scholarly journals Role of the Alkyl Hydroperoxide Reductase (ahpCF) Gene in Oxidative Stress Defense of the Obligate Anaerobe Bacteroides fragilis

1999 ◽  
Vol 181 (18) ◽  
pp. 5701-5710 ◽  
Author(s):  
Edson R. Rocha ◽  
C. Jeffrey Smith
Keyword(s):  
Fusidic Acid ◽  
Resistant Strain ◽  
Parent Strain ◽  
Bacteroides Fragilis ◽  
Northern Blot Hybridization ◽  
Obligate Anaerobe ◽  
Alkyl Hydroperoxide Reductase ◽  
Oxygen Exposure ◽  
Alkyl Hydroperoxide

ABSTRACT In this study we report the identification and role of the alkyl hydroperoxide reductase (ahp) gene in Bacteroides fragilis. The two components of ahp,ahpC, and ahpF, are organized in an operon, and the deduced amino acid sequences revealed that B. fragilisAhpCF shares approximately 60% identity to orthologues in other gram-positive and gram-negative bacteria. Northern blot hybridization analysis of total RNA showed that the ahpCF genes were transcribed as a polycistronic 2.4-kb mRNA and that ahpCalso was present as a 0.6-kb monocistronic mRNA. ahpC andahpCF mRNAs were induced approximately 60-fold following H2O2 treatment or oxygen exposure of the parent strain but were constitutive in a peroxide-resistant strain. Further investigation using an ahpCF′::β-xylosidase gene transcriptional fusion confirmed that ahpCF had lost normal regulation in the peroxide-resistant strain compared to the parent. The ahpCF mutant was more sensitive to growth inhibition and mutagenesis by organic peroxides than the parent strain, as determined by disk inhibition assays and the frequency of mutation to fusidic acid resistance. This finding suggests that theahp genes play an important role in peroxide resistance inB. fragilis. Under anaerobic conditions, we observed increases in the number of spontaneous fusidic acid-resistant mutants of five- and sevenfold in ahpCF and ahpF strain backgrounds, respectively, and eightfold in the ahpCF katBdouble mutant strain compared to the parent and katBstrains. In addition, ahpCF, ahpF, andahpCF katB mutants were slightly more sensitive to oxygen exposure than the parent strain. Moreover, the isolation of a strain with enhanced aerotolerance and high-level resistance to alkyl hydroperoxides from an ahpCF katB parent suggests that the physiological responses to peroxide toxicity and to the toxic effects of molecular oxygen are overlapping and complex in this obligate anaerobe.

scholarly journals Roles of Alkyl Hydroperoxide Reductase Subunit C (AhpC) in Viable but Nonculturable Vibrio parahaemolyticus

2013 ◽  
Vol 79 (12) ◽  
pp. 3734-3743 ◽  
Author(s):  
Hen-Wei Wang ◽  
Chun-Hui Chung ◽  
Tsung-Yong Ma ◽  
Hin-chung Wong
Keyword(s):  
Vibrio Parahaemolyticus ◽  
Parent Strain ◽  
Vbnc State ◽  
Alkyl Hydroperoxide Reductase ◽  
Subunit C ◽  
Viable But Nonculturable ◽  
Content Type ◽  
Alkyl Hydroperoxide ◽  
The Times

ABSTRACTAlkyl hydroperoxide reductase subunit C (AhpC) is the catalytic subunit responsible for the detoxification of reactive oxygen species that form in bacterial cells or are derived from the host; thus, AhpC facilitates the survival of pathogenic bacteria under environmental stresses or during infection. This study investigates the role of AhpC in the induction and maintenance of a viable but nonculturable (VBNC) state inVibrio parahaemolyticus. In this investigation,ahpC1(VPA1683) andahpC2(VP0580) were identified in chromosomes II and I of this pathogen, respectively. Mutants with deletions of these twoahpCgenes and their complementary strains were constructed from the parent strain KX-V231. The growth of these strains was monitored on tryptic soy agar–3% NaCl in the presence of the extrinsic peroxides H2O2andtert-butyl hydroperoxide (t-BOOH) at different incubation temperatures. The results revealed that bothahpCgenes were protective againstt-BOOH, whileahpC1was protective against H2O2. The protective function ofahpC2at 4°C was higher than that ofahpC1. The times required to induce the VBNC state (4.7 weeks) at 4°C in a modified Morita mineral salt solution with 0.5% NaCl and then to maintain the VBNC state (4.7 weeks) in anahpC2mutant and anahpC1 ahpC2double mutant were significantly shorter than those for the parent strain (for induction, 6.2 weeks; for maintenance, 7.8 weeks) and theahpC1mutant (for induction, 6.0 weeks; for maintenance, 8.0 weeks) (P< 0.03). Complementation with anahpC2gene reversed the effects of theahpC2mutation in shortening the times for induction and maintenance of the VBNC state. This investigation identified the different functions of the twoahpCgenes and confirmed the particular role ofahpC2in the VBNC state ofV. parahaemolyticus.

scholarly journals Thioredoxin Reductase Is Essential for Thiol/Disulfide Redox Control and Oxidative Stress Survival of the Anaerobe Bacteroides fragilis

2007 ◽  
Vol 189 (22) ◽  
pp. 8015-8023 ◽  
Author(s):  
Edson R. Rocha ◽  
Arthur O. Tzianabos ◽  
C. Jeffrey Smith
Keyword(s):  
Parent Strain ◽  
Culture Media ◽  
Blot Hybridization ◽  
Thioredoxin Reductase ◽  
Bacteroides Fragilis ◽  
Redox System ◽  
Opportunistic Pathogen ◽  
Abscess Formation ◽  
Infection Model ◽  
Northern Blot Hybridization

ABSTRACT Results of this study showed that the anaerobic, opportunistic pathogen Bacteroides fragilis lacks the glutathione/glutaredoxin redox system and possesses an extensive number of putative thioredoxin (Trx) orthologs. Analysis of the genome sequence revealed six Trx orthologs and an absence of genes required for synthesis of glutathione and glutaredoxins. In addition, it was shown that the thioredoxin reductase (TrxB)/Trx system is the major or sole redox system for thiol/disulfide cellular homeostasis in this anaerobic bacterium. Expression of the B. fragilis trxB gene was induced following treatment with diamide or H2O2 or exposure to oxygen. This inducible trxB expression was OxyR independent. Northern blot hybridization analysis showed that the trxB mRNA was cotranscribed with lolA as a bicistronic transcript or was present as a monocistronic transcript that was also highly induced under the same conditions. The role of LolA, a prokaryotic periplasmic lipoprotein-specific molecular chaperone in the thiol/disulfide redox system, is unknown. A trxB deletion mutant was more sensitive to the effects of diamide and oxygen than the parent strain. In addition, the trxB mutant was unable to grow in culture media without addition of a reductant. Furthermore, the trxB mutant was not able to induce intraabdominal abscess formation in a mouse model, whereas the parent strain was. Taken together, these data strongly suggest that TrxB/Trx is the major, if not the sole, thiol/disulfide redox system in this anaerobe required for survival and abscess formation in a peritoneal cavity infection model.

scholarly journals Multi-step resistance to Chloramphenicol in RC-stringent Escherichia coli K12—its effect on the induction of RNA synthesis by antibiotics under amino acid starvation

1965 ◽  
Vol 6 (2) ◽  
pp. 304-309 ◽  
Author(s):  
E. C. R. Reeve ◽  
J. O. Bishop
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Resistant Strain ◽  
Parent Strain ◽  
Rna Synthesis ◽  
Amino Acid Starvation ◽  
Sensitive Strain ◽  
Escherichia Coli K12 ◽  
High Doses

A multi-step Chloramphenicol (CM)-resistant derivative of an RC-stringent strain of Escherichia coli auxotrophic for threonine and leucine was resistant also to Aureomycin (AM) and Puromycin (PM). All three antibiotics released the repression of RNA synthesis due to amino acid starvation in the CM-sensitive parent strain, their relative activities being about 1:10:100 for AM: CM: PM. High doses of AM and CM failed to induce RNA synthesis. The CM-resistant strain required greater concentrations of each antibiotic than the sensitive strain to induce the same level of RNA synthesis, and appeared to be about one hundred times, ten times and five times more resistant to CM, AM and PM, respectively, than the sensitive strain.

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