Folate Production Research Articles

Foxtail millet [Setaria italica (L.) P. Beauv.] grown under nitrogen deficiency exhibits a lower folate contents.

Foxtail millet [Setaria italica (L.) P. Beauv.], as a rich source of folates, has been cultivated on arid infertile lands, for which N deficiency is one of the major issues. Growing environments might have a significant influence on cereal folate levels. However, little is known whether N deficiency modulates cereal folate levels. In order to obtain enriched folate foxtail millet production in nutrient-poor soil, we conducted a study investigating the content of folate derivatives of 29 diverse foxtail millet cultivars under two N regimes (0 and 150 kg N ha-1) for 2 years to explore folate potential grown under low N. The contents of total folate and most derivatives were reduced by N deficiency. The effect on total folate content caused by N was stronger than cultivar genotype did. Folate content of enriched folate cultivars was prone to be reduced by N deficiency. Structural equation models (SEMs) revealed that N fertilization had a positive indirect effect on grain folate content through influencing plant N and K accumulation. Collectively, the results indicate much more attention should be paid to N management when foxtail millet is cultivated in infertile soil, to improve foxtail millet folate contents.

Correlation of gene content in selected bifidobacteria with folate supplier or scavenger status during growth in laboratory media

Five different bifidobacterial cultures were screened to investigate if their de novo folate production ability could be predicated based on their folate biosynthetic gene content. As previously developed folate free media could not adequately support the growth of these five strains, even with added folate, an alternative medium (YNB+) was developed. Consistent with its genome prediction, Bifidobacterium animalis subsp. lactis strain Bb-12 required folate for its growth even in the presence of p-aminobenzoic acid (PABA). Although B. longum DJO10A and B. breve ATCC 15701 lack an alkaline phosphatase gene for the formation of one pathway intermediate (DHPPP), both were found to be able to grow independent from folate and PABA suggesting that lack of these genes in genome annotations cannot be used to predict folate dependency in bifidobacteria. Similarly, having all required genes cannot always predict folate production, as B. longum subsp. infantis RECb4 contained a full set of pathway genes, but required folate or PABA for growth. While many bifidobacteria can produce folate, this also cannot be used to predicate their ability to supply folate as some can act as scavengers when folate is readily available in the environment, such as B. breve ATCC 15701 in this study. As bifidobacterial cultures are sometimes considered for use in foods based on their folate production abilities, it is important to consider whether they function as folate scavengers or suppliers when growing in folate containing media.

Scheffersomyces stipitis ability to valorize different residual biomasses for vitamin B9 production.

Sugar beet pulp (SBP), sugar beet molasses (SBM) and unfermented grape marcs (UGM) represent important waste in the agro-food sector. If suitably pre-treated, hexose and pentose sugars can be released in high quantities and can subsequently be used by appropriate cell factories as growth media and for the production of (complex) biomolecules, accomplishing the growing demand for products obtained from sustainable resources. One example is vitamin B9 or folate, a B-complex vitamin currently produced by chemical synthesis, almost exclusively in the oxidized form of folic acid (FA). It is therefore desirable to develop novel competitive strategies for replacing its current fossil-based production with a sustainable bio-based process. In this study, we assessed the production of natural folate by the yeast Scheffersomyces stipitis, investigating SBM, SBP and UGM as potential growth media. Pre-treatment of SBM and SBP had previously been optimized in our laboratory; thus, here we focused only on UGM pre-treatment and hydrolysis strategies for the release of fermentable sugars. Then, we optimized the growth of S. stipitis on the three media formulated from those biomasses, working on inoculum pre-adaptation, oxygen availability and supplementation of necessary nutrients to support the microorganism. Folate production, measured with a microbiological assay, reached 188.2 ± 24.86 μg/L on SBM, 130.6 ± 1.34 μg/L on SBP and 101.9 ± 6.62 μg/L on UGM. Here, we demonstrate the flexibility of S.stipitis in utilizing different residual biomasses as growth media. Moreover, we assessed the production of folate from waste, and to the best of our knowledge, we obtained the highest production of folate from residual biomasses ever reported, providing the first indications for the future development of this microbial production process.

Phenotypic characterization and genome analysis reveal the probiotic potential of a banyan endophyte Bacillus velezensis K1.

The current study aimed to scrutinize the probiotic traits and safety aspects of Bacillus velezensis K1 through experimental and supporting genome studies. The seven cultures previously isolated from the aerial roots of Ficus benghalensis were initially screened for their antibacterial activity as well as acid and bile tolerance. The isolate K1 was found to be the most potent and was further investigated for probiotic traits and safety. K1 showed tolerance to simulated digestive juices and 0.3% bile. It showed notable aggregation, cell surface hydrophobicity, and adherence to HT-29 cells. K1 significantly prevented the adhesion of E. coli O157: H7 and S. enterica ATCC 13076 to HT-29 in cell culture assays. K1 could hydrolyze phytate and complex polysaccharides. The genes related to stress tolerance, adhesion, antimicrobial activity, and production of vitamins, viz. thiamine, riboflavin, pyridoxine, pantothenic acid, folate, and biotin were annotated in the K1 genome. K1 was found to be non-hemolytic, noncytotoxic, as well as susceptible to antibiotics. No virulence or toxin-encoding genes were identified in its genome. B. velezensis K1 is a prospective probiotic with the ability to tolerate gastrointestinal stress, adhere to intestinal surfaces, and inhibit enteropathogens.

Lactobacillus spp. isolated from prebiotic-derived raw goat milk: probiotic characteristics, cholesterol assimilation and folate production.

This work provides to evaluate cholesterol assimilation and folic acid production by determining the probiotic properties of Lactobacillus spp. from raw goat milk with prebiotic properties. We isolated Lactobacilli from goat milk and identified API 50, CHL, and 16sRNA. Probiotic properties were determined according to bile salt and acidic tolerance, hydrophobicity, hemolytic activity, antibiotic sensitivity, antagonistic effect, and exopolysaccharide production. In addition, the cholesterol assimilation and folate production of cultures were determined. L. plantarum GM-12 and L. plantarum GM-15 showed the highest folate production and the highest cholesterol assimilation.These two strains are strong candidates for use as potential probiotics and starter cultures.

First report on Vitamin B9 production including quantitative analysis of its vitamers in the yeast Scheffersomyces stipitis

BackgroundThe demand for naturally derived products is continuously growing. Nutraceuticals such as pre- and post-biotics, antioxidants and vitamins are prominent examples in this scenario, but many of them are mainly produced by chemical synthesis. The global folate market is expected to register a CAGR of 5.3% from 2019 to 2024 and reach USD 1.02 billion by the end of 2024. Vitamin B9, commonly known as folate, is an essential micronutrient for humans. Acting as a cofactor in one-carbon transfer reactions, it is involved in many biochemical pathways, among which the synthesis of nucleotides and amino acids. In addition to plants, many microorganisms can naturally produce it, and this can pave the way for establishing production processes. In this work, we explored the use of Scheffersomyces stipitis for the production of natural vitamin B9 by microbial fermentation as a sustainable alternative to chemical synthesis.ResultsGlucose and xylose are the main sugars released during the pretreatment and hydrolysis processes of several residual lignocellulosic biomasses (such as corn stover, wheat straw or bagasse). We optimized the growth conditions in minimal medium formulated with these sugars and investigated the key role of oxygenation and nitrogen source on folate production. Vitamin B9 production was first assessed in shake flasks and then in bioreactor, obtaining a folate production up to 3.7 ± 0.07 mg/L, which to date is the highest found in literature when considering wild type microorganisms. Moreover, the production of folate was almost entirely shifted toward reduced vitamers, which are those metabolically active for humans.ConclusionsFor the first time, the non-Saccharomyces yeast S. stipitis was used to produce folate. The results confirm its potential as a microbial cell factory for folate production, which can be also improved both by genetic engineering strategies and by fine-tuning the fermentation conditions and nutrient requirements.

Modelling of the nutritional behaviour of cowpea seeds during soaking, germination and cooking process

Two modelling approaches previously developed and describing separately the diffusion–reaction of folates and alpha-galactosides during cowpea seed soaking, germination and cooking processes were combined here to simulate the effect of some key processing parameters on the nutritional value of cowpea seeds. The simulator was upgraded, considering thermal-pH dependency of both diffusivities and reactivities of folates and alpha-galactosides together with water-to-seed ratio and starch gelatinization. The simulations showed that soaking and cooking processes were deleterious for folates whereas germination promoted the production of folates while reducing alpha-galactosides concentration. This study suggests that a long germination (96 h), followed by a short cooking seems to be optimal both in terms of nutritional value and degree of cooking.

Potential for increasing folate contents of traditional African fermented sorghum gruel (Motoho) using presumptive probiotic lactic acid bacteria

Folate deficiency is commonly observed in most developing countries. Lactic acid bacteria (LAB) fermentation of cereal-based foods could be an alternative to improving folate intake. This study evaluated the possibility of improving the folate contents of traditional African fermented sorghum gruel (motoho) using indigenous potential probiotic LAB. A total of 220 LAB strains isolated from maize gruel were screened for extracellular and intracellular folate production. Strains were further examined for in vitro probiotic characteristics and antimicrobial activity. Sixteen (16) LAB strains exhibited high production of total folate which ranged between 44 and 180 µg/100 mL with the lowest and highest value in L. plantarum S8 and L. plantarum S49, respectively. Fermentation of sorghum gruel with folate producing probiotic LAB strains for in situ folate production in motoho was determined. The folate contents of sorghum motoho fermented with LAB ranged between 13 and 20 µg/100 mL while the control was below 2.5 µg/100 mL. Indigenous LAB strains from traditional African fermented gruel possess desirable in vitro probiotic properties and ability to produce folates, thus, could be used for natural in situ folate fortification. This study presents a strategy for future application of indigenous probiotic LAB cultures as natural means of fortifying cereal foods with folates.

Characterization of Two Highly Arsenic-Resistant Caulobacteraceae Strains of Brevundimonas nasdae: Discovery of a New Arsenic Resistance Determinant.

Arsenic (As), distributed widely in the natural environment, is a toxic substance which can severely impair the normal functions in living cells. Research on the genetic determinants conferring functions in arsenic resistance and metabolism is of great importance for remediating arsenic-contaminated environments. Many organisms, including bacteria, have developed various strategies to tolerate arsenic, by either detoxifying this harmful element or utilizing it for energy generation. More and more new arsenic resistance (ars) determinants have been identified to be conferring resistance to diverse arsenic compounds and encoded in ars operons. There is a hazard in mobilizing arsenic during gold-mining activities due to gold- and arsenic-bearing minerals coexisting. In this study, we isolated 8 gold enrichment strains from the Zijin gold and copper mine (Longyan, Fujian Province, China) wastewater treatment site soil, at an altitude of 192 m. We identified two Brevundimonas nasdae strains, Au-Bre29 and Au-Bre30, among these eight strains, having a high minimum inhibitory concentration (MIC) for As(III). These two strains contained the same ars operons but displayed differences regarding secretion of extra-polymeric substances (EPS) upon arsenite (As(III)) stress. B. nasdae Au-Bre29 contained one extra plasmid but without harboring any additional ars genes compared to B. nasdae Au-Bre30. We optimized the growth conditions for strains Au-Bre29 and Au-Bre30. Au-Bre30 was able to tolerate both a lower pH and slightly higher concentrations of NaCl. We also identified folE, a folate synthesis gene, in the ars operon of these two strains. In most organisms, folate synthesis begins with a FolE (GTP-Cyclohydrolase I)-type enzyme, and the corresponding gene is typically designated folE (in bacteria) or gch1 (in mammals). Heterologous expression of folE, cloned from B. nasdae Au-Bre30, in the arsenic-hypersensitive strain Escherichia coli AW3110, conferred resistance to As(III), arsenate (As(V)), trivalent roxarsone (Rox(III)), pentavalent roxarsone (Rox(V)), trivalent antimonite (Sb(III)), and pentavalent antimonate (Sb(V)), indicating that folate biosynthesis is a target of arsenite toxicity and increased production of folate confers increased resistance to oxyanions. Genes encoding Acr3 and ArsH were shown to confer resistance to As(III), Rox(III), Sb(III), and Sb(V), and ArsH also conferred resistance to As(V). Acr3 did not confer resistance to As(V) and Rox(V), while ArsH did not confer resistance to Rox(V).

Alterations in Bacterial Metabolism Contribute to the Lifespan Extension Exerted by Guarana in Caenorhabditis elegans.

Guarana (Paullinia cupana) is a widely consumed nutraceutical with various health benefits supported by scientific evidence. However, its indirect health impacts through the gut microbiota have not been studied. Caenorhabditis elegans is a useful model to study both the direct and indirect effects of nutraceuticals, as the intimate association of the worm with the metabolites produced by Escherichia coli is a prototypic simplified model of our gut microbiota. We prepared an ethanoic extract of guarana seeds and assessed its antioxidant capacity in vitro, with a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, and in vivo, utilizing C. elegans. Additionally, we studied the impact of this extract on C. elegans lifespan, utilizing both viable and non-viable E. coli, and assessed the impact of guarana on E. coli folate production. The extract showed high antioxidant capacity, and it extended worm lifespan. However, the antioxidant and life-extending effects did not correlate in terms of the extract concentration. The extract-induced life extension was also less significant when utilizing dead E. coli, which may indicate that the effects of guarana on the worms work partly through modifications on E. coli metabolism. Following this observation, guarana was found to decrease E. coli folate production, revealing one possible route for its beneficial effects.

Delineating the role of gldcin nephron patterning during kidney development

Glycine is a simple amino acid that is vital to the proper functioning of the body, but its roles during ontogeny are not well understood. Glycine levels are precisely regulated through the glycine cleavage system (GCS), a complex molecular machine that produces one‐carbon units for later metabolism and folate production. Elevated glycine levels due to congenital mutations in GCS components, such as glycine dehydrogenase (gldc), cause devastating human birth defects and the rare disease nonketotic hyperglycinemia (NKH). NKH patients suffer from pleiotropic symptoms including seizures, apnea, lethargy, severe mental retardation, and early death; therefore, it is imperative to elucidate the downstream pathological mechanisms of glycine accumulation, which are to date largely unknown. Our lab, and others, have recently reported gldcdeficiency and glycine addition cause developmental changes in various tissues. Here, we interrogate the role of gldcin renal ontogeny. Whole mount in situ hybridization (WISH) revealed that gldc transcripts were highly expressed in multiple tissues, including the central nervous system and embryonic kidney, recapitulating both mouse and human expression studies. We found that formation of nephron cell populations in the kidney was disrupted in gldcdeficient and glycine treated embryos, where the distal early (DE) segment was increased at the expense of the distal late (DL) segment. These alterations led us to hypothesize that gldc is essential for distal segment patterning, a process that is regulated by interplay between the Iroquois (irx) transcription factors and transcription factor AP‐2a (tfap2a). To test this hypothesis, we have begun to analyze the spatiotemporal expression domains of irx3b and irx1a, which are conserved regulators of intermediate and distal segment development. Using WISH, we found that gldcdeficient and glycine treated animals exhibited an increase in the expression domain of irx1a, coinciding with the changes in DE specific solute transporter gene expression patterns. Taken together, these studies indicate that gldc has essential roles in regulating segment pattern during distal nephron development.

Ex vivo folate production by fecal bacteria does not predict human blood folate status: Associations between dietary patterns, gut microbiota, and folate metabolism

Although some bacteria inhabiting the human gut synthesize folates, it has not yet been established whether bacterial folate biosynthesis can impact human folate status. The main objectives of this study were to evaluate associations between different lifestyle factors and the potential of fecal microbiota to produce folates, and to investigate whether this potential is associated with circulating folate and total homocysteine (tHcy) levels in humans. To this end, we carried out an observational study of two hundred adult participants, with high variance in dietary habits. Diet was determined using three-day food records. Fecal microbiota composition was assessed by 16S rRNA gene amplicon sequencing. To establish the folate-production potential of fecal bacteria, cultures containing feces were incubated under anaerobic conditions for 24 h, and the folate concentration was measured before and after incubation. The folate concentration in cultures was 185.4 ± 228.1 pg/ml/log(CFU/g) (2125.4 ± 2454.3 pg/ml) higher after incubation. This change in concentration was not associated with the healthy eating index that measures diet quality (r = -0.11, p = 0.11), but it was positively associated with low α-diversity (r = -0.18, p < 0.01), and high relative abundance of the Bacteroides, as well as Sutterella and Parasutterella genera. The gut microbiota’s folate producing potential was associated neither with serum folate nor with plasma tHcy levels.In conclusion, some taxa of the native gut microbiota have the ability to synthesize folates under culture conditions, but this bacterial folate biosynthesis capacity does not predict human folate status.

The genomic basis of the Streptococcus thermophilus health-promoting properties

BackgroundStreptococcus thermophilus is a Gram-positive bacterium widely used as starter in the dairy industry as well as in many traditional fermented products. In addition to its technological importance, it has also gained interest in recent years as beneficial bacterium due to human health-promoting functionalities. The objective of this study was to inventory the main health-promoting properties of S. thermophilus and to study their intra-species diversity at the genomic and genetic level within a collection of representative strains.ResultsIn this study various health-related functions were analyzed at the genome level from 79 genome sequences of strains isolated over a long time period from diverse products and different geographic locations. While some functions are widely conserved among isolates (e.g., degradation of lactose, folate production) suggesting their central physiological and ecological role for the species, others including the tagatose-6-phosphate pathway involved in the catabolism of galactose, and the production of bioactive peptides and gamma-aminobutyric acid are strain-specific. Most of these strain-specific health-promoting properties seems to have been acquired via horizontal gene transfer events. The genetic basis for the phenotypic diversity between strains for some health related traits have also been investigated. For instance, substitutions in the galK promoter region correlate with the ability of some strains to catabolize galactose via the Leloir pathway. Finally, the low occurrence in S. thermophilus genomes of genes coding for biogenic amine production and antibiotic resistance is also a contributing factor to its safety status.ConclusionsThe natural intra-species diversity of S. thermophilus, therefore, represents an interesting source for innovation in the field of fermented products enriched for healthy components that can be exploited to improve human health. A better knowledge of the health-promoting properties and their genomic and genetic diversity within the species may facilitate the selection and application of strains for specific biotechnological and human health-promoting purpose. Moreover, by pointing out that a substantial part of its functional potential still defies us, our work opens the way to uncover additional health-related functions through the intra-species diversity exploration of S. thermophilus by comparative genomics approaches.

Probiotic Potential of a Folate-Producing Strain Latilactobacillus sakei LZ217 and Its Modulation Effects on Human Gut Microbiota.

Folate is a B-vitamin required for DNA synthesis, methylation, and cellular division, whose deficiencies are associated with various disorders and diseases. Currently, most folic acid used for fortification is synthesized chemically, causing undesirable side effects. However, using folate-producing probiotics is a viable option, which fortify folate in situ and regulate intestinal microbiota. In this study, the folate production potential of newly isolated strains from raw milk was analyzed by microbiological assay. Latilactobacillus sakei LZ217 showed the highest folate production in Folic Acid Assay Broth, 239.70 ± 0.03 ng/μL. The folate produced by LZ217 was identified as 5-methyltetrahydrofolate. LZ217 was tolerant to environmental stresses (temperature, pH, NaCl, and ethanol), and was resistant to gastrointestinal juices. Additionally, the in vitro effects of LZ217 on human gut microbiota were investigated by fecal slurry cultures. 16S rDNA gene sequencing indicated that fermented samples containing LZ217 significantly increased the abundance of phylum Firmicutes and genus Lactobacillus, Faecalibacterium, Ruminococcus 2, Butyricicoccus compared to not containing. Short-chain fatty acids (SCFAs) analysis revealed that LZ217 also increased the production of butyric acid by fermentation. Together, L. sakei LZ217 could be considered as a probiotic candidate to fortify folate and regulate intestinal microecology.

Effects of cultivation conditions on biofortification of yogurt with natural folate by Propionibacterium freudenreichii

In this study, Propionibacterium (P.) freudenreichii was used for the folate production by co-culturing with traditional yogurt starter cultures in synbiotic yogurt and then it was compared to control yogurt containing only LAB starters. First, effects of process variables, including two probiotic strains, inoculum size, inulin concentration, milk fat percentage, sunflower oil content, fermentation temperature and storage time of incubation at 4 °C on folate production by P. freudenreichii were investigated using screening method of Plackett-Burman design. Then, optimization of folate production process was carried out using three main effects of inulin concentration, fermentation temperature and storage time at 4 °C using central composite design. Analysis of variance showed that the models were highly significant (p ≤ 0.05) and folate production affected by the three factors. Optimized folate production by P. freudenreichii ssp. shermanii in yogurts was achieved after 12 d of storage at 4 °C in 2% (w/v) skim-milk inulin as prebiotic and fermentation temperature of 38.6 °C. Reconfirmation assays showed that the highest produced folate included 39.2 ± 0.7 μg 100 mL-1 in FW yogurt containing P. freudenreichii ssp. shermanii under the highlighted optimum conditions, which increased by 49% of the total folate production, compared to control yogurt, which contained only LAB starters. Furthermore, textural properties were assessed using texture analysis. Coculture of P. freudenreichii with traditional yogurt starter culture and addition of inulin to yogurt samples increased hardness, consistency and cohesiveness values in yogurt samples. Results showed that by co-culturing of P. freudenreichii ssp. shermanii in yogurt, in addition to producing a probiotic product, also increases the amount of folate production compared to control yogurt, which contained only LAB starters.

Elucidation of the biosynthetic pathway of B-group vitamins via genome mining of food-derived \(\textit{Bacillus velezensis}\) VTX20

B-vitamins are micronutrients that play an important role in various cellular processes of organisms, which are only synthesized by plants, yeasts, and bacteria. Since animals and humans lack the ability to synthesize B-vitamins, supplements of vitamins from dietary and the B-vitamin producing bacteria are required. In this study, we, for the first time, shed some light on biosynthetic pathways involved in folate (vitamin B9), riboflavin (vitamin B2), and biotin (vitamin B7) production in Bacillus velezensis VTX20. The genome-wide comparison revealed that B. velezensis VTX20 shared high similarities with B. tequilensis KCTC 13622, B. subtilis 168, B. amyloliquefaciens DSM 7. Genomic analysis revealed the presence of a complete folate biosynthesis pathway in which some core components were not found in most Bacillus species. Moreover, strain VTX20 also had the metabolic pathways for riboflavin and biotin that are important probiotic traits. These results highlighted that B. velezensis VTX20 is a producer of B-vitamins, which can be applied further in the agricultural biotechnology industry.

Folate in Milk Fermented by Lactic Acid Bacteria from Different Food Sources.

Folates are essential micronutrients, and folate deficiency still occurs in many countries. Lactic acid bacteria (LAB) are known to be able to synthesize folates during fermentation, but the folate production is strain-dependent and influenced by the fermentation medium, presence of a folate precursor, and fermentation time. This study aimed to screen extracellular folate-producing LAB from local food sources and evaluate the factors influencing their folate biosynthesis during milk fermentation. The selection of folate-producing LAB was based on their ability to grow in folate-free medium (FACM), with folate concentrations quantified by microbiological assay. Growth of the 18 LAB in FACM varied between isolates, with only 8 isolates growing well and able to synthesize extracellular folate at relatively high concentrations (up to 24.27 ng/mL). The isolates with highest extracellular folate levels, Lactobacillus fermentum JK13 from kefir granules, Lactobacillus plantarum 4C261 from salted mustard, and Lactobacillus rhamnosus R23 from breast milk, were applied to milk fermentation. The last two isolates were probiotic candidates. The three isolates consumed folate when it was present in the milk, and its consumption was in line with their growth. The availability of folate precursors affected the amount of folate consumed, but did not lead to increased folate concentrations in the medium after 72 h fermentation. The results of this study indicate that these isolates cannot be utilized for producing folate in folate-containing milk, as it shows feedback inhibition on folate biosynthesis.

Succinylsulfathiazole modulates the mTOR signaling pathway in the liver of c57BL/6 mice via a folate independent mechanism.

Researchers studying the effect of folate restriction on rodents have resorted to the use of the antibiotic succinylsulfathiazole (SST) in the folate depleted diet to induce a folate deficient status. SST has been used extensively in rodent studies since the 1940s. Its localized effect on the gut bacteria as well as its effectiveness in reducing folate producing species is well documented. The possible overlap between the pathways affected by folate depletion and SST could potentially produce a confounding variable in such studies. In our novel study, we analyzed the effect of SST on folate levels in c57Bl/6 male mice fed folate supplemented and deficient diets. We did not observe any significant difference on growth and weight gain at 21weeks. SST did not significantly affect folate levels in the plasma, liver and colon tissues; however, it did alter energy metabolism and expression of key genes in the mTOR signaling pathway in the liver. This research sheds light on a possible confounding element when using SST to study folate depletion due to the potential overlap with multiple critical pathways such as mTOR. SUMMARY: The antibiotic succinylsulfathiazole (SST) is used to reduce folate producing bacteria in rodent folate depletion studies. SST can modulate critical energy and nutrient sensing pathways converging onto mTOR signaling, and potentially confounding cancer studies.

Gldc is essential for embryonic development and kidney organogenesis

Glycine is a simple amino acid that is vital to the proper functioning of the body, but its roles during ontogeny are not well understood. Glycine levels are precisely regulated through the glycine cleavage system (GCS), a complex molecular machine that produces one-carbon units for later metabolism and folate production. Elevated glycine levels due to congenital mutations in GCS components, such as glycine dehydrogenase (gldc), cause devastating human birth defects and the rare disease nonketotic hyperglycinemia (NKH). The exact pathological implications of glycine accumulation are unknown, but NKH patients suffer from pleiotropic symptoms including seizures, apnea, lethargy, severe mental retardation, and early death. Animal models utilizing mice and zebrafish have shown increased ventriculomegaly and motor dysfunction in gldc deficient animals, but have not analyzed organogenesis. Here, we have utilized zebrafish as a genetic platform to explore the roles of gldc during early development. Whole mount in situ hybridization revealed that gldc transcripts were highly expressed in multiple tissues, including the central nervous system and embryonic kidney, recapitulating both mouse and human expression studies. gldc deficiency caused hydrocephalus and pericardial edema, suggesting defects in brain and renal ontogeny. Elevated apoptosis was detected in locations within the brain, kidney, and cloaca. Furthermore, formation of nephron cell populations in the kidney was disrupted in gldc deficient embryos, where the proximal straight tubule was increased at the expense of the distal tubule. Morphological and expression studies revealed severe alterations in brain patterning and morphogenesis as well. Taken together, these studies indicate that gldc has essential roles during early development and have revealed novel functions for pattern formation and cell survival in tissues such as the brain and kidney. Our evidence further emphasizes the zebrafish as a faithful model to examine NKH. Ongoing work with the zebrafish model is poised to expand our medical understanding about the developmental defects consequent to NKH and can be used to perform chemical screens to identify potential therapeutic interventions.

Genome Characterization of Lactiplantibacillus plantarum Strain UTNGt2 Originated from Theobroma grandiflorum (White Cacao) of Ecuadorian Amazon: Antimicrobial Peptides from Safety to Potential Applications.

The genome characterization of the Lactiplantibacillus plantarum strain UTNGt2, isolated from wild copoazu or white cacao (Theobroma grandiflorum), is described. A total of 31 contigs is assembled with a total length of 3,264,448 bases, with all contigs matching the core genome of different groups in the database. The genome size is 3,540,752 bases with GC content of 44.53% and the genome repeat sequences constitute around 457,386 bases of the assembly. The UTNGt2 matches the Lactiplantibacillus plantarum genome with 99% identity. The genome contains 3115 genes, 3052 protein-coding genes, assigned with the EggNOG database. On the basis of the results, 745 proteins are classified with an unknown function, from which 128 proteins have no match in the BLASTN database. It also contains 57 tRNAs, 5 copies of 5S rRNA, and 1 copy of tmRNA. Based on gene prediction and annotation results, 9.4% of proteins are involved in carbohydrate transport and metabolism and 8.46% in transcription, 2.36% are responsible for defense mechanisms, 0.5% are responsible for the biosynthesis of secondary metabolites, transport, and catabolism, while 25.11% have an unknown function. The genome revealed the presence of genes involved in riboflavin and folate production, the presence of CRISPR/Cas genes, phage sequences, the absence of acquired antibiotics resistance genes, virulence, and pathogenic factors, suggesting that UTNGt2 is a safe strain. Its highly antimicrobial capacity is related to the presence of two bacteriocin clusters (class IIc) of the sactipeptide class (contig 4) and plantaricin E class (contig 22), as detected by the BAGEL 4 webserver. Several RiPP-like peptides (non-bactericidal ribosomally produced and post-translationally modified peptides), polyketides (PKs), and terpenes were predicted. Whole-genome sequencing analysis revealed that the UTNGt2 strain has diverse bacteriocins with a high inhibitory capacity, thus it is a bacteriocinogenic strain. Considering the safety profile, UTNGt2 is a nonpathogenic, nonvirulent strain with valuable biotechnological traits and can be further exploited for its probiotic and antimicrobial potential in the food industry or as a potential producer strain of antimicrobial peptides as an alternative to conventional antibiotics.