High-throughput screening of food additives with synergistic effects on high hydrostatic pressure inactivation of budding yeast

ABSTRACTThe synergistic effects of high hydrostatic pressure (HHP), mild heating, and amino acids on the germination ofClostridium sporogenesspores were examined by determining the number of surviving spores that returned to vegetative growth after pasteurization following these treatments. Pressurization at 200 MPa at a temperature higher than 40°C and treatment with some of the 19l-amino acids at 10 mM or higher synergistically facilitated germination. When one of these factors was omitted, the level of germination was insignificant. Pressures of 100 and 400 MPa were less effective than 200 MPa. The spores were effectively inactivated by between 1.8 and 4.8 logs by pasteurization at 80°C after pressurization at 200 MPa at 45°C for 120 min with one of the amino acids with moderate hydrophobicity, such as Leu, Phe, Cys Met, Ala, Gly, or Ser. However, other amino acids showed poor inactivation effects of less than 0.9 logs. Spores in solutions containing 80 mM of either Leu, Phe, Cys, Met, Ala, Gly, or Ser were successfully inactivated by pasteurization by more than 5.4 logs after pressurization at 200 MPa at 70°C for 15 to 120 min. Ala and Met reduced the spore viability by 2.8 and 1.8 logs, respectively, by pasteurization at a concentration of 1 mM under 200 MPa at 70°C. These results indicate that germination of the spores is facilitated by a combination of high hydrostatic pressure, mild heating, and amino acids.

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Synergistic effect of high hydrostatic pressure treatment and food additives on the inactivation of Salmonella enteritidis

Food Control ◽

10.1016/j.foodcont.2008.11.004 ◽

2009 ◽

Vol 20 (11) ◽

pp. 963-966 ◽

Cited By ~ 29

Author(s):

Hirokazu Ogihara ◽

Maya Yatuzuka ◽

Naoko Horie ◽

Soichi Furukawa ◽

Makai Yamasaki

Keyword(s):

Hydrostatic Pressure ◽

Synergistic Effect ◽

High Hydrostatic Pressure ◽

Salmonella Enteritidis ◽

Food Additives ◽

Pressure Treatment ◽

High Hydrostatic Pressure Treatment

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Genetically encoded multimode reporter of adaptor protein 3 (AP-3) traffic in budding yeast

10.1101/314286 ◽

2018 ◽

Author(s):

Rachael L. Plemel ◽

Greg Odorizzi ◽

Alexey J. Merz

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Budding Yeast ◽

Agar Plate ◽

Adaptor Protein ◽

Epileptic Encephalopathy ◽

Early Endosomes ◽

Hermansky Pudlak Syndrome ◽

Functional Deficiency

SYNOPSISThe AP-3 (adaptor complex 3) mediates traffic from the late Golgi or early endosomes to late endosomal compartments. Here, a synthetic reporter is presented that allows convenient monitoring of AP-3 traffic, and direct screening or selection for mutants with defects in the pathway. The reporter can be assayed by fluorescence microscopy or in liquid or agar plate formats and is adaptable to high-throughput screening.SUMMARYAP-3 (adaptor complex 3) mediates traffic from the late Golgi or early endosomes to late endosomal compartments. In mammals, mutations in AP-3 cause Hermansky-Pudlak Syndrome type 2, cyclic neutropenias, and a form of epileptic encephalopathy. In budding yeast, AP-3 carries cargo directly from the trans-Golgi to the lysosomal vacuole. Despite the pathway’s importance and its discovery two decades ago, rapid screens and selections for AP-3 mutants have not been available. We now report GNSI, a synthetic, genetically encoded reporter that allows rapid plate-based assessment of AP-3 functional deficiency, using either chromogenic or growth phenotype readouts. This system identifies defects in both the formation and consumption of AP-3 carrier vesicles and is adaptable to high-throughput screening or selection in both plate array and liquid batch culture formats. Episomal and integrating plasmids encoding GNSI have been submitted to the Addgene repository.

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Computationally guided high-throughput design of self-assembling drug nanoparticles

10.1101/786251 ◽

2019 ◽

Author(s):

Daniel Reker ◽

Yulia Rybakova ◽

Ameya R. Kirtane ◽

Ruonan Cao ◽

Jee Won Yang ◽

...

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Ex Vivo ◽

Drug Loading ◽

Food Additives ◽

Drug Nanoparticles ◽

Self Assembling ◽

Wide Range ◽

Approved Drugs

AbstractNanoformulations are transforming our capacity to effectively deliver and treat a myriad of conditions. However, many nanoformulation approaches still suffer from high production complexity and low drug loading. One potential solution relies on harnessing co-assembly of drugs and small molecular excipients to facilitate nanoparticle formation through solvent exchange without the need for chemical synthesis, generating nanoparticles with up to 95% drug loading. However, there is currently no understanding which of the millions of possible combinations of small molecules can result in the formation of these nanoparticles. Here we report the development of a high-throughput screening platform coupled to machine learning to enable the rapid evaluation of such nanoformulations. Our platform identified 101 novel self-assembling drug nanoparticles from 2.1 million pairings derived from 788 candidate drugs with one of 2686 excipients, spanning treatments for multiple diseases and often harnessing well-known food additives, vitamins, or approved drugs as carrier materials – with potential for accelerated approval and translation. Given their long-term stability and potential for clinical impact, we further characterize novel sorafenib-glycyrrhizin and terbinafine-taurocholic acid nanoparticles ex vivo and in vivo. We anticipate that this platform could accelerate the development of safer and more efficacious nanoformulations with high drug loadings for a wide range of therapeutics.

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Enabling cost-effective screening for antimicrobials against Listeria monocytogenes in ham

Journal of Food Protection ◽

10.4315/jfp-20-435 ◽

2020 ◽

Author(s):

Cristina Resendiz-Moctezuma ◽

Shannon D Rezac ◽

Michael J Miller ◽

Matthew J Stasiewicz

Keyword(s):

Listeria Monocytogenes ◽

High Throughput ◽

High Throughput Screening ◽

Synergistic Effects ◽

Cost Effective ◽

Severe Illness ◽

Lytic Bacteriophage ◽

Cell Counts ◽

Slice Method ◽

Antimicrobial Combinations

Ready-to-eat (RTE) meat products, such as deli ham, can support the growth of Listeria monocytogenes (LM) which can cause severe illness in immunocompromised individuals. The objectives of this study were to validate a miniature ham model (MHM) against the ham slice method and screen antimicrobial combinations to control LM on ham using response surface methodology (RSM) as a time- and cost-effective high-throughput screening tool. The effect of nisin (Ni), potassium lactate sodium acetate (PLSDA), lauric arginate (LAG), lytic bacteriophage (P100), and Ɛ-polylysine (EPL) added alone, or in combination, was determined on the MHM over 12 days of storage. Results showed the MHM accurately mimics the ham slice method since no statistical differences were found (p=0.526) in the change of LM cell counts in MHM and slice counts after 12 days of storage at 4°C for treated and untreated hams. The MHM was then used to screen antimicrobial combinations using an on-face design and three center points in a central composite design. The RSM was tested using a cocktail of five LM strains isolated from foodborne disease outbreaks. Three levels of the above mentioned antimicrobials were used in combination for a total of 28 runs performed in triplicate. The change of LM cell counts were determined after 12 days of storage at 4°C. All tested antimicrobials were effective on reducing LM cell counts on ham when added alone. A significant antagonistic interaction (p=0.002) was identified by the RSM between LAG and P100, where this antimicrobial combination caused a 2.2 logCFU/g change of LM cell counts after 12 days of storage. Two interactions, between Ni and EPL (p=0.058), and Ni and P100 (p=0.068), showed possible synergistic effects against LM on the MHM. Other interactions were clearly non-significant, suggesting additive effects. In future work, the developed MHM in combination with RSM can be used as a high-throughput method to analyze novel antimicrobial treatments against LM

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High-throughput screening of high protein producer budding yeast using gel microdrop technology

10.1101/830596 ◽

2019 ◽

Author(s):

Hirotsugu Fujitani ◽

Soichiro Tsuda ◽

Tomoko Ishii ◽

Masayuki Machida

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Protein Production ◽

Budding Yeast ◽

Screening Method ◽

Screening Strategy ◽

Secreted Proteins ◽

Cell Sorter ◽

Emulsion Method ◽

Mutagenized Cell

AbstractThe need for protein production has been growing over the years in various industries. We here present a high-throughput screening strategy to isolate high producer budding yeast clones from a mutagenized cell population using gel microdrop (GMD) technology. We use a microfluidic water-in-oil (W/O) emulsion method to produce monodisperse GMDs and a microfluidic cell sorter for damage-free sorting of GMDs by fluorescently quantifying secreted proteins. As a result, this high-throughput GMD screening method effectively selects high producer clones and improves protein production up to five-fold. We speculate that this screening strategy can be applied, in principle, to select any types of high producer cells (bacterial, fungal, mammalian, etc.) which produce arbitrary target protein as it does not depend on enzymes to be produced.

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A New High-Throughput-Screening-Assay for Photoantimicrobials Based on EUCAST Revealed Unknown Photoantimicrobials in Cortinariaceae

Frontiers in Microbiology ◽

10.3389/fmicb.2021.703544 ◽

2021 ◽

Vol 12 ◽

Author(s):

Johannes Fiala ◽

Harald Schöbel ◽

Pamela Vrabl ◽

Dorothea Dietrich ◽

Fabian Hammerle ◽

...

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Best Practice ◽

Synergistic Effects ◽

Light Emitting Diode ◽

Microtiter Plate ◽

Screening Assay ◽

Innovative Strategies ◽

Awareness Campaigns ◽

High Throughput Screening Assay

Antimicrobial resistance is one of the biggest health and subsequent economic threat humanity faces. Next to massive global awareness campaigns, governments and NGOs alike stress the need for new innovative strategies to treat microbial infections. One of such innovative strategies is the photodynamic antimicrobial chemotherapy (PACT) in which the synergistic effects of photons and drugs are exploited. While many promising reports are available, PACT – and especially the drug-design part behind – is still in its infancy. Common best-practice rules, such as the EUCAST or CLSI protocols for classic antibiotics as well as high-throughput screenings, are missing, and this, in turn, hampers the identification of hit structures. Hit-like structures might come from synthetic approaches or from natural sources. They are identified via activity-guided synthesis or isolation strategies. As source for new antimicrobials, fungi are highly ranked. They share the same ecological niche with many other microbes and consequently established chemical strategies to combat with the others. Recently, in members of the Cortinariaceae, especially of the subgenus Dermocybe, photoactive metabolites were detected. To study their putative photoantimicrobial effect, a photoantimicrobial high-throughput screening (HTS) based on The European Committee on Antimicrobial Susceptibility Testing (EUCAST) was established. After validation, the established HTS was used to evaluate a sample set containing six colorful representatives from the genus Cortinarius (i.e., Cortinarius callisteus, C. rufo-olivaceus, C. traganus, C. trivialis, C. venetus, and C. xanthophyllus). The assay is built on a uniform, light-emitting diode (LED)-based light irradiation across a 96-well microtiter plate, which was achieved by a pioneering arrangement of the LEDs. The validation of the assay was accomplished with well-known photoactive drugs, so-called photosensitizers, utilizing six distinct emission wavelengths (λexc = 428, 478, 523, 598, or 640 nm) and three microbial strains (Candida albicans, Staphylococcus aureus, and Escherichia coli). Evaluating the extracts of six Cortinarius species revealed two highly promising species, i.e., C. rufo-olivaceus and C. xanthophyllus. Extracts from the latter were photoactive against the Gram-positive S. aureus (c = 7.5 μg/ml, H = 30 J/cm2, λ = 478 nm) and the fungus C. albicans (c = 75 μg/ml, H = 30 J/cm2, λ = 478 nm).

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