Optimized expression and purification of a human adenosine deaminase in E. coli and characterization of its Asp8Asn variant.

Homo sapiens adenosine deaminase isoform 1 (HsADA1) hydrolyzes adenosine and 2-deoxyadenosine as a key step in the purine nucleoside salvage pathway. Some HsADA1 mutations have severe deleterious effects, as is the case in a severe combined immunodeficiency resulting from loss of enzyme activity (ADA-SCID). Other mutations that reduce enzyme activity, for instance the Asp8Asn (D8N) variant, do not cause ADA-SCID but are correlated with other consequences to health. To ease further study of HsADA1 and its variants, we optimized an inexpensive, recombinant expression process in an Escherichia coli host through multiplexed parameter testing enabled by a lysate-based microtiter plate assay. We demonstrate the importance of gene codon usage, induction time and temperature, and alcohol supplementation towards improving enzyme yield to a final titer of 5 mg per liter of culture. We further show that use of a double-histidine-tag (his-tag) system greatly improves purity. We then utilize our expression and purification framework to produce the HsADA1 D8N variant, which had previously not been purified to homogeneity. We confirm that the D8N variant is ∼30% less active than the wildtype HsADA1 and show that it better retains its activity in human serum. Additionally, we show that both HsADA1 and the D8N variant have heightened activity in serum, driven in part by a previously undescribed phenomenon involving albumin. Therefore, this work presents a valuable process to produce HsADA1 that allows for insights into it and its variants' behavior. We also confirm the utility of lysate-based activity assays towards finding optimal E. coli expression conditions for enzymes and show how fusing his-tags in tandem can enhance product purity.

Preventive effects of prenatal administration of OM-85/BV on asthma and respiratory infection risk in the offspring: A review of animal models.

Asthma is the most common chronic disease in childhood affecting the daily lives of many patients despite current treatment regimens. Therefore, the need for new therapeutic approaches is evident, where a primary prevention strategy is the ultimate goal. Studies of children born to mothers in farming environments have shown a lower risk of respiratory infections and asthma development. Already at birth, these newborns have demonstrated accelerated maturation and upregulation of host defense immune functions suggesting a prenatal transplacental training of the innate immune system through maternal microbial exposure. This mechanism could possibly be utilized to help prevent both respiratory infections and asthma in young children. Human studies exploring the potential preventative effects of pregnancy bacterial lysate treatment on asthma and respiratory infections are lacking, however, this has been studied in experimental studies using mice through administrations of the bacterial lysate OM-85. This review will present the current literature on the immunomodulatory effects relevant for respiratory infections and asthma in the offspring of mice treated with OM-85 throughout pregnancy. Further, the review will discuss the cellular and molecular mechanisms behind these effects. In conclusion, we found promising results of an accelerated immune competence and improved resistance to airway challenges as a result of prenatal bacterial lysate treatment that may pave the way for implementing this in human trials to prevent asthma and respiratory infections.

Assessment of the Effect on Periodontitis of Antibiotic Therapy and Bacterial Lysate Treatment.

Periodontitis is an inflammatory process that starts with soft tissue inflammation caused by the intervention of oral bacteria. By modulating local immunity, it is possible to supplement or replace current therapeutic methods. The aim of this study was to compare the effects of an immunostimulatory treatment with the antibiotherapy usually applied to periodontitis patients. On a model of periodontitis induced in 30 rats (divided into three equal groups) with bacterial strains selected from the human oral microbiome (Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum and Streptococcus oralis), we administered antibiotics, bacterial lysates and saline for 10 days. Clinically, no significant lesions were observed between the groups, but hematologically, we detected a decrease in lymphocyte and neutrophil counts in both the antibiotic and lysate-treated groups. Immunologically, IL-6 remained elevated compared to the saline group, denoting the body's effort to compensate for bone loss due to bacterial action. Histopathologically, the results show more pronounced oral tissue regeneration in the antibiotic group and a reduced inflammatory reaction in the lysate group. We can conclude that the proposed bacterial lysate has similar effects to antibiotic therapy and can be considered an option in treating periodontitis, thus eliminating the unnecessary use of antibiotics.

Airway administration of OM-85, a bacterial lysate, blocks experimental asthma by targeting dendritic cells and the epithelium/IL-33/ILC2 axis.

BACKGROUND: Microbial interventions against allergic asthma have robust epidemiologic underpinnings and the potential to recalibrate disease-inducing immune responses. Oral administration of OM-85, a standardized lysate of human airways bacteria, is widely used empirically to prevent respiratory infections and a clinical trial is testing its ability to prevent asthma in high-risk children. We previously showed that intranasal administration of microbial products from farm environments abrogates experimental allergic asthma. OBJECTIVES: We sought to investigate whether direct administration of OM-85 to the airway compartment protects against experimental allergic asthma; and to identify protective cellular and molecular mechanisms activated through this natural route. METHODS: Different strains of mice sensitized and challenged with ovalbumin or Alternaria received OM-85 intranasally, and cardinal cellular and molecular asthma phenotypes were measured. Airway transfer experiments assessed whether OM-85-treated dendritic cells protect allergen-sensitized, OM-85-naive mice against asthma. RESULTS: Airway OM-85 administration suppressed allergic asthma in all models acting on multiple innate and adaptive immune targets: the airway epithelium/IL-33/ILC2 axis, lung allergen-induced type 2 responses, and dendritic cells whose Myd88/Trif-dependent tolerogenic reprogramming was sufficient to transfer OM-85-induced asthma protection. CONCLUSIONS: We provide the first demonstration that administering a standardized bacterial lysate to the airway compartment protects from experimental allergic asthma by engaging multiple immune pathways. Because protection required a cumulative dose 27- to 46-fold lower than the one reportedly active through the oral route, the efficacy of intranasal OM-85 administration may reflect its direct access to the airway mucosal networks controlling the initiation and development of allergic asthma.

The OM-85 bacterial lysate inhibits SARS-CoV-2 infection of epithelial cells by downregulating SARS-CoV-2 receptor expression.

BACKGROUND: Treatments for coronavirus disease 2019, which is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), are urgently needed but remain limited. SARS-CoV-2 infects cells through interactions of its spike (S) protein with angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) on host cells. Multiple cells and organs are targeted, particularly airway epithelial cells. OM-85, a standardized lysate of human airway bacteria with strong immunomodulating properties and an impeccable safety profile, is widely used to prevent recurrent respiratory infections. We found that airway OM-85 administration inhibits Ace2 and Tmprss2 transcription in the mouse lung, suggesting that OM-85 might hinder SARS-CoV-2/host cell interactions. OBJECTIVES: We sought to investigate whether and how OM-85 treatment protects nonhuman primate and human epithelial cells against SARS-CoV-2. METHODS: ACE2 and TMPRSS2 mRNA and protein expression, cell binding of SARS-CoV-2 S1 protein, cell entry of SARS-CoV-2 S protein-pseudotyped lentiviral particles, and SARS-CoV-2 cell infection were measured in kidney, lung, and intestinal epithelial cell lines, primary human bronchial epithelial cells, and ACE2-transfected HEK293T cells treated with OM-85 in vitro. RESULTS: OM-85 significantly downregulated ACE2 and TMPRSS2 transcription and surface ACE2 protein expression in epithelial cell lines and primary bronchial epithelial cells. OM-85 also strongly inhibited SARS-CoV-2 S1 protein binding to, SARS-CoV-2 S protein-pseudotyped lentivirus entry into, and SARS-CoV-2 infection of epithelial cells. These effects of OM-85 appeared to depend on SARS-CoV-2 receptor downregulation. CONCLUSIONS: OM-85 inhibits SARS-CoV-2 epithelial cell infection in vitro by downregulating SARS-CoV-2 receptor expression. Further studies are warranted to assess whether OM-85 may prevent and/or reduce the severity of coronavirus disease 2019.

[Efficacy and safety of bacterial lysate OM-85 in the treatment of uncomplicated acute respiratory infections: a double-blind, placebo-controlled, multicenter, randomized trial].

AIM: To evaluate the efficacy and safety of OM-85 in the treatment of uncomplicated acute respiratory infections (ARI) in adults. MATERIALS AND METHODS: A double-blind, placebo-controlled, multicenter, randomized trial included 556 patients (18-60 years old) with mild and moderate ARI and negative results of polymerase chain reaction analysis for SARS-CoV-2 RNA and rapid test for influenza A and B viruses. Patients were randomized into two groups: in the first group (n=278), patients received OM-85 (Broncho-munal®) one capsule 7 mg/day for 10 days, while the second group (n=278) was treated with placebo in the same regimen. The primary endpoint was the dynamics of the severity of symptoms over 3, 5, 7 and 10 days of treatment according to the 21-item Wisconsin Upper Respiratory Symptom Survey (WURSS-21), which was assessed by the area under the curve. Secondary efficacy criteria were the dynamics of the severity of symptoms according to the Common Cold Questionnaire (CCQ), the time to the resolution of symptoms according to WURSS-21 and CCQ, the proportion of patients with body temperature below 37°C on each day of treatment, frequency of the need for systemic antibacterial therapy. RESULTS: The superiority of OM-85 over placebo by primary endpoint was observed on the 5th, 7th and 10th days of treatment. OM-85 efficacy has also been proven by secondary criteria. OM-85 shortened the time until the symptoms of ARI resolved according to the WURSS-21 and CCQ, increased the proportion of patients with body temperature below 37°C by 2-9 days. The time needed to resolve the symptoms of disease in 20% of patients according to WURSS-21 was 7 and 9 days in patients taking OM-85 and placebo, respectively. Bacterial lysate increased the probability of complete disappearance of symptoms according to CCQ by 45.7% compared to placebo. The analysis of the frequency and severity of adverse events, laboratory tests, physical and instrumental examination results during treatment confirmed the good tolerability and safety of OM-85. CONCLUSION: The study confirmed the efficacy and safety of OM-85 in the complex treatment of ARI in adults.

Evaluation of a Gel Containing a Propionibacterium Extract in an In Vivo Model of Wound Healing.

Inappropriate wound healing (WH) management can cause significant comorbidities, especially in patients affected by chronic and metabolic diseases, such as diabetes. WH involves several different, partially overlapping processes, including hemostasis, inflammation, cell proliferation, and remodeling. Oxidative stress in WH contributes to WH impairment because of the overexpression of radical oxygen species (ROS) and nitrogen species (RNS). This study aimed to evaluate the in vitro antioxidative action of a gel containing a Propionibacterium extract (Emorsan® Gel) and assess its skin re-epithelialization properties in a mouse model of WH. The scavenging effects of the bacterial extract were assessed in vitro through the ABTS and DPPH assays and in L-929 murine fibroblasts. The effects of the Emorsan® Gel were studied in vivo in a murine model of WH. After WH induction, mice were treated daily with vehicle or Emorsan® Gel for 6 or 12 days. According to the in vitro tests, the Propionibacterium extract exerted an inhibitory effect on ROS and RNS, consequently leading to the reduction in malondialdehyde (MDA) and nitrite levels. Before proceeding with the in vivo study, the Emorsan® Gel was verified to be unabsorbed. Therefore, the observed effects could be ascribed to a local action. The results obtained in vivo showed that through local reduction of oxidative stress and inflammation (IL-1ß, TNF-α), the Emorsan® Gel significantly reduced the infiltration of mast cells into the injured wound, leading to the amelioration of symptoms such as itch and skin irritation. Therefore, the Emorsan® Gel improved the speed and percentage of wound area closure by improving the tissue remodeling process, prompting vascular-endothelial growth factor (VEGF) and transforming growth factor (TGF)- ß production and reducing the expression of adhesion molecules. Emorsan® Gel, by its ability to inhibit free radicals, could reduce local inflammation and oxidative stress, thus enhancing the speed of wound healing.

Bacterial Lysate Complex Administered Intranasally Suppresses Inflammation in an In Vivo Model of Aseptic Lymphadenitis.

The effect of a bacterial lysate complex IRS 19 on local and systemic manifestations of inflammation was studied in a rat model of aseptic lymphadenitis. Injection of λ-carrageenan into the cervical lymph node via surgical approach caused an increase in the thickness of the capsule, disturbances in histoarchitecture, the appearance of necrosis foci, histiocytosis of subcapsular and cerebral sinuses in the lymph node, as well as an increase in the level of TNFα in the blood serum and the number of circulating leukocytes and neutrophils. Course intranasal administration of bacterial lysate complex in this model dose-dependently reduced the level of these markers of the systemic inflammatory response and the severity of microstructural disorders in the affected lymph nodes. Thus, bacterial lysate complex after intranasal administration produces a systemic anti-inflammatory effect that goes beyond the respiratory tract.

Bacterial lysate treatment in allergic disease: A systematic review and meta-analysis.

OBJECTIVE: The aim of this review was to assess the efficacy of bacterial lysate treatment in patients with allergic disease. METHOD: Randomized controlled trials (RCTs) of bacterial lysate therapy for patients with allergic diseases (asthma, atopic dermatitis, and allergic rhinitis) were searched using PubMed, EMBASE, Cochrane, China National Knowledge Infrastructure, Chinese Biomedical literature, and Wanfang databases up to March 2020. Based on the guidelines of the Cochrane collaboration, risk of bias was assessed. RESULTS: This meta-analysis based on 19 studies comparing bacterial lysate-treated patients with a control group showed a 24% (RR: 1.24, 95% CI [1.19, 1.30]) increase in improvement of allergy symptom control. In addition, the improvement of asthma symptom control was 22% (RR: 1.22, 95% CI [1.14, 1.26]) higher in the bacterial lysate treatment group. Moreover, the levels of immunoglobulin (IgA and IgG), T lymphocyte subtype (CD3+, CD4+, CD4+/CD8+, Th1), and cytokines (IFN-γ, IL-2, and IL-12) were increased in the treated group compared with controls. There was no significant difference in adverse event rate between the two groups. CONCLUSION: Treatment with bacterial lysate improves symptom control in patients with allergic diseases on the basis of routine therapy. No adverse risk was found in this meta-analysis.

Screening Collagenase Activity in Bacterial Lysate for Directed Enzyme Applications.

Collagenases are essential enzymes capable of digesting triple-helical collagen under physiological conditions. These enzymes play a key role in diverse physiological and pathophysiological processes. Collagenases are used for diverse biotechnological applications, and it is thus of major interest to identify new enzyme variants with improved characteristics such as expression yield, stability, or activity. The engineering of new enzyme variants often relies on either rational protein design or directed enzyme evolution. The latter includes screening of a large randomized or semirational genetic library, both of which require an assay that enables the identification of improved variants. Moreover, the assay should be tailored for microplates to allow the screening of hundreds or thousands of clones. Herein, we repurposed the previously reported fluorogenic assay using 3,4-dihydroxyphenylacetic acid for the quantitation of collagen, and applied it in the detection of bacterial collagenase activity in bacterial lysates. This enabled the screening of hundreds of E. coli colonies expressing an error-prone library of collagenase G from C. histolyticum, in 96-well deep-well plates, by measuring activity directly in lysates with collagen. As a proof-of-concept, a single variant exhibiting higher activity than the starting-point enzyme was expressed, purified, and characterized biochemically and computationally. This showed the feasibility of this method to support medium-high throughput screening based on direct evaluation of collagenase activity.

[Clinical efficacy of mechanical bacterial lysate in the prevention of infectious exacerbations of chronic obstructive pulmonary disease].

AIM: To evaluate the efficacy of mechanical bacterial lysate on the prevention of infectious exacerbations of chronic obstructive pulmonary disease in patients with frequent exacerbations. MATERIALS AND METHODS: The study included patients (n=60) with frequent exacerbations of COPD (groups C and D according to the GOLD classification). All COPD patients were divided into two groups by blind method. The first group (n=30) received conventional therapy for COPD plus MBL (the course included 3 cycles of 10 days therapy with 20-day intervals between them). The second group of patients (control, n=30) received conventional therapy for COPD without MBL.We evaluated the severity of symptoms, frequency of recurrence of COPD exacerbations, readmissions, need for emergency care and changes in basic therapy of COPD. Evaluations were done on 10 days, 1, 3 and 6 months from the start of the study. RESULTS: Adding of MBL to the therapy list of COPD resulted in a significant decrease of biomarkers of systemic inflammation and sputum purulence during compared to the control group. After 6 months of observation MBL group demonstrated statistically significant improvement of respiratory function, decrease in frequency of COPD exacerbations, needs for emergency medical service, reduced changes in basic therapy and hospitalization for exacerbation of COPD. Therapy with MBL showed a high degree of safety and low incidence of adverse events. CONCLUSION: The results of the study indicate that MBL may be used for the prevention of severe infectious exacerbations of COPD.

A randomized, placebo-controlled, double-blinded, single-centre, phase IV trial to assess the efficacy and safety of OM-85 in children suffering from recurrent respiratory tract infections.

BACKGROUND: Over many years, OM-85, a lysate of 21 common bacterial respiratory pathogens, has been demonstrated to prevent respiratory recurrences in children. However, further studies are needed to explore the true importance of OM-85 in the prevention of respiratory tract infections (RTIs) in children. This study was planned to further contribute to the evaluation of the role played by OM-85 in prevention of recurrent RTIs in children. METHODS: This study was a randomized (3:3:1), placebo-controlled, double-blind, single-centre, phase IV trial carried out in Italy to assess the efficacy of OM-85 (Broncho-Vaxom®; Vifor Pharma; Meyrin 2/Geneva, Switzerland) in reducing the number of new RTI episodes in 288 children aged 1 to 6 years with a history of recurrent RTIs and to compare the efficacy of the standard 3-month regimen with that of administration of OM-85 for 6 months during a 6-month study period. RESULTS: The number of RTIs and of children who experienced at least one RTI were significantly lower among patients receiving OM-85 for 3 months than among those given placebo (33% vs 65.1%, p < 0.0001). Differences were statistically significant for upper RTIs (i.e., common cold/viral pharyngitis and acute otitis media; p < 0.0001 and p = 0.006, respectively). Days of absence from day-care for children and working days lost by parents were significantly lower in the group with children treated with OM-85 for 3 months than in the placebo group (p = 0.007 and p = 0.004, respectively). No difference was seen between children who received OM-85 for 3 and those who received OM-85 for 6 months. The prevalence of atopy as well as the history of recurrent wheezing and age of the study child did not influence the results. Benefit was maximally evident among children with a history of frequent recurrences. OM-85 was well tolerated and safe, even in children who received an influenza vaccination. CONCLUSIONS: The use of OM-85 for 3 months in 3 series of 10 consecutive days each time reduces the risk of recurrent RTIs in children, with a favourable safety profile. The greater effect observed in children prone to several respiratory episodes than in non-prone children seems to indicate that this lysate should be administered especially to children with a proven high susceptibility to RTIs.

Clinical and Immunological Benefits of OM-85 Bacterial Lysate in Patients with Allergic Rhinitis, Asthma, and COPD and Recurrent Respiratory Infections.

PURPOSE: The aim of this study was to evaluate the efficacy of OM-85 in reducing the incidence of respiratory tract infections (RTIs) in patients with allergic rhinitis, asthma, or chronic obstructive pulmonary disease (COPD), and its effect on immunological parameters, namely serum and secretory IgA levels. METHODS: This was an open-label, prospective, sequential study which included 84 consecutive patients aged 16-65 years, who presented with recurrent (three or more) respiratory infections during the year prior to study entry. In the first year of the study, patients received standard optimized care (SOC), according to their underlying disease condition (asthma, allergic rhinitis, or COPD). In the following year, patients received treatment with OM-85 oral bacterial lysate (one 7 mg capsule daily for ten consecutive days per month, for 3 months), with a 6-month follow-up. Medical history, clinical symptoms, serum, and secretory IgA levels, and the number of infections and exacerbations were evaluated before and after treatment. RESULTS: There was a decrease in the total number of RTIs before the OM-85 treatment period (SOC only) compared to the year before the study start [69/266 (corresponding to a 74 % reduction)] and an additional decrease [38/69 (corresponding to a 45 % reduction)] after OM-85 treatment; p < 0.05. There was also a significant reduction in the total number of exacerbations related to the patients' underlying medical conditions, which decreased from 55 to 35 during OM-85 (+SOC) treatment, corresponding to a reduction of 36 %. In addition, an increase in serum and secretory IgA levels which coincided with the administration of OM-85 was observed. CONCLUSIONS: Our results showed the clinical benefits of OM-85 in reducing RTIs and exacerbations of the underlying medical condition, in patients with allergic rhinitis, asthma, or COPD.

Sub-lingual administration of a polyvalent mechanical bacterial lysate (PMBL) in patients with moderate, severe, or very severe chronic obstructive pulmonary disease (COPD) according to the GOLD spirometric classification: A multicentre, double-blind, randomised, controlled, phase IV study (AIACE study: Advanced Immunological Approach in COPD Exacerbation).

Polyvalent mechanical bacterial lysates (PMBLs) have been shown to reduce the number of infectious episodes in patients with recurrent infections of the respiratory tract. Some previous investigations have also shown the effectiveness of PMBLs in reducing exacerbations of chronic obstructive pulmonary disease (COPD). The AIACE study, which was developed according to criteria of evidence-based medicine, evaluated whether the administration of PMBLs to COPD patients, in addition to the recommended treatment, was able to reduce the number of exacerbations by 25%. Two hundred eighty-eight patients with moderate to very severe COPD were recruited and randomly assigned to either placebo or PMBLs. The placebo or PMBLs were administered according to the standard scheme. The primary outcome of the study was not achieved. However, the number of days with fever (21 days per year versus 40.15; p < 0.001), the days of hospitalisation (65 days vs 162 days; p < 0.001), the interval between the first and second exacerbations (123.89 days vs 70.36; p = 0.03) and the number of days in poor health (109 days/year vs 171 days/year; p < 0.001) were significantly better in the PMBL group than in the placebo group. In conclusion, the results of this trials showed that Ismigen, in addition to guideline-suggested treatment, could not significantly reduce the number of exacerbations in the considered population; nevertheless, the secondary outcome results demonstrated potential benefits of this compound for relevant clinical outcomes.

The administration of a polyvalent mechanical bacterial lysate in elderly patients with COPD results in serological signs of an efficient immune response associated with a reduced number of acute episodes.

The administration of a polyvalent mechanical bacterial lysate (PMBL) in elderly patients with COPD has been shown to reduce the number of exacerbation. This is largely related to the involvement of cells belonging to the innate and the adaptive immune system (including dendritic cells, granulocytes, T and B lymphocytes and NK cells) that actively cooperate inducing the production of specific opsonizing antibodies directed to the antigens of PMBL. We have evaluated the production of antibodies directed to respiratory and systemic pathogens in a group of elderly COPD patients, recruited in a clinical trial, ancillary to a larger multicenter double blind, placebo-controlled, parallel-designed clinical trial in which patients were randomized to daily receive either PMBL or placebo. The treated group not only experienced a reduced number of seroconversion, but also, better controlled the number of infectious episodes and COPD exacerbations. It was thus evident that the administration of PMBL resulted not only effective in inducing the secretion of specific antibodies, but also effective in reducing the infectious episodes trough the potentiation of the antibody-mediated arm of the immune response.

Endotype and Phenotype Related Postoperative Effects of Bacterial Lysate in Chronic Rhinosinusitis.

OBJECTIVES: Chronic rhinosinusitis (CRS) endotypes have demonstrated clinical value in guiding treatment decisions. Bacterial lysates are immunomodulators that have shown beneficial effects in various respiratory inflammatory diseases. This study aimed to evaluate the effect of postoperative bacterial lysate therapy on different CRS endotypes. METHODS: Patients diagnosed with CRS who underwent endoscopic sinus surgery were recruited. Bacterial lysates were administered postoperatively for 10 days per month for 3 months to the experimental group comprising patients with a history of frequent upper respiratory infections without adverse reactions. The remaining participants were allocated to the control group. The results of the postoperative 3-, 6-, and 12-month assessments, including the modified Lund-Kennedy (mLK) endoscopic and Sinonasal Outcome Test (SNOT) 22 scores, for the groups were compared. The tissue samples obtained from the participants were evaluated to detect the presence of relevant inflammatory mediators. RESULTS: Among the 92 participants, 47 started bacterial lysate therapy 2 weeks after the surgery. The tissue cytokine profiles and clinical parameters, such as the disease severity and blood eosinophil percentage, of the bacterial lysate and control groups were comparable before treatment. The mLK endoscopic and SNOT-22 scores did not differ after 3, 6, and 12 months of follow-up. The subgroup analysis revealed that the bacterial lysate group had significantly lower mLK endoscopic scores than the control group for CRS without nasal polyps, while there was a tendency toward significance for the interleukin (IL)-5 negative group after 6 months. CONCLUSION: Postoperative bacterial lysate therapy has some beneficial effects on the endoscopic findings of patients with CRS without nasal polyps or those who are negative for IL-5.

The Use of Bacterial Lysate for the Prevention of Wheezing Episodes in Preschool Children: A Cost-Utility Analysis.

BACKGROUND: Although increasing recent evidence has shown the efficacy of bacterial lysate therapy for the prevention of wheezing episodes and asthma exacerbations in pediatric patients, evidence of its cost-effectiveness in preschool patients is scarce. OBJECTIVES: To evaluate the cost-utility of bacterial lysate therapy as an add-on to standard care of preschool children with recurrent wheezing. METHODS: To achieve the objectives of the study, we used a Markov simulation model with 3 mutually exclusive nonabsorbent states (regular Markov chain). Effectiveness parameters were obtained from a recent systematic review of the literature with meta-analyses (5 randomized controlled trials, 433 children). Cost data were obtained from hospital bills and from the national manual of drug prices in Colombia. The study was carried out from the perspective of the national health care system in Colombia. The main outcome of the model was quality-adjusted life-years. To assess the robustness of the model's results, we performed deterministic and probabilistic sensitivity analysis. RESULTS: Compared with standard care, bacterial lysate add-on therapy to standard care was associated with lower overall treatment costs (US $694.03 vs $830.71 average cost per patient) and the greatest gain in QALYs (0.9211 vs 0.9154 QALYs on average per patient), thus showing dominance. CONCLUSIONS: In Colombia, compared with standard care, bacterial lysate add-on therapy to standard care for treating preschool children with recurrent wheezing is a dominant strategy because it showed a greater gain in QALYs at lower total treatment costs.

Use of immunomodulatory therapy as part of comprehensive treatment of non-severe community-acquired pneumonia and its long-term results.

Background: This study investigates the efficiency of two different types of immunomodulators for the treatment of non-severe community-acquired pneumonia (CAP) and assesses their long-term effects. Methods: The study included 55 patients with non-severe CAP. Group 1 (control) received only standard CAP therapy; the other two groups received immunomodulators simultaneously with the standard therapy: bacterial lysate for group 2 and azoximer bromide (AzB) for group 3. TNF and IL-6 concentrations were determined on the day of hospitalization as well as on days 13 and 60 of follow-up. For 2 years, we monitored the incidence of low respiratory tract infections (LRTIs) in the same patients with CAP (n=55). Results: The overall duration of all symptoms was lower in the immunomodulator groups compared with the control group. During treatment, TNF and IL-6 concentrations decreased on days 13 and 60 in all patients; in patients who received immunomodulators, TNF and IL-6 were reliably lower than in control patients. IL-6 concentration decreased on day 60 in the bacterial lysate and AzB treatment groups and did not differ (p=0.72). The odds ratio for the development of LRTIs in the AzB group was 0.15 (0.02-0.93) (p=0.04), suggesting its protective effect. Conclusion: Inclusion of immunomodulators in the basic treatment of non-severe CAP reduces the duration of symptoms and is associated with improvement of the pro-inflammatory cytokine profile. In 2 years of follow-up, the long-term effects of the immunomodulatory therapy showed a statistically significant lower incidence of LRTIs in the AzB group only. However, given the small sample size of this study, further clinical studies are needed.

Antimicrobial activity of dietary supplements based on bacterial lysate of Lactobacillus rhamnosus DV.

Introduction: According to WHO, antibiotic resistance is increasing to hazardous levels worldwide. Candidiasis often occurs after taking antibiotics. Therefore, antibiotic resistance is a global problem and searching for antibacterial agents is necessary. Aim: To determine the antimicrobial activity of bacterial lysate of Lactobacillus (L.) rhamnosus DV separately and with plant extracts against bacterial and yeast test cultures. Material and methods: Antimicrobial activity of Del-Immune V® (cell wall and DNA fragments from a L. rhamnosus DV) separately and with cinnamon, beetroot, and blackcurrant extracts was determined by the minimum inhibitory concentration (MIC). Twofold serial dilutions determined the MIC in previously prepared meat-peptone broth (MPB) for bacteria and liquid wort for yeast. In the study, gram-negative (Escherichia coli IEM-1, Proteus vulgaris PА-12, Pseudomonas sp. MI-2, L. rhamnosus 13/2) and gram-positive (Bacillus (B.) subtilis BТ-2, Staphylococcus aureus BМС-1) bacteria, as well as yeast (Candida (C.) albicans D-6, C. tropicalis PE-2, C. utilis BVS-65) were used as test cultures. Results: The MIC for the studied bacterial test cultures after application of L. rhamnosus DV bacterial lysates was from 1.0 ± 0.05 mg/mL to 12.5 ± 0.63 mg/mL, which was significantly less than that of the thermally inactivated control (MIC from 125.0 ± 6.25 mg/mL to 250.0 ± 12.5 mg/mL). B. subtilis BT-2 culture was the least sensitive to the action of the bacterial lysate (MIC-12.5 ± 0.63 mg/mL). It showed the best antibacterial and antifungal effect bacterial lysate with the phytonutrient blackcurrant. Conclusions: It was demonstrated that bacterial lysate of lactic acid bacteria L. rhamnosus DV exhibits antibacterial and antifungal properties during direct contact with pathogenic agents.

Effect of immunostimulation with bacterial lysate on the clinical course of allergic rhinitis and the level of γδT, iNKT and cytotoxic T cells in children sensitized to grass pollen allergens: A randomized controlled trial.

Background: There are many drugs for allergic rhinitis (AR), however, these drugs show variable clinical effectiveness and some side effects. Therefore, new methods of AR pharmacotherapy are being sought. Objectives: The objectives of this study were to evaluate the efficacy of polyvalent mechanical bacterial lysate (PMBL) therapy in improving the clinical course of grass pollen-induced AR (seasonal AR, SAR) in children and its effect on changes in the blood level of the γδT, iNKT and cytotoxic T cell subsets. Methods: Fifty children with SAR were enrolled in this study and were randomly assigned to either the PMBL group or the placebo group. The severity of SAR symptoms was assessed using the total nasal symptom score (TNSS) and visual analogue scale (VAS). During two visits (V1, V2), peak nasal inspiratory flow (PNIF) was measured and peripheral blood was collected for immunological analyses. The study also included 2 telephone contacts (TC1, TC2). Results: The severity of the nasal symptoms of SAR on the TNSS scale was revealed to have a significantly lower impact in the PMBL group vs the placebo group at measuring points TC1 and V2 (p = 0.01, p = 0.009, respectively). A statistically significantly lower mean severity of nasal symptoms of SAR on the VAS scale was recorded for children in the PMBL group compared to the placebo group at measuring points TC1, V2 and TC2 (p = 0.04, p = 0.04, p = 0.03, respectively). The compared groups do not show significant differences in terms of PNIF values at individual measuring points. There were no statistically significant changes in immune variables. For both groups, there was a statistically significant association between the level of Th1-like γδT cells and the severity of SAR symptoms expressed on the TNSS scale (p = 0.03) - the lower the level of Th1-like γδT cells, the higher the TNSS value. Conclusion: Administration of sublingual PMBL tablets during the grass pollen season proves to have a high efficacy in alleviating SAR symptoms in children sensitized to grass pollen allergens. Th1-like γδT cells may be used as potential markers for SAR severity in children. Clinical trial registration: ClinicalTrials.gov, identifier (NCT04802616).