Recent advances in bio-based production of top platform chemical, succinic acid: an alternative to conventional chemistry.

Succinic acid (SA) is one of the top platform chemicals with huge applications in diverse sectors. The presence of two carboxylic acid groups on the terminal carbon atoms makes SA a highly functional molecule that can be derivatized into a wide range of products. The biological route for SA production is a cleaner, greener, and promising technological option with huge potential to sequester the potent greenhouse gas, carbon dioxide. The recycling of renewable carbon of biomass (an indirect form of CO2), along with fixing CO2 in the form of SA, offers a carbon-negative SA manufacturing route to reduce atmospheric CO2 load. These attractive attributes compel a paradigm shift from fossil-based to microbial SA manufacturing, as evidenced by several commercial-scale bio-SA production in the last decade. The current review article scrutinizes the existing knowledge and covers SA production by the most efficient SA producers, including several bacteria and yeast strains. The review starts with the biochemistry of the major pathways accumulating SA as an end product. It discusses the SA production from a variety of pure and crude renewable sources by native as well as engineered strains with details of pathway/metabolic, evolutionary, and process engineering approaches for enhancing TYP (titer, yield, and productivity) metrics. The review is then extended to recent progress on separation technologies to recover SA from fermentation broth. Thereafter, SA derivatization opportunities via chemo-catalysis are discussed for various high-value products, which are only a few steps away. The last two sections are devoted to the current scenario of industrial production of bio-SA and associated challenges, along with the author's perspective.

Arabinose as an overlooked sugar for microbial bioproduction of chemical building blocks.

The circular economy is anticipated to bring a disruptive transformation in manufacturing technologies. Robust and industrial scalable microbial strains that can simultaneously assimilate and valorize multiple carbon substrates are highly desirable, as waste bioresources contain substantial amounts of renewable and fermentable carbon, which is diverse. Lignocellulosic biomass (LCB) is identified as an inexhaustible and alternative resource to reduce global dependence on oil. Glucose, xylose, and arabinose are the major monomeric sugars in LCB. However, primary research has focused on the use of glucose. On the other hand, the valorization of pentose sugars, xylose, and arabinose, has been mainly overlooked, despite possible assimilation by vast microbial communities. The present review highlights the research efforts that have explicitly proven the suitability of arabinose as the starting feedstock for producing various chemical building blocks via biological routes. It begins by analyzing the availability of various arabinose-rich biorenewable sources that can serve as potential feedstocks for biorefineries. The subsequent section outlines the current understanding of arabinose metabolism, biochemical routes prevalent in prokaryotic and eukaryotic systems, and possible products that can be derived from this sugar. Further, currently, exemplar products from arabinose, including arabitol, 2,3-butanediol, 1,2,3-butanetriol, ethanol, lactic acid, and xylitol are discussed, which have been produced by native and non-native microbial strains using metabolic engineering and genome editing tools. The final section deals with the challenges and obstacles associated with arabinose-based production, followed by concluding remarks and prospects.

Fermentative valorisation of xylose-rich hemicellulosic hydrolysates from agricultural waste residues for lactic acid production under non-sterile conditions.

Lactic acid (LA) is a platform chemical with diverse industrial applications. Presently, commercial production of LA is dominated by microbial fermentation using sugary or starch-based feedstocks. Research pursuits emphasizing towards sustainable production of LA using non-edible and renewable feedstocks have accelerated the use of lignocellulosic biomass (LCB). The present study focuses on the valorisation of xylose derived from sugarcane bagasse (SCB) and olive pits (OP) through hydrothermal and dilute acid pretreatment, respectively. The xylose-rich hydrolysate obtained was used for LA production by homo-fermentative and thermophilic Bacillus coagulans DSM2314 strain under non-sterile conditions. The fed-batch mode of fermentation resulted in maximum LA titers of 97.8, 52.4 and 61.3 g/L with a yield of 0.77, 0.66 and 0.71 g/g using pure xylose, xylose-rich SCB and OP hydrolysates, respectively. Further, a two-step aqueous two-phase system (ATPS) extraction technique was employed for the separation and recovery of LA accumulated on pure and crude xylose. The LA recovery was 45 - 65% in the first step and enhanced to 80-90% in the second step.The study demonstrated an efficient integrated biorefinery approach to valorising the xylose-rich stream for cost-effective LA production and recovery.

Strategies and tools for the biotechnological valorization of glycerol to 1, 3-propanediol: Challenges, recent advancements and future outlook.

Global efforts towards decarbonization, environmental sustainability, and a growing impetus for exploiting renewable resources such as biomass have spurred the growth and usage of bio-based chemicals and fuels. In light of such developments, the biodiesel industry will likely flourish, as the transport sector is taking several initiatives to attain carbon-neutral mobility. However, this industry would inevitably generate glycerol as an abundant waste by-product. Despite being a renewable organic carbon source and assimilated by several prokaryotes, presently realizing glycerol-based biorefinery is a distant reality. Among several platform chemicals such as ethanol, lactic acid, succinic acid, 2, 3-butanediol etc., 1, 3-propanediol (1, 3-PDO) is the only chemical naturally produced by fermentation, with glycerol as a native substrate. The recent commercialization of glycerol-based 1, 3-PDO by Metabolic Explorer, France, has revived research interests in developing alternate cost-competitive, scalable and marketable bioprocesses. The current review outlines natural glycerol assimilating and 1, 3-PDO-producing microbes, their metabolic pathways, and associated genes. Later, technical barriers are carefully examined, such as the direct use of industrial glycerol as input material and genetic and metabolic issues related to microbes alleviating their industrial use. Biotechnological interventions exploited in the past five years, which can substantially circumvent these challenges, such as microbial bioprospecting, mutagenesis, metabolic, evolutionary and bioprocess engineering, including their combinations, are discussed in detail. The concluding section sheds light on some of the emerging and most promising breakthroughs which have resulted in evolving new, efficient, and robust microbial cell factories and/or bioprocesses for glycerol-based 1, 3-PDO production.

Development of Hypertolerant Strain of Yarrowia lipolytica Accumulating Succinic Acid Using High Levels of Acetate.

Acetate is emerging as a promising feedstock for biorefineries as it can serve as an alternate carbon source for microbial cell factories. In this study, we expressed acetyl-CoA synthase in Yarrowia lipolytica PSA02004PP, and the recombinant strain grew on acetate as the sole carbon source and accumulated succinic acid or succinate (SA). Unlike traditional feedstocks, acetate is a toxic substrate for microorganisms; therefore, the recombinant strain was further subjected to adaptive laboratory evolution to alleviate toxicity and improve tolerance against acetate. At high acetate concentrations, the adapted strain Y. lipolytica ACS 5.0 grew rapidly and accumulated lipids and SA. Bioreactor cultivation of ACS 5.0 with 22.5 g/L acetate in a batch mode resulted in a maximum cell OD600 of 9.2, with lipid and SA accumulation being 0.84 and 5.1 g/L, respectively. However, its fed-batch cultivation yielded a cell OD600 of 23.5, SA titer of 6.5 g/L, and lipid production of 1.5 g/L with an acetate uptake rate of 0.2 g/L h, about 2.86 times higher than the parent strain. Cofermentation of acetate and glucose significantly enhanced the SA titer and lipid accumulation to 12.2 and 1.8 g/L, respectively, with marginal increment in cell growth (OD600: 26.7). Furthermore, metabolic flux analysis has drawn insights into utilizing acetate for the production of metabolites that are downstream to acetyl-CoA. To the best of our knowledge, this is the first report on SA production from acetate by Y. lipolytica and demonstrates a path for direct valorization of sugar-rich biomass hydrolysates with elevated acetate levels to SA.

Emerging trends in high-solids enzymatic saccharification of lignocellulosic feedstocks for developing an efficient and industrially deployable sugar platform.

For the techno-commercial success of any lignocellulosic biorefinery, the cost-effective production of fermentable sugars for the manufacturing of bio-based products is indispensable. High-solids enzymatic saccharification (HSES) is a straightforward approach to develop an industrially deployable sugar platform. Economic incentives such as reduced capital and operational expenditure along with environmental benefits in the form of reduced effluent discharge makes this strategy more lucrative for exploitation. However, HSES suffers from the drawback of non-linear and disproportionate sugar yields with increased substrate loadings. To overcome this bottleneck, researchers tend to perform HSES at high enzyme loadings. Nonetheless, the production costs of cellulases are one of the key contributors that impair the entire process economics. This review highlights the relentless efforts made globally to attain a high-titer of sugars and their fermentation products by performing efficient HSES at low cellulase loadings. In this context, technical innovations such as advancements in new pretreatment strategies, next-generation cellulase cocktails, additives, accessory enzymes, novel reactor concepts and enzyme recycling studies are especially showcased. This review further covers new insights, learnings and prospects in the area of lignocellulosic bioprocessing.

Indian Academy of Pediatrics (IAP) Task Force Recommendations for Incorporating Nurturing Care for Early Childhood Development (NC-ECD) in Medical Education in India.

BACKGROUND: The World Health Organization (WHO) recommends promotion of nurturing care for early childhood development (NC-ECD) by focusing on five essential components viz., good health, adequate nutrition, promotion of early childhood learning, responsive caregiving, and safety and security. Indian medical graduates and pediatricians are the keys to successful delivery and propagation of NC-ECD in the community. Their training therefore needs to include skills and knowledge needed to promote and practice ECD. OBJECTIVE: To evaluate the existing undergraduate (UG) and postgraduate (PG) curricula of pediatrics for components related to early childhood development, assess gaps in the training essential to practice and promote ECD, and suggest recommendations to incorporate NC-ECD in the UG and PG curricula. PROCESS: Indian Academy of Pediatrics created a task force to review the UG/PG medical curricula, consisting of experts from pediatrics and medical education. The task force deliberated on 20 March, 2021 and identified the gaps in current curricula and provided suggestions to strengthen it. The recommendations of the task force are presented here. RECOMMENDATIONS: Taskforce identified that the UG/PG medical curricula are lacking training for propagating early childhood learning, responsive caregiving, caregiver support, and ensuring safety and security of children. The taskforce provided a list of competencies related to ECD that need to be included in both UG and PG curriculum. NC-ECD should also be included in topics for integrated teaching. Postgraduates also need to be exposed to hands-on-training at anganwadis, creches, and in domestic setting.

Tracheostomy Tube as Foreign Body in Right Main Bronchus: A Case Report.

Tracheostomy is a commonly performed procedure to secure airway in patients who need prolonged airway support (Marchese et al. in Respir Med 104(5):749-753, 2010). It is relatively safe procedure but associated with few early and late complications (Fernandez-Bussy et al. in J Bronchol Interv Pulmonol 22(4):357-364, 2015). Metallic tracheostomy tubes were used in patients in the past. Those tubes were associated with fracture and dislodgement due to corrosive injury. Few case reports have been published in the past (Lynrah et al. in Int J Pediatr Otorhinolaryngol 76(11):1691-1695, 2012; Agarwal and Agarwal in Indian J Chest Dis Allied Sci 53(2):111, 2011). PVC tubes are less prone for fracture, but wear and tear associated with prolonged use can lead to break in the tube and dislodgement. We successfully diagnosed and managed such a case of broken PVC Tracheostomy tube in right main bronchus and share our experiences.

India's Policy and Program Landscape for Early Childhood Development.

Despite continuous improvement in health and nutrition-related child outcomes in India in the last decade, sub-optimal developmental outcomes still prevail due to multiple risk factors. Overcoming these risks depends on a supportive policy environment and implementation across relevant sectors to achieve universal coverage for all children and their families, especially those at-risk and those already affected by developmental disorders/disability. Several national policies articulate the multi-dimensional and multisectoral vision for achieving early childhood development (ECD), focusing on the first 1000 days and reaching the most vulnerable children. The enactment of various 'Acts' entitles children to their right to nutrition, safety, and security. Flagship programs of various Ministries translate this vision into action through various schemes that provide services in each of the five domains of nurturing care. Public spending is the largest source of funding for ECD programs while Corporate Social Responsibility is emerging as a promising funding opportunity. Ensuring effective implementation of ECD by developing a shared framework for implementation across sectors, establishing a robust governance mechanism, and sustainable financing strategies for universal access is the recommended way forward.

Promoting Nurturing Care for Early Childhood Development Through India's Public Health System.

Implementing the nurturing care framework (NCF) for early childhood development (ECD) is essentially multisectoral, requiring coordination amongst all sectors and harmoniously integrating it within the existing contact opportunities in the health sector. This paper discusses the relative strengths, persisting gaps, challenges, and the way forward to implement nurturing care for ECD through the public health system. The vast network of frontline health workers and health facilities; community, home, and center-based service delivery; health and wellness centers located close to the communities have the potential to promote nurturing care. Persisting gaps include limited capacities of health workers in the nurturing care domains, lack of community engagement for ECD, weak referral linkages, inability to reach the most vulnerable children, missed opportunities for early identification of children at risk, and early intervention for children developmental delays and difficulties. Moving forward, incorporating nurturing care components into essential services packages, enhancing competencies of health workers, engaging with parents, establishing a mechanism for tracking children at risk, and developmental surveillance by trained service providers can provide the much-needed impetus to ECD.

Intersectoral Action for Early Childhood Development in India: Opportunities, Challenges and the Way Forward.

Nurturing care framework for early childhood development (ECD) focuses on five essential aspects of the holistic development of a child, which are interrelated and inseparable. This multidimensional approach to child development is dependent on contributions from multiple sectors, requiring the 'whole of government' approach. In India, the lack of a single multisectoral framework for ECD, narrow accountability to sector-specific outcomes, overlapping responsibilities of frontline workers, lack of leadership for coordination, and limited supervisory mechanisms result in fragmented service delivery. In recent years, there is high-level political commitment to intersectoral action, which promote holistic health. Better results and developmental outcomes are possible with different sectors working closely by converging their resources under the Sustainable Development Goals strategic action plans, POSHAN Abhiyaan, and the Aspirational districts program. Leveraging opportunities for intersectoral action requires a deliberate and consistent effort towards alignment of goals, favorable conditions of partnerships, leadership and governance, and capacity at every level.

Family-Centered Care for Newborns: From Pilot Implementation to National Scale-up in India.

Family-centered care (FCC) is a well-evidenced approach, recognized as the standard of care for newborns and children. This approach promotes a mutually beneficial partnership between health professionals and infant/young children's families, providing an opportunity for developing trustful relations and transparency of care. Implementation experience from our newborn intensive care unit highlighted three prerequisites for successful implementation, which include adequate infrastructure and basic amenities for the primary caregiver; attitudinal change amongst healthcare providers accepting parents/family as partners in the care of the newborn; and active involvement of primary caregivers in baby care activities alongside the nurses. Healthcare providers played a crucial role in empowering parents, improving their confidence and competence to transition into the role of primary caregivers after discharge. FCC contributes to all domains of nurturing care and has relevance in newborn care and pediatric care at all levels, with scope for being expanded to the antenatal and postnatal period to promote early childhood development. With national scale-up in progress, addressing actual or perceived barriers to implementation requires context-specific adaptation and the best use of opportunities and funding support available under the national health program.

High yield recovery of 2,3-butanediol from fermented broth accumulated on xylose rich sugarcane bagasse hydrolysate using aqueous two-phase extraction system.

Downstream processing of chemicals obtained from fermentative route is challenging and cost-determining factor of any bioprocess. 2,3-Butanediol (BDO) is a promising chemical building block with myriad applications in the polymer, food, pharmaceuticals, and fuel sector. The current study focuses on the recovery and purification of BDO produced (68.2 g/L) from detoxified xylose-rich sugarcane bagasse hydrolysate by a mutant strain of Enterobacter ludwigii. Studies involving screening and optimization of aqueous-two phase system (ATPS) revealed that 30% w/v (NH4)2SO4 addition to clarified fermented broth facilitated BDO extraction in isopropanol (0.5 v/v), with maximum recovery and partition coefficient being 97.9 ± 4.6% and 45.5 ± 3.5, respectively. The optimized protocol was repeated with unfiltered broth containing 68.2 g/L BDO, cell biomass, and unspent protein, which led to the partitioning of 66.7 g/L BDO, 2.0 g/L xylose and 9.0 g/L acetic acid into organic phase with similar BDO recovery (97%) and partition coefficient (45).

Indian Academy of Pediatrics Position Paper on Nurturing Care for Early Childhood Development.

Early childhood development (ECD) refers to the physical, motor, socio-emotional, cognitive, and linguistic development of a young child. The 'Countdown to 2030' global distribution of 'children at risk of poor development' indicates the need for urgent action and investment in ECD. Nurturing care enhances ECD, even in the presence of adversities. Strategic actions should exist at multiple levels: the family, community, health care providers and government. Previously, child health related policies and programs of the Government of India functioned in isolation, but have recently started demonstrating multi-sectoral collaboration. Nonetheless, the status of ECD in India is far from optimal. There is strong evidence that parenting programs improve outcomes related to ECD. This is dependent on key programmatic areas (timing, duration, frequency, intensity, modality, content, etc.), in addition to political will, funding, partnership, and plans for scaling up. Each country must implement its unique ECD program that is need-based and customized to their stakeholder community. Barriers like inadequate sensitization of the community and low competency of health care providers need to be overcome. IAP firmly believes that responsive parenting interventions revolving around nurturing care should be incorporated in office practice. This paper outlines IAP's position on ECD, and its recommendations for pediatricians and policy makers. It also presents the roadmap in partnership with other stakeholders in maternal, neonatal, and child health; Federation of Obstetric and Gynaecological Societies of India (FOGSI), National Neonatology Forum (NNF), World Health Organization (WHO), and United Nation Children Fund (UNICEF).

MLKL promotes cellular differentiation in myeloid leukemia by facilitating the release of G-CSF.

The blockade of cellular differentiation represents a hallmark of acute myeloid leukemia (AML), which is largely attributed to the dysfunction of lineage-specific transcription factors controlling cellular differentiation. However, alternative mechanisms of cellular differentiation programs in AML remain largely unexplored. Here we report that mixed lineage kinase domain-like protein (MLKL) contributes to the cellular differentiation of transformed hematopoietic progenitor cells in AML. Using gene-targeted mice, we show that MLKL facilitates the release of granulocyte colony-stimulating factor (G-CSF) by controlling membrane permeabilization in leukemic cells. Mlkl-/- hematopoietic stem and progenitor cells released reduced amounts of G-CSF while retaining their capacity for CSF3 (G-CSF) mRNA expression, G-CSF protein translation, and G-CSF receptor signaling. MLKL associates with early endosomes and controls G-CSF release from intracellular storage by plasma membrane pore formation, whereas cell death remained unaffected by loss of MLKL. Of note, MLKL expression was significantly reduced in AML patients, specifically in those with a poor-risk AML subtype. Our data provide evidence that MLKL controls myeloid differentiation in AML by controlling the release of G-CSF from leukemic progenitor cells.

Acetate as a potential feedstock for the production of value-added chemicals: Metabolism and applications.

Acetate is regarded as a promising carbon feedstock in biological production owing to its possible derivation from C1 gases such as CO, CO2 and methane. To best use of acetate, comprehensive understanding of acetate metabolisms from genes and enzymes to pathways and regulations is needed. This review aims to provide an overview on the potential of acetate as carbon feedstock for industrial biotechnology. Biochemical, microbial and biotechnological aspects of acetate metabolism are described. Especially, the current state-of-the art in the production of value-added chemicals from acetate is summarized. Challenges and future perspectives are also provided.

Valorisation of xylose to renewable fuels and chemicals, an essential step in augmenting the commercial viability of lignocellulosic biorefineries.

Biologists and engineers are making tremendous efforts in contributing to a sustainable and green society. To that end, there is growing interest in waste management and valorisation. Lignocellulosic biomass (LCB) is the most abundant material on the earth and an inevitable waste predominantly originating from agricultural residues, forest biomass and municipal solid waste streams. LCB serves as the renewable feedstock for clean and sustainable processes and products with low carbon emission. Cellulose and hemicellulose constitute the polymeric structure of LCB, which on depolymerisation liberates oligomeric or monomeric glucose and xylose, respectively. The preferential utilization of glucose and/or absence of the xylose metabolic pathway in microbial systems cause xylose valorization to be alienated and abandoned, a major bottleneck in the commercial viability of LCB-based biorefineries. Xylose is the second most abundant sugar in LCB, but a non-conventional industrial substrate unlike glucose. The current review seeks to summarize the recent developments in the biological conversion of xylose into a myriad of sustainable products and associated challenges. The review discusses the microbiology, genetics, and biochemistry of xylose metabolism with hurdles requiring debottlenecking for efficient xylose assimilation. It further describes the product formation by microbial cell factories which can assimilate xylose naturally and rewiring of metabolic networks to ameliorate xylose-based bioproduction in native as well as non-native strains. The review also includes a case study that provides an argument on a suitable pathway for optimal cell growth and succinic acid (SA) production from xylose through elementary flux mode analysis. Finally, a product portfolio from xylose bioconversion has been evaluated along with significant developments made through enzyme, metabolic and process engineering approaches, to maximize the product titers and yield, eventually empowering LCB-based biorefineries. Towards the end, the review is wrapped up with current challenges, concluding remarks, and prospects with an argument for intense future research into xylose-based biorefineries.

Enhanced xylitol production using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse by newly isolated Pichia fermentans.

BACKGROUND: Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable lignocellulosic biorefineries. In this aspect, microbial cell factories are harnessed for the sustainable production of commercially viable biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, in the terrestrial ecosystem, microbial systems can efficiently consume glucose. On the contrary, pentose sugars are less preferred carbon source as most of the microbes lack metabolic pathway for their utilization. The effective utilization of both pentose and hexose sugars is key for economical biorefinery. RESULTS: Bioprospecting the food waste and selective enrichment on xylose-rich medium led to screening and isolation of yeast which was phylogenetically identified as Pichia fermentans. The newly isolated xylose assimilating yeast was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with > 40% conversion yield. Chemical mutagenesis of isolated yeast with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min EMS exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titer and yield of 34.0 g/L and 0.68 g/g, respectively. However, under the same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of the mutant for xylitol accumulation. The xylitol titer and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively. In comparison, xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g. CONCLUSION: This study demonstrates the potential of newly isolated P. fermentans in successfully valorising the hemicellulosic fraction for the sustainable xylitol production.

MCL-1 gains occur with high frequency in lung adenocarcinoma and can be targeted therapeutically.

Evasion of programmed cell death represents a critical form of oncogene addiction in cancer cells. Understanding the molecular mechanisms underpinning cancer cell survival despite the oncogenic stress could provide a molecular basis for potential therapeutic interventions. Here we explore the role of pro-survival genes in cancer cell integrity during clonal evolution in non-small cell lung cancer (NSCLC). We identify gains of MCL-1 at high frequency in multiple independent NSCLC cohorts, occurring both clonally and subclonally. Clonal loss of functional TP53 is significantly associated with subclonal gains of MCL-1. In mice, tumour progression is delayed upon pharmacologic or genetic inhibition of MCL-1. These findings reveal that MCL-1 gains occur with high frequency in lung adenocarcinoma and can be targeted therapeutically.