scholarly journals Ethidium Binding to Salmonella enterica ser. Typhimurium Cells and Salmon Sperm DNA

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3386
Author(s):  
Sandra Sakalauskaitė ◽  
Valeryia Mikalayeva ◽  
Rimantas Daugelavičius
Keyword(s):  
Salmonella Enterica ◽  
Bacterial Resistance ◽  
Polymyxin B ◽  
Medium Composition ◽  
High Ionic Strength ◽  
Factors Affecting ◽  
Sperm Dna ◽  
Spectrofluorimetric Analysis ◽  
Assay Conditions ◽  
Salmon Sperm Dna

Bacterial resistance to antibiotics due to increased efficiency of the efflux is a serious problem in clinics of infectious diseases. Knowledge of the factors affecting the activity of efflux pumps would help to find the solution. For this, fast and trustful methods for efflux analysis are needed. Here, we analyzed how the assay conditions affect the accumulation of efflux indicators ethidium (Et+) and tetraphenylphosphonium in Salmonella enterica ser. Typhimurium cells. An inhibitor phenylalanyl-arginyl-β-naphtylamide was applied to evaluate the input of RND family pumps into the total efflux. In parallel to spectrofluorimetric analysis, we used an electrochemical assessment of Et+ concentration. The results of our experiments indicated that Et+ fluorescence increases immediately after the penetration of this indicator into the cells. However, when cells bind a high amount of Et+, the intensity of the fluorescence reaches the saturation level and stops reacting to the accumulated amount of this indicator. For this reason, electrochemical measurements provide more trustful information about the efficiency of efflux when cells accumulate high amounts of Et+. Measurements of Et+ interaction with the purified DNA demonstrated that the affinity of this lipophilic cation to DNA depends on the medium composition. The capacity of DNA to bind Et+ considerably decreases in the presence of Mg2+, Polymyxin B or when DNA is incubated in high ionic strength media.

scholarly journals Ethidium binding to Salmonella enterica ser. Typhimurium Cells and Salmon sperm DNA

2021 ◽  
Author(s):  
Sandra Sakalauskaite ◽  
Valeryia Mikalayeva ◽  
Rimantas Daugelavičius
Keyword(s):  
Salmonella Enterica ◽  
Bacterial Resistance ◽  
Polymyxin B ◽  
Medium Composition ◽  
High Ionic Strength ◽  
Factors Affecting ◽  
Sperm Dna ◽  
Spectrofluorimetric Analysis ◽  
Assay Conditions ◽  
Salmon Sperm Dna

Bacterial resistance to antibiotics due to an increased efficiency of the efflux is a serious problem in clinics of infectious diseases. Knowledge of the factors affecting the activity of efflux pumps would help to find the solution. For this, fast and trustful methods for the efflux analysis are needed. Here we analyzed how the assay conditions affect the accumulation of efflux indicators ethidium (Et+) and tetraphenylphosphonium in Salmonella enterica ser. Typhimurium cells. An inhibitor phenyl-alanyl-arginyl-β-naphtylamide was applied to evaluate the input of RND family pumps into the total efflux. In parallel to spectrofluorimetric analysis, we used an electrochemical assessment of Et+ concentration. Results of our experiments indicated that Et+ fluorescence increases immediately after the penetration of this indicator into the cells. However, when cells bind a high amount of Et+, intensity of the fluorescence reaches the saturation level and stops reacting to the accumulated amount of this indicator. For this reason, electrochemical measurements provide more trustful information about the efficiency of efflux when cells accumulate high amounts of Et+. Measure-ments of Et+ interaction with the purified DNA demonstrated that affinity of this lipophilic cation to DNA depends on the medium composition. The capacity of DNA to bind Et+ considerably de-creases in presence of Mg2+, Polymyxin B or when DNA is incubated in high ionic strength media.

scholarly journals Release of the Lipopolysaccharide Deacylase PagL from Latency Compensates for a Lack of Lipopolysaccharide Aminoarabinose Modification-Dependent Resistance to the Antimicrobial Peptide Polymyxin B in Salmonella enterica

2007 ◽  
Vol 189 (13) ◽  
pp. 4911-4919 ◽  
Author(s):  
Kiyoshi Kawasaki ◽  
Kotaro China ◽  
Masahiro Nishijima
Keyword(s):  
Salmonella Enterica ◽  
Lipid A ◽  
Bacterial Resistance ◽  
Biological Significance ◽  
Regulatory System ◽  
Polymyxin B ◽  
Expression Plasmid ◽  
Loss Of Resistance ◽  
Anionic Charge

ABSTRACT Salmonella enterica modifies its lipopolysaccharide (LPS), including the lipid A portion, to adapt to its environments. The lipid A 3-O-deacylase PagL exhibits latency; deacylation of lipid A is not usually observed in vivo despite the expression of PagL, which is under the control of a two-component regulatory system, PhoP-PhoQ. In contrast, PagL is released from latency in pmrA and pmrE mutants, both of which are deficient in aminoarabinose-modified lipid A, although the biological significance of this is not clear. The attachment of aminoarabinose to lipid A decreases the net anionic charge at the membrane's surface and reduces electrostatic repulsion between neighboring LPS molecules, leading to increases in bacterial resistance to cationic antimicrobial peptides, including polymyxin B. Here we examined the effects of the release of PagL from latency on resistance to polymyxin B. The pmrA pagL and pmrE pagL double mutants were more susceptible to polymyxin B than were the parental pmrA and pmrE mutants, respectively. Furthermore, introduction of the PagL expression plasmid into the pmrA pagL double mutant increased the resistance to polymyxin B. In addition, PagL-dependent deacylation of lipid A was observed in a mutant in which lipid A could not be modified with phosphoethanolamine, which partly contributes to the PmrA-dependent resistance to polymyxin B. These results, taken together, suggest that the release of PagL from latency compensates for the loss of resistance to polymyxin B that is due to a lack of other modifications to LPS.

QUINACRINE FLUORESCENCE AND Q-BANDING PATTERNS OF HUMAN CHROMOSOMES.: I. Effects of Varying Factors

1975 ◽  
Vol 17 (1) ◽  
pp. 81-92 ◽  
Author(s):  
C. C. Lin ◽  
H. van de Sande ◽  
W. K. Smink ◽  
D. R. Newton
Keyword(s):  
Exposure Time ◽  
Uv Light ◽  
Human Chromosomes ◽  
Concave Mirror ◽  
Banding Patterns ◽  
Fluorescent Microscope ◽  
Sperm Dna ◽  
Quinacrine Fluorescence ◽  
Well Differentiated ◽  
Salmon Sperm Dna

Various factors involved in the production of "Q-bands" have been studied. It was found that a Zeiss standard WL fluorescent microscope required a shorter exposure time for photography as compared to a Zeiss photomicroscope. The minimal exposure time was obtained when the standard WL microscope was equipped with a UV light source containing a DC powered mercury burner and a concave mirror. Further, the pH and type of water used in the staining, washing and mounting of the slide were also important factors in producing clear and well differentiated "Q-bands". It also appears that the factors involved in the production of "Q-bands" effect the enhancement or quenching of fluorescence by poly d(A-T).poly d(A-T) and salmon sperm DNA or poly dG∙poly dC respectively. This preliminary report also suggests that DNA or polynucleotides with a specific base sequence may play an important role in Q-banding patterns on chromosomes.

scholarly journals Survival characteristics of Salmonella enterica serovar Enteritidis in chicken egg albumen

2006 ◽  
Vol 134 (5) ◽  
pp. 967-976 ◽  
Author(s):  
H. KANG ◽  
C. LOUI ◽  
R. I. CLAVIJO ◽  
L. W. RILEY ◽  
S. LU
Keyword(s):  
Salmonella Enterica ◽  
Iron Acquisition ◽  
Foodborne Pathogen ◽  
Human Infection ◽  
Salmonella Enterica Serovar Enteritidis ◽  
Factors Affecting ◽  
Chicken Egg ◽  
Egg Albumen ◽  
High Concentrations ◽  
Incubation Temperatures

Salmonella enterica serovar Enteritidis (SE) is a major foodborne pathogen primarily causing human infection through contaminated chicken eggs. To understand how SE survives in chicken egg albumen, we systematically and quantitatively analysed the survival properties of SE in egg albumen and identified factors affecting its survival. Survival assays of SE in egg indicate that egg albumen restricted the growth of SE. A major factor that controlled SE's growth in egg albumen was iron restriction, since egg albumen supplemented with iron allowed SE to grow, and iron acquisition mutants of SE showed decreased survival in egg albumen. In addition, low pH of albumen, high concentrations of bacteria and low incubation temperatures of bacteria with albumen facilitates the survival of SE. Our results suggest that egg albumen uses multiple mechanisms to control SE including iron limitation, surface interaction and possible enzymatic activities.

Pseudomonas aeruginosa Oligoribonuclease Controls Susceptibility to Polymyxin B by Regulating Pel Exopolysaccharide Production

2022 ◽  
Author(s):  
Baopeng Yang ◽  
Yujun Jiang ◽  
Yongxin Jin ◽  
Fang Bai ◽  
Zhihui Cheng ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Resistance ◽  
Defense Mechanism ◽  
Extracellular Polysaccharide ◽  
Opportunistic Pathogen ◽  
Polymyxin B ◽  
Guanosine Monophosphate ◽  
Extracellular Dna ◽  
Bacterial Cells ◽  
Bacterial Survival

Polymyxins are considered as the last resort antibiotics to treat infections caused by multidrug-resistant Gram negative pathogens. Pseudomonas aeruginosa is an opportunistic pathogen that causes various infections in humans. Proteins involved in lipopolysaccharide modification and maintaining inner and outer membrane integrities have been found to contribute to the bacterial resistance to polymyxins. Oligoribonuclease (Orn) is an exonuclease that regulates the homeostasis of intracellular (3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), thereby regulating the production of extracellular polysaccharide in P. aeruginosa . Previously, we demonstrated that Orn affects the bacterial resistance to fluoroquinolone, β-lactam and aminoglycoside antibiotics. In this study, we found that mutation of orn increased the bacterial survival following polymyxin B treatment in a wild type P. aeruginosa strain PA14. Overexpression of c-di-GMP degradation enzymes in the orn mutant reduced the bacterial survival. By using a fluorescence labeled polymyxin B, we found that mutation of orn increased the bacterial surface bound polymyxin B. Deletion of the Pel synthesis genes or treatment with a Pel hydrolase reduced the surface bound polymyxin B and bacterial survival. We further demonstrated that Pel binds to extracellular DNA (eDNA), which traps polymyxin B and thus protects the bacterial cells. Collectively, our results revealed a novel defense mechanism against polymyxin in P. aeruginosa .

scholarly journals Role of Extracellular DNA during Biofilm Formation by Listeria monocytogenes

2010 ◽  
Vol 76 (7) ◽  
pp. 2271-2279 ◽  
Author(s):  
Morten Harmsen ◽  
Martin Lappann ◽  
Susanne Kn�chel ◽  
S�ren Molin
Keyword(s):  
Listeria Monocytogenes ◽  
Biofilm Formation ◽  
Extracellular Dna ◽  
Central Component ◽  
High Molecular Weight Dna ◽  
Food Borne ◽  
Attachment Sites ◽  
Sperm Dna ◽  
Salmon Sperm Dna

ABSTRACT Listeria monocytogenes is a food-borne pathogen that is capable of living in harsh environments. It is believed to do this by forming biofilms, which are surface-associated multicellular structures encased in a self-produced matrix. In this paper we show that in L. monocytogenes extracellular DNA (eDNA) may be the only central component of the biofilm matrix and that it is necessary for both initial attachment and early biofilm formation for 41 L. monocytogenes strains that were tested. DNase I treatment resulted in dispersal of biofilms, not only in microtiter tray assays but also in flow cell biofilm assays. However, it was also demonstrated that in a culture without eDNA, neither Listeria genomic DNA nor salmon sperm DNA by itself could restore the capacity to adhere. A search for additional necessary components revealed that peptidoglycan (PG), specifically N-acetylglucosamine (NAG), interacted with the DNA in a manner which restored adhesion. If a short DNA fragment (less than approximately 500 bp long) was added to an eDNA-free culture prior to addition of genomic or salmon sperm DNA, adhesion was prevented, indicating that high-molecular-weight DNA is required for adhesion and that the number of attachment sites on the cell surface can be saturated.

Oligodeoxyribonucleotides derived from salmon sperm DNA: An alternative to defibrotide

Biologicals ◽  
2013 ◽  
Vol 41 (3) ◽  
pp. 190-196
Author(s):  
Chang-Ye Hui ◽  
Yan Guo ◽  
Xi Zhang ◽  
Jian-Hua Shao ◽  
Xue-Qin Yang ◽  
...  
Keyword(s):  
Sperm Dna ◽  
Salmon Sperm Dna
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