Aluminium oxide nanoparticles inhibit EPS production,adhesion and biofilm formation by multidrug resistant Acinetobacter baumannii

Abstract

Acinetobacter baumannii is a biofilm forming multidrug resistant (MDR) pathogen responsible for respiratory tract infections. In this study, aluminium oxide nanoparticles (Al₂O₃ NPs) were synthesized and characterized by TEM and EDX and shown to be spherical shaped nanoparticles with a diameter < 10?nm. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) for the Al₂O₃ NPs ranged between 125 and 1,000?µg ml^?1. Exposure to NPs caused cellular membrane disruption, indicated by an increase in cellular leakage of the contents. Biofilm inhibition was 11.64 to 70.2%, whereas attachment of bacteria to polystyrene surfaces was reduced to 48.8 to 51.9% in the presence of NPs. Nanoparticles also reduced extracellular polymeric substance production and the biomass of established biofilms. The data revealed the non-toxic nature of Al₂O₃ NPs up to a concentrations of 120?µg ml^?1 in HeLa cell lines. These results demonstrate an effective and safer use of Al₂O₃ NPs against the MDR A. baumannii by targeting biofilm formation, adhesion and EPS production.     

Domain swapping reveals complement control protein modules critical for imparting cofactor and decay-accelerating activities in vaccinia virus complement control protein

Vaccinia virus encodes a structural and functional homolog of human complement regulators named vaccinia virus complement control protein (VCP). This four-complement control protein domain containing secretory protein is known to inhibit complement activation by supporting the factor I-mediated inactivation of complement proteins, proteolytically cleaved form of C3 (C3b) and proteolytically cleaved form of C4 (C4b) (termed cofactor activity), and by accelerating the irreversible decay of the classical and to a limited extent of the alternative pathway C3 convertases (termed decay-accelerating activity [DAA]). In this study, we have mapped the VCP domains important for its cofactor activity and DAA by swapping its individual domains with those of human decay-accelerating factor (CD55) and membrane cofactor protein (MCP; CD46). Our data indicate the following: 1) swapping of VCP domain 2 or 3, but not 1, with homologous domains of decay-accelerating factor results in loss in its C3b and C4b cofactor activities; 2) swapping of VCP domain 1, but not 2, 3, or 4 with corresponding domains of MCP results in abrogation in its classical pathway DAA; and 3) swapping of VCP domain 1, 2, or 3, but not 4, with homologous MCP domains have marked effect on its alternative pathway DAA. These functional data together with binding studies with C3b and C4b suggest that in VCP, domains 2 and 3 provide binding surface for factor I interaction, whereas domain 1 mediates dissociation of C2a and Bb from the classical and alternative pathway C3 convertases, respectively.     

Myocardial infarction and intramyocardial injection models in swine

Sustainable and reproducible large animal models that closely replicate the clinical sequelae of myocardial infarction (MI) are important for the translation of basic science research into bedside medicine. Swine are well accepted by the scientific community for cardiovascular research, and they represent an established animal model for preclinical trials for US Food and Drug Administration (FDA) approval of novel therapies. Here we present a protocol for using porcine models of MI created with a closed-chest coronary artery occlusion-reperfusion technique. This creates a model of MI encompassing the anteroapical, lateral and septal walls of the left ventricle. This model infarction can be easily adapted to suit individual study design and enables the investigation of a variety of possible interventions. This model is therefore a useful tool for translational research into the pathophysiology of ventricular remodeling and is an ideal testing platform for novel biological approaches targeting regenerative medicine. This model can be created in approximately 8-10 h.     

Impact of sequential exposure of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> and imidacloprid against susceptible and resistant strains of <Emphasis Type="Italic">Musca domestica</Emphasis>

Insecticide resistance in the housefly Musca domestica is hampering pest management. However, entomopathogens, possibly in combination with insecticides, may have control potential against resistant houseflies. This study investigates the combination of the entomopathogenic fungus Beauveria bassiana and the neonicotinoid insecticide, imidacloprid against a susceptible and a resistant housefly strain, respectively under laboratory conditions. The fungus and insecticide were tested alone and in combinations at LC₃₀. Significant and synergistic interactions between B. bassiana and imidacloprid were observed with increased mortality rates of the combined treatment as compared to individual treatment in housefly strains 772a (susceptible) and 766b (resistant). Significant differences in the GST and P450 activities for both strains were found. Female 766b flies caused 15- to 237-fold increases in gene expression of xenobiotic response genes for B. bassiana and 23- to 120-fold changes for imidacloprid. The combination of B. bassiana and imidacloprid caused significant synergistic interaction when applied against two housefly strains irrespective of order of application. The effect was highest when the insecticide was applied first. The resistant housefly strain had elevated detoxification enzymes and higher expression of detoxification genes, but showed the same level of susceptibility to the combined fungus/insecticide treatment as the susceptible strain.     

Pomegranate (Punica granatum) purified polyphenol extract inhibits influenza virus and has a synergistic effect with oseltamivir

Influenza epidemics cause numerous deaths and millions of hospitalizations each year. Because of the alarming emergence of resistance to anti-influenza drugs, there is a need to identify new naturally occurring antiviral molecules. We tested the hypothesis that pomegranate polyphenol extract (PPE) has anti-influenza properties. Using real time PCR, plaque assay, and TCID 50% hemagglutination assay, we have shown that PPE suppresses replication of influenza A virus in MDCK cells. PPE inhibits agglutination of chicken red blood cells (cRBC) by influenza virus and is virucidal. The single-cycle growth conditions indicated that independent of the virucidal effect PPE also inhibits viral RNA replication. PPE did not alter virus ribonucleoprotein (RNP) entry into nucleus or translocation of virus RNP from nucleus to cytoplasm in MDCK cells. We evaluated four major Polyphenols in PPE (ellagic acid, caffeic acid, luteolin, and punicalagin) and demonstrated that punicalagin is the effective, anti-influenza component of PPE. Punicalagin blocked replication of the virus RNA, inhibited agglutination of chicken RBC's by the virus and had virucidal effects. Furthermore, the combination of PPE and oseltamivir synergistically increased the anti-influenza effect of oseltamivir. In conclusion, PPE inhibited the replication of human influenza A/Hong Kong (H3N2) in vitro. Pomegranate extracts should be further studied for therapeutic and prophylactic potential especially for influenza epidemics and pandemics.     

In vitro antibiofilm and anti‐adhesion effects of magnesium oxide nanoparticles against antibiotic resistant bacteria

The aim of the current investigation was to determine the antibacterial and antibiofilm potential of MgO nanoparticles (NPs) against antibiotic‐resistant clinical strains of bacteria. MgO NPs were synthesized by a wet chemical method and further characterized by scanning electron microscopy and energy dispersive X‐ray. Antibacterial activity was determined by broth microdilution and agar diffusion methods. The Bradford method was used to assess cellular protein leakage as a result of loss of membrane integrity. Microtiter plate assay following crystal violet staining was employed to determine the effect of MgO NPs on biofilm formation and removal of established biofilms. MIC values ranged between 125 and 500 μg/mL. Moreover, treatment with MgO NPs accelerated rate of membrane disruption, measured as a function of leakage of cellular proteins. Leakage of cellular protein content was greater among gram‐negative bacteria. Cell adherence assay indicated 25.3–49.8% inhibition of bacterial attachment to plastic surfaces. According to a static biofilm method, MgO NPs reduced biofilm formation potential from 31% to 82.9% in a time‐dependent manner. Moreover, NPs also significantly reduced the biomass of 48, 72, 96 and 120 hr old biofilms (P < 0.05). Cytotoxicity experiments using a neutral red assay revealed that MgO NPs are non‐toxic to HeLa cells at concentrations of 15–120 μg/mL. These data provide in vitro scientific evidence that MgO NPs are effective and safe antibiofilm agents that inhibit adhesion, biofilm formation and removal of established biofilms of multidrug‐resistant bacteria.

     

Visualization of grapevine root colonization by the Saharan soil isolate Saccharothrix algeriensis NRRL B-24137 using DOPE-FISH microscopy

Background and aim

There is currently a gap of knowledge regarding whether some beneficial bacteria isolated from desert soils can colonize epi- and endophytically plants of temperate regions. In this study, the early steps of the colonization process of one of these bacteria, Saccharothrix algeriensis NRRL B-24137, was studied on grapevine roots to determine if this beneficial strain can colonize a non-natural host plant. An improved method of fluorescence in situ hybridization (FISH), the double labeling of oligonucleotide probes (DOPE)-FISH technique was used to visualize the colonization behavior of such bacteria as well as to determine if the method could be used to track microbes on and inside plants.

Methods

A probe specific to Saccharothrix spp. was firstly designed. Visualization of the colonization behavior of S. algeriensis NRRL B-24137 on and inside roots of grapevine plants was then carried out with DOPE-FISH microscopy.

Results

The results showed that 10 days after inoculation, the strain could colonize the root hair zone, root elongation zone, as well as root emergence sites by establishing different forms of bacterial structures as revealed by the DOPE-FISH technique. Further observations showed that the strain could be also endophytic inside the endorhiza of grapevine plants.

Conclusions

Taking into account the natural niches of this beneficial strain, this study exemplifies that, in spite of its isolation from desert soil, the strain can establish populations as well as subpopulations on and inside grapevine plants and that the DOPE-FISH tool can allow to detect it.     

Microbiota analysis revealed vertical transmission and microbial adjusting function of symbiotic fungus in the attelabid weevil fungiculture

<正>Dear Editor,Microbial symbionts are crucial participants in the life history of fungus-farming insects (Mueller et al., 2005;Youngsteadt, 2008). The attelabid weevil (Euops chinensis)feeding on Fallopia japonica in southern China, has evolved a special proto-farming bipartite symbiosis with the fungus(Penicillium herquei), which produces antibiotic (+)-scleroderolide in the leaf-rolls ("cradle" for offspring development) to protect the offspring from potential microbial parasites and pathogens (Sakurai, 1985; Wang et al., 2015).     

The Saharan isolate Saccharothrix algeriensis NRRL B-24137 induces systemic resistance in Arabidopsis thaliana seedlings against Botrytis cinerea

Background and aim

Saccharothrix algeriensis NRRL B-24137, isolated from a Saharan soil, has been described as a potential biocontrol agent against Botrytis cinerea and other phytopathogens. However, the plant protection mechanisms involved still need to be described. The aim of this study was to determine this protection phenomenon as well as parts of the mechanisms involved, using Arabidopsis thaliana seedlings and B. cinerea.

Methods

The bacterial colonization process was evaluated on A. thaliana seedlings using fluorescence in situ hybridization. Protection of A. thaliana seedlings inoculated with NRRL B-24137 against B. cinerea was then evaluated. Parts of the mechanisms involved in the systemic protection against B. cinerea were evaluated using known mutants of genes involved in jasmonate (JA)/ethylene (ET)/salicylic acid (SA) signaling. Other Arabidopsis mutants, AtrhbohD-3, AtrhbohF-3, and ups1-1 were also screened to determine other parts of the mechanisms involved.

Results

The results showed that the strain NRRL B-24137 colonized, epi- and endophytically, the roots of Arabidopsis seedlings but the strain was not a systemic colonizer during the time of the experiment. The strain NRRL B-24137 also reduced B. cinerea symptoms and the protection was linked to known mechanisms of induced systemic resistance (ISR; JA/ET signaling), as well as to functionality of AtrbohF oxidase and of UPS1. Crosstalk between ET/JA and SA signaling could also be involved.

Conclusions

The isolate NRRL B-24137, after colonizing the root systems of A. thaliana, induces an ISR against B. cinerea, which is JA/ET dependent, but could also require SA crosstalk and protection could also require NAPDH oxidases and UPS1 functionalities.