Original Strain Research Articles

Protein rational design and modification of erythrose reductase for the improvement of erythritol production in Yarrowia lipolytica.

Erythritol is a natural non-caloric sweetener, which is produced by fermentation and extensively applied in food, medicine and chemical industries. The final step of the erythritol synthesis pathway is involved in erythritol reductase, whose activity and NADPH-dependent become the limiting node of erythritol production efficiency. Herein, we implemented a strategy combining molecular docking and thermal stability screening to construct an ER mutant library. And we successfully obtained a double mutant ERK26N/V295M (ER*) whose catalytic activity was 1.48 times that of wild-type ER. Through structural analysis and MD analysis, we found that the catalytic pocket and the enzyme stability of ER* were both improved. We overexpressed ER* in the engineered strain ΔKU70 to obtain the strain YLE-1. YLE-1 can produce 39.47 g/L of erythritol within 144 h, representing a 35% increase compared to the unmodified strain, and a 10% increase compared to the strain overexpressing wild-type ER. Considering the essentiality of NADPH supply, we further co-expressed ER* with two genes from the oxidative phase of PPP, ZWF1 and GND1. This resulted in the construction of YLE-3, which exhibited a significant increase in production, producing 47.85 g/L of erythritol within 144 h, representing a 63.90% increase compared to the original chassis strain. The productivity and the yield of the engineered strain YLE-3 were 0.33 g/L/h and 0.48 g/g glycerol, respectively. This work provided an ER mutation with excellent performance, and also proved the importance of cofactors in the process of erythritol synthesis, which will promote the industrial production of erythritol by metabolic engineering of Y. lipolytica.

Genome-wide comparative analysis of CC1 Staphylococcus aureus between colonization and infection

BackgroundStaphylococcus aureus is one of the most important bacteria in human colonization and infection. Clonal complex1 (CC1) is one of the largest and most important S. aureus CCs, and it is a predominant clone in S. aureus colonization and can cause a series of S. aureus infections including bloodstream infections. No studies on the relationship of CC1 S. aureus between colonization and infection have been published.MethodsTo figure out if there are some significant factors in CC1 S. aureus help its colonization or infection, 15 CC1 S. aureus isolates including ten from colonization and five from bloodstream infections were enrolled in this study. Whole-genome sequencing and bioinformatics analysis were performed.ResultsVirulence factor regulators XdrA, YSIRK signal peptide, CPBP family and OmpR family specifically found in infection isolates can promote virulence factors and enhance the pathogenicity of S. aureus. In addition, some significant differences in metabolism and human diseases were discovered between colonization and infection. Fst family of type I toxin–antitoxin system that mainly maintains stable inheritance was specifically found in CC1 S. aureus colonization isolates and might help S. aureus survive for colonization. No significant differences in genomic evolutionary relationship were found among CC1 S. aureus isolates between colonization and infection.ConclusionsVirulence factor regulators and metabolic state can promote CC1 S. aureus pathogenic process compared with colonization, and it seems that the strains of colonization origin cannot have pathogenic potential. Experimental confirmation and a bigger number of CC1 S. aureus strains are necessary for further study about the details and mechanism between colonization and infection.

Enhanced S-adenosyl-L-methionine synthesis in Saccharomyces cerevisiae using metabolic engineering strategies

S-adenosyl-L-methionine (SAM) plays pivotal roles in various physiological processes. With its increasing application in the treatment of diseases such as liver disease, depression, osteoarthritis and Alzheimer's, interest in SAM production aroused. Currently, Saccharomyces cerevisiae is the main industrial producer of SAM. With the surge in demand for SAM, improving the SAM biosynthesis is of importance. In this study, a multimodule engineering strategy was employed to improving SAM production: 1) Enhancing the gene expression of the sulfur assimilation pathway; 2) Strengthening the metabolic flux of the SAM synthesis pathway; 3) Weakening the SAM degradation pathway; 4) Increasing ATP supply. The resulting engineered mutant SC06 (S. cerevisiae CEN.PK2–1CΔgal80∷Tcyc1-sam2-Pgal1-Pgal10-met14-Tadh1, Δlsc2∷Tcyc1-hom6-Pgal1-Pgal10-met6-Tadh1, Δsah1Δmls1) displayed the highest SAM titer of 240.86 mg/L, which was 10.22-fold increase compared with the original strain. With optimized conditions, the SAM titer of mutant SC06 in shake flask fermentation reached 473.02 mg/L with a specific yield of 127.18 mg/g dry cell weight (DCW). In a 5 L fermenter with fed-batch fermentation, the maximal SAM yield of mutant SC06 reached 1.25 g/L with a specific yield of 166.67 mg/g DCW after 58 h cultivation. Therefore, the established metabolic engineering strategies displayed promising efficiency in improving the SAM productivity of S. cerevisiae CEN.PK2–1C, which may provide a useful tool for the improvement of SAM-producing strains.

Exploring AMR and virulence in Klebsiella pneumoniae isolated from humans and pet animals: A complement of phenotype by WGS-derived profiles in a One Health study in Egypt

Klebsiella pneumoniae is a ubiquitous nosocomial pathogen associated with various types of infections in hospitalized patients and different animal species. In the current study, 49 Klebsiella strains isolated from humans, dogs, and cats were investigated using NGS technology. MALDI-TOF failed to identify newly discovered K. variicola and K. quasipneumoniae isolates correctly. MLST analysis revealed different sequence types among K. pneumoniae isolates, and the most frequent STs were ST29, ST219, and ST37. Three ST23 that are generally known as hypervirulent type were identified but they lacked major discriminatory determinants for hypervirulent K. pneumoniae (hvKp). K. pneumoniae isolates showed high diversity, and several isolates from humans and animals were assigned to the same ST and were almost identical. Isolates from humans exhibited more pronounced resistance patterns compared to the animal isolates. High levels of resistance were observed for piperacillin, trimethoprim/sulfamethoxazole, and cephalosporins, and resistance to carbapenem compounds was only found in isolates of human origin. Three strains of human origin were extensively drug-resistant (XDR). A diverse range of resistance genes primarily confer resistance to beta-lactams., phenicol/quinolone, aminoglycoside, macrolide, sulfonamides, and fosfomycin were identified in silico. However, there were inconsistencies between the phenotypic characterization of isolates and the set of resistance genes detected in silico in this set of Klebsiella isolates. Further research using a larger number of isolates from various sources is necessary to fully comprehend the relationship between the presence of antimicrobial resistance determinants and phenotypic data. It is also necessary to monitor the spread of K. pneumoniae from a One Health perspective in Egypt.

Hot Spots of Site-Specific Integration into the Sinorhizobium meliloti Chromosome.

The diversity of phage-related sequences (PRSs) and their site-specific integration into the genomes of nonpathogenic, agriculturally valuable, nitrogen-fixing root nodule bacteria, such as Sinorhizobium meliloti, were evaluated in this study. A total of 314 PRSs, ranging in size from 3.24 kb to 88.98 kb, were identified in the genomes of 27 S. meliloti strains. The amount of genetic information foreign to S. meliloti accumulated in all identified PRSs was 6.30 Mb. However, more than 53% of this information was contained in prophages (Phs) and genomic islands (GIs) integrated into genes encoding tRNAs (tRNA genes) located on the chromosomes of the rhizobial strains studied. It was found that phiLM21-like Phs were predominantly abundant in the genomes of S. meliloti strains of distant geographical origin, whereas RR1-A- and 16-3-like Phs were much less common. In addition, GIs predominantly contained fragments of phages infecting bacteria of distant taxa, while rhizobiophage-like sequences were unique. A site-specific integration analysis revealed that not all tRNA genes in S. meliloti are integration sites, but among those in which integration occurred, there were "hot spots" of integration into which either Phs or GIs were predominantly inserted. For the first time, it is shown that at these integration "hot spots", not only is the homology of attP and attB strictly preserved, but integrases in PRSs similar to those of phages infecting the Proteobacteria genera Azospirillum or Pseudomonas are also present. The data presented greatly expand the understanding of the fate of phage-related sequences in host bacterial genomes and also raise new questions about the role of phages in bacterial-phage coevolution.

High Throughput Screening of Transcription Factor LysG for Constructing a Better Lysine Biosensor.

The biosensors based on transcription factors (TFs) are widely used in high throughput screening of metabolic overproducers. The unsatisfactory performances (narrow detection and dynamic ranges) of biosensors limit their practical application and need more improvement. In this study, using the TF LysG (sensing lysine) as an example, a biosensor optimization method was constructed by growth-coupled screening of TF random mutant libraries. The better the performance of the biosensor, the faster the strain grows under screening pressure. A LysGE15D, A54D, and I164V-based biosensors were obtained, which were about 2-fold of the control in the detection and dynamic ranges. A lysine high-producer was screened effectively using the optimized biosensor with the production at 1.51 ± 0.30 g/L in flasks (2.22-fold of the original strain). This study provided a promising strategy for optimizing TF-based biosensors and was of high potential to be applied in the lysine high-producers screening process.

Industrial-Level Brewing Using Oenological Saccharomyces cerevisiae and Schizosaccharomyces pombe as Mixed-Inoculum

The development of new food processes and formulations begins at the laboratory stage, progresses through pilot plant trials, and culminates in industrial production. Although the positive effects in terms of sensory characteristics and qualitative differentiation have been widely studied at laboratory level, fermentations conducted at the industrial level by oenological Saccharomyces cerevisiae and non-Saccharomyces strains have not been thoroughly investigated. Scaling up to the industrial level is a critical process that involves more than simply increasing the dimensions of the process itself. The purpose of our research was to compare laboratory and industrial-level brewing of a novel craft beer produced with the addition of common unmalted wheat and fermented by Schizosaccharomyces pombe and S. cerevisiae strains. Fermentation was carried out using a S. cerevisiae strain either of oenological origin alone or through sequential inoculations with S. pombe. Beers produced with the mixed starter showed greater reproducibility between the two production levels than those fermented by S. cerevisiae alone. According to the results, the main differences highlighted between laboratory and industrial-level trials with S. cerevisiae alone concerned the extent of starch degradation, fermentation efficiency, and alcohol production, which were higher in brewing at the laboratory level. In contrast, beers produced at industrial level using sequential inoculation received significantly higher scores for foam quantity and persistence, as well as overall olfactory intensity, while scoring significantly lower scores for saltiness and sourness. To our knowledge, this research is the first to explore the use of Sc. pombe for industrial beer production.

Antibiotic-resistant mutants of lactic acid bacteria: potential food control agents

Background: One of the most pressing challenges facing global agriculture today is the restoration of degraded farmland. Organic fertilisers are increasingly being used as a safer alternative to synthetic forms. The most promising form of environmentally friendly fertiliser is a bioconsortium based on different groups of microorganisms. Lactic acid bacteria (LAB) with high antifungal activity, isolated from Armenian dairy products, can form part of such a bioconsortium and protect plants from phytopathogens. In addition, LAB can be used as a preservative in the food industry due to its GRAS (generally recognised as safe) status. Objective: To obtain antibiotic-resistant pleiotropic lactic acid bacterial mutants with higher antifungal activity and broad-spectrum activity compared to natural strains. Methods: The objects of the study were LAB, whose antifungal activity was studied against phytopathogenic fungi of the genera Penicillium, Aspergillus, Alternaria and Fusarium, which cause crop/food spoilage. To increase antifungal activity, spontaneous pleiotropic mutants resistant to the streptomycin, kanamycin or rifampicin were obtained on the basis of original strains. Results: Among all the LAB tested, Lactobacillus bovis MDC 1061, L. paracasei MDC 10898 and L. buchneri BKM 1599 were selected for their antifungal activity. Based on the latter, 78 antibiotic-resistant mutants with different levels of antifungal activity were obtained. L. bovis MDC 1061 STR-25 and L. buchneri BKM 1599 RIF-7 mutants with up to 98% and 95% inhibition of fungal growth, respectively, were selected as potential food control agents. Conclusion: Antibiotic-resistant mutants L. bovis MDC 1061 STR-25 and L. buchneri BKM 1599 RIF-7 with high antifungal ability and a wide range of its effects were obtained. For the first time, the phenomenon of pleiotropic effect has been used in LAB to improve antifungal activity. After extensive study, these mutants can be used as promising components for creating an effective bioconsortium, as well as a preservative in food production, especially in the case of plant-based functional foods to protect against contamination by fungi. Keywords: lactic acid bacteria, pleiotropic mutants, antifungal activity

Potential of Trichoderma strains to positively modulate plant growth processes and bulb yield in Rabi onion

The use of beneficial microbes is hitherto known and constantly increasing in agriculture due to their positive impact on crop growth and yield, and their minimal negative impact on the environment. The objective of this study was to evaluate the impact of eight Trichoderma strains of diverse origin on crop growth and yield of onion under field conditions. The identity of the strains used in the current study was confirmed by ITS and Tef1 gene sequencing. Field experiments were conducted in the Rabi season for 2 years (2020–21, and 2022–23) to evaluate the effect of the application of eight different Trichoderma strains that were applied individually and separately as eight different treatments (T1–T8) in experimental plots. In the plant growth promotion assay conducted in vitro, all strains showed the ability to produce IAA (indole-3-acetic acid), with levels ranging from 23.52 μg/mL (T6) to 45.54 μg/mL (T3). Our results revealed that Trichoderma treated experimental plots displayed better growth indices (plant height, pseudostem diameter), RWC (Relative water content), leaf chlorophyll content, and yield-attributing features like biomass (bulb and root dry mass), bulb diameter, and harvested bulb yield compared to the untreated control plants. In terms of yield, the T2 strain exhibited the highest bulb yield consistently for both the years (2020–21 and 2022–23) followed by T3 being statistically at par with T5. Among all the evaluated Trichoderma strains, the strain T2 (OGRDT2) and T3 (GRDT1), taxonomically identified as Trichoderma longibrachiatum, registered bulb yield of 32.24 t/ha and 30.76 t/ha, respectively while T5 (GRDT3), identified as Trichoderma asperellum, registered 30.55 t/ha average yield for 2 years compared to 24.08 t/ha average yield recorded for untreated control plants with an increase of 34, 28 and 27%, respectively. Based on our findings, it is concluded that the T. longibrachiatum strains OGRDT2 (T2) and GRDT1 (T3), T. asperellum strain GRDT3 (T5) are the best inducers of the onion crop growth and yield in the Rabi season and would be explored further for its commercial application in onion farming.

PSI-22 Evaluating colonization success of Bacteroidetes from different host origin in broiler chickens

Abstract Current broiler production practices that aim to maximize biosecurity have limited chick exposure to beneficial microbes. Previous research highlights a notable disparity in Bacteroidetes abundance and taxonomy between birds reared under intensive and extensive systems. Building upon this, we hypothesized that Bacteroidetes strains populating intensively raised birds may originate from human sources, potentially leading to varied colonization efficiencies. The objective of this study was to evaluate the colonization patterns of diverse Bacteroidetes strains within the chicken gastrointestinal tract. One day-old broiler birds (n = 63) were raised in individually ventilated isolators accommodating 3 birds per cage. The chicks were subjected to inoculation either via oral gavage with 150 µL of glycerol stock containing 1x104 colony forming units (CFU)/mL; or spray method (2 mL per cage) using chicken-derived, human-derived, and a combination of Bacteroidetes strains. Notably, birds administered via oral gavage displayed an increase in body weight gain by d 07 compared with their counterparts treated via spray and the control group (P < 0.01). Subsequent quantification of Bacteroidetes, Enterobacteriaceae, and total bacteria using qPCR revealed that oral gavage resulted in more successful Bacteroidetes colonization at d 14 (P < 0.01). Moreover, discernible variations were noted in total bacterial abundance across treatment cohorts. It can be concluded that the method of inoculation appears to influence the initial colonization patterns of Bacteroidetes in the chicken gut, or that inoculation in the environment requires a higher dose. Amplicon sequencing that is underway will reveal the success of the Bacteroidetes from different host origin and the most successful strains of chicken origin. Overall, enhanced growth coincided with more successful Bacteroidetes colonization, identifying them as potential targets for enhanced broiler performance. Further investigation is warranted to determine if Bacteroidetes from different sources demonstrate varying colonization efficiencies.

Aurantiochytrium mutant strains exhibiting different colony colors altered the contents of squalene.

Aurantiochytrium sp. 18W-13a, a marine heterotrophic protist belonging to the genus thraustochytrid, is known to accumulate high levels of squalene and carotenoids. Nowadays, the mutagenesis breeding of microorganisms is still widely practiced because the induced mutations of DNA do not involve the permanent integration of heterologous DNA sequences. Therefore, in this study, we focused on the improvement of squalene yield by mutagenesis breeding using Aurantiochytrium sp. 18W-13a. To bypass the massively laborious screening, we propose to use colony colors as the first criterion to screen mutants with high squalene accumulation, since the carotenoid and squalene synthetic pathways share an intermediate. We selected pale (white)-colored mutants after carbon ion irradiation. The white mutants exhibited larger squalene yields than twice as much of the original strain. The results clearly indicate that the present screening method with colony colors promises to obtain productive strains of squalene.

Analysis of oil accumulation mechanisms in plasma induced mutant Scenedesmus strains compared to original Scenedesmus strains

Scenedesmus sp. is a species of the Scenedesmus genus within the phylum Chlorophyta, commonly found as a planktonic algal species in freshwater and known for its rapid growth rate. This study employs room-temperature, atmospheric-pressure plasma mutagenesis for the breeding of Scenedesmus sp., utilizing transcriptomic analysis to investigate the biosynthesis mechanism of triglycerides. Further analysis of differentially expressed genes in transcriptome by measuring the macroscopic biological indicators of mutant and original algal strains. The findings of the study suggest that the mutant strain's photosynthesis has been enhanced, leading to improved light energy utilization and CO2 fixation, thereby providing more carbon storage and energy for biomass and lipid production. The intensification of glycolysis and the TCA (tricarboxylic acid) cycle results in a greater shift in carbon flux towards lipid accumulation. An elevated expression level of related enzymes in starch and protein degradation pathways may enhance acetyl CoA accumulation, facilitating a larger substrate supply for fatty acid production and thereby increasing lipid yield.

Evolving patterns of COVID-19 mortality in US counties: A longitudinal study of healthcare, socioeconomic, and vaccination associations.

The COVID-19 pandemic emphasized the need for pandemic preparedness strategies to mitigate its impacts, particularly in the United States, which experienced multiple waves with varying policies, population response, and vaccination effects. This study explores the relationships between county-level factors and COVID-19 mortality outcomes in the U.S. from 2020 to 2023, focusing on disparities in healthcare access, vaccination coverage, and socioeconomic characteristics. We conduct multi-variable rolling regression analyses to reveal associations between various factors and COVID-19 mortality outcomes, defined as Case Fatality Rate (CFR) and Overall Mortality to Hospitalization Rate (OMHR), at the U.S. county level. Each analysis examines the association between mortality outcomes and one of the three hierarchical levels of the Social Vulnerability Index (SVI), along with other factors such as access to hospital beds, vaccination coverage, and demographic characteristics. Our results reveal persistent and dynamic correlations between various factors and COVID-19 mortality measures. Access to hospital beds and higher vaccination coverage showed persistent protective effects, while higher Social Vulnerability Index was associated with worse outcomes persistently. Socioeconomic status and vulnerable household characteristics within the SVI consistently associated with elevated mortality. Poverty, lower education, unemployment, housing cost burden, single-parent households, and disability population showed significant associations with Case Fatality Rates during different stages of the pandemic. Vulnerable age groups demonstrated varying associations with mortality measures, with worse outcomes predominantly during the Original strain. Rural-Urban Continuum Code exhibited predominantly positive associations with CFR and OMHR, while it starts with a positive OMHR association during the Original strain. This study reveals longitudinal persistent and dynamic factors associated with two mortality rate measures throughout the pandemic, disproportionately affecting marginalized communities. The findings emphasize the urgency of implementing targeted policies and interventions to address disparities in the fight against future pandemics and the pursuit of improved public health outcomes.

Booster vaccination using bivalent DS-5670a/b is safe and immunogenic against SARS-CoV-2 variants in children aged 5-11 years: a phase 2/3, randomized, active-controlled study.

DS-5670 is a messenger ribonucleic acid (mRNA) vaccine platform targeting the receptor-binding domain (RBD) of the spike protein derived from severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2). Booster vaccination against coronavirus disease 2019 (COVID-19) with monovalent DS-5670a (incorporating mRNA encoding the RBD from the original SARS-CoV-2 strain) or bivalent DS-5670a/b (original and omicron BA.4-5 RBD antigens) is effective and safe in adults. Data from a phase 2/3 active-controlled, non-inferiority, pediatric study evaluating a third booster dose of DS-5670a/b are reported here. Children aged 5-11 years who had completed the two-dose primary vaccination series with monovalent BNT162b2 (original strain) at least 3 months prior to enrolment were randomly assigned to receive DS-5670a/b (20 µg of mRNA) or bivalent BNT1 62b2 (original/omicron BA.4-5; 10 µg of mRNA) on Day 1. The primary efficacy endpoint was blood neutralization geometric mean titer (GMT) against SARS-CoV-2 (omicron variant BA.5.2.1) and immune response rate (≥ 4-fold increase in post-vaccination circulating anti-SARS-CoV-2 neutralizing activity) on Day 29. Among evaluable participants (DS-5670a/b, n = 74; bivalent BNT162b2, n = 75), the adjusted GMT ratio of DS-5670a/b to bivalent BNT162b2 on Day 29 was 1.636 (95% CI, 1.221, 2.190). Immune response rates were ≥ 89% with both study vaccines; adjusted difference 2.6% (95% CI, -7.8, 13.8). The prespecified non-inferiority margins were exceeded, and the study met the primary endpoint. DS-5670a/b also demonstrated broad neutralization activity across recent omicron sublineages and no cases of COVID-19 between Days 8-29 post-administration were reported. There were no novel safety concerns in the pediatric population at data cut-off. Bivalent DS-5670a/b was non-inferior to bivalent BNT162b2 in terms of immunogenicity, and had a manageable safety profile, when administered as a heterologous booster in children aged 5-11 years. https://jrct.niph.go.jp/, identifier jRCT2031220665.

Potential Probiotic Properties and Complete Genome Analysis of Limosilactobacillus reuteri LRA7 from Dogs.

This study aimed to isolate and screen canine-derived probiotics with excellent probiotic properties. Strain characterization was conducted using a combination of in vitro and in vivo probiotic characterization and safety assessments, as well as complete genome analysis. The results showed that Limosilactobacillus reuteri LRA7 exhibited excellent bacteriostatic and antioxidant activities. The survival rate at pH 2.5 was 79.98%, and the viable counts after exposure to gastrointestinal fluid and 0.5% bile salts were 7.77 log CFU/mL and 5.29 log CFU/mL, respectively. The bacterium also exhibited high hydrophobicity, self-coagulation, and high temperature tolerance, was negative for hemolysis, and was sensitive to clindamycin. In vivo studies in mice showed that the serum superoxide dismutase activity level was 53.69 U/mL higher in the MR group of mice compared to that of the control group, the malondialdehyde content was 0.53 nmol/mL lower in the HR group, and the highest jejunal V/C value was 4.11 ± 1.05 in the HR group (p < 0.05). The L. reuteri LRA7 gene is 2.021 megabases in size, contains one chromosome and one plasmid, and is annotated with 1978 functional genes. In conclusion, L. reuteri LRA7 has good probiotic potential and is safe. It can be used as an ideal probiotic candidate strain of canine origin.

Development of a green Komagataella phaffii cell factory for sustainable production of plant-derived sesquiterpene (–)-α-bisabolol

(–)-α-Bisabolol is a plant-derived sesquiterpene derived from Eremanthus erythropappus, which can be used as a raw material in cosmetics and has anti-inflammatory function. In this study, we designed six mutation sites of the (–)-α-bisabolol synthase BOS using the plmDCA algorithm. Among these, the F324Y mutation demonstrated exceptional performance, increasing the product yield by 73 %. We constructed a de novo (–)-α-bisabolol biosynthesis pathways through systematic synthetic biology strategies, including the enzyme design of BOS, selection of different linkers in fusion expression, and optimization of the mevalonate pathway, weakening the branching metabolic flow and multi-copy strategies, the yield of (–)-α-bisabolol was significantly increased, which was nearly 35-fold higher than that of the original strain (2.03 mg/L). The engineered strain was capable of producing 69.7 mg/L in shake flasks. To the best of our knowledge, this is the first report on the biosynthesis of (–)-α-bisabolol in Komagataella phaffii, implying this is a robust cell factory for sustainable production of other terpenoids.

In vitro Synergistic and Bactericidal Effects of Aztreonam in Combination with Ceftazidime/ Avibactam, Meropenem/Vaborbactam and Imipenem/Relebactam Against Dual-Carbapenemase-Producing Enterobacterales.

Our aim was to elucidate the resistance mechanisms and assess the combined synergistic and bactericidal activities of aztreonam in combination with ceftazidime/avibactam (CZA), meropenem/vaborbactam (MEV), and imipenem/relebactam (IMR) against Enterobacterales strains producing dual carbapenemases. Species identification, antimicrobial susceptibility testing and determination of carbapenemase type were performed for these strains. Plasmid sizes, plasmid conjugation abilities and the localization of carbapenemase genes were investigated. Whole-genome sequencing was performed for all strains and their molecular characteristics were analyzed. In vitro synergistic and bactericidal activities of the combination of aztreonam with CZA, MEV and IMR against these strains were determined using checkerboard assay and time-kill curve assay. A total of 12 Enterobacterales strains producing dual-carbapenemases were collected, including nine K. pneumoniae, two P. rettgeri, and one E. hormaechei. The most common dual-carbapenemase gene pattern observed was bla (KPC-2+NDM-5) (n=4), followed by bla KPC-2+IMP-26 (n=3), bla (KPC-2+NDM-1) (n=2), bla (KPC-2+IMP-4) (n=1), bla (NDM-1+IMP-4) (n=1) and bla (KPC-2+KPC-2) (n=1). In each strain, the carbapenemase genes were found to be located on two distinct plasmids which were capable of conjugating from the original strain to the receipt strain E. coli J53. The results of the checkerboard synergy analysis consistently revealed good synergistic effects of the combination of ATM with CZA, MEV and IMR. Except for one strain, all strains exhibited significant synergistic activity and bactericidal activity between 2 and 8 hours. Dual-carbapenemase-producing Enterobacterales posed a significant threat to clinical anti-infection treatment. However, the combination of ATM with innovative β-lactam/β-lactamase inhibitor compounds had proven to be an effective treatment option.

Lactiplantibacillus plantarum IS-10506 Supplementation Improves Clinical Outcome and Immunology Markers in Psoriasis Vulgaris Patients: A Randomized Controlled Trial

BACKGROUND: Probiotics may modify the gut microbiome and have been proven to improve psoriasis vulgaris. Lactiplantibacillus plantarum IS-10506 is a probiotic strain of Indonesian origin. It offers a safe and effective probiotic for psoriasis patients in Indonesia. This study was conducted to evaluate the effect of L. plantarum IS-10506 on clinical and immunology markers in psoriasis vulgaris.METHODS: This randomized, placebo-controlled, and double-blind trial compared L. plantarum IS-10506 (2×1010 CFU/day) and placebo in 49 patients mild-moderate psoriasis vulgaris, which were divided into intervention (n=24) and control groups (n=25). The interventions were given twice daily for 12 weeks. Both groups received topical corticosteroid and emollient as standard treatment. Psoriasis area and severity index (PASI), dermatology life quality index (DLQI), interleukin (IL)-10, IL-17, and forkhead box protein (Foxp3) were then assessed.RESULTS: Mean PASI score for the the subjects in probiotic group was significantly reduced compared to placebo at week-6 (p=0.024), and was sustained until week-12 (p=0.049). At week-12, DLQI scores in the probiotic group were lower than placebo (7.57±5.77 vs. 7.79±5.48). IL-17 level was significantly decreased (p=0.013), while the IL-10 and Foxp3 were significantly increased (p≤0.001 and p=0.048, respectively) in probiotic group. Six months after the completion of study, subjects in probiotic group had a lower probability of flares (52.2%) compared to placebo (79.2%). Two subjects receiving probiotics and one receiving placebo noticed changes in defecation frequency, while another subject in the placebo group complained of mild nausea.CONCLUSION: L. plantarum IS-10506 might effectively improve clinical outcomes and immune biomarkers in psoriasis vulgaris patients, potentially acting as an adjuvant therapy.KEYWORDS: psoriasis, probiotic, clinical severity, immune marker, human and health

Improvement of Saccharomyces cerevisiae strain tolerance to vanillin through heavy ion radiation combined with adaptive laboratory evolution

Vanillin is an inhibitor of lignocellulose hydrolysate, which can reduce the ability of Saccharomyces cerevisiae to utilize lignocellulose, which is an important factor limiting the development of the ethanol fermentation industry. In this study, mutants of vanillin-tolerant yeast named H6, H7, X3, and X8 were bred by heavy ion irradiation (HIR) combined with adaptive laboratory evolution (ALE). Phenotypic tests revealed that the mutants outperformed the original strain WT in tolerance, growth rate, genetic stability and fermentation ability. At 1.6 g/L vanillin concentration, the average OD600 value obtained for mutant strains was 0.95 and thus about 3.4-fold higher than for the wild-type. When the concentration of vanillin was 2.0 g/L, the glucose utilization rate of the mutant was 86.3 % within 96 h, while that of the original strain was only 70.0 %. At this concentration of vanillin, the mitochondrial membrane potential of the mutant strain recovered faster than that of the original strain, and the ROS scavenging ability was stronger. We analyzed the whole transcriptome sequencing map and the whole genome resequencing of the mutant, and found that DEGs such as FLO9, GRC3, PSP2 and SWF1, which have large differential expression multiples and obvious mutation characteristics, play an important role in cell flocculation, rDNA transcription, inhibition of DNA polymerase mutation and protein palmitoylation. These functions can help cells resist vanillin stress. The results show that combining HIR with ALE is an effective mutagenesis strategy. This approach can efficiently obtain Saccharomyces cerevisiae mutants with improved vanillin tolerance, and provide reference for obtaining robust yeast strains with lignocellulose inhibitor tolerance.

The Clinical Severity of COVID-19 Variants of Concern: Retrospective Population-Based Analysis.

SARS-CoV-2 variants of concern (VOCs) emerged and rapidly replaced the original strain worldwide. The increased transmissibility of these new variants led to increases in infections, hospitalizations, and mortality. However, there is a scarcity of retrospective investigations examining the severity of all the main VOCs in presence of key public health measures and within various social determinants of health (SDOHs). This study aims to provide a retrospective assessment of the clinical severity of COVID-19 VOCs in the context of heterogenous SDOHs and vaccination rollout. We used a population-based retrospective cohort design with data from the British Columbia COVID-19 Cohort, a linked provincial surveillance platform. To assess the relative severity (hospitalizations, intensive care unit [ICU] admissions, and deaths) of Gamma, Delta, and Omicron infections during 2021 relative to Alpha, we used inverse probability treatment weighted Cox proportional hazard modeling. We also conducted a subanalysis among unvaccinated individuals, as assessed severity differed across VOCs and SDOHs. We included 91,964 individuals infected with a SARS-CoV-2 VOC (Alpha: n=20,487, 22.28%; Gamma: n=15,223, 16.55%; Delta: n=49,161, 53.46%; and Omicron: n=7093, 7.71%). Delta was associated with the most severe disease in terms of hospitalization, ICU admissions, and deaths (hospitalization: adjusted hazard ratio [aHR] 2.00, 95% CI 1.92-2.08; ICU: aHR 2.05, 95% CI 1.91-2.20; death: aHR 3.70, 95% CI 3.23-4.25 relative to Alpha), followed generally by Gamma and then Omicron and Alpha. The relative severity by VOC remained similar in the unvaccinated individual subanalysis, although the proportion of individuals infected with Delta and Omicron who were hospitalized was 2 times higher in those unvaccinated than in those fully vaccinated. Regarding SDOHs, the proportion of hospitalized individuals was higher in areas with lower income across all VOCs, whereas among Alpha and Gamma infections, 2 VOCs that cocirculated, differential distributions of hospitalizations were found among racially minoritized groups. Our study provides robust severity estimates for all VOCs during the COVID-19 pandemic in British Columbia, Canada. Relative to Alpha, we found Delta to be the most severe, followed by Gamma and Omicron. This study highlights the importance of targeted testing and sequencing to ensure timely detection and accurate estimation of severity in emerging variants. It further sheds light on the importance of vaccination coverage and SDOHs in the context of pandemic preparedness to support the prioritization of allocation for resource-constrained or minoritized groups.