SARS-CoV-2 lineage B.1.1.7 is associated with greater disease severity among hospitalised women but not men: multicentre cohort study.

BACKGROUND: SARS-CoV-2 lineage B.1.1.7 has been associated with an increased rate of transmission and disease severity among subjects testing positive in the community. Its impact on hospitalised patients is less well documented. METHODS: We collected viral sequences and clinical data of patients admitted with SARS-CoV-2 and hospital-onset COVID-19 infections (HOCIs), sampled 16 November 2020 to 10 January 2021, from eight hospitals participating in the COG-UK-HOCI study. Associations between the variant and the outcomes of all-cause mortality and intensive therapy unit (ITU) admission were evaluated using mixed effects Cox models adjusted by age, sex, comorbidities, care home residence, pregnancy and ethnicity. FINDINGS: Sequences were obtained from 2341 inpatients (HOCI cases=786) and analysis of clinical outcomes was carried out in 2147 inpatients with all data available. The HR for mortality of B.1.1.7 compared with other lineages was 1.01 (95% CI 0.79 to 1.28, p=0.94) and for ITU admission was 1.01 (95% CI 0.75 to 1.37, p=0.96). Analysis of sex-specific effects of B.1.1.7 identified increased risk of mortality (HR 1.30, 95% CI 0.95 to 1.78, p=0.096) and ITU admission (HR 1.82, 95% CI 1.15 to 2.90, p=0.011) in females infected with the variant but not males (mortality HR 0.82, 95% CI 0.61 to 1.10, p=0.177; ITU HR 0.74, 95% CI 0.52 to 1.04, p=0.086). INTERPRETATION: In common with smaller studies of patients hospitalised with SARS-CoV-2, we did not find an overall increase in mortality or ITU admission associated with B.1.1.7 compared with other lineages. However, women with B.1.1.7 may be at an increased risk of admission to intensive care and at modestly increased risk of mortality.This report was produced by members of the COG-UK-HOCI Variant substudy consortium. COG-UK-HOCI is part of COG-UK. COG-UK is supported by funding from the Medical Research Council (MRC) part of UK Research & Innovation (UKRI), the National Institute of Health Research (NIHR) and Genome Research Limited, operating as the Wellcome Sanger Institute

Global age-sex-specific mortality, life expectancy, and population estimates in 204 countries and territories and 811 subnational locations, 1950–2021, and the impact of the COVID-19 pandemic: a comprehensive demographic analysis for the Global Burden of Disease Study 2021

Background: Estimates of demographic metrics are crucial to assess levels and trends of population health outcomes. The profound impact of the COVID-19 pandemic on populations worldwide has underscored the need for timely estimates to understand this unprecedented event within the context of long-term population health trends. The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 provides new demographic estimates for 204 countries and territories and 811 additional subnational locations from 1950 to 2021, with a particular emphasis on changes in mortality and life expectancy that occurred during the 2020–21 COVID-19 pandemic period. Methods: 22 223 data sources from vital registration, sample registration, surveys, censuses, and other sources were used to estimate mortality, with a subset of these sources used exclusively to estimate excess mortality due to the COVID-19 pandemic. 2026 data sources were used for population estimation. Additional sources were used to estimate migration; the effects of the HIV epidemic; and demographic discontinuities due to conflicts, famines, natural disasters, and pandemics, which are used as inputs for estimating mortality and population. Spatiotemporal Gaussian process regression (ST-GPR) was used to generate under-5 mortality rates, which synthesised 30 763 location-years of vital registration and sample registration data, 1365 surveys and censuses, and 80 other sources. ST-GPR was also used to estimate adult mortality (between ages 15 and 59 years) based on information from 31 642 location-years of vital registration and sample registration data, 355 surveys and censuses, and 24 other sources. Estimates of child and adult mortality rates were then used to generate life tables with a relational model life table system. For countries with large HIV epidemics, life tables were adjusted using independent estimates of HIV-specific mortality generated via an epidemiological analysis of HIV prevalence surveys, antenatal clinic serosurveillance, and other data sources. Excess mortality due to the COVID-19 pandemic in 2020 and 2021 was determined by subtracting observed all-cause mortality (adjusted for late registration and mortality anomalies) from the mortality expected in the absence of the pandemic. Expected mortality was calculated based on historical trends using an ensemble of models. In location-years where all-cause mortality data were unavailable, we estimated excess mortality rates using a regression model with covariates pertaining to the pandemic. Population size was computed using a Bayesian hierarchical cohort component model. Life expectancy was calculated using age-specific mortality rates and standard demographic methods. Uncertainty intervals (UIs) were calculated for every metric using the 25th and 975th ordered values from a 1000-draw posterior distribution. Findings: Global all-cause mortality followed two distinct patterns over the study period: age-standardised mortality rates declined between 1950 and 2019 (a 62·8% [95% UI 60·5–65·1] decline), and increased during the COVID-19 pandemic period (2020–21; 5·1% [0·9–9·6] increase). In contrast with the overall reverse in mortality trends during the pandemic period, child mortality continued to decline, with 4·66 million (3·98–5·50) global deaths in children younger than 5 years in 2021 compared with 5·21 million (4·50–6·01) in 2019. An estimated 131 million (126–137) people died globally from all causes in 2020 and 2021 combined, of which 15·9 million (14·7–17·2) were due to the COVID-19 pandemic (measured by excess mortality, which includes deaths directly due to SARS-CoV-2 infection and those indirectly due to other social, economic, or behavioural changes associated with the pandemic). Excess mortality rates exceeded 150 deaths per 100 000 population during at least one year of the pandemic in 80 countries and territories, whereas 20 nations had a negative excess mortality rate in 2020 or 2021, indicating that all-cause mortality in these countries was lower during the pandemic than expected based on historical trends. Between 1950 and 2021, global life expectancy at birth increased by 22·7 years (20·8–24·8), from 49·0 years (46·7–51·3) to 71·7 years (70·9–72·5). Global life expectancy at birth declined by 1·6 years (1·0–2·2) between 2019 and 2021, reversing historical trends. An increase in life expectancy was only observed in 32 (15·7%) of 204 countries and territories between 2019 and 2021. The global population reached 7·89 billion (7·67–8·13) people in 2021, by which time 56 of 204 countries and territories had peaked and subsequently populations have declined. The largest proportion of population growth between 2020 and 2021 was in sub-Saharan Africa (39·5% [28·4–52·7]) and south Asia (26·3% [9·0–44·7]). From 2000 to 2021, the ratio of the population aged 65 years and older to the population aged younger than 15 years increased in 188 (92·2%) of 204 nations. Interpretation: Global adult mortality rates markedly increased during the COVID-19 pandemic in 2020 and 2021, reversing past decreasing trends, while child mortality rates continued to decline, albeit more slowly than in earlier years. Although COVID-19 had a substantial impact on many demographic indicators during the first 2 years of the pandemic, overall global health progress over the 72 years evaluated has been profound, with considerable improvements in mortality and life expectancy. Additionally, we observed a deceleration of global population growth since 2017, despite steady or increasing growth in lower-income countries, combined with a continued global shift of population age structures towards older ages. These demographic changes will likely present future challenges to health systems, economies, and societies. The comprehensive demographic estimates reported here will enable researchers, policy makers, health practitioners, and other key stakeholders to better understand and address the profound changes that have occurred in the global health landscape following the first 2 years of the COVID-19 pandemic, and longer-term trends beyond the pandemic

Global age-sex-specific mortality, life expectancy, and population estimates in 204 countries and territories and 811 subnational locations, 1950–2021, and the impact of the COVID-19 pandemic: a comprehensive demographic analysis for the Global Burden of Disease Study 2021

BACKGROUND: Estimates of demographic metrics are crucial to assess levels and trends of population health outcomes. The profound impact of the COVID-19 pandemic on populations worldwide has underscored the need for timely estimates to understand this unprecedented event within the context of long-term population health trends. The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 provides new demographic estimates for 204 countries and territories and 811 additional subnational locations from 1950 to 2021, with a particular emphasis on changes in mortality and life expectancy that occurred during the 2020–21 COVID-19 pandemic period. METHODS: 22 223 data sources from vital registration, sample registration, surveys, censuses, and other sources were used to estimate mortality, with a subset of these sources used exclusively to estimate excess mortality due to the COVID-19 pandemic. 2026 data sources were used for population estimation. Additional sources were used to estimate migration; the effects of the HIV epidemic; and demographic discontinuities due to conflicts, famines, natural disasters, and pandemics, which are used as inputs for estimating mortality and population. Spatiotemporal Gaussian process regression (ST-GPR) was used to generate under-5 mortality rates, which synthesised 30 763 location-years of vital registration and sample registration data, 1365 surveys and censuses, and 80 other sources. ST-GPR was also used to estimate adult mortality (between ages 15 and 59 years) based on information from 31 642 location-years of vital registration and sample registration data, 355 surveys and censuses, and 24 other sources. Estimates of child and adult mortality rates were then used to generate life tables with a relational model life table system. For countries with large HIV epidemics, life tables were adjusted using independent estimates of HIV-specific mortality generated via an epidemiological analysis of HIV prevalence surveys, antenatal clinic serosurveillance, and other data sources. Excess mortality due to the COVID-19 pandemic in 2020 and 2021 was determined by subtracting observed all-cause mortality (adjusted for late registration and mortality anomalies) from the mortality expected in the absence of the pandemic. Expected mortality was calculated based on historical trends using an ensemble of models. In location-years where all-cause mortality data were unavailable, we estimated excess mortality rates using a regression model with covariates pertaining to the pandemic. Population size was computed using a Bayesian hierarchical cohort component model. Life expectancy was calculated using age-specific mortality rates and standard demographic methods. Uncertainty intervals (UIs) were calculated for every metric using the 25th and 975th ordered values from a 1000-draw posterior distribution. FINDINGS: Global all-cause mortality followed two distinct patterns over the study period: age-standardised mortality rates declined between 1950 and 2019 (a 62·8% [95% UI 60·5–65·1] decline), and increased during the COVID-19 pandemic period (2020–21; 5·1% [0·9–9·6] increase). In contrast with the overall reverse in mortality trends during the pandemic period, child mortality continued to decline, with 4·66 million (3·98–5·50) global deaths in children younger than 5 years in 2021 compared with 5·21 million (4·50–6·01) in 2019. An estimated 131 million (126–137) people died globally from all causes in 2020 and 2021 combined, of which 15·9 million (14·7–17·2) were due to the COVID-19 pandemic (measured by excess mortality, which includes deaths directly due to SARS-CoV-2 infection and those indirectly due to other social, economic, or behavioural changes associated with the pandemic). Excess mortality rates exceeded 150 deaths per 100 000 population during at least one year of the pandemic in 80 countries and territories, whereas 20 nations had a negative excess mortality rate in 2020 or 2021, indicating that all-cause mortality in these countries was lower during the pandemic than expected based on historical trends. Between 1950 and 2021, global life expectancy at birth increased by 22·7 years (20·8–24·8), from 49·0 years (46·7–51·3) to 71·7 years (70·9–72·5). Global life expectancy at birth declined by 1·6 years (1·0–2·2) between 2019 and 2021, reversing historical trends. An increase in life expectancy was only observed in 32 (15·7%) of 204 countries and territories between 2019 and 2021. The global population reached 7·89 billion (7·67–8·13) people in 2021, by which time 56 of 204 countries and territories had peaked and subsequently populations have declined. The largest proportion of population growth between 2020 and 2021 was in sub-Saharan Africa (39·5% [28·4–52·7]) and south Asia (26·3% [9·0–44·7]). From 2000 to 2021, the ratio of the population aged 65 years and older to the population aged younger than 15 years increased in 188 (92·2%) of 204 nations. INTERPRETATION: Global adult mortality rates markedly increased during the COVID-19 pandemic in 2020 and 2021, reversing past decreasing trends, while child mortality rates continued to decline, albeit more slowly than in earlier years. Although COVID-19 had a substantial impact on many demographic indicators during the first 2 years of the pandemic, overall global health progress over the 72 years evaluated has been profound, with considerable improvements in mortality and life expectancy. Additionally, we observed a deceleration of global population growth since 2017, despite steady or increasing growth in lower-income countries, combined with a continued global shift of population age structures towards older ages. These demographic changes will likely present future challenges to health systems, economies, and societies. The comprehensive demographic estimates reported here will enable researchers, policy makers, health practitioners, and other key stakeholders to better understand and address the profound changes that have occurred in the global health landscape following the first 2 years of the COVID-19 pandemic, and longer-term trends beyond the pandemic. FUNDING: Bill & Melinda Gates Foundation

Investigation of hospital discharge cases and SARS-CoV-2 introduction into Lothian care homes

Background The first epidemic wave of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Scotland resulted in high case numbers and mortality in care homes. In Lothian, over one-third of care homes reported an outbreak, while there was limited testing of hospital patients discharged to care homes. Aim To investigate patients discharged from hospitals as a source of SARS-CoV-2 introduction into care homes during the first epidemic wave. Methods A clinical review was performed for all patients discharges from hospitals to care homes from 1st March 2020 to 31st May 2020. Episodes were ruled out based on coronavirus disease 2019 (COVID-19) test history, clinical assessment at discharge, whole-genome sequencing (WGS) data and an infectious period of 14 days. Clinical samples were processed for WGS, and consensus genomes generated were used for analysis using Cluster Investigation and Virus Epidemiological Tool software. Patient timelines were obtained using electronic hospital records. Findings In total, 787 patients discharged from hospitals to care homes were identified. Of these, 776 (99%) were ruled out for subsequent introduction of SARS-CoV-2 into care homes. However, for 10 episodes, the results were inconclusive as there was low genomic diversity in consensus genomes or no sequencing data were available. Only one discharge episode had a genomic, time and location link to positive cases during hospital admission, leading to 10 positive cases in their care home. Conclusion The majority of patients discharged from hospitals were ruled out for introduction of SARS-CoV-2 into care homes, highlighting the importance of screening all new admissions when faced with a novel emerging virus and no available vaccine

SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway

Vaccines based on the spike protein of SARS-CoV-2 are a cornerstone of the public health response to COVID-19. The emergence of hypermutated, increasingly transmissible variants of concern (VOCs) threaten this strategy. Omicron (B.1.1.529), the fifth VOC to be described, harbours multiple amino acid mutations in spike, half of which lie within the receptor-binding domain. Here we demonstrate substantial evasion of neutralization by Omicron BA.1 and BA.2 variants in vitro using sera from individuals vaccinated with ChAdOx1, BNT162b2 and mRNA-1273. These data were mirrored by a substantial reduction in real-world vaccine effectiveness that was partially restored by booster vaccination. The Omicron variants BA.1 and BA.2 did not induce cell syncytia in vitro and favoured a TMPRSS2-independent endosomal entry pathway, these phenotypes mapping to distinct regions of the spike protein. Impaired cell fusion was determined by the receptor-binding domain, while endosomal entry mapped to the S2 domain. Such marked changes in antigenicity and replicative biology may underlie the rapid global spread and altered pathogenicity of the Omicron variant

Global burden of 288 causes of death and life expectancy decomposition in 204 countries and territories and 811 subnational locations, 1990–2021: a systematic analysis for the Global Burden of Disease Study 2021

BACKGROUND Regular, detailed reporting on population health by underlying cause of death is fundamental for public health decision making. Cause-specific estimates of mortality and the subsequent effects on life expectancy worldwide are valuable metrics to gauge progress in reducing mortality rates. These estimates are particularly important following large-scale mortality spikes, such as the COVID-19 pandemic. When systematically analysed, mortality rates and life expectancy allow comparisons of the consequences of causes of death globally and over time, providing a nuanced understanding of the effect of these causes on global populations. METHODS The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 cause-of-death analysis estimated mortality and years of life lost (YLLs) from 288 causes of death by age-sex-location-year in 204 countries and territories and 811 subnational locations for each year from 1990 until 2021. The analysis used 56 604 data sources, including data from vital registration and verbal autopsy as well as surveys, censuses, surveillance systems, and cancer registries, among others. As with previous GBD rounds, cause-specific death rates for most causes were estimated using the Cause of Death Ensemble model-a modelling tool developed for GBD to assess the out-of-sample predictive validity of different statistical models and covariate permutations and combine those results to produce cause-specific mortality estimates-with alternative strategies adapted to model causes with insufficient data, substantial changes in reporting over the study period, or unusual epidemiology. YLLs were computed as the product of the number of deaths for each cause-age-sex-location-year and the standard life expectancy at each age. As part of the modelling process, uncertainty intervals (UIs) were generated using the 2·5th and 97·5th percentiles from a 1000-draw distribution for each metric. We decomposed life expectancy by cause of death, location, and year to show cause-specific effects on life expectancy from 1990 to 2021. We also used the coefficient of variation and the fraction of population affected by 90% of deaths to highlight concentrations of mortality. Findings are reported in counts and age-standardised rates. Methodological improvements for cause-of-death estimates in GBD 2021 include the expansion of under-5-years age group to include four new age groups, enhanced methods to account for stochastic variation of sparse data, and the inclusion of COVID-19 and other pandemic-related mortality-which includes excess mortality associated with the pandemic, excluding COVID-19, lower respiratory infections, measles, malaria, and pertussis. For this analysis, 199 new country-years of vital registration cause-of-death data, 5 country-years of surveillance data, 21 country-years of verbal autopsy data, and 94 country-years of other data types were added to those used in previous GBD rounds. FINDINGS The leading causes of age-standardised deaths globally were the same in 2019 as they were in 1990; in descending order, these were, ischaemic heart disease, stroke, chronic obstructive pulmonary disease, and lower respiratory infections. In 2021, however, COVID-19 replaced stroke as the second-leading age-standardised cause of death, with 94·0 deaths (95% UI 89·2-100·0) per 100 000 population. The COVID-19 pandemic shifted the rankings of the leading five causes, lowering stroke to the third-leading and chronic obstructive pulmonary disease to the fourth-leading position. In 2021, the highest age-standardised death rates from COVID-19 occurred in sub-Saharan Africa (271·0 deaths [250·1-290·7] per 100 000 population) and Latin America and the Caribbean (195·4 deaths [182·1-211·4] per 100 000 population). The lowest age-standardised death rates from COVID-19 were in the high-income super-region (48·1 deaths [47·4-48·8] per 100 000 population) and southeast Asia, east Asia, and Oceania (23·2 deaths [16·3-37·2] per 100 000 population). Globally, life expectancy steadily improved between 1990 and 2019 for 18 of the 22 investigated causes. Decomposition of global and regional life expectancy showed the positive effect that reductions in deaths from enteric infections, lower respiratory infections, stroke, and neonatal deaths, among others have contributed to improved survival over the study period. However, a net reduction of 1·6 years occurred in global life expectancy between 2019 and 2021, primarily due to increased death rates from COVID-19 and other pandemic-related mortality. Life expectancy was highly variable between super-regions over the study period, with southeast Asia, east Asia, and Oceania gaining 8·3 years (6·7-9·9) overall, while having the smallest reduction in life expectancy due to COVID-19 (0·4 years). The largest reduction in life expectancy due to COVID-19 occurred in Latin America and the Caribbean (3·6 years). Additionally, 53 of the 288 causes of death were highly concentrated in locations with less than 50% of the global population as of 2021, and these causes of death became progressively more concentrated since 1990, when only 44 causes showed this pattern. The concentration phenomenon is discussed heuristically with respect to enteric and lower respiratory infections, malaria, HIV/AIDS, neonatal disorders, tuberculosis, and measles. INTERPRETATION Long-standing gains in life expectancy and reductions in many of the leading causes of death have been disrupted by the COVID-19 pandemic, the adverse effects of which were spread unevenly among populations. Despite the pandemic, there has been continued progress in combatting several notable causes of death, leading to improved global life expectancy over the study period. Each of the seven GBD super-regions showed an overall improvement from 1990 and 2021, obscuring the negative effect in the years of the pandemic. Additionally, our findings regarding regional variation in causes of death driving increases in life expectancy hold clear policy utility. Analyses of shifting mortality trends reveal that several causes, once widespread globally, are now increasingly concentrated geographically. These changes in mortality concentration, alongside further investigation of changing risks, interventions, and relevant policy, present an important opportunity to deepen our understanding of mortality-reduction strategies. Examining patterns in mortality concentration might reveal areas where successful public health interventions have been implemented. Translating these successes to locations where certain causes of death remain entrenched can inform policies that work to improve life expectancy for people everywhere. FUNDING Bill & Melinda Gates Foundation

Μικροβιακές κοινότητες της Ανατολικής Μεσογείου και η δυνατότητά τους στην αποδόμηση υδρογονανθράκων

The Eastern Mediterranean Sea (EMS) is a semi-enclosed basin, usually referred to as a “miniature ocean” because of the complex oceanic activities that take place there. Increased bottom sea temperatures, salinity levels and ultraoligotrophic conditions are some of its unique characteristics that shape microbial communities. This sub-basin of the Mediterranean Sea has become a hotspot for oil and gas activities in the recent decades. Large reservoirs have already been discovered and exploited in depths reaching ~1500m while several others, including ultra-deep marine areas, have been committed and are currently under exploration (South-Southwest Crete). In the aftermath of Deepwater Horizon (DWH) blowout in 2010, deep-sea oil biodegradation studies flourished providing the scientific community with valuable information about the fate of hydrocarbons (HC) in the deep sea, microbial succession patterns and key oil-degrading taxa. One of the major lessons learned from the DWH accident was the importance of conducting site-specific research under in situ conditions to provide policy makers with realistic data for the construction of efficient bioremediation protocols. The ongoing activities, which are progressing in deeper and more challenging waters, increase the risk for a potential oil spill accident in the deep EMS with many environmental and financial consequences for the surrounding countries. This PhD thesis attempts to evaluate the microbial response and self-healing capability (natural attenuation) in the event of an accidental hydrocarbon release scenario in the deep EMS. For this purpose, seawater was retrieved from the EMS water column, down to 1000 m below sea level, in stations south of Crete (Cretan Passage) using Niskin bottles (decompressed) and a high-pressure sampling apparatus (in situ pressure). Hydrocarbon-biodegradation experiments were conducted in Erlenmeyer flasks and in high-pressure bottles or high-pressure bioreactor for incubations at 0.1MPa and 10MPa respectively, using Iranian light crude oil as carbon source. Data on microbial community analysis and hydrocarbon-degradation rates were produced via high throughput sequencing and GC-MS analysis. Prior to any hydrocarbon exposure experiments, the synthesis of the pristine microbial community along with interspecies associations were analysed across the water column in this understudied marine region and background levels of known hydrocarbon degraders were recorded. Interestingly, even though natural seepages were not present near the sampling stations, notable abundances of taxa involved in oil bioremediation were found, especially in the deep-water layers. The known hydrocarbonoclastic genus, Alcanivorax, was included in the significant nodes of the deep network. Comparison of a timeseries hydrocarbon-degradation experiment at in situ temperature conditions between surface and deep EMS-collected microbial communities indicated that the latter (deep community) responded faster to oil contamination than the surface one despite the incubation at lower in situ temperature of 14 ºC. Furthermore, incubation of the deep consortium at a higher temperature of 25 ºC (for direct comparison with the surface community) did not affect oil biodegradation levels suggesting a microbial community that is acclimatized for HC biodegradation at the lower in situ temperature. Monitoring of microbial succession patterns resulted in different key hydrocarbon-degrading taxa in each treatment. The deep consortium at 14 ºC, was dominated primarily by the generalist Vibrio and substituted later on by the slow-growing specialist Alcanivorax whereas incubation at 25 ºC led to the dominance of Pseudomonas and Pseudoalteromonas in the deep community. On the other hand, the surface consortium was enriched in Thalassospira, Halomonas, Alteromonas and Idiomarina genera. The efficacy of the deep EMS consortia in hydrocarbon bioremediation, was further tested under in situ high-pressure conditions. In particular, the impact of decompression was evaluated during deep-sea sampling and in enrichment incubations for the isolation of oil degraders. Decompression upon sampling resulted in a drastic decrease in deep microbial diversity. In addition, subjection of HC-degrading consortia for enrichment in ONR7 medium caused further decrease in biodiversity and taxa correlated with HC removal were outcompeted by Pseudoalteromonas, Halomonas, Thalassomonas and Alcanivorax which were favored under all treatments tested. Dispersant application had no significant effect in microbial community composition at any stage of the experimental process. Biodiversity loss impacted the community functionality in terms of biodegradation of the more recalcitrant oil compounds (PAHs, Heavy Alkanes). A strains of Alcanivorax venustensis (recently emended as Alloalcanivorax venustensis) that dominated the communities was isolated. Furthermore, a deep EMS hydrocarbon plume emulation experiment was conducted under in situ EMS conditions to address primary microbial responders and succession patterns in the absence and presence of dispersant. Genera belonging to Gammaproteobacteria (Oleispira, Thalassomonas, Thalassotalea, Ralstonia) were among the first responders to oil-only contamination similarly to studies in the aftermath of the DWH accident. Those were then succeeded by Alcanivorax and Methylophaga, followed by Marinobacter and Thalassospira in the late phases of exposure to crude oil. The presence of dispersant favored members of Bacteroidota along with Hyphomonas and Alcanivorax, with the latter dominating the community. In conclusion, marine areas of interest for oil and gas exploration and exploitation (off-Crete) present a microbial “seed bank” of species related to hydrocarbon removal especially in deep-water layers. This could explain why the deep EMS community responds faster to oil contamination even though at lower temperature conditions than the surface consortium. Moreover, the results of this dissertation suggest an important role for Alcanivorax in hydrocarbon bioremediation in the deep EMS along with other known obligate oil degraders such as Oleispira and Marinobacter. Overall, this PhD thesis underlines the importance of maintaining in situ pressure and temperature conditions during sampling and experimentation when conducting experiments with deep-seawater microbial communities.Η Ανατολική Μεσόγειος (ΑΜ) είναι μια ημίκλειστη θαλάσσια λεκάνη με πολύπλοκες ωκεάνιες διεργασίες. Η αυξημένη θερμοκρασία στα βαθιά της νερά, τα υψηλά επίπεδα αλατότητας και οι υπερολιγοτροφικές συνθήκες είναι μερικά από τα μοναδικά χαρακτηριστικά της ΑΜ τα οποία επηρεάζουν την σύνθεση και τις μεταβολικές διεργασίες των μικροβιακων κοινοτήτων. Τις τελευταίες δεκαετίες, αυτή η περιοχή έχει αποκτήσει ιδιαίτερο ενδιαφέρον λόγω των ερευνών-εξορύξεων πετρελαίου και φυσικού αερίου. Μεγάλα υποθαλάσσια κοιτάσματα βρίσκονται ήδη υπό εκμετάλλευση σε βάθη ~1500 μέτρων από την επιφάνεια της θάλασσας, ενώ άλλες περιοχές, σε ακόμα μεγαλύτερα βάθη, έχουν δεσμευτεί και βρίσκονται ήδη υπό εξερεύνηση (Νότια-Νοτιοδυτικά της Κρήτης). Στον απόηχο του ατυχήματος του Deepwater Horizon (DWH) το 2010, πραγματοποιήθηκαν πολλές μελέτες βιοαποδόμησης πετρελαίου στα βαθιά νερά παρέχοντας πολύτιμες πληροφορίες σχετικά με την τύχη των υδρογονανθράκων (Υ/Α), τα μοτίβα μικροβιακής διαδοχής και ανέδειξαν σημαντικά είδη ως προς την αποδόμηση πετρελαίου. Ένα από τα σημαντικότερα διδάγματα του ατυχήματος DWH είναι η ανάγκη διεξαγωγής πειραμάτων αποδόμησης Υ/Α στις περιβαλλοντικές συνθήκες της περιοχής ενδιαφέροντος προκειμένου να καταγράφονται ρεαλιστικά δεδομένα που να μπορούν να χρησιμοποιηθούν για την δημιουργία αποτελεσματικών πρωτοκόλλων βιοαποκατάστασης. Οι συνεχιζόμενες δραστηριότητες στην ΑΜ, οι οποίες προχωρούν σε ολοένα και βαθύτερα ύδατα με αυξανόμενες τεχνικές δυσκολίες, ενισχύουν τον κίνδυνο για ένα ατύχημα έκλυσης υδρογονανθράκων με πολλές περιβαλλοντικές και οικονομικές συνέπειες για τις γύρω χώρες. Αυτή η διδακτορική διατριβή επιχειρεί να αξιολογήσει την μικροβιακή απόκριση και την ικανότητα αυτοεξυγίανσης μέσω φυσικής εξασθένησης σε ένα ενδεχόμενο ατύχημα απελευθέρωσης υδρογονανθράκων στα βαθιά νερά της ΑΜ. Για το σκοπό αυτό, ανακτήθηκε θαλασσινό νερό από τη στήλη νερού της ΑΜ, μέχρι τα 1000 m, σε σταθμούς νότια της Κρήτης (Κρητικό Πέρασμα) με τη χρήση φιαλών Niskin (αποσυμπιεσμένο) και με συσκευή δειγματοληψίας υψηλής πίεσης (διατήρηση in situ πίεσης). Πειράματα βιοαποδόμησης υδρογονανθράκων διεξήχθησαν σε φλάσκες Erlenmeyer και σε δοχεία ή σε βιοαντιδραστήρα υψηλής πίεσης για επωάσεις στα 0,1 MPa και 10 MPa αντίστοιχα, χρησιμοποιώντας ελαφρύ Ιρανικό αργό πετρέλαιο ως πηγή άνθρακα. Δεδομένα σχετικά με την ανάλυση μικροβιακής κοινότητας και τους ρυθμούς αποδόμησης υδρογονανθράκων παρήχθησαν μέσω αλληλούχισης υψηλής απόδοσης και ανάλυσης αέριας χρωματογραφίας (GC-MS).Πριν από οποιαδήποτε πειράματα έκθεσης σε υδρογονάνθρακες, η σύνθεση της φυσικής μικροβιακής κοινότητας και οι συσχετίσεις μεταξύ των ειδών αναλύθηκαν σε όλη τη στήλη του νερού σε αυτήν την θαλάσσια περιοχή που δεν έχει μελετηθεί σχεδόν καθόλου ενώ καταγράφηκαν και οι συγκεντρώσεις υπόβαθρου γνωστών αποδομητών Υ/Α. Ενδιαφέρον αποτελεί το γεγονός πως, παρόλο που δεν υπάρχουν φυσικές πηγές διαρροής Υ/Α κοντά στους σταθμούς δειγματοληψίας, εντοπίστηκαν αξιοσημείωτες αφθονίες μικροοργανισμών που εμπλέκονται στη βιοαποδόμηση του πετρελαίου ειδικά στα βαθύτερα ύδατα. Το είδος Alcanivorax εντοπίστηκε ανάμεσα στους σημαντικούς κόμβους του δικτύου συσχετίσεων στα βαθιά νερά της ΑΜ.Η σύγκριση ενός πειράματος αποδόμησης Υ/Α σε συνθήκες in situ θερμοκρασίας μεταξύ επιφανειακών και βαθιών μικροβιακών κοινοτήτων της ΑΜ έδειξε ότι η βαθιά κοινότητα αποκρίθηκε ταχύτερα από την επιφανειακή στη μόλυνση του πετρελαίου παρά την επώαση της σε χαμηλότερη θερμοκρασία (14 ºC). Επιπλέον, η επώαση της βαθιάς μικροβιακής κοινότητας στους 25 ºC (για άμεση σύγκριση με την επιφανειακή) δεν επηρέασε τα επίπεδα βιοαποδόμησης πετρελαίου υποδηλώνοντας μια εγκλιματισμένη κοινότητα με ικανότητα βιοαποδόμησης Υ/Α στη χαμηλότερη in situ θερμοκρασία. Η ανάλυση των μοτίβων μικροβιακής διαδοχής ανέδειξε διαφορετικά είδη σε κάθε πειραματική επεξεργασία. Στη βαθιά κοινότητα που επωάστηκε στους 14 ºC, κυριάρχησε αρχικά το γένος Vibrio που αντικαταστάθηκε αργότερα από το Alcanivorax, ενώ η επώαση στους 25 ºC οδήγησε στην κυριαρχία των Pseudomonas και Pseudoalteromonas στη βαθιά κοινότητα. Από την άλλη, η επιφανειακή κοινότητα εμπλουτίστηκε στα γένη Thalassospira, Halomonas, Alteromonas και Idiomarina.Η αποτελεσματικότητα της μικροβιακής κοινότητας από τα βαθιά νερά της ΑΜ στη βιοεξυγίανση υδρογονανθράκων, ελέγχθηκε περαιτέρω σε in situ συνθήκες υψηλής πίεσης. Ειδικότερα, αξιολογήθηκε ο αντίκτυπος της αποσυμπίεσης κατά τη δειγματοληψία από τα βαθιά ύδατα όπως και σε πειράματα εμπλουτισμού για την απομόνωση αποδομητών πετρελαίου. Η αποσυμπίεση κατά τη δειγματοληψία οδήγησε σε δραστική μείωση της μικροβιακής ποικιλότητας από τα βαθιά νερά. Επιπλέον, η υποβολή των εγκλιματισμένων κοινοτήτων στο μέσο ONR7 για τον εμπλουτισμό και την απομόνωση αποδομητών Υ/Α προκάλεσε περαιτέρω μείωση της ποικιλομορφίας της κοινότητας και την κυριαρχία των Pseudoalteromonas, Halomonas, Thalassomonas και Alcanivorax σε όλες τις πειραματικές επεξεργασίες. Η εφαρμογή του διασκορπιστικού μέσου δεν είχε σημαντική επίδραση στη μικροβιακή σύνθεση της κοινότητας σε οποιοδήποτε στάδιο της πειραματικής διαδικασίας. Η απώλεια βιοποικιλότητας επηρέασε την ικανότητα της κοινότητας ως προς την αποδόμηση των πιο ανθεκτικών ενώσεων του πετρελαίου. Στέλεχος του είδους Alcanivorax venustensis (πρόσφατα τροποποιήθηκε σε Alloalcanivorax venustensis) που κυριαρχούσε στις κοινότητες απομονώθηκε στο τέλος του πειράματος.Επιπλέον, διεξήχθη ένα πείραμα εξομοίωσης πλουμίου υδρογονανθράκων στη βαθιά θάλασσα σε in situ συνθήκες της ΑΜ για την διερεύνηση των μικροβιακων ειδών που αποκρίνονται πρώτα στην παρουσία πετρελαίου στη βαθιά θάλασσα καθώς και των μοτίβων διαδοχής απουσία και παρουσία διασκορπιστικού μέσου. Τα γένη που ανήκουν στα γ-Proteobacteria (Oleispira, Thalassomonas, Thalassotalea, Ralstonia) ήταν μεταξύ των πρώτων που αποκρίθηκαν στη μόλυνση, αντίστοιχα με τις μελέτες που πραγματοποιήθηκαν στον απόηχο του DWH. Στη συνέχεια, η κοινότητα εμπλουτίστηκε στα είδη Alcanivorax και Methylophaga, ενώ έπειτα ακολούθησε η ενίσχυση στα γένη Marinobacter και Thalassospira στις όψιμες φάσεις της έκθεσης στο αργό πετρέλαιο. Η παρουσία του διασκορπιστικού μέσου ευνόησε μικροοργανισμούς που ανήκουν στο φύλο Bacteroidota μαζί με τα γένη Hyphomonas και Alcanivorax, με το τελευταίο να κυριαρχεί στην κοινότητα.Συμπερασματικά, οι θαλάσσιες περιοχές ενδιαφέροντος για εξερεύνηση και εκμετάλλευση πετρελαίου και φυσικού αερίου (νότια της Κρήτης) παρουσιάζουν φυσική παρουσία μικροβιακών ειδών που σχετίζονται με την απομάκρυνση υδρογονανθράκων ειδικά στα βαθιά νερά. Αυτό θα μπορούσε να ερμηνεύσει και το γεγονός πως η βαθιά μικροβιακή κοινότητα αποκρίνεται ταχύτερα στη ρύπανση πετρελαίου ακόμα και σε συνθήκες χαμηλότερης θερμοκρασίας από την αντίστοιχη επιφανειακή κοινότητα. Επιπλέον, τα αποτελέσματα αυτής της μελέτης υποδηλώνουν τον σημαντικό ρόλο του γένους Alcanivorax στη βιοξυγίανση υδρογονανθράκων στη βαθιά θάλασσα της Ανατολικής Μεσογείου μαζί με τα Oleispira και Marinobacter. Συνολικά, αυτή η διατριβή υπογραμμίζει τη σημασία της διατήρησης των περιβαλλοντικών συνθηκών πίεσης και θερμοκρασίας τόσο κατά τη δειγματοληψία όσο και κατά την διεξαγωγή πειραμάτων με μικροβιακές κοινότητες από τα βαθιά νερά

Emulating Deep-Sea Bioremediation: Oil Plume Degradation by Undisturbed Deep-Sea Microbial Communities Using a High-Pressure Sampling and Experimentation System

Hydrocarbon biodegradation rates in the deep-sea have been largely determined under atmospheric pressure, which may lead to non-representative results. In this work, we aim to study the response of deep-sea microbial communities of the Eastern Mediterranean Sea (EMS) to oil contamination at in situ environmental conditions and provide representative biodegradation rates. Seawater from a 600 to 1000 m depth was collected using a high-pressure (HP) sampling device equipped with a unidirectional check-valve, without depressurization upon retrieval. The sample was then passed into a HP-reactor via a piston pump without pressure disruption and used for a time-series oil biodegradation experiment at plume concentrations, with and without dispersant application, at 10 MPa and 14 °C. The experimental results demonstrated a high capacity of indigenous microbial communities in the deep EMS for alkane degradation regardless of dispersant application (>70%), while PAHs were highly degraded when oil was dispersed (>90%) and presented very low half-lives (19.4 to 2.2 days), compared to published data. To our knowledge, this is the first emulation study of deep-sea bioremediation using undisturbed deep-sea microbial communities

Emulating Deep-Sea Bioremediation: Oil Plume Degradation by Undisturbed Deep-Sea Microbial Communities Using a High-Pressure Sampling and Experimentation System

Hydrocarbon biodegradation rates in the deep-sea have been largely determined under atmospheric pressure, which may lead to non-representative results. In this work, we aim to study the response of deep-sea microbial communities of the Eastern Mediterranean Sea (EMS) to oil contamination at in situ environmental conditions and provide representative biodegradation rates. Seawater from a 600 to 1000 m depth was collected using a high-pressure (HP) sampling device equipped with a unidirectional check-valve, without depressurization upon retrieval. The sample was then passed into a HP-reactor via a piston pump without pressure disruption and used for a time-series oil biodegradation experiment at plume concentrations, with and without dispersant application, at 10 MPa and 14 °C. The experimental results demonstrated a high capacity of indigenous microbial communities in the deep EMS for alkane degradation regardless of dispersant application (>70%), while PAHs were highly degraded when oil was dispersed (>90%) and presented very low half-lives (19.4 to 2.2 days), compared to published data. To our knowledge, this is the first emulation study of deep-sea bioremediation using undisturbed deep-sea microbial communities

Unconventional targeting of a thiol peroxidase to the mitochondrial intermembrane space facilitates oxidative protein folding

Thiol peroxidases are conserved hydrogen peroxide scavenging and signaling molecules that contain redox-active cysteine residues. We show here that Gpx3, the major H2O2 sensor in yeast, is present in the mitochondrial intermembrane space (IMS), where it serves a compartment-specific role in oxidative metabolism. The IMS-localized Gpx3 contains an 18-amino acid N-terminally extended form encoded from a non-AUG codon. This acts as a mitochondrial targeting signal in a pathway independent of the hitherto known IMS-import pathways. Mitochondrial Gpx3 interacts with the Mia40 oxidoreductase in a redox-dependent manner and promotes efficient Mia40-dependent oxidative protein folding. We show that cells lacking Gpx3 have aberrant mitochondrial morphology, defective protein import capacity, and lower inner membrane potential, all of which can be rescued by expression of a mitochondrial-only form of Gpx3. Together, our data reveal a novel role for Gpx3 in mitochondrial redox regulation and protein homeostasis