Population-based genomic newborn screening identifying newborns at risk for early-onset cancers has not been evaluated. Here, within a Michigan birth cohort (1987-2020), we identify all children developing a solid or central nervous system malignancy by age 8 years (n = 1948). We perform targeted sequencing of 11 cancer predisposition genes using archived newborn dried blood spot DNA. We find pathogenic or likely-pathogenic germline variants (P/LP) in 6.8% of cases (n = 132): RB1 (n = 69), TP53 (n = 24), SMARCB1 (n = 8), WT1 (n = 7), RET (n = 6), SUFU (n = 6), PTCH1 (n = 4), DICER1 (n = 4), APC (n = 3) and PHOX2B (n = 1). We show approximately 1/27,000 newborns develop an early-onset malignancy with an associated pathogenic or likely-pathogenic variant. Germline variant prevalence is 100% in medullary thyroid carcinoma, 40% in retinoblastoma, and 11-30% across five additional diagnoses, with strong gene-tumor specificity (p < 0.001). P/LP variants are rare in comparison datasets from healthy newborns and gnomAD. Our data support newborn screening for selected cancer-risk genes.

With the day to day increase in energy consumption due to increase in urbanization production of bioethanol is highly in demand. At this point where the traditional methods are not able to suffice the demands due to its high cost and low productivity, new methods need to be developed. This review aims to understand the importance and the regulation of ADH2 in Saccharomyces cerevisiae because Adh2p is the only enzyme that initiates the reaction for the conversion of ethanol, the end product of fermentation to acetaldehyde. The effect of glucose on regulatory mechanisms of Alcohol dehydrogenase II (ADH2) with respect to Snf1 kinase, Target of Rapamycin (TOR) and CCR4 (Carbon Catabolite Repression) pathway on S. cerevisiae are discussed. Snf1 is a serine threonine kinase which is inactive in presence of high glucose concentrations and gets activated in low glucose environments which in turn affects the transcription of ADH2 by controlling the upstream TFs (Transcription Factors). TOR pathway is an essential signalling network that senses the availability of nutrients, mostly glucose and amino acids. This gets activated in presence of glucose. TORC1 regulates the transcription of ADH2 via various downstream transcription factors like Sch9p, Rim15, etc. Another global transcription factor CCR4, regulates ADH2 by acting directly upon its promoter region. The unique function of Adh2p in yeast metabolism, has directed numerous research work making it a vital target. Genetic manipulation of ADH2 gene has proved to be beneficial for food, bioethanol industry.

Nitrosative stress is a phenomenon where reactive nitrogen species (RNS), oxidizes different cellular macromolecules. In this study we investigated the effect of sub-toxic dose of different nitrosative stress agents on S. cerevisiae grown with 2% ethanol as sole carbon source. Our SEM analysis showed significant increase in biofilm production under stress with changes in cellular morphology. Genes responsible for biofilm formation like FLO11, BSC2 and MAC1 in S. cerevisiae upon treatment with ac. NaNO2 and SNP were also found to be upregulated. Redox enzymes like glutathione reductase (GR) showed an increase in specific activity in treated sets but catalase activity had no significant difference. Utilization of ethanol as sole carbon source was followed primarily by assessing the specific activity of Adh2p, which showed a significant 4-fold increase in both treated sets. This was also corroborated with gene expression analysis of ADH2. Together with this estimation of important enzymes of other associated metabolic pathways were also done to understand the changes in metabolic flux. Importantly both glyoxylate cycle and TCA cycle were found to be partially blocked under stress condition whereas, aldehyde reductase (Bdh2p) an important enzyme for 2,3-Butanediol production was found to have upregulated significantly. Altogether our study provides the first report on the effect of nitrosative stress on S. cerevisiae grown on ethanol as a carbon source with possibility to produce 2,3-Butanediol, an industrially important compound, that has a huge demand in the paint, drug and cosmetic industries.

Very early onset rare pediatric cancers such as retinoblastoma are not included in current public health newborn screening (NBS), even though early detection can prevent serious health problems. Currently, babies are screened for retinoblastoma using the “red-reflex” eye test, but this method often misses cancers, leading to delayed diagnoses and poor outcomes. Modern genomic technologies such as targeted Next-Generation Sequencing (tNGS) can detect causal variants in the RB1 gene and other cancer predisposition syndrome (CPS) genes, but applying tNGS individually to every newborn remains cost-prohibitive for NBS implementation. We propose that DNA pooling strategies can further reduce per-sample costs, making CPS gene testing feasible for NBS enabling earlier detection and monitoring of high-risk infants. As part of a proof-of-concept cancer predisposition screening study, we obtained tNGS data from archived dried blood spots (DBS) — the standard NBS sample — from the Michigan BioTrust for Health, from 1,948 children diagnosed with CPS before age eight. To evaluate the pooling approach, we organized DBS DNA extracted from 28 different newborns into groups: a 4×4 grid of 16 samples and one additional row and column of 10 samples overlapping with the 4 x 4 grid. The samples were pooled across rows and columns (16 samples in 4 x 4 grid resulted in 8 pools) and the remaining two row and column sample pools were in pools of 10. We performed custom tNGS and bioinformatics processing for these 10 pools (with either 4- or 10-sample pools) targeting RB1 and other cancer-associated genes (TP53, WT1, APC, SMARCB1, DICER1, PTCH1, SUFU, RET). A custom algorithm deconvoluted the pooled sequencing data to pinpoint which individual DBS sample carried a causal rare (pathogenic) variant. Among the 28 samples, two carried pathogenic RB1 variants (c.751C>T [p.Arg251Ter] and c.1333C>T [p.Arg454Ter]). Both variants were within the 4 x 4 grid and were correctly assigned to the appropriate individual. Within the larger 10-sample pools, only one RB1 variant was pinpointed (as anticipated), highlighting the success of both pooling approaches. This approach preserved sensitivity and specificity while demonstrating an estimated 2–5-fold library prep cost reduction versus individual tNGS. Preliminary testing across additional algorithm settings, CPS genes and cohorts will be presented. Pooling DBS for tNGS in public health NBS is feasible, accurate, and more cost-effective for detecting rare early-onset pediatric cancer risks. This supports a transformative “cancer-risk NBS” paradigm, enabling proactive surveillance and earlier intervention for at-risk infants. These findings provide a strong basis for pilot implementation in real-world NBS programs, with potential to expand to additional actionable conditions. Arindam Bhattacharjee, Mads C Nielsen, Ulrik K Stoltze, Lisa Diller, Richard B Parad. Feasibility of Low-Cost Genomic Newborn Screening for Pediatric Cancers via Targeted NGS Pooling [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Discovery and Innovation in Pediatric Cancer— From Biology to Breakthrough Therapies; 2025 Sep 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85(18_Suppl_2):Abstract nr B029.

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Nitrosative stress is an adverse physiological condition mediated by an excessive level of reactive nitrogen species (RNS). RNS react with the different macromolecules in vivo and result in the inactivation of these molecules. But the mechanism to counteract the effect of nitrosative stress is poorly understood. Escherichia coli is one of the best understood and well-studied microorganism. Although several studies have been reported on Escherichia coli to characterize the effect of various stress response but fewer works are there to see the effect of nitrosative stress. Escherichia coli encounter numerous stresses during its growth, survival, and infection. They respond to various stress conditions by activating common regulator proteins and thiols. These stress conditions result in the accumulation of these regulator proteins and thiols that allow cells to adjust to specific stress situations, conferring stress tolerance and survival. In this review, different enzymatic and non-enzymatic mechanisms to counteract the effect of nitrosative stress in Escherichia coli have been discussed and a hypothesis for the working mechanism of hybrid cluster protein that helps to combat nitrosative stress has been proposed. Here, we have tried to give a clear scenario about the mode of action of stress-responsive elements present in Escherichia coli .

AIMS:The work aimed to understand the important changes during glucose metabolism in Saccharomyces cerevisiae under acidified sodium nitrite (ac.NaNO2 ) mediated nitrosative stress.METHODS AND RESULTS:Confocal microscopy and fluorescence-activated cell sorting analysis were performed to investigate the generation of reactive nitrogen and oxygen species, and redox homeostasis under nitrosative stress was also characterized. Quantitative PCR analysis revealed that the expression of ADH genes was upregulated under such condition, whereas the ACO2 gene was downregulated. Some of the enzymes of the tricarboxylic acid cycle were partially inhibited, whereas malate metabolism and alcoholic fermentation were increased under nitrosative stress. Kinetics of ethanol production was also characterized. A network analysis was conducted to validate our findings. In the presence of ac.NaNO2 , in vitro protein tyrosine nitration formation was checked by western blotting using pure alcohol dehydrogenase and aconitase.CONCLUSIONS:Alcoholic fermentation rate was increased under stress condition and this altered metabolism might be conjoined with the defence machinery to overcome the nitrosative stress.SIGNIFICANCE AND IMPACT OF THE STUDY:This is the first work of this kind where the role of metabolism under nitrosative stress has been characterized in S. cerevisiae and it will provide a base to develop an alternative method of industrial ethanol production.

The inspection of molecular interaction widespread in [L-Tert-leucine] and in aqueous solutions of αand cyclodextrins (Cyclic Oligosaccharides) have been probed by thermo physical properties.Tensiometry and conductivity studies establish development of 1:1 inclusion complex.Interaction coverage is articulated by Limiting apparent molar volume ( 0 V) and Viscosity B-coefficients.Solvent plays a major role in 'solute-solute' and 'solute-solvent' interactions in complex formed, which are examined via stability constant-NMR titration, Steady state Fluorescence, HRMS, SEM, HRTEM, Hydrogen bonds, hydrophobic and structural results for construction of inclusion complexes.Cell viability and Cytotoxicity showed potential effectiveness in the studied drug delivery systems.

When exposed to nitmsative stress, the redox status of Saccharomyces cerevisiae changes significantly in vivo. Under nitrosative stress, aconitase, which catalyzes the conversion of citrate to isocitrate in the tricarboxylic acid (TCA) cycle, is known to be vulnerable. In this study, aconitase was completely repressed in the presence of 0.25 mM S-nitrosoglutathione (GSNO) as the nitrosative stress agent. Furthermore, a similar to 1.5 fold increase in ethanol production and a 3.5 fold increase in alcohol dehydmgenase (ADH) activity were observed in the presence of 0.25 mM GSNO when compared to the control (untreated). Furthermore, we supported our findings with a gene expression study of the adh3 gene, which showed a 4 fold increase in the presence of 0.25 mM GSNO. This is the first report of its kind to characterize ethanol production under GSNO stress. This study may prove to be industrially significant in ethanol production.

Background: The Himalayas have always been an enigma and, being biodiversity hotspots, are considered extremely important from an ecological point of view. Recent advances in studies regarding high-altitude lakes have garnered relevant importance as these habitats could harbor potential psychrophilic and psychrotrophic microbes with bio-prospective applications. Contemplating the above scenario, the present study has been undertaken to understand the diversity and the functional capacities of the microbes thriving in this lake. Results: In our present study on Samiti Lake, the abundance of Proteobacteria as the major phylum was seen in both the soil and water samples. Incase of the ABSLW (water) and ABS1 (soil) sample, 148,066 and 239,754 predicted genes, were taken for functional analysis. The KEGG analysis showed that ABSLW and ABS1 had 122,911 and 160,268, genes assigned to KO terms respectively. Whereas in case of COG functional analysis, 104,334 and 130,191 genes were assigned to different COG classes for ABSLW and ABS1 respectively. Further, on studying the glycoside hydrolases, an abundance of GH13, GH2, GH3, GH43, and GH23 in both the soil and water samples were seen. Conclusion: Our study has provided a comprehensive report about the bacterial diversity and functional capacities of microbes thriving in Samiti Lake. It has also thrown some light on the occurrence of glycoside hydrolases in this region, as they have numerous biotechnological applications in different sectors.

Duchenne muscular dystrophy (DMD) is not currently part of mandatory newborn screening, despite the availability of a test since 1975. In the absence of screening, a DMD diagnosis is often not established in patients until 3–6 years of age. During this time, irreversible muscle degeneration takes place, and clinicians agree that the earlier therapy is initiated, the better the long-term outcome. With recent availability of FDA-approved DMD therapies, interest has renewed for adoption by state public health programs, but such implementation is a multiyear process. To speed access to approved therapies, we implemented a unique, hospital-based program offering parents of newborns an optional, supplemental DMD newborn screen (NBS) via a two-tiered approach: utilizing a creatine kinase (CK) enzyme assay coupled with rapid targeted next-generation sequencing (tNGS) for the DMD gene (using a Whole-Exome Sequencing (WES) assay). The tNGS/WES assay integrates the ability to detect both point mutations and large deletion/duplication events. This tiered newborn screening approach allows for the opportunity to improve treatment and outcomes, avoid the diagnostic delays, and diminish healthcare disparities. To implement this screening algorithm through hospitals in a way that would ultimately be acceptable to public health laboratories, we chose an FDA-approved CK-MM immunoassay to avoid the risks of false-negative/-positive results. Because newborn CK values can be affected due to non-DMD-related causes such as birth trauma, a confirmatory repeat CK assay on a later dried blood spot (DBS) collection has been proposed. Difficulties associated with non-routine repeat DBS collection, including the tracking and recall of families, and the potential creation of parental anxiety associated with false-positive results, can be avoided with this algorithm. Whereas a DMD diagnosis is essentially ruled out by the absence of detected DMD sequence abnormalities, a subsequent CK would still be warranted to confirm resolution of the initial elevation, and thus the absence of non-DMD muscular dystrophy or other pathologies. To date, we have screened over 1500 newborns (uptake rate of ~80%) by a CK-MM assay, and reflexed DMD tNGS in 29 of those babies. We expect the experience from this screening effort will serve as a model that will allow further expansion to other hospital systems until a universal public health screening is established.

The present study is the first of its kind which is focused on Tsomgo lake, a high-altitude lake, located in the Eastern Himalayas of Sikkim. To get a major insight into the bacterial diversity, the shotgun sequencing was carried out in Illumina platform. Our results showed that both the samples TLSS1 (soil) and TLSW1 (water), had Proteobacteria as the most abundant taxa. Cluster of Orthologous group (COG) functional category of TLSS1 has 1,46,965 predicted functions. Cluster of Orthologous Group (COG) functional category of TLSW1 has 1,34,773 predicted functions. Kyoto Encyclopedia of Gene and Genomes (KEGG) functional category of TLSS1 has 1,76,825 predicted functions, most of the sequence fall in metabolism followed by Environmental information processing function. (KEGG) functional category of TLSW1 has 1,62,696 predicted functions and it follows the same pattern as TLSS1. Our studies also provide insight into the presence of distribution of different carbohydrate-active enzymes (CAZymes) present in Tsomgo lake. We have found that in case of both the samples TLSW1 and TLSS1, GlycosylTransferases were active followed by GlycosylHydrolase. The result found, represents for the first time very important findings related to the microbial diversity and the abundance of CAZymes in Tsomgo lake one of the pristine high-altitude lakes in Sikkim.

Krabbe disease (KD) results from galactocerebrosidase (GALC) deficiency. Infantile KD symptoms include irritability, progressive stiffness, developmental delay, and death. The only potential treatment is hematopoietic stem cell transplantation. New York State (NYS) implemented newborn screening for KD in 2006.Dried blood spots from newborns were assayed for GALC enzyme activity using mass spectrometry, followed by molecular analysis for those with low activity (≤12% of the daily mean). Infants with low enzyme activity and one or more mutations were referred for follow-up diagnostic testing and neurological examination.Of >1.9 million screened, 620 infants were subjected to molecular analysis and 348 were referred for diagnostic testing. Five had enzyme activities and mutations consistent with infantile KD and manifested clinical/neurodiagnostic abnormalities. Four underwent transplantation, two are surviving with moderate to severe handicaps, and two died from transplant-related complications. The significance of many sequence variants identified is unknown. Forty-six asymptomatic infants were found to be at moderate to high risk for disease.The positive predictive value of KD screening in NYS is 1.4% (5/346) considering confirmed infantile cases. The incidence of infantile KD in NYS is approximately 1 in 394,000, but it may be higher for later-onset forms.

With rapid advancements in medical treatments, it is critical that screening assays for early detection of rare pediatric genetic disease are piloted in a hospital setting before initiation of state public health newborn screening (SPHNBS). The nomination–initiation of new tests for SPHNBS takes several years. The delay in implementation of state screening may be addressed by hospital screening of newborns for early identification, follow up, and treatment. Duchenne muscular dystrophy (DMD), a rare genetic disease that affects approximately 1 in 3,500 boys, is of increasing interest due to the availability of new therapies; yet, state programs may not adopt DMD screening for several years. Most of the current conditions screened in state laboratories from DBS are collected by nurses in hospital well-baby nurseries and NICUs. Testing can be performed from only one or two extra drops of blood, collected at the same time as samples for state screening. DMD newborn screening in hospitals is an interim step (referred to as supplementary screening) and can identify newborn patients with DMD by biochemical and molecular testing, which allows medical care to be initiated in a timely fashion, before symptom onset. Elevated creatine kinase (CK) enzyme activity establishes suspicion of DMD, and targeted next-generation sequencing (tNGS) is used as a second-tier or confirmatory reflexed test. The combined algorithm, if subsidized, avoids disparities and initiates testing several years ahead of public health state-mandated testing. Several hospitals or clinic sites will be involved in a clinical testing program. Currently, we are considering implementations in California, North Carolina, Massachusetts, and Pennsylvania health networks. We will observe caregiver and parental attitude toward enrollment, testing, and research. We will assess DMD-screening feasibility, follow up, and medical implementation economics. Pilot studies will also include new biochemical and molecular tests, algorithms, improvements, and potential methods for identifying additional muscular dystrophies under research consent. Neonatal nurses are frontline implementers of routine state-mandated and hospital-based testing for newborns, and it is advantageous to increase awareness of innovations in the field for optimized care.

Pompe disease (PD) is screened by a two tier newborn screening (NBS) algorithm, the first tier of which is an enzymatic assay performed on newborn dried blood spots (DBS). As first tier enzymatic screening tests have false positive results, an immediate second tier test on the same sample is critical in resolving newborn health status. Two methodologies have been proposed for second tier testing: (a) measurement of enzymatic activities such as of Creatine/Creatinine over alpha-glucosidase ratio, and (b) DNA sequencing (a molecular genetics approach), such as targeted next generation sequencing. (tNGS). In this review, we discuss the tNGS approach, as well as the challenges in providing second tier screening and follow-up care. While tNGS can predict genotype-phenotype effects when known, these advantages may be diminished when the variants are novel, of unknown significance or not discoverable by current test methodologies. Due to the fact that criticisms of screening algorithms that utilize tNGS are based on perceived complexities, including variant detection and interpretation, we clarify the actual limitations and present the rationale that supports optimizing a molecular genetic testing approach with tNGS. Second tier tNGS can benefit clinical decision-making through the use of the initial NBS DBS punch and rapid turn-around time methodology for tNGS, that includes copy number variant analysis, variant effect prediction, and variant 'cut-off' tools for the reduction of false positive results. The availability of DNA sequence data will contribute to the improved understanding of genotype-phenotype associations and application of treatment. The ultimate goal of second tier testing should enable the earliest possible diagnosis for the earliest initiation of the most effective clinical interventions in infants with PD.

Purpose: Population-based newborn screening (NBS) allows early detection and treatment of inherited disorders. For certain medically-actionable conditions, however, NBS is limited by the absence of reliable biochemical signatures amenable to detection by current platforms. We sought to assess the analytic validity of an ATP7A targeted next generation DNA sequencing assay as a potential newborn screen for one such disorder, Menkes disease. Methods: Dried blood spots from control or Menkes disease subjects (n = 22) were blindly analyzed for pathogenic variants in the copper transport gene, ATP7A. The analytical method was optimized to minimize cost and provide rapid turnaround time. Results: The algorithm correctly identified pathogenic ATP7A variants, including missense, nonsense, small insertions/deletions, and large copy number variants, in 21/22 (95.5%) of subjects, one of whom had inconclusive diagnostic sequencing previously. For one false negative that also had not been detected by commercial molecular laboratories, we identified a deep intronic variant that impaired ATP7A mRNA splicing. Conclusions: Our results support proof-of-concept that primary DNA-based NBS would accurately detect Menkes disease, a disorder that fulfills Wilson and Jungner screening criteria and for which biochemical NBS is unavailable. Targeted next generation sequencing for NBS would enable improved Menkes disease clinical outcomes, establish a platform for early identification of other unscreened disorders, and complement current NBS by providing immediate data for molecular confirmation of numerous biochemically screened conditions.

S.cerevisiae is an industrially important organism known for its ability to produce ethanol as the demand for ethanol is increasing day by day all over the world, the need to find better and alternative ways to increase ethanol production is also rising. In this work we have proposed such alternative but effective method for producing ethanol by S.cerevisiae. Here, we are reporting for the first time the effect of nitrosative stress on ethanol production. Under in vivo condition, nitrosative stress is marked by the modification of macromolecules in the presence of reactive nitrogen species (RNS). Our result showed that treated cells were more capable for ethanol production compared with untreated cells. Our result also showed enhanced alcohol dehydrogenase activity under stressed condition. Further ethanol production was also optimized by using Response Surface Methodology (RSM) with stressed cells. Further, production of ethanol with immobilized beads of stress affected Saccharomyces cerevisiae was also determined. Overall, the obtained data showed that under nitrosative stress, the maximum ethanol production is 34.4 g/l after 24 h and such higher production was observed even after several cycles of fermentation. This is the first report of this kind showing the relation between nitrosative stress and ethanol production in Saccharomyces cerevisiae which may have important industrial application.

Isovalcric acidemia (IVA) is an inborn error of metabolism caused by deficiency of isovaleryl-CoA dehydrogenase. IVA clinical picture includes gastroenterological and progressive neurological symptoms which can lead to permanent disability and death. Early detection by newborn screening (NBS) and treatment promotes normal development. In this study, clinical summaries, biochemical measurements and targeted next generation sequencing (tNGS) data from the IVD gene were compared in 13 Mexican patients. The main symptoms were vomiting, feeding refusal, abdominal pain, impaired alertness, lethargy, stupor, coma; hypotonia, ataxia, hallucinations, seizures; anemia, neutropenia and pancytopenia. Mean blood concentration of isovalerylcarnintine was above the reference value (0.5 mu M) in symptomatic patients (8.78 mu M), as well as in the screen positive newborns (2.23 mu M). The molecular spectrum of this cohort was heterogeneous, with 14 different variants identified, seven were previously-described, and seven were novel. The most frequent variant was c.158G > C (p.R53P). In this study, we found a long diagnostic delay (average of 44 months). Thus, it is essential to increase physician awareness of this treatable condition. Biochemical IVA NBS accompanied by molecular studies (e.g. tNGS) will permit identification of potentially asymptomatic forms of the disease, and improve genotype-phenotype relationship, management decisions and follow-up.