Regulation of the floral transition at the shoot apical meristem of Arabidopsis as studied by genetics and next generation sequencing

Der Vorgang bei welchem Angiospermen von vegetativem Wachstum zur Bildung von Blüten übergehen wird als Übergang zur Blüte („floral transition“) bezeichnet. Dieser entwicklungsbiologische Vorgang ist von großer Bedeutung und wird streng durch ein genetisches Netzwerk reguliert, wobei einige Komponenten des Netzwerkes auf Umweltfaktoren reagieren. Arabidopsis thaliana blüht früher unter Langtagbedingungen (LD) des Frühlings als unter den Kurztagbedingungen (SD) des Winters. Die Tageslänge oder Photoperiode ist einer der wichtigsten Umweltfaktoren welcher die Blühantwort beeinflusst. Die Photoperiode wird über die Blätter wahrgenommen während der Übergang zur Blüte im apikalen Sprossmeristem (SAM) stattfindet. Unter Langtagbedingungen wird eine genetische Kaskade im Leitgewebe des Blattes angestoßen, woraufhin ein Schlüsseltranskriptionsregulator mit dem Namen CONSTANS das Gen mit dem Namen FLOWERING LOCUS T (FT) und sein Homolog TWIN SISTER OF FT (TSF) aktiviert. Das FT Protein wird daraufhin durch das Phloem transportiert und erreicht schlussendlich das SAM wo es den Übergang zur Blüte auslöst. Durch Bildung eines Komplexes mit FD, einem bZIP Transkriptionsfaktor, aktiviert FT Zielgene wie SUPPRESSOR OF OVEREXPRESSION OF CO (SOC1), FRUITFULL (FUL) und später APETALA1 (AP1), welche für MADS-Box Transkriptionsfaktoren codieren. Der Übergang zur Blüte erfordert jedoch im SAM eine dramatische Neuprogrammierung der Transkriptionsvorgänge. Ein vollständiges Bild der Genexpression welche spezifisch im SAM stattfindet fehlt bislang. Daher wurden im ersten Teil dieser Arbeit apikale Sprossmeristeme durch Lasersezierung aus Pflanzen ausgeschnitten, welche von SD nach LD überführt worden waren. RNA, welche aus den Meristemen isoliert worden war, wurde in cDNA umgeschrieben und die Genexpression durch Next-Generation Sequencing durch RNA-seq quantifiziert. Die Gene wurden nach gesteigerter oder verringerter Expression sortiert, wobei ein besonderes Augenmerk auf neue Gene gelegt wurde deren Expression ähnlich der von SOC1 und FUL gesteigert wurde. Ein Teil dieser Gene wurde über in situ Hybridisierung in Wildtyp-Apizes getestet um ihre Aktivierung im SAM zu bestätigen und ihr räumliches Expressionsmuster aufzuklären. Für mehrere neue Gene konnte die Induktion durch Transfer der Pflanzen in LD bestätigt werden; auch zeigten sie spezifische räumliche Expressionsmuster in zahlreichen Regionen des apikalen Sprossmeristems. Es wurden auch Apizes von ft tsf Doppelmutanten hybridisiert um aufzudecken ob die neuen Gene an der von der Photoperiode abhängige Kaskade folgend auf FT/TSF beteiligt sind. Während viele Gene ähnlich wie SOC1 nur in Anwesenheit von FT/TSF induziert wurden, fanden sich erstaunlicherweise auch Gene welche auch in ft tsf Doppelmutanten noch auf die Photoperiode reagierten. Dies legt nahe, dass zusätzliche unbekannte Signale eine Rolle in der Antwort auf eine induzierende Tageslänge unabhängig von FT und TSF spielen. In der vorliegenden Arbeit werden auch weitere Untersuchungen neuer Gene beschrieben. Im zweiten Teil der vorliegenden Arbeit wurden genetische Ansätze verwendet um die Funktion von SHORT VEGETATIVE PHASE (SVP), einem Blührepressor aus der MADS-Box Familie, zu untersuchen. Hierbei wurden neue Interaktionen mit die Blüte fördernden Genen und spezifische Rollen des SVP Gens in Blättern und Meristem aufgedeckt

I Like the Way You Eat It: Lemur (Indri indri) Gut Mycobiome and Geophagy

Here, we investigated the possible linkages among geophagy, soil characteristics, and gut mycobiome of indri (Indri indri), an endangered lemur species able to survive only in wild conditions. The soil eaten by indri resulted in enriched secondary oxide-hydroxides and clays, together with a high concentration of specific essential micronutrients. This could partially explain the role of the soil in detoxification and as a nutrient supply. Besides, we found that soil subject to geophagy and indris’ faeces shared about 8.9% of the fungal OTUs. Also, several genera (e.g. Fusarium, Aspergillus and Penicillium) commonly associated with soil and plant material were found in both geophagic soil and indri samples. On the contrary, some taxa with pathogenic potentials, such as Cryptococcus, were only found in indri samples. Further, many saprotrophs and plant-associated fungal taxa were detected in the indri faeces. These fungal species may be involved in the digestion processes of leaves and could have a beneficial role in their health. In conclusion, we found an intimate connection between gut mycobiome and soil, highlighting, once again, the potential consequent impacts on the wider habitat. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00248-020-01677-5

Disentangling the Possible Drivers of Indri indri Microbiome: A Threatened Lemur Species of Madagascar

Research on the gut microbiome may help with increasing our understanding of primate health with species’ ecology, evolution, and behavior. In particular, microbiome-related information has the potential to clarify ecology issues, providing knowledge in support of wild primates conservation and their associated habitats. Indri (Indri indri) is the largest extant living lemur of Madagascar. This species is classified as “critically endangered” by the IUCN Red List of Threatened Species, representing one of the world’s 25 most endangered primates. Indris diet is mainly folivorous, but these primates frequently and voluntarily engage in geophagy. Indris have never been successfully bred under human care, suggesting that some behavioral and/or ecological factors are still not considered from the ex situ conservation protocols. Here, we explored gut microbiome composition of 18 indris belonging to 5 different family groups. The most represented phyla were Proteobacteria 40.1 ± 9.5%, Bacteroidetes 28.7 ± 2.8%, Synergistetes 16.7 ± 4.5%, and Firmicutes 11.1 ± 1.9%. Further, our results revealed that bacterial alpha and beta diversity were influenced by indri family group and sex. In addition, we investigated the chemical composition of geophagic soil to explore the possible ecological value of soil as a nutrient supply. The quite acidic pH and high levels of secondary oxide-hydroxides of the soils could play a role in the folivorous diet’s gut detoxification activity. In addition, the high contents of iron and manganese found the soils could act as micronutrients in the indris’ diet. Nevertheless, the concentration of a few elements (i.e., calcium, sulfur, boron, nickel, sodium, and chromium) was higher in non-geophagic than in geophagic soils. In conclusion, the data presented herein provide a baseline for outlining some possible drivers responsible for the gut microbiome diversity in indris, thus laying the foundations for developing further strategies involved in indris’ conservation

Superinfections caused by carbapenem-resistant Enterobacterales in hospitalized patients with COVID-19: a multicentre observational study from Italy (CREVID Study)

Objectives To describe clinical characteristics and outcomes of COVID-19 patients who developed secondary infections due to carbapenem-resistant Enterobacterales (CRE). Methods Retrospective observational study including COVID-19 patients admitted to 12 Italian hospitals from March to December 2020 who developed a superinfection by CRE. Superinfection was defined as the occurrence of documented bacterial infection >48 h from admission. Patients with polymicrobial infections were excluded. Demographic, clinical characteristics and outcome were collected. Isolates were classified as KPC, metallo-beta-lactamase (MBL) and OXA-48-producing CRE. A Cox regression analysis was performed to identify factors independently associated with 30 day mortality. Results Overall, 123 patients (median age 66 years, IQR 59-75) were included. The majority of infections occurred in the ICU (81, 65.9%), while 42 (34.1%) in medical wards. The most common types of infection were bloodstream infections (BSI) (n = 64, 52%), followed by urinary-tract infections (UTI) (n = 28, 22.8%), hospital-acquired/ventilator-associated pneumonia (HAP/VAP) (n = 28, 22.8%), intra-abdominal infections (n = 2, 1.6%) and skin infections (n = 1, 0.8%). Sixty-three (51.2%) infections were caused by KPC-, 54 (43.9%) by MBL-, and 6 (4.8%) by OXA-48-producing CRE. Thirty-day mortality was 33.3% (41/123). On Cox regression analysis, HAP/VAP compared with UTI (HR 7.23, 95% CI 2.09-24.97, P = 0.004), BSI compared with UTI (HR 3.96, 95% CI, 1.33-11.77, P = 0.004), lymphopenia on admission (HR 3, 95% CI 1.44-6.26, P = 0.003) and age (HR 1.05, 95% CI 1.02-1.08, P = 0.002) were predictors of 30 day mortality. Conclusions Superinfections by CRE were associated with high risk of 30 day mortality in patients with COVID-19. HAP/VAP was the strongest predictor of death in these patients

Transient reprogramming of crop plants for agronomic performance

The development of a new crop variety is a time-consuming and costly process due to the reliance of plant breeding on gene shuffling to introduce desired genes into elite germplasm, followed by backcrossing. Here, we propose alternative technology that transiently targets various regulatory circuits within a plant, leading to operator-specified alterations of agronomic traits, such as time of flowering, vernalization requirement, plant height or drought tolerance. We redesigned techniques of gene delivery, amplification and expression around RNA viral transfection methods that can be implemented on an industrial scale and with many crop plants. The process does not involve genetic modification of the plant genome and is thus limited to a single plant generation, is broadly applicable, fast, tunable and versatile, and can be used throughout much of the crop cultivation cycle. The RNA-based reprogramming may be especially useful in plant pathogen pandemics but also for commercial seed production and for rapid adaptation of orphan crops

Enhanced immunological recovery with early start of antiretroviral therapy during acute or early HIV infection–results of Italian Network of ACuTe HIV InfectiON (INACTION) retrospective study

ABSTRACT Background: Viral load peak and immune activation occur shortly after exposure during acute or early HIV infection (AEHI). We aimed to define the benefit of early start of antiretroviral treatment (ART) during AEHI in terms of immunological recovery, virological suppression, and treatment discontinuation. Setting: Patients diagnosed with AEHI (Fiebig stages I-V) during 2008-2014 from an analysis of 20 Italian centers. Methods: This was an observational, retrospective, and multicenter study. We investigated the ef- fect of early ART (defined as initiation within 3 months from AEHI diagnosis) on time to virolog- ical suppression, optimal immunological recovery (defined as CD4 count ≥ 500/μL, CD4 ≥ 30%, and CD4/CD8 ≥ 1), and first-line ART regimen discontinuation by Cox regression analysis. Results: There were 321 patients with AEHI included in the study (82.9% in Fiebig stage III-V). At diagnosis, the median viral load was 5.67 log10 copies/mL and the median CD4 count was 456 cells/μL. Overall, 70.6% of patients started early ART (median time from HIV diagnosis to ART initiation 12 days, IQR 6-27). Higher baseline viral load and AEHI diagnosis during 2012-2014 were independently associated with early ART. HBV co-infection, baseline CD4/CD8 ≥ 1, lower baseline HIV-RNA, and AEHI diagnosis in recent years (2012-2014) were independently associ- ated with a shorter time to virological suppression. Early ART emerged as an independent predic- tor of optimal immunological recovery after adjustment for baseline CD4 (absolute and percent- age count) and CD4/CD8 ratio. The only independent predictor of first-line ART discontinuation was an initial ART regimen including > 3 drugs. Conclusions: In a large cohort of well-characterized patients with AEHI, we confirmed the ben- eficial role of early ART on CD4+ T-cell recovery and on rates of CD4/CD8 ratio normalization. Moreover, we recognized baseline CD4/CD8 ratio as an independent factor influencing time to virological response in the setting of AEHI, thus giving new insights into research of immunolog- ical markers associated with virological control

Trends and correlates of HIV-1 resistance among subjects failing an antiretroviral treatment over the 2003-2012 decade in Italy

BACKGROUND: Despite a substantial reduction in virological failures following introduction of new potent antiretroviral therapies in the latest years, drug resistance remains a limitation for the control of HIV-1 infection. We evaluated trends and correlates of resistance in treatment failing patients in a comprehensive database over a time period of relevant changes in prescription attitudes and treatment guidelines. METHODS: We analyzed 6,796 HIV-1 pol sequences from 49 centres stored in the Italian ARCA database during the 2003-2012 period. Patients (n = 5,246) with viremia > 200 copies/mL received a genotypic test while on treatment. Mutations were identified from IAS-USA 2013 tables. Class resistance was evaluated according to antiretroviral regimens in use at failure. Time trends and correlates of resistance were analyzed by Cochran-Armitage test and logistic regression models. RESULTS: The use of NRTI backbone regimens slightly decreased from 99.7% in 2003-2004 to 97.4% in 2010-2012. NNRTI-based combinations dropped from 46.7% to 24.1%. PI-containing regimens rose from 56.6% to 81.7%, with an increase of boosted PI from 36.5% to 68.9% overtime. In the same reference periods, Resistance to NRTIs, NNRTIs and PIs declined from 79.1% to 40.8%, from 77.8% to 53.8% and from 59.8% to 18.9%, respectively (p < .0001 for all comparisons). Dual NRTI + NNRTI and NRTI + PI resistance decreased from 56.4% to 33.3% and from 36.1% to 10.5%, respectively. Reduced risk of resistance over time periods was confirmed by a multivariate analysis. CONCLUSIONS: Mutations associated with NRTIs, NNRTIs and PIs at treatment failure declined overtime regardless of specific class combinations and epidemiological characteristics of treated population. This is likely due to the improvement of HIV treatment, including both last generation drug combinations and prescription guidelines