Impact of sarA Mutation on Immune System Evasion and Stress Response in Staphylococcus aureus

2020 ◽  
Vol 29 (3) ◽  
pp. 105-112
Author(s):  
Yomna A. Hagag ◽  
Abdelaziz Elgaml ◽  
Ramadan Hassan ◽  
Hany I. Kenawy
Keyword(s):  
Staphylococcus Aureus ◽  
Immune System ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Human Neutrophils ◽  
Bacterial Survival ◽  
Wild Type ◽  
The Impact

Background: Staphylococcus aureus is a major human pathogen responsible for a large number of infections. In S. aureus, SarA is an important global locus responsible for the regulation of virulence factors, as well as biofilm formation. Objectives: The aim of this work is to clarify the impact of SarA on biofilm formation, immune system evasion, as well as the survival of S. aureus under stress conditions. Methodology: A comparative study between S. aureus wild type strain, sarA mutant and complemented strains was established addressing the biofilm formation, opsonization, phagocytosis, as well as ability of the bacterium to survive in stressful environments including acidic pH, hyperosmotic and oxidative stress. The in vitro experiments were confirmed by challenging of mice via intraperitoneal injection with the wild type strain, sarA mutant and complemented strains. Results: Mutation of sarA diminished significantly biofilm formation. Moreover, this mutation resulted in a slight decrease in the deposition of the most important opsonin in complement-mediated immunity, named C3 on S. aureus cells. However, this mutation was associated with a significant enhancement of bacterial phagocytosis and killing by human neutrophils. Furthermore, this mutation altered bacterial survival in stressful conditions. It is also noteworthy that sarA mutation resulted in a significant higher survival rates during the challenging of mice. Conclusion: SarA plays a role as a key regulator of biofilm formation, which in turn has a great impact on immune system evasion through affecting opsonization and phagocytosis. In addition, SarA improves the ability of S. aureus to survive in stressful conditions.

scholarly journals Hyaluronan Modulation Impacts Staphylococcus aureus Biofilm Infection

2016 ◽  
Vol 84 (6) ◽  
pp. 1917-1929 ◽  
Author(s):  
Carolyn B. Ibberson ◽  
Corey P. Parlet ◽  
Jakub Kwiecinski ◽  
Heidi A. Crosby ◽  
David K. Meyerholz ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Infection Model ◽  
Wild Type ◽  
Content Type ◽  
Joint Infections ◽  
The Impact ◽  
Biofilm Infection ◽  
Biofilm Matrix

Staphylococcus aureusis a leading cause of chronic biofilm infections. Hyaluronic acid (HA) is a large glycosaminoglycan abundant in mammalian tissues that has been shown to enhance biofilm formation in multiple Gram-positive pathogens. We observed that HA accumulated in anS. aureusbiofilm infection using a murine implant-associated infection model and that HA levels increased in a mutant strain lacking hyaluronidase (HysA).S. aureussecretes HysA in order to cleave HA during infection. Throughin vitrobiofilm studies with HA, thehysAmutant was found to accumulate increased biofilm biomass compared to the wild type, and confocal microscopy showed that HA is incorporated into the biofilm matrix. Exogenous addition of purified HysA enzyme dispersed HA-containing biofilms, while catalytically inactive enzyme had no impact. Additionally, induction ofhysAexpression prevented biofilm formation and also dispersed an established biofilm in the presence of HA. These observations were corroborated in the implant model, where there was decreased dissemination from anhysAmutant biofilm infection compared to theS. aureuswild type. Histopathology demonstrated that infection with anhysAmutant caused significantly reduced distribution of tissue inflammation compared to wild-type infection. To extend these studies, the impact of HA andS. aureusHysA on biofilm-like aggregates found in joint infections was examined. We found that HA contributes to the formation of synovial fluid aggregates, and HysA can disrupt aggregate formation. Taken together, these studies demonstrate that HA is a relevant component of theS. aureusbiofilm matrix and HysA is important for dissemination from a biofilm infection.

scholarly journals Identification of a Novel Two-Component System in Streptococcus gordonii V288 Involved in Biofilm Formation

2004 ◽  
Vol 72 (6) ◽  
pp. 3489-3494 ◽  
Author(s):  
Yongshu Zhang ◽  
Yu Lei ◽  
Ali Khammanivong ◽  
Mark C. Herzberg
Keyword(s):  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Response Regulator ◽  
Streptococcus Gordonii ◽  
Wild Type ◽  
Two Component System ◽  
Component System ◽  
Two Component

ABSTRACT Streptococcus gordonii is a pioneer colonizer of the teeth, contributing to the initiation of the oral biofilm called dental plaque. To identify genes that may be important in biofilm formation, a plasmid integration library of S. gordonii V288 was used. After screening for in vitro biofilm formation on polystyrene, a putative biofilm-defective mutant was isolated. In this mutant, pAK36 was inserted into a locus encoding a novel two-component system (bfr [biofilm formation related]) with two cotranscribed genes that form an operon. bfrA encodes a putative response regulator, while bfrB encodes a receptor histidine kinase. The bfr mutant and wild-type strain V288 showed similar growth rates in Todd-Hewitt broth (THB). A bfr-cat fusion strain was constructed. During growth in THB, the reporter activity (chloramphenicol acetyltransferase) was first detected in mid-log phase and reached a maximum in stationary phase, suggesting that transcription of bfr was growth stage dependent. After being harvested from THB, the bfr mutant adhered less effectively than did wild-type strain V288 to saliva-coated hydroxyapatite (sHA). To simulate pioneer colonization of teeth, S. gordonii V288 was incubated with sHA for 4 h in THB with 10% saliva to develop biofilms. RNA was isolated, and expression of bfrAB was estimated. In comparison to that of cells grown in suspension (free-growing cells), bfr mRNA expression by sessile cells on sHA was 1.8-fold greater and that by surrounding planktonic cells was 3.5-fold greater. Therefore, bfrAB is a novel two-component system regulated in association with S. gordonii biofilm formation in vitro.

scholarly journals Cryptococcus neoformanssecretes small molecules that inhibit IL-1β inflammasome-dependent secretion

2019 ◽  
Author(s):  
Pedro Henrique Bürgel ◽  
Clara Luna Marina ◽  
Pedro H. V. Saavedra ◽  
Patrícia Albuquerque ◽  
Paulo Henrique Holanda ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Cryptococcus Neoformans ◽  
Type Strain ◽  
Wild Type Strain ◽  
Conditioned Media ◽  
Inflammasome Activation ◽  
Wild Type ◽  
Fungal Virulence ◽  
The Impact

AbstractCryptococcus neoformansis an encapsulated yeast that causes disease mainly in immunosuppressed hosts. It is considered a facultative intracellular pathogen because of its capacity to survive and replicate inside phagocytes, especially macrophages. This capacity is heavily dependent on various virulence factors, particularly the glucuronoxylomannan (GXM) component of the polysaccharide capsule, that render the non- or poorly-activated macrophage ineffective against phagocytosed yeast. Strategies utilized by macrophages to prevent this scenario include pyroptosis (a rapid highly inflammatory cell death) and vomocytosis (the expulsion of the pathogen from the intracellular environment without lysis). Inflammasome activation in phagocytes is usually protective against fungal infections, including cryptococcosis. Nevertheless, recognition ofC. neoformansby inflammasome receptors requires specific changes in morphology or the opsonization of the yeast, impairing a proper inflammasome function. In this context, we analyzed the impact of molecules secreted byC. neoformansB3501 strain and its acapsular mutantΔcap67in an inflammasome activationin vitromodel. Our results showed that conditioned media derived from B3501 was capable of inhibiting inflammasome dependent events (i. e. IL-1β secretion and LDH release via pyroptosis) more strongly than conditioned media fromΔcap67, regardless of GXM presence. We also demonstrated that macrophages treated with conditioned media were less responsive against infection with the virulent strain H99, exhibiting lower rates of phagocytosis, increased fungal burdens and enhanced vomocytosis. Moreover, we showed that the aromatic metabolite DL-Indole-3-lactic acid (ILA) was present in B3501’s conditioned media and that this fungal metabolite is involved in the regulation of inflammasome activation byC. neoformans. Overall, the results presented show that conditioned media from a wild-type strain can inhibit an important recognition pathway and subsequent fungicidal functions of macrophages, contributing to fungal survivalin vitroand suggesting that this serves as an important role for secreted molecules during cryptococcal infections.Author’s SummaryCryptococcus neoformansis the agent of cryptococcal meningitis, a disease that can be life-threatening in immunocompromised hosts such as those infected with HIV. The infection thrives in hosts that poorly activate their immune system, mainly because of the yeast’s ability to survive inside macrophages and migrate towards the central nervous system. Emerging data indicate that cryptococci modulate the host immune response, but the underlying mechanisms remain largely uncharacterized. Here we show that secreted molecules from a wild-type strain ofC. neoformansimpair inflammatory responses driven by inflammasome activation, which in turn impact the macrophage antifungal activity. We further show that this inhibition does not involve GXM, the main constituent of the fungal capsule, but rather is partially dependent on DL-Indole-3-lactic acid (ILA), a metabolite not previously implicated in fungal virulence.

scholarly journals Bactericidal Activities of BMS-284756, a Novel Des-F(6)-Quinolone, against Staphylococcus aureus Strains with Topoisomerase Mutations

2002 ◽  
Vol 46 (1) ◽  
pp. 191-195 ◽  
Author(s):  
Laura E. Lawrence ◽  
MaryBeth Frosco ◽  
Brenda Ryan ◽  
Susan Chaniewski ◽  
Hyekyung Yang ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
In Vivo Efficacy ◽  
Mutant Strains ◽  
Bactericidal Activities ◽  
Time Kill

ABSTRACT The antistaphylococcal activities of BMS-284756 (T-3811ME), levofloxacin, moxifloxacin, and ciprofloxacin were compared against wild-type and grlA and grlA/gyrA mutant strains of Staphylococcus aureus. BMS-284756 was the most active quinolone tested, with MICs and minimal bactericidal concentrations against S. aureus wild-type strain MT5, grlA mutant MT5224c4, and grlA/gyrA mutant EN8 of 0.03 and 0.06, 0.125 and 0.125, and 4 and 4 μg/ml, respectively. In the time-kill studies, BMS-284756 and levofloxacin exhibited rapid killing against all strains. Ciprofloxacin, however, was not bactericidal for the double mutant, EN8. BMS-284756 and levofloxacin were bactericidal (3 log10 decrease in CFU/ml) against the MT5 and MT5224c4 strains at two and four times the MIC within 2 to 4 h. Against EN8, BMS-284756 was bactericidal within 4 h at two and four times the MIC, and levofloxacin achieved similar results within 4 to 6 h. Both the wild-type strain MT5 and grlA mutant MT5224c4 should be considered susceptible to both BMS-284756 and levofloxacin, and both quinolones are predicted to have clinical efficacy. The in vivo efficacy of BMS-284756, levofloxacin, and moxifloxacin against S. aureus strain ISP794 and its single mutant 2C6(1)-1 directly reflected the in vitro activity: increased MICs correlated with decreased in vivo efficacy. The 50% protective doses of BMS-284756 against wild-type and mutant strains were 2.2 and 1.6 mg/kg of body weight/day, respectively, compared to the levofloxacin values of 16 and 71 mg/kg/day and moxifloxacin values of 4.7 and 61.6 mg/kg/day. BMS-284756 was more potent than levofloxacin and equipotent with moxifloxacin against ISP794 both in vitro and in vivo, while BMS-284756 was more potent than levofloxacin and moxifloxacin against 2C6(1)-1.

scholarly journals Impact of Inorganic Carbon Availability on Microcystin Production by Microcystis aeruginosa PCC 7806

2007 ◽  
Vol 73 (21) ◽  
pp. 6994-7002 ◽  
Author(s):  
Sabine J�hnichen ◽  
Tilo Ihle ◽  
Thomas Petzoldt ◽  
J�rgen Benndorf
Keyword(s):  
Microcystis Aeruginosa ◽  
Type Strain ◽  
Inorganic Carbon ◽  
Wild Type Strain ◽  
Light Cycle ◽  
Variable Fluorescence ◽  
Wild Type ◽  
Dark Light ◽  
Sodium Dependent ◽  
The Impact

ABSTRACT Batch culture experiments with the cyanobacterium Microcystis aeruginosa PCC 7806 were performed in order to test the hypothesis that microcystins (MCYSTs) are produced in response to a relative deficiency of intracellular inorganic carbon (Ci,i). In the first experiment, MCYST production was studied under increased Ci,i deficiency conditions, achieved by restricting sodium-dependent bicarbonate uptake through replacement of sodium bicarbonate in the medium with its potassium analog. The same experimental approach was used in a second experiment to compare the response of the wild-type strain M. aeruginosa PCC 7806 with its mcyB mutant, which lacks the ability to produce MCYSTs. In a third experiment, the impact of varying the Ci,i status on MCYST production was examined without suppressing the sodium-dependent bicarbonate transporter; instead, a detailed investigation of a dark-light cycle was performed. In all experiments, a relative Ci,i deficiency was indicated by an elevated variable fluorescence signal and led to enhanced phycocyanin cell quotas. Higher MCYST cell quotas (in the first and third experiments) and increased total (intracellular plus extracellular) MCYST production (in the first experiment) were detected with increased Ci,i deficiency. Furthermore, the MCYST-producing wild-type strain and its mcyB mutant showed basically the same response to restrained inorganic carbon uptake, with elevated variable fluorescence and phycocyanin cell quotas with increased Ci,i deficiency. The response of the wild type, however, was distinctly stronger and also included elevated chlorophyll a cell quotas. These differences indicate the limited ability of the mutant to adapt to low-Ci,i conditions. We concluded that MCYSTs may be involved in enhancing the efficiency of the adaptation of the photosynthetic apparatus to fluctuating inorganic carbon conditions in cyanobacterial cells.

Identification of pilin pools in the membranes of Pseudomonas aeruginosa

1982 ◽  
Vol 152 (2) ◽  
pp. 687-691
Author(s):  
T H Watts ◽  
E A Worobec ◽  
W Paranchych
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Gel Electrophoresis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Wild Type ◽  
Outer Membranes

The proteins of purified inner and outer membranes obtained from Pseudomonas aeruginosa strains PAK and PAK/2Pfs were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and treated with antiserum raised against pure pili. Bound antipilus antibodies were visualized by reaction with 125I-labeled protein A from Staphylococcus aureus. The results showed that there are pools of pilin in both the inner and outer membranes of P. aeruginosa and that the pool size in the multipiliated strain is comparable with that of the wild-type strain.

scholarly journals Construction and Characterization of aMycobacterium tuberculosis Mutant Lacking the Alternate Sigma Factor Gene, sigF

2000 ◽  
Vol 68 (10) ◽  
pp. 5575-5580 ◽  
Author(s):  
Ping Chen ◽  
Rafael E. Ruiz ◽  
Qing Li ◽  
Richard F. Silver ◽  
William R. Bishai
Keyword(s):  
Stationary Phase ◽  
Sigma Factor ◽  
Type Strain ◽  
Wild Type Strain ◽  
Axenic Culture ◽  
Wild Type ◽  
Intracellular Growth ◽  
Factor Gene ◽  
Time To Death

ABSTRACT The alternate RNA polymerase sigma factor gene, sigF, which is expressed in stationary phase and under stress conditions in vitro, has been deleted in the virulent CDC1551 strain ofMycobacterium tuberculosis. The growth rate of the ΔsigF mutant was identical to that of the isogenic wild-type strain in exponential phase, although in stationary phase the mutant achieved a higher density than the wild type. The mutant showed increased susceptibility to rifampin and rifapentine. Additionally, the ΔsigF mutant displayed diminished uptake of chenodeoxycholate, and this effect was reversed by complementation with a wild-type sigF gene. No differences in short-term intracellular growth between mutant and wild-type organisms within human monocytes were observed. Similarly, the organisms did not differ in their susceptibilities to lymphocyte-mediated inhibition of intracellular growth. However, mice infected with the ΔsigF mutant showed a median time to death of 246 days compared with 161 days for wild-type strain-infected animals (P < 0.001). These data indicate that M. tuberculosis sigF is a nonessential alternate sigma factor both in axenic culture and for survival in macrophages in vitro. While the ΔsigF mutant produces a lethal infection of mice, it is less virulent than its wild-type counterpart by time-to-death analysis.

scholarly journals Mutations Conferring Aminoglycoside and Spectinomycin Resistance in Borrelia burgdorferi

2006 ◽  
Vol 50 (2) ◽  
pp. 445-452 ◽  
Author(s):  
Daniel Criswell ◽  
Virginia L. Tobiason ◽  
J. Stephen Lodmell ◽  
D. Scott Samuels
Keyword(s):  
16S Rrna ◽  
Borrelia Burgdorferi ◽  
Type Strain ◽  
Wild Type Strain ◽  
Small Subunit ◽  
Wild Type ◽  
Spectinomycin Resistance ◽  
E Coli ◽  
Resistant Mutants

ABSTRACT We have isolated and characterized in vitro mutants of the Lyme disease agent Borrelia burgdorferi that are resistant to spectinomycin, kanamycin, gentamicin, or streptomycin, antibiotics that target the small subunit of the ribosome. 16S rRNA mutations A1185G and C1186U, homologous to Escherichia coli nucleotides A1191 and C1192, conferred >2,200-fold and 1,300-fold resistance to spectinomycin, respectively. A 16S rRNA A1402G mutation, homologous to E. coli A1408, conferred >90-fold resistance to kanamycin and >240-fold resistance to gentamicin. Two mutations were identified in the gene for ribosomal protein S12, at a site homologous to E. coli residue Lys-87, in mutants selected in streptomycin. Substitutions at codon 88, K88R and K88E, conferred 7-fold resistance and 10-fold resistance, respectively, to streptomycin on B. burgdorferi. The 16S rRNA A1185G and C1186U mutations, associated with spectinomycin resistance, appeared in a population of B. burgdorferi parental strain B31 at a high frequency of 6 × 10−6. These spectinomycin-resistant mutants successfully competed with the wild-type strain during 100 generations of coculture in vitro. The aminoglycoside-resistant mutants appeared at a frequency of 3 × 10−9 to 1 ×10−7 in a population and were unable to compete with wild-type strain B31 after 100 generations. This is the first description of mutations in the B. burgdorferi ribosome that confer resistance to antibiotics. These results have implications for the evolution of antibiotic resistance, because the 16S rRNA mutations conferring spectinomycin resistance have no significant fitness cost in vitro, and for the development of new selectable markers.

scholarly journals SdiA Aids Enterohemorrhagic Escherichia coli Carriage by Cattle Fed a Forage or Grain Diet

2013 ◽  
Vol 81 (9) ◽  
pp. 3472-3478 ◽  
Author(s):  
Haiqing Sheng ◽  
Y. N. Nguyen ◽  
Carolyn J. Hovde ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Deletion Mutant ◽  
Wild Type Strain ◽  
Rumen Fluid ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Dependent Manner ◽  
Bacterial Survival ◽  
Wild Type

ABSTRACTEnterohemorrhagicEscherichia coli(EHEC) causes hemorrhagic colitis and life-threatening complications. The main reservoirs for EHEC are healthy ruminants. We reported that SdiA senses acyl homoserine lactones (AHLs) in the bovine rumen to activate expression of the glutamate acid resistance (gad) genes priming EHEC's acid resistance before they pass into the acidic abomasum. Conversely, SdiA represses expression of the locus of enterocyte effacement (LEE) genes, whose expression is not required for bacterial survival in the rumen but is necessary for efficient colonization at the rectoanal junction (RAJ) mucosa. Our previous studies show that SdiA-dependent regulation was necessary for efficient EHEC colonization of cattle fed a grain diet. Here, we compared the SdiA role in EHEC colonization of cattle fed a forage hay diet. We detected AHLs in the rumen of cattle fed a hay diet, and these AHLs activatedgadgene expression in an SdiA-dependent manner. The rumen fluid and fecal samples from hay-fed cattle were near neutrality, while the same digesta samples from grain-fed animals were acidic. Cattle fed either grain or hay and challenged with EHEC orally carried the bacteria similarly. EHEC was cleared from the rumen within days and from the RAJ mucosa after approximately one month. In competition trials, where animals were challenged with both wild-type and SdiA deletion mutant bacteria, diet did not affect the outcome that the wild-type strain was better able to persist and colonize. However, the wild-type strain had a greater advantage over the SdiA deletion mutant at the RAJ mucosa among cattle fed the grain diet.

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