology, in which morphotypic variants are common in laboratory grown biofilm populations, but are rare in diverse, environmental biofilm communities.     

Patient Experience and Satisfaction with Inpatient Service: Development of Short Form Survey Instrument Measuring the Core Aspect of Inpatient Experience

Patient experience reflects quality of care from the patients’ perspective; therefore, patients’ experiences are important data in the evaluation of the quality of health services. The development of an abbreviated, reliable and valid instrument for measuring inpatients’ experience would reflect the key aspect of inpatient care from patients’ perspective as well as facilitate quality improvement by cultivating patient engagement and allow the trends in patient satisfaction and experience to be measured regularly. The study developed a short-form inpatient instrument and tested its ability to capture a core set of inpatients’ experiences. The Hong Kong Inpatient Experience Questionnaire (HKIEQ) was established in 2010; it is an adaptation of the General Inpatient Questionnaire of the Care Quality Commission created by the Picker Institute in United Kingdom. This study used a consensus conference and a cross-sectional validation survey to create and validate a short-form of the Hong Kong Inpatient Experience Questionnaire (SF-HKIEQ). The short-form, the SF-HKIEQ, consisted of 18 items derived from the HKIEQ. The 18 items mainly covered relational aspects of care under four dimensions of the patient’s journey: hospital staff, patient care and treatment, information on leaving the hospital, and overall impression. The SF-HKIEQ had a high degree of face validity, construct validity and internal reliability. The validated SF-HKIEQ reflects the relevant core aspects of inpatients’ experience in a hospital setting. It provides a quick reference tool for quality improvement purposes and a platform that allows both healthcare staff and patients to monitor the quality of hospital care over time.     

Myeloma with involvement of the serous cavities. Cytologic and immunochemical diagnosis and literature review

The significance of serous cavity involvement by myeloma was evaluated in two patients with pleural involvement and two with peritoneal involvement. The involvement occurred at presentation in two patients and after the diagnosis of myeloma in two. The effusions were bloody exudates containing numerous atypical plasma cells. The diagnosis of cavitary involvement was made by morphologic examination of air-dried smears of the effusions, supplemented by the immunocytochemical demonstration of monoclonal proliferation of the plasma cells. In all four cases, these cells contained cytoplasmic kappa light chain immunoglobulins; many of them also stained positively for epithelial membrane antigen. It was best to interpret these immunocytochemical findings with those of the morphologic and additional immunocytochemical studies; the best results for studies for cytoplasmic immunoglobulins were obtained only if the cells in the effusions were washed before they were used for smear preparations and staining. The four patients responded poorly to treatment, dying 12 days, 16 months, 1 month and 10 days after cavitary involvement was recognized. Review of the literature confirmed that the findings in these cases were similar to those in other cases. Cavitary involvement by myeloma carries an ominous prognosis; an accurate recognition of the plasma cells by morphologic and immunocytochemical studies provides the best method of diagnosing cavitary involvement of myeloma and of predicting the poor outcome in such patients.     

"The orchids have been a splendid sport"--an alternative look at Charles Darwin's contribution to orchid biology

Charles Darwin's work with orchids and his thoughts about them are of great interest and not a little pride for those who are interested in these plants, but they are generally less well known than some of his other studies and ideas. Much has been published on what led to his other books and views. However, there is a paucity of information in the general literature on how Darwin's orchid book came about. This review will describe how The Various Contrivances by Which Orchids Are Fertilised by Insects came into being and will discuss the taxonomy of the orchids he studied. It also will concentrate on some of the less well-known aspects of Darwin's work and observations on orchids-namely, rostellum, seeds and their germination, pollination effects, and resupination-and their influence on subsequent investigators, plant physiology, and orchid science.     

Dissection and Culture of Commissural Neurons from Embryonic Spinal Cord

Commissural neurons have been widely used to investigate the mechanisms underlying axon guidance during embryonic spinal cord development. The cell bodies of these neurons are located in the dorsal spinal cord and their axons follow stereotyped trajectories during embryonic development. Commissural axons initially project ventrally towards the floorplate. After crossing the midline, these axons turn anteriorly and project towards the brain. Each of these steps is regulated by the action of several guidance cues. Cultures highly enriched in commissural neurons are ideally suited for many experiments addressing the mechanisms of axon pathfinding, including turning assays, immunochemistry and biochemistry. Here, we describe a method to dissect and culture commissural neurons from E13 rat dorsal spinal cord. First, the spinal cord is isolated and dorsal strips are dissected out. The dorsal tissue is then dissociated into a cell suspension by trypsinization and mechanical disruption. Neurons are plated onto poly-L-lysine-coated glass coverslips or tissue-culture dishes. After 30 hours in vitro, most neurons have extended an axon. The purity of the culture (Yam et al. 2009), typically over 90%, can be assessed by immunolabeling with the commissural neuron markers DCC, LH2 and TAG1 (Helms and Johnson, 1998). This neuronal preparation is a useful tool for in vitro studies of the cellular and molecular mechanisms of commissural axon growth and guidance during spinal cord development.Download video file.^{(145M, mp4)}