Moraxella catarrhalis uses a twin-arginine translocation system to secrete the β-lactamase BRO-2
© Balder et al.; licensee BioMed Central Ltd. 2013
Received: 4 April 2013
Accepted: 10 June 2013
Published: 19 June 2013
Moraxella catarrhalis is a human-specific gram-negative bacterium readily isolated from the respiratory tract of healthy individuals. The organism also causes significant health problems, including 15-20% of otitis media cases in children and ~10% of respiratory infections in adults with chronic obstructive pulmonary disease. The lack of an efficacious vaccine, the rapid emergence of antibiotic resistance in clinical isolates, and high carriage rates reported in children are cause for concern. Virtually all Moraxella catarrhalis isolates are resistant to β-lactam antibiotics, which are generally the first antibiotics prescribed to treat otitis media in children. The enzymes responsible for this resistance, BRO-1 and BRO-2, are lipoproteins and the mechanism by which they are secreted to the periplasm of M. catarrhalis cells has not been described.
Comparative genomic analyses identified M. catarrhalis gene products resembling the TatA, TatB, and TatC proteins of the well-characterized Twin Arginine Translocation (TAT) secretory apparatus. Mutations in the M. catarrhalis tatA, tatB and tatC genes revealed that the proteins are necessary for optimal growth and resistance to β-lactams. Site-directed mutagenesis was used to replace highly-conserved twin arginine residues in the predicted signal sequence of M. catarrhalis strain O35E BRO-2, which abolished resistance to the β-lactam antibiotic carbanecillin.
Moraxella catarrhalis possesses a TAT secretory apparatus, which plays a key role in growth of the organism and is necessary for secretion of BRO-2 into the periplasm where the enzyme can protect the peptidoglycan cell wall from the antimicrobial activity of β-lactam antibiotics.
Moraxella catarrhalis is a Gram-negative bacterium primarily associated with otitis media in children and respiratory infections in adults with compromised lung function, particularly patients with Chronic Obstructive Pulmonary Disease (COPD). The organism is also readily isolated from the upper respiratory tract of healthy individuals and thus was considered a commensal bacterium until relatively recently. The rate of colonization by M. catarrhalis varies depending on many factors such as age, socioeconomic status, geography, and overall health condition. It has been reported that ~2/3 of children are colonized in their first year of life and 3-5% of adults carry the organism asymptomatically. Following initial colonization, there is a high rate of turnover, indicating continual clearance and re-colonization by new strains [1–27].
Moraxella catarrhalis possesses several virulence determinants that enable it to persist in the human respiratory tract. A number of molecules in the outer membrane have been shown to contribute to adherence, allowing M. catarrhalis to bind and colonize the host mucosa. These include LOS, UspA1, UspA2H, McaP, OMPCD, Hag/MID, MhaB1, MhaB2, MchA1, MchA2, and the type IV pilus [28–37]. In order to persist following colonization, M. catarrhalis possesses several mechanisms to evade the host immune system including resistance to complement. The best studied of these being UspA2 and UspA2H, which bind the C4-binding protein, C3 and vitronectin [38–41], as well as CopB, OMPCD, OmpE, and LOS [31, 37, 42, 43].
Moraxella catarrhalis is often refractory to antibiotic treatment. Over 90% of isolates have been shown to possess a beta-lactamase, making them resistant to penicillin-based antibiotics [44–51], which are typically prescribed first to treat otitis media. The genes specifying this resistance appear to be of Gram-positive origin [52, 53], suggesting that M. catarrhalis can readily acquire genes conferring resistance to additional antibiotics via horizontal transfer. Additionally, recent evidence has shown that M. catarrhalis persists as a biofilm in vivo, giving it further protection from antibiotic treatment and the host immune response [54–58].
The bacterial twin-arginine translocation (TAT) system mediates secretion of folded proteins across the cytoplasmic membrane. The TAT apparatus typically consists of three integral membrane proteins, namely TatA, TatB, and TatC. TatA forms the pore through which TAT substrates are secreted whereas TatB and TatC are important for binding and directing the substrates to the TatA pore. TatC acts as the gatekeeper for the secretion apparatus and specifically recognizes TAT substrates via a well-conserved signal sequence [59–62]. This signal sequence is similar to that of the general secretory system in that it possesses N- (neutral), H- (hydrophobic), and C- (charged) regions, but also contains a TAT-recognition motif (S/T-R-R-x-F-L-K) exhibiting nearly invariable twin arginines (RR) followed by two uncharged residues [59–62]. Proteins secreted via the TAT system are often, but not limited to, proteins that bind cofactors in the cytoplasm prior to transport, such as those involved in respiration and electron transport, and proteins that bind catalytic metal ions [59–62]. The TAT system has also been shown to secrete several factors important for bacterial pathogenesis including iron acquisition, flagella synthesis, toxins, phospholipases, and beta-lactamases [59, 62–74]. In this study, we identified genes encoding a TAT system in M. catarrhalis and mutated these genes in order to elucidate the role of this translocase in the secretion of proteins that may be important for pathogenesis.
Results and discussion
Identification of tatA, tatB and tatC genes in M. catarrhalis
Analysis of the patented genomic sequence of M. catarrhalis strain ATCC43617 using NCBI’s tblastn service (http://blast.ncbi.nlm.nih.gov/Blast.cgi) identified an ORF (nucleotides 267,266 to 266,526 of GenBank accession number AX06766.1) that encodes a protein similar to the tatC gene product of Pseudomonas stutzeri (expect value of 7e-56). TatC is the most highly-conserved component of the TAT system among organisms known (or predicted) to utilize this particular secretion apparatus [59–62]. TatC is located in the cytoplasmic membrane, typically contains 6 membrane-spanning regions, and plays a key role in recognizing the twin-arginine motif in the signal sequence of molecules secreted by the TAT system. The M. catarrhalis ATCC43617 tatC-like ORF specifies a 27-kDa protein of 247 amino acids, and analysis using the TMPred server (http://www.ch.embnet.org/software/TMPRED_form.html) revealed that it contains 6 potential membrane-spanning domains (data not shown).
To assess the presence and conservation of the tat genes in other M. catarrhalis isolates, we amplified and sequenced these genes from strains O35E, O12E, McGHS1, V1171, and TTA37. The encoded gene products were then compared using ClustalW (http://www.ebi.ac.uk/Tools/msa/clustalw2/). Of note, the annotated genomic sequence of the M. catarrhalis isolate BBH18 has been published  and the predicted aa sequence of the TatA (MCR_0127, GenBank accession number ADG60399.1), TatB (MCR_0126, GenBank accession number ADG60398.1) and TatC (MCR_0125, GenBank accession number ADG60397.1) proteins were included in our comparative analyses. Overall, the TatA and TatC proteins are perfectly conserved. The TatB proteins divide the strains into two groups where O35E, McGHS1, TTA37, ATCC43617, and BBH18 are 100% identical to each other, while O12E and V1171 both contain the same aa substitution at residue 38 (S in lieu of G). We also noted that in all isolates examined, the tatA and tatB ORFs overlap by one nucleotide. A similar one-nucleotide overlap is also observed for the tatB and tatC coding regions. This observation suggests that the M. catarrhalis tatA, tatB, and tatC genes are transcriptionally and translationally linked.
The M. catarrhalis tatA, tatB and tatCgenes are necessary for optimal growth
To study the functional properties of the Tat proteins in M. catarrhalis, we constructed a panel of isogenic mutant strains of isolate O35E in which the tatA, tatB and tatC genes were disrupted with a kanamycin resistance (kanR) marker. Each mutant was also complemented with a plasmid encoding a wild-type (WT) copy of the mutated tat gene and/or with a plasmid specifying the entire tatABC locus. A growth defect was immediately noted in the tat mutants as ~40-hr of growth at 37°C was necessary for isolated colonies of appreciable size to develop on agar plates, compared to ~20-hr for the parent strain O35E. Hence, we compared the growth of the tat mutants to that of the WT isolate O35E in liquid medium under aerobic conditions. This was accomplished by measuring the optical density (OD) of cultures over a 7-hr period. In some of these experiments, we also plated aliquots of the cultures to enumerate colony forming units (CFU) as a measure of bacterial viability.
The tatA, tatB and tatC genes are necessary for the secretion of β-lactamase by M. catarrhalis
Identification of other M. catarrhalisgene products potentially secreted by the TAT system
To identify other M. catarrhalis molecules that may be secreted via the TAT apparatus, the annotated genomic sequences of strains ATCC43617  and BBH18  were analyzed with prediction algorithms available through the TatFind 1.4  and TatP 1.0  servers. Although these servers are designed for the same purpose (i.e. identify proteins secreted by the TAT system), the algorithms used for each differ and as such proteins identified as TAT substrates do not overlap 100% between the two prediction algorithms.
Six putative TAT substrates were identified in the genome of M. catarrhalis strain BBH18, five of which overlapping those predicted in ATCC43617 (Figure 8). Strain BBH18 specifies the unique TAT substrate MCR_920, which is predicted to be a highly-conserved phosphatase (Figure 8). The MCORF 1659 of strain ATCC43617 encodes a gene product that is 96.8% identical to this putative phosphatase, but neither of the TatFind 1.4 and TatP 1.0 servers identified the ORF as a TAT substrate, likely due to significant amino acid divergence in the signal sequence (data not shown). Strain BBH18 specifies a putative C-terminal processing peptidase (MCR_1063) that is 98.1% identical to the putative TAT substrate MCORF 838 of ATCC43617. Like the MCORF 838 of ATCC43617, the BBH18 gene product lacked a TAT motif in its signal sequence (data not shown). Despite these similarities, the BBH18 MCR_1063 C-terminal processing peptidase was not identified as a being potential TAT substrate by the TatFind 1.4 or TatP 1.0 algorithms.
This report is the first characterization of a secretory apparatus for M. catarrhalis. Our data demonstrate that the TAT system mediates secretion of β-lactamase and is necessary for optimal growth of the bacterium. Moraxella catarrhalis is a leading cause of otitis media worldwide along with Streptococcus pneumoniae and non-typeable Haemophilus influenzae (NTHi), and is often found in mixed infections with these organisms [1–8, 89]. In contrast to M. catarrhalis, most S. pneumoniae and NTHi isolates are susceptible to β-lactam antibiotics . In a set of elegant studies, Schaar et al. demonstrated that outer membrane vesicles produced by M. catarrhalis contain β-lactamase and function as a long-distance delivery system to confer antimicrobial resistance for β-lactamase negative isolates of S. pneumoniae and NTHi . This constitutes a novel mechanism by which M. catarrhalis promotes survival and infection by other pathogens in the context of polymicrobial disease. Hence, a greater understanding of the TAT secretion system of M. catarrhalis is a key area of future study as it may lead to the development of innovative strategies to improve the efficacy of existing antimicrobials used to treat bacterial infections by common childhood pathogens. Small molecular weight compounds that selectively inhibit TAT secretion in M. catarrhalis could be used in concert with β-lactam antibiotics as β-lactamase inhibitors. This hypothesis is supported by the recent discovery that the compounds N-phenyl maleimide and Bay 11–7782 specifically interfere with TAT-dependent secretion of the Pseudomonas aeruginosa phospholipase C PlcH .
Bacterial strains, plasmids, and growth conditions
Strains and plasmids used in this study
WT isolate from middle ear effusion (Dallas, TX)
tatA isogenic mutant of strain O35E, kanR
tatB isogenic mutant of strain O35E, kanR
tatC isogenic mutant of strain O35E, kanR
bro-2 isogenic mutant of strain O35E, kanR
WT isolate from middle ear effusion (Dallas, TX)
tatC isogenic mutant of strain O12E, kanR
WT isolate from patient with respiratory infection (Toledo, OH)
WT isolate from transtracheal aspirate (Johnson City, TN)
WT isolate from nasopharynx of a healthy child (Chapel Hill, NC)
Host strain for cloning experiments with pWW115
Cloning vector, camR
pCC1-based plasmid containing kanR marker, camRkanR
Contains 886-nt insert specifying O35E tatA in pCC1, camR
Contains 858-nt insert specifying O35E tatB in pCC1, camR
Contains 1,018-nt insert specifying O12E tatC in pCC1, camR
pRB.TatC.2 in which a kanR marker disrupts the tatC ORF, camR kanR
Contains 2,083-nt insert specifying O35E tatABC locus in pCC1, camR
pRB.Tat.1 in which a kanR marker disrupts the tatA ORF, camR kanR
pRB.Tat.1 in which a kanR marker disrupts the tatB ORF, camR kanR
Contains 994-nt insert specifying O35E bro-2 in pCC1, camR
pRN.Bro11 in which a kanR marker disrupts the bro-2 ORF, camR kanR
pRN.Bro11 in which 2 arginines in the signal sequence of the bro-2 gene product were replaced with 2 lysines by site-directed mutagenesis, camR
M. catarrhalis-H. influenzae shuttle cloning vector, spcR
pWW115 into which the tatA insert of pRB.TatA.5 was subcloned, spcR
pWW115 into which the tatB insert of pRB.TatB.1 was subcloned, spcR
pWW115 into which the tatB insert of pRB.TatC.2 was subcloned, spcR
pWW115 into which the tatABC locus of pRB.Tat.1 was subcloned, spcR
pWW115 into which the bro-2 insert of pRN.Bro11 was subcloned, spcR
pWW115 into which the bro-2 insert of pTS.BroKK.Ec was subcloned, spcR
Recombinant DNA techniques
Standard molecular biology techniques were performed as described elsewhere . Moraxella catarrhalis genomic DNA was obtained using the Easy-DNA™ kit (Invitrogen™ Life Technologies™) per the manufacturer’s instructions. Plasmid DNA was purified with the QIAprep Spin Miniprep system (QIAGEN).
Polymerase chain reactions were performed using Taq DNA Polymerase (Invitrogen™ Life Technologies™) unless otherwise specified. A 1,018-nt fragment containing the tatC gene was amplified with primers P1 (5′- AAAGCCAAGCCAACGGACTT-3′) and P2 (5′-ACCTCCAAGAAACCCACGCTATCA-3′) using genomic DNA from M. catarrhalis strain O12E (see Figure 1 for more details regarding primers). This PCR product was cloned into the vector pCC1 using the Copy Control™ PCR Cloning Kit (Epicentre® Illumina®) per the manufacturer’s instructions. This process yielded plasmid pRB.TatC.2, which was sequenced to verify that mutations were not introduced in the tatC gene during cloning. PCR products comprising tatA (886-nt in length), tatB (858-nt in length) and the entire tatABC locus (2,083-nt in length) were amplified with primers P3 (5′-AGGGCAACTGGCAAATTACCAACC-3′) and P4 (5′-AAACATGCCATACCATCGCCCAAG-3′), P5 (5′-CAAAGACTTGGGCAGTGCGGTAAA-3′) and P6 (5′-ATTCATTGGGCAGTAGAGCGACCA-3), and P7 (5′-CATCATTGCGGCCAAAGAGCTTGA-3′) and P8 (5′-AGCTTGCCGATCCAAACAGCTTTC-3′), respectively, using genomic DNA from M. catarrhalis strain O35E (see Figure 1 for more details regarding primers). These amplicons were cloned in the vector pCC1 as described above, producing plasmids pRB.TatA.5, pRB.TatB.1, and pRB.Tat.1. These constructs were sequenced to verify that mutations were not introduced in the tat genes during PCR. To examine conservation of the TatABC gene products, genomic DNA from M. catarrhalis strains O35E, O12E, McGHS1, V1171, and TTA37 was used to amplify 2.1-kb DNA fragments containing the entire tatABC locus with primer P7 and P8. These amplicons were sequenced in their entirety and the sequences were deposited in GenBank under accession numbers HQ906880 (O35E), HQ906881 (O12E), HQ906882 (McGHS1), HQ906883 (V1171), and HQ906884 (TTA37).
The bro-2 gene specifying the β-lactamase of M. catarrhalis strain O35E was amplified with primers P9 (5′-TAATGATGCAACGCCGTCAT-3′) and P10 (5′-GCTTGTTGGGTCATAAATTTCC-3′) using Platinum® Pfx DNA Polymerase (Invitrogen™ Life Technologies™). This 994-nt PCR product was cloned into pCC1 as described above, generating the construct pRN.Bro11. Upon sequencing, the bro-2 gene contained by pRN.Bro11 was found to be free of mutation. The nucleotide sequence of O35E bro-2 was deposited in GenBank under the accession number JF279451.
To create a tatC mutation in M. catarrhalis, the plasmid pRB.TatC.2 was mutagenized with the EZ-TN5™ < KAN-2 > Insertion Kit (Epicentre® Illumina®) and introduced into Transformax™ EPI300™ electrocompetent cells. Chloramphenicol resistant (camR, specified by the vector pCC1) and kanamycin resistant (kanR, specified by the EZ-TN5 < KAN-2 > TN) colonies were selected and plasmids were analyzed by PCR using the pCC1-specific primer, P11 (5′-TACGCCAAGCTATTTAGGTGAGA-3′), and primers specific for the kanR marker, P12 (5′-ACCTACAACAAAGCTCTCATCAACC-3′) and P13 (5′-GCAATGTAACATCAGAGATTTTGAG-3′). This strategy identified plasmid pRB.TatC:kan, in which the EZ-TN5 < KAN-2 > TN was inserted near the middle of the tatC ORF. The disrupted tatC gene was then amplified from pRB.TatC:kan with the pCC1-specific primers P11 and P14 (5′-TAATACGACTCACTATAGGG-3′) using Platinum® Pfx DNA Polymerase. This 2.3-kb PCR product was purified and electroporated into M. catarrhalis strains O12E and O35E to create the kanR isogenic mutant strains O12E.TC and O35E.TC via homologous recombination. Allelic replacement was confirmed by PCR with the tatC primers P1 and P2 using Platinum® Pfx DNA Polymerase. These primers yielded PCR products in the mutant strains that were 1.2-kb larger than the amplicons obtained in the wild-type (WT) isolates O35E and O12E due to the presence of the EZ-TN5 < KAN-2 > TN in tatC.
To construct mutations in the tatA and tatB genes of M. catarrhalis O35E, the plasmid pRB.Tat.1 was first mutagenized with the EZ-TN5™ In-Frame Linker Insertion Kit (Epicentre® Illumina®) and introduced into Transformax™ EPI300™ electrocompetent cells. Plasmid DNA was isolated from several camR (specified by the vector pCC1) and kanR (specified by the EZ-TN5 < Not I/KAN-3 > TN) clones and sequenced to determine the sites of insertion of the TN. This approach identified the plasmids pRB.TatA:kan and pRB.TatB:kan, which contained the EZ-TN5 < Not I/KAN-3 > TN at nt 90 of the tatA ORF and nt 285 of the tatB ORF, respectively. These plasmids were then introduced into M. catarrhalis strain O35E by natural transformation as previously described . The resulting kanR strains were screened by PCR using primers specific for tatA (P3 and P4) and tatB (P5 and P6), which produced DNA fragments that were 1.2-kb larger in size in mutant strains when compared to the WT strain O35E because of the insertion of the EZ-TN5 < Not I/KAN-3 > TN in tatA and tatB. This strategy yielded the mutant strains O35E.TA and O35E.TB.
To construct a mutation in the bro-2 gene of M. catarrhalis O35E, plasmid pRN.Bro11 was mutagenized with the EZ-TN5™ In-Frame Linker Insertion Kit as described above. Plasmids were isolated from kanR camR colonies and sequenced to identify constructs containing the EZ-TN5 < Not I/KAN-3 > TN near the middle of the bro-2 ORF. This approach yielded the construct pRB.Bro:kan, which was introduced in M. catarrhalis O35E by natural transformation. Transformants were selected for resistance to kanamycin and then tested for their ability to grow on agar plates containing the β-lactam antibiotic carbenicillin. KanR and carbenicillin sensitive (cabS) strains were further analyzed by PCR using primers P9 and P10 to verify allelic exchange of the bro-2 gene. These primers produced a 1-kb DNA fragment in the WT strain O35E and a 2.2-kb in the mutant O35E.Bro, which is consistent with insertion of the 1.2-kb EZ-TN5 < Not I/KAN-3 > TN in bro-2.
Site-directed mutagenesis of the M. catarrhalis bro-2gene
The bro-2 ORF of M. catarrhalis O35E harbored by plasmid pRN.Bro11 was mutated using the QuikChange Lightning Site-Directed Mutagenesis Kit (Agilent Technologies) according to the manufacturer’s instructions. The mutagenesis primers, P15 (5′- AAGGGGATAATGATGCAAAAGAAGCATTTTTTA-3′) and P16 (5′-GGTTTTTTGTAAAAAATGCTTCTTTTGCAT CAT-3′), were used to replace two arginine residues at position 4 and 5 of BRO-2 with two lysines, yielding plasmid pTS.BroKK.Ec. This plasmid was sequenced to verify that only the intended mutations were introduced in the bro-2 ORF.
Complementation of mutants
The construction of plasmids to complement tat and bro2 mutant strains was achieved as follows. Plasmid DNA (pRB.TatA.5, pRB.TatB.1, pRB.TatC.2, pRB.Tat.1, pRN.Bro11, pTS.BroKK.Ec) was digested with BamHI to release the cloned M. catarrhalis genes from the vector pCC1. Gene fragments were purified from agarose gel slices using the High Pure PCR Product Purification Kit (Roche Applied Science), ligated into the BamHI site of the M. catarrhalis/Haemophilus influenza-compatible shuttle vector pWW115 , and electroporated into H. influenzae strain DB117. Spectinomycin resistant (spcR) colonies were screened by PCR using the pWW115-specific primers P17 (5′-TACGCCCTTTTATACTGTAG-3′) and P18 (5′-AACGACAGGAGCACGATCAT-3′), which flank the BamHI cloning site, to identify clones containing inserts of the appropriate size for the tat and bro2 genes. This process produced plasmids pRB.TatA, pRB.TatB, pRB.TatC, pRB.TAT, pTS.Bro, and pTS.BroKK. The O35E.TA mutant was naturally transformed with plasmids pWW115, pRB.TatA, and pRB.TatABC. The plasmids pWW115, pRB.TatB, and pRB.TAT were introduced in the O35E.TB mutant by natural transformation. The tatC mutants O35E.TC and O12E.TC were naturally transformed with the vector pWW115 and plasmid pRB.TatC. The plasmids pWW115, pTS.Bro, and pTS.BroKK were electroporated into the bro-2 mutant strain O35E.Bro. The successful introduction of these plasmids into the indicated strains was verified by PCR analysis of spcR transformants with the pWW115-specific primers P17 and P18, and by restriction endonuclease analysis of plasmid DNA purified from each strain.
Growth rate experiments
Moraxella. catarrhalis strains were first cultured onto agar plates supplemented with appropriate antibiotics. These plate-grown bacteria were used to inoculate 500-mL sidearm flasks containing 20-mL of broth (without antibiotics) to an optical density (OD) of 50 Klett units. The cultures were then incubated with shaking (225-rpm) at a temperature of 37°C for 7-hr. The OD of each culture was determined every 60-min using a Klett™ Colorimeter (Scienceware®). These experiments were repeated on at least three separate occasions for each strain. In some experiments, aliquots were taken out of each culture after recording the optical density, diluted, and spread onto agar plates to determine the number of viable colony forming units (CFU).
Carbenicillin sensitivity assays
Moraxella catarrhalis strains were first cultured onto agar plates supplemented with the appropriate antibiotics. These plate-grown bacteria were used to inoculate sterile Klett tubes containing five-mL of broth (without antibiotics) to an OD of 40 Klett units. Portions of these suspensions (25 μL) were spotted onto agar medium without antibiotics as well on plates supplemented with carbenicillin, and incubated at 37°C for 48-hr to evaluate growth. Each strain was tested in this manner a minimum of three times.
Moraxella catarrhalis and H. influenzae strains were tested for their ability to cleave the chromogenic β-lactamase substrate nitrocefin as previously described . Bacterial strains were first cultured onto agar plates supplemented with appropriate antibiotics. These plate-grown cells were suspended to an OD of 300 Klett units in 5-mL of broth, and aliquots (50 μL, ~107 CFU) were transferred to duplicate wells of a 48-well tissue culture plate; control wells were seeded with broth only. To each of these wells, 325 μL of a nitrocefin (Calbiochem®) solution (250 μg/mL in phosphate buffer) was added and the absorbance at a wavelength of 486 nm (A486) was immediately measured using a μQuant™ Microplate Spectrophotometer (BioTek®) and recorded as time “0”. The A486 of the samples was then measured after a 30-min incubation at room temperature. These experiments were repeated a minimum of three times for each strain.
Sequence analyses and TAT prediction Programs
Sequencing results were analyzed and assembled using Sequencher® 4.9 (Gene Codes Corporation). Sequence analyses and comparisons were performed using the various tools available through the ExPASy Proteomics Server (http://au.expasy.org/) and NCBI (http://blast.ncbi.nlm.nih.gov). To identify potential TAT substrates of M. catarrhalis, annotated nucleotide sequences from strain ATCC43617  were translated and analyzed with the prediction algorithms available through the TatFind 1.4 (http://signalfind.org/tatfind.html)  and TatP 1.0 (http://www.cbs.dtu.dk/services/TatP/)  servers using the default settings. The published genomic sequence of M. catarrhalis strain BBH18  was analyzed in the same manner.
The GraphPad Prism Software was used for all statistical analyses. Growth rate experiments and nitrocefin assays were analyzed by a two-way analysis of variants (ANOVA), followed by the Bonferroni post-test of the means of each time point. Asterisks indicate statistically significant differences where P < 0.05.
Open reading frame
Absorbance at a wavelength of 486 nm, TAT, twin-arginine translocation
Colony forming units
This study was supported by a grant from NIH/NIAID (AI051477) and startup funds from the University of Georgia College of Veterinary Medicine to ERL.
- Cripps AW, Otczyk DC, Kyd JM: Bacterial otitis media: a vaccine preventable disease?. Vaccine. 2005, 23 (17–18): 2304-2310.PubMedView ArticleGoogle Scholar
- Giebink GS, Kurono Y, Bakaletz LO, Kyd JM, Barenkamp SJ, Murphy TF, Green B, Ogra PL, Gu XX, Patel JA, et al: Recent advances in otitis media. 6. Vaccine. Ann Otol Rhinol Laryngol Suppl. 2005, 194: 86-103.PubMedGoogle Scholar
- Karalus R, Campagnari A: Moraxella catarrhalis: a review of an important human mucosal pathogen. Microbes Infect. 2000, 2 (5): 547-559. 10.1016/S1286-4579(00)00314-2.PubMedView ArticleGoogle Scholar
- Murphy TF: Vaccine development for non-typeable Haemophilus influenzae and Moraxella catarrhalis: progress and challenges. Expert Rev Vaccines. 2005, 4 (6): 843-853. 10.1586/147605184.108.40.2063.PubMedView ArticleGoogle Scholar
- Pichichero ME, Casey JR: Otitis media. Expert Opin Pharmacother. 2002, 3 (8): 1073-1090. 10.1517/146565220.127.116.113.PubMedView ArticleGoogle Scholar
- Verduin CM, Hol C, Fleer A, van Dijk H, van Belkum A: Moraxella catarrhalis: from emerging to established pathogen. Clin Microbiol Rev. 2002, 15 (1): 125-144. 10.1128/CMR.15.1.125-144.2002.PubMedPubMed CentralView ArticleGoogle Scholar
- Faden H: The microbiologic and immunologic basis for recurrent otitis media in children. Eur J Pediatr. 2001, 160 (7): 407-413. 10.1007/s004310100754.PubMedView ArticleGoogle Scholar
- Enright MC, McKenzie H: Moraxella (Branhamella) catarrhalis–clinical and molecular aspects of a rediscovered pathogen. J Med Microbiol. 1997, 46 (5): 360-371. 10.1099/00222615-46-5-360.PubMedView ArticleGoogle Scholar
- Arguedas A, Kvaerner K, Liese J, Schilder AG, Pelton SI: Otitis media across nine countries: disease burden and management. Int J Pediatr Otorhinolaryngol. 2010, 74 (12): 1419-1424. 10.1016/j.ijporl.2010.09.022.PubMedView ArticleGoogle Scholar
- Subcommittee on Management of Acute Otitis Media: Diagnosis and management of acute otitis media. Pediatrics. 2004, 113 (5): 1451-1465.View ArticleGoogle Scholar
- Del Beccaro MA, Mendelman PM, Inglis AF, Richardson MA, Duncan NO, Clausen CR, Stull TL: Bacteriology of acute otitis media: a new perspective. J Pediatr. 1992, 120 (1): 81-84. 10.1016/S0022-3476(05)80605-5.PubMedView ArticleGoogle Scholar
- Faden H, Duffy L, Wasielewski R, Wolf J, Krystofik D, Tung Y: Relationship between nasopharyngeal colonization and the development of otitis media in children. Tonawanda/Williamsville Pediatrics. J Infect Dis. 1997, 175 (6): 1440-1445. 10.1086/516477.PubMedView ArticleGoogle Scholar
- Faden H, Stanievich J, Brodsky L, Bernstein J, Ogra PL: Changes in nasopharyngeal flora during otitis media of childhood. Pediatr Infect Dis J. 1990, 9 (9): 623-626.PubMedGoogle Scholar
- Ruuskanen O, Heikkinen T: Otitis media: etiology and diagnosis. Pediatr Infect Dis J. 1994, 13 (1 Suppl 1): S23-S26. discussion S50-S54PubMedView ArticleGoogle Scholar
- Stool SE, Field MJ: The impact of otitis media. Pediatr Infect Dis J. 1989, 8 (1 Suppl): S11-S14.PubMedGoogle Scholar
- Klein JO: Otitis media. Clin Infect Dis. 1994, 19 (5): 823-833. 10.1093/clinids/19.5.823.PubMedView ArticleGoogle Scholar
- Klein JO: The burden of otitis media. Vaccine. 2000, 19 (Suppl 1): S2-S8.PubMedView ArticleGoogle Scholar
- Klein JO, Teele DW, Pelton SI: New concepts in otitis media: results of investigations of the Greater Boston Otitis Media Study Group. Adv Pediatr. 1992, 39: 127-156.PubMedGoogle Scholar
- Murphy TF: Branhamella catarrhalis: epidemiology, surface antigenic structure, and immune response. Microbiol Rev. 1996, 60 (2): 267-279.PubMedPubMed CentralGoogle Scholar
- Murphy TF, Brauer AL, Grant BJ, Sethi S: Moraxella catarrhalis in chronic obstructive pulmonary disease: burden of disease and immune response. Am J Respir Crit Care Med. 2005, 172 (2): 195-199. 10.1164/rccm.200412-1747OC.PubMedPubMed CentralView ArticleGoogle Scholar
- Sethi S, Evans N, Grant BJ, Murphy TF: New strains of bacteria and exacerbations of chronic obstructive pulmonary disease. N Engl J Med. 2002, 347 (7): 465-471. 10.1056/NEJMoa012561.PubMedView ArticleGoogle Scholar
- Sethi S, Murphy TF: Bacterial Infection in Chronic Obstructive Pulmonary Disease in 2000: a State-of-the-Art Review. Clin Microbiol Rev. 2001, 14 (2): 336-363. 10.1128/CMR.14.2.336-363.2001.PubMedPubMed CentralView ArticleGoogle Scholar
- (NHLBI) NIoH: Morbidity and Mortality: 2009 Chart Book on Cardiovascular, Lung, and Blood Diseases. 2009, http://wwwnhlbinihgov/resources/docs/2009_ChartBookpdf,Google Scholar
- Strassels SA, Smith DH, Sullivan SD, Mahajan PS: The costs of treating COPD in the United States. Chest. 2001, 119 (2): 344-352. 10.1378/chest.119.2.344.PubMedView ArticleGoogle Scholar
- Sullivan SD, Ramsey SD, Lee TA: The economic burden of COPD. Chest. 2000, 117 (2 Suppl): 5S-9S.PubMedView ArticleGoogle Scholar
- Hunter MH, King DE: COPD: management of acute exacerbations and chronic stable disease. Am Fam Physician. 2001, 64 (4): 603-612.PubMedGoogle Scholar
- Hurd S: The impact of COPD on lung health worldwide: epidemiology and incidence. Chest. 2000, 117 (2 Suppl): 1S-4S.PubMedView ArticleGoogle Scholar
- Lafontaine ER, Cope LD, Aebi C, Latimer JL, McCracken GH, Hansen EJ: The UspA1 protein and a second type of UspA2 protein mediate adherence of Moraxella catarrhalis to human epithelial cells in vitro. J Bacteriol. 2000, 182 (5): 1364-1373. 10.1128/JB.182.5.1364-1373.2000.PubMedPubMed CentralView ArticleGoogle Scholar
- Lipski SL, Akimana C, Timpe JM, Wooten RM, Lafontaine ER: The Moraxella catarrhalis autotransporter McaP is a conserved surface protein that mediates adherence to human epithelial cells through its N-terminal passenger domain. Infect Immun. 2007, 75 (1): 314-324. 10.1128/IAI.01330-06.PubMedPubMed CentralView ArticleGoogle Scholar
- Timpe JM, Holm MM, Vanlerberg SL, Basrur V, Lafontaine ER: Identification of a Moraxella catarrhalis outer membrane protein exhibiting both adhesin and lipolytic activities. Infect Immun. 2003, 71 (8): 4341-4350. 10.1128/IAI.71.8.4341-4350.2003.PubMedPubMed CentralView ArticleGoogle Scholar
- Holm MM, Vanlerberg SL, Foley IM, Sledjeski DD, Lafontaine ER: The Moraxella catarrhalis porin-like outer membrane protein CD is an adhesin for human lung cells. Infect Immun. 2004, 72 (4): 1906-1913. 10.1128/IAI.72.4.1906-1913.2004.PubMedPubMed CentralView ArticleGoogle Scholar
- Balder R, Krunkosky TM, Nguyen CQ, Feezel L, Lafontaine ER: Hag mediates adherence of Moraxella catarrhalis to ciliated human airway cells. Infect Immun. 2009, 77 (10): 4597-4608. 10.1128/IAI.00212-09.PubMedPubMed CentralView ArticleGoogle Scholar
- Bullard B, Lipski SL, Lafontaine ER: Hag directly mediates the adherence of Moraxella catarrhalis to human middle ear cells. Infect Immun. 2005, 73 (8): 5127-5136. 10.1128/IAI.73.8.5127-5136.2005.PubMedPubMed CentralView ArticleGoogle Scholar
- Balder R, Hassel J, Lipski S, Lafontaine ER: Moraxella catarrhalis strain O35E expresses two filamentous hemagglutinin-like proteins that mediate adherence to human epithelial cells. Infect Immun. 2007, 75 (6): 2765-2775. 10.1128/IAI.00079-07.PubMedPubMed CentralView ArticleGoogle Scholar
- Plamondon P, Luke NR, Campagnari AA: Identification of a novel two-partner secretion locus in Moraxella catarrhalis. Infect Immun. 2007, 75 (6): 2929-2936. 10.1128/IAI.00396-07.PubMedPubMed CentralView ArticleGoogle Scholar
- Luke NR, Jurcisek JA, Bakaletz LO, Campagnari AA: Contribution of Moraxella catarrhalis type IV pili to nasopharyngeal colonization and biofilm formation. Infect Immun. 2007, 75 (12): 5559-5564. 10.1128/IAI.00946-07.PubMedPubMed CentralView ArticleGoogle Scholar
- Peng D, Hu WG, Choudhury BP, Muszynski A, Carlson RW, Gu XX: Role of different moieties from the lipooligosaccharide molecule in biological activities of the Moraxella catarrhalis outer membrane. FEBS J. 2007, 274 (20): 5350-5359. 10.1111/j.1742-4658.2007.06060.x.PubMedView ArticleGoogle Scholar
- Attia AS, Ram S, Rice PA, Hansen EJ: Binding of vitronectin by the Moraxella catarrhalis UspA2 protein interferes with late stages of the complement cascade. Infect Immun. 2006, 74 (3): 1597-1611. 10.1128/IAI.74.3.1597-1611.2006.PubMedPubMed CentralView ArticleGoogle Scholar
- Hallstrom T, Nordstrom T, Tan TT, Manolov T, Lambris JD, Isenman DE, Zipfel PF, Blom AM, Riesbeck K: Immune evasion of Moraxella catarrhalis involves ubiquitous surface protein A-dependent C3d binding. J Immunol. 2011, 186 (5): 3120-3129. 10.4049/jimmunol.1002621.PubMedView ArticleGoogle Scholar
- Nordstrom T, Blom AM, Forsgren A, Riesbeck K: The emerging pathogen Moraxella catarrhalis interacts with complement inhibitor C4b binding protein through ubiquitous surface proteins A1 and A2. J Immunol. 2004, 173 (7): 4598-4606.PubMedView ArticleGoogle Scholar
- Nordstrom T, Blom AM, Tan TT, Forsgren A, Riesbeck K: Ionic binding of C3 to the human pathogen Moraxella catarrhalis is a unique mechanism for combating innate immunity. J Immunol. 2005, 175 (6): 3628-3636.PubMedView ArticleGoogle Scholar
- Murphy TF, Brauer AL, Yuskiw N, Hiltke TJ: Antigenic structure of outer membrane protein E of Moraxella catarrhalis and construction and characterization of mutants. Infect Immun. 2000, 68 (11): 6250-6256. 10.1128/IAI.68.11.6250-6256.2000.PubMedPubMed CentralView ArticleGoogle Scholar
- Helminen ME, Maciver I, Paris M, Latimer JL, Lumbley SL, Cope LD, McCracken GH, Hansen EJ: A mutation affecting expression of a major outer membrane protein of Moraxella catarrhalis alters serum resistance and survival in vivo. J Infect Dis. 1993, 168 (5): 1194-1201. 10.1093/infdis/168.5.1194.PubMedView ArticleGoogle Scholar
- Jacobs MR, Bajaksouzian S, Windau A, Good CE, Lin G, Pankuch GA, Appelbaum PC: Susceptibility of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis to 17 oral antimicrobial agents based on pharmacodynamic parameters: 1998–2001 U S Surveillance Study. Clin Lab Med. 2004, 24 (2): 503-530. 10.1016/j.cll.2004.03.008.PubMedView ArticleGoogle Scholar
- Klugman KP: The clinical relevance of in-vitro resistance to penicillin, ampicillin, amoxycillin and alternative agents, for the treatment of community-acquired pneumonia caused by Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis. J Antimicrob Chemother. 1996, 38 (Suppl A): 133-140. 10.1093/jac/38.suppl_A.133.PubMedView ArticleGoogle Scholar
- Manninen R, Huovinen P, Nissinen A: Increasing antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in Finland. J Antimicrob Chemother. 1997, 40 (3): 387-392. 10.1093/jac/40.3.387.PubMedView ArticleGoogle Scholar
- Richter SS, Winokur PL, Brueggemann AB, Huynh HK, Rhomberg PR, Wingert EM, Doern GV: Molecular characterization of the beta-lactamases from clinical isolates of Moraxella (Branhamella) catarrhalis obtained from 24 U.S. medical centers during 1994–1995 and 1997–1998. Antimicrob Agents Chemother. 2000, 44 (2): 444-446. 10.1128/AAC.44.2.444-446.2000.PubMedPubMed CentralView ArticleGoogle Scholar
- Kadry AA, Fouda SI, Elkhizzi NA, Shibl AM: Correlation between susceptibility and BRO type enzyme of Moraxella catarrhalis strains. Int J Antimicrob Agents. 2003, 22 (5): 532-536. 10.1016/S0924-8579(03)00158-4.PubMedView ArticleGoogle Scholar
- Schmitz FJ, Beeck A, Perdikouli M, Boos M, Mayer S, Scheuring S, Kohrer K, Verhoef J, Fluit AC: Production of BRO beta-lactamases and resistance to complement in European Moraxella catarrhalis isolates. J Clin Microbiol. 2002, 40 (4): 1546-1548. 10.1128/JCM.40.4.1546-1548.2002.PubMedPubMed CentralView ArticleGoogle Scholar
- Johnson DM, Sader HS, Fritsche TR, Biedenbach DJ, Jones RN: Susceptibility trends of haemophilus influenzae and Moraxella catarrhalis against orally administered antimicrobial agents: five-year report from the SENTRY Antimicrobial Surveillance Program. Diagn Microbiol Infect Dis. 2003, 47 (1): 373-376. 10.1016/S0732-8893(03)00089-0.PubMedView ArticleGoogle Scholar
- Esel D, Ay-Altintop Y, Yagmur G, Gokahmetoglu S, Sumerkan B: Evaluation of susceptibility patterns and BRO beta-lactamase types among clinical isolates of Moraxella catarrhalis. Clin Microbiol Infect. 2007, 13 (10): 1023-1025. 10.1111/j.1469-0691.2007.01776.x.PubMedView ArticleGoogle Scholar
- Bootsma HJ, Aerts PC, Posthuma G, Harmsen T, Verhoef J, van Dijk H, Mooi FR: Moraxella (Branhamella) catarrhalis BRO beta-lactamase: a lipoprotein of gram-positive origin?. J Bacteriol. 1999, 181 (16): 5090-5093.PubMedPubMed CentralGoogle Scholar
- Bootsma HJ, van Dijk H, Vauterin P, Verhoef J, Mooi FR: Genesis of BRO beta-lactamase-producing Moraxella catarrhalis: evidence for transformation-mediated horizontal transfer. Mol Microbiol. 2000, 36 (1): 93-104. 10.1046/j.1365-2958.2000.01828.x.PubMedView ArticleGoogle Scholar
- Torretta S, Drago L, Marchisio P, Gaffuri M, Clemente IA, Pignataro L: Topographic distribution of biofilm-producing bacteria in adenoid subsites of children with chronic or recurrent middle ear infections. Ann Otol Rhinol Laryngol. 2013, 122 (2): 109-113.PubMedView ArticleGoogle Scholar
- Bakaletz LO: Bacterial biofilms in the upper airway - evidence for role in pathology and implications for treatment of otitis media. Paediatr Respir Rev. 2012, 13 (3): 154-159. 10.1016/j.prrv.2012.03.001.PubMedPubMed CentralView ArticleGoogle Scholar
- Armbruster CE, Hong W, Pang B, Weimer KE, Juneau RA, Turner J, Swords WE: Indirect Pathogenicity of Haemophilus influenzae and Moraxella catarrhalis in Polymicrobial Otitis Media Occurs via Interspecies Quorum Signaling. MBio. 2010, 1 (3): e00102-e00110.PubMedPubMed CentralView ArticleGoogle Scholar
- Hoa M, Tomovic S, Nistico L, Hall-Stoodley L, Stoodley P, Sachdeva L, Berk R, Coticchia JM: Identification of adenoid biofilms with middle ear pathogens in otitis-prone children utilizing SEM and FISH. Int J Pediatr Otorhinolaryngol. 2009, 73 (9): 1242-1248. 10.1016/j.ijporl.2009.05.016.PubMedView ArticleGoogle Scholar
- Hall-Stoodley L, Hu FZ, Gieseke A, Nistico L, Nguyen D, Hayes J, Forbes M, Greenberg DP, Dice B, Burrows A, et al: Direct detection of bacterial biofilms on the middle-ear mucosa of children with chronic otitis media. JAMA. 2006, 296 (2): 202-211. 10.1001/jama.296.2.202.PubMedPubMed CentralView ArticleGoogle Scholar
- Palmer T, Berks BC: The twin-arginine translocation (Tat) protein export pathway. Nat Rev Microbiol. 2012, 10 (7): 483-496.PubMedGoogle Scholar
- Sargent F: The twin-arginine transport system: moving folded proteins across membranes. Biochem Soc Trans. 2007, 35 (Pt 5): 835-847.PubMedView ArticleGoogle Scholar
- Berks BC, Palmer T, Sargent F: Protein targeting by the bacterial twin-arginine translocation (Tat) pathway. Curr Opin Microbiol. 2005, 8 (2): 174-181. 10.1016/j.mib.2005.02.010.PubMedView ArticleGoogle Scholar
- Lammertyn E, Anne J: Protein secretion in Legionella pneumophila and its relation to virulence. FEMS Microbiol Lett. 2004, 238 (2): 273-279.PubMedGoogle Scholar
- He H, Wang Q, Sheng L, Liu Q, Zhang Y: Functional characterization of Vibrio alginolyticus twin-arginine translocation system: its roles in biofilm formation, extracellular protease activity, and virulence towards fish. Curr Microbiol. 2011, 62 (4): 1193-1199. 10.1007/s00284-010-9844-6.PubMedView ArticleGoogle Scholar
- Truan D, Vasil A, Stonehouse M, Vasil ML, Pohl E: High-level over-expression, purification, and crystallization of a novel phospholipase C/sphingomyelinase from Pseudomonas aeruginosa. Protein Expr Purif. 2013, 90 (1): 40-46. 10.1016/j.pep.2012.11.005.PubMedPubMed CentralView ArticleGoogle Scholar
- Sitkiewicz I, Stockbauer KE, Musser JM: Secreted bacterial phospholipase A2 enzymes: better living through phospholipolysis. Trends Microbiol. 2007, 15 (2): 63-69. 10.1016/j.tim.2006.12.003.PubMedView ArticleGoogle Scholar
- Shi ZY, Wang H, Gu L, Cui ZG, Wu LF, Kan B, Pang B, Wang X, Xu JG, Jing HQ: Pleiotropic effect of tatC mutation on metabolism of pathogen Yersinia enterocolitica. Biomed Environ Sci. 2007, 20 (6): 445-449.PubMedGoogle Scholar
- Lavander M, Ericsson SK, Broms JE, Forsberg A: Twin arginine translocation in Yersinia. Adv Exp Med Biol. 2007, 603: 258-267. 10.1007/978-0-387-72124-8_23.PubMedView ArticleGoogle Scholar
- Caldelari I, Mann S, Crooks C, Palmer T: The Tat pathway of the plant pathogen Pseudomonas syringae is required for optimal virulence. Mol Plant Microbe Interact. 2006, 19 (2): 200-212. 10.1094/MPMI-19-0200.PubMedView ArticleGoogle Scholar
- Bronstein PA, Marrichi M, Cartinhour S, Schneider DJ, DeLisa MP: Identification of a twin-arginine translocation system in Pseudomonas syringae pv. tomato DC3000 and its contribution to pathogenicity and fitness. J Bacteriol. 2005, 187 (24): 8450-8461. 10.1128/JB.187.24.8450-8461.2005.PubMedPubMed CentralView ArticleGoogle Scholar
- Ochsner UA, Snyder A, Vasil AI, Vasil ML: Effects of the twin-arginine translocase on secretion of virulence factors, stress response, and pathogenesis. Proc Natl Acad Sci USA. 2002, 99 (12): 8312-8317. 10.1073/pnas.082238299.PubMedPubMed CentralView ArticleGoogle Scholar
- Feltcher ME, Sullivan JT, Braunstein M: Protein export systems of Mycobacterium tuberculosis: novel targets for drug development?. Future Microbiol. 2010, 5 (10): 1581-1597. 10.2217/fmb.10.112.PubMedPubMed CentralView ArticleGoogle Scholar
- McDonough JA, McCann JR, Tekippe EM, Silverman JS, Rigel NW, Braunstein M: Identification of functional Tat signal sequences in Mycobacterium tuberculosis proteins. J Bacteriol. 2008, 190 (19): 6428-6438. 10.1128/JB.00749-08.PubMedPubMed CentralView ArticleGoogle Scholar
- McCann JR, McDonough JA, Pavelka MS, Braunstein M: Beta-lactamase can function as a reporter of bacterial protein export during Mycobacterium tuberculosis infection of host cells. Microbiology. 2007, 153 (Pt 10): 3350-3359.PubMedPubMed CentralView ArticleGoogle Scholar
- McDonough JA, Hacker KE, Flores AR, Pavelka MS, Braunstein M: The twin-arginine translocation pathway of Mycobacterium smegmatis is functional and required for the export of mycobacterial beta-lactamases. J Bacteriol. 2005, 187 (22): 7667-7679. 10.1128/JB.187.22.7667-7679.2005.PubMedPubMed CentralView ArticleGoogle Scholar
- Heikkila MP, Honisch U, Wunsch P, Zumft WG: Role of the Tat ransport system in nitrous oxide reductase translocation and cytochrome cd1 biosynthesis in Pseudomonas stutzeri. J Bacteriol. 2001, 183 (5): 1663-1671. 10.1128/JB.183.5.1663-1671.2001.PubMedPubMed CentralView ArticleGoogle Scholar
- Stevenson LG, Strisovsky K, Clemmer KM, Bhatt S, Freeman M, Rather PN: Rhomboid protease AarA mediates quorum-sensing in Providencia stuartii by activating TatA of the twin-arginine translocase. Proc Natl Acad Sci USA. 2007, 104 (3): 1003-1008. 10.1073/pnas.0608140104.PubMedPubMed CentralView ArticleGoogle Scholar
- Chen L, Hu B, Qian G, Wang C, Yang W, Han Z, Liu F: Identification and molecular characterization of twin-arginine translocation system (Tat) in Xanthomonas oryzae pv. oryzae strain PXO99. Arch Microbiol. 2009, 191 (2): 163-170. 10.1007/s00203-008-0440-0.PubMedView ArticleGoogle Scholar
- de Vries SP, van Hijum SA, Schueler W, Riesbeck K, Hays JP, Hermans PW, Bootsma HJ: Genome analysis of Moraxella catarrhalis strain RH4, a human respiratory tract pathogen. J Bacteriol. 2010, 192 (14): 3574-3583. 10.1128/JB.00121-10.PubMedPubMed CentralView ArticleGoogle Scholar
- Stanley NR, Findlay K, Berks BC, Palmer T: Escherichia coli strains blocked in Tat-dependent protein export exhibit pleiotropic defects in the cell envelope. J Bacteriol. 2001, 183 (1): 139-144. 10.1128/JB.183.1.139-144.2001.PubMedPubMed CentralView ArticleGoogle Scholar
- Saint-Joanis B, Demangel C, Jackson M, Brodin P, Marsollier L, Boshoff H, Cole ST: Inactivation of Rv2525c, a substrate of the twin arginine translocation (Tat) system of Mycobacterium tuberculosis, increases beta-lactam susceptibility and virulence. J Bacteriol. 2006, 188 (18): 6669-6679. 10.1128/JB.00631-06.PubMedPubMed CentralView ArticleGoogle Scholar
- Wang W, Reitzer L, Rasko DA, Pearson MM, Blick RJ, Laurence C, Hansen EJ: Metabolic analysis of Moraxella catarrhalis and the effect of selected in vitro growth conditions on global gene expression. Infect Immun. 2007, 75 (10): 4959-4971. 10.1128/IAI.00073-07.PubMedPubMed CentralView ArticleGoogle Scholar
- Rose RW, Bruser T, Kissinger JC, Pohlschroder M: Adaptation of protein secretion to extremely high-salt conditions by extensive use of the twin-arginine translocation pathway. Mol Microbiol. 2002, 45 (4): 943-950. 10.1046/j.1365-2958.2002.03090.x.PubMedView ArticleGoogle Scholar
- Bendtsen JD, Nielsen H, Widdick D, Palmer T, Brunak S: Prediction of twin-arginine signal peptides. BMC Bioinformatics. 2005, 6: 167-10.1186/1471-2105-6-167.PubMedPubMed CentralView ArticleGoogle Scholar
- Sturm A, Schierhorn A, Lindenstrauss U, Lilie H, Bruser T: YcdB from Escherichia coli reveals a novel class of Tat-dependently translocated hemoproteins. J Biol Chem. 2006, 281 (20): 13972-13978. 10.1074/jbc.M511891200.PubMedView ArticleGoogle Scholar
- van Bloois E, Torres Pazmino DE, Winter RT, Fraaije MW: A robust and extracellular heme-containing peroxidase from Thermobifida fusca as prototype of a bacterial peroxidase superfamily. Appl Microbiol Biotechnol. 2010, 86 (5): 1419-1430. 10.1007/s00253-009-2369-x.PubMedPubMed CentralView ArticleGoogle Scholar
- Bachmann J, Bauer B, Zwicker K, Ludwig B, Anderka O: The Rieske protein from Paracoccus denitrificans is inserted into the cytoplasmic membrane by the twin-arginine translocase. FEBS J. 2006, 273 (21): 4817-4830. 10.1111/j.1742-4658.2006.05480.x.PubMedView ArticleGoogle Scholar
- Sanders C, Wethkamp N, Lill H: Transport of cytochrome c derivatives by the bacterial Tat protein translocation system. Mol Microbiol. 2001, 41 (1): 241-246. 10.1046/j.1365-2958.2001.02514.x.PubMedView ArticleGoogle Scholar
- Webb DC, Rosenberg H, Cox GB: Mutational analysis of the Escherichia coli phosphate-specific transport system, a member of the traffic ATPase (or ABC) family of membrane transporters. A role for proline residues in transmembrane helices. J Biol Chem. 1992, 267 (34): 24661-24668.PubMedGoogle Scholar
- Krishnamurthy A, McGrath J, Cripps AW, Kyd JM: The incidence of Streptococcus pneumoniae otitis media is affected by the polymicrobial environment particularly Moraxella catarrhalis in a mouse nasal colonisation model. Microbes Infect. 2009, 11 (5): 545-553. 10.1016/j.micinf.2009.03.001.PubMedView ArticleGoogle Scholar
- Darabi A, Hocquet D, Dowzicky MJ: Antimicrobial activity against Streptococcus pneumoniae and Haemophilus influenzae collected globally between 2004 and 2008 as part of the Tigecycline Evaluation and Surveillance Trial. Diagn Microbiol Infect Dis. 2010, 67 (1): 78-86. 10.1016/j.diagmicrobio.2009.12.009.PubMedView ArticleGoogle Scholar
- Schaar V, Nordstrom T, Morgelin M, Riesbeck K: Moraxella catarrhalis outer membrane vesicles carry beta-lactamase and promote survival of Streptococcus pneumoniae and Haemophilus influenzae by inactivating amoxicillin. Antimicrob Agents Chemother. 2011, 55 (8): 3845-3853. 10.1128/AAC.01772-10.PubMedPubMed CentralView ArticleGoogle Scholar
- Vasil ML, Tomaras AP, Pritchard AE: Identification and evaluation of twin-arginine translocase inhibitors. Antimicrob Agents Chemother. 2012, 56 (12): 6223-6234. 10.1128/AAC.01575-12.PubMedPubMed CentralView ArticleGoogle Scholar
- Holm MM, Vanlerberg SL, Sledjeski DD, Lafontaine ER: The Hag protein of Moraxella catarrhalis strain O35E is associated with adherence to human lung and middle ear cells. Infect Immun. 2003, 71 (9): 4977-4984. 10.1128/IAI.71.9.4977-4984.2003.PubMedPubMed CentralView ArticleGoogle Scholar
- Aebi C, Lafontaine ER, Cope LD, Latimer JL, Lumbley SL, McCracken GH, Hansen EJ: Phenotypic effect of isogenic uspA1 and uspA2 mutations on Moraxella catarrhalis 035E. Infect Immun. 1998, 66 (7): 3113-3119.PubMedPubMed CentralGoogle Scholar
- Wang W, Hansen EJ: Plasmid pWW115, a cloning vector for use with Moraxella catarrhalis. Plasmid. 2006, 56 (2): 133-137. 10.1016/j.plasmid.2006.03.002.PubMedView ArticleGoogle Scholar
- Setlow JK, Brown DC, Boling ME, Mattingly A, Gordon MP: Repair of deoxyribonucleic acid in Haemophilus influenzae. I. X-ray sensitivity of ultraviolet-sensitive mutants and their behavior as hosts to ultraviolet-irradiated bacteriophage and transforming deoxyribonucleic acid. J Bacteriol. 1968, 95 (2): 546-558.PubMedPubMed CentralGoogle Scholar
- Sambrook J, Russell DW: Molecular Cloning: A Laboratory Manual. 2001, Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press, ThirdGoogle Scholar
- Pearson MM, Hansen EJ: Identification of Gene Products Involved in Biofilm Production by Moraxella catarrhalis ETSU-9 In Vitro. Infect Immun. 2007, 75 (9): 4316-4325. 10.1128/IAI.01347-06.PubMedPubMed CentralView ArticleGoogle Scholar