November 30, 2024

Bioengineered uterus

Towards a bioengineered uterus: bioactive sheep uterus scaffolds are effectively recellularized by enzymatic preconditioning



Uterine factor infertility was considered incurable until recently when we reported the first successful live birth after uterus transplantation. However, risky donor surgery and immunosuppressive therapy are factors that may be avoided with bioengineering.

 For example, transplanted recellularized constructs derived from decellularized tissue restored fertility in rodent models and mandate translational studies. In this study, we decellularized whole sheep uterus with three different protocols using 0.5% sodium dodecyl sulfate, 2% sodium deoxycholate (SDC) or 2% SDC, and 1% Triton X-100. Scaffolds were then assessed for bioactivity using the dorsal root ganglion and chorioallantoic membrane assays, and we found that all the uterus scaffolds exhibited growth factor activity that promoted neurogenesis and angiogenesis. Extensive recellularization optimization was conducted using multipotent sheep fetal stem cells and we report results from the following three in vitro conditions; (a) standard cell culturing conditions, (b) constructs cultured in transwells, and (c) scaffolds preconditioned with matrix metalloproteinase 2 and 9. 

The recellularization efficiency was improved short-term when transwells were used compared with standard culturing conditions. However, the recellularization efficiency in scaffolds preconditioned with matrix metalloproteinases was 200–300% better than the other strategies evaluated herein, independent of decellularization protocol. Hence, a major recellularization hurdle has been overcome with the improved recellularization strategies and in vitro platforms described herein. These results are an important milestone and should facilitate the production of large bioengineered grafts suitable for future in vivo applications in the sheep, which is an essential step before considering these principles in a clinical setting.

Bioengineered uterus, sheep uterus scaffold, bioactive scaffold, enzymatic preconditioning, tissue engineering, regenerative medicine, uterine recellularization, extracellular matrix, decellularization, recellularization techniques, organ scaffolding, biomaterials, biocompatibility, fertility restoration, uterine transplantation, cell-seeded scaffolds, reproductive tissue repair, bioengineering, scaffold bioactivity, enzymatic treatment, organ regeneration, uterine repair, stem cell integration, fertility research, translational medicine, biomedical innovation.

#BioengineeredUterus, #TissueEngineering, #RegenerativeMedicine, #UterineScaffold, #FertilityRestoration, #Biomaterials, #Decellularization, #Recellularization, #UterineTransplantation, #OrganRegeneration, #ReproductiveHealth, #StemCellResearch, #Bioengineering, #ExtracellularMatrix, #TranslationalMedicine, #BiomedicalInnovation, #EnzymaticPreconditioning, #ScaffoldTechnology, #UterineRepair, #Biocompatibility, #FertilityResearc

    at No comments:

    November 29, 2024

    Pathogen Genomic Surveillance

    WHO and partners announce grants to boost pathogen genomic surveillance


    The World Health Organization (WHO) and partners announced 10 projects that will receive almost US$ 2 million in grants to improve capacities in pathogen genomic surveillance.

    The catalytic grant fund was established by the International Pathogen Surveillance Network (IPSN) to support partners from low- and middle-income countries to build their capacities in pathogen genomic analysis. This technology analyses the genetic code of viruses, bacteria and other disease-causing organisms to understand, in conjunction with other data, how easily they spread, and how sick they can make people. This data allows scientists and public health teams to track and respond to infectious disease threats, supports the development of vaccines and treatments and empowers countries to take faster decisions.

    The fund is hosted by the United Nations Foundation and supported by the Bill & Melinda Gates Foundation, The Rockefeller Foundation and Wellcome.

    "The IPSN catalytic grant fund recipients will accelerate the benefits of pathogen genomic surveillance in low- and middle-income settings, as well as explore new applications for genomic surveillance, such as wastewater surveillance," said Manisha Bhinge, Vice President of the Health Initiative at The Rockefeller Foundation. "Pandemics and epidemics continue to be a global threat, further amplified by climate change. There is urgent need for equitable access to these tools and capabilities to protect lives in vulnerable communities."

    One of the recipients, the American University of Beirut, will use wastewater surveillance to study how diseases spread in refugee populations, helping to ensure that people can quickly receive the care and support they need in migration settings. Another grantee, the Pasteur Institute of Laos, will use the funding to develop new methods to track avian flu in live-bird markets, a setting that is often overlooked but vital to millions of people worldwide.

    "If we are to protect vulnerable populations from the devastating impacts of disease, we first need to better understand how these pathogens spread, evolve and cause illness. These projects, developed in-country and tailored to local priorities, will generate new insights, knowledge and evidence that will help track global pathogen trends and inform evidence-based decisions to implement effective interventions" said Titus Divala, Interim Head of Epidemics and Epidemiology at Wellcome.

    The Federal University of Rio de Janeiro in Brazil will use the funding to develop an open-source bioinformatics tool that can be used to conduct offline analyses. The tool will be piloted in Latin America with potential for global use, especially in low-resource settings.

    "SARS-CoV-2 and subsequent regional disease outbreaks have underscored the importance of access to genomic surveillance tools in all countries. The IPSN's catalytic investments will generate data and innovative methods to support the much-needed scale-up in LMICs," said Simon Harris of the Gates Foundation.

    The grantees were announced at the IPSN Global Partners Forum held in Bangkok, Thailand, from 21–22 November. The event was co-hosted by the WHO Regional Offices for South-East Asia and the Western Pacific and the Centre for Pathogen Genomics at the Doherty Institute in Australia.

    Pathogen genomic surveillance, infectious disease monitoring, genomic epidemiology, whole-genome sequencing, outbreak investigation, pathogen evolution, genetic diversity, molecular epidemiology, antimicrobial resistance, viral genomics, bacterial genomics, metagenomics, phylogenetic analysis, public health genomics, data sharing, genomic databases, bioinformatics tools, surveillance networks, epidemic preparedness, genomic sequencing platforms, pathogen tracking, genomic mutations, genomic data analysis, early outbreak detection, global health security, pandemic response

    #PathogenGenomics, #GenomicSurveillance, #InfectiousDisease, #OutbreakResponse, #MolecularEpidemiology, #PublicHealth, #AntimicrobialResistance, #Phylogenetics, #DataSharing, #GlobalHealth, #DiseaseMonitoring, #ViralGenomics, #BacterialGenomics, #Metagenomics, #PandemicPreparedness, #HealthSecurity, #Bioinformatics, #GenomicEpidemiology, #PathogenEvolution, #SurveillanceScience, #EpidemicControl, #WholeGenomeSequencing, #EarlyDetection, #GenomicData, #HealthInnovation

    International Conference on Genetics and Genomics of Diseases 




    For Enquiries: genetics@healthcarek.com 

    Get Connected Here 
    --------------------------------- 
    --------------------------------- 
    at No comments:

    November 28, 2024

    Transgenic Arabidopsis Plants

    The Maize Gene ZmGLYI-8 Confers Salt and Drought Tolerance in Transgenic Arabidopsis Plants




    Methylglyoxal (MG), a highly reactive and cytotoxic α-oxoaldehyde compound, can over-accumulate under abiotic stress, consequently injuring plants or even causing death. Glyoxalase I (GLYI), the first enzyme of the glyoxalase pathway, plays multiple roles in the detoxification of MG and in abiotic stress responses. However, the GLY1 gene in maize has been little studied in response to abiotic stress. In this study, we screened a glyoxalase I gene (ZmGLYI-8) and overexpressed in Arabidopsis. This gene was localized in the cytoplasm and can be induced in maize seedlings under multiple stress treatments, including salt, drought, MG, ABA, H2O2 and high temperature stress. Phenotypic analysis revealed that after MG, salt and drought stress treatments, overexpression of ZmGLYI-8 increased the tolerance of transgenic Arabidopsis to MG, salt and drought stress. Furthermore, we demonstrated that the overexpression of ZmGLYI-8 scavenges accumulated reactive oxygen species, detoxifies MG and enhances the activity of antioxidant enzymes to improve the resistance of transgenic Arabidopsis plants to salt and drought stress. In summary, this study preliminarily elucidates the molecular mechanism of the maize ZmGLYI-8 gene in transgenic Arabidopsis and provides new insight into the breeding of salt- and drought-tolerant maize varieties.

    Materials and Methods

    Gene Isolation and Bioinformatics Analysis of ZmGLYI-8
    Based on the B73 genome in the MaizeGDB database, Primer 5.0 software was used to design specific primers for ZmGLYI-8. Total RNA was extracted from the leaves of ten-day-old maize seedlings with TRIzol reagent. The concentration and integrity of total RNA were determined by using formaldehyde gel electrophoresis and a NanoDrop 2000 spectrophotometer (Thermo Scientifi, Shanghai, China). CDNA from B73 leaves was synthesized by using HiScript®II Q Select RT SuperMix (Vazyme, Nanjing, China). The pMD19-T cloning vectors with the PCR products of ZmGLYI-8 were transformed into Escherichia coli DH5α, which was subsequently sequenced by Boshang (Shanghai, China). The molecular weight and isoelectric point of ZmGLYI-8 were evaluated by ProtParam (http://web.expasy.org/protparam/ (accessed on 1 October 2024)), and the conserved functional domains of this gene were predicted by using NCBI CD-Search (https://www.ncbi.nlm.nih.gov/cdd/ (accessed on 1 October 2024)). The similarity of amino acid sequences was determined by using DNAMAN v6 software for multiple sequence alignments. MEGA 4.0 software was used for generating the phylogenetic tree.

    Stress Treatment of B73

    The maize inbred line B73 was used for investigating the expression pattern of ZmGLYI-8 in different tissues and under various stress treatments. The seeds of inbred line B73 were preserved in our laboratory. Roots, stems and leaves of B73 at the V6 stage were collected for the expression pattern analysis of different tissues. B73 seeds were germinated in quartz sand, and the plants were cultivated at 25 °C under a 14 h light/10 h dark cycle. After ten days, the seedlings were transferred to Hoagland solution supplemented with 150 mM NaCl, 10% (w/v) PEG-6000, 10 mM MG, 0.5 mM H2O2, 50 µM ABA, 50 µM MeJA or 5 mM ET. Other seedlings were subjected to 38 °C for the high-temperature (HT) treatment. Leaf samples were collected 0, 1, 6, 12 and 24 h after the different treatments. Each biological replicate contained three plants, and all ZmGLYI-8 expression pattern experiments were performed by using three biological replicates. All samples were frozen directly in liquid nitrogen and then stored at −80 °C before RNA extraction. The specific gene primers used for expression pattern analysis.

    Subcellular Localization

    The subcellular localization of ZmGLYI-8 was predicted by the SubLoc program. Moreover, the expression vector PROKII-GFP was used to investigate the subcellular localization of ZmGLYI-8. The coding sequence (CDS) of ZmGLYI-8 without a termination codon was amplified by polymerase chain reaction (PCR) and then fused to the PROKII-GFP vector and driven by the CaMV35S promoter. The ZmGLYI-8 primers used for plasmid construction are listed in Table S1. The Agrobacterium tumefaciens strains GV3101 with the 35S::ZmGLYI-8-GFP plasmid or 35S::GFP plasmid were injected into four-leaf-stage Nicotiana benthamiana plants. Three days after infiltration, GFP fluorescence was observed by using an LSM 880 confocal laser scanning microscope.

    Prokaryotic Expression of ZmGLYI-8

    The full-length CDS of ZmGLYI-8 was fused to the pET-30a vector and then transformed into Escherichia coli Rosetta. PCR identification and sequencing analysis (Boshang) were used for selecting positive clones. The IPTG-induced bacteria were diluted with LB liquid medium in gradients of 10−2, 10−4 and 10−6. Three microliters of different concentrations of bacteria was added dropwise to LB solid medium supplemented with 0.5 mM MG, 200 mM NaCl or 200 mM mannitol. The growth of the recombinant strain pET-30a-ZmGLYI-8 and the control strain pET-30a was observed after culture at 37 °C for 12 h.

    Development of Transgenic Arabidopsis

    The coding sequence of the ZmGLYI-8 gene was cloned and inserted into pBI121 with the CaMV35S promoter. The resulting plasmid was subsequently introduced into Agrobacterium tumefaciens GV3101, which was subsequently used for floral dip transformation of Arabidopsis Columbia-0 (WT) plants. Transgenic Arabidopsis plants were screened on 1/2 MS solid medium supplemented with 50 mg/L kanamycin. The expression level of ZmGLYI-8 in positive transgenic plants was further measured by qRT–PCR. T3 transgenic Arabidopsis plants were used for subsequent experimentation. Fifty seeds of WT and ZmGLYI-8 transgenic Arabidopsis plants were sown on 1/2 MS solid medium supplemented with 5 mM MG. These petri dishes were placed in a growth chamber at 22 °C with a 16-h light/8-h dark cycle. The germination rate of the seeds was calculated every day, and photos were taken on the 7th day after sowing.

    Expression Analysis by qRT–PCR

    Total RNA from maize and Arabidopsis plants was extracted for qRT–PCR using the TRIzol reagent (TaKaRa, Beijing, China). HiScript®II Q Select RT SuperMix (Vazyme, China) was used for synthesizing cDNA from the total RNA. According to the manufacturers’ protocols, qRT–PCR experiments were performed by using a Bio-Rad CFX96 real-time system with AceQ®qPCR SYBR Green Master Mix (Vazyme, China). The inner reference gene Actin was used for normalizing the expression levels of genes. The gene expression levels were calculated by using the 2−ΔΔCt method. Triplicate independent biological assays of each cDNA sample were performed to ensure accurate statistical analysis.

    Assays for Salt and Drought Tolerance

    To detect seed germination of ZmGLY1-8 transgenic Arabidopsis lines under MG stress, the sterile seeds of the WT and transgenic Arabidopsis lines (OE3, OE4 and OE5) were sown on 1/2 MS solid medium with MG (0 and 2.5 mM). The number of seed germinations was recorded every day, and the photos were taken on the 7th day after the seeds were sown. For the root length experiment, the sterilized WT and transgenic line seeds were sown on 1/2 MS medium for vernalization at 4 °C for 3 days. Then, these seeds were vertically cultivated at 22 °C under a 16 h light/8 h dark cycle. After five days, the seedlings were transplanted into 1/2 MS solid medium supplemented with 0.5 mM MG, 150 mM NaCl and 200 mM mannitol. The primary root lengths of the transgenic Arabidopsis lines were analyzed by using ImageJ 1.50I software after 7 days of stress treatment. The experiment was repeated with three biological replicates.

    Floating Leaf Disc and Chlorophyll Extraction

    Leaf discs (1 cm in diameter) were excised from the third and fourth true rosette leaves of the WT and ZmGLY1-8 transgenic plants. The discs were placed in water (simulated control) or under multiple stress conditions (simulated stress treatment) for 48 h, including salt stress in 200 mM and 400 mM NaCl, drought stress in 200 mM and 400 mM mannitol and MG stress in 1 mM and 2.5 mM. The chlorophyll content was determined by an ultraviolet spectrophotometer after extraction of all the leaf discs with 95% ethanol. The experiment was repeated with three biological replicates.

    Physiological Analysis and MG Content Determination

    Four-week-old Arabidopsis plants were treated with 400 mM NaCl or 200 mM 10% PEG-6000 for 7 days, after which leaves from the same parts were removed for physiological measurements. The contents of O2•− and H2O2 were measured using kits purchased from Suzhou Grace Biotechnology Co., Ltd., Suzhou, China. About 0.1g of the sample was taken, the extract was added, and the supernatant was collected as the solution to be measured after centrifugation. The reaction solution was added according to the kit instructions, and O2•− and H2O2 were measured at 540 nm and 415 nm, respectively. The MDA content was also measured using kits from the same company. MDA was measured according to the kit instructions. First, 0.1 g of sample was taken, 1 mL of extraction solution was added, and the supernatant was collected after centrifuged at 12,000× g for 10 min at 4 °C. Next, 600 mL of working solution was added, and the supernatant was removed from the water bath at 90 °C for 30 min. MDA was measured at 532 nm and 600 nm. The experiment was conducted using three biological replicates. The activities of CAT, POD and SOD were determined by Wu’s method. In order to determine the CAT, POD and SOD, fresh samples were taken, and 5 mL pre-cooled phosphoric acid buffer was added. Then, the mixture was ground mechanically and centrifuged at 10,000× g for 20 min at 4 °C. The collected supernatant was used for subsequent assays. SOD was measured at 560 nm using the nitrogen blue tetrazolium (NBT) method. POD and CAT activities were determined at 470 nm and 240 nm, respectively, with absorbance changes every 10 s. The methylglyoxal content was determined according to previously described methods. MG content was measured from 300 mg of fresh leaf tissue. First, 0.5 M of 3 mL perchloric acid was added, ice-bathed for 15 min and centrifuged at 12,000× g for 10 min at 4 °C. Second, the colored supernatant was transferred to a new centrifuge tube, bleached with charcoal (10 mg/mL) for 15 min at room temperature and centrifuged at 11,000× g for 10 min. Then, the supernatant was transferred to a new centrifuge tube, and the reaction was neutralized by the addition of saturated potassium carbonate at room temperature for 15 min and centrifuged at 11,000× g for 10 min; 650 μL of supernatant was collected. Finally, the reaction mixture (1 mL), containing 250 mL of 7.2 mM 1,2-diaminobenzene, 100 mL of 5 M perchloric acid, 10 mL of 100 mM NaN3 and 650 μL of neutralized supernatant, was incubated for 3 h at room temperature, and the absorbance was measured at 336 nm.

    Statistical Analysis

    All data are presented as the mean of three biological replicates ± SD. One-way ANOVA and Student’s t-test were used for statistical analysis, and p < 0.05 is considered significant. One-way ANOVA was performed using SPSS 22.0, and the other analyses were performed using Microsoft Office Excel 2010.

    Transgenic Arabidopsis plants, genetic engineering, gene expression, plant biotechnology, CRISPR-Cas9, plant transformation, Arabidopsis thaliana, genome editing, transgenic crops, molecular biology, plant genetics, Agrobacterium-mediated transformation, transgene expression, promoter analysis, gene silencing, abiotic stress tolerance, biotic stress resistance, metabolic engineering, functional genomics, RNA interference, overexpression, marker genes, floral dip method, plant tissue culture, stress-responsive genes, transgenic research,

    #TransgenicPlants, #Arabidopsis, #PlantBiotechnology, #GeneEditing, #CRISPR, #PlantTransformation, #ArabidopsisThaliana, #GeneticEngineering, #MolecularBiology, #FunctionalGenomics, #AbioticStress, #BioticStress, #PlantGenetics, #Genomics, #Agrobacterium, #PromoterAnalysis, #GeneSilencing, #RNAi, #PlantResearch, #TissueCulture, #GenomeEditing, #BiotechInnovation, #FloralDipMethod, #PlantScience, #TransgenicResearch

    at No comments:

    November 27, 2024

    Escherichia coli

    Antibacterial Efficacy of Feline-Derived Lactic Acid Bacteria against Enteropathogenic Escherichia coli: A Comprehensive In Vitro Analysis



    Introduction

    Enteropathogenic Escherichia coli (EPEC) is a significant cause of diarrhea in cats, particularly affecting kittens. EPEC leads to severe intestinal damage, resulting in symptoms such as vomiting, lethargy, and dehydration, which can be fatal if untreated. Kittens with EPEC have significantly greater intestinal damage and higher quantities of the pathogen compared to those without diarrhea. Additionally, EPEC is known to cause similar severe diarrheal disease and intestinal damage in other animals, indicating its broad pathogenic potential. Infection with EPEC results in significant fluid and electrolyte losses, exacerbating the risk of dehydration and necessitating medical intervention. This infection poses a substantial economic burden due to the costs of veterinary care, decreased productivity in breeding operations, and emotional distress experienced by pet owners. The conventional treatment for EPEC-induced diarrhea includes antibiotics; however, the growing prevalence of antibiotic-resistant EPEC strains complicates these treatments. This resistance necessitates the search for alternative solutions, with probiotics emerging as a promising approach due to their natural and sustainable benefit.

    Probiotics, which are live microorganisms that confer health benefits to the host, have demonstrated effectiveness in treating diarrhea caused by Escherichia coli (E. coli) in various animal models. These benefits are derived from mechanisms such as the competitive exclusion of pathogens, enhancement of the host immune response, and production of antimicrobial substances. Several studies have highlighted the efficacy of probiotics in addressing E. coli-induced diarrhea. For instance, research has shown that exopolysaccharides produced by Bifidobacterium animalis can mitigate E. coli-induced damage in intestinal epithelial cells by inhibiting apoptosis and restoring autophagy. In another study, a screening of over 1100 Lactobacillus plantarum strains identified several with potent inhibitory effects against enterotoxigenic Escherichia coli (ETEC) K88 in weaned piglets, suggesting the potential of these strains to reduce E. coli infections. Furthermore, strains of Lactobacillus isolated from various sources have demonstrated significant antimicrobial activity against uropathogenic Escherichia coli (UPEC), effectively reducing biofilm formation by up to 50%. Comparative studies on different probiotics also revealed that a multi-strain synbiotic that contained various Lactobacillus and Bifidobacterium strains exhibited significantly stronger inhibition of E. coli and other pathogens compared with single-strain probiotics.

    Given the unique gut microbiota of cats, it is crucial to develop probiotics from cat-derived strains. Host-specific probiotics are more likely to survive in, colonize, and exert beneficial effects on the feline gastrointestinal tract. Research has shown that these species-specific probiotics tend to be more effective than those derived from non-host sources. For example, studies have demonstrated that probiotics isolated from the same species provide better colonization and health benefits compared with commercial probiotics from different species. Further supporting this concept, genetic variation and host-specific adaptation studies revealed that Lactobacillus johnsonii strains exhibit host-specific genetic variations when isolated from different animal hosts, suggesting a co-evolution with their hosts that enhances their effectiveness. Additionally, Enterococcus hirae F2, a strain isolated from the gut of Catla catla fish, showed significant probiotic potential by surviving under highly acidic and bile salt conditions and exhibiting strong antimicrobial activity against pathogens. These findings highlight the importance of using host-specific probiotics for achieving maximum efficacy in treating and maintaining the health of different species.

    This study focused on isolating and screening lactic acid bacteria (LAB) strains from healthy cats’ feces to find probiotics that effectively combat EPEC. Initially, 700 LAB strains were isolated. From these, 200 randomly selected strains underwent 16S rRNA sequencing. These isolates were then tested for antibacterial activity against EPEC. The selected strains were further evaluated for their physiological and biochemical properties, such as their tolerance to different temperatures, salt concentrations, and pH levels, which ensured they can be easily propagated and maintained during production and storage. The safety assessments included testing for hemolytic activity, antibiotic susceptibility, and the absence of virulence and biogenic amine genes, which confirmed the strains’ safety. This research is significant for developing effective, host-specific probiotics for cats, thus offering a natural and sustainable solution to manage EPEC-induced diarrhea and reducing reliance on antibiotics.

    2. Materials and Methods

    2.1. Sample Collection and LAB Isolation

    Three hundred fresh fecal samples from healthy cats were collected from pet stores and catteries in Luohe, China, between September and December 2023. Fecal samples were collected using sterile sampling spoons from the upper part of naturally expelled fecal pellets to avoid contamination. Each sample was placed in a sterile EP tube, labeled, and immediately transported to the laboratory for analysis in a cold chain box with sufficient dry ice. The time from sampling to analysis did not exceed 3 h, ensuring each sample’s integrity. All procedures involving the collection and handling of fecal samples were conducted in compliance with ethical guidelines approved by the Institutional Animal Care and Use Committee of China Agricultural University (AW20704202-5-4). In the laboratory, the fecal samples were serially diluted with sterile distilled water and plated on de Man, Rogosa, and Sharpe (MRS) agar (Merck, Darmstadt, Germany). The plates were incubated at 37 °C for 48 h under anaerobic conditions to culture single LAB colonies. Colonies that were round, raised or flat, creamy white or slightly yellow, moist, medium sized, and neatly edged were preliminarily identified as LAB. These isolates were stored at −80 °C for further testing and analysis.

    2.2. Preliminary Identification of Strains

    Among the approximately 700 preserved LAB strains, 200 strains were randomly selected for a diversity analysis.

    2.2.1. Morphological Characteristics

    The morphological characteristics of the LAB were identified using Gram staining. A drop of sterile water was placed on a slide, and a small amount of a single colony was smeared onto the slide and then air-dried over an alcohol lamp. The smear was stained with ammonium oxalate crystal violet for 1 min and rinsed with sterile water, followed by air-drying over an alcohol lamp. An iodine solution was then added to the slide for approximately 1 min for mordanting, rinsed with sterile water, and air dried. The slide was decolorized with 95% alcohol for 20 s, rinsed with sterile water, and air-dried. Subsequently, the smear was counterstained with safranin for 1 min, rinsed with sterile water, and air-dried. Finally, the slide was examined under a light microscope.

    2.2.2. Physiological Experiments

    The experiments used to identify the physiological characteristics of LAB mainly included the catalase and glucose gas production tests. 

    Catalase test: Place 100 μL of 3% H2O2 on a blank culture dish. Using an inoculating loop, pick a single colony and place it into the H2O2. Observe whether gas bubbles are produced. If bubbles are formed, the result is catalase-positive; if no bubbles are formed, the result is catalase-negative.
    Glucose gas production test: Culture the lactic acid bacteria by inoculating a single colony into MRS broth and incubating at 30 °C for 24 h. Invert a Durham tube in a test tube containing the cultured lactic acid bacteria inoculated at 1% into MRS broth and incubate statically at 30 °C for 7 days. Observe the Durham tube for the presence of gas bubbles. The presence of gas bubbles indicates heterofermentative fermentation, while the absence of bubbles indicates homofermentative fermentation.

    2.2.3. 16 S rRNA Gene Analysis

    The selected 200 LAB strains were identified through genetic analysis using PCR and 16S rRNA gene sequencing. The universal primers 27 F (5′-AGAGTTTGATCCTGGCTC AG-3′) and 1492 R (5′-GGTTACCTTGTTACGACTT-3′) were utilized for the PCR amplification of the 16S rRNA gene. The amplified products were then analyzed by Sangon Biotech Co., Ltd., Shanghai, China. The sequence similarities of each contig were assessed by comparing their homologies in the GenBank database using BLAST (http://www.ncbi.nlm.nih.gov accessed on 12 January 2024). A phylogenetic tree of the selected high-performance strains was subsequently constructed using the neighbor-joining method with MEGA-X software (version 10.1.5).

    2.3. Screening of LAB with Strong Antibacterial Activity against EPEC

    The well diffusion technique was adapted from the method described by Sirichokchatchawan et al., with modifications to suit this study. The target bacterium, EPEC, was cultured in a nutrient liquid medium and incubated at 37 °C with shaking at 180 rpm for 12 h until the concentration reached 1 × 108 CFU/mL. Then, 200 µL of the bacterial suspension was spread on LB agar plates. Separately, 200 µL of cultures of different LAB strains, each at a concentration of 1 × 108 CFU/mL, that had been incubated for 16 h was added to wells (10 mm in diameter) punched into the LB agar plates. Uninoculated MRS broth and penicillin served as the negative and positive controls, respectively. The EPEC strain was kindly provided by China Agricultural University. The LAB strains that produced inhibition zones greater than 18 mm in diameter were selected for further physiological and biochemical characterization.

    2.4. Physiological and Biochemical Characteristics of Selected LAB Isolates

    Following the methodology described by Zhang et al., the physiological and biochemical characteristics of the selected LAB strains, including pH, salt, and temperature tolerance, were evaluated using MRS broth. Single LAB colonies were initially picked and inoculated into 20 mL of sterile MRS liquid medium. The cultures were incubated at 37 °C for 16 h, after which the OD was adjusted to 0.8 at 600 nm using sterile water. Then, 100 µL of each LAB suspension was mixed with 9.9 mL of MRS broth.
    Acid and alkaline resistance: The LAB strains were tested for acid and alkaline resistance by culturing them in MRS broth adjusted to various pH levels (3.0, 3.5, 4.0, 4.5, 5.0, 6.0, 9.0, and 10.0). These cultures were incubated at 37 °C for 7 days.

    Salt tolerance: The salt tolerance was assessed by culturing the LAB strains in MRS broth containing 3.0% and 6.5% NaCl, with the incubation at 37 °C for 48 h.

    Temperature tolerance: The temperature tolerance of the LAB strains was determined by incubating them in MRS broth at different temperatures (5 °C, 10 °C, 45 °C, and 50 °C) for 7 days.
    Post-incubation, the growth rates of the LAB strains were measured using the turbidimetry method by recording absorbance values using the OD at 600 nm. The visual turbidity was also noted. A sterile MRS medium without inoculation served as the control, with its OD600 value recorded as 0. The growth was categorized as follows based on the OD600 readings: 0 ≤ OD600 ≤ 0.2 indicated no growth (“−”); 0.2 < OD600 ≤ 0.6 indicated weak growth (“w”); and OD600 > 0.6 indicated growth (“+”).

    2.5. Hydrophobicity and Auto-Aggregation Ability of Selected LAB Isolates

    The adhesion ability of the selected LAB strains, as indicated by the cell surface hydrophobicity and auto-aggregation, was tested using the methods described by Wang et al.. The strains that exhibited high cell surface hydrophobicity and auto-aggregation were further analyzed for gastrointestinal tolerance.

    2.6. Survival of Representative Strains in GI Fluids

    The simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) tests were performed following the methods described by Zhang et al. The LAB strains were exposed to SGF for 3 h and SIF for 4 h. The strains that demonstrated strong tolerance to both SGF and SIF were selected for further safety analysis.

    2.7. Safety Evaluation

    2.7.1. Hemolytic Activity

    Hemolytic activity was assessed using blood agar plates according to the manufacturer’s instructions. Fresh bacterial strains ZY25, ZY33, and ZY35 were streaked on Columbia blood agar plates. Staphylococcus aureus ATCC 29213T was used as the positive control. The plates were incubated, and the hemolytic activity was evaluated by observing the clear zones around the bacterial colonies, which indicated hemolysis.

    2.7.2. Antibiotic Susceptibility

    The antibiotic susceptibility of LAB isolates was determined using a modified disk diffusion method based on the protocol described by Niu et al. The antibiotics tested included gentamicin (GEN, 10 μg/disk), ciprofloxacin (CIP, 5 μg/disk), ceftriaxone (CTR, 30 μg/disk), erythromycin (E, 15 μg/disk), ampicillin (AMP, 10 μg/disk), tetracycline (TET, 30 μg/disk), compound sulfamethoxazole (SXT, 25 μg/disk), chloramphenicol (C, 30 μg/disk), lincomycin (MY, 2 μg/disk), and penicillin (PEN, 10 μg/disk). The results were expressed in millimeter diameters of the inhibition zones. The susceptibility of the isolates was classified as resistant, intermediate resistant, or susceptible according to the cutoff values proposed by de Souza et al. Each test was conducted in triplicate to ensure accuracy and reproducibility.

    2.7.3. PCR Screening of Strains for Virulence Factors, Biogenic Amines, and Antibiotic Resistance Genes

    The genetic traits related to virulence factors, biogenic amines, and antibiotic resistance in strains ZY25 and ZY35 were screened using PCR protocols following the method outlined by Wang et al. Enterococcus faecalis ATCC 29212T, which harbors the target virulence genes (ace, cylA, and gelE), was used as the positive control, while Milli-Q water was used as the negative control.

    The antimicrobial activity of ZY25 and ZY35 against various pathogenic bacteria was evaluated using the agar well diffusion method, as referenced from Section 2.3 of the standard protocol. The broad-spectrum antimicrobial efficacy of ZY25 and ZY35 were tested against the following bacterial strains: Pseudomonas aeruginosa CICC 23694T, Staphylococcus aureus ATCC 29213T, Listeria monocytogenes CICC 23929T, Escherichia coli CICC 24189T, Bacillus subtilis CICC 10275T, and Shigella dysenteriae CICC 23829T.

    The antimicrobial substances produced by strains ZY25 and ZY35 were investigated using the method described by Ni et al. The experiments aimed to eliminate the effects of acid, hydrogen peroxide, and protease hydrolysis. First, the pH of the fermentation broths was adjusted within a range of 2.5 to 10.0 using 0.2 M hydrochloric acid and 0.2 M sodium hydroxide solutions. To remove the hydrogen peroxide, the broths were treated with a 0.5 mg/mL catalase solution and incubated at 37 °C for 2 h. For the protease treatment, the pH was adjusted to 6.0, and the broths were mixed with 1 mg/mL of proteinase K, trypsin, and pepsin, respectively, and then incubated at 37 °C for 2 h. Following these treatments, the broths were centrifuged at 8000 rpm for 10 min, and the supernatants were collected. The antimicrobial activity of the supernatants against EPEC was assessed using the agar well diffusion method, with untreated MRS broth supernatants as controls. Each experiment was performed in triplicate to ensure accuracy.

    Each LAB colony was isolated and cultured in 20 mL of sterile MRS broth. The optical density at 600 nm (OD600) and colony forming units (CFU/mL) were measured at 2-h intervals up to 24 h post-inoculation at 37 °C. Additionally, the pH of each fermentation solution was recorded at 6-h intervals up to 48 h post-inoculation at 37 °C.

    2.8.4. Determination of Organic Acid Content by High-Performance Liquid Chromatography (HPLC)
    Chemicals and reagents (methanol, acetonitrile, formic acid, 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide, chloroform, and 3-nitrophenylhydrazine) were sourced from ANPEL (Shanghai, China), with all solvents being LC-MS grade. Ultra-pure water was prepared using a Milli-Q system (Merck Millipore, Burlington, MA, USA). The samples were extracted with 1 mL of a methanol–chloroform (7:3) solution on ice for 30 min; then, 600 μL of H2O was added, and the mixture was centrifuged at 12,000 rpm at 4 °C for 10 min. The supernatant was collected, and the process was repeated. For the derivatization, 10 μL of 0.1 M EDC and 10 μL of 0.1 M 3NPH were added, and the reaction proceeded for 30 min at 40 °C. The extracts were analyzed using a UPLC-Orbitrap-MS system with a Waters BEH C18 column (50 × 2.1 mm, 1.8 μm) at 40 °C, a flow rate of 0.35 mL/min, and an injection volume of 2 μL. The solvent system was water (0.1% formic acid)–acetonitrile (0.1% formic acid) with a gradient program from 90:10 to 10:90 and back to 90:10. HRMS data were recorded on a Q Exactive hybrid Q-Orbitrap mass spectrometer using Fullms-ms2 methods with the following parameters: −2.8 kV spray voltage, 40 arb sheath gas, 10 arb aux gas, 320 °C capillary temperature, and 350 °C aux gas heater temperature. The data were processed with Xcalibur 4.1 (Thermo Scientific, Waltham, MA, USA) and TraceFinder™ 4.1 Clinical (Thermo Scientific, Waltham, MA, USA), and the results were outputted in Excel format.

    Statistical Analyses

    Each test was performed in triplicate. The data analysis was conducted using one-way ANOVA or a paired t-test in SPSS 22.0 (IBM Corp., Armonk, NY, USA). The results are presented as the mean ± standard error of the mean (SEM), with p < 0.05 indicating statistical significance.

    3. Results

    Isolation and Screening of Lactic Acid Bacteria in Healthy Cat Feces
    Approximately 700 LAB strains were isolated from the feces of 300 healthy cats. A random selection of 200 LAB strains underwent 16S rRNA sequencing. Statistical analysis was performed on the successfully sequenced strains, with the results. Among the 200 LAB strains, the top six strains identified were Ligilactobacillus animalis, Ligilactobacillus salivarius, Enterococcus hirae, Ligilactobacillus agilis, Enterococcus faecium, and Pediococcus acidilactici, with respective proportions of 34.5%, 19%, 13%, 7.5%, 6.5%, and 4%. The physiological characteristics of the strains. Among the LAB strains, 75% were rod-shaped, while 25% were cocci. Additionally, 96% of the strains were homofermentative, i.e., unable to produce gas from glucose, whereas 4% were heterofermentative, i.e., capable of producing gas from glucose. All the strains were Gram-positive and catalase-negative.

    4. Conclusions

    This study successfully identified and characterized feline-derived LAB strains, specifically ZY25 and ZY35, which demonstrated significant antibacterial activity against EPEC. These strains exhibited strong tolerance to various stress conditions, including low pH, bile salts, and gastrointestinal fluids, alongside high hydrophobicity and auto-aggregation abilities, underscoring their potential as probiotics. Safety evaluations further confirmed the absence of hemolytic activity, virulence factors, and antibiotic resistance genes, reinforcing their suitability for safe probiotic use. The antimicrobial efficacy of ZY25 and ZY35 is primarily attributed to the production of organic acids, particularly lactic and acetic acids, which effectively inhibited the growth of EPEC and other pathogenic bacteria. Additionally, their robust growth and acid production capabilities under simulated gastrointestinal conditions suggest their potential in vivo efficacy. These findings highlight ZY25 and ZY35 as promising natural alternatives to conventional antibiotics for managing EPEC-induced diarrhea in cats, warranting further in vivo trials to validate their probiotic benefits and explore their commercial application in the veterinary field.


    antibacterial activity, gut microbiota, feline-derived lactic acid bacteria, Escherichia coli (E. coli), enteropathogens, biofilm inhibition, pathogen colonization, antimicrobial peptides, competitive exclusion, adhesion capacity, gastrointestinal health, probiotic mechanisms, Lactobacillus spp., cat microbiome, zoonotic bacteria, in vitro analysis, antimicrobial resistance, host-pathogen interactions, bacterial growth inhibition, immune modulation, cytokine response, intestinal epithelial cells, symbiotic balance

    #probiotics #antibacterialactivity #gutmicrobiota #lacticacidbacteria #felineprobiotics #Ecoli #enteropathogens #biofilminhibition #pathogencolonization #antimicrobialpeptides #competitiveexclusion #adhesioncapacity #gastrointestinalhealth #probioticmechanisms #Lactobacillus #catmicrobiome #zoonoticbacteria #invitroanalysis #antimicrobialresistance #hostpathogeninteractions #bacterialgrowthinhibition #immunemodulation #cytokineresponse #intestinalhealth #microbiomerestoration

    at No comments:

    November 26, 2024

    Viking settlers of the Faroe Islands

    Genetic evidence points to distinct Viking settlers of the Faroe Islands and Iceland


    Geneticists have studied the distribution of Y-chromosome haplogroups on the Faroe Islands, known to have been colonized by Vikings around the year 900 CE, and compared these to distributions of haplogroups in today's Scandinavia.

    They showed with novel analysis methods that the haplotype distribution in the Faroe Islands most closely resembled that in Norway and Denmark, and to a lesser extent that in Sweden, but differed from that in Iceland. They concluded that a band of Viking men from all over Scandinavia colonized the Faroe Islands, which differed in their geographical origin and genetic make-up from those who settled Iceland.

    The ancient Vikings certainly had the travel bug. Between the late eighth century and approximately 1050 CE, they roamed the Atlantic in their longships all the way to Newfoundland, Labrador, and Greenland, as well as exploring the Mediterranean and continental Eurasia.

    Among the places the Vikings are known to have settled were the Faroe Islands, an archipelago of 18 islands in the North Atlantic. They probably weren't the first to do so: Archaeologists have found evidence that these islands had been inhabited since approximately 300 CE, possibly by Celtic monks or others from the British Isles. But according to the Færeyinga Saga, written about 1200, a Viking chief called Grímur Kamban settled in the Faroe Islands between approximately 872 and 930 CE.

    But where in Scandinavia did Grímur and his followers come from?

    "Here we provide strong evidence that the Faroe Islands were colonized by a diverse group of male settlers from multiple Scandinavian populations," said Dr. Christopher Tillquist, an associate professor at the University of Louisville in Kentucky and the lead author of a new study published in Frontiers in Genetics.

    Tillquist's co-authors were Dr. Allison Mann from the University of Wyoming and Dr. Eyðfinn Magnussen from the University of the Faroe Islands.

    The scientists determined the genotype at 12 "short tandem repeat" (STR) loci on the Y-chromosome of 139 men from the Faroese islands of Borðoy, Streymoy, and Suðuroy. They assigned each man to the most likely haplogroup, each of which has different known distribution across today's Europe.

    The researchers compared the distribution of genotypes to those found in 412 men from Norway, Sweden, Denmark, Iceland, and Ireland. This allowed them to reconstruct the source population of the Viking population founders.

    Advanced analyses showed that the range of Faroese samples resembled the range of genotypes from broader Scandinavian, whereas the Icelandic genotypes were distinct.

    The authors also developed a powerful innovative genetic method, called "Mutational Distance from Modal Haplotype" to analyze variation in SNPs (single-nucleotide polymorphisms) within the STRs. This allowed them to reveal a "founder effect"—traces of random loss of diversity during historical colonization by a small number of people—persisting in the genetic make-up of today's Faroese and Icelandic male populations.

    "Scientists have long assumed that the Faroe Islands and Iceland were both settled by similar Norse people. Yet our novel analysis has shown that these islands were founded by men from different gene pools within Scandinavia," said Tillquist.

    "One group, diverse in their Scandinavian origins, established themselves in the Faroe Islands, while another and more genetically divergent band of Vikings colonized Iceland. They have separate genetic signatures that persist to this day."

    "There doesn't seem to have been any interbreeding afterwards between these two populations, despite their geographic proximity. Our results demonstrate that Viking expansion into the North Atlantic was more complex than previously thought."

    "Each longship that set sail for these distant islands carried not just Vikings, but distinct genetic legacies. We can now trace these separate journeys of conquest and settlement, revealing a more nuanced story of Viking exploration than told by the history books."

    Vikings, Faroe Islands, Norse settlers, Viking history, Viking Age, Faroese culture, Norse expansion, seafaring, Viking heritage, Norse mythology, Faroese archaeology, medieval Scandinavia, Viking trade, Norse farming, Viking longships, Faroese settlement, Norse exploration, Viking artifacts, Faroe Islands geography, Norse language, Viking influence, Faroe Islands history, Nordic heritage, Viking society, medieval Norse.

    #Vikings, #FaroeIslands, #NorseSettlers, #VikingAge, #FaroeseCulture, #NorseExpansion, #Seafaring, #VikingHeritage, #NorseMythology, #FaroeseArchaeology, #ScandinavianHistory, #VikingTrade, #NorseFarming, #VikingLongships, #FaroeseSettlement, #NorseExploration, #VikingArtifacts, #FaroeGeography, #NorseLanguage, #VikingInfluence, #NordicHeritage, #FaroeHistory, #VikingSociety, #MedievalNorse, #NorseLegacy.

    International Conference on Genetics and Genomics of Diseases 




    For Enquiries: genetics@healthcarek.com 

    Get Connected Here 
    --------------------------------- 
    --------------------------------- 
    at No comments:

    November 25, 2024

    Cancer Genomics Research

    Cancer Genomics Research


    The Importance of Cancer Genomics Research

    The study of cancer genomes (all the DNA in cancer cells) has revealed the mind-boggling complexity of genomic changes that drive cancer growth and survival. This knowledge has greatly expanded our understanding of how cancer develops and progresses. Ultimately, it has led to new ways of diagnosing and treating cancer, as well as new ways of identifying those at high risk of developing cancer. In short, genomics research has changed the way we see cancer.

    Large-scale research projects such as The Cancer Genome Atlas (TCGA), and its pediatric counterpart Therapeutically Applicable Research to Generate Effective Treatments (TARGET), have surveyed and cataloged the genomic changes in multiple types of cancer. The discovery of novel cancer-associated genetic changes has led to an explosion of therapies that target specific changes and tests that identify patients whose cancers harbor those changes—an approach to cancer treatment known as precision medicine.

    Genomics studies have also revealed unexpected genetic similarities across different types of tumors, shifting the way we define cancer. Treatments are typically given based on the tumor’s location in the body, such as the brain or lung. But a handful of new cancer drugs are used to treat tumors with specific genetic or molecular features, no matter where in the body the cancer started growing.

    On the other hand, genomics research has also shown how different cancers can be, even when they started growing in the same organ. This finding has helped explain why some subtypes of tumors grow at different rates and why cancer treatments don’t work the same way in every patient.

    NCI plays a key role in large-scale genomics research by facilitating collaborations between scientists of different disciplines, providing funding, and providing access to state-of-the-art technologies, data sets, and other resources.

    In addition to genomics, cancer researchers are also creating large-scale catalogs of other kinds of molecular changes in cancer, including epigenomic, transcriptomic, proteomic, and metabolomic changes. These studies have further expanded our understanding of cancer and have led to new ideas for treating it.

    The results of these projects illustrate the wide-ranging variation of genetic and molecular changes in cancer, and provide a better understanding of what causes cancer at the molecular level.



    Selected NCI Activities in Cancer Genomics Research

    Exploring the genetic and molecular foundations of cancer is a vital part of NCI’s research efforts. The following list highlights NCI-led or NCI-funded activities related to genomics and other “omics” research.

    Building Genomics Data Sets

    Large-scale research projects use omics technologies to catalog the molecular changes in multiple types of cancer.The Human Tumor Atlas Network is constructing 3-dimensional atlases of human cancers as they change over time, including single-cell analyses of genomics, epigenomics, and transcriptomics. The network is generating adult and pediatric atlases and strives to include minority and underserved patients across cancer types and stages of disease.

    The Clinical Proteomic Tumor Analysis Consortium is using large-scale proteomic and genomic analyses, or proteogenomics, to characterize multiple types of cancer. The consortium is also using proteogenomics to answer questions about toxicity and resistance in clinical trials of new cancer drugs.
    NCI leads several projects to molecularly characterize large collections of cancers as well as collecting a rich profile of clinical features and outcomes. The Genomic Data Analysis Network develops cutting-edge computational tools and performs an initial analysis of the data from these projects, providing a launching pad for a broad community of researchers.

    The Participant Engagement and Cancer Genome Sequencing (PE-CGS) Research Network Exit Disclaimer, a Cancer Moonshot initiative, is using direct participant engagement approaches to promote cancer genome sequencing programs for rare cancers, highly lethal cancers, cancers that occur at an early age, cancers that disproportionately affect certain populations, and cancers that are prevalent in understudied populations.
     
    The Childhood Cancer Data Initiative (CCDI) is gathering clinical care and research data—including genomics data—from children, teens, and young adults with cancer. CCDI has also created infrastructure to share this data with researchers to help them learn faster and on a larger scale than is possible for any single institution.

    Connecting Genomics Data to Cancer Biology

    NCI conducts and supports research to find out how genetic and other molecular changes in cancer cells affect cancer development, cancer progression, and treatment response.


    The Cancer Target Discovery and Development Program is a network of research centers devoted to studying how molecular changes found in cancers affect cancer cells and explore opportunities to target those changes with new therapies.

    The Epidemiology and Genomics Research Program supports research in human populations to understand genomic determinants of cancer occurrence and cancer outcomes, and translate findings to clinical and public health interventions.

    NCI researchers use molecular and genomic profiling to study the causes of cancer. This enables a better understanding of how normal cells transform into cancer cells, and to pinpoint internal processes and external exposures associated with specific molecular or genomic subtypes of cancer.

    The Cancer Systems Biology Consortium integrates and analyzes large omics data sets to get a birds-eye view of the molecular changes in cancer, how these changes interact with one another, and how they change over time and space.

    Using Genomics in Clinical Trials

    NCI clinical trials are among the first to use genomic testing to match patients to cancer treatments based on the genetic changes in their tumors.ComboMATCH is a group of clinical trials that are using genetic testing to find genetic changes in patients’ cancers and potentially match them to combinations of cancer treatments that target those genetic changes.

    ALCHEMIST is a set of precision medicine lung cancer trials designed to evaluate whether adding targeted therapy based on patients' tumor genetics can help prevent lung cancer from returning after surgery.

    NCI-COG Pediatric MATCH is an international cancer treatment clinical trial for children, teens, and young adults that is testing the use of precision medicine for childhood cancers.

    at No comments:

    November 23, 2024

    Reproductive genetic carrier screening

    Reproductive genetic carrier screening: A tool for reproductive decision-making


    Couple-based reproductive genetic carrier screening is an accessible and effective method of reproductive decision making, according to a recent study published in the New England Journal of Medicine.1

    Childhood-onset autosomal recessive and X-linked conditions have been linked to over 2500 genes, with most parents carrying these genes only learning about their carrier status after childbirth. Reproductive decision making may be improved through the use of reproductive genetic carrier screening.

    The availability of reproductive genetic carrier screening has been improved by commercial providers, but health care professionals must remain aware about how to best offer screening and how their patients may view this option.

    This critical information has been detailed by the Australian Reproductive Genetic Carrier Screening Project.2 The project included over 10,000 reproductive couples and provided reproductive genetic carrier screening for at least 1281 genes.

    Investigators conducted a study to evaluate screening uptake, the rate of couples with an increased risk of having an affected child, reproductive decisions, psychosocial outcomes, and screening acceptability.1 Participating health care centers offered free reproductive genetic carrier screening to their patients.

    Reproductive couple included 2 individuals of the opposite sex who were the genetic parents in pregnancy. Those in the analysis were aged 18 years or older and planning to conceive or at under 10 weeks’ gestation. Demographic data was obtained from participants at enrollment.

    The panel covered 1300 genes linked to 750 childhood-onset autosomal recessive or X-linked conditions. Most of these conditions had available treatment, but this treatment was often limited in availability or burdensome. After revision, 1281 genes were included in the final panel.

    Autosomal genes were assessed in male and female partners while X-linked genes were only assessed in female partners. Exome sequencing and a targeted gene panel were used during testing.

    Increased risks were determined based on the presence of a pathogenic or likely pathogenic variant in the same autosomal gene across both partners and a pathogenic or likely pathogenic variant in an X-linked gene for the female partner. When these criteria were not met, the odds were considered “low chance.”

    Participants with an increased risk of having an offspring with a genetic condition received genetic counseling, with referral to a subspecialist given when necessary. During genetic counseling, participants were told about reproductive options available through the study.

    Reproductive genetic carrier screening was given to 9107 couples. Couples who were invited to participate but did not undergo screening were more often residing in the most socioeconomic disadvantaged area, had a lower education level, and had 2 or more children.

    Low odds of having a child with a genetic condition were reported in 96.1% of couples who received reproductive genetic carrier screening, while new identification of an increased chance was reported in 1.9% and knowledge prior to screening in 2%. Of the 180 couples with knowledge prior to the screening, 4 learned about an increased risk of a different condition.

    Having previously been assumed to have the same level of risk as the general population before screening was reported in 78.3% of couples with a newly identified risk. The remaining 21.7% had family history or consanguinity leading them to be considered at an increased risk.

    Of newly identified couples with current pregnancy when receiving the result, 64% had the fetus genetically tested, with 24 of these 29 couples receiving normal test results. In impacted pregnancy was found in 5, with 4 deciding to terminate the pregnancy.

    Sixteen of the newly identified couples did not elect to receive testing. Of these, 1 had a miscarriage, 2 had babies with a condition identified in genetic screening after birth, 4 had unaffected babies, 4 had ongoing pregnancy during the assessment, and 5 had unknown outcomes.

    These results highlighted the efficacy and availability of providing reproductive genetic carrier screening to a diverse population. Investigators noted reproductive genetic carrier screening is acceptable for patients and providers when delivered using this method.

    carrier screening, genetic testing, reproductive health, inherited disorders, preconception care, prenatal testing, family planning, genetic counseling, rare diseases, autosomal recessive, X-linked disorders, carrier status, cystic fibrosis, thalassemia, Tay-Sachs disease, sickle cell anemia, genetic risk, next-generation sequencing, personalized medicine

    #CarrierScreening, #GeneticTesting, #ReproductiveHealth, #InheritedDisorders, #PreconceptionCare, #PrenatalTesting, #FamilyPlanning, #GeneticCounseling, #RareDiseases, #AutosomalRecessive, #XLinkedDisorders, #CarrierStatus, #CysticFibrosis, #Thalassemia, #TaySachsDisease, #SickleCellAnemia, #GeneticRisk, #NGS, #PersonalizedMedicine, #ReproductiveGenetics

    International Conference on Genetics and Genomics of Diseases 




    For Enquiries: genetics@healthcarek.com 

    Get Connected Here 
    --------------------------------- 
    --------------------------------- 
    at No comments:

    November 22, 2024

    Genetic Discovery

    Novel method goes 'below the limit of quantification' to enhance genetic discovery


    Research scientist Yosuke Tanigawa and Professor Manolis Kellis at MIT Computer Science and Artificial Intelligence Laboratory (CSAIL) have developed a novel methodology in human genetics to address an often-overlooked problem: how to handle clinical measurements that fall "below the limit of quantification" (BLQ).

    Published in the American Journal of Human Genetics, their new approach, "hypometric genetics," utilizes these typically discarded measurements to enhance genetic discovery, with significant implications for personalized genomic medicine and drug development.

    Imagine trying to weigh a feather on a standard bathroom scale. The scale might detect that something is there. However, it will not give an exact measurement value because the feather is too light to be weighed accurately, falling below the scale's sensitivity.

    In scientific research, measurements that fall below the calibrated range for reliable quantification of a measurement device are marked as BLQ. Traditionally, scientists often discard these data points to ensure that their analysis is not affected by unreliable information. However, the new approach shows that such quality control information can still provide valuable genetic insights.

    "We often treat below-quantification-limit data points as missing or uninformative, but our approach shows that these flags hold critical information for genetic analysis," says Kellis, senior and co-corresponding author of the study.

    "By leveraging both the flags and standard quantitative traits, we not only rescue discarded data but also improve our ability to discover genetic associations, revealing key biological insights that would otherwise be missed."

    The researchers analyzed more than 220,000 de-identified participants in UK Biobank and found that genetic factors influence the presence or absence of BLQ flags for specific lipid molecules in the blood.

    By integrating information from BLQ flags with conventional quantitative data, they identified genetic associations with 2.8 times as many candidate genes as the standard approach, benefiting especially from rare genetic variants with large effects. This work lays a foundation for discovering new trait-related genes and could accelerate therapeutic target discovery.

    "Our approach builds on the previous work focusing on individuals with extreme trait values," says Tanigawa, the lead and co-corresponding author of the study. "Instead of recruiting individuals with extreme trait values, we used BLQ—as an indicator of extremely low measurement values—and demonstrated that it offers valuable information for genetic discovery."

    The study has attracted substantial attention. "The work by Tanigawa resolved an overlooked but important issue of how to assess the 'missing' clinical values in human genomics," says Yukinori Okada, professor of genome informatics at the Graduate School of Medicine at the University of Tokyo.

    "By showing how binarized BLQ successfully empowers genotype-phenotype associations, their work should be valuable for implementing personalized genomic medicine."

    Tanigawa envisions that this technique could aid pharmaceutical companies in translating genetic findings to prioritize therapeutic targets more effectively.

    "Human genetic evidence increases the success rate of therapeutic development, but genetic discovery typically requires a large number of individuals," he explains. "Our results indicate that tapping into typically discarded information can maximize findings, making it a cost-effective strategy."

    Tanigawa also notes that he and his colleagues intend to apply the methodology to other biological data types, such as proteomics and RNA expression, potentially leading to even more genetic insights.

    genomics, epigenetics, CRISPR technology, genome sequencing, genetic mutations, hereditary diseases, gene editing, DNA repair, gene expression, precision medicine, bioinformatics, synthetic biology, genetic engineering, population genetics, molecular genetics, gene therapy, functional genomics, pharmacogenomics, genetic testing, evolutionary genetics.

    #GeneticDiscovery, #Genomics, #CRISPR, #GeneTherapy, #PrecisionMedicine, #GeneEditing, #Epigenetics, #GenomeSequencing, #GeneticMutations, #HereditaryDiseases, #SyntheticBiology, #Bioinformatics, #DNARepair, #MolecularGenetics, #FunctionalGenomics, #Pharmacogenomics, #GeneticTesting, #PopulationGenetics, #GeneticEngineering, #EvolutionaryGenetics

    International Conference on Genetics and Genomics of Diseases 




    For Enquiries: genetics@healthcarek.com 

    Get Connected Here 
    --------------------------------- 
    --------------------------------- 
    at No comments:
    Subscribe to: Posts (Atom)

    Wonder Drug

    Scientists Finally Crack 60-Year Mystery Behind Diabetes ‘Wonder Drug’ Metformin The antidiabetic drug metformin, widely prescribed for mana...

    • Fruitful innovation
      Fruitful innovation: Transforming watermelon genetics with advanced base editors The development of new adenine base editors (ABE) and adeni...
    • Genetic factors with clinical trial stoppage
      Genetic factors associated with reasons for clinical trial stoppage Many drug discovery projects are started but few progress fully through ...
    • Asexual Ants maintain Genetic Diversity
      How do asexual ants maintain genetic diversity? It’s commonly accepted wisdom that genetic diversity is vital for the survival of a species...