Antimicrobial and toxicological evaluation of Origanum vulgare: an in vivo study

Origanum vulgare has been of great interest in academia and pharma industry due to its antioxidant, antifungal and antitumor properties. The present study aimed to find the anti-MRSA potential and in vivo toxicity assessments of O. vulgare. O. vulgare extract was used to monitor anti-MRSA activity in mice. Following MRSA established infection in mice (Mus musculus), treatment with O. vulgare was continued for 7 days. Autopsies were performed and re-isolation, gross lesion scoring and bacterial load in various organs were measured. Additionally, blood sample was analysed for hematological assays. Toxicity assessment of O. vulgare potential as medicine was done at 200 mg/kg and 400 mg/kg by evaluating liver and kidney functions. Bacterial load and gross lesion in lungs and heart were significantly low compared to positive control following O. vulgare treatment. Likewise, O. vulgare treated groups had hematological, neutrophil and TLC values similar to control groups. Increased AST, ALP and total bilirubin alongwith marked hepatocellular degeneration and distortion around the central vein, inflammatory cell infiltration, and cytoplasmic vacuolization of hepatic cells was observed at higher dose. It is concluded that crude extract of O. vulgare may contain beneficial secondary metabolites and in future may be explored for curing infectious diseases.

Isolation, antimicrobial susceptibility patterns, and risk factors assessment of non-typhoidal Salmonella from apparently healthy and diarrheic dogs

Background Dogs are one of the important asymptomatic carriers of antimicrobial resistant and potentially pathogenic strains of Salmonella. They can harbor large bacterial load in the intestines and mesenteric lymph nodes which can be shed in their feces with the possibility of transmission to humans. Therefore, a cross-sectional study was conducted with the objectives of estimating the prevalence of non-typhoidal Salmonella, assessing the risk factors for dog’s Salmonella carriage, and profiling the antimicrobial resistance pattern of Salmonella isolates among housed dogs in Harar town, Eastern Ethiopia. A total of 415 rectal swab samples were collected from randomly selected dogs. Samples were examined for non-typhoidal Salmonella using standard bacteriologic culture and biochemical tests. The disk diffusion method (Kirby-Bauer test) was employed to evaluate the isolates for their susceptibility against five antimicrobials. Results Non-typhoidal Salmonella were isolated from 26 (6.3%) of the rectal swab samples, with significantly higher occurrence in diarrheic (15.2%) than non-diarrheic (5.5%) dogs. The risk of Salmonella harboring was significantly higher in female dogs than in male dogs (OR = 2.5, p = 0.027). Dogs fecal shedding of Salmonella was relatively higher in households who used offal as a main feed type for their dogs (23.1%; 95% CI = 5–53.8) than those who used leftover food (10.1%; 95% CI = 5.7–16.1) and practiced mixed feeding system (17%; 95% CI = 7.6–30.8). Salmonella isolates showed higher resistance to ampicillin (41.7%), while all isolates were fully susceptible to gentamicin. Moreover, 58.3% of Salmonella isolates showed resistance to at least one of the tested antimicrobials. Majorities (72.7%) of the dog owners had no awareness on the risk of zoonotic salmonellosis from dog and all of the respondents use bare hand to clean dog kennel. Conclusion Our study reveals the importance of both diarrheic and apparently healthy housed dogs in the harboring and shedding of antimicrobial resistant non-typhoidal Salmonella. The risk of non-typhoidal Salmonella spread among pet owners is not negligible, especially in households who use offal as main feed type. Therefore, an integrated approach such as: proper dog handling practices; continuous evaluation of antimicrobial resistance; and rational use of antimicrobials in the field of veterinary sector are necessary to tackle the problem.

Human Amnion-Derived Mesenchymal Stromal Cells: A New Potential Treatment for Carbapenem-Resistant Enterobacterales in Decompensated Cirrhosis

Background: Spontaneous bacterial peritonitis (SBP) is a severe and often fatal infection in patients with decompensated cirrhosis and ascites. The only cure for SBP is antibiotic therapy, but the emerging problem of bacterial resistance requires novel therapeutic strategies. Human amniotic mesenchymal stromal cells (hA-MSCs) possess immunomodulatory and anti-inflammatory properties that can be harnessed as a therapy in such a context. Methods: An in vitro applications of hA-MSCs in ascitic fluid (AF) of cirrhotic patients, subsequently infected with carbapenem-resistant Enterobacterales, was performed. We evaluated the effects of hA-MSCs on bacterial load, innate immunity factors, and macrophage phenotypic expression. Results: hA-MSCs added to AF significantly reduce the proliferation of both bacterial strains at 24 h and diversely affect M1 and M2 polarization, C3a complement protein, and ficolin 3 concentrations during the course of infection, in a bacterial strain-dependent fashion. Conclusion: This study shows the potential usefulness of hA-MSC in treating ascites infected with carbapenem-resistant bacteria and lays the foundation to further investigate antibacterial and anti-inflammatory roles of hA-MSC in in vivo models.

Characterization of phage resistance and phages capable of intestinal decolonization of carbapenem-resistant Klebsiella pneumoniae in mice

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has emerged as a severe global health challenge. We isolate and characterize two previously unidentified lytic phages, P24 and P39, with large burst sizes active against ST11 KL64, a major CRKP lineage. P24 and P39 represent species of the genera Przondovirus (Studiervirinae subfamily) and Webervirus (Drexlerviridae family), respectively. P24 and P39 together restrain CRKP growth to nearly 8 h. Phage-resistant mutants exhibit reduced capsule production and decreased virulence. Modifications in mshA and wcaJ encoding capsule polysaccharide synthesis mediate P24 resistance whilst mutations in epsJ encoding exopolysaccharide synthesis cause P39 resistance. We test P24 alone and together with P39 for decolonizing CRKP using mouse intestinal colonization models. Bacterial load shed decrease significantly in mice treated with P24 and P39. In conclusion, we report the characterization of two previously unidentified lytic phages against CRKP, revealing phage resistance mechanisms and demonstrating the potential of lytic phages for intestinal decolonization.

High Mycobacterium tuberculosis Bacillary Loads Detected by Tuberculosis Molecular Bacterial Load Assay in Patient Stool: a Potential Alternative for Nonsputum Diagnosis and Treatment Response Monitoring of Tuberculosis

This paper highlights the value of stool as a sample type for diagnosis of tuberculosis. While other studies have used DNA-based assays like the Xpert MTB/RIF and culture to detect Mycobacterium tuberculosis in stool, this is the first study that has applied TB-MBLA, an RNA-based assay, to quantify TB bacteria in stool.

Investigation of the Potential of Heterophil/Lymphocyte Ratio as a Biomarker to Predict Colonization Resistance and Inflammatory Response to Salmonella enteritidis Infection in Chicken

Salmonella causes significant economic loss to the poultry industry and represents a real threat to human health. The region of difference 21 (ROD21) pathogenicity island removal is a genetic mechanism by which Salmonellaenteritidis (SE) invades the intestinal epithelium and induces systemic infection in mice. The heterophil/lymphocyte (H/L) ratio reflects the chicken’s robustness and immune system status. The H/L ratio is considered a disease resistance trait, and it could be used as a marker for selecting Salmonella resistance in live chickens. However, the association of the H/L ratio with Salmonella resistance and the inflammatory response remains to be elucidated. Moreover, the kinetics of ROD21 excision in the intestine and immune organs of chickens is unknown. Therefore, this study aimed to investigate the bacterial load, the ROD21 excision, the IL-1β, IL-8, and INF-γ blood serum concentration kinetics, and the association with the H/L ratio in chicken at 1, 3, 7, and 21 days post-SE infection. The results showed a significant correlation between the H/L ratio and the bacterial load in the ileum and caecum at 7 dpi. The ROD21 pathogenicity island absolute and relative excision in the caecum were positively correlated at 1 dpi but negatively correlated at 7 dpi with the H/L ratio. However, in the liver, we found the opposite tendency. The association of the H/L ratio with IL-1β, IL-8, and INF-γ blood serum concentrations showed that a low H/L ratio is correlated with increased IL-1β and INF-γ at 21 dpi. This study confirmed that the H/L ratio is associated with robustness and Salmonella-resistance in chicken. The methodology used in this study can separate individuals into susceptible and resistant and can help in the selection and breeding of Salmonella-resistant chickens.

Fusobacterium nucleatum and Bacteroides fragilis detection in colorectal tumours: Optimal target site and correlation with total bacterial load

Background Mucosal infiltration by certain bacterial species may contribute to the development and progression of colorectal cancer (CRC). There is considerable variation in reported detection rates in human CRC samples and the extent to which bacterial infiltration varies across regions of the primary tumour is unknown. This study aimed to determine if there is an optimal site for bacterial detection within CRC tumours. Methods Presence of target bacterial species was assessed by quantitative real-time PCR (qPCR) in 42 human CRC tumours. Abundance in primary tumour regions, normal epithelium and at metastatic sites was investigated in an expanded cohort of 51 patients. Species presence/absence was confirmed by diversity profiling in five patients. Correlation with total bacterial load and clinicopathological features was assessed. Results Fusobacterium nucleatum and Bacteroides fragilis were detected in tumours from 43% and 24% of patients, respectively (17% positive for both species). The optimal detection site was the tumour luminal surface (TLS). Patients testing positive at the TLS frequently tested negative at other sites, including central tumour and invasive margin. F. nucleatum was detected at a higher frequency in tumour versus normal epithelium (p < 0.01) and was associated with more advanced disease (p = 0.01). Detection of both species correlated with total bacterial load. However, corroboration of qPCR results via diversity profiling suggests detection of these species may indicate a specific microbial signature. Conclusions This study supports a role for F. nucleatum in CRC development. Presence of F. nucleatum and B. fragilis varies across primary tumour regions, with the TLS representing the optimal site for bacterial detection.

Treatment of wound infections in a mouse model using Zn2+-releasing phage bound to gold nanorods

Infections caused by drug-resistant bacteria, particularly gram-negative organisms, are increasingly difficult to treat using antibiotics. A potential alternative is phage therapy, in which phages infect and lyse the bacterial host. However, phage therapy poses serious drawbacks and safety concerns, such as the risk of genetic transduction of antibiotic resistance genes, inconsistent pharmacokinetics, and unknown evolutionary potential. In contrast, metallic nanoparticles possess precise, tunable properties, including efficient conversion of electronic excitation into heat. In this work, we demonstrate that engineered phage-nanomaterial conjugates that target the gram-negative pathogen P. aeruginosa, are highly effective as a treatment of infected wounds in mice. Photothermal heating, performed as a single treatment (15 min) or as two treatments on consecutive days, rapidly reduced the bacterial load and released Zn2+ to promote wound healing. The phage-nanomaterial treatment was significantly more effective than systemic fluoroquinolone antibiotics in reducing both bacterial load and wound size, and was notably effective against a P. aeruginosa strain resistant to polymyxins, a last-line antibiotic therapy. Unlike these antibiotics, the phage-nanomaterial showed no detectable toxicity or systemic effects in mice, consistent with the short duration and localized nature of phage-nanomaterial treatment. Our results demonstrate that phage therapy controlled by inorganic nanomaterials can be a safe and effective antimicrobial strategy in vivo.

Natural Killer Cells Regulate Pulmonary Macrophages Polarization in Host Defense Against Chlamydial Respiratory Infection

NK cells and pulmonary macrophages both are important components of innate immunity. The interaction between NK cells and pulmonary macrophages during chlamydial infection is poorly understood. In this study, we explored the effect of NK cells on regulation of pulmonary macrophage function during chlamydial respiratory infection. We found that NK depletion led to polarization of pulmonary macrophages from M1 to M2 phenotype, and it is related to reduced miR-155 expression in lung macrophage. Using adoptive transfer approach, we found that the recipients receiving lung macrophages isolated from C. muridarum-infected NK-cell-depleted mice exhibited an increased bacterial load and severe inflammation in the lung upon chlamydial challenge infection when compared with the recipients of lung macrophages from infected isotype control antibody treated mice. Herein, the effects of NK cells on macrophage polarization were examined in vitro. We found that NK cells from chlamydial-infected mice (iNK) significantly induced M1 polarization compared to that from uninfected mice (uNK). Inhibition of miR-155 expression in macrophages reduced M1 polarization induced by iNK, while miR-155 over-expression enhanced it. Furthermore, neutralization of IFN-γ in the coculture system decreased the expression of miR-155 by macrophages, and resulted in weakened M1 polarization. The data indicates that NK cells promote M1 polarization through up-regulation of miR-155 in macrophages by producing IFN-γ during chlamydial infection, and NK-regulated macrophage polarization is functionally relevant to host defense against the infection.

Tertiary hospital sewage as reservoir of bacteria expressing MDR phenotype in Brazil

High doses of antibiotics used in hospitals can affect the microbial composition of sewers, selecting resistant bacteria. In this sense, we evaluated the antibiotic resistance profile and the multiresistant phenotype of bacteria isolated in sewage from a tertiary hospital in the interior São Paulo state, Brazil. For bacteria isolation, 10 µL of sewage samples were sown in selective culture media and the isolates were identified using VITEK-2 automatized system. The antibiotic sensitivity test was performed by disk diffusion. High percentages of resistance were found for amoxicillin, ampicillin, ceftazidime, clindamycin, vancomycin and the multidrug-resistant phenotype (MDR) was attributed to 60.7% of the isolates. Our results show bacteria classified as critical/high priority by WHO List of Priority Pathogens (Enterococcus and Staphylococcus aureus resistant to vancomycin and Enterobacteriaceae resistant to carbapenems) in hospital sewage. Therefore, the implementation of disinfection technologies for hospital sewage would reduce the bacterial load in the sewage that will reach urban wastewater treatment plants, minimizing superficial water contamination and bacterial resistance spread in the environment.