Cytotype variation and clonal diversity in polyploid apomictic populations of Pilosella (Compositeae, Cichorieae) introduced to southern Patagonia

Introduction and objectives: The members of the genus Pilosella are native in Europe and Asia, but they are successful invasive species on most continents. These species form an agamic complex with common apomixis. Apomictic species hybridize, they have different degree of residual sexuality. Main aim of this paper was to determine if the interspecific hybridization already occurred in Patagonia. M&M: This study is based on analysis of seed progeny collected at thirteen populations of Pilosella in southern Argentina and Chile. The plants were examined for their taxonomic identity, DNA ploidy level (using flow cytometry), chromosome number, reproduction, formation of parthenogenetic seeds and clonal identity (using isozyme phenotypes). Results: No mixed-species population was recorded. Two apomictic clones of P. officinarum (one pentaploid and the other hexaploid) were found in populations: eight were hexaploid and one was mixed in cytotype composition. A new species for Patagonia, the apomictic pentaploid P. caespitosa, was represented by plants from two populations in Argentina. Some of the progeny plants cultivated from seeds sampled at three localities represented seed-fertile aneuploids the morphology of which implied a hybrid origin and indicated P. officinarum as one of the parents. Conclusions: The presence of seed-fertile, aneuploid and parthenogenetic hybrids among the cultivated plants signifies an increased risk of the formation of new hybridogeneous genotypes of Pilosella in southern Patagonia.

Visual Barcodes for Multiplexing Live Microscopy-Based Assays

While multiplexing samples using DNA barcoding revolutionized the pace of biomedical discovery, multiplexing of live imaging-based applications has been limited by the number of fluorescent proteins that can be deconvoluted using common microscopy equipment. To address this limitation we developed visual barcodes that discriminate the clonal identity of single cells by targeting different fluorescent proteins to specific subcellular locations. We demonstrate that deconvolution of these barcodes is highly accurate and robust to many cellular perturbations. We then used visual barcodes to generate ‘Signalome’ cell-lines by multiplexing live reporters to monitor the simultaneous activity in 12 branches of signaling, in live cells, at single cell resolution, over time. Using the ‘Signalome’ we identified two distinct clusters of signaling pathways that balance growth and proliferation, emphasizing the importance of growth homeostasis as a central organizing principle in cancer signaling. The ability to multiplex samples in live imaging applications, both in vitro and in vivo may allow better high-content characterization of complex biological system

Longitudinal monitoring of sinonasal and oral bacterial reservoirs to prevent chronic lung infection in people with cystic fibrosis

BackgroundParanasal sinuses act as bacterial reservoirs and contribute to transmitting bacteria to the lower airway of patients with cystic fibrosis (CF). Also, passage of bacteria from the oral cavity to the lungs may occur.MethodsWe evaluated the presence of Pseudomonas aeruginosa, Staphylococcus aureus, Stenotrophomonas maltophilia, Achromobacter xylosoxidans and Serratia marcescens in sputum and nasal lavage of 59 patients with CF, and also collected saliva and used toothbrushes from 38 of them. We assessed the clonal identity of the strains isolated from the different samples by pulsed-field gel electrophoresis.ResultsAbout 80% of the patients were positive for at least one of the bacterial species examined in nasal lavage and sputum. Among the subjects with positive sputum, 74% presented the same species in the nasal lavage and saliva, and 26% on their toothbrush. S. aureus was the most abundant species in all samples. Clonal identity (≥80% similarity) of the strains isolated among the different samples from each patient was confirmed in almost all cases. Longitudinal observation helped to identify five patients who were colonised in the lower airways after an initial period of nasal or oral colonisation.ConclusionNasal and oral sites act as bacterial reservoirs, favouring the transmission of potentially pathogenic microorganisms to the lower airway. The lack of eradication from these sites might undermine the antibiotic therapy applied to treat the lung infection, allowing the persistence of the bacteria within the patient if colonisation of these sites is not assessed, and no specific therapy is performed.

Validating Blackberry Seedling Pedigrees and Developing an Improved Multiplexed Microsatellite Fingerprinting Set

Confirming parentage and clonal identity is an important aspect of breeding and managing germplasm collections of clonally propagated, outcrossing crops, like blackberry (Rubus subgenus Rubus). DNA fingerprinting sets are used to identify off-cross progeny and confirm clonal identity. Previously, a six-simple sequence repeat (6-SSR) fingerprinting set was developed for blackberry using a small number of samples. The usefulness of the 6-SSR fingerprinting set for pedigree confirmation had not been evaluated. Therefore, it was used in this study to validate parentage for 6 and 12 biparental populations from the University of Arkansas (UA) and US Department of Agriculture Agricultural Research Service (USDA-ARS), Horticultural Crops Research Unit (HCRU) breeding programs, respectively. Twenty-seven of the 489 individuals in these breeding populations were identified as off-cross. The 6-SSR fingerprinting set was sufficient for parentage confirmation; however, a total of 61 plants distributed across 28 sets of genotypes could not be distinguished from each other. An 8-SSR fingerprinting set with improved resolution was subsequently developed and used to evaluate 177 Rubus accessions from the USDA-ARS National Clonal Germplasm Repository, UA, and USDA-ARS HCRU programs. The 8-SSR fingerprinting set distinguished all samples expected to have unique genotypes and identified differing DNA fingerprints for two sets of accessions suspected to have identical fingerprints. Cluster analysis grouped the accessions from the eastern and western US breeding programs based on geography and descent. Future work will focus on establishing a database of DNA fingerprints for germplasm identification and for determining pedigree relationships between blackberry accessions.

Persisting transmission of carbapenemase-producing Klebsiella pneumoniae due to an environmental reservoir in a university hospital, France, 2012 to 2014

In France, the proportion of episodes of carbapenemase-producing Enterobacteriaceae (CPE) with no recent stay or hospitalisation abroad is increasing. In this study, we investigate epidemiological links between apparently unrelated cases of OXA-48-producing Klebsiella pneumoniae (Kp OXA-48) colonisation or infection. We genotyped detected organisms by repetitive sequence-based PCR, and used a dynamic registry of cases and contacts to cross-reference patients' hospital stays. Between 1 November 2012 and 28 February 2014, 23 Kp OXA-48 cases were detected in a university hospital in Montpellier, of which 15 were involved in three outbreaks: outbreaks I and II occurred in November 2012 and outbreak III in October 2013. Molecular comparison of bacterial strains revealed clonal identity between cases involved in outbreaks II and III and four single cases. Cross-referencing of hospital stays revealed that these single cases and the index case of outbreak III had occupied the same room. Active case search among former occupants of that room found an additional Kp OXA-48 carrier. A clonal strain was isolated from the sink of that room. The epidemiological link between the contaminated room and outbreak II remained undetected. This study is a reminder that environmental reservoirs should be considered as a source of CPE transmission.

Increase in paratyphoid fever cases in Japanese travellers returning from Cambodia in 2013

SUMMARYIn 2013, an unusual increase of paratyphoid fever cases in travellers returning from Cambodia was reported in Japan. From December 2012 to September 2013, 18 cases ofSalmonellaParatyphi A infection were identified. Microbiological analyses revealed that most isolates had the same clonal identity, although the epidemiological link between these cases remains unclear. It was inferred that the outbreak was caused by a common and persistent source in Cambodia that was likely to have continued during 2014. The information of surveillance and laboratory data from cases arising in travellers from countries with limited surveillance systems should be timely shared with the country of origin.

Transdifferentiation of Malignant B-Cells into Macrophages in a Murine Model of Burkitt´s Lymphoma

B-cell lymphomas, such as Burkitt's lymphoma, are malignant diseases of the hematopoietic system. They arise from transformed B-cells originating from diverse primary and secondary lymphatic tissues. Like in other cancers, the stroma of lymphomas is typically infiltrated by so-called tumor-associated macrophages (TAMs), which execute manifold tumor-specific functions. Interestingly it was shown, that mature B-cells could be efficiently re-programmed into macrophages by the overexpression of myeloid-specific transcription factors. Moreover, other studies observed in vitro that this lymphoid/myeloid plasticity might be also caused by oncogenes in cultured B-cells of murine lymphoma models. Therefore we consider, if lymphoma B-cells themselves might be a source of TAMs, besides the well-known infiltration of monocytic cells into the tumor environment. Based on the system of CD45.1/2 allotypes, a murine model of lymphoma was thus developed, which allows tracking of the conversion of lymphoma B-cells into TAMs. It could be shown, that some lymphoma B-cells of the established model in fact spontaneously switched into a macrophage-like phenotype. Accordingly, they start to express typical macrophage markers, but seem to perform a transition into a myeloid-like expression profile on the level of transcription as well. Furthermore, analysis of the recombined immunoglobulin heavy chain confirmed the clonal identity of lymphoma b-cells and TAMs. Although these cells do not exhibit an overt immunological phenotype, they show elementary functional properties of macrophages, such as phagocytosis as well as a post-mitotic character. Moreover, transdifferentiated B-cells cells were resistant to chemotherapy and persist after treatment, while lymphoma B-cells were eradicated. Which consequences the lymphoid/myeloid plasticity of lymphoma B-cells has for tumor development, progression, as well as for the outcome of therapy, and if these observations could be transferred to human tumors, will be object of future investigations. Disclosures No relevant conflicts of interest to declare.

Transient Abnormal Myelopoiesis in Neonates: GATA Get the Diagnosis

Transient abnormal myelopoiesis occurs exclusively in patients with Down syndrome (constitutional trisomy 21), manifests in the neonatal period, and is characterized by circulating megakaryoblasts with varied degrees of multisystem organ involvement. In most cases, this process resolves spontaneously by 3 to 6 months of age, but for some, the disease can be fatal. Affected patients are particularly prone to develop acute megakaryoblastic leukemia in early childhood. Somatic GATA1 mutations are believed to be pivotal in the development of transient abnormal myelopoiesis and have proven to be a marker of clonal identity in its evolution to megakaryoblastic leukemia. We describe a study case of transient abnormal myelopoiesis and review the clinical manifestations, laboratory features, natural history, molecular genetics, and postulated disease pathogenesis of this disorder.