Altered Amino Acid Research Articles

Ferroptosis and aerobic training in ageing.

Ferroptosis is a form of programmed cell death that plays a significant role in causing several diseases such as heart attack and heart failure, through alterations in fat, amino acid, and iron metabolism. Comprehending the regulatory mechanisms of ferroptosis signaling is critical because it has a considerable effect on the elderly's mortality. Conversely, age-related changes in substrate metabolism and metabolite levels are recognized to give rise to obesity. Furthermore, research has proposed that aging and obesity-related changes in substrate metabolism may aggravate ferroptosis. The suppression of ferroptosis holds potential as a successful therapeutic approach for managing different diseases, including sarcopenia, cardiovascular diseases, and central nervous system diseases. However, the pathologic and biological mechanisms behind the function of ferroptosis are not fully comprehended yet. Physical activity could affect lipid, amino acid, and iron metabolism to modulate ferroptosis. The aim of this study is to showcase the current understanding of the molecular mechanisms leading to ferroptosis and discuss the role of aging and physical activity in this phenomenon.

Signals from intestinal microbiota mediate the crosstalk between the lung-gut axis in an influenza infection mouse model.

Introduction: The influenza virus primarily targets the respiratory tract, yet both the respiratory and intestinal systems suffer damage during infection. The connection between lung and intestinal damage remains unclear. Our experiment employs 16S rRNA technology and Liquid Chromatography-Mass Spectrometry (LC-MS) to detect the impact of influenza virus infection on the fecal content and metabolites in mice. Additionally, it investigates the effect of influenza virus infection on intestinal damage and its underlying mechanisms through HE staining, Western blot, Q-PCR, and flow cytometry. Our study found that influenza virus infection caused significant damage to both the lungs and intestines, with the virus detected exclusively in the lungs. Antibiotic treatment worsened the severity of lung and intestinal damage. Moreover, mRNA levels of Toll-like receptor 7 (TLR7) and Interferon-b (IFN-b) significantly increased in the lungs post-infection. Analysis of intestinal microbiota revealed notable shifts in composition after influenza infection, including increased Enterobacteriaceae and decreased Lactobacillaceae. Conversely, antibiotic treatment reduced microbial diversity, notably affecting Firmicutes, Proteobacteria, and Bacteroidetes. Metabolomics showed altered amino acid metabolism pathways due to influenza infection and antibiotics. Abnormal expression of indoleamine 2,3-dioxygenase 1 (IDO1) in the colon disrupted the balance between helper T17 cells (Th17) and regulatory T cells (Treg cells) in the intestine. Mice infected with the influenza virus and supplemented with tryptophan and Lactobacillus showed reduced lung and intestinal damage, decreased Enterobacteriaceae levels in the intestine, and decreased IDO1 activity. Overall, influenza infection caused damage to lung and intestinal tissues, disrupted intestinal microbiota and metabolites, and affected Th17/Treg balance. Antibiotic treatment exacerbated these effects. Supplementation with tryptophan and Lactobacillus improved lung and intestinal health, highlighting a new understanding of the lung-intestine connection in influenza-induced intestinal disease.

Adenocarcinoma of unknown primary with TP53 gene polymorphism: a rare case report with literature review

BackgroundCancer of unknown primary (CUP) is an orphan disease generally presented by undifferentiated and aggressive morphological phenotype. The treatment of CUP is solely dependent upon the origin of cancer. Despite extensive diagnostic testing, in most of the cases the primary site remains unidentifiable.Case presentationThis case demonstrates a 75-year-old male patient, who initially presented with the complaints of swelling over right side of the neck since 2 months. A cervical lymph node biopsy was taken for immunohistochemistry, which revealed cytokeratin (CK) and CK7 markers to be positive. Computerized tomography (CT) of Thorax showed subcentimetric subpleural nodules in bilateral lungs fields, predominantly in lower lobes (metastatic in nature). A subsequent pulmoCORE 12 gene panel test was recommended, and patient was discharged with tablet gefitinib 250mg and capsule containing vitamins plus minerals. After one month, patient revisited with the pulmoCORE 12 gene test report which revealed polymorphism in TP53. A pathogenic variant of tumor protein p53 (TP53), i.e., p.Glu198Ter (amino acid alteration) and c.592G > T (coding) variant, was detected, which has 17.2% variant allele frequency. There are no treatment guidelines for TP53 mutation; therefore, the patient was treated with injection paclitaxel 70mg and carboplatin 100mg for 12 cycles along with palliative radiotherapy of 20 Gy for 5 fractions. The overall prognosis of patient was found to be favorable.ConclusionsThere is a need for development of comprehensive guidelines and new molecularly targeted therapies for treatment of CUP which can be tailored for each patient and achieve precise therapeutic outcome.

PCR-based detection and mutation dynamics of fusion protein gene of orthoaviula viruses sequestered during 2023 field outbreaks in Pakistan

: To isolate and detect a Newcastle disease virus in commercial poultry, Molecular characterization and phylogenetic analysis of the confirmed isolate and Multiple sequence alignment and achievement of accession numbers against our submissions in NCBI bankit.: Genetic and antigenic diversity in the fusion protein gene of New Castle disease virus strains has been recognized and the progressive changes over sequential years indicate that it is a continuously evolving virus. The current vaccines containing conventional vaccinal strains can protect birds to a certain level but do not prevent infection and virus shedding. : The partial fusion protein gene of the 14 NDV isolates during the 2023 outbreaks from different areas of Pakistan was determined and analyzed. The antigenic protein translational segment of the fusion gene nucleotide fragment was targeted with a specifically designed primer executed 202 bp size of predictable amplicon during PCR amplification. The nucleotide sequence analysis of studied isolates showed closed similarity to the NCBI bankit numbers. Phylogenetic analysis revealed that 3 isolates belong to genotype II while, 2 isolates positions near genotype VIII of class II. The 6 isolates were located near genotype XVII and only 1 was presented on genotype V branch in calss II. Mutation analysis results revealed various mutations at nucleotide intervals and even found altered amino acids during translation. The results revealed that nucleotide mutation at various positions attributes amino acid substitution that enables wild prevailing strains to evade artificial active immunity. In such a scenario Chimeric and genotype match vaccines prepared from indigenous isolates may be useful in developing candidate vaccines to prevent virus shedding and infection. Further studies are suggested at molecular level to determine the consensus amino acid sequence for virulent, mesogenic, and avirulent prevailing NDV strains.

Rational Design of a Flavoenzyme for Aerobic Nicotine Catabolism.

Enzymatic therapy with nicotine-degrading enzyme is a new strategy in treating nicotine addiction, which can reduce nicotine concentrations and weaken withdrawal in the rat model. However, when O2 is used as the electron acceptor, no satisfactory performance has been achieved with one of the most commonly studied and efficient nicotine-catabolizing enzymes, NicA2. To obtain more efficient nicotine-degrading enzyme, we rationally designed and engineered a flavoenzyme Pnao, which shares high structural similarity with NicA2 (RMSD = 1.143 Å) and efficiently catalyze pseudooxynicotine into 3-succinoyl-semialdehyde pyridine using O2. Through amino acid alterations with NicA2, five Pnao mutants were generated, which can degrade nicotine in Tris-HCl buffer and retained catabolic activity on its natural substrate. Nicotine-1'-N-oxide was identified as one of the reaction products. Four of the derivative mutants showed activity in rat serum and Trp220 and Asn224 were found critical for enzyme specificity. Our findings offer a novel avenue for research into aerobic nicotine catabolism and provides a promising method of generating additional nicotine-catalytic enzymes.

Genome-wide analysis of histone modifications can contribute to the identification of candidate cis-regulatory regions in the threespine stickleback fish

BackgroundCis-regulatory mutations often underlie phenotypic evolution. However, because identifying the locations of promoters and enhancers in non-coding regions is challenging, we have fewer examples of identified causative cis-regulatory mutations that underlie naturally occurring phenotypic variations than of causative amino acid-altering mutations. Because cis-regulatory elements have epigenetic marks of specific histone modifications, we can detect cis-regulatory elements by mapping and analyzing them. Here, we investigated histone modifications and chromatin accessibility with cleavage under targets and tagmentation (CUT&Tag) and assay for transposase-accessible chromatin-sequencing (ATAC-seq).ResultsUsing the threespine stickleback (Gasterosteus aculeatus) as a model, we confirmed that the genes for which nearby regions showed active marks, such as H3K4me1, H3K4me3, and high chromatin accessibility, were highly expressed. In contrast, the expression levels of genes for which nearby regions showed repressive marks, such as H3K27me3, were reduced, suggesting that our chromatin analysis protocols overall worked well. Genomic regions with peaks of histone modifications showed higher nucleotide diversity within and between populations. By comparing gene expression in the gills of the marine and stream ecotypes, we identified several insertions and deletions (indels) with transposable element fragments in the candidate cis-regulatory regions.ConclusionsThus, mapping and analyzing histone modifications can help identify cis-regulatory elements and accelerate the identification of causative mutations in the non-coding regions underlying naturally occurring phenotypic variations.

Impact of Maternal Pre-Pregnancy Underweight on Cord Blood Metabolome: An Analysis of the Population-Based Survey of Neonates in Pomerania (SNiP).

Intrauterine growth restriction leads to an altered lipid and amino acid profile in the cord blood at the end of pregnancy. Pre-pregnancy underweight is an early risk factor for impaired fetal growth. The aim of this study was to investigate whether a pre-pregnancy body mass index (ppBMI) of <18.5 kg/m2, as early as at the beginning of pregnancy, is associated with changes in the umbilical cord metabolome. In a sample of the Survey of Neonates in Pomerania (SNIP) birth cohort, the cord blood metabolome of n = 240 newborns of mothers with a ppBMI of <18.5 kg/m2 with n = 208 controls (ppBMI of 18.5-24.9 kg/m2) was measured by NMR spectrometry. A maternal ppBMI of <18.5 kg/m2 was associated with increased concentrations of HDL4 cholesterol, HDL4 phospholipids, VLDL5 cholesterol, HDL 2, and HDL4 Apo-A1, as well as decreased VLDL triglycerides and HDL2 free cholesterol. A ppBMI of <18.5 kg/m2 combined with poor intrauterine growth (a gestational weight gain (GWG) < 25th percentile) was associated with decreased concentrations of total cholesterol; cholesterol transporting lipoproteins (LDL4, LDL6, LDL free cholesterol, and HDL2 free cholesterol); LDL4 Apo-B; total Apo-A2; and HDL3 Apo-A2. In conclusion, maternal underweight at the beginning of pregnancy already results in metabolic changes in the lipid profile in the cord blood, but the pattern changes when poor GWG is followed by pre-pregnancy underweight.

Characterization of Inflammatory Mediators and Metabolome in Interstitial Fluid Collected with Dermal Open Flow Microperfusion before and at the End of Dupilumab Treatment in Atopic Dermatitis.

Dupilumab is a monoclonal antibody approved for the treatment of atopic dermatitis (AD); however, its effects on molecular, cellular, and immunological levels remain to be elucidated. In this study, blood and dermal interstitial fluid (ISF) from nonlesional (NL) and lesional (L) skin were collected from eight patients with moderate to severe AD, before (visit 2-v2) and at the end of a 16-week treatment with dupilumab (visit 10-v10). Clinical treatment effect was demonstrated by significantly decreased AD severity scores at the end of treatment. At v10 versus v2, the percentages of CD4+ interleukin-producing cells showed a decreasing trend in ISF L and NL, unbound IL-4 levels in plasma were increased, IL-5 levels in ISF L reduced, and levels of factors involved in anti-inflammatory pathways and re-epithelization increased. At v2, ISF L showed that AD lesions might have altered amino acid pathways and lipid signaling compared to ISF NL. At v10, ISF L exhibited raised levels of long- and very-long-chain fatty acids and lipids compared to v2. Furthermore, dupilumab administration caused reduced expression of miR-155-5p and miR-378a-3p in ISF L. In conclusion, results from the present study provided novel knowledge by linking local immune and metabolic alterations to AD pathogenesis and treatment response.

Discovery of urinary biosignatures for tuberculosis and nontuberculous mycobacteria classification using metabolomics and machine learning

Nontuberculous mycobacteria (NTM) infection diagnosis remains a challenge due to its overlapping clinical symptoms with tuberculosis (TB), leading to inappropriate treatment. Herein, we employed noninvasive metabolic phenotyping coupled with comprehensive statistical modeling to discover potential biomarkers for the differential diagnosis of NTM infection versus TB. Urine samples from 19 NTM and 35 TB patients were collected, and untargeted metabolomics was performed using rapid liquid chromatography-mass spectrometry. The urine metabolome was analyzed using a combination of univariate and multivariate statistical approaches, incorporating machine learning. Univariate analysis revealed significant alterations in amino acids, especially tryptophan metabolism, in NTM infection compared to TB. Specifically, NTM infection was associated with upregulated levels of methionine but downregulated levels of glutarate, valine, 3-hydroxyanthranilate, and tryptophan. Five machine learning models were used to classify NTM and TB. Notably, the random forest model demonstrated excellent performance [area under the receiver operating characteristic (ROC) curve greater than 0.8] in distinguishing NTM from TB. Six potential biomarkers for NTM infection diagnosis, including methionine, valine, glutarate, 3-hydroxyanthranilate, corticosterone, and indole-3-carboxyaldehyde, were revealed from univariate ROC analysis and machine learning models. Altogether, our study suggested new noninvasive biomarkers and laid a foundation for applying machine learning to NTM differential diagnosis.

Effects of High Dietary Starch Levels on the Growth Performance, Liver Function, and Metabolome of Largemouth Bass (Micropterus salmoides)

In this study, we conducted a 16-week feeding trial to investigate the effects of a high-cassava starch diet on growth performance, liver function, and metabolism in largemouth bass (Micropterus salmoides). We formulated five diets containing varying levels of cassava starch: 12%, 9%, 6%, 3%, and 0% (termed M12, M9, M6, M3, and M0, respectively). We distributed these diets among largemouth bass with the initial body weight of 83.33 ± 0.55 g via an in-pond “raceway” aquaculture system. Our findings suggest that high level (12%) of cassava starch dietary inclusion adversely affected growth performance metrics such as weight gain rate and specific growth rate, along with feed utilization efficiency indicators, including protein efficiency, protein deposition rate, and the apparent digestibility of dry matter and protein. This negative impact was accompanied by a decrease in intestinal amylase activity. Through further transcriptomic analysis, we identified several key genes associated with carbohydrate metabolism, which underwent changes influencing liver function. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed the involvement of these differentially expressed genes (DEGs) in the tricarboxylic acid cycle (TCA cycle). Comparative metabolomics analysis further indicated that the M9 group showed significant enrichment in pathways related to amino acid metabolism and alterations in the levels of metabolites involved in carbohydrate metabolism. In conclusion, our study demonstrates that incorporating up to 9% cassava starch in the diet can enhance growth performance in largemouth bass by stimulating digestive enzyme activities and promoting glucose utilization.

Unveiling the impact of glutathione (GSH) and p53 gene deletion on tumor cell metabolism by amino acid and proteomics analysis.

Tumor cell inhibition is a pivotal focus in anti-cancer research, and extensive investigations have been conducted regarding the role of p53. Numerous studies have highlighted its close association with reactive oxygen species (ROS). However, the precise impact of the antioxidant glutathione (GSH) in this context remains inadequately elucidated. Here, we will elucidate the anti-cancer mechanisms mediated by p53 following treatment with GSH. In this study, we employed a p53 gene knockout approach in SW480 colorectal cells and conducted comprehensive analyses of 20 amino acids and proteomics using liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). These analyses unveiled profound alterations in amino acids and proteins triggered by GSH treatment, shedding light on novel phenomena and delineating the intricate interplay between GSH and cellular proteins. The deletion of the p53 gene exerts a profound influence on tumor cell proliferation. Moreover, tumor cell proliferation is significantly affected by elevated GSH levels. Importantly, in the absence of the p53 gene, cells exhibit heightened sensitivity to GSH, leading to inhibited cell growth. The combined therapeutic approach involving GSH and p53 gene deletion expedites the demise of tumor cells. It is noteworthy that this treatment leads to a marked decline in amino acid metabolism, particularly affecting the down-regulation of methionine (Met) and phenylalanine (Phe) amino acids. Among the 41 proteins displaying significant changes, 8 exhibit consistent alterations, with 5 experiencing decreased levels and 3 demonstrating increased quantities. These proteins primarily participate in crucial cellular metabolic processes and immune functions. In conclusion, the concurrent administration of GSH treatment and p53 gene deletion triggers substantial modifications in the amino acid and protein metabolism of tumor cells, primarily characterized by down-regulation. This, in turn, compromises cell metabolic activity and immune function, ultimately culminating in the demise of tumor cells. These newfound insights hold promising implications and could pave the way for the development of straightforward and efficacious anti-cancer treatments.

Amino acid metabolic reprogramming in the tumor microenvironment and its implication for cancer therapy.

Amino acids are essential building blocks for proteins, crucial energy sources for cell survival, and key signaling molecules supporting the resistant growth of tumor cells. In tumor cells, amino acid metabolic reprogramming is characterized by the enhanced uptake of amino acids as well as their aberrant synthesis, breakdown, and transport, leading to immune evasion and malignant progression of tumor cells. This article reviews the altered amino acid metabolism in tumor cells and its impact on tumor microenvironment, and also provides an overview of the current clinical applications of amino acid metabolism. Innovative drugs targeting amino acid metabolism hold great promise for precision and personalized cancer therapy.

PHACTboost: A Phylogeny-Aware Pathogenicity Predictor for Missense Mutations via Boosting.

Most algorithms that are used to predict the effects of variants rely on evolutionary conservation. However, a majority of such techniques compute evolutionary conservation by solely using the alignment of multiple sequences while overlooking the evolutionary context of substitution events. We had introduced PHACT, a scoring-based pathogenicity predictor for missense mutations that can leverage phylogenetic trees, in our previous study. By building on this foundation, we now propose PHACTboost, a gradient boosting tree-based classifier that combines PHACT scores with information from multiple sequence alignments, phylogenetic trees, and ancestral reconstruction. By learning from data, PHACTboost outperforms PHACT. Furthermore, the results of comprehensive experiments on carefully constructed sets of variants demonstrated that PHACTboost can outperform 40 prevalent pathogenicity predictors reported in the dbNSFP, including conventional tools, metapredictors, and deep learning-based approaches as well as more recent tools such as AlphaMissense, EVE, and CPT-1. The superiority of PHACTboost over these methods was particularly evident in case of hard variants for which different pathogenicity predictors offered conflicting results. We provide predictions of 215 million amino acid alterations over 20,191 proteins. PHACTboost is available at https://github.com/CompGenomeLab/PHACTboost. PHACTboost can improve our understanding of genetic diseases and facilitate more accurate diagnoses.

Non-synonymous Single Nucleotide Polymorphism on TLR1A gene as a candidate for immune function of Indonesian indigenous chicken

Abstract. Indrijani H, Hilmia N, Anang A, Pangestu MSP. 2024. Non-synonymous Single Nucleotide Polymorphism on TLR1A gene as a candidate for immune function of Indonesian indigenous chicken. Biodiversitas 25: 2434-2441. TLR1A is a candidate gene related to the immune system in chickens, which contributes to disease resilience, which could impact their productivity. This study aimed to determine mutations in the TLR1A gene in three indigenous Indonesian chickens namely, Pelung, Black Kedu and Sentul. A total of 127 DNA samples were used and consisted of 38, 41 and 48 respectively for Pelung, Black Kedu and Sentul chickens. DNA was isolated from white blood cells and amplification of TLR1A gene was conducted in part of exon 4 using PCR with specific primer. The sequencing results for the TLR1A gene were analyzed to identify mutations using the BioEdit and MEGA X programs. The results showed genetic polymorphisms in the TLR1A gene in Indonesian indigenous chicken. One synonymous mutation, g.710C&gt;T, in which amino acid coded leusine at 207th, and four non-synonymous mutation, i.e., g.822A&gt;G, altered amino acids coding from threonine to alanine (T245A), g.835C&gt;T, which altered amino acids coding 249th from threonine to methionine (T249M), g.1015G&gt;A that altered amino acids coding 309th from serine to aspargine (S309N) and g.1165T&gt;G that changed amino acid from arginine to proline R359P. All mutation frequencies for the alleles are lower than those for the non-mutation alleles. The non-synonymous mutation of the TLR1A gene might alter the metabolism of this hormone related to the immune system in chickens.

Revealing the complete mtDNA genome sequence of Cemani chicken (Gallus gallus) by using Nanopore sequencing analysis.

This study aimed to identify, discover and explore the characteristics of the mtDNA genomes of Cemani chicken (Gallus gallus). This study used gDNA of Cemani chicken isolated from liver tissue. mtDNA sequencing was performed using WGS mtDNA analysis with nanopore technology by Oxford Nanopore Technologies GridION. Bioinformatics and data analysis were then performed. This study showed that the length of the mtDNA genome is 16,789 bp, consisting of two ribosomal RNA (12S rRNA, 16S rRNA), 22 transfer RNA genes (trnR, trnG, trnK, trnD, trnS, trnY, trnC, trnN, trnA, trnW, trnM, trnQ, trnl, trnL, trnV, trnF, trnP, trnT, trnE, trnL, trnS, trnH), 13 protein-coding genes (PCGs) (ND4l, ND3, COX3, ATP6, ATP8, COX2, COX1, ND2, ND1, CYTB, ND6, ND5, ND4), and a noncoding control region (Dloop). Furthermore, analysis showed there were polymorphic sites and amino acid alterations when mtDNA Cemani chicken was aligned with references from GenBank. Site (988T>*) in Dloop genes and (328A>G) in ND3 genes which alter glycine to stop codon, were specific markers found only in Cemani chicken.

Modulation of mercaptopurine intestinal toxicity and pharmacokinetics by gut microbiota

The interaction between the gut microbiota and mercaptopurine (6-MP), a crucial drug used in pediatric acute lymphoblastic leukemia (ALL) treatment, has not been extensively studied. Here we reveal the significant perturbation of gut microbiota after 2-week 6-MP treatment in beagles and mice followed by the functional prediction that showed impairment of SCFAs production and altered amino acid synthesis. And the targeted metabolomics in plasma also showed changes in amino acids. Additionally, targeted metabolomics analysis of feces showed changes in amino acids and SCFAs. Furthermore, ablating the intestinal microbiota by broad-spectrum antibiotics exacerbated the imbalance of amino acids, particularly leading to a significant decrease in the concentration of S-adenosylmethionine (SAM). Importantly, the depletion of gut microbiota worsened the damage of small intestine caused by 6-MP, resulting in increased intestinal permeability. Considering the relationship between toxicity and 6-MP metabolites, we conducted a pharmacokinetic study in pseudo germ-free rats to confirm that gut microbiota depletion altered the methylation metabolites of 6-MP. Specifically, the concentration of MeTINs, a secondary methylation metabolite, showed a negative correlation with SAM, the pivotal methyl donor. Additionally, we observed a strong correlation between Alistipes and SAM levels in both feces and plasma. In conclusion, our study demonstrates that 6-MP disrupts the gut microbiota, and depleting the gut microbiota exacerbates 6-MP-induced intestinal toxicity. Moreover, SAM derived from microbiota plays a crucial role in influencing plasma SAM and the methylation of 6-MP. These findings underscore the importance of comprehending the role of the gut microbiota in 6-MP metabolism and toxicity.

Mucosal organs exhibit distinct response signatures to hydrogen sulphide in Atlantic salmon (Salmo salar)

Hydrogen sulphide (H2S) is considered an immunotoxicant, and its presence in the water can influence the mucosal barrier functions of fish. However, there is a significant knowledge gap on how fish mucosa responds to low environmental H2S levels. The present study investigated the consequences of prolonged exposure to sub-lethal levels of H2S on the mucosal defences of Atlantic salmon (Salmo salar). Fish were continuously exposed to two levels of H2S (low: 0.05 µM; and high: 0.12 µM) for 12 days. Unexposed fish served as control. Molecular and histological profiling focused on the changes in the skin, gills and olfactory rosette. In addition, metabolomics and proteomics were performed on the skin and gill mucus. The gene expression profile indicated that the gills and olfactory rosette were more sensitive to H2S than the skin. The olfactory rosette showed a dose-dependent response, but not the gills. Genes related to stress responses were triggered at mucosal sites by H2S. Moreover, H2S elicited strong inflammatory responses, particularly in the gills. All mucosal organs demonstrated the key molecular repertoire for sulphide detoxification, but their temporal and spatial expression was not substantially affected by sub-lethal H2S levels. Mucosal barrier integrity was not considerably affected by H2S. Mucus metabolomes of the skin and gills were unaffected, but a matrix-dependent response was identified. Comparing the high-concentration group's skin and gills mucus metabolomes identified altered amino acid biosynthesis and metabolism pathways. The skin and gill mucus exhibited distinct proteomic profiles. Enrichment analysis revealed that proteins related to immunity and metabolism were affected in both mucus matrices. The present study expands our knowledge of the defence mechanisms against H2S at mucosal sites in Atlantic salmon. The findings offer insights into the health and welfare consequences of sub-lethal H2S, which can be incorporated into the risk assessment protocols in salmon land-based farms.

Lipid extraction alters amino acid composition and bulk, but not amino acid, carbon and nitrogen isotope values.

Concerns exist over observed shifts in value and variance of nitrogen isotopes following physicochemical extraction of lipids from organic matter. The mechanisms behind these apparent changes in bulk tissue δ15N values are not fully understood yet have major implications for analytical costs and integrity of data interpretations. Changes in proximate analysis, amino acid composition, C:N ratios, bulk tissue and amino acid δ13C and δ15N values, and resulting isotope-based food web metrics were compared between lipid-intact and lipid-extracted muscle tissue of fishes spanning <1% to >20% muscle fat content to identify mechanisms of nitrogen isotope fractionation associated with physicochemical lipid extraction. Bulk δ13C and δ15N values increased and %N, C:N ratios and crude protein content decreased following lipid extraction. Resulting bulk isotope niche spacing and overlap varied significantly between lipid-intact and lipid-extracted tissues. While amino acid composition significantly changed during lipid extraction, particularly for lipid-associated amino acids (e.g., Glu, Lys, Ser), individual amino acid δ13C and δ15N values, and their associated compound-specific isotope analysis of amino acids (CSIA-AA)-based food web metrics, did not. Physicochemical lipid extraction caused significant tissue composition changes (e.g., leaching of amino acids and 15N-deplete nitrogenous waste) that affected δ13C and δ15N values and tissue %C and %N beyond simply removing lipids. However, lipid extraction did not alter individual amino acid δ13C or δ15N values or their associated CSIA-AA-based food web metrics.

Effects of A1 Milk, A2 Milk and the Opioid-like Peptide β-Casomorphin-7 on the Proliferation of Human Peripheral Blood Mononuclear Cells.

Special attention is given to cow's milk and its variants, with ongoing discussions about health-related impacts primarily focusing on the A1 variant in contrast to the A2 variant. The difference between these variants lies in a single amino acid alteration at position 67 of β-casein. This alteration is presumed to make the A1 variant more susceptible to enzymatic breakdown during milk digestion, leading to an increased release of the peptide β-casomorphin-7 (BCM-7). BCM-7 is hypothesized to interact with µ-opioid receptors on immune cells in humans. Although BCM-7 has demonstrated both immunosuppressive and inflammatory effects, its direct impact on the immune system remains unclear. Thus, we examined the influence of A1 and A2 milk on Concanavalin A (ConA)-stimulated human peripheral blood mononuclear cells (PBMCs), as well as the effect of experimentally digested A1 and A2 milk, containing different amounts of free BCM-7 from β-casein cleavage. Additionally, we evaluated the effects of pure BCM-7 on the proliferation of ConA-stimulated PBMCs and purified CD4+ T cells. Milk fundamentally inhibited PBMC proliferation, independent of the β-casein variant. In contrast, experimentally digested milk of both variants and pure BCM-7 showed no influence on the proliferation of PBMCs or isolated CD4+ T cells. Our results indicate that milk exerts an anti-inflammatory effect on PBMCs, regardless of the A1 or A2 β-casein variant, which is nullified after in vitro digestion. Consequently, we deem BCM-7 unsuitable as a biomarker for food-induced inflammation.

A Coiled-Coil Nucleotide-Binding Domain Leucine-Rich Repeat Receptor Gene MeRPPL1 Plays a Role in the Replication of a Geminivirus in Cassava.

Disease resistance gene (R gene)-encoded nucleotide-binding leucine-rich repeat proteins (NLRs) are critical players in plant host defence mechanisms because of their role as receptors that recognise pathogen effectors and trigger plant effector-triggered immunity (ETI). This study aimed to determine the putative role of a cassava coiled-coil (CC)-NLR (CNL) gene MeRPPL1 (Manes.12G091600) (single allele) located on chromosome 12 in the tolerance or susceptibility to South African cassava mosaic virus (SACMV), one of the causal agents of cassava mosaic disease (CMD). A transient protoplast system was used to knock down the expression of MeRPPL1 by clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9 (CRISPR-Cas9). The MeRPPL1-targeting CRISPR vectors and/or SACMV DNA A and DNA B infectious clones were used to transfect protoplasts isolated from leaf mesophyll cells from the SACMV-tolerant cassava (Manihot esculenta) cultivar TME3. The CRISPR/Cas9 silencing vector significantly reduced MeRPPL1 expression in protoplasts whether with or without SACMV co-infection. Notably, SACMV DNA A replication was higher in protoplasts with lower MeRPPL1 expression levels than in non-silenced protoplasts. Mutagenesis studies revealed that protoplast co-transfection with CRISPR-MeRPPL1 silencing vector + SACMV and transfection with only SACMV induced nucleotide substitution mutations that led to altered amino acids in the highly conserved MHD motif of the MeRPPL1-translated polypeptide. This may abolish or alter the regulatory role of the MHD motif in controlling R protein activity and could contribute to the increase in SACMV-DNA A accumulation observed in MeRPPL1-silenced protoplasts. The results herein demonstrate for the first time a role for a CNL gene in tolerance to a geminivirus in TME3.