Qac Genes Research Articles

Resistance pattern and distribution of carbapenemase and antiseptic resistance genes among multidrug-resistant Acinetobacter baumannii isolated from intensive care unit patients.

Nosocomial infections caused by multidrug resistant Acinetobacter baumannii have emerged as a serious problem in healthcare settings worldwide. A total of 100 A. baumannii clinical isolates from immunocompromised patients hospitalized in ICUs in Iran were investigated for antimicrobial susceptibility and the presence of carbapenemase and antiseptic resistance genes. All isolates were resistant to one or more antibiotics, with the most frequent resistance found against ciprofloxacin and imipenem (100 %) and piperacillin (99 %). The MICs of biocides were determined by the agar dilution method. No apparent resistance to biocides was seen among the 100 A. baumannii isolates. All isolates were effectively inhibited by the user's defined concentrations of cetrimide, benzalkonium chloride and glutardaldehyde. The intrinsic β-lactamase gene, blaOXA-51-like, was detected in all A. baumannii isolates. Coexistence of blaOXA-51 andblaOXA-23 was encountered in 89 % of isolates. However, genes blaOXA-58, blaSIM and blaIMP were not detected in any isolates. While A. baumannii isolates were sensitive to biocides, they carried qac genes with the qacEΔ1 gene being the most common, at a frequency of 91 %. Our study revealed the high frequency of multidrug- and carbapenem-resistant isolates of A. baumannii in ICU patients, with a high prevalence of the genes blaOXA-23 and blaOXA-51. However, no apparent biocide resistance was seen in A. baumannii isolates. It appears that appropriate surveillance and control measures are essential to prevent the emergence and transmission of MDR A. baumannii in Iran.

Genetic relatedness of Staphylococcus aureus isolates obtained from cystic fibrosis patients at a tertiary academic hospital in Pretoria, South Africa

Cystic fibrosis (CF) is an inherited recessive disease that affects mucocillary clearance in the lung, allowing it to be colonised with bacteria such as Staphylococcus aureus. To survive in the CF lung S. aureus adapts both phenotypically and genotypically, through various mechanisms. In this study, multiple specimens were collected from the participants and were processed routinely and were additionally cultured in chromogenic media. Multiplex PCR assays were employed to detect methicillin resistance and selected virulence and quaternary ammonium compound (qac) genes. Genetic relatedness of the S. aureus was determined using agr, SCCmec and spa typing as well as pulsed field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). Thirty-three S. aureus isolates were isolated, of which 51% (17/33) were methicillin resistant S. aureus (MRSA). The virulence and qac genes were more prevalent in MRSA than the methicillin sensitive S. aureus (MSSA) isolates. The PFGE analysis showed nine distinct pulsotypes while MLST showed eight sequence types. All the STs detected in this study, except for ST508 have been previously isolated from CF patients according to the literature. This study showed a genetically diverse S. aureus population with a high prevalence of virulence genes among the MRSA isolates from the CF clinic.

The widespread use of topical antimicrobials enriches for resistance in Staphylococcus aureus isolated from patients with atopic dermatitis.

SummaryBackgroundCarriage rates of Staphylococcus aureus on affected skin in atopic dermatitis (AD) are approximately 70%. Increasing disease severity during flares and overall disease severity correlate with increased burden of S. aureus. Treatment in AD therefore often targets S. aureus with topical and systemic antimicrobials.ObjectivesTo determine whether antimicrobial sensitivities and genetic determinants of resistance differed in S. aureus isolates from the skin of children with AD and healthy child nasal carriers.MethodsIn this case–control study, we compared S. aureus isolates from children with AD (n = 50) attending a hospital dermatology department against nasal carriage isolates from children without skin disease (n = 49) attending a hospital emergency department for noninfective conditions. Using whole genome sequencing we generated a phylogenetic framework for the isolates based on variation in the core genome, then compared antimicrobial resistance phenotypes and genotypes between disease groups.Results Staphylococcus aureus from cases and controls had on average similar numbers of phenotypic resistances per isolate. Case isolates differed in their resistance patterns, with fusidic acid resistance (FusR) being significantly more frequent in AD (P = 0·009). The genetic basis of FusR also differentiated the populations, with chromosomal mutations in fusA predominating in AD (P = 0·049). Analysis revealed that FusR evolved multiple times and via multiple mechanism in the population. Carriage of plasmid‐derived qac genes, which have been associated with reduced susceptibility to antiseptics, was eight times more frequent in AD (P = 0·016).ConclusionsThe results suggest that strong selective pressure drives the emergence and maintenance of specific resistances in AD.

Distribution of biocide resistant genes and biocides susceptibility in multidrug-resistant Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii - A first report from the Kingdom of Saudi Arabia.

The aim of this study was to determine the frequency of biocide resistant genes, qacA, qacE and cepA in multidrug resistant (MDR) bacteria: Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii and to correlate the presence or absence of resistant genes with biocides susceptibility. The study included 44 MDR K. pneumoniae, P. aeruginosa and A. baumannii microorganisms. The bacteria were screened for the presence of biocide resistant genes by the polymerase chain reaction (PCR) method. The test organisms were isolated from various clinical specimens in the Qassim region, Saudi Arabia. The in vitro susceptibility tests of the three biocides (benzalkonium chloride, cetrimide and chlorhexidine gluconate) were studied against the test isolates by broth microdilution method following Clinical and Laboratory Standards Institute guidelines. With the distribution of biocide resistant genes in K. pneumoniae, all 9 isolates (100%) possessed cepA; 4 (44.4%) and 1 (11.1%) isolate contained qacA and qacE genes respectively. Among 24 isolates of A. baumannii tested, cepA, qacA and qacE genes were found in 54.2%, 16.7% and 33.3% of isolates respectively. Among 11 P. aeruginosa isolates, 63.6% contained cepA gene, 18.2% contained qacE genes, and none of the isolates harboured qacA gene. There was no significant correlation between presence or absence of biocide resistant genes and high MIC values of the test isolates (p≥0.2). Our observations imply that there was no significant correlation between presence or absence of biocide resistant genes and MICs observed in MDR K. pneumoniae, P. aeruginosa and A. baumannii. Further studies are required to find to confirm the trend of reduced susceptibility to biocides of problematic nosocomial pathogens.

Increased Usage of Antiseptics Is Associated with Reduced Susceptibility in Clinical Isolates of Staphylococcus aureus.

ABSTRACTHospital-acquired infection is a major cause of morbidity and mortality, and regimes to prevent infection are crucial in infection control. These include the decolonization of vulnerable patients with methicillin-resistant Staphylococcus aureus (MRSA) carriage using antiseptics, including chlorhexidine and octenidine. Concern has been raised, however, regarding the possible development of biocide resistance. In this study, we assembled a panel of S. aureus isolates, including isolates collected before the development of chlorhexidine and octenidine and isolates, from a major hospital trust in the United Kingdom during a period when the decolonization regimes were altered. We observed significant increases in the MIC and minimum bactericidal concentration (MBC) of chlorhexidine in isolates from periods of high usage of chlorhexidine. Isolates with increased MICs and MBCs of octenidine rapidly emerged after octenidine was introduced in the trust. There was no apparent cross-resistance between the two biocidal agents. A combination of variable-number tandem repeat (VNTR) analysis, PCR for qac genes, and whole-genome sequencing was used to type isolates and examine possible mechanisms of resistance. There was no expansion of a single strain associated with decreased biocide tolerance, and biocide susceptibility did not correlate with carriage of qac efflux pump genes. Mutations within the NorA or NorB efflux pumps, previously associated with chlorhexidine export, were identified, however, suggesting that this may be an important mechanism of biocide tolerance. We present evidence that isolates are evolving in the face of biocide challenge in patients and that changes in decolonization regimes are reflected in changes in susceptibility of isolates.

Investigating antibiotics, antibiotic resistance genes, and microbial contaminants in groundwater in relation to the proximity of urban areas.

Groundwater is an essential public and drinking water supply and its protection is a goal for global policies. Here, we investigated the presence and prevalence of antibiotic residues, antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), and microbial contamination in groundwater environments at various distances from urban areas. Antibiotic concentrations ranged from below detection limit to 917 ng/L, being trimethoprim, macrolide, and sulfonamide the most abundant antibiotic classes. A total of eleven ARGs (aminoglycoside, β-lactam, chloramphenicol, Macrolide-Lincosamide-Streptogramin B - MLSB, sulfonamide, and tetracycline), one antiseptic resistance gene, and two MGEs were detected by qPCR with relative abundances ranging from 6.61 × 10-7 to 2.30 × 10-1 copies/16S rRNA gene copies. ARGs and MGEs were widespread in the investigated groundwater environments, with increased abundances not only in urban, but also in remote areas. Distinct bacterial community profiles were observed, with a higher prevalence of Betaproteobacteria and Bacteroidetes in the less-impacted areas, and that of Firmicutes in the contaminated groundwater. The combined characteristics of increased species diversity, distinct phylogenetic composition, and the possible presence of fecal and/or pathogenic bacteria could indicate different types of contamination. Significant correlations between ARGs, MGEs and specific taxa within the groundwater bacterial community were identified, revealing the potential hosts of resistance types. Although no universal marker gene could be determined, a co-selection of int1, qacEΔ1 and sulI genes, a proxy group for anthropogenic pollution, with the tetC, tetO, tetW resistance genes was identified. As the tet group was observed to follow the pattern of environmental contamination for the groundwater samples investigated in this study, our results strongly support the proposal of this group of genes as an environmental tracer of human impact. Overall, the present study investigated several emerging contaminants in groundwater habitats that may be included in monitoring programs to enable further regulatory and protection measures.

Qac genes and biocide tolerance in clinical veterinary methicillin-resistant and methicillin-susceptible Staphylococcus aureus and Staphylococcus pseudintermedius

Qac genes are associated with increased tolerance to quaternary ammonium compounds and other cationic biocides such as chlorhexidine. This study aimed to determine whether qac genes and increased biocide tolerance were present in 125 clinical methicillin-resistant and susceptible veterinary staphylococci. A total of 125 methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant and -susceptible Staphylococcus pseudintermedius (MRSP and MSSP) from three archived Australian veterinary staphylococci collections underwent whole genome sequencing, multilocus sequence typing and qac gene screening. Two MRSA isolates (12%) harboured qacA/B genes; both isolates were ST8 from horses. QacJ, qacG and smr genes were identified in 28/90 (31%) MRSP and 1/18 (6%) MSSP isolates. ST71 MRSP was significantly more likely to harbour qac genes than other MRSP clones (p < 0.05). A random subset of 31 isolates underwent minimum bactericidal concentration (MBC) testing against F10SCTM (benzalkonium chloride and biguanide), and HexaconTM (chlorhexidine gluconate), with and without the addition of bovine serum albumin (BSA) as an in vitro substitute for organic matter contamination. Qac genes were not associated with increased phenotypic biocide tolerance but biocide efficacy was significantly affected by the presence of BSA. In the absence of BSA, all MBC values were well below the recommended usage concentration. When BSA was present, regardless of qac gene presence, 50% of MRSA and 43% of MRSP had an F10SCTM MBC above the recommended concentration for general disinfection. Qac genes did not confer increased in vitro biocide tolerance to veterinary staphylococci. Organic matter contamination must be minimized to ensure the efficacy of biocides against MRSA and MRSP.

Effect of nano-silver, nano-copper, deconex and benzalkonium chloride on biofilm formation and expression of transcription regulatory quorum sensing gene (rh1R) in drug-resistance Pseudomonas aeruginosa burn isolates

BackgroundBiofilm forming drug-resistant Pseudomonas aeruginosa are responsible for major death in burn center of different hospitals across the globe. ObjectiveThe aims of this study were to evaluate the effect of nano-silver (Ag), nano-copper (Cu), and two hospital disinfectants (deconex and benzalkonium chloride) on biofilm formation and expression of transcription regulatory quorum sensing gene rh1R in P. aeruginosa burn isolates. Methods28 multidrug-resistant P. aeruginosa (MDRPA) strains were isolated from patients hospitalized in the burn center of a referral hospital in Kerman, Iran. Sizes and purities of nanoparticles were checked by TEM and X-ray diffraction (XRD) analysis. The Minimal Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of the nanoparticles (NPs), deconex and benzalkonium chloride were determined by broth microdilution method. Antibiofilm activities of these compounds were measured by microtiter assay. Polymerase chain reaction (PCR) was used for detection of qacEΔ1, cepA, copA and rhlR genes. Quantification of rhlR gene expression in presence and absence of the above compounds was carried out by relative quantitative real-time PCR (qRT-PCR). ResultsBenzalkonium chloride had a potent antimicrobial activity and inhibited growth of all the isolates at MIC 0.06±0.2mg/mL, while nano-Ag was effective at MIC 20±0.2mg/mL. Furthermore, 28.5% of the isolates showed strong, 25% moderate, 14% weak and 32% demonstrated no biofilm activity. Ag NPs exerted highest antibiofilm activity, follow by deconex and benzalkonium chloride. The qacEΔ1 was absent in this study, whereas 17.8% and 60.8% of the isolates were positive for cepA and copA genes. Benzalkonium chloride, Ag NPs and deconex increased the expression of rhlR gene 64, 2 and 7 folds, respectively. ConclusionOur results suggest that, there is direct relationship between decrease in antibiofilm activity and increase in expression of the rhlR gene in the presence of benzalkonium chloride. Absence of qacEΔ1 gene may be contributed in sensitivity of the isolates to the above agents.

Examination of Quaternary Ammonium Compound Resistance in Proteus mirabilis Isolated from Cooked Meat Products in China.

The aim of this study was to examine the presence of genes responsible for resistance to quaternary ammonium compounds (QACs) and the association of qac genes with class 1 integrons in Proteus mirabilis isolated from cooked meat products. A total of 52 P. mirabilis isolates (29.2%) were detected from 178 samples, and their minimum inhibitory concentrations (MICs) of benzalkonium chloride (BC) ranged from 4 to >32 μg/mL. The isolates with BC MICs of 24 μg/mL were observed most frequently. PCR assays indicated that mdfA, ydgE/ydgF, qacE, qacEΔ1, emrE, sugE(c), and sugE(p) were commonly present (32.7%–100%) in these isolates, but qacH was less prevalent (3.8%). Five groups of resistance gene cassettes were identified in 10 intI1-positive isolates. An unusual gene cassette array dfrA32-ereA-aadA2 was found in one foodborne isolate of P. mirabilis. Two isolates harbored qacH- and sul3- associated non-classic integrons: aadA2-cmlA1-aadA1-qacH-IS440-sul3 and a new arrangement dfrA32-ereA1-aadA2-cmlA1-aadA1-qacH-IS440-sul3, which is first reported in P. mirabilis. Non-classic class 1 integrons were located on conjugative plasmids of 100 kb in two tested isolates. Our data showed that the QAC resistance genes were commonly present among P. mirabilis isolates from cooked meats and qacH was associated with non-classic class 1 integrons. The creation of transconjugants demonstrated that qacH-associated non-classic class 1 integrons were located on conjugative plasmids and therefore could facilitate the co-dissemination of disinfectant and antimicrobial resistance genes among bacteria, an increasing area of concern.

Characterization and Horizontal Transfer of Antimicrobial Resistance Genes and Integrons in Bacteria Isolated from Cooked Meat Products in China

The aim of this study was to investigate antimicrobial resistance and the presence and transferability of corresponding resistance genes and integrons in bacteria isolated from cooked meat samples in the People's Republic of China. A total of 150 isolates (22 species belonging to 15 genera) were isolated from 49 samples. Resistance of these isolates to antimicrobials was commonly observed; 42.7, 36.0, and 25.3% of the isolates were resistant to tetracycline, streptomycin, and ampicillin, respectively. Multidrug resistance was observed in 41 (27.3%) of the isolates. Sixteen resistance genes, i.e., blaTEM-1 and blaCTX-M-14 (β-lactams), aac(3)-IIa (gentamicin), strA and strB (streptomycin), qnrB and qnrS (fluoroquinolone), sul1, sul2, and sul3 (sulfamethoxazole), cat1 and cat2 (chloramphenicol), and tetM, tetA, tetS, and tetB (tetracycline), were found in 54 isolates. One isolate of Pseudomonas putida carried qnrB, and sequence analysis of the PCR product revealed 96% identity to qnrB2. The qnr genes were found coresiding and were cotransferred with bla genes in two isolates. Twelve isolates were positive for the class 1 integrase gene, and four isolates carried the class 2 integrase gene. However, no class 3 integrase gene was detected. One isolate of Proteus mirabilis carried dfrA32-ereA-aadA2, and this unusual array could be transferred to Escherichia coli. Nonclassic class 1 integrons lacking qacEΔ1 and sul1 genes were found in 2 of the 12 intI1-positive isolates. Our results revealed the presence of multidrug-resistant bacteria in cooked meats and the presence and transferability of resistance genes in some isolates, suggesting that cooked meat products may act as reservoirs of drug-resistant bacteria and may facilitate the spread of resistance genes.

Antibiotic Resistance in Salmonella from Retail Foods of Animal Origin and Its Association with Disinfectant and Heavy Metal Resistance.

This study aims to demonstrate the antibiotic resistance and its association with disinfectant and heavy metal resistance in 152 Salmonella isolates recovered from retail foods of animal origins. Susceptibility testing demonstrated that 92.8% isolates were resistant to at least one antibiotic, and the resistance was highest to oxytetracycline (80.9%), followed by trimethoprim (64.5%), amoxicillin (28.9%), ampicillin (28.3%), levofloxacin (21.7%), ciprofloxacin (16.4%), and gentamicin (10.5%), respectively. The blaTEM and tetA genes (44.7%) were commonly present. The qacF and qacEΔ1 genes were detected in 18.4% and 8.6% of all isolates. The Cu-resistance genes pcoR, pcoC, and pcoA were the most prevalent (20.4-40.8%), followed by Hg-resistance gene merA (17.8%) and As-resistance genes arsB (6.6%). The antibiotic resistance was highly associated with disinfectant or certain heavy metal resistance genes. Most notably, the association among Cu-resistance genes (pcoC, pcoR), disinfectant resistance genes (qacF, qacEΔ1), and tetracycline and sulfonamide resistance genes (tet, sul) was significant (p < 0.05). Pulsed-field gel electrophoresis revealed that Salmonella isolates was associated with supermarkets indicating the possibility of crosscontamination in farms or processing environment. This study indicated that retail meats may be a reservoir for the dissemination of antibiotic-resistant Salmonella and using disinfectants for decontamination or metals in livestock may provide a pressure for coselecting strains with acquired resistance to other antimicrobials.

Characterization of Heterogeneous MRSA and MSSA with Reduced Susceptibility to Chlorhexidine in Kuwaiti Hospitals.

The objective of this investigation was to identify the lineages of MRSA and MSSA with reduced susceptibility to chlorhexidine in Kuwaiti hospitals. 121 clinical MRSA and 56 MSSA isolates were included in this study. Antimicrobial susceptibility testing was performed for a selection of agents including chlorhexidine and resistance genes were amplified and sequenced. PFGE, spa typing, and MLST were completed for a selection of isolates. The results showed SCCmec II, III, IV, and V were present in 0.8, 21.5, 69.4, and 8.3% of the MRSA isolates. agr-1Sa was the most prevalent type in both MSSA (48%) and MRSA (54%). Forty-five percentage of MRSA contained pvl and 39% contained lukE-lukD, however, as many as 86% of MSSA contained pvl and 96.4% contained lukE-lukD. qac A-C genes were identified in 12.3% of MRSA, norA was present in 82.6% and blaZ in 94.2%. Among MSSA only 5.4% harbored qacA, 83% contained norA, and 91% blaZ. Multi-drug resistant ST239/t945 lineage containing a qac gene was the most identified S. aureus. However, other lineages, including ST772-MRSA-V/t4867/pvl(+)qacC/smr and non-qac harboring lineages of ST217-MRSAIV/t3244/pvl(–), ST34-MSSA/t161/pvl(+), ST5-MSSA/t688/pvl(+), ST5-MSSA/t4867/norA(+), and ST672-MSSA/t003/pvl(–), also showed reduced susceptibility to chlorhexidine. The observed reduced susceptibility of non-qac dependent MSSA isolates to chlorhexidine suggests the involvement of other elements in promoting higher MBC (≥30 mg/L). Our results confirm that monitoring MSSA is essential as they may have the potential to survive low level biocide exposure.

Characterization and horizontal transfer of qacH-associated class 1 integrons in Escherichia coli isolated from retail meats

The aim of this study was to examine the presence of genes responsible for resistance to quaternary ammonium compounds (QACs) and the association of qac genes with class 1 integrons in Escherichia coli isolated from retail meats. Among the 179 E. coli isolates tested, the minimum inhibitory concentrations (MICs) of benzalkonium chloride (BC) ranged from 4 to 64μg/mL. PCR assays indicated that QAC-resistance genes sugE(c), ydgE/ydgF, mdfA, emrE and qacEΔ1 were commonly present (40.2%–88.3%) in these isolates, but qacE, qacF, qacH and sugE(p) were less prevalent (2.2%–28.5%). Seven different gene cassette arrangements were identified in 31 intI1-positive isolates. Three types of qacH-sul3-associated non-classic integrons were observed in four isolates: dfrA12-orfF-aadA2-cmlA1-aadA1-qacH-IS440-sul3, aadA2-cmlA1-aadA1-qacH-IS440-sul3 and dfrA1-aadA1-qacH-IS440-sul3. Non-classic class 1 integrons were located on plasmids of 100–150kb in these four isolates. Our results demonstrated that the qacH-associated integrons located on 100 kb plasmids in two isolates could be transferred to an E. coli recipient, indicating the co-existence and co-dissemination of disinfectant and antimicrobial resistance genes among bacterial species.

Evolution of class 1 integrons: Mobilization and dispersal via food-borne bacteria.

Class 1 integrons have played a major role in the global dissemination of antibiotic resistance. Reconstructing the history of class 1 integrons might help us control further spread of antibiotic resistance by understanding how human activities influence microbial evolution. Here we describe a class 1 integron that represents an intermediate stage in the evolutionary history of clinical integrons. It was embedded in a series of nested transposons, carried on an IncP plasmid resident in Enterobacter, isolated from the surface of baby spinach leaves. Based on the structure of this integron, we present a modified hypothesis for integron assembly, where the ancestral clinical class 1 integron was captured from a betaproteobacterial chromosome to form a Tn402-like transposon. This transposon then inserted into a plasmid-borne Tn21-like ancestor while in an environmental setting, possibly a bacterium resident in the phyllosphere. We suggest that the qacE gene cassette, conferring resistance to biocides, together with the mercury resistance operon carried by Tn21, provided a selective advantage when this bacterium made its way into the human commensal flora via food. The integron characterized here was located in Tn6007, which along with Tn6008, forms part of the larger Tn6006 transposon, itself inserted into another transposable element to form the Tn21-like transposon, Tn6005. This element has previously been described from the human microbiota, but with a promoter mutation that upregulates integron cassette expression. This element we describe here is from an environmental bacterium, and supports the hypothesis that the ancestral class 1 integron migrated into anthropogenic settings via foodstuffs. Selection pressures brought about by early antimicrobial agents, including mercury, arsenic and disinfectants, promoted its initial fixation, the acquisition of promoter mutations, and subsequent dissemination into various species and pathogens.

High Prevalence of blaNDM-1, blaVIM, qacE, and qacEΔ1 Genes and Their Association with Decreased Susceptibility to Antibiotics and Common Hospital Biocides in Clinical Isolates of Acinetobacter baumannii.

The objective of this study was to evaluate the susceptibility of metallo-β-lactamase (MBL)-producing Acinetobacter baumannii (A. baumannii) clinical isolates to biocides. We also determined the prevalence and correlation of efflux pump genes, class 1 integron and MBL encoding genes. In addition, blaVIM, blaNDM-1, qacE and qacEΔ1 nucleotide sequence analysis was performed and compared to sequences retrieved from GenBank at the National Center for Biotechnology Information database. A. baumannii had a resistance rate to carbapenem of 71.4% and 39.3% and was found to be a MBL producer. The minimum inhibitory concentrations (MICs) of chlorhexidine and cetrimide were higher than the recommended concentrations for disinfection in 54.5% and 77.3% of MBL-positive isolates respectively and their MICs were significantly higher among qac gene-positive isolates. Coexistence of qac genes was detected in 68.1% and 50% of the isolates with blaVIM and blaNDM-1 respectively. There was a significant correlation between the presence of qac genes and MBL-encoding blaVIM and blaNDM-1 genes. Each of the blaNDM-1, blaVIM, qacE and qacEΔ1 DNA sequences showed homology with each other and with similar sequences reported from other countries. The high incidence of Verona integron-encoded metallo-β-lactamases (VIM) and New-Delhi-metallo-β-lactamase (NDM) and qac genes in A. baumannii highlights emerging therapeutic challenges for being readily transferable between clinically relevant bacteria. In addition reduced susceptibility to chlorhexidine and cetrimide and the potential for cross resistance to some antibiotics necessitates the urgent need for healthcare facilities to periodically evaluate biocides efficacy, to address the issue of antiseptic resistance and to initiate a “biocidal stewardship”.

Culture-independent detection of chlorhexidine resistance genes qacA/B and smr in bacterial DNA recovered from body sites treated with chlorhexidine-containing dressings.

Dressings containing chlorhexidine gluconate (CHG) are increasingly used in clinical environments for prevention of infection at central venous catheter insertion sites. Increased tolerance to this biocide in staphylococci is primarily associated with the presence of qacA/B and smr genes. We used a culture-independent method to assess the prevalence of these genes in 78 DNA specimens recovered from the skin of 43 patients at catheter insertion sites in the arm that were covered with CHG dressings. Of the 78 DNA specimens analysed, 52 (67 %) possessed qacA/B and 14 (18 %) possessed smr; all samples positive for smr were also positive for qacA/B. These prevalence rates were not statistically greater than those observed in a subsample of specimens taken from non-CHG treated contralateral arms and non-CHG-dressing exposed arms. A statistically greater proportion of specimens with greater than 72 h exposure to CHG dressings were qac-positive (P=0.04), suggesting that the patients were contaminated with bacteria or DNA containing qacA/B during their hospital stay. The presence of qac genes was not positively associated with the presence of DNA specific for Staphylococcusepidermidis and Staphylococcusaureus in these specimens. Our results show that CHG genes are highly prevalent on hospital patients' skin, even in the absence of viable bacteria.

Characterization of Staphylococcus epidermidis strains isolated from industrial cleanrooms under regular routine disinfection.

The purpose of this study was the genotypic and phenotypic characterization of 57 strains of Staphylococcus epidermidis isolated from cleanroom environments, based on their biofilm formation and antimicrobial resistance profiles. Biofilm formation was investigated using real-time PCR (icaA, aap, bhp genes), the Congo red agar method and the crystal violet assay. The majority of the strains (59·7%; 34/57) did not form biofilms according to the crystal violet assay, although the biofilm-associated genes were present in 94·7% (54/57) of the strains. Of the biofilm formers (40·4%; 23/57), 39·1% (9/23) have been identified as strong biofilm formers (>4× crystal violet absorbance cut-off). Resistance to a commercial disinfectant and its quaternary ammonium active component, didecyl-dimethyl-ammonium chloride (DDAC), was determined according to minimum inhibitory concentrations (MICs) and the presence of the qac (quaternary ammonium compound) genes. More than 95% (55/57) of the Staph. epidermidis strains had the qacA/B and qacC genes, but not the other qac genes. The MICs for the disinfectant and DDAC varied among the Staph. epidermidis strains, although none were resistant. Although 59·6% of the Staph. epidermidis strains did not form biofilms and none were resistant to DDAC, more than 94% had the genetic basis for development of resistance to quaternary ammonium compounds, and among them at least 14·0% (8/57) might represent a high risk to cleanroom hygiene as strong biofim formers with qacA/B and qacC genes. To assure controlled cleanroom environments, bacterial strains isolated from cleanroom environments need to be characterized regularly using several investigative methods.

Abundance of antibiotics, antibiotic resistance genes and bacterial community composition in wastewater effluents from different Romanian hospitals

Antimicrobial resistance represents a growing and significant public health threat, which requires a global response to develop effective strategies and mitigate the emergence and spread of this phenomenon in clinical and environmental settings. We investigated, therefore, the occurrence and abundance of several antibiotics and antibiotic resistance genes (ARGs), as well as bacterial community composition in wastewater effluents from different hospitals located in the Cluj County, Romania. Antibiotic concentrations ranged between 3.67 and 53.05 μg L−1, and the most abundant antibiotic classes were β-lactams, glycopeptides, and trimethoprim. Among the ARGs detected, 14 genes confer resistance to β-lactams, aminoglycosides, chloramphenicol, macrolide-lincosamide-streptogramin B (MLSB) antibiotics, sulfonamides, and tetracyclines. Genes encoding quaternary ammonium resistance and a transposon-related element were also detected. The sulI and qacEΔ1 genes, which confer resistance to sulfonamides and quaternary ammonium, had the highest relative abundance with values ranging from 5.33 × 10−2 to 1.94 × 10−1 and 1.94 × 10−2 to 4.89 × 10−2 copies/16 rRNA gene copies, respectively. The dominant phyla detected in the hospital wastewater samples were Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria. Among selected hospitals, one of them applied an activated sludge and chlorine disinfection process before releasing the effluent to the municipal collector. This conventional wastewater treatment showed moderate removal efficiency of the studied pollutants, with a 55–81% decrease in antibiotic concentrations, 1–3 order of magnitude lower relative abundance of ARGs, but with a slight increase of some potentially pathogenic bacteria. Given this, hospital wastewaters (raw or treated) may contribute to the spread of these emerging pollutants in the receiving environments. To the best of our knowledge, this study quantified for the first time the abundance of antibiotics and ARGs in wastewater effluents from different Romanian hospitals.

Integron occurrence is linked to reduced biocide susceptibility in multidrug resistant Pseudomonas aeruginosa

Objective: Integrons are gene acquisition systems commonly found in bacterial genomes that play a major role in the dissemination of resistance to antibiotics. This work aimed to study the relationship between the presence of integrons and the reduced susceptibility of multidrug-resistant (MDR) Pseudomonas aeruginosa isolates towards different groups of biocides.Methods: The antimicrobial susceptibility patterns of 104 clinical isolates were determined against different antibiotics by the disk diffusion method. The isolates were also tested for their susceptibility to six biocides (glutaraldehyde, benzalkonium chloride, cetrimide, chlorhexidine gluconate, chlorocresol and gluconate, and phenyl mercuric nitrate) by agar dilution. The presence of integrons and resistance genes in MDR isolates were detected by polymerase chain reaction.Results: Thirty-six Pseudomonas isolates were MDR, and the majority of these isolates showed reduced susceptibility to biocides. In the MDR isolates, Class I integron was detected in 22 isolates (61.1%), while Class II and III integrons were identified in only four isolates (11.1%), In addition, aacA4 and qacE genes were detected in 22 (61.1%) and 11 (30.5%) isolates, respectively. Integron I-positive isolates showed reduced susceptibility to tested biocides.Conclusions: The current study reveals the presence of different classes of integrons, with class I being predominant. Class I integron may be responsible for generating MDR P. aeruginosa isolates with reduced susceptibility to biocides. This linkage between integrons and biocide resistance in MDR-Pseudomonas isolates is notable and could be clinically important. Strict antibiotic prescription policies and the adequate use of biocides could help in controlling this problem.

Prevalence of eaeA and qacEΔ1 genes in Escherichia coli isolated from omphalitis in baby chicks

A total of two hundred diseased Saso chicks with omphalitis were examined (1400 samples) for the isolation of E. colifrom different organs (liver, caecum, spleen, heart, lung, yolk sac and cloacal swab). Results showed that 64 cases werepositive with an incidence of 32%. Fifty isolates (25%) of E. coli were recovered from chicks could be sero-grouped in19 O groups with the most predominant serotype was O91 20 % (10 out of 50 isolates), O26 10% (5 strains out of 50) O788% (4 strains out of 50), and 7 strains untypable E. coli 14%. Among E. Coli O groups were found to be resistant toAmoxicillin 93.02% followed by Tetracycline 74.42%, Enrofloxacin 46.51%, Erythromycin 30.32%, Ciprofloxacin27.91%, Norofloxacin and Streptomycin 20.93% and Gentamycin 6.98%. While they were found to be sensitive forGentamycin, Streptomycin, Ciprofloxacin, Tetracycline, Erythromycin, Norofloxacin, Enrofloxacin, and Amoxicillin asthe following: 88.37%, 46. 51%, 34.89%, 23.26%, 18.60%, 9.30%, 9.30% and 6.98%, respectively. The incidence rateof eaeA gene of E. coli was 15.79%. Quaternnary ammonium compound (qacEΔ1) gene also was detected in E. coli withincidence rate 63.16%.