American Type Research Articles

Luteibacter mycovicinus sp. nov., a yellow-pigmented gammaproteobacterium found as an endohyphal symbiont of endophytic Ascomycota.

We isolated and described a yellow-pigmented strain of bacteria (strain 9143T), originally characterized as an endohyphal inhabitant of an endophytic fungus in the Ascomycota. Although the full-length sequence of its 16S rRNA gene displays 99 % similarity to Luteibacter pinisoli, genomic hybridization demonstrated <30 % genomic similarity between 9143T and its closest named relatives, further supported by average nucleotide identity results. This and related endohyphal strains form a well-supported clade separate from L. pinisoli and other validly named species including the most closely related Luteibacter rhizovicinus. The name Luteibacter mycovicinus sp. nov. is proposed, with type strain 9143T (isolate DBL433), for which a genome has been sequenced and is publicly available from the American Type Culture Collection (ATCC TSD-257T) and from the Leibniz Institute DSMZ (DSM 112764T). The type strain reliably forms yellow colonies across diverse media and growth conditions (lysogeny broth agar, King's Medium B, potato dextrose agar, trypticase soy agar and Reasoner's 2A (R2A) agar). It forms colonies readily at 27 °C on agar with a pH of 6-8, and on salt (NaCl) concentrations up to 2 %. It lacks the ability to utilize sulphate as a sulphur source and thus only forms colonies on minimal media if supplemented with alternative sulphur sources. It is catalase-positive and oxidase-negative. Although it exhibits a single polar flagellum, motility was only clearly visible on R2A agar. Its host range and close relatives, which share the endohyphal lifestyle, are discussed.

Influence of Five Different Commercially Available Mouthwashes on the Growth of Candida albicans Adhered to Customized Prefabricated Heat-Cured Denture Base Acrylic Resin Sheets: An In Vitro Study.

Background The purpose of this in vitro investigation was to evaluate the impact of five distinct commercial mouthwashes on the development of Candida albicans that had been adhered to heat-cured acrylic resin sheets. Methods This in vitro investigation was carried out at the MES Medical College's Microbiology Department in Perinthalmanna, Kerala, India. A total of 72 heat-cured acrylic resin sheets, size 10 × 10 × 2 mm, were fabricated. After disinfection, all 72 acrylic sheets were placed in a flask containing a suspension of the standard strain of Candida species (American Type Culture Collection) and incubated at 37ºC for 24 hours. Then, the acrylic sheets were randomly divided into six groups, with each group containing 12 acrylic sheets. Group 1 was the control group to which no mouthwash was added. In group 2, Colgate Plax was added. In group 3, Hiora Himalaya was added. In group 4, Oral B was added. In group 5, Listerine was added. In group 6, Pepsodent was added. Colony-forming units (CFUs) were assessed using a colony counter every six, 24, 48, and 120 hours. After obtaining the pH and CFU of all 72 specimens, software known as the Statistical Package for Social Sciences (SPSS) (IBM Corp., Armonk, NY) was used to analyze the data. Results Candida albicans adhered to heat-cured denture base acrylic resin sheets differed significantly in response to commercially available mouthwashes (Oral B, Colgate Plax, and Pepsodent) and non-commercial mouthwashes (Hiora Himalaya and Listerine) that contained cetylpyridinium chloride. Conclusions Compared to other mouthwashes that do not contain cetylpyridinium chloride (Listerine and Hiora Himalaya), mouthwashes with cetylpyridinium chloride as the active ingredient (Oral B, Pepsodent, and Colgate Plax) have shown good antifungal properties against the adhering Candida albicans on denture base resin.

Metabolic Comparison and Molecular Networking of Antimicrobials in Streptomyces Species.

Streptomyces are well-known for producing bioactive secondary metabolites, with numerous antimicrobials essential to fight against infectious diseases. Globally, multidrug-resistant (MDR) microorganisms significantly challenge human and veterinary diseases. To tackle this issue, there is an urgent need for alternative antimicrobials. In the search for potent agents, we have isolated four Streptomyces species PC1, BT1, BT2, and BT3 from soils collected from various geographical regions of the Himalayan country Nepal, which were then identified based on morphology and 16S rRNA gene sequencing. The relationship of soil microbes with different Streptomyces species has been shown in phylogenetic trees. Antimicrobial potency of isolates was carried out against Staphylococcus aureus American Type Culture Collection (ATCC) 43300, Shigella sonnei ATCC 25931, Salmonella typhi ATCC 14028, Klebsiella pneumoniae ATCC 700603, and Escherichia coli ATCC 25922. Among them, Streptomyces species PC1 showed the highest zone of inhibition against tested pathogens. Furthermore, ethyl acetate extracts of shake flask fermentation of these Streptomyces strains were subjected to liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis for their metabolic comparison and Global Natural Products Social Molecular Networking (GNPS) web-based molecular networking. We found very similar metabolite composition in four strains, despite their geographical variation. In addition, we have identified thirty-seven metabolites using LC-MS/MS analysis, with the majority belonging to the diketopiperazine class. Among these, to the best of our knowledge, four metabolites, namely cyclo-(Ile-Ser), 2-n-hexyl-5-n-propylresorcinol, 3-[(6-methylpyrazin-2-yl) methyl]-1H-indole, and cyclo-(d-Leu-l-Trp), were detected for the first time in Streptomyces species. Besides these, other 23 metabolites including surfactin B, surfactin C, surfactin D, and valinomycin were identified with the help of GNPS-based molecular networking.

In vitro evaluation of the binding activity of novel mouse IgG1 opsonic monoclonal antibodies to Mycobacterium tuberculosis and other selected mycobacterial species

Antimicrobial resistance alongside other challenges in tuberculosis (TB) therapeutics have stirred renewed interest in host-directed interventions, including the role of antibodies as adjunct therapeutic agents. This study assessed the binding efficacy of two novel IgG1 opsonic monoclonal antibodies (MABs; GG9 & JG7) at 5, 10, and 25 µg/mL to live cultures of Mycobacterium tuberculosis, M. avium, M. bovis, M. fortuitum, M. intracellulare, and M. smegmatis American Type Culture Collection laboratory reference strains, as well as clinical susceptible, multi-drug resistant, and extensively drug resistant M. tuberculosis strains using indirect enzyme-linked immunosorbent assays. These three MAB concentrations were selected from a range of concentrations used in previous optimization (binding and functional) assays. Both MABs bound to all mycobacterial species and sub-types tested, albeit to varying degrees. Statistically significant differences in MAB binding activity were observed when comparing the highest and lowest MAB concentrations (p < 0.05) for both MABs GG9 and JG7, irrespective of the M. tuberculosis resistance profile. Binding affinity increased with an increase in MAB concentration, and optimal binding was observed at 25 µg/mL. JG7 showed better binding activity than GG9. Both MABs also bound to five MOTT species, albeit at varied levels. This non-selective binding to different mycobacterial species suggests a potential role for GG9 and JG7 as adjunctive agents in anti-TB chemotherapy with the aim to enhance bacterial killing.

Pharmaceutical Properties and Phytochemical Profile of Extract Derived from Purple Leaf Graptophyllum pictum (L.) Griff.

Graptophyllum pictum (L.) Griff is a medicinal shrub belonging to the Acanthaceae family and is traditionally used to treat various diseases. Therefore, this study aimed to evaluate the pharmaceutical properties and phytochemical profiles of the methanolic extract of G. pictum. G. pictum leaves was extracted using methanol. Antioxidant, cytotoxic on Michigan Cancer Foundation-7 (MCF-7) and HepG2, antidiabetic, and antibacterial properties were evaluated in vitro. Chemical profile of the extract was identified through qualitative (for phytochemicals), quantitative (for phenolic and flavonoid content), and gas chromatography-mass spectrometry (GC-MS) analysis. The results showed that the extract had potent antioxidant activity against 2,2-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) and 2,2-diphenyl-1-picrylhydrazyl radicals with IC50 values of 49.00 ± 3.20 µg/mL and 70.18 ± 3.27 µg/mL, respectively. It also exhibited cytotoxic effects on human breast (MCF-7) and liver (HepG2) carcinoma cells with growth inhibition percentages of 74.29 ± 1.53% and 64.90 ± 1.94%, respectively. The antidiabetic assay showed that the extract had inhibitory effects on α-glucosidase activity with IC50 value 194.59 ± 15.59 µg/mL, indicating its potential to be developed as an antidiabetic agent. Furthermore, it had antibacterial properties against four test strains, and the highest activity was found against Bacillus subtilis American Type Culture Collection 19659, with minimum inhibitory concentration and minimum bactericidal concentration values of 625 µg/mL and 1250 µg/mL, respectively. Phytochemical tests indicated the presence of alkaloids, flavonoid and terpenoids in the extract, with total phenolic content and total flavonoid content of 41.17 ± 2.38 mg gallic acid equivalents/g and 26.52 ± 0.61 mg quercetin equivalent/g, respectively. GC-MS analysis revealed that it contained several active compounds, including eicosane, 2,4-Di-tert-butylphenol, hentriacontane, tetracosane, octacosane, sulfurous acid, 2-methylhexacosane, docosane, heneicosane, 1-propene-1,2,3-tricarboxylic acid, tributyl ester, and pentacosane. The extract derived from G. pictum leaves was a potential source of therapeutic compounds, particularly for antioxidant, antidiabetic, anticancer, and antibacterial agents.

The Role of Graphene and BiodentineTM on Proliferation and Odontoblastic Differentiation of Pulp Stem Cells

Objective: The need for new biomaterials with biocompatibility, mechanical, and antimicrobial properties continues in regenerative endodontic clinical applications in dentistry. Biodentine™ is successfully used in vital pulp therapies and has regenerative effects. Additionally, graphene, which possesses good physicochemical, mechanical, and biological properties, promotes cellular biomineralization and osteogenic differentiation in dentistry, along with its neuroregenerative effect. This study aims to investigate the effects of graphene and Biodentine™ on cell proliferation, oxidative stress, and odontogenic differentiation in human dental pulp stem cells (hDPSCs). Methods: Cryopreserved hDPSCs purchased from American Type Culture Collection (ATCC) were used in our study (Cat No: PT-5025). hDPSCs were seeded into the E-plate wells and subsequently four different doses of graphene (12.5, 25, 50 and 100 µg/ml) and Biodentine™ (2, 4, 8 and 16 µg/ml) were added. Results of MTT, total antioxidant capacity (TAC), total oxidant status (TOS) and alkaline phosphatase (ALP) tests were obtained at the end of the 24th hr. 96 hr-real time cell index data were collected with xCELLigence® system. Resulting data were compared using one-way analysis of variance (ANOVA). Results: 12.5 µg/ml graphene and 2 µg/ml Biodentine™ were found to be the subgroups with the highest levels of cell proliferation and the lowest oxidative stress. Antioxidative effect was determined in all Biodentine™ doses but only in 12.5 µg/ml graphene. Odontogenic differentiation was observed in all doses of graphene and Biodentine™. Conclusion: 12.5 µg/ml graphene and 2 µg/ml Biodentine™ were observed to have positive impacts on the proliferation, oxidative stress and odontogenic differentiations of hDPSCs. Keywords: Cell proliferation. Graphene. Oxidative stress. Odontogenic differentiation. Tricalcium silicate

Abstract 5892: SCUBE3 promotes therapy resistance in triple negative breast cancer

Abstract Introduction: The development of novel targeted therapies is urgently required for improving the outcome of triple negative breast cancer (TNBC) patients. Chemotherapy is the common treatment option for TNBC. However, resistance and tumor relapse remain the major obstacles hindering the effectiveness of chemotherapeutic agents in cancer patients. Therefore, identifying genes/factors that sensitize breast cancer cells to chemotherapeutic agents could improve treatment outcome in patients. Here we report Signal peptide CUB domain EGF-like 3 (SCUBE3) genes as a novel therapeutic adjuvant that can improve the efficacy of doxorubicin, a chemotherapeutic agent commonly used in treating breast cancer patients. Methods: Unbiased high-throughput loss of function genomic screen was performed on breast cancer cells in the presence or absence of doxorubicin. Breast cancer cells lines (MDA-MB-231 and MDA-MB-468) were purchased from the American Type Culture Collection and cultured in standard medium. Breast cancer cells were transfected either with SCUBE3 overexpression plasmid or shRNA specific to SCUBE3. These overexpression/knockdown breast cancer cells were analyzed for cell viability, migration, invasion, colony formation, cell cycle, apoptosis assays, western blotting, and in vivo tumor xenograft study. Results: Our finding demonstrated that SCUBE3 promotes breast cancer cell progression as knockdown of SCUBE3 inhibited the ability of breast cancer cells to form colony, migrate, and invade, while overexpression of SCUBE3 promoted tumor growth in preclinical mouse models. Our results revealed that SCUBE3 mediates its protumor effects by regulating genes involved in growth and survival in the MAPK pathway, DNA damage surveillance pathway including RAD51 and FOXM1, and apoptotic pathway including Mcl-1. We demonstrated that SCUBE3 promotes tumor growth and progression by interacting with and activating several oncoproteins. Conclusions: Overall, our data suggests that SCUBE3 can be a potent therapeutic target for treating TNBC patients. Citation Format: Deepika Singh, Daisy Medina, Benjamin Onyeagucha, Rahul Mojidra, Panneerdoss Subbarayallu, Alex Taylor, Dongwen Lyu, Santosh Timilsina, Prabhakar Pitta Venkata, Saif S R Nirzhor, Shahad M. Abdulsahib, Trong Phat Do, Yidong Chen, Ratna Vadlamudi, Manjeet K. Rao. SCUBE3 promotes therapy resistance in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5892.

Abstract 3156: Unveiling the functional role of HDGFRP3 in pediatric medulloblastoma: Implications for growth, progression, and radioresistance

Abstract Introduction: Medulloblastoma (MB) is the most common malignant brain cancer in children. Although long-term survival has improved in recent years, only a modest percentage of patients survive high-risk disease. The quality of life for those who survive is substantially reduced due to the high toxicity associated with the radiation and chemotherapy. Therefore, identification of safe and potent therapeutic approaches that can sensitize the therapy response in MB without inducing toxicity will lead to more efficient treatment of MB patients. We recently discovered that targeting HDGFRP3, a protein highly expressed in MB, and can block MB growth and improve the efficacy of radiation therapy. HDGFRP3 is a multifaceted protein that plays important roles in diverse cellular processes. Our investigation sheds light on the functional role of HDGFRP3 in MB, focusing on its involvement in growth, progression, and radioresistance where we report HDGFRP3 as a novel oncogenic driver of these processes. Methods: Human MB cell lines (DAOY, D556, D425, and HDMB03) were purchased from the American Type Culture Collection and cultured in standard medium. MB cancer cells were transfected with siRNA specific to HDGFRP3. These knockdown cells were analyzed for cell viability, migration, invasion, colony formation, stemness, cell cycle, apoptosis assays for functional role assessment. Real-time qPCR and western blotting were performed to validate the downstream targets of HDGFRP3 in MB models. Results: Our findings demonstrate that HDGFRP3 functions as an oncogene in MB, as its overexpression promotes cancer growth, while its depletion results in decreased tumor growth in MB models. Furthermore, we provide evidence that HDGFRP3 knockdown results in decreased migratory potential of MB cells as well as a reduction in stemness. Interestingly, HDGFRP3 depletion improved the efficacy of standard of care radiation in multiple MB models. At present, in vivo studies are validate the oncogenic role of HDGFRP3 in preclinical animal models of MB. Conclusions: In conclusion, our data suggests HDGFRP3 facilitates MB growth, progression, stemness as well as resistance to radiation therapy. Our comprehensive investigation into HDGFRP3's functional role in MB presented here provides valuable insights into its potential as a therapeutic target and opens avenues for further research into novel treatment strategies for the pediatric brain tumors. Citation Format: Saif S. Nirzhor, Prabhakar Pitta Venkata, Shahad Abdulsahib, Panneerdoss Subbarayalu, Santosh Timilsina, Manjeet K. Rao. Unveiling the functional role of HDGFRP3 in pediatric medulloblastoma: Implications for growth, progression, and radioresistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3156.

Abstract 628: RNA binding protein Matrin3 acts as a tumor suppressor by inhibiting microtubule nucleation

Abstract Introduction: Microtubule-associated proteins (MAPs) regulate microtubule dynamics, which is critical for controlling cell division, proliferation, migration, and intracellular transport. Due to their critical role in maintaining cellular integrity, microtubules have served as potent therapeutic targets for cancer treatment. Here, we report “Matrin 3 (MATR3)” as a novel MAP that interacts with β- tubulin and γ-tubulin, which constitutes microtubule organizing center (MTOC) essential for proper spindle formation and chromosomal segregation. Methods: Breast cancer cell lines (MDA-MB-231, MDA-MB-468, SKBR3, MCF7, BT549, HMEC, and MCF10A) were purchased from the American Type Culture Collection and cultured in standard medium. Breast cancer cells were transfected either with MATR3 overexpression plasmid or siRNA specific to MATR3. These overexpression/knockdown breast cancer cells were analyzed for cell viability, migration, invasion, colony formation, cell cycle, apoptosis assays, RNA sequencing, RT-qPCR, western blotting, RNA immunoprecipitation, and in vivo tumor xenograft study. Results: Our findings show that MATR3 binds to RNA encoding MAPs including β-tubulin and γ-tubulin, and regulate their stabilization. MATR3 also interacted with β-tubulin protein. Our data demonstrate that MATR3 functions as a tumor suppressor, as its overexpression inhibits cancer growth, while its depletion increased tumor growth in vivo. Mechanistic studies confirmed that MATR3 mediates its tumor suppressor role by regulating the expression of the MTOC-associated protein-encoding gene MZT2B (Mitotic Spindle Organizing Protein 2B), which is overexpressed in cancers and promotes cancer growth. Further, loss of MATR3 expression or function results in dysregulated microtubule dynamics and uncontrolled expression of the oncogenic proteins including MZT2B. Conclusions: In conclusion, our data strongly suggests that MATR3 serves as a novel RNA-binding protein with tumor suppressor properties. This discovery holds significant importance as MATR3 becomes the second protein, following Adenomatous Polyposis Coli (APC), to be identified for its binding to both MAP RNA and protein. Citation Format: Panneerdoss Subbarayalu, Subapriya Rajamanickam, Santosh Timilsina, Saif Nirzhor, Daisy Medina, Shahad Abdulsahib, Pitta V Prabhakar, Deepika Singh, Fu-Yang Li, Pooja Yadav, Esha Reddy, Krishna Evani, Vijay Eedunuri, Trong Phat Do, Benjamin C. Onyeagucha, Tabrez A. Mohammad, Jae-Hoon Ji, Yidong Chen, Nourhan Abdelfattah, Nicholas Dybdal-Hargreaves, Li-Ju Wang, Yu-Chiao Chiu, Suryavathi Viswanadhapalli, Ratna K. Vadlamudi, Manjeet K. Rao. RNA binding protein Matrin3 acts as a tumor suppressor by inhibiting microtubule nucleation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 628.

ISOLATION, CHARACTERIZATION AND ANTIMYCOBACTERIAL POTENCY OF A FURAN DERIVATIVE FROM THE CHLOROFORM CRUDE EXTRACT OF Icacina trichantha oliv TUBER.

Spectroscopic analysis of a pure isolate (OAU 2, a furan derivative) by column chromatography from the chloroform crude extract of Icacina trichantha tuber was done using FT-IR, 1HNMR, 13CNMR, COSY and was established by literature as well as the investigation of antimycobacterial potency. This anitmycobacterial activity revealed that this isolate actually has activity against DR-TB Strain and H37RV Strain which are American type culture collection strain and multi-drug resistant strains of M. tuberculosis being the test organisms. Hence, confirming the folk believe of the use of I. trichantha tuber in the treatment of “serious cough”

Establishment of Big Bone chicken fibroblast cell bank and study of its biological characteristics

A fibroblast bank of Big Bone chicken was established using tissue adherent culture method. This cell bank included 29 embryo samples, had stocks of 147 cryogenically-preserved vials each containing 2~3Å~106 cells, and met all the cell line quality control standards established by the American Type Culture Collection (ATCC). The cells cultured in vitro showed the typical morphology of fibroblasts. The growth curve assumed the “S” shape, and the cell population doubling time (PDT) was about 30h. Cell viability was 98.1% before cryopreservation and 96.6% after recovery. All tests for microbial contamination were negative. The isoenzyme pattern was of species specificity. The frequency of diploid cells was 91%. The transfection efficiency of three fluorescent protein genes fluctuated between 10.6% and 26.5%. These results showed that the cells cultured in vitro grew well and had stable genetic properties. This research thus does not only preserve the poultry genetic resources of Big Bone chicken at the cell level, but also opens new ways for preserving important genetic resources of endangered animals in the form of somatic cells.

Pathogenomes and virulence profiles of representative big six non-O157 serogroup Shiga toxin-producing Escherichia coli.

Shiga toxin (Stx)-producing Escherichia coli (STEC) of non-O157:H7 serotypes are responsible for global and widespread human food-borne disease. Among these serogroups, O26, O45, O103, O111, O121, and O145 account for the majority of clinical infections and are colloquially referred to as the "Big Six." The "Big Six" strain panel we sequenced and analyzed in this study are reference type cultures comprised of six strains representing each of the non-O157 STEC serogroups curated and distributed by the American Type Culture Collection (ATCC) as a resource to the research community under panel number ATCC MP-9. The application of long- and short-read hybrid sequencing yielded closed chromosomes and a total of 14 plasmids of diverse functions. Through high-resolution comparative phylogenomics, we cataloged the shared and strain-specific virulence and resistance gene content and established the close relationship of serogroup O26 and O103 strains featuring flagellar H-type 11. Virulence phenotyping revealed statistically significant differences in the Stx-production capabilities that we found to be correlated to the strain's individual stx-status. Among the carried Stx1a, Stx2a, and Stx2d phages, the Stx2a phage is by far the most responsive upon RecA-mediated phage mobilization, and in consequence, stx2a + isolates produced the highest-level of toxin in this panel. The availability of high-quality closed genomes for this "Big Six" reference set, including carried plasmids, along with the recorded genomic virulence profiles and Stx-production phenotypes will provide a valuable foundation to further explore the plasticity in evolutionary trajectories in these emerging non-O157 STEC lineages, which are major culprits of human food-borne disease.

Machine Learning-Based Virtual Screening of Antibacterial Agents against Methicillin-Susceptible and Resistant Staphylococcus aureus.

The application of computer-aided drug discovery (CADD) approaches has enabled the discovery of new antimicrobial therapeutic agents in the past. The high prevalence of methicillin-resistantStaphylococcus aureus(MRSA) strains promoted this pathogen to a high-priority pathogen for drug development. In this sense, modern CADD techniques can be valuable tools for the search for new antimicrobial agents. We employed a combination of a series of machine learning (ML) techniques to select and evaluate potential compounds with antibacterial activity against methicillin-susceptible S. aureus (MSSA) and MRSA strains. In the present study, we describe the antibacterial activity of six compounds against MSSA and MRSA reference (American Type Culture Collection (ATCC)) strains as well as two clinical strains of MRSA. These compounds showed minimal inhibitory concentrations (MIC) in the range from 12.5 to 200 μM against the different bacterial strains evaluated. Our results constitute relevant proven ML-workflow models to distinctively screen for novel MRSA antibiotics.

Epigallocatechin Gallate (EGCG) ve Yeşil Çay (Camellia Sinensis) Yağının Çeşitli Patojenler Üzerinde İn Vitro Antimikrobiyal Etkinliğinin Değerlendirilmesi

Aim: EGCG, the polyphenolic component of Camellia sinesis catechins, and Camellia sinesis extract have broad antimicrobial activity. To investigate the invitro antibacterial and antifungal activity of the Epigallocatechin gallate and Camellia sinesis extract. Method: The present study tested the antibacterial and antifungal activity of Epigallocatechin gallate and Camellia sinesis extract against some gram-negative, gram-positive, and fungal isolates of American Type Culture Collection (ATCC). The minimum inhibitory concentration (MIC) of Epigallocatechin gallate (EGCG) and Camellia sinensis extract were determined for each test microorganism. In general, EGCG and Camellia sinensis extract results were found to be compatible. Epigallocatechin gallate and Camellia sinesis extract exhibited antibacterial and antifungal activity against all test organisms. MIC was determined visually after 16-20 hours of incubation at 37°C according to broth microdilution methods recommended by the Clinical and Laboratory Standards Institute (CLSI). Result: The Camellia sinesis extract and EGCG MIC for gram-positive bacteria was found to be lower than the MIC for gram-negative bacteria and the Camellia sinesis extract MIC for C. albicans was found to be higher than the EGCG MIC. Conclusions: However, EGCG was found to be more effective on C. albicans isolate than Camellia sinensis extract. Camellia sinesis extract Epigallocatechin gallate and Camellia sinesis extract may be a promising sources of antibacterial and antifungal for further studies.

Moraxella oculi sp. nov., isolated from a cow with infectious bovine keratoconjunctivitis.

A novel species of the genus Moraxella was isolated from an ocular swab from a cow with infectious bovine keratoconjunctivitis. 16S rRNA gene sequencing suggested this species was Moraxella bovis (99.59 % nucleotide identity). Average nucleotide identity was calculated using a draft whole genome sequence of this strain compared with type strains of closely related Moraxella species and results established that it represents a new species. The genome size was 2 006 474 nucleotides and the G+C content was 42.51 mol%. The species could not be identified by matrix assisted laser desorption/ionization-time of flight mass spectrometry using a commercial database, confirming the novelty of the strain. We propose the name Moraxella oculi sp. nov. for this new species. The type strain is Tifton1T and has been deposited into the American Type Culture Collection (TSD-373T) and the National Collection of Type Cultures (NCTC), UK Health Security Agency (NCTC 14942T).

Antioxidant and antimicrobial effect of sodium alginate nanoemulsion coating enriched with oregano essential oil (Origanum vulgare L.) and Trachyspermum ammi oil (Carum cupticum) on food pathogenic bacteria.

Today, microbial contamination in food is one of the major problems of the food industry and public health in general around the world. Foodborne illnesses, such as diarrheal diseases, kill many people around the world each year. The general objective of this study was to evaluate the antioxidant and antibacterial activity of sodium alginate nanoemulsion coating incorporated with oregano essential oil (Origanum vulgare L.) and Trachyspermum ammi oil (Carum cupticum) on Escherichia coli, and Listeria monocytogenes. To achieve this study, fresh chicken meat was used for this experiment. Listeria monocytogenes ATCC 19111 and Escherichia coli ATCC 35218 were obtained from the American Type Culture Collection (Manassas, VA, USA). After the preparation of the essential oil, the chemical composition of this essential oil was determined by using (GC-MS). The physicochemical properties of the nanoemulsion essential oil prepared were characterized and their antimicrobial activity was evaluated. The results showed that the GC-MS analysis of the volatile constituents of the Origanum vulgare essential oil compounds allowed the identification of 19 compounds representing 93.72% of the total oil. The major components detected in Origanum vulgare essential oil were pulegone (49.25%), eucalyptol (18.23%), and menthone (12.37%). About the Carum cupticum essential oil, 21 compounds representing 98.5% of the total oil were identified. The major components detected in Origanum vulgare essential oil were thymol (23.3%), p-cymene (17.5%), and γ-terpinene (16.8%). The best z-average (d.nm) is 483.4 nm (Carum cupticum essential oil + nano) followed by 470.1 nm (nanochitosan). The results of the antimicrobial test showed that the different preparations have a good inhibitory activity for the invitro growth of Escherichia coli and Listeria monocytogenes. According to the MIC and MBC results of this study, the nanoemulsion also presented a good bacteriostatic activity against the two pathogenic bacteria tested in this study.

Histopathological Association of Non-alcoholic Fatty Liver Disease With Coronary Artery Atherosclerosis Grade

Background: The association between the severity of coronary atherosclerosis and histopathologic findings in patients with non-alcoholic fatty liver disease (NAFLD) is not entirely understood. Considering the gold standard method, this study evaluates the histopathologic association between the severity of NAFLD and the grades of coronary atherosclerosis. Methods: In this descriptive-analytical study, data from 205 cadavers who were referred to an Iranian (Tabriz) forensic medicine organization between 2015 and 2017 and underwent simultaneous liver and coronary artery biopsies were examined. Finally, 168 cases were entered based on the inclusion criteria. First, pathological slides of these cadavers were extracted from the forensic medicine archive and re-examined. Then, the selected cases’ blocks were extracted from the tissue block bank, and again, after preparing a new slide, they were stained with trichrome for accurate estimation of liver fibrosis. Results: The assessment of NAFLD histological status in the studied cases revealed that 75.6% of the cases were classified as severity I, 18.4% as severity II, and 6% as severity III. Most cases with coronary atherosclerosis were classified as American Heart Association staging (AHA), type V (19.6%), and normal (19.6%). There was no statistically significant relationship between the severity of simple steatosis, steatohepatitis, and NAFLD, with coronary atherosclerosis. In subjects with higher severity of coronary atherosclerosis, the liver fibrosis rate is also higher, but no statistically significant difference was observed. Conclusion: The present study revealed no significant histopathological association between NAFLD and coronary artery atherosclerosis grade.

Investigation of proapoptotic and cytotoxic effects of 2-aminobenzothiazole on human laryngeal carcinoma cells.

In the present study, we investigated the effects of 2-aminobenzothiazole application on human laryngeal carcinoma cells. Human larynx epidermoid carcinoma (HEp-2) (ATCC® CCL-23™) cells were purchased from American Type Culture Collection (ATCC, USA). Human larynx epidermoid carcinoma HEp-2 cells were cultured in complete Dulbecco's Modified Eagle's Medium (DMEM) supplemented with fetal bovine serum (FBS) (10%) and penicillin/streptomycin (1%) in a CO2 (5%) incubator under standard cell culture conditions. 2-aminobenzothiazole was prepared, and further dilutions ranging from 3.13 to 100 μM were prepared in fresh culture DMEM. HEp-2 cells on 96 well plates were incubated with the prepared dilutions of 2-aminobenzothiazole for 24, 48, and 72 hours. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test performed cytotoxicity evaluation and viability percentages. The annexin-V staining technique detected 2-aminobenzothiazole-triggered apoptosis of HEp-2 cells. The activated caspases 3/7 on HEp-2 cells after 2-aminobenzothiazole exposure were evaluated with flow cytometric analysis. The membrane potential changing of HEp-2 cells was measured following the Muse™ MitoPotential kit manufacturer instructions. MTT cytotoxicity test results showed that the viability of human laryngeal carcinoma cells decreased with an increase in the application of 2-aminobenzothiazole for 24 hours. The highest growth inhibition by 2-aminobenzothiazole for short-term application of 24 hours was detected at the highest concentration of 2-aminobenzothiazole (100 µM). The results underline that the cytotoxic effect of 2-aminobenzothiazole is dose-dependent. Cytotoxicity test results for an application time of 48 hours showed that the cytotoxicity of 2-aminobenzothiazole is dose-dependent on HEp-2 cells. The required dose of 2-aminobenzothiazole to decrease the cell viability to 50 percent has been 9-fold augmented. Annexin-V findings showed that after exposure to IC50 concentration of 2-aminobenzothiazole for 24 hours, HEp-2 cells underwent the early apoptotic stage (25.99%) and late apoptotic (16.69%), whereas 56.93% of the treated cells were alive. Only 0.39% of 2-aminobenzothiazole treated cells were necrotic. All study results showed that 2-aminobenzothiazole triggered apoptosis on HEp-2 cells with a percentage of total apoptotic cells 42.62 compared to untreated HEp-2 cells. Caspase 3/7 activation results showed that only 0.65% of control HEp-2 cells were with activated caspase 3/7, and 99.35% live cells. The analysis data from the Muse cell analyzer revealed that the percentage of cells with intact mitochondrial membranes was 21.30 after 2-aminobenzothiazole application, and 79.9% were cells with depolarized mitochondrial membranes. It has been understood that the depolarization of the inner mitochondrial membrane has been considered a dysfunction in mitochondria as a sign of apoptosis and drug toxicity. Based on all study findings, 2-aminobenzothiazole has cytotoxicity on human laryngeal carcinoma cells in a dose and time-dependent manner. That means that it decreased viability via inducing caspase-dependent apoptosis. Consequently, it was concluded that 2-aminobenzothiazole has good potential to lead to cytotoxicity and apoptosis on human laryngeal carcinoma cells and, after deeper in vitro and in vivo investigations, can be a good candidate for designing anticancer drugs with high efficiency.

The Bear: New (stereotypical) representations of Italian Americans in contemporary television series

This article investigates the representations of Italian Americans in the successful TV series The Bear (2022–), following Ryan Calabretta-Sajder and Alan J Gravano's innovative critical framework according to which the field of Italian American cinema, TV and media studies needs to go beyond the strict notion of ethnicity and be, instead, reconsidered as an intersectional area including issues as class, gender, sexuality, power and production. The Bear portrays Carmen (‘Carmy’) Berzatto, a young top chef who returns to his hometown of Chicago to take over his family's Italian sandwich shop and who is living through an emotional breakdown because of his brother's death. The series received great praise for its ground-breaking narration style and nonconformist characters. Still, while the narrative arc of the series does not focus on the Italian roots of the protagonist and his family, many of the climatic moments recurrently exploit the commonplace repertoire of Italian American type: loud, quickly bursting into fits of rage, family obsessed, food enthusiast, somehow involved in dirty business. This analysis will explore how, in this contemporary depiction of Italian Americans, the author paradoxically departs and at the same time relies on the stereotypical dominant cliche usually employed for these characters. As a result, a transitional narration emerges, portraying figures precariously balancing between innovation and conservatism. With a discursive approach, this article will examine how The Bear follows this contradictory pattern, seemingly dismissing crystalized representational myths of Italian descent in the USA and simultaneously validating them.

Building in vitro tools for livestock genomics: chromosomal variation within the PK15 cell line

BackgroundCultured porcine cell lines are powerful tools for functional genomics and in vitro phenotypic testing of candidate causal variants. However, to be utilised for genomic or variant interrogation assays, the genome sequence and structure of cultured cell lines must be realised. In this work, we called variants and used read coverage in combination with within-sample allele frequency to detect potential aneuploidy in two immortalised porcine kidney epithelial (PK15) cell lines and in a pig embryonic fibroblast line.ResultsWe compared two PK15 cultured cells samples: a new American Type Culture Collection (ATCC) sample and one that has been utilised and passaged within the laboratory for an extended period (> 10 years). Read coverage and within-sample allele frequencies showed that several chromosomes are fully or partially aneuploid in both PK15 lines, including potential trisomy of chromosome 4 and tetrasomy of chromosome 17. The older PK15 line showed evidence of additional structural variation and potentially clonal variation. By comparison, the pig embryonic fibroblast line was free from the gross aneuploidies seen in the PK15s.ConclusionsOur results show that the PK15 cell lines examined have aneuploidies and complex structural variants in their genomes. We propose that screening for aneuploidy should be considered for cell lines, and discuss implications for livestock genomics.