Culture Medium Research Articles

Multicenter study of the performance of NTM Elite agar for the detection of nontuberculous mycobacteria from patients with cystic fibrosis.

The performance of a novel selective agar was evaluated against the performance of conventional mycobacterial cultures, i.e., a combination of the mycobacterial growth indicator tube (MGIT) with Löwenstein-Jensen (LJ), for the detection of nontuberculous mycobacteria (NTM) in sputum samples from people with cystic fibrosis (pwCF). Two hundred eighty-three sputum samples (231 fresh sputum and 52 spiked sputum) from 143 pwCF were collected. They were inoculated without prior decontamination on NTM Elite agar (30°C ± 2°C for 28 days) and inoculated on both MGIT and LJ (35°C-37°C for 6-8 weeks) after N-acetyl-L-cysteine-2% sodium hydroxide decontamination. NTM were identified by Matrix-Assisted Laser Desorption Ionization/Time of Flight Mass Spectrometry and/or PCR, and whole-genome sequencing. A total of 67 NTM were recovered overall by the combination of all culture media. NTM Elite agar allowed the recovery of 65 NTM (97%), compared to 22 for the conventional MGIT and LJ media combination (32.8%), including 22 NTM for MGIT (32.8%) and 3 NTM with the LJ medium (4.5%). For Mycobacterium abscessus complex, the sensitivity of NTM Elite agar was 95% compared with a sensitivity of 30% for the conventional MGIT and LJ media combination. Overall, 17.3% of cultures on NTM Elite agar were contaminated with other micro-organisms vs 46.3% on MGIT and 77% on LJ. This study shows that the novel selective agar (NTM Elite agar) significantly outperforms the conventional MGIT and LJ media combination in terms of sensitivity, selectivity, and ease of culture, without the requirement of an L3 laboratory.IMPORTANCENontuberculous mycobacteria (NTM) are significant pulmonary pathogens in patients with pre-existing structural lung conditions such as cystic fibrosis, bronchiectasis, or chronic obstructive pulmonary disease. Mycobacterium avium complex and Mycobacterium abscessus complex (MABSC) are the most frequently isolated organisms. Compared to the recommended culture method for NTM, which combines solid and liquid culture media, NTM Elite agar enables a faster/easier diagnosis and speeds up identification and susceptibility testing as the final reading is at 28 days instead of 6-8 weeks for the conventional mycobacterial cultures. In addition, for the NTM Elite agar, no decontamination stage before inoculation is necessary, unlike the conventional mycobacterial cultures. NTM Elite agar is derived from a formulation of medium adapted to rapidly growing mycobacteria (RGM). The medium enables the growth of RGM while suppressing other flora. It is supported with published clinical data showing the benefits of this medium.

Volatile Organic Compounds Produced by Trichoderma asperellum with Antifungal Properties against Colletotrichum acutatum.

Managing plant diseases caused by phytopathogenic fungi, such as anthracnose caused by Colletotrichum species, is challenging. Different methods have been used to identify compounds with antibiotic properties. Trichoderma strains are a source of novel molecules with antifungal properties, including volatile organic compounds (VOCs), whose production is influenced by the nutrient content of the medium. In this study, we assessed the VOCs produced in dual confrontation systems performed in two culture media by Trichoderma strains (T. atroviride IMI206040, T. asperellum T1 and T3, and Trichoderma sp. T2) on Colletotrichum acutatum. We analysed the VOC profiles using gas chromatography coupled with mass spectrometry. The Luria Bertani (LB) medium stimulated the production of VOCs with antifungal properties in most systems. We identified 2-pentyl furan, dimethyl disulfide, and α-phellandrene and determined their antifungal activity in vitro. The equimolar mixture of those VOCs (250 µM ea.) resulted in 14% C. acutatum diametral growth inhibition. The infective ability and disease severity caused by the mycelia exposed to the VOCs mixture were notably diminished in strawberry leaves. Application of these VOCs as biofumigants may contribute to the management of anthracnose. LB represents a feasible strategy for identifying novel VOCs produced by Trichoderma strains with antifungal properties.

Modified culture media development using locally available fertilizers and vitamin supplements for continuous mass culture of Chaetoceros gracilis

Microalgae culture is a vital nutritional resource for hatchery development in aquaculture and has gained significant interest as a sustainable food ingredient, versatile feedstock for biofuels, with broad industrial applications. The high cost of microalgae culture media hinders mass production, necessitating the development of affordable, simpler alternatives for commercial viability. The aim of this study was to develop a modified culture media for Chaetoceros gracilis utilizing locally available fertilizers and vitamins. In the first experiment, the cell density of C. gracilis across four fertilizers combinations (T1, T2, T3 and T4) with three replicates was evaluated to identify the best performing fertilizers recipe. The significantly higher cell density was observed in T1, where a combination of urea, TSP, mineral mix, EDTA, silicate gel, NPK, compost were used, than the other treatments where one or more ingredients were missing. The growth curve showed that with 1 ml/L fertilizers combination of T1 cell density peaked (21.8×10^5 cell/ml) at day three with the death phase starting at day 04. In the second experiment, the effect of vitamin addition was evaluated with T1 considering as control (V0-without vitamin supplement) and, the Multivitamin-Zinc B and Neobion addition with T1 as treatment V1 and V2, respectively. While Neobion addition significantly increased cell density, multivitamin-Zinc B addition reduced the cell density. Finally, the potentiality of continuous culture of C. gracilis with the modified fertilizer media (V2) was evaluated. The cell density in 300 L tank culture starting from 6.6×10^5 cell/ml peaked 23.8 ×10^5 cell/ml at day two and remained relatively static until investigation finished with the supplement of V2 media two days interval. The present study suggest that development of modified fertilizer media using affordable, locally sourced ingredients for algae cultivation in the hatcheries and aquaculture industry could create an alternative to relying on pure laboratory-grade media.

Predominance of enterotoxigenic Escherichia coli among ESBL/plasmid-mediated AmpC-producing strains isolated from diarrheic foals: a public health concern

BackgroundThe upsurge of diarrheagenic E. coli pathotypes carrying extended-spectrum beta-lactamases (ESBLs)/plasmid-mediated AmpC β-lactamase (pAmpC) among animals constitutes an emerging threat for humans and animals. This study investigated the burden of ESBL-/pAmpC-producing diarrheagenic E. coli among diarrheic foals and its potential public health implications. Rectal swabs were collected from 80 diarrheic foals. These swabs were processed to isolate and identify ESBL/pAmpC-producing E. coli using a selective culture medium, biochemical tests, phenotypic identification, and molecular identification of ESBL- and pAmpC-encoding genes. Moreover, all ESBL-/pAmpC-producing E. coli isolates were examined for different virulence genes related to diarrheagenic E. coli pathotypes.ResultsOut of 80 examined foals, 26 (32.5%) were confirmed as ESBL-/pAmpC-producing E. coli, of which 14 (17.5%) animals carried only ESBL-producing E. coli, whereas 12 (15%) animals possessed ESBL-pAmpC-producing E. coli. The only detected diarrheagenic pathotype was enterotoxigenic, encoded by the heat-stable enterotoxin gene (ST) with a prevalence rate of 80.8% (21/26). The ST gene was further characterized where STa, STb, and STa + STb were found in one, four, and 16 strains, respectively. Moreover, all enterotoxigenic E. coli (ETEC) isolates exhibited a multidrug-resistance pattern. The phylogenetic analysis of 3 obtained partial STb sequences revealed high genetic relatedness to ETEC isolates retrieved from humans, conferring such sequences' public health significance.ConclusionsThese findings highlight that diarrheic foals could serve as a potential reservoir for multidrug-resistant ESBL-/pAmpC-producing enterotoxigenic E. coli.

Organogenesis in a Broad Spectrum of Grape Genotypes and Agrobacterium-Mediated Transformation of the Podarok Magaracha Grapevine Cultivar.

We present data on the ability for organogenesis in 22 genotypes of grapevine and developed a direct organogenesis protocol for the cultivar Podarok Magaracha and the rootstock Kober 5BB. The protocol does not require replacement of culture media and growth regulators, and the duration is 11 weeks. The cultivation of explants occurs on modified MS medium with the addition of 2.0 mg L-1 benzyladenine and indole-3-butyric acid (0.15 mg L-1 for the rootstock Kober 5BB or 0.05 mg L-1 for the cultivar Podarok Magaracha). The direct organogenesis protocol consists of three time periods: (1) culturing explants for 2 weeks in dark conditions for meristematic bulk tissue, (2) followed by 4 weeks of cultivation in light conditions for regeneration, and (3) 5 weeks of cultivation in dark conditions for shoot elongation. Based on this protocol, conditions for the Agrobacterium-mediated transformation of the Podarok Magaracha cultivar were developed with an efficiency of 2.0% transgenic plants per 100 explants. Two stably transformed lines with integration into the genome of the pBin35SGFP plasmid construction, confirmed by Southern blotting, were obtained.

Effects of a bioengineered allogeneic cellular construct on burn-related macrophage phenotype.

Bioengineered allogeneic cellularised constructs (BACC) exert pro-healing effects in burn wounds and skew macrophage phenotype towards a predominately reparative phenotype. However, whether BACC can modulate the phenotype of dysregulated macrophages, like those present in burn wounds, is not known. To better understand the macrophage modulatory characteristics of the BACC, primary human macrophages were polarised to the M2b phenotype, an immunosuppressive phenotype relevant to burn wounds, by simultaneously exposing macrophages to polystyrene plate-coated immunoglobulin G and the endotoxin lipopolysaccharide (LPS). The resulting macrophage phenotype upregulated both inflammatory and reparative genes, and increased secretion of the M2b marker CCL1 compared to five different in vitro macrophage phenotypes. M2b macrophages were cultured with the BACC in the presence or absence of LPS to mimic infection, which is a common occurrence in burn wounds. The BACC caused up-regulation of reparative gene sets and down-regulation of pro-inflammatory gene sets, even when LPS was present in the cell culture media. Co-cultures were maintained for 1, 3, or 5 days in the presence of LPS, and by day 1 both non-activated macrophages and M2b macrophages exhibited signs of endotoxin tolerance, as demonstrated by a reduced secretion of tumour necrosis factor α (TNFα) in response to fresh LPS stimulus. The BACC was not able to prevent endotoxin tolerance, but reparative genes were upregulated in macrophages chronically exposed to LPS. These results suggest that the BACC can promote a reparative phenotype in dysregulated macrophages relevant to the pathophysiology of burns.

Adipose tissue-derived extracellular vesicles from obese mice suppressed splenocyte-mediated pancreatic cancer cell death.

Obesity is a risk factor for pancreatic cancer and negatively contributes to the immune system. However, the mechanisms by which obesity mediates these actions are still poorly understood. Recent studies have demonstrated that extracellular vesicles (EVs) are key mediators of communication between cells and may influence various aspects of cancer progression. We aim to explore the influence of EVs derived from adipose tissue of obese mice on cytokine production within the interactions between cancer cells and immune cells. We isolated EVs from the adipose tissue of both C57BL6/J mice and Ob/Ob mice. Subsequently, we treated EVs with Panc02 cells, the murine ductal pancreatic cancer cell line, which were co-cultured with splenocytes. Viability and SMAD4 gene expression were examined in Panc02 cells, and cytokine concentrations of IL-6, IL-4, IL-12, and IL-12p70 were measured in the cultured medium. Interestingly, we observed a significant reduction in splenocyte-mediated Panc02 cell death when treated with EVs derived from the adipose tissue of Ob/Ob mice, compared to those from C57BL6/J mice. Additionally, EVs from Ob/Ob mice-derived adipose tissue significantly increased the levels of IL-4, IL-2, and IL-12p70 in the culture media of Panc02 cells co-cultured with splenocytes, compared to EVs from C57BL6/J mice-derived adipose tissue. Adipose tissue-derived EVs from obese mice suppressed splenocyte-mediated Panc02 cell death and upregulated IL-4, IL-2, and IL-12p70 in cultured medium.

The Potential of Biotic Elicitors to Increase Isoflavone Production (Pachyrhizus erosus L.) Callus

Abstract The isoflavones contained in the Pachyrhizus plant (Pachyrhizus erosus Linn) have multiple functions, including serving as raw materials for the health/pharmacology or cosmetic industries. Isoflavones are beneficial as antioxidants, and cholesterol reducers and are capable of neutralizing free radicals and delaying aging. Tissue culture techniques are widely used to produce secondary metabolite compounds. Isoflavones can be made in vitro through callus production by tissue culture. By using elicitation techniques with the addition of various glucose concentrations to the culture media (media Murashige and Skoog or Vacin and Went), it is expected to increase the quantity and quality of callus and the content of secondary metabolites (Isoflavones) in Pachyrhizus callus. The research aims to increase the production of Isoflavones in Pachyrhizus callus by adding elicitors with various concentrations of glucose. The research method used a completely randomized factorial design. Factor I: two levels of basic media: M1 (Murashige and Skoog Media) and M2 (Vacin and Went Media). Factor II: Addition of Glucose concentrations with seven levels: 0%, 5%, 10%, 15%, 20%, 25%, 30%. The plant material used was the young leaves of Pachyrhizus. The results of the study showed that (1) all formed callus exhibited compact quality; (2) the addition of 30% glucose concentration resulted in the best growth of callus; (3) the highest Isoflavone content (0.15%) was found in the callus of Pachyrhizus erosus L. due to the treatment with both MS and VW media with 20% Glucose Elicitor

Mechanism of action and impact of thiol homeostasis on efficacy of an enzyme replacement therapy for classical homocystinuria

Homocystinuria (HCU) due to cystathionine beta-synthase (CBS) deficiency is characterized by elevated plasma and tissue homocysteine levels. There is no cure, but HCU is typically managed by methionine/protein restriction and vitamin B6 supplementation. Enzyme replacement therapy (ERT) based on human CBS has been developed and has shown significant efficacy correcting HCU phenotype in several mouse models by bringing plasma total homocysteine below the clinically relevant 100 μM threshold. As the reactive nature of homocysteine promotes disulfide formation and protein binding, and ERT is unable to normalize plasma total homocysteine levels, the mechanism of action of ERT in HCU remains to be further characterized. Here we showed that only a reduced homocysteine serves as a substrate for CBS and its availability restricts the homocysteine-degrading capacity of CBS. We also demonstrated that cells export homocysteine in its reduced form, which is efficiently metabolized by CBS in the culture medium. Availability of serine, a CBS co-substrate, was not a limiting factor in our cell-based model. Biological reductants, such as N-acetylcysteine, MESNA or cysteamine, increased the availability of the reduced homocysteine and thus promoted its subsequent CBS-based elimination. In a transgenic I278T mouse model of HCU, administration of biological reductants significantly increased the proportion of protein-unbound homocysteine in plasma, which improved the efficacy of the co-administered CBS-based ERT, as evidenced by significantly lower plasma total homocysteine levels. These results clarify the mechanism of action of CBS-based ERT and unveil novel pharmacological approaches to further increase its efficacy.

Proteomics fingerprinting reveals importance of iron and oxidative stress in Streptomyces scabies-Solanum tuberosum interactions.

The Gram-positive actinobacterium Streptomyces scabies is the major causal agent of potato common scab. The main pathogenicity factor is thaxtomin A, a phytotoxin that causes atypical cell death, although other secondary metabolites have been described to play a role in S. scabies virulence. Despite this, many aspects of the interaction between S. scabies and its primary host Solanum tuberosum L. remain to be elucidated. Intracellular proteins of S. scabies EF-35 grown in the presence of in vitro produced tubers (microtubers) of the Russet Burbank and Yukon Gold potato cultivars were extracted and analysed by electrospray mass spectrometry (ES MS/MS). Based on the results of proteomic analysis, iron quantification by ICP-MS and nitrite quantification using Griess reagent in growth media as well as RT-qPCR analysis of the siderophore pyochelin gene expression were performed in the presence and absence of microtubers. Hydrogen peroxide accumulation was also determined in the nutrient medium used for co-cultivation of bacteria and potato microtubers. Potato microtubers caused an increase in the content of bacterial proteins involved in stress and defense, secondary metabolism, and cell differentiation, as well as secreted proteins. Co-cultivation with potato microtubers induced the accumulation of S. scabies proteins implicated in siderophore pyochelin biosynthesis, nitrite production and oxidative stress perception and response. The increase in the abundance of proteins related to pyochelin biosynthesis was consistent with a significant decrease in the iron content in the culture medium, as well as with induction of expression of pyochelin biosynthesis genes. Elevated nitrite/sulfite reductase protein levels were associated with increased nitrite excretion by S. scabies cells in the presence of host microtubers. The increase in the levels of proteins associated with signaling and oxidative stress response could have been caused by the accumulation of ROS, in particular hydrogen peroxide, detected in the studied system. These findings show that interactions of S. scabies with living potato microtubers induce the production of secondary metabolites, defense responses, and protection from oxidative stress. This study suggests the importance of iron during host - S. scabies interactions, resulting in competition between pathogen and its host.

Evaluation of 3β-hydroxysteroid dehydrogenase activity using progesterone and androgen receptors-mediated transactivation.

3β-Hydroxysteroid dehydrogenases (3β-HSDs) catalyze the oxidative conversion of delta (5)-ene-3-beta-hydroxy steroids and ketosteroids. Human 3β-HSD type 2 (HSD3B2) is predominantly expressed in gonadal and adrenal steroidogenic cells for producing all classes of active steroid hormones. Mutations in HSD3B2 gene cause a rare form of congenital adrenal hyperplasia with varying degree of salt wasting and incomplete masculinization, resulting from reduced production of corticoids and androgens. Therefore, evaluation of the HSD3B2 enzymatic activity in both pathways for each steroid hormone production is important for accurately understanding and diagnosing this disorder. Using progesterone receptor (PR)- and androgen receptor (AR)-mediated transactivation, we adapted a method that easily evaluates enzymatic activity of HSD3B2 by quantifying the conversion from substrates [pregnenolone (P5) and dehydroepiandrosterone (DHEA)] to (progesterone and androstenedione). HEK293 cells were transduced to express human HSD3B2, and incubated medium containing P5 or DHEA. Depending on the incubation time with HSD3B2-expressing cells, the culture media progressively increased luciferase activities in CV-1 cells, transfected with the PR/AR expression vector and progesterone-/androgen-responsive reporter. Culture media from human and other mammalian HSD3B1-expressing cells also increased the luciferase activities. HEK293 cells expressing various missense mutations in the HSD3B2 gene revealed the potential of this system to evaluate the relationship between the enzymatic activities of mutant proteins and patient phenotype.

Pengembangan Media Pop Up Book dalam Meningkatkan Budaya Literasi Program Pojok Baca di SMP Negeri 1 Berastagi

The research and development is titled “Development of Pop-Up Book Media in Enhancing Literacy Culture in the Reading Corner Program at SMP Negeri 1 Berastagi.”This development research aims to determine: (1) The design of pop-up book media; (2) The development of pop-up book media in enhancing student literacy; (3) The feasibility of pop-up book media in improving student literacy. This research is conducted using a Research and Development (R&D) model with the 4D model, which includes Define (definition), Design (design), Development (development), and Dissemination (dissemination). The subjects of this research are students of SMP Negeri 1 Berastagi, totaling 35 individuals. The instruments used in this research include questionnaires to assess the development, feasibility, and effectiveness of the media. Data analysis techniques include product data analysis, initial data analysis, and hypothesis testing.The results show that the pop-up book media received evaluations of 81% from media experts, 92% from material experts, and 65% from language experts. There was an average increase in student scores of 83%, indicating a significant change between pretest and posttest scores. The conclusion of the research is that the pop-up book media for drama content is suitable for use in literacy activities of the reading corner program. Recommendations for future research include being more meticulous in planning the development of pop-up book media, ensuring it aligns with the needs and characteristics of students.

Physiological acclimatization in a medium amended with colloidal chitin improves the nematophagous capacity of a fungus on the tomato root-knot nematode

Purpureocillium lilacinum is a nematophagous fungus whose ability to control the plant parasitic nematode Nacobbus aberrans sensu lato has been demonstrated. In this study, physiological acclimatization was performed using different nutrient sources and water activity levels of the culture medium to improve the nematophagous activity of two strains of P. lilacinum (SR14 and SR38). The development of the fungal inoculum in a medium amended with colloidal chitin and reduced levels of water activity (0.95) stimulates the production of conidia and pathogenicity in vitro. This condition was selected to produce the fungal inoculum for conducting antagonism studies against N. aberrans s.l. in tomato plants. The nematode population was significantly reduced (94–96%) with the application of SR38 and SR14 + SR38. The physiological acclimatization in medium with colloidal chitin and 0.95 water activity stimulates the pathogenicity mechanisms (sporulation and infectivity) of P. lilacinum SR38, improving its nematophagous capacity.

B-019 Reduction in Culture Media Costs, without Impacting Turnaround Time, Through Implementation of an Automated Microbiology Platform and Gram Stain Interpretation

Abstract Background Automated plating instruments have been shown to improve quality in Microbiology laboratories in a number of ways including consistent streaking patterns that provide better isolation of bacterial colonies compared to manual methods. The time per culture set up and automated camera imaging is dependent upon the amount of media set, such that more individual plates slow down turn around time. Conversely, smaller media surface area and the use of non-selective media may result in mixed colonies that require sub-culture for isolation before identification and susceptibilities can be performed. Based upon these findings, we attempted to reduce our media usage per culture with the implementation of automation, with the goal of using as few plates as possible while at the same time not negatively impacting the turn around time of the culture via the need to subculture poorly isolated colonies. We conducted a pre-and post-implementation study evaluating the costs per culture and turn around time delays related to subculturing in a subset. Methods In the pre-implementation period, from April-January 2023, three individual media plates (sheep blood agar, chocolate agar and CNA/MacConkey bi-plate) were routinely set for general aerobic bacterial culture sources including swabs and body fluids. In the post implementation period, from April-January 2024, a single sheep blood/chocolate agar bi-plate was set unless the gram stain showed evidence of mixed organisms (defined as more than one morphology) at which time a secondary selective media (CNA/MacConkey) was manually set. For urine cultures, in the pre-implementation period 2 plates were set (sheep blood and CNA/MacConkey bi-plate) and in the post-implementation period a single chromogenic agar/sheep blood bi-plate was set. Total numbers of cultures in both periods were collected as well as the number of cultures for which additional media were set based on gram stain results. Orders of subculture media in the laboratory information system were compared per positive cultures. Statistical analysis was performed in GraphPad Prism. Results In the pre- versus post-implementation periods, a total of 16,454 vs 17,782 general aerobic cultures and 50,680 vs 53,916 urine cultures were set, respectively. Only 3.6% of general cultures required additional media set based on gram stain. The difference in cost between pre- and post-implementation was approximately a two-thirds reduction in cost for general cultures and a 25% reduction for urine cultures, with a total savings of approximately $34,238 in media avoidance in the post-implementation period. The frequency of positive cultures requiring subculture was not significantly different between the pre- (average 36% of positive growth) and post-implementation (average 22% of positive growth) periods for both culture types and may actually have been reduced due to automation. Conclusions The implementation of reduced media concomitant with streaking via automation resulted in significant cost savings while not negatively impacting isolation, which in turn impacts culture turn around times. The automation may have reduced need for subculture, perhaps due to better isolation, thus subsequent analyses will explore this additional finding.

An Overview of Leishmania In Vitro Cultivation and Implications for Antileishmanial Screenings against Promastigotes

The quest for new drug candidates targeting neglected parasitic diseases has become increasingly urgent over the past decades. Advancements in formulating and optimizing drug delivery systems begin with basic research, including direct assays to evaluate the activity of molecules against parasitic stages maintained in laboratories; i.e., promastigotes. In the context of leishmaniasis, an endemic disease worldwide, the cultivation of Leishmania parasites can vary significantly across different laboratories. Factors such as culture media composition, pH, supplementation, and temperature can lead to varied drug responses in in vitro activity assays. This study aims to compile the parameters used in Leishmania spp. promastigotes cultivation protocols described in scientific articles published in indexed journals over the past ten years. The data reveal a lack of uniformity among Leishmania culture protocols, suggesting a potential bottleneck in comparing the leishmanicidal potential of in vitro drug candidates reported by different research groups. This condition is crucial to consider, because viability/inhibition assays should begin with fully-grown, healthy promastigote cultures capable of homogeneous division, thereby producing more reproducible results.

Enhancement of mono-acylated MEL-D production in an acyltransferase gene-deleted strain of Pseudozyma tsukubaensis by supplementation with di-acylated MEL-B in culture medium

Mannosylerythritol lipids (MELs) are glycolipid biosurfactants produced by various yeasts. MEL producers produce mainly di-acylated MELs (consisting of two fatty acid chains). Among them, Pseudozyma tsukubaensis is a di-acylated MEL-B (d-MEL-B) producer. In a previous study, we generated an acyltransferase-deleted strain of P. tsukubaensis (ΔPtMAC2), which selectively produced mono-acylated MEL-D (m-MEL-D, consisting of one fatty acid chain), but not d-MEL-B. However, m-MEL-D productivity in ΔPtMAC2 was low, and oil consumption was significantly reduced compared to the parent strain. Based on these findings, we hypothesized that the d-MEL-B produced by the parent strain may act as an emulsifier in the culture medium, leading to easier utilization of the oil. By contrast, the m-MEL-D produced by ΔPtMAC2 may not have the ability to emulsify oil, thus the oil is used inefficiently and productivity of m-MEL-D is low. Therefore, we expected that adding d-MEL-B to the culture medium during ΔPtMAC2 cultivation would increase m-MEL-D production. To enhance the oil consumption and m-MEL-D production of ΔPtMAC2, d-MEL-B and chemical surfactants were added to the culture medium as emulsifiers during ΔPtMAC2 cultivation. Adding d-MEL-B enhanced both the oil consumption and m-MEL-D production of ΔPtMAC2; Tween 20 and Triton X-100 also showed enhancement effects. As expected, d-MEL-B, Tween20 and TritonX-100, showed marked olive oil emulsification activity, whereas m-MEL-D did not. These results strongly support our hypothesis and significantly improve m-MEL-D productivity.

Detection of Salmonella by competitive ELISA of lipopolysaccharide secreted into the culture medium

Detection of Salmonella in food is topical due to known cases of salmonellosis epidemics. Immunochemical methods including ELISA are widely used for Salmonella detection. Traditionally, commercial ELISA kits are based on sandwich technique and detect lipopolysaccharide (LPS), which is considered to be the component of the outer membrane of Gram-negative bacteria. Our aim was elaboration of competitive ELISA test for Salmonella detection in food with improved parameters. It was shown that in the Salmonella culture after the standard sample preparation procedure LPS is present mainly outside cells as a component of outer membrane vesicles. Improved sample preparation procedure includes separation of bacteria from the medium and analysis of the medium, which increases analytical sensitivity. Immobilization of the bovine serum albumin (BSA)-LPS conjugate in microplate wells allows to obtain a more homogeneous coating than immobilization of LPS itself. Thus, we have developed test system for Salmonella detection in food by competitive ELISA of LPS secreted into the culture medium with the immobilized BSA-LPS conjugate and monoclonal antibodies (mAb) to LPS core in the liquid phase. New competitive ELISA test is high sensitive, give reproducible results, allows the detection of any Salmonella serotype and is important for the protection of human health.

Effects of different culture media on growth, composition, quality and palatability of the green algae Ulva sp. cultivated in cylindrical photobioreactors

Ulva spp. are valuable seaweeds with recognized commercial applications, including food, feed, and ecosystem services. Ensuring a sustainable and consistent supply of biomass with desirable profiles aligned with intended uses is fundamental for the successful applications of this seaweed. In this study, the growth rate, morphology, physiology, and composition of Ulva sp. produced by propagation in indoor cylindrical photobioreactors using four different culture media (lagoon water - LW, lagoon water enriched with Guillard medium (LF), with sea urchin wastewater - LU, and cow digestate - LD) was assessed; moreover, the nutrient uptake potential of the species was evaluated. The palatability and attractivity of the produced biomass towards the sea urchin Paracentrotus lividus were investigated. It was found that the media influenced all the parameters examined, the LF biomass weight was double compared to the other treatments and showed a slightly higher absorbance. Colorimetric analyses reported a significant darker color in Ulva sp. grown under enriched media. Ulva sp. showed higher nutrient removal potential in LF. The lipid content did not vary (2–3 % dry weight, DW), while the protein content ranged from 21 % in LF to 6–9 % in the other treatments. Carbohydrates and fiber content were significantly lower in LF (16 % and 30 %) compared to the other treatments, 27–34 %, and 41–48 %, respectively. Pigment content significantly varied, being higher in biomass grown in LF and LU. Sea urchins showed preferences for biomass grown under LU, followed by LD. This study shows how different nutrient sources affect the biochemical composition, growth, quality, and palatability of Ulva sp.. When cultivated under the synthetic enriched media (LF) the species exhibits characteristics better suitable for human consumption, although requiring a higher economic investment for production, while biomass derived from wastewater nutrients (LD, LU) confirms potential applications of the seaweed as valuable feed and for bioremediation services.

Comparison of the anti-inflammatory effects of carotenoids-rich product from microalgal mass-cultured between natural seawater and magma seawater

Microalgae can be mass-produced in a short period of time and are rich in high value-added product such as carotenoids, thereby it is widely used in a variety industrial field. However, the production cost expensed a lot due to high price of culture ingredient, vitamins. Additionally, > 80% of synthetic carotenoids are used in industrial field, and their cost is also highly expensed. Therefore, in this study, a microalga, Tetraselmis sp. was mass-cultured using natural seawater (NS) and magma seawater (MS) as vitamins. As a result of comparing the general composition between the extracts cultured in two culture media, crude carbohydrate and crude lipid contents were 2.68% and 4.38% higher, respectively, while crude protein content was 4.1% lower in the MS culture media. The extraction yield was 8.6% higher in the MS than the NS culture media. Also, lutein and zeaxanthin were predominantly presented in the species and detected in the microalgal extract cultured in MS was 2- and 8-folds higher, respectively, than in NS. Consequently, the carotenoids-rich extracts cultured with two seawaters were exhibited prominent anti-inflammatory effects by suppressing nitric oxide (NO) overproduction and pro-inflammatory mediator such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, inducible nitric oxide synthase (iNOS), and cyclooxygenase (COX)-2 expression through MAPKSs and NF-κB pathway. Interestingly, extract of microalgal cultured in MS showed a more efficient anti-inflammatory effect. These results suggest that the value of MS in increasing the carotenoids contents with beneficial effects in mass cultivation of microalgae.

Blood culture bottles meet the operating room: enhancing the diagnostic accuracy of infectious spondylitis through open microsurgical biopsy and intraoperative inoculation.

Infectious spondylitis is caused by hematogenous seeding or adjacent soft tissue infection. No study has provided evidence that incubating biopsy specimens in blood culture bottles could enhance detection rates, nor has any study compared this method with conventional culture techniques. We aimed to assess the diagnostic yield of open microsurgical biopsies for infectious spondylitis and the efficacy of various culture media in the presence and absence of pre-biopsy antibiotic therapy. This retrospective study, which was conducted at a university-affiliated teaching hospital in Korea, enrolled 165 adult patients with suspected infectious spondylitis between February 2014 and September 2020. The diagnostic yield of open biopsy was compared among three culture media, namely, blood culture bottles, swab culture using transport media, and tissue culture using plain tubes, while considering preoperative antibiotic exposure. Causative bacteria were identified in 84.2% of all cases. Blood culture bottles had the highest positivity rate (83.5%), followed by swab cultures (64.4%) and tissue cultures (44.9%). The differences in positivity rates were significant (P < 0.001). Preoperative antibiotic therapy reduced detection rates across all media, particularly in tissue cultures. We established the high diagnostic yield of open microsurgical biopsy using blood culture bottles, suggesting that pre-biopsy antibiotic therapy significantly affects bacterial detection, thereby underscoring the importance of culture medium selection in the diagnosis of infectious spondylitis.