Clinical Impact of Serious Respiratory Disease in Children Under the Age of 2 Years During the 2021-2022 Bronchiolitis Season in England, Scotland, and Ireland.

BACKGROUND: Interventions introduced to reduce the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to a widespread reduction in childhood infections. However, from spring 2021 onwards the United Kingdom and Ireland experienced an unusual out-of-season epidemic of respiratory disease. METHODS: We conducted a prospective observational study (BronchStart), enrolling children 0-23 months of age presenting with bronchiolitis, lower respiratory tract infection, or first episode of wheeze to 59 emergency departments across England, Scotland, and Ireland from May 2021 to April 2022. We combined testing data with national admissions datasets to infer the impact of respiratory syncytial virus (RSV) disease. RESULTS: The BronchStart study collected data on 17 899 presentations for 17 164 children. Risk factors for admission and escalation of care included prematurity and congenital heart disease, but most admissions were for previously healthy term-born children. Of those aged 0-11 months who were admitted and tested for RSV, 1907 of 3912 (48.7%) tested positive. We estimate that every year in England and Scotland 28 561 (95% confidence interval, 27 637-29 486) infants are admitted with RSV infection. CONCLUSIONS: RSV infection was the main cause of hospitalizations in this cohort, but 51.3% of admissions in infants were not associated with the virus. The majority of admissions were in previously healthy term-born infants.

Biovalorisation of agro-industrial wastes into astaxanthin by Xanthophyllomyces dendrorhous.

Astaxanthin is a red xanthophyll with high economic and industrial value in the pharmaceutical, nutraceutical, cosmetic and food industries. In recent years, the biotechnological production of astaxanthin has attracted much attention as a sustainable alternative to the predominating petrochemical-dependent chemical synthesis. In this regard, Xanthophyllomyces dendrorhous is regarded as a promising microorganism for industrial production of astaxanthin. Unfortunately, biotechnological production of the carotenoid is currently expensive. The present study investigated soy molasses (SM) and residual brewers' yeast as cheap fermentation feedstocks for the cultivation of X. dendrorhous and astaxanthin production. Yeast extract was obtained from residual brewers' yeast using various techniques and then combined with SM to formulate a two-component growth medium which was subsequently used to cultivate X. dendrorhous. Generally, the yeast extract produced from residual brewers' yeast supported X. dendrorhous growth and astaxanthin production at levels comparable to those seen with commercial yeast extract. Overall, cultivating X. dendrorhous in an SM-based medium containing 5% SM and 0.2% yeast extract obtained from residual brewers' yeast resulted in significantly higher (> 20% more) biomass accumulation compared to the control media (YPD). A similar slightly higher astaxanthin output (up to 14% more) was recorded in the SM-based medium compared to YPD. The formulated cultivation medium in this study provides an opportunity to reduce the production cost of astaxanthin from X. dendrorhous while simultaneously reducing the environmental impact related to the disposal of the industrial waste used as feedstock. KEY POINTS: • Cheap culture media were formulated from soy molasses and brewers' spent yeast • The formulated medium resulted in at least 20% more biomass than the control • Up to 14% more astaxanthin was produced in molasses-based medium.

Respiratory Syncytial Virus in Adult Patients at a Tertiary Care Hospital in Germany: Clinical Features and Molecular Epidemiology of the Fusion Protein in the Severe Respiratory Season of 2022/2023.

Following an interseasonal rise in mainly pediatric respiratory syncytial virus (RSV) cases in Germany in 2021, an exceptionally high number of adult cases was observed in the subsequent respiratory season of 2022/2023. The aim of this study was to compare the clinical presentation of RSV infections in the pre- and post-SARS-CoV-2 pandemic periods. Additionally, the local epidemiology of the RSV fusion protein was analyzed at a molecular genetic and amino acid level. RSV detections in adults peaked in calendar week 1 of 2023, 8 weeks earlier than the earliest peak observed in the three pre-pandemic seasons. Although the median age of the adult patients was not different (66.5 vs. 65 years), subtle differences between both periods regarding comorbidities and the clinical presentation of RSV cases were noted. High rates of comorbidities prevailed; however, significantly lower numbers of patients with a history of lung transplantation (p = 0.009), chronic kidney disease (p = 0.013), and immunosuppression (p = 0.038) were observed in the 2022/2023 season. In contrast, significantly more lower respiratory tract infections (p < 0.001), in particular in the form of pneumonia (p = 0.015) and exacerbations of obstructive lung diseases (p = 0.008), were detected. An ICU admission was noted for 23.7% of all patients throughout the study period. Sequence analysis of the fusion protein gene revealed a close phylogenetic relatedness, regardless of the season of origin. However, especially for RSV-B, an accumulation of amino acid point substitutions was noted, including in antigenic site Ø. The SARS-CoV-2 pandemic had a tremendous impact on the seasonality of RSV, and the introduction of new vaccination and immunization strategies against RSV warrants further epidemiologic studies of this important pathogen.

Insights on kraft lignin degradation in an anaerobic environment.

Lignin is an aromatic macromolecule and one of the main constituents of lignocellulosic materials. Kraft lignin is generated as a residual by-product of the lignocellulosic biomass industrial process, and it might be used as a feedstock to generate low molecular weight aromatic compounds. In this study, we seek to understand and explore the potential of ruminal bacteria in the degradation of kraft lignin. We established two consortia, KLY and KL, which demonstrated significant lignin-degrading capabilities. Both consortia reached maximum growth after two days, with KLY showing a higher growth and decolorization rate. Additionally, SEM analysis revealed morphological changes in the residual lignin from both consortia, indicating significant degradation. This was further supported by FTIR spectra, which showed new bands corresponding to the C-H vibrations of guaiacyl and syringyl units, suggesting structural transformations of the lignin. Taxonomic analysis showed enrichment of the microbial community with members of the Dickeya genus. Seven metabolic pathways related to lignin metabolism were predicted for the established consortia. Both consortia were capable of consuming aromatic compounds such as 4-hydroxybenzoic acid, syringaldehyde, acetovanillone, and syringic acid, highlighting their capacity to convert aromatic compounds into commercially valuable molecules presenting antifungal activity and used as food preservatives as 4-hydroxyphenylacetic, 3-phenylacetic, and phenylacetic acids. Therefore, the microbial consortia shown in the present work are models for understanding the process of lignin degradation and consumption in bacterial anaerobic communities and developing biological processes to add value to industrial processes based on lignocellulosic biomass as feedstock.

Epidemiology and Management of Pediatric Group A Streptococcal Pneumonia With Parapneumonic Effusion: An Observational Study.

BACKGROUND: During autumn/winter 2022, UK pediatricians reported an unseasonal increase in invasive group A streptococcal infections; a striking proportion presenting with pneumonia with parapneumonic effusion. METHODS: Clinicians across the United Kingdom were requested to submit pseudonymized clinical data using a standardized report form for children (<16 years) admitted between September 30, 2022 and February 17, 2023, with microbiologically confirmed group A streptococcal pneumonia with parapneumonic effusion. RESULTS: From 185 cases submitted, the median patient age was 4.4 years, and 163 (88.1%) were previously healthy. Respiratory viral coinfection was detected on admission for 101/153 (66.0%) children using extended respiratory pathogen polymerase chain reaction panel. Molecular testing was the primary method of detecting group A streptococcus on pleural fluid (86/171; 50.3% samples). Primary surgical management was undertaken in 171 (92.4%) children; 153/171 (89.4%) had pleural drain inserted (96 with fibrinolytic agent), 14/171 (8.2%) had video-assisted thoracoscopic surgery. Fever duration after admission was prolonged (median, 12 days; interquartile range, 9-16). Intravenous antibiotic courses varied in length (median, 14 days; interquartile range, 12-21), with many children receiving multiple broad-spectrum antibiotics, although evidence for additional bacterial infection was limited. CONCLUSIONS: Most cases occurred with viral coinfection, a previously well-recognized risk with influenza and varicella zoster, highlighting the need to ensure routine vaccination coverage and progress on vaccines for other common viruses (eg, respiratory syncytial virus, human metapneumovirus) and for group A streptococcus. Molecular testing is valuable to detect viral coinfection and confirm invasive group A streptococcal diagnosis, expediting the incorporation of cases into national reporting systems. Range and duration of intravenous antibiotics administered demonstrated the need for research on the optimal duration of antimicrobials and improved stewardship.

Robust and sensitive amplicon-based whole-genome sequencing assay of respiratory syncytial virus subtype A and B.

Prevention of respiratory syncytial virus (RSV) infection is now a global health priority, with a long-acting monoclonal antibody and two RSV vaccines recently licenced for clinical use. Most licenced and candidate interventions target the RSV fusion (RSV-F) protein. New interventions may be associated with the spread of mutations, reducing susceptibility to antibody neutralization in RSV-F. There is a need for ongoing longitudinal global surveillance of circulating RSV strains. To achieve this large-scale genomic surveillance, a reliable, high-throughput RSV sequencing assay is required. Here we report an improved high-throughput RSV whole-genome sequencing (WGS) assay performed directly on clinical samples without additional enrichment, using a 4-primer-pool, short-amplicon PCR-tiling approach that is suitable for short-read sequencing platforms. Using upper respiratory tract (URT) RSV-positive clinical samples obtained from a sentinel network of primary care providers and from hospital patients (29.7% and 70.2%, respectively; n = 1,037), collected over the period 2019 to 2023, this assay had a threshold of approximately 4 × 103 to 8 × 103 copies/mL (RSV-B and RSV-A sub-types, respectively) as the lowest amount of virus needed in the sample to achieve >96% of whole-genome coverage at a high-quality level. Using a Ct value of 31 as an empirical cut-off, the overall assay success rate of obtaining >90% genome coverage at a read depth minimum of 20 was 96.83% for clinical specimens successfully sequenced from a total of 1,071. The RSV WGS approach described in this study has increased sensitivity compared to previous approaches and can be applied to clinical specimens without the requirement for enrichment. The updated approach produces sequences of high quality consistently and cost-effectively, suitable for implementation to underpin national programs for the surveillance of RSV genomic variation. IMPORTANCE: In this paper, we report an improved high-throughput respiratory syncytial virus (RSV) whole-genome sequencing (WGS) assay performed directly on clinical samples, using a 4-primer-pool, short-amplicon PCR-tiling approach that is suitable for short-read sequencing platforms. The RSV WGS approach described in this study has increased sensitivity compared to previous approaches and can be applied to clinical specimens without the requirement for enrichment. The updated approach produces sequences of high quality consistently and cost-effectively, suitable for implementation to underpin national and global programs for the surveillance of RSV genomic variation. The quality of sequence produced is essential for preparedness for new interventions in monitoring antigenic escape, where a single point mutation might lead to a reduction in antibody binding effectiveness and neutralizing activity, or indeed in the monitoring of retaining susceptibility to neutralization by existing and new interventions.

Uptake of intra-muscular vitamin K administration after birth: A national cohort study.

AIM: A long-acting monoclonal antibody against RSV (nirsevimab), given as an injection shortly after birth, is currently being rolled out globally. Carer acceptance of intra-muscular (IM) vitamin K, another injection given shortly after birth, could serve to indicate the acceptability of nirsevimab. METHODS: We analysed a national dataset of postnatal health visitor visits in Scotland; individual-level data on gestation were not available. The primary outcome measure was the modality of administration of vitamin K; potential explanatory variables were maternal age, infant ethnicity, English as a first language, and measures of socio-economic deprivation. We examined associations between IM vitamin K administration or oral/no vitamin K and each explanatory variable. RESULTS: From 2019 to 2021, questionnaires were available for 142 857 infants; data was missing for 2.7%. IM Vitamin K uptake was high: 95.5% of carers consented, with 1.1% requesting oral vitamin K and 0.9% refusing vitamin K altogether. Infant ethnicity, use of English as a first language, socio-economic status and maternal age were not associated with reduced uptake of IM vitamin K. CONCLUSION: If IM Vitamin K administration is a valid proxy measure for nirsevimab acceptance, we did not identify groups that might require increased engagement prior to nirsevimab roll-out.

The clinical impact of serious respiratory disease in children under the age of two years during the 2021-2022 bronchiolitis season in England, Scotland and Ireland.

BACKGROUND: Interventions introduced to reduce the spread of SARS-CoV-2 led to a widespread reduction in childhood infections. However, from spring 2021 onwards the United Kingdom and Ireland experienced an unusual out-of-season epidemic of respiratory disease. METHODS: We conducted a prospective observational study (BronchStart), enrolling children 0-23 months of age presenting with bronchiolitis, lower respiratory tract infection or first episode of wheeze to 59 Emergency Departments across England, Scotland and Ireland from May 2021 to April 2022. We combined testing data with national admissions datasets to infer the impact of respiratory syncytial virus (RSV) disease. RESULTS: The BronchStart study collected data on 17,899 presentations for 17,164 children. Risk factors for admission and escalation of care included prematurity and congenital heart disease, but most admissions were for previously healthy term-born children. Of those aged 0-11 months who were admitted and tested for RSV, 1,907/3,912 (48.7%) tested positive. We estimate that every year in England and Scotland 28,561 (95% confidence interval 27,637-29,486) infants are admitted with RSV infection. CONCLUSIONS: RSV infection was the main cause of hospitalisations in this cohort, but 51.3% of admissions in infants were not associated with the virus. The majority of admissions were in previously healthy term-born infants.

Parallel laboratory evolution and rational debugging reveal genomic plasticity to S. cerevisiae synthetic chromosome XIV defects.

Synthetic chromosome engineering is a complex process due to the need to identify and repair growth defects and deal with combinatorial gene essentiality when rearranging chromosomes. To alleviate these issues, we have demonstrated novel approaches for repairing and rearranging synthetic Saccharomyces cerevisiae genomes. We have designed, constructed, and restored wild-type fitness to a synthetic 753,096-bp version of S. cerevisiae chromosome XIV as part of the Synthetic Yeast Genome project. In parallel to the use of rational engineering approaches to restore wild-type fitness, we used adaptive laboratory evolution to generate a general growth-defect-suppressor rearrangement in the form of increased TAR1 copy number. We also extended the utility of the synthetic chromosome recombination and modification by loxPsym-mediated evolution (SCRaMbLE) system by engineering synthetic-wild-type tetraploid hybrid strains that buffer against essential gene loss, highlighting the plasticity of the S. cerevisiae genome in the presence of rational and non-rational modifications.

Have you tried turning it off and on again? Oscillating selection to enhance fitness-landscape traversal in adaptive laboratory evolution experiments.

Adaptive Laboratory Evolution (ALE) is a powerful tool for engineering and understanding microbial physiology. ALE relies on the selection and enrichment of mutations that enable survival or faster growth under a selective condition imposed by the experimental setup. Phenotypic fitness landscapes are often underpinned by complex genotypes involving multiple genes, with combinatorial positive and negative effects on fitness. Such genotype relationships result in mutational fitness landscapes with multiple local fitness maxima and valleys. Traversing local maxima to find a global maximum often requires an individual or sub-population of cells to traverse fitness valleys. Traversing involves gaining mutations that are not adaptive for a given local maximum but are necessary to 'peak shift' to another local maximum, or eventually a global maximum. Despite these relatively well understood evolutionary principles, and the combinatorial genotypes that underlie most metabolic phenotypes, the majority of applied ALE experiments are conducted using constant selection pressures. The use of constant pressure can result in populations becoming trapped within local maxima, and often precludes the attainment of optimum phenotypes associated with global maxima. Here, we argue that oscillating selection pressures is an easily accessible mechanism for traversing fitness landscapes in ALE experiments, and provide theoretical and practical frameworks for implementation.

Trimming the genomic fat: minimising and re-functionalising genomes using synthetic biology.

Naturally evolved organisms typically have large genomes that enable their survival and growth under various conditions. However, the complexity of genomes often precludes our complete understanding of them, and limits the success of biotechnological designs. In contrast, minimal genomes have reduced complexity and therefore improved engineerability, increased biosynthetic capacity through the removal of unnecessary genetic elements, and less recalcitrance to complete characterisation. Here, we review the past and current genome minimisation and re-functionalisation efforts, with an emphasis on the latest advances facilitated by synthetic genomics, and provide a critical appraisal of their potential for industrial applications.

Comparison of UK paediatric SARS-CoV-2 admissions across the first and second pandemic waves.

BACKGROUND: We hypothesised that the clinical characteristics of hospitalised children and young people (CYP) with SARS-CoV-2 in the UK second wave (W2) would differ from the first wave (W1) due to the alpha variant (B.1.1.7), school reopening and relaxation of shielding. METHODS: Prospective multicentre observational cohort study of patients <19 years hospitalised in the UK with SARS-CoV-2 between 17/01/20 and 31/01/21. Clinical characteristics were compared between W1 and W2 (W1 = 17/01/20-31/07/20,W2 = 01/08/20-31/01/21). RESULTS: 2044 CYP < 19 years from 187 hospitals. 427/2044 (20.6%) with asymptomatic/incidental SARS-CoV-2 were excluded from main analysis. 16.0% (248/1548) of symptomatic CYP were admitted to critical care and 0.8% (12/1504) died. 5.6% (91/1617) of symptomatic CYP had Multisystem Inflammatory Syndrome in Children (MIS-C). After excluding CYP with MIS-C, patients in W2 had lower Paediatric Early Warning Scores (PEWS, composite vital sign score), lower antibiotic use and less respiratory and cardiovascular support than W1. The proportion of CYP admitted to critical care was unchanged. 58.0% (938/1617) of symptomatic CYP had no reported comorbidity. Patients without co-morbidities were younger (42.4%, 398/938, <1 year), had lower PEWS, shorter length of stay and less respiratory support. CONCLUSIONS: We found no evidence of increased disease severity in W2 vs W1. A large proportion of hospitalised CYP had no comorbidity. IMPACT: No evidence of increased severity of COVID-19 admissions amongst children and young people (CYP) in the second vs first wave in the UK, despite changes in variant, relaxation of shielding and return to face-to-face schooling. CYP with no comorbidities made up a significant proportion of those admitted. However, they had shorter length of stays and lower treatment requirements than CYP with comorbidities once those with MIS-C were excluded. At least 20% of CYP admitted in this cohort had asymptomatic/incidental SARS-CoV-2 infection. This paper was presented to SAGE to inform CYP vaccination policy in the UK.

Studying the Long-term Impact of COVID-19 in Kids (SLICK). Healthcare use and costs in children and young people following community-acquired SARS-CoV-2 infection: protocol for an observational study using linked primary and secondary routinely collected healthcare data from England, Scotland and Wales.

INTRODUCTION: SARS-CoV-2 infection rarely causes hospitalisation in children and young people (CYP), but mild or asymptomatic infections are common. Persistent symptoms following infection have been reported in CYP but subsequent healthcare use is unclear. We aim to describe healthcare use in CYP following community-acquired SARS-CoV-2 infection and identify those at risk of ongoing healthcare needs. METHODS AND ANALYSIS: We will use anonymised individual-level, population-scale national data linking demographics, comorbidities, primary and secondary care use and mortality between 1 January 2019 and 1 May 2022. SARS-CoV-2 test data will be linked from 1 January 2020 to 1 May 2022. Analyses will use Trusted Research Environments: OpenSAFELY in England, Secure Anonymised Information Linkage (SAIL) Databank in Wales and Early Pandemic Evaluation and Enhanced Surveillance of COVID-19 in Scotland (EAVE-II). CYP aged ≥4 and <18 years who underwent SARS-CoV-2 reverse transcription PCR (RT-PCR) testing between 1 January 2020 and 1 May 2021 and those untested CYP will be examined.The primary outcome measure is cumulative healthcare cost over 12 months following SARS-CoV-2 testing, stratified into primary or secondary care, and physical or mental healthcare. We will estimate the burden of healthcare use attributable to SARS-CoV-2 infections in the 12 months after testing using a matched cohort study of RT-PCR positive, negative or untested CYP matched on testing date, with adjustment for confounders. We will identify factors associated with higher healthcare needs in the 12 months following SARS-CoV-2 infection using an unmatched cohort of RT-PCR positive CYP. Multivariable logistic regression and machine learning approaches will identify risk factors for high healthcare use and characterise patterns of healthcare use post infection. ETHICS AND DISSEMINATION: This study was approved by the South-Central Oxford C Health Research Authority Ethics Committee (13/SC/0149). Findings will be preprinted and published in peer-reviewed journals. Analysis code and code lists will be available through public GitHub repositories and OpenCodelists with meta-data via HDR-UK Innovation Gateway.

Harnessing bioengineered microbes as a versatile platform for space nutrition.

Human enterprises through the solar system will entail long-duration voyages and habitation creating challenges in maintaining healthy diets. We discuss consolidating multiple sensory and nutritional attributes into microorganisms to develop customizable food production systems with minimal inputs, physical footprint, and waste. We envisage that a yeast collection bioengineered for one-carbon metabolism, optimal nutrition, and diverse textures, tastes, aromas, and colors could serve as a flexible food-production platform. Beyond its potential for supporting humans in space, bioengineered microbial-based food could lead to a new paradigm for Earth's food manufacturing that provides greater self-sufficiency and removes pressure from natural ecosystems.

Toward Methanol-Based Biomanufacturing: Emerging Strategies for Engineering Synthetic Methylotrophy in Saccharomyces cerevisiae.

The global expansion of biomanufacturing is currently limited by the availability of sugar-based microbial feedstocks, which require farmland for cultivation and therefore cannot support large increases in production without impacting the human food supply. One-carbon feedstocks, such as methanol, present an enticing alternative to sugar because they can be produced independently of arable farmland from organic waste, atmospheric carbon dioxide, and hydrocarbons such as biomethane, natural gas, and coal. The development of efficient industrial microorganisms that can convert one-carbon feedstocks into valuable products is an ongoing challenge. This review discusses progress in the field of synthetic methylotrophy with a focus on how it pertains to the important industrial yeast, Saccharomyces cerevisiae. Recent insights generated from engineering synthetic methylotrophic xylulose- and ribulose-monophosphate cycles, reductive glycine pathways, and adaptive laboratory evolution studies are critically assessed to generate novel strategies for the future engineering of methylotrophy in S. cerevisiae.

Construction of a synthetic Saccharomyces cerevisiae pan-genome neo-chromosome.

The Synthetic Yeast Genome Project (Sc2.0) represents the first foray into eukaryotic genome engineering and a framework for designing and building the next generation of industrial microbes. However, the laboratory strain S288c used lacks many of the genes that provide phenotypic diversity to industrial and environmental isolates. To address this shortcoming, we have designed and constructed a neo-chromosome that contains many of these diverse pan-genomic elements and which is compatible with the Sc2.0 design and test framework. The presence of this neo-chromosome provides phenotypic plasticity to the Sc2.0 parent strain, including expanding the range of utilizable carbon sources. We also demonstrate that the induction of programmable structural variation (SCRaMbLE) provides genetic diversity on which further adaptive gains could be selected. The presence of this neo-chromosome within the Sc2.0 backbone may therefore provide the means to adapt synthetic strains to a wider variety of environments, a process which will be vital to transitioning Sc2.0 from the laboratory into industrial applications.

Differences in the metabolic and functional mechanisms used to tolerate flooding in Guazuma ulmifolia (Lam.) from flood-prone Amazonian and dry Cerrado savanna populations.

Flood tolerance is crucial to the survival of tree species subject to long periods of flooding, such as those present in the Amazonian várzea. Tolerance can be mediated by adjustments of metabolism, physiology and morphology, reinforcing the need to investigate the physiological and biochemical mechanisms used by tropical tree species to survive this stress. Moreover, such mechanisms may vary between populations that are subjected to differences in the frequency of flooding events. Here, we aimed to identify the mechanisms used by two populations of the tropical tree Guazuma ulmifolia (Lam.) to tolerate flooding: an Amazonian population frequently exposed to flooding and a Cerrado population, adapted to a dry environment. Young plants were subjected to a flooding of the roots and lower stem for 32 days, followed by 17 days of recovery. Amazonian plants exhibited greater increases in shoot length and higher maximum photosynthetic rate (Amax) compared with non-flooded plants from 7 days of flooding onwards, whereas increased Amax occurred later in flooded Cerrado plants and was not accompanied by increased shoot length. Lactate accumulated in roots of Cerrado plants after 24 h flooding, together with transcripts coding for lactate dehydrogenase in roots of both Cerrado and Amazonian plants. After 7 days of flooding, lactate decreased and alcohol dehydrogenase activity increased transiently, together with concentrations of alanine, γ-aminobutyric acid and succinate, indicating activation of metabolic processes associated with low oxygen availability. Other amino acids also increased in flooded Cerrado plants, revealing more extensive metabolic changes than in Amazonian plants. Wetland and dryland populations of G. ulmifolia revealed the great capacity to tolerate flooding stress through a suite of alterations in photosynthetic gas exchange and metabolism. However, the integrated physiological, biochemical and molecular analyses realized here indicated that wetland plants acclimatized more efficiently with increased shoot elongation and more rapid restoration of normal metabolism.