Gonococcal resistance to antimicrobial hydrophobic agents (HAs) is due to energy-dependent removal ofHAs from the bacterial cell by the MtrCDE membrane-associated efflux pump. The mtrR (multiple transferrable resistance Regulator) gene encodes a putative transcriptional repressor protein (MtrR) believed to be responsible for regulation of mtrCDE gene expression. Gel mobility shift and DNase I footprint assays that used a maltose-binding protein (MBP)-MtrR fusion protein demonstrated that the MtrR repressor is capable of specifically binding the DNA sequence between the mtrR and mtrC genes. This binding site was localized to a 26-nucleotide stretch that includes the promoter utilized for mtrCDE transcription and, on the complementary strand, a 22-nucleotide stretch that contains the ؊35 region of the mtrR promoter. A single transition mutation (A3G) within the MtrR-binding site decreased the affinity of the target DNA for MtrR and enhanced gonococcal resistance to HAs when introduced into HA-susceptible strain FA19 by transformation. Since this mutation enhanced expression of the mtrCDE gene complex but decreased expression of the mtrR gene, the data are consistent with the notion that MtrR acts as a transcriptional repressor of the mtrCDE efflux pump protein genes.Resistance of Neisseria gonorrhoeae to structurally diverse hydrophobic agents (HAs) has been the subject of several recent investigations (5,6,10,20,23). Although earlier studies indicated that the mtr (multiple transferrable resistance) locus modifies the permeability barrier of the gonococcal cell envelope (3,4,13,16,22,24), more recent studies revealed that mtr encodes an energy-dependent efflux system (5,6,10,20,23). The membrane proteins (MtrC, MtrD, and MtrE) forming the efflux pump share considerable amino acid sequence similarity with other efflux proteins in Escherichia coli (12,17) and Pseudomonas aeruginosa (21). Active removal of toxic compounds by bacteria is an important mechanism of multiple antibiotic resistance (8, 17) and may confer a selective advantage on organisms, such as gonococci, when they colonize mucosal sites bathed in fluids containing antibacterial fatty acids and bile salts or encounter other environmental stresses (8,11,12,(16)(17)(18)23).Production of the MtrCDE efflux proteins is controlled at the level of transcription by both cis-and trans-acting factors involving the mtrR gene (5,6,20,23). The mtrR gene is located 250 bp upstream of the mtrCDE gene complex and transcribed divergently (5,6,20). It encodes a 210-amino-acid protein with a molecular mass of approximately 23 kDa that contains a putative helix-turn-helix motif near its N terminus (5,20,23). MtrR has considerable amino acid sequence similarities to several transcriptional repressors, most notably, the tetracycline repressor of pSC101 (1,7,20). Recent investigations have revealed that missense or deletion mutations within the mtrR coding region result in enhanced mtrCDE transcription and gonococcal resistance to HAs (6, 23). Thus, the available genetic dat...

The mfr (multiple transferable resistance) system of Neisseria gonorrhoeae mediates resistance of gonococci to structurally diverse hydrophobic agents (HAS) through an energy-dependent efflux process. Recently, complete or partial ORFs that encode membrane proteins (MtrG MtrD, MtrE) forming an efflux pump responsible for removal of HAS from gonococci were identified and appeared to constitute a single transcriptional unit. In this study, the complete nucleotide sequence of the mtrD gene was determined, permitting the characterization of the MtrD protein. The full-length MtrD protein has a predicted molecular mass of nearly 114 kDa, putatively containing a 56 amino acid signal peptide. MtrD displays significant amino acid sequence similarity to a f am i I y of cytoplasmic membrane proteins, termed resistancehodu lat ion/ division (RND) proteins, which function as energy-dependent transporters of antibacterial agents and secrete bacterial products to the extracellular fluid. The predicted topology of the MtrD transporter protein revealed 12 potential membrane-spanning domains, which were clustered within the central and Cterminal regions of the primary sequence. Loss of MtrD due to insertional inactivation of the mtrD gene rendered gonococci hypersusceptible to several structurally diverse HAS, including two fatty acids (capric acid and palmitic acid) and a bile salt (cholic acid), but not hydrophilic antibiotics such as ciprof loxacin and streptomycin. Since gonococci often infect mucosal sites rich in toxic fatty acids and bile salts, the expression of the mtr efflux system may promote growth of gonococci under hostile conditions encountered in vivo.

No abstract

Background The number of reported cases of Legionnaires’ disease, a severe pneumonia caused by the bacterium Legionella, is increasing in the United States. During 2000–2014, the rate of reported legionellosis cases increased from 0.42 to 1.62 per 100 000 persons; 4% of reported cases were outbreak‐associated. Legionella is transmitted through aerosolization of contaminated water. A new industry standard for prevention of Legionella growth and transmission in water systems in buildings was published in 2015. CDC investigated outbreaks of Legionnaires’ disease to identify gaps in building water system maintenance and guide prevention efforts. Methods Information from summaries of CDC Legionnaires’ disease outbreak investigations during 2000–2014 was systematically abstracted, and water system maintenance deficiencies from land‐based investigations were categorized as process failures, human errors, equipment failures, or unmanaged external changes. Results During 2000–2014, CDC participated in 38 field investigations of Legionnaires’ disease. Among 27 land‐based outbreaks, the median number of cases was 10 (range = 3–82) and median outbreak case fatality rate was 7% (range = 0–80%). Sufficient information to evaluate maintenance deficiencies was available for 23 (85%) investigations. Of these, all had at least one deficiency; 11 (48%) had deficiencies in ≥2 categories. Fifteen cases (65%) were linked to process failures, 12 (52%) to human errors, eight (35%) to equipment failures, and eight (35%) to unmanaged external changes. Conclusions and Implications for Public Health Practice Multiple common preventable maintenance deficiencies were identified in association with disease outbreaks, highlighting the importance of comprehensive water management programs for water systems in buildings. Properly implemented programs, as described in the new industry standard, could reduce Legionella growth and transmission, preventing Legionnaires’ disease outbreaks and reducing disease.