- Volume 61, Issue 6, 2012
Volume 61, Issue 6, 2012
- Review
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Granulicatella infection: diagnosis and management
More LessGranulicatella species, along with the genus Abiotrophia, were originally known as ‘nutritionally variant streptococci’. They are a normal component of the oral flora, but have been associated with a variety of invasive infections in man and are most noted as a cause of bacterial endocarditis. It is often advised that Granulicatella endocarditis should be treated in the same way as enterococcal endocarditis. We review here the published data concerning diagnosis and treatment of Granulicatella infection, and include some observations from local cases, including four cases of endocarditis.
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- Pathogenicity and virulence
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Colicin insensitivity correlates with a higher prevalence of extraintestinal virulence factors among Escherichia coli isolates from skin and soft-tissue infections
More LessColicins are toxic proteins with a narrow killing spectrum that are produced by colicinogenic Escherichia coli strains. The aim of this study was to analyse systematically whether extra-intestinal virulence potential is linked to colicin (in)sensitivity. In total, 102 well-characterized E. coli isolates from skin and soft-tissue infections (SSTIs) were exposed to 17 single-colicin-producing strains, and the correlation between insensitivity to colicin and phylogenetic group as well as the extra-intestinal virulence potential of the SSTI strains was examined. The results showed that SSTI strains belonging to the B2 phylogenetic group were statistically significantly associated with insensitivity to at least ten colicins, and several colicin insensitivities were correlated with virulence factors. As far as is known, this is the first study to report such correlations.
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- Host response
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Identification of conserved antigens from staphylococcal and streptococcal pathogens
The design of vaccines containing epitopes shared between different human pathogens may lead to cross-species protection. In order to identify potentially conserved bacterial antigens, bacteriophage expression libraries of genomic DNA from Streptococcus agalactiae, Streptococcus pneumoniae and Streptococcus pyogenes were probed with human sera from Staphylococcus aureus-infected and healthy individuals. By comparison with previous screening data from Staphylococcus epidermidis and Staph. aureus, putative antigenic, conserved domains across the genera were identified. In particular, three potentially antigenic conserved regions were identified based on the N-terminal domain of SACOL0609 (SdrD), the C-terminal domain of SACOL0723 (ScaB) and the C-terminus of SACOL1140 (IsdA) from Staph. aureus. The three domains were overexpressed, recombinant proteins were purified and polyclonal antisera raised against them recognized cell surface-located proteins from both staphylococcal and streptococcal species. The antisera were also able to opsonize both Staph. aureus and Strep. agalactiae thereby increasing their phagocytic uptake by human neutrophils. The conserved antigenic domains therefore represent potential cross-protective vaccine candidates.
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- Diagnostics, typing and identification
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High-resolution melting analysis of the single nucleotide polymorphism hot-spot region in the rpoB gene as an indicator of reduced susceptibility to rifaximin in Clostridium difficile
Clostridium difficile, a Gram-positive, spore-forming, anaerobic bacterium, is the main causative agent of hospital-acquired diarrhoea worldwide. In addition to metronidazole and vancomycin, rifaximin, a rifamycin derivative, is a promising antibiotic for the treatment of recurring C. difficile infections (CDI). However, exposure of C. difficile to this antibiotic has led to the development of rifaximin-resistance due to point mutations in the β-subunit of the RNA polymerase (rpoB) gene. In the present study, 348 C. difficile strains with known PCR-ribotypes were investigated for respective single nucleotide polymorphisms (SNPs) within the proposed rpoB hot-spot region by using high-resolution melting (HRM) analysis. This method allows the detection of SNPs by comparing the altered melting behaviour of dsDNA with that of wild-type DNA. Discrimination between wild-type and mutant strains was enhanced by creating heteroduplexes by mixing sample DNA with wild-type DNA, leading to characteristic melting curve shapes from samples containing SNPs in the respective rpoB section. In the present study, we were able to identify 16 different rpoB sequence-types (ST) by sequencing analysis of a 325 bp fragment. The 16 PCR STs displayed a total of 24 different SNPs. Fifteen of these 24 SNPs were located within the proposed 151 bp SNP hot-spot region, resulting in 11 different HRM curve profiles (CP). Eleven SNPs (seven of which were within the proposed hot-spot region) led to amino acid substitutions associated with reduced susceptibility to rifaximin and 13 SNPs (eight of which were within the hot-spot region) were synonymous. This investigation clearly demonstrates that HRM analysis of the proposed SNP hot-spot region in the rpoB gene of C. difficile is a fast and cost-effective method for the identification of C. difficile samples with reduced susceptibility to rifaximin and even allows simultaneous SNP subtyping of the respective C. difficile isolates.
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- Antimicrobial agents and chemotherapy
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Effect of 1-(1-naphthylmethyl)-piperazine on antimicrobial agent susceptibility in multidrug-resistant isogenic and veterinary Escherichia coli field strains
More LessThe objective of this study was to evaluate the interaction of the efflux pump inhibitor 1-(1-naphthylmethyl)-piperazine (NMP) when combined with different families of antimicrobial agents against isogenic strains and multidrug-resistant (MDR) Escherichia coli field strains isolated from animals. Laboratory isogenic strains of E. coli with different levels of expression of efflux pumps were used as quality controls. Ten MDR E. coli strains were collected from healthy animals in a cross-sectional study in four commercial dairy farms. The MICs of florfenicol, ciprofloxacin, tetracycline and ampicillin were determined by a serial microdilution method in Luria–Bertani broth in the presence or absence of NMP. NMP used with ampicillin exerted no effect on the isogenic or field strains. In most of the field MDRE. coli strains and in an acrAB-overexpressing (AG112) isogenic strain, the MICs of florfenicol, ciprofloxacin and tetracycline decreased at least fourfold when the antimicrobial was combined with the highest NMP concentrations. In the wild-type strain (AG100), there were no decreases of more than twice the MIC, whilst in strain AG100A, an efflux pump-deficient strain, the MIC did not change, regardless of the concentration of NMP used with these three antimicrobials. Thus, ampicillin was not affected by the efflux pump mechanism, whereas ciprofloxacin, tetracycline and florfenicol were shown to be substrates of efflux pumps, with a consequent significant reduction in MICs. Resistance could not be completely reversed in the E. coli field strains by NMP, probably because other resistance mechanisms were also present. However, in strain AG112, the MIC results demonstrated that NMP expressed an important synergistic activity with florfenicol. The reduction in florfenicol MIC value was sufficient to reverse antimicrobial resistance completely for AG112.
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Non-thermal argon plasma is bactericidal for the intracellular bacterial pathogen Chlamydia trachomatis
Svetlana A. Ermolaeva, Elena V. Sysolyatina, Natalia I. Kolkova, Petr Bortsov, Amir I. Tuhvatulin, Mikhail M. Vasiliev, Andrey Y. Mukhachev, Oleg F. Petrov, Shimizu Tetsuji, Boris S. Naroditsky, Gregor E. Morfill, Vladimir E. Fortov, Anatoly I. Grigoriev, Nelly A. Zigangirova and Alexander L. GintsburgNon-thermal plasma (NTP) is a flow of partially ionized argon gas at an ambient macroscopic temperature and is microbicidal for bacteria, viruses and fungi. Viability of the Gram-negative obligate intracellular bacterial parasite Chlamydia trachomatis and its host cells was investigated after NTP treatment. NTP treatment of C. trachomatis extracellular elementary bodies (EBs) diminished the concentration of infectious bacteria by a factor of 9×104, as established by the parallel infection of murine fibroblast McCoy cells with treated and control EBs. NTP treatment of infected McCoy cells caused disruption of membrane-restricted vacuoles (inclusions), where C. trachomatis intracellular reticulate bodies (RBs) multiply, and a 2×106-fold reduction in the concentration of infectious bacteria. When the samples were covered with magnesium fluoride glass to obstruct plasma particles and UV rays alone were applied, the bactericidal effect was reduced 1.4×101-fold and 5×104-fold for EBs and RBs, respectively. NTP treatment caused the viability of host McCoy cells to diminish by 19 %. Therefore, the results obtained demonstrated that (i) both extracellular and intracellular forms ofC. trachomatis are sensitive to NTP treatment; (ii) the reduction in concentration of infectious bacteria after NTP treatment of infected cells is superior to the reduction in viability of host cells; and (iii) the effect of NTP on intracellular bacteria does not depend on UV rays.
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Involvement of efflux mechanisms in biocide resistance of Campylobacter jejuni and Campylobacter coli
More LessActive efflux has an important role in the antimicrobial resistance of Campylobacter jejuni and Campylobacter coli. The effects of two putative efflux pump inhibitors (EPIs), phenylalanine-arginine β-naphthylamide and 1-(1-naphthylmethyl)-piperazine, and the effects of inactivation of the cmeB,cmeF and cmeR genes on resistance to a broad range of antimicrobials were studied using the broth microdilution method. The antimicrobials tested in C. jejuni and C. coli were the biocides triclosan, benzalkonium chloride, chlorhexidine diacetate, cetylpyridinium chloride and trisodium phosphate, along with the anionic surfactant SDS and the antibiotics erythromycin and ciprofloxacin. Both EPIs partially reversed the resistance to all of these antimicrobials. Differences between these EPIs were seen for substrate preference and reductions in MIC. The MICs of the antimicrobials were reduced in the cmeB and cmeF mutants and increased in the cmeR mutant, with few exceptions. Both of these putative EPIs further decreased the MICs of the antimicrobials in these mutant strains. These data confirm that active efflux is an important mechanism in biocide resistance in C. jejuni and C. coli. At least one non-CmeABC efflux system or reduced uptake is responsible for resistance to biocides.
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Detection of carbapenemase-producing Enterobacteriaceae with a commercial DNA microarray
More LessThe Check-MDR CT102 DNA microarray enables detection of the most prevalent carbapenemases (NDM, VIM, KPC, OXA-48 and IMP) and extended-spectrum β-lactamase (ESBL) gene families (SHV, TEM and CTX-M). The test performance of this microarray was evaluated with 95 Enterobacteriaceae isolates suspected of being carbapenemase producers, i.e. with meropenem MICs ≥0.5 mg l−1. The collection of isolates contained 70 carbapenemase-producing isolates, including 37 bla KPC-, 20 bla VIM-, five bla OXA-48-, four bla KPC/bla VIM- and four bla NDM-positive isolates; and 25 carbapenemase-gene-negative isolates. ESBLs were produced by 51 of the isolates. PCR and sequencing of β-lactamase genes was used as reference test. For detection of carbapenemases, the sensitivity of the microarray was 97 % (68/70), with 100 % specificity. The two negative isolates tested positive when the microarray test was repeated; these isolates were an OXA-48- and a KPC-producing isolate. For ESBL detection, the sensitivity was 100 % (51/51) and the specificity was 98 % (43/44), although 20 % of the SHV-12 ESBLs were categorized as SHV-2-like ESBLs. In conclusion, the CDT102 microarray is a rapid and accurate tool for the detection of carbapenemase and ESBL genes, although the array seems less suitable for epidemiology of ESBL genes.
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Transcription factor Efg1 contributes to the tolerance of Candida albicans biofilms against antifungal agents in vitro and in vivo
We investigated the molecular basis of the tolerance of Candida albicans biofilms to antifungals using the miconazole as a model compound, and translated the resulting data to other antifungals. Sessile cells of C. albicans Δefg1, lacking the transcription factor Efg1, showed increased susceptibility to miconazole, amphotericin B and caspofungin, whereas these sessile cells were equally resistant to fluconazole. The increased sensitivity to miconazole was, at least, partly due to an increased accumulation of miconazole in the cells as compared to wild-type or reintegrant Δefg1(EFG1) sessile cells. By using a rat biofilm model, we further confirmed the role of Efg1 in the tolerance of C. albicans biofilms to miconazole when grown in vivo.
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- Epidemiology
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Molecular epidemiology of the attachment glycoprotein (G) gene in respiratory syncytial virus in children with acute respiratory infection in Japan in 2009/2010
This study performed a detailed genetic analysis of the glycoprotein (G) gene of respiratory syncytial virus (RSV) detected in 50 Japanese children with acute respiratory infection (ARI) in the 2009/2010 season. A phylogenetic tree constructed by the neighbour-joining method showed that 34 and 16 of the RSV strains could be classified into subgroups A and B, respectively. Strains belonging to subgroups A and B were further subdivided into GA2 and BA, respectively. The nucleotide and deduced amino acid sequence identities were relatively high among these strains (>90 %). The deduced amino acid sequences implied that a relatively high frequency of amino acid substitutions occurred in the C-terminal 3rd hypervariable region of the G protein in these strains. In addition, some positively selected sites were estimated. The results suggest that RSV with genotypes GA2 and BA was associated with ARI in Japanese children in 2009/2010.
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Diarrhoeagenic Escherichia coli as a predominant cause of paediatric nosocomial diarrhoea in India
More LessIntestinal nosocomial infections remain a major concern in paediatric wards leading to increased morbidity and mortality. This study determined the aetiological and epidemiological profile of nosocomial diarrhoea (ND) among children admitted to a hospital in India. During the period of January 2008 to June 2009, we consecutively enrolled 100 children between the age of 2 months and 14 years who developed ND as defined by the Centers for Disease Control and Prevention. A control group of patients matched for age and severity score but with no diarrhoea at admission or during their hospital stay (n = 50) were also enrolled. Stool samples were cultured for various pathogens using standard protocols. Clostridium difficile toxins and rotavirus antigen were detected using commercial ELISAs. Detection of diarrhoeagenic Escherichia coli was carried out by multiplex PCR assay. All patient details were noted. In this study, males predominated (77 %), and 56 % children were <1 year of age and 96 % were <5 years. The mean duration of diarrhoea and hospitalization in the case group was 3.2 days and 27.5 days, respectively. Malignancy and nasogastric tube usage were significant underlying factors for the development of ND. Diarrhoeagenic E. coli was the commonest agent (47 %: enterotoxigenic E. coli, enteroaggregative E. coli and enteropathogenic E. coli were isolated in 22, 18 and 7 % of patients, respectively). C. difficile toxin was seen in 9 % of cases, whilst rotavirus was found in 8 % of cases. Although rotavirus and C. difficile are major causative agents of hospital-acquired diarrhoea in the developed world, in this setting diarrhoeagenic E. coli was responsible for the majority of cases of hospital-acquired diarrhoea. ND was most common in children aged <5 years.
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Wild birds as biological indicators of environmental pollution: antimicrobial resistance patterns of Escherichia coli and enterococci isolated from common buzzards (Buteo buteo)
A total of 36 Escherichia coli and 31 enterococci isolates were recovered from 42 common buzzard faecal samples. The E. coli isolates showed high levels of resistance to streptomycin and tetracycline. The following resistance genes were detected: bla TEM (20 of 22 ampicillin-resistant isolates), tet(A) and/or tet(B) (16 of 27 tetracycline-resistant isolates), aadA1 (eight of 27 streptomycin-resistant isolates), cmlA (three of 15 chloramphenicol-resistant isolates), aac(3)-II with/without aac(3)-IV (all seven gentamicin-resistant isolates) and sul1 and/or sul2 and/or sul3 [all eight sulfamethoxazole/trimethoprim-resistant (SXT) isolates]. intI1 and intI2 genes were detected in four SXT-resistant isolates. The virulence-associated genes fimA (type 1 fimbriae), papC (P fimbriae) and aer (aerobactin) were detected in 61.1, 13.8 and 11.1 % of the isolates, respectively. The isolates belonged to phylogroups A (47.2 %), B1 (8.3 %), B2 (13.9 %) and D (30.5 %). For the enterococci isolates, Enterococcus faecium was the most prevalent species (48.4 %). High levels of tetracycline and erythromycin resistance were found among our isolates (87 and 81 %, respectively). Most of the tetracycline-resistant strains carried the tet(M) and/or tet(L) genes. The erm(B) gene was detected in 80 % of erythromycin-resistant isolates. The vat(D) and/or vat(E) genes were found in nine of the 17 quinupristin–dalfopristin-resistant isolates. The enterococcal isolates showing high-level resistance for kanamycin, gentamicin and streptomycin contained the aph(3′)-IIIa, aac(6′)-aph(2″) and ant(6)-Ia genes, respectively. This report reveals that common buzzards seem to represent an important reservoir, or at least a source, of multi-resistant E. coli and enterococci isolates, and consequently may represent a considerable hazard to human and animal health by transmission of these isolates to waterways and other environmental sources via their faecal deposits.
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- Clinical microbiology and virology
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A comparison of molecular methods for hepatitis B virus (HBV) DNA detection from oral fluid samples
More LessThe objective of the present study was to evaluate four commercial DNA extraction methods and three PCR protocols for hepatitis B virus (HBV) detection in artificially contaminated oral fluid samples. The extraction protocols were selected based on ease of use and cost, and were also compared with respect to sensitivity and cost. Prior PCR optimization was conducted, in which the sample volume for DNA extraction and the concentrations of DNA and Taq DNA polymerase in the PCR were adjusted. One-round PCR, used to amplify the core region of the HBV genome, achieved high levels of sensitivity in comparison with nested and semi-nested PCR experiments that were designed for the amplification of HBV surface protein genes. Of the four extraction protocols evaluated, the RTP DNA/RNA Virus Mini kit and the QIAamp DNA Mini kit gave the highest recovery rates, presenting 20 copies of HBV DNA ml−1 as the limit of detection. These results suggest that HBV DNA can be detected from oral fluid samples but that the optimization of the PCR assays and the choice of extraction methods must be determined by laboratories before the implementation of this method in routine diagnostics.
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- Models of infection
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Shiga toxin-producing Escherichia coli O157 : H7 shows an increased pathogenicity in mice after the passage through the gastrointestinal tract of the same host
Haemolytic uraemic syndrome (HUS) is a rare but life-threatening complication of Shiga toxin (Stx)-producing Escherichia coli (STEC) infections, characterized by acute renal failure, thrombocytopenia and haemolytic anaemia. Although the main infection route is the consumption of contaminated food or water, person-to-person transmission has been suggested in several situations. Moreover, epidemiological data indicate that the horizontal transmission of several pathogens, including STEC, among individuals of the same species requires significantly lower doses than those used in animal models infected with laboratory-cultured bacteria. Thus, the aim of this study was to evaluate whether the passage of a clinically isolated STEC strain through the gastrointestinal tract of mice affects its pathogenicity in mice. To test this, weaned mice were orally inoculated by gavage with either an E. coli O157 : H7 isolate from an HUS patient, or the same strain recovered from stools after one or two successive passages through the gastrointestinal tract of the mice. We show that stool-recovered strains are able to induce a more generalized and persistent colonization than the parent strain. Furthermore, a 104-fold-reduced inoculum of the stool-recovered strains still causes gut colonization and mouse mortality, which are not observed with the parent strain. These results indicate an increased pathogenicity in stool-recovered strains that may be associated with an increased ability to colonize the mouse intestine.
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- Case reports
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Corynebacterium pseudodiphtheriticum septic arthritis secondary to intra-articular injection – a case report and literature review
More LessThis is believed to be the first report of a case of septic arthritis, secondary to intra-articular injection, caused by Corynebacterium pseudodiphtheriticum – a skin commensal micro-organism. Review of the literature highlights the rarity of this pathogen in osteoarticular infections and a potential for delayed diagnosis and inadequate treatment due to subtle initial presentation.
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Abdominal abscess due to NDM-1-producing Klebsiella pneumoniae in Spain
We describe a clinical case of an abdominal abscess due to NDM-1-producing Klebsiella pneumoniae in a 35-year-old Spanish patient after hospitalization in India for perforated appendicitis and peritonitis. The strain belonged to the MLST type 231 and had multiple additional antibiotic resistance genes such as bla CTX-M-15, armA methylase, aac(6′)-Ib-cr, dfrA12, sul1 and qnrB and lack of porin genes ompK35 and ompK36. The patient was cured after abscess drainage.
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Catheter-associated bloodstream infection caused by Leifsonia aquatica in a haemodialysis patient: a case report
Leifsonia aquatica is an aquatic coryneform rod that is capable of forming biofilms in environmental water sources. It has rarely been associated with human infections and its pathogenicity and clinical significance are uncertain. We describe a case of catheter-related bloodstream infection in a haemodialysis patient. The isolate grew on conventional media as a yellow-pigmented colony, but identification required molecular methods. Although the strain displayed reduced sensitivity to vancomycin, the clinical outcome was favourable after catheter removal and intravenous treatment with this antibiotic. Our report gives further evidence of the capability of this aquatic bacterium to cause human infection.
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Prosthetic valve endocarditis caused by Bordetella holmesii, an Acinetobacter lookalike
We report a case of fulminant endocarditis on a prosthetic homograft aortic valve caused by Bordetella holmesii, which was successfully managed by surgical valve replacement and antibiotic treatment. B. holmesii, a strictly aerobic, small, Gram-negative coccobacillus, has been implicated as an infrequent cause of a pertussis-like syndrome and other respiratory illnesses. However, B. holmesii is also a rare cause of septicaemia and infective endocarditis, mostly in immunocompromised patients. To our knowledge, this is the first report of B. holmesii endocarditis on a prosthetic aortic valve. Routine laboratory testing initially misidentified the strain as Acinetobacter sp. Correct identification was achieved by 16S rRNA gene and outer-membrane protein A (ompA) gene sequencing. Interestingly, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry also produced an accurate species-level identification. Subsequent susceptibility testing and review of the literature revealed ceftazidime, cefepime, carbapenems, aminoglycosides, fluoroquinolones, piperacillin/tazobactam, tigecycline and colistin as possible candidates to treat infections caused by B. holmesii.
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- Correspondence
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Volumes and issues
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Volume 73 (2024)
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Volume 72 (2023 - 2024)
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Volume 71 (2022)
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