Endotoxin Production Research Articles

Cleaning methods for biosafety cabinet to eliminate residual mycoplasmas, viruses, and endotoxins after changeover

IntroductionCell-processing operations can potentially contaminate biosafety cabinets, which should be maintained sterile. However, unintended contamination can occur owing to the presence of viruses, mycoplasmas, and bacteria in the raw materials. Moreover, although several methods for expunging these contaminants have been proposed, an optimal method has not yet been determined. Additionally, the effectiveness of conventional methods for eliminating these contaminants remains unclear owing to their unique characteristics and potential resistances to cleaning. Therefore, this paper proposes a risk-based approach to identify appropriate cleaning methods and reduce the likelihood of cross-contamination in biosafety cabinets by these contaminants. MethodsVarious cleaning methods for eliminating mycoplasmas, viruses, and endotoxins from biosafety cabinets were evaluated, including ultraviolet (UV) irradiation at 200 mJ/cm2 for 20 min and wiping with disinfectants such as distilled water, benzalkonium chloride (BKC), and 70 % ethanol (ETH). The effectiveness of each method was evaluated by applying the contaminants on stainless steel plates and cleaning them using each method. Mycoplasma orale was cultured for 2 weeks in a liquid medium after cleaning. Feline calicivirus (FCV) was used for evaluating the virus-cleaning effectiveness and its presence was tested using the TCID50 test, whereas endotoxins obtained from the dried extract of Escherichia coli were measured via endotoxin testing. ResultsUV irradiation and wiping with BKC inhibited the growth of mycoplasma and significant decreased their presence compared with the other cleaning methods. Notably, mycoplasma were detected after wiping all SUS304 plates with ETH, which is a widely used cleaning method. Additionally, the cleaning efficacy for virus showed that the TCID50 of the wet group was 132,000 TCID50/plate, whereas those after UV irradiation or cleaning with BKC or DW were below the detection limit. Finally, UV irradiation did not significantly reduce the endotoxin production compared with that in the dry group. Additionally, wiping with ETH did not significantly reduce endotoxins compared with the dry group and their residues were higher than those detected after wiping with BKC or DW. ConclusionsThe changeover protocols currently employed in most cell-processing facilities may be ineffective as pathogenic or nonpathogenic materials may remain even after ETH wiping, leading to unintended cross-contamination. To the best of our knowledge, this is the first study to provide reference data of different cleaning methods for mycoplasmas, viruses, and endotoxins in cell-product manufacturing facilities, and can potentially support the development of evidence-based management strategies for ensuring safe cell-product processing.

Impacts of a highly pathogenic ovine Eimeria ovinoidalis on the growth of Hu lambs

The apicomplexan Eimeria ovinoidalis is distributed worldwide. It can cause clinical coccidiosis, which is one of the most pathogenic species in sheep, reducing growth rates and resulting in significant economic losses in the industry. Its principal clinical sign is profuse diarrhoea in young animals. In this study, we established a model of E. ovinoidalis infection in lambs to understand its pathogenicity and evaluate the gut microbiota and fecal metabolite profiles. Specifically, we observed a significant shift in the abundance of bacteria and disrupted metabolism in lambs. Especially during the peak period of excrete oocysts, it promoted the reproduction of some harmful bacteria in Proteobacteria and Actinobacteriota, and reduced the abundance of beneficial bacteria such as Lachnospiraceae and Rikenellaceae. In the later stage of the patent period, the abundance of harmful bacteria in the intestine decreased, the abundance of beneficial bacteria which could produce anti-inflammatory substances began to increase, and the abundance and diversity of intestinal flora also tended to parallel with the control group. Coccidia infection could lead to the increase of differential metabolites and metabolic pathways between infected and control group, but the difference decreased with time. During the peak period of excrete oocysts, although the antimicrobial metabolites such as Lividamine were up-regulated, the excess of these metabolites could still induce the production of endotoxin, while Butanoic acid and other anti-inflammatory metabolites decreased significantly. A metabolomics analysis showed that E. ovinoidalis infection altered metabolites and metabolic pathways, with biosynthesis of unsaturated fatty acids, Teichoic acid biosynthesis and Butanoate metabolism as the major disrupted metabolic pathways. Details of the gut microbiota and the metabolome after infection with E. ovinoidalis may aid in the discovery of specific diagnostic markers and help us understand the changes in parasite metabolic pathways.

Inhalation of ferrate-disinfected Escherichia coli caused lung injury via endotoxin-induced oxidative stress and inflammation response

Ferrate (Fe(VI)) is an environmentally friendly disinfectant that is widely used to eradicate microbes in reclaimed water. However, the potential health risks associated with inhalation of Fe(VI)-treated bacteria-laden reclaimed water remains uncertain. We aimed to explore the inhalation hazards and potential mechanisms of K2FeO4-treated Escherichia coli (E. coli, ATCC 25922). Our findings indicated that Fe(VI) disinfection induced a dose- and time-dependent E. coli inactivation, accompanied by a rapid release of the bacterial endotoxin, lipopolysaccharide (LPS). Scanning electron microscopy (SEM) observations indicate that Fe(VI)-induced endotoxin production consists of at least two stages: initial binding of endotoxin to bacteria and subsequent dissociation to release free endotoxin. Furthermore, Fe(VI) disinfection was not able to effectively eliminate pure or E. coli-derived endotoxins. The E. coli strain used in this study lacks lung infection capability, thus the inhalation of bacteria alone failed to induce severe lung injury. However, mice inhaled exposure to Fe(VI)-treated E. coli showed severe impairment of lung structure and function. Moreover, we observed an accumulation of neutrophil/macrophage recruitment, cell apoptosis, and ROS generation in the lung tissue of mice subjected to Fe(VI)-treated E. coli. RNA sequencing (RNA-seq) and PCR results revealed that genes involved with endotoxin stimuli, cell apoptosis, antioxidant defence, inflammation response, chemokines and their receptors were upregulated in response to Fe(VI)-treated E. coli. In conclusion, Fe(VI) is ineffective in eliminating endotoxins and can trigger secondary hazards owing to endotoxin release from inactivated bacteria. Aerosol exposure to Fe(VI)-treated E. coli causes considerable damage to lung tissue by inducing oxidative stress and inflammatory responses.

Importance of Nano-Sized Feed Additives in Animal Nutrition

"Nano", which derives from the Latin word nanus and means dwarf, refers to a very small unit of measurement equal to one billionth of a meter. Nanotechnology, which deals with the manipulation of matter at the atomic and molecular level, has an application area in animal husbandry as well as in many fields. Nano-sized feed additives, which have come to the forefront in the livestock sector in recent years, have become an innovative application used to increase the nutritional value of feeds and optimize animal health and performance. Since these additives are nano-sized particles with increased specific surface area, they can have a positive effect on a number of factors such as digestibility, nutrient absorption, immune system, growth and development. Minerals in the form of nanoparticles used as feed additives can increase bioavailability by passing through the intestinal wall to body cells faster compared to larger particles. The nano level of the substance not only increases the productivity of animals, but also brings the potential to improve the functionality of feed molecules. Nano feed additives increase the digestion and absorption of feed, allowing animals to benefit from feed more effectively. However, there are several challenges associated with this approach. These include the potential for endotoxin production, reduced nutrient absorption due to interaction with natural nutrients, the possibility of nanoparticle accumulation in the animal body, health risks, ethical considerations, environmental concerns and some negative effects such as interference with natural nutrients that can be avoided by encapsulation. This article discusses recent studies on nano-sized feed additives that offer potential benefits in animal nutrition.

Unraveling the microbial puzzle: exploring the intricate role of gut microbiota in endometriosis pathogenesis.

Endometriosis (EMs) is a prevalent gynecological disorder characterized by the growth of uterine tissue outside the uterine cavity, causing debilitating symptoms and infertility. Despite its prevalence, the exact mechanisms behind EMs development remain incompletely understood. This article presents a comprehensive overview of the relationship between gut microbiota imbalance and EMs pathogenesis. Recent research indicates that gut microbiota plays a pivotal role in various aspects of EMs, including immune regulation, generation of inflammatory factors, angiopoietin release, hormonal regulation, and endotoxin production. Dysbiosis of gut microbiota can disrupt immune responses, leading to inflammation and impaired immune clearance of endometrial fragments, resulting in the development of endometriotic lesions. The dysregulated microbiota can contribute to the release of lipopolysaccharide (LPS), triggering chronic inflammation and promoting ectopic endometrial adhesion, invasion, and angiogenesis. Furthermore, gut microbiota involvement in estrogen metabolism affects estrogen levels, which are directly related to EMs development. The review also highlights the potential of gut microbiota as a diagnostic tool and therapeutic target for EMs. Interventions such as fecal microbiota transplantation (FMT) and the use of gut microbiota preparations have demonstrated promising effects in reducing EMs symptoms. Despite the progress made, further research is needed to unravel the intricate interactions between gut microbiota and EMs, paving the way for more effective prevention and treatment strategies for this challenging condition.

Effects of low-frequency ultrasound combined with anti-MRSA agents on the mouse model of pulmonary infection.

The treatment of methicillin-resistant Staphylococcus aureus (MRSA)-induced pneumonia with antibiotics alone poses considerable challenges. To address these challenges, low-frequency ultrasound (LFU) emerges as a promising approach. In this study, a mouse pneumonia model was established through intratracheal injection of MRSA to investigate the therapeutic efficacy of LFU in combination with antibiotics. Minimal inhibitory concentration was assessed, and the distribution of antibiotics in the lung and plasma was determined using liquid chromatography coupled with mass spectrometry. Various parameters, including the survival rate, histopathology, lung bacterial clearance, and the expressions of cytokines and inflammation-related genes, were evaluated before and after treatment. Compared with the infection group, both the antibiotic-alone groups [vancomycin (VCM), linezolid, and contezolid (CZD)] and the groups in combination with LFU demonstrated an improvement in the survival status of mice. The average colony-forming units of lung tissue in the LFU combination groups were lower compared with the antibiotic-alone groups. While no significant changes in C-reactive protein and procalcitonin in plasma and bronchoalveolar lavage fluid were observed, histopathological results revealed reduced inflammatory damage in LFU combination groups. The secretion of interleukin-6 and tumor necrosis factor-alpha was decreased by the combination treatment, particularly in the VCM + LFU group. Furthermore, the expressions of MRSA virulence factors (hla and agrA) and inflammation-related genes (Saa3, Cxcl9, and Orm1) were further reduced by the combinations of LFU and antibiotics. Additionally, LFU treatment facilitated the distribution of VCM and CZD in mouse lung tissue at 30 and 45 min, respectively, after dosage.IMPORTANCETreating pneumonia caused by methicillin-resistant Staphylococcus aureus (MRSA) with antibiotics alone poses significant challenges. In this in vivo study, we present compelling evidence supporting the efficacy of low-frequency ultrasound (LFU) as a promising approach to overcome these obstacles. Our findings demonstrated that LFU enhanced the effectiveness of vancomycin, linezolid, and contezolid in an MRSA pneumonia model. The combination of LFU with anti-MRSA agents markedly improved the survival rate of mice, accelerated the clearance of pulmonary bacteria, reduced inflammatory injury, inhibited the production of MRSA endotoxin, and enhanced the distribution of antibiotics in lung tissue. The application of LFU in the treatment of pulmonary infections held substantial significance. We believe that readers of your journal will find this topic of considerable interest.

Microencapsulation as a Protective Strategy for Sialylated Immunoglobulin G: Efficacy in Alleviating Symptoms of Dextran Sulfate Sodium-Induced Colitis in Mice and Potential Mechanisms.

Sialylated immunoglobulin G (IgG) is a vital glycoprotein in breast milk with the ability to promote the growth of Bifidobacterium in gut microbiota and relieve inflammatory bowel disease (IBD) symptoms in vitro. Here, it was found that the microcapsules with sialylated IgG could protect and release sialylated IgG with its structure and function in the intestine. Furthermore, the sialylated IgG microcapsules alleviated the clinical symptoms (body weight, feed quantity, and colon length loss), decreased disease activity index score, suppressed the production of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β, IFN-γ, and MCP-1) and endotoxin (lipopolysaccharide), and enhanced the intestinal mucosal barrier (Claudin1, Muc2, Occludin, and ZO-1) in dextran sulfate sodium (DSS)-induced colitis mice. Additionally, the sialylated IgG microcapsules improved the gut microbiota by increasing the relative abundance of critical microbe Bifidobacterium bifidum and promoted the production of short-chain fatty acids (SCFAs). Correlation analysis indicated that the key microbes were strongly correlated with pro-inflammatory factors, clinical symptoms, tight junction protein, and SCFAs. These findings suggest that the sialylated IgG microcapsules have the potential to be used as a novel therapeutic approach for treating IBD.

Engineering Bacillus subtilis for production of 3-hydroxypropanoic acid.

3-Hydroxypropionic acid (3-HP) is a valuable platform chemical that is used as a precursor for several higher value-added chemical products. There is an increased interest in development of cell factories as a means for the synthesis of 3-HP and various other platform chemicals. For more than a decade, concentrated effort has been invested by the scientific community towards developing bio-based approaches for the production of 3-HP using primarily Escherichia coli and Klebsiella pneumoniae as production hosts. These hosts however might not be optimal for applications in e.g., food industry due primarily to endotoxin production and the pathogenic origin of particularly the K. pneumoniae. We have previously demonstrated that the generally recognized as safe organism Bacillus subtilis can be engineered to produce 3-HP using glycerol, an abundant by-product of the biodiesel industry, as substrate. For commercial exploitation, there is a need to substantially increase the titer. In the present study, we optimized the bioprocess conditions and further engineered the B. subtilis 3-HP production strain. Thereby, using glycerol as substrate, we were able to improve 3-HP production in a 1-L bioreactor to a final titer of 22.9g/L 3-HP.

Characterization of Bacillus thuringiensis from Cotton Fields and its Effectiveness against Spodoptera litura

Cotton is a cash crop of many countries as it serves as a source of fiber, edible oil, and seedcake. It is grown in a diverse range of environment. Bacillus thuringiensis (Bt) is a gram positive bacterium found in different habitats but mostly found in soil. This bacterium produces endotoxin which is harmful for various insects. Numerous crystal-forming strains of Bt exist in nature, but not all of them are efficient against insect pests. Bt cotton is prevalent internationally because of less insect attack. The key objective of this research was to analyze different Bacillus thuringiensis isolates residing in soil of different cotton fields of Multan and Bahawalpur districts and to identify them on the basis of morphology. To attain this, different soil samples were collected from Multan (Gup wala, Mosey Wala, Taloki Wala, Hafiz Wala, Choudary Wala, Basti Mangla Mari, Sharkha Wala, Lawain Wala, Ghulamo Wala) and Bahawalpur district (Basti Deewan Wali, Check No 8, Basti Tariqabad, Basti Khaji Wala, Basti Khandin, Basti Rammo Wali, Sultanpur, Khurampur, Jahanpur). About 178 colonies of different bacteria from eighteen soil samples appeared on Nutrient Agar plates. The screening of colonies based on morphology yielded 18 colonies of Bt. We performed endo-toxin production experiment and confirmed the presence of parasporal crystals in identified B. thuringiensis colonies. Colony forming units of soil samples collected from Multan and Bahawalpur represented statistically significant results. Based on morphology, eight colonies of B. thuringiensis were confirmed out of eighteen colonies. Results of the current study revealed that out of 18 isolates, 8 bacterial isolates that were grown on the media had an increased elevation, a round, white, slimy morphology, and smooth edges. Toxicity test for resistance to cotton leaf worm second instar larvae revealed that two strains (BtS2, BtS7) of B. thuringiensis were toxic and showed 39.25% mortality. This study will help to identify the local Bt isolates that can be used for the indigenous insect pest control.

Gut microbiota interactions with anti-diabetic medications and pathogenesis of type 2 diabetes mellitus.

Microorganisms including bacteria, viruses, protozoa, and fungi living in the gastrointestinal tract are collectively known as the gut microbiota. Dysbiosis is the imbalance in microbial composition on or inside the body relative to healthy state. Altered Firmicutes to Bacteroidetes ratio and decreased abundance of Akkermansia muciniphila are the predominant gut dysbiosis associated with the pathogenesis of type 2 diabetes mellitus (T2DM) and metabolic syndrome. Pathophysiological mechanisms linking gut dysbiosis, and metabolic diseases and their complications include altered metabolism of short-chain fatty acids and bile acids, interaction with gut hormones, increased gut microbial metabolite trimethylamine-N-oxide, bacterial translocation/Leaky gut syndrome, and endotoxin production such as lipopolysaccharides. The association between the gut microbiota and glycemic agents, however, is much less understood and is the growing focus of research and conversation. Recent studies suggest that the gut microbiota and anti-diabetic medications are interdependent on each other, meaning that while anti-diabetic medications alter the gut microbiota, the gut microbiota also alters the efficacy of anti-diabetic medications. With increasing evidence regarding the significance of gut microbiota, it is imperative to review the role of gut microbiota in the pathogenesis of T2DM. This review also discusses the interaction between gut microbiota and the various medications used in the treatment of T2DM.

Klebsiella pneumoniae utilizes intestinal mucus to increase fitness in the gastrointestinal tract

BackgroundKlebsiella pneumoniae is a growing concern in the healthcare setting, as carbapenem‐resistant Enterobacteriaceae are considered an urgent health threat by the Center of Disease Control (CDC). K. pneumoniae is found in the mucosa of the human gastrointestinal tract, but upon intestinal disruption K. pneumoniae can migrate into the blood stream and cause infection. One of the barriers against bacterial translocation is the intestinal mucus layer. Mucus proteins are covered with branching glycans which can serve as a fuel source for microbes that harbor mucus‐specific glycosyl hydrolases.HypothesisWe hypothesized that K. pneumoniae could degrade and utilize mucus as an energy source to increase its fitness in the GI tract.Methods & ResultsUsing the Carbohydrate‐Active enZYme Database (CAZY), we identified that 274 K. pneumoniae genomes harbored at least 3 mucin‐degrading glycosyl hydrolase families. To confirm the ability of K. pneumoniae to degrade mucus, we grew five commercially available K. pneumoniae strains and six clinical isolates of K. pneumoniaein a chemically defined minimal media (CDMM) lacking glucose supplemented with mucin‐associated oligosaccharides or dialyzed porcine intestinal MUC2 mucus. All K. pneumoniae strains grew well is sialic acid, GalNAc, fucose and galactose. Interestingly, K. pneumoniae strains exhibited robust growth during log phase with glucose, but were unable to sustain long term growth. Surprisingly, some clinical isolates showed similar growth with GluNAC as with glucose, while other strains could not utilize GluNAC as an energy source. All but one strain of K. pneumoniae could grow with dialyzed mucus as the sole carbon source, indicating that K. pneumoniae can enzymatically degrade mucus and utilize the liberated sugars for growth. To determine how mucus influenced the pathogenesis of K. pneumoniae, we grew one commercially available and one clinical isolate of K. pneumoniae in CDMM with fucose or mucus, isolated the RNA and examined the expression of genes associated with K. pneumoniae virulence by qPCR. We examined virulence factors related to iron uptake (iroD), nucleic acid metabolism (allS), endotoxin production (clbA), fatty acid biosynthesis (clbB), type 1‐ fimbriae (fimH), and flagella (fliC). Both K. pneumoniae strains had increased iroD expression in the presence of mucus. Interestingly, we observed that the clinical isolate, but not the commercially available strain of K. pneumoniae had increased expression of virulence genes clbB, allS and fliC when grown with mucus when compared to being grown with fucose. K. pneumoniae is traditionally considered a nonmotile bacterium, however, we confirmed the expression of flagella in our clinical isolate by performing a chemotaxis assay using mucus‐associated sugars and mucus. We indeed saw that our commercially available strain was unable to migrate towards the mucus or mucus‐associated sugars. However, our clinical isolate was able to swim towards mucus‐associated sugars.ConclusionsThese results suggest that K. pneumoniae utilizes intestinal mucus as an energy source in the GI tract, which in turn influences the pathogenicity of clinical isolates of K. pneumoniae.

Intestinal Flora Changes Induced by a High-Fat Diet Promote Activation of Primordial Follicles through Macrophage Infiltration and Inflammatory Factor Secretion in Mouse Ovaries.

Obesity induced by a high-fat diet (HFD) leads to the excessive consumption of primordial follicles (PFs) in the ovaries. There is systemic chronic inflammation under HFD conditions, but no previous studies have explored whether there is a certain causal relationship between HFD-induced chronic inflammation and the overactivation of PFs. Here, we showed that HFD causes disorders of intestinal microflora in mice, with five Gram-negative bacteria showing the most profound increase at the genus level compared to the normal diet (ND) groups and contributes to the production of endotoxin. Endotoxin promotes M1 macrophage infiltration in the ovaries, where they exhibit proinflammatory actions by secreting cytokines IL-6, IL-8, and TNFα. These cytokines then boost the activation of PFs by activating Signal Transducer and Activator of Transcription 3 (STAT3) signaling in follicles. Interestingly, transplantation of the HFD intestinal microflora to the ND mice partly replicates ovarian macrophage infiltration, proinflammation, and the overactivation of PFs. Conversely, transplanting the ND fecal microbiota to the HFD mice can alleviate ovarian inflammation and rescue the excessive consumption of PFs. Our findings uncover a novel and critical function of gut microbes in the process of PF overactivation under HFD conditions, and may provide a new theoretical basis for the microbial treatment of patients with premature ovarian insufficiency caused by HFD.

Mechanistic Insights Into Gut Microbiome Dysbiosis-Mediated Neuroimmune Dysregulation and Protein Misfolding and Clearance in the Pathogenesis of Chronic Neurodegenerative Disorders.

The human gut microbiota is a complex, dynamic, and highly diverse community of microorganisms. Beginning as early as in utero fetal development and continuing through birth to late-stage adulthood, the crosstalk between the gut microbiome and brain is essential for modulating various metabolic, neurodevelopmental, and immune-related pathways. Conversely, microbial dysbiosis – defined as alterations in richness and relative abundances – of the gut is implicated in the pathogenesis of several chronic neurological and neurodegenerative disorders. Evidence from large-population cohort studies suggests that individuals with neurodegenerative conditions have an altered gut microbial composition as well as microbial and serum metabolomic profiles distinct from those in the healthy population. Dysbiosis is also linked to psychiatric and gastrointestinal complications – comorbidities often associated with the prodromal phase of Parkinson’s disease (PD) and Alzheimer’s disease (AD). Studies have identified potential mediators that link gut dysbiosis and neurological disorders. Recent findings have also elucidated the potential mechanisms of disease pathology in the enteric nervous system prior to the onset of neurodegeneration. This review highlights the functional pathways and mechanisms, particularly gut microbe-induced chronic inflammation, protein misfolding, propagation of disease-specific pathology, defective protein clearance, and autoimmune dysregulation, linking gut microbial dysbiosis and neurodegeneration. In addition, we also discuss how pathogenic transformation of microbial composition leads to increased endotoxin production and fewer beneficial metabolites, both of which could trigger immune cell activation and enteric neuronal dysfunction. These can further disrupt intestinal barrier permeability, aggravate the systemic pro-inflammatory state, impair blood–brain barrier permeability and recruit immune mediators leading to neuroinflammation and neurodegeneration. Continued biomedical advances in understanding the microbiota-gut-brain axis will extend the frontier of neurodegenerative disorders and enable the utilization of novel diagnostic and therapeutic strategies to mitigate the pathological burden of these diseases.

Application of encapsulated nano materials as feed additive in livestock and poultry: a review.

Livestock plays a significant role in the socio-economic development of developing countries by providing livelihood and nutritional security, transport, contingency during crop failure and day-to-day earning to the farm family. The human population is projected to reach 9.9 billion by 2050, which along with increasing urbanization and growing income is expected to increase the demand for livestock products. Global livestock feed demand will be almost doubled, and 1.3 billion tons of grain will be required for feeding farm animals alone. To increase the livestock production efficiency, feed additives have been used to improve the production and growth of high-quality livestock products. Even though feed additives have a pertinent role in animal growth performance, disadvantages like endo toxin production and reduced absorption of nutrients by interaction of additives with naturally occurring nutrients limits the use of feed additives. A majority of the macronutrients in the rumen are metabolized or transformed by microbes, hence nutrients should be preserved and made available in the small intestine. Encapsulation technology, which has been employed in a broad range of applications in various fields of science, can address this challenge and rejuvenate livestock nutrition research. The present review explores the importance of encapsulated nano material in livestock and poultry production.

Gut Microbiome in Stress-related Disorders: The New Approaches to Neuroinflamation syndrome

The review presents modern ideas about changes in the quantitative and qualitative composition of the human intestinal microbiome and their role in the development of stress-induced mental and neurological disorders, eating disorders, autism, etc. The dualism of the role of the commensal representatives of the microbiome, which have the ability to modulate metabolic and signaling reactions in conditionally healthy people and patients suffering from various neurological, psychoemotional and cognitive disorders associated with the development of neuroinflammation, is shown. The favorable and negative effects established by foreign researchers are associated with the presence of specific surface membrane proteins in the intestinal microbiota, the production of certain short-chain fatty acids, mucin degradation, changes in the intestinal barrier function, endotoxin production, as well as the synthesis of certain neurotransmitters. The prospects and difficulties of searching for new microbial biomarkers for predicting the development of stress-induced diseases, as well as for creating new microbial nutraceuticals and new-generation medicines based on living bacteria are considered.

Hyperproduction of PHA copolymers containing high fractions of 4-hydroxybutyrate (4HB) by outer membrane-defected Halomonas bluephagenesis grown in bioreactors.

SummaryBacterial outer membrane (OM) is a self‐protective and permeable barrier, while having many non‐negligible negative effects in industrial biotechnology. Our previous studies revealed enhanced properties of Halomonas bluephagenesis based on positive cellular properties by OM defects. This study further expands the OM defect on membrane compactness by completely deleting two secondary acyltransferases for lipid A modification in H. bluephagenesis, LpxL and LpxM, and found more significant advantages than that of the previous lpxL mutant. Deletions on LpxL and LpxM accelerated poly(3‐hydroxybutyrate) (PHB) production by H. bluephagenesis WZY229, leading to a 37% increase in PHB accumulation and 84‐folds reduced endotoxin production. Enhanced membrane permeability accelerates the diffusion of γ‐butyrolactone, allowing H. bluephagenesis WZY254 derived from H. bluephagenesis WZY229 to produce 82wt% poly(3‐hydroxybutyrate‐co‐23mol%4‐hydroxybutyrate) (P(3HB‐co‐23mol%4HB)) in shake flasks, showing increases of 102% and 307% in P(3HB‐co‐4HB) production and 4HB accumulation, respectively. The 4HB molar fraction in copolymer can be elevated to 32 mol% in the presence of more γ‐butyrolactone. In a 7‐l bioreactor fed‐batch fermentation, H. bluephagenesis WZY254 supported a 84 g l−1 dry cell mass with 81wt% P(3HB‐co‐26mol%4HB), increasing 136% in 4HB molar fraction. This study further demonstrated that OM defects generate a hyperproduction strain for high 4HB containing copolymers.

Endotoxin production in the early stages of urgent abdominal pathology and correction ways

To study the features of the development of endogenous intoxication in association with damage to the organs of the detoxification system, the severity of catabolic processes associated with the activity of peroxidation of membrane lipids and phospholipases in various urgent pathology of the abdomen, on the basis of which to develop a new pathogenetically based approach to the prevention of the progression of surgical endotoxicosis. Clinical section. The studies were conducted in 162 patients with acute urgent pathology (seven groups) with predominant aseptic inflammation (acute severe pancreatitis, acute intestinal obstruction, acute catarrhal cholecystitis) and pronounced purulent-necrotic phenomena (acute peritonitis, acute destructive cholecystitis with empyema of the gallbladder). The severity of endogenous intoxication, the functional state of detoxification system organs (liver, kidneys and intestines), the activity of peroxidation of membrane lipids and phospholipases were evaluated in dynamics. In the last two groups of patients with pancreatitis and peritonitis, complex therapy included remaxol. Experimental section. The experiments were conducted on mongrel dogs that were modeled with acute peritonitis (n=15) or biliary pancreatitis (n=15). A complex of studies similar to those in the clinic was applied in dynamics, but in the tissue structures of the liver, kidneys and intestines, the activity of peroxidation of membrane lipids and phospholipases, as well as the phospholipid composition, was studied. Experimental and clinical studies have shown that in the studied diseases with a different nature of the inflammatory process (aseptic or purulent) in the initial periods, the production of toxins is largely associated with catabolic phenomena associated with a significant activation of peroxidation of membrane lipids and phospholipases. A noticeable increase in the activity of these processes was noted regardless of the nature and severity of the inflammatory process and is associated with the severity of endogenous intoxication and the dysfunctional state of the detoxification system organs. Subsequently, there is a decrease in the detoxification ability of the body against the background of stabilization or even reduction of catabolic phenomena. It is shown that a significant increase in the activity of trigger agents of membrane-stabilizing phenomena occurs regardless of the nature and severity of the inflammatory process. Their significant activity was experimentally established in the tissue structures of the detoxification system organs themselves, which caused the development of pronounced membrane-stabilizing phenomena. It has been shown that the use of remaxol in the complex therapy of patients leads to a noticeable decrease in the activity of membrane modulating agents, which leads to a significant decrease in the severity of endogenous inetoxification syndrome, regardless of the nature of inflammation. This fact was an additional evidence base for the conceptual rethinking of the significance of catabolic phenomena in the pathogenesis of endotoxin production in the early stages of the studied diseases. Experimental and clinical grounds have been obtained for a conceptual rethinking of the role of membrane-stabilizing agents in flooding the body with toxins at the beginning of the disease. One of the leading roles of peroxidation of membrane lipids and phospholipase systems in catabolic processes in the earliest periods of urgent abdominal diseases and in the defeat of detoxification system organs in the subsequent ones is shown.

Plant Extracts in Obesity: A Role of Gut Microbiota.

Obesity has become one of the most serious chronic diseases threatening human health. Its occurrence and development are closely associated with gut microbiota since the disorders of gut microbiota can promote endotoxin production and induce inflammatory response. Recently, numerous plant extracts have been proven to mitigate lipid dysmetabolism and obesity syndrome by regulating the abundance and composition of gut microbiota. In this review, we summarize the potential roles of different plant extracts including mulberry leaf extract, policosanol, cortex moutan, green tea, honokiol, and capsaicin in regulating obesity via gut microbiota. Based on the current findings, plant extracts may be promising agents for the prevention and treatment of obesity and its related metabolic diseases, and the mechanisms might be associated with gut microbiota.

Effect of lon Protease Overexpression on Endotoxin Production and Stress Resistance in Bacillus thuringiensis.

Lon protease, an intracellular protease, plays a key role in cell homeostasis in bacteria and is involved in numerous physiological processes. In this work, we aimed to study the impact of Lon on the production of endotoxins and stress response in Bacillus thuringiensis, which is an important bioinsecticide alternative for toxic chemicals. For this purpose, lon gene was cloned into a multi-copy vector with its original promoter and transcriptional terminator and expressed in B. thuringiensis serovar israelensis ATCC 35,646. Our results showed that the recombinant lon gene transcribed and translated efficiently and the resulting protein was active. Although the sporulation efficiency of the recombinant strain was found to be reduced and its mobility impaired, overexpression of the lon gene triggered the production of endotoxin. Together with increased biofilm formation, recombinant strain exhibited significantly better adaptation to osmotic and heat shock stresses and UV exposure compared to wild type and the control strain with empty plasmid. This study suggested a possible link between Lon protease and the production of insecticide and stress response in B. thuringiensis and provides a platform for future studies focusing on enhancing bio-insecticidal production using this bacterium.

Survival of viable but nonculturable Cronobacter sakazakii in macrophages contributes to infections

Cronobacter sakazakii (C. sakazakii), a pathogen that exists in dry and low-moisture environments, such as powder infant formula (PIF), can enter a viable but nonculturable (VBNC) state under harsh conditions, which enables it to escape traditional detection methods and thus poses a potential public health risk. This study aimed at assessing the virulent nature of VBNC C. sakazakii. Our results showed that VBNC C. sakazakii induced intestinal inflammation in neonatal rats. However, the degree of inflammation was significantly lower than that of culturable bacteria due to decreasing endotoxin production, motility, adhesion, and invasion ability in the VBNC state. From the perspective of bacterial translocation, the numbers of C. sakazakii in the blood, liver, and spleen of rats treated with VBNC cells were in the same order of magnitude as those treated with its culturable counterpart and may lead to the same degree of bacteremia. According to the macrophage survival assays, the survival rate of VBNC C. sakazakii within macrophages was 4.7 times higher than that of culturable cells. Based on these findings, we hypothesize that VBNC C. sakazakii evaded the host immune defense system, penetrated the tissue barrier, and translocated to the bloodstream, liver, and spleen through macrophages. Thus, our study reveals that VBNC C. sakazakii could be a potential risk for infants’ health.