Qac Genes Research Articles

A TaqMan real-time PCR assay for detection of qacEΔ1 gene in Gram-negative bacteria

The transfer of biocide and antibiotic resistance genes by mobile genetic elements is the most common mechanism for rapidly acquiring and spreading resistance among bacteria. The qacEΔ1 gene confers the resistance to quaternary ammonium compounds (QACs). It has also been considered a genetic marker for the presence of class 1 integrons associated with multidrug-resistant (MDR) phenotypes in Gram-negative bacteria. In this study, a TaqMan real-time PCR assay was developed to detect the qacEΔ1 gene in Gram-negative bacteria. The assay has a detection limit of 80 copies of the qacEΔ1 gene per reaction. No false-positive or false-negative results have been observed. Simultaneous amplification and detection of the 16S rRNA gene is performed as an endogenous internal amplification control (IAC). The TaqMan real-time PCR assay developed is a rapid, sensitive, and specific method that could be used to monitor resistance to QACs, the spread of class 1 integrons, and the prediction of associated MDR phenotypes in Gram-negative bacteria.

Evaluation of Biocides Susceptibility and Expression of Qace-Δ1 And Suge1 Genes of Pseudomonas Aeruginosa in Exposure to Biocides and Meropenem

Pseudomonas aeruginosa is a species of medical significance that plays a role in nosocomial infections, particularly those resulting from burns and wounds. Bacterial antibiotic resistance may have developed as a result of the abuse of biocides in hospitals. This study aimed to determine the minimum inhibitory concentrations (MICs) of carbapenem-resistant P. aeruginosa isolates in relation to ethylenediaminetetraacetic acid (EDTA), chlorohexidine (CHX), and resazurin dye using a microtiter plate assay. Additionally, it evaluated the expression of the efflux pump genes (qacE-Δ1 and sugE1) at sub-inhibitory concentrations of the biocides using RT-qPCR (delta delta Ct method) to improve their efflux pump system. The MICs of 24 P. aeruginosa isolates (14 isolates were resist to carbapenems and 10 isolates sensitive to carbapenems) against Chloroxylenol (Dettol) revealed that the range of MICs were (0. 125-2%). The results of MICs of Chlorohexidine (CHX) revealed that the range of inhibitory activity was (0.8-102.4 µg/ml), also the findings of MICs of EDTA demonstrated that the range of inhibitory activity was (125-2000 µg/ml). The genes involved in efflux pump (qacE -Δ1 and sugE1) were found to be significantly up-regulated in the local isolates, according to our findings.. The four isolates' sugE1 gene expression folds ranged from 1.81 to 5.39; this result was more than the fold of the control (the fold change is 1), with the meropenem isolate having the highest fold value (5.39) and sub-MIC value (8 µg/ml). Furthermore, the four isolates' qacE-Δ1 gene expression folds ranged from 2.11 to 5.01, which was higher than the fold of the control, where the highest fold value was recorded for the EDTA (5.01) with the sub-MIC value (125-500 µg/ml). Also, a high value of upregulation of the gene qacE –Δ1, during the treatment with dettol with sub-MIC (0.25-0.5%), and the fold change was 4.19. In conclusion, the present findings indicated to the significant relationship between the treatments with meropenem and the high values of gene expression of the gene sugE1 in comparison with the control and other agents, Furthermore, compared to the antibiotic and control, there was a strong correlation found between the high fold changes of the gene qacE -Δ1 with the presence of dettol and EDTA.

Prevalence of methicillin-resistant Staphylococcus aureus (MRSA) harboring mupirocin and biocide resistance genes in a large health care system

We aimed to determine the prevalence of genes associated with high-level mupirocin and biocide resistance in methicillin-resistant Staphylococcus aureus (MRSA) isolates among hospitalized patients and to characterize their genomic and epidemiologic features. Study conducted on an integrated health system. Clinical cultures with MRSA from hospitalized patients collected between March 1, 2023, and January 20, 2024 underwent prospective whole-genome sequencing (WGS), including assessment for the presence of markers of resistance against mupirocin (mupA) and biocides (qac). Demographic and clinical characteristics were reviewed. We analyzed 463 MRSA isolates. The overall prevalence of mupA(+), qacA(+), and qacC(+) genes was 22.0%, 2.4%, and 19.0%, respectively. Most mupA(+) isolates belonged to ST8, but ST8732 (a novel variant of ST8) had the highest prevalence of mupA(+) isolates at 95%. Patients mupA(+) were older, and none of the isolates from pediatric patients harbored this gene. Through prospective WGS of MRSA isolates we detected a prevalence of genes conferring mupirocin resistance considerably higher than previously reported, particularly among MRSA ST8 variants. Our findings highlight the need for monitoring resistance to agents used for the prevention of Staphylococcus aureus infections, as these trends have implications for infection prevention programs and public health at large.

Complex Infection-Control Measures with Disinfectant Switch Help the Successful Early Control of Carbapenem-Resistant Acinetobacter baumannii Outbreak in Intensive Care Unit.

A carbapenem-resistant Acinetobacter baumannii (CRAB) outbreak in an intensive care unit (ICU) was contained by an improved infection-control measure that included a disinfectant policy. In our retrospective cohort study, we describe the epidemiological investigations and infection-control measures during this outbreak. Descriptive analysis was used to summarize patient demographics, neurological diseases, surgical treatment, underlying diseases, infection, and outcomes. In December 2023, two CARB-positive patients were observed in the ICU, and four more patients became CRAB-positive in January. During this outbreak, there was an overlap of hospitalization periods among the CRAB-positive patients, and CRAB was isolated from the environment; the isolated CRAB strain was identical. Infection-control measures, including hand hygiene, contact precautions and isolation, surveillance, decolonization, environmental cleaning, and disinfection, were reviewed and modified. The aim of this study was to examine the molecular background of the effectiveness of the disinfectant shift used during successful outbreak control. Experiments were carried out to study the phenotypic sensitivity and genetic background of different disinfectant agents. A thorough analysis of the detected CRAB strain included whole-genome sequencing (WGS), investigation of the qacE and qacEΔ1 genes' relative expression by qPCR after exposure to different disinfectant solutions, as well as an analysis of biofilm formation. WGS analysis of the CRAB strain identified that an ST2 high-risk clone was responsible for the outbreak, which produced OXA-83 and ADC-30 beta-lactamases; in addition, qacE and qacEΔ1 genes were also detected, which confer resistance to disinfectants containing quaternary ammonium compounds (QACs). A qPCR analysis demonstrated that after exposure to different disinfectants, the gene expression levels of qacE and qacEΔ1 increased and correlated with concentrations of QACs of disinfectants. During the outbreak, the standard-of-care QAC-based disinfectant was changed to a mainly alcohol-based agent in the ICU, which contributed to the successful control of this outbreak, and no additional patients were identified with CRAB. We conclude that continuous surveillance and hand hygiene training combined with fast identification and reaction to new cases, as well as an in-depth analysis of multidrug-resistant outbreak strains and investigation of their disinfectant tolerance/resistance during an outbreak, are essential to effectively control the spread of nosocomial pathogens. The smart policy of disinfectant agent selection played a crucial role in controlling the outbreak and ensuring patient safety in the ICU.

Prevalence of qacEΔ1, qacE, oqxA, oqxB, acrA, cepA and zitB genes among multidrug-resistant Klebsiella pneumoniae isolated in a cardiac hospital

Background. Infections caused by multidrug-resistant (MDR) Klebsiella pneumoniae are the leading cause of mortality worldwide. The widespread use of disinfectants and antiseptics has caused the emergence of K. pneumoniae with reduced sensitivity to them, which, in combination with MDR, can pose a significant epidemiological threat. The aim of the study was to assess the prevalence of efflux pump and transporter genes associated with biocide resistance and their association with antibiotic resistance among K. pneumoniae isolated in a cardiac surgical hospital. Materials and methods. K. pneumoniae isolates (n = 50) from the patients and medical equipment were tested by polymerase chain reaction for the presence of genes of 4 types of efflux pumps (qacEΔ1, qacE, oqxA, oqxB, acrA) and 2 transporters involved in the outflow of cations (cepA) and zinc ions (zitB). Spearman's rank correlation test was used to assess the strength of the association between the efflux pumps, beta-lactamase genes and mobile genetic elements. Results. The occurrence of K. pneumoniae containing qacEΔ1, qacE, oqxA, oqxB, acrA, cepA and zitB was high: 54, 62, 100, 84, 100, 72 и 96% respectively. K. pneumoniae with a combination of all the studied pumps was most often detected (32%), and these isolates were MDR in 100% of cases. The qacE, qacEΔ1 genes were closely associated with resistance to cephalosporins, carbapenems, fluoroquinolones, carbapenemase genes, and integrons. The results of the study showed that the genes of various efflux pumps associated with biocide resistance and their combinations were widely represented among the clinical isolates of MDR K. pneumoniae. Conclusion. The high prevalence of efflux pump genes associated with resistance to quaternary ammonium compounds, chlorhexidine and zinc salts and their significant association with antibiotic resistance in nosocomial K. pneumoniae underlines the importance of further studying the mechanisms of cross-resistance to biocides to improve methods of combating MDR nosocomial pathogens.

Phenotypic and genotypic determination of resistance to common disinfectants among strains of Acinetobacter baumannii producing and non-producing biofilm isolated from Iran

BackgroundNosocomial infections are a global problem in hospitals all around the world. It is considered a major health problem, especially in developing countries. The increase in the patient’s stay in hospitals has increased the mortality rate, and consequently, the costs drastically increase. The main purpose of using disinfectants in the hospital environment is to reduce the risk of nosocomial infections. Ethylene diamine tetra acetic acid (EDTA) causes lysis and increases susceptibility to antimicrobial agents in the planktonic form of bacteria. This substance affects the permeability of the outer membrane of bacteria. It also prevents the formation of biofilms by bacteria.Materials and methodsIn the current study, 120 isolates of Acinetobacter baumannii (A. baumannii) were confirmed by phenotypic and genotypic methods. Antibiogram was performed and then the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of isolates against 5% sodium hypochlorite, ethanol %70, sayasept-HP 2%, chlorhexidine 2%, dettol 4/8% were evaluated. In addition, the disinfectant effect was re-evaluated with the mixture of EDTA solution. All isolates were examined for biofilm presence by crystal violet staining method in triplicates and repeated three times for each strain. Also for all isolates detection of efflux pump genes (Qac-E, qacE-Δ1, SUG-E) by PCR technique was done.ResultsAntibiogram results of A. baumannii showed that 6.7% were Multi-drug-resistant (MDR), and 89.2% were Extensively drug-resistant (XDR) isolates. The highest effect of disinfectants was related to 5% sodium hypochlorite, and the least effect was 70% ethanol. EDTA increases the efficacy of selected disinfectants significantly. The highest prevalence of the efflux pump genes was related to SUG-E (95%) and Qac-E (91.7%), and, the qacE-Δ1 gene with 12.5%. The biofilm production rate was 91.3% among all isolates.ConclusionThe best and safest way to disinfect hospital floors and surfaces is to choose the right disinfectants, and learn how to use them properly. In this study, a mixture of disinfectants and EDTA had a significant effect on bactericidal activity. it was found that improper use of disinfectants, especially the use of sub-inhibitory dilutions, increases the resistance of bacteria to disinfectants.

Increasing Fluroquinolone Susceptibility and Genetic Diversity of ESBL-Producing E. coli from the Lower Respiratory Tract during the COVID-19 Pandemic.

Lower respiratory tract infections (LRTIs) are the fourth leading cause of death worldwide, among which Escherichia coli (E. coli) pneumonia is considered a rare phenomenon. Treatment options for LRTIs have become limited, especially for extended-spectrum β-lactamase-producing E. coli (ESBL-EC), which are usually resistant to other groups of antimicrobials as well. The aim of our study was to compare the phenotypic resistance profiles and genotypes of ESBL-EC isolates associated with LRTIs before (pre-COVID-19) and during (COVID-19) the COVID-19 pandemic. All isolates were screened for antimicrobial resistance genes (ARGs) and virulence-associated genes (VAGs) and assigned to phylogenetic groups, sequence types and clonal groups by PCR. During the pandemic, a significantly lower proportion of ciprofloxacin-, levofloxacin- and trimethoprim-sulfamethoxazole-resistant ESBL-EC isolates was retrieved from lower respiratory tract (LRT) samples. PCR-based genotypization revealed greater clonal diversity and a significantly lower proportion of isolates with blaTEM, aac(6')-Ib-cr and qacEΔ1 genes. In addition, a higher proportion of isolates with the integrase gene int1 and virulence genes sat and tsh was confirmed. The lower prevalence of fluoroquinolone resistance and greater genetic diversity of ESBL-EC isolated during the COVID-19 period may have been due to the introduction of new bacterial strains into the hospital environment, along with changes in clinical establishment guidelines and practices.

Prevalence of Antibiotic Resistance Genes in Bacterial Isolates in Iran: A Systematic Review

Context: The widespread use of biocides containing quaternary ammonium compounds (QACs) in medical centers has led to resistant strains of microorganisms. This study aimed to evaluate the frequency of genes resistant to antiseptics in Iranian bacterial isolates. Methods: The present study was conducted as a systematic review. The keywords in the titles or abstracts of articles published in Persian and English were monitored. The search was performed in databases without a time limit. Results: The results showed a total of 975 bacterial isolates. The most common disinfectant-resistant genes in gram-positive isolates were QacAB genes, with an average of 28.48; SMR gene, with an average of 19.8; and qacE and qacEΔ1 genes, with an average of 45 in gram-negative isolates. Gram-negative isolates containing qacE and qacEΔ1 genes were mainly detected in patients hospitalized in intensive care, internal diseases, and infectious diseases wards and from trachea and urine samples. Conclusions: The unique conditions of the intensive care units and the use of invasive tools such as urinary catheters were the leading causes of infection with resistant pathogens. In addition, diversity in infection control measures like the type, amount, and concentration of disinfectants used in different hospital departments were other reasons for resistance to antiseptics in medical environments. Indiscriminate use of antibiotics caused microbial resistance in the same way antibiotics did not affect many microbial diseases, and arbitrary use caused the highest microbial resistance.

Assessing chlorhexidine resistance in MRSA isolates from hospitals in Cleveland, OH and Detroit, MI

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most common causes of procedure-related, skin, and soft-tissue infections. Hospitalized patients who are colonized with MRSA are at a higher risk of developing invasive infections after discharge. Chlorhexidine, an antiseptic/disinfectant, has been used to reduce carriage and prevent infections in these patients. Studies have shown chlorhexidine resistance among MRSA strains. Chlorhexidine resistance is associated with qac genes, which encode multidrug efflux pumps that increase bacterial tolerance to disinfectant agents. The global distribution and prevalence of qacA and qacB genes are highly variable. One study reported that qacA and qacB genes could be found in 0.9% - 83.3% of clinical MRSA isolates worldwide. The goal of this study was to determine the prevalence of chlorhexidine resistance and identify the resistance-associated genes from our MRSA samples using whole genome sequencing (WGS). Methods: 474 MRSA samples were obtained from hospitals in Detroit, MI (287) and Cleveland, OH (187). Whole genome sequencing was performed using the NextSeq (Illumina Inc., CA) platform. The sequencing data was analyzed using ResFinder 4.1, a publicly available database that can be used to identify acquired genes and chromosomal mutations mediating antimicrobial resistance. The output was organized into a data sheet to visualize the presence of the genes of interest. Results: The qacA gene was present in only one MRSA sample from the Cleveland area hospital. In the samples from Detroit, 14 out of 287 showed disinfectant resistance genes. The qacA, qacB, and qacD were present in 1, 6, and 7 samples, respectively. The prevalence of any qac gene in the Cleveland area samples was 0.5%. Meanwhile, the prevalence of any qac gene in Detroit area samples was 4.9%. Among the 7 samples that have qacD gene, 6 samples have more than one copy of qacD. Conclusions: The prevalence of the “qac” gene varied widely based on the origin of the samples. Detroit area samples had more qac genes prevalence than Cleveland area samples. Chlorhexidine is a widely used antiseptic/disinfectant, and it plays a vital role in reducing carriage and preventing infection among hospitalized patients colonized with MRSA. Monitoring and addressing MRSA-reduced susceptibility to chlorhexidine is imperative for maintaining the effectiveness of infection control practices such as decolonization.

Association analysis of antibiotic and disinfectant resistome in human and foodborne E. coli in Beijing, China

The widespread use of chemical disinfectants and antibiotics poses a major threat to food safety and human health. However, the mechanisms of co-transmission of antimicrobial resistance genes (ARGs) and biocides and metal resistance genes (BMRGs) of foodborne pathogens in the food chain is still unclear. This study isolated 343 E. coli strains from animal-derived foods in Beijing and incorporated online data of human-derived E. coli strains from NCBI. Our results demonstrated a relatively uniform distribution of strains from various regions in Beijing, indicating a lack of region-specific clustering. Additionally, predominant sequence types varied between food- and human-derived strains, suggesting a preference for different hosts and environments. Phenotypic association analysis showed that the chlorine disinfectants peroxides had a significant positive correlation with tetracyclines. Many more ARGs and BMRGs were enriched in human-associated E. coli compared with those in chicken- and pork-origin. The quaternary ammonium compounds (QACs) resistance gene qacEΔ1 had a strong correlation with aminoglycoside resistance gene aadA5, folate pathway antagonist resistance gene dfrA17, sul1 and macrolide resistance gene mph(A). The correlation results indicated a significant association between the copper resistance gene cluster pco and the silver resistance gene cluster sil. Coexistence of many resistance genes was observed within the qacEΔ1 gene structure, with qacEΔ1-sul1 being the most common combination. Our findings demonstrated that the epidemiological spread of resistance is affected by a combination of heavy metals, disinfectants and antibiotic use, suggesting that the prevention and control strategies of antimicrobial resistance need to be multifaceted and comprehensive.

Recombination-mediated dissemination of Methicillin-resistant S. aureus clonal complex 1 in the Egyptian health care settings

BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) is a rapidly evolving pathogen that is frequently associated with outbreaks and sustained epidemics. This study investigated the population structure, resistome, virulome, and the correlation between antimicrobial resistance determinants with phenotypic resistance profiles of 36 representative hospital-acquired MRSA isolates recovered from hospital settings in Egypt.ResultsThe community-acquired MRSA lineage, clonal complex 1 (CC1) was the most frequently detected clone, followed by three other globally disseminated clones, CC121, CC8, and CC22. Most isolates carried SCCmec type V and more than half of isolates demonstrated multi-drug resistant phenotypes. Resistance to linezolid, a last resort antibiotic for treating multidrug resistant MRSA, was observed in 11.11% of the isolates belonging to different genetic backgrounds. Virulome analysis indicated that most isolates harboured a large pool of virulence factors and toxins. Genes encoding aureolysin, gamma hemolysins, and serine proteases were the most frequently detected virulence encoding genes. CC1 was observed to have a high pool of AMR resistance determinants including cfr, qacA, and qacB genes, which are involved in linezolid and quaternary ammonium compounds resistance, as well as high content of virulence-related genes, including both of the PVL toxin genes. Molecular clock analysis revealed that CC1 had the greatest frequency of recombination (compared to mutation) among the four major clones, supporting the role of horizontal gene transfer in modulating AMR and hypervirulence in this clone.ConclusionsThis pilot study provided evidence on the dissemination success of CA-MRSA clone CC1 among Egyptian hospitals. Co-detection of multiple AMR and virulence genes in this lineage pose a broad public health risk, with implications for successful treatment. The results of this study, together with other surveillance studies in Egypt, should be used to develop strategies for controlling MRSA infections in Egyptian health-care settings.

Characterization of disinfectant susceptibility profiles among clinical isolates of Acinetobacter baumannii in Ardabil, Iran

AbstractAntimicrobial disinfectants have been extensively used to control hospital-acquired infections worldwide. Prolonged exposure to bacteria could promote resistance to antimicrobial disinfectants. This study evaluated the antimicrobial activity of four commonly used disinfectants; triclosan, chlorhexidine digluconate, benzalkonium chloride, and formaldehyde against Acinetobacter baumannii clinical isolates. This study also determined the prevalence and association of efflux pumps encoding genes qacE, qacED1, emrA, and aceI with tolerance to disinfectants. A total of 100 A. baumannii isolates were included in the current study. The antimicrobial disinfectants' minimum inhibitory concentration (MIC) was determined using an agar dilution method. Genes involved in resistance to disinfectants were investigated by PCR method. The benzalkonium chloride MICs ranged between 32 and 128 μg mL−1, chlorhexidine digluconate 8–64 μg mL−1, triclosan 1–32 μg mL−1, and formaldehyde 128 μg mL−1. Overall, the highest MIC90 value was identified for formaldehyde (128 μg mL−1), followed by benzalkonium chloride and chlorhexidine digluconate (64 μg mL−1, each one) and triclosan (4 μg mL−1). In the present study, the qacE, qacED1, emrA, and aceI genes were found in 91%, 55%, 100%, and 88% of isolates, respectively. The qacG gene was not identified in our A. baumannii isolates. The qacED1 gene was associated with higher MICs for all disinfectants tested (P < 0.05), while the qacE and aceI genes were associated with higher MICs for benzalkonium chloride and chlorhexidine. This study indicated that triclosan is the most effective disinfectant against A. baumannii isolates.

713. Prevalence of Microbicide Resistance Genes in Clinical Isolates of Acinetobacter baumannii

Abstract Background The persistence of Acinetobacter on different surfaces and the continuous exposure of bacteria to disinfectants such as quaternary ammonium compounds (QACs) in the healthcare environment have led to microbicide tolerance. Acinetobacter baumannii have also rapidly acquired carbapenem resistance. Here we determined the prevalence of microbicide resistance genes in carbapenem susceptible and resistant clinical isolates of Acinetobacter. Methods All clinical isolates were collected from two separate hospitals in Detroit between 2017-2021. Following DNA isolation using Qiagen kit, Nextera Flex kit was used for library preparation of Acinetobacter clinical isolates. Whole genome sequencing (WGS) was performed via Illumina NextSeq 550 using the prepared DNA library. The FASTQ files from the sequencing run were then subjected to bioinformatic analysis with Bionumerics software v7.6. ResFinder was used for determination of presence or absence of QAC resistant genes and results were tabulated by Carbapenem-Resistant Acinetobacter baumannii (CRAb) or Carbapenem-Susceptible Acinetobacter baumannii (CSAb) with or without the QAC resistant genes. Results Out of the 139 Acinetobacter patient isolates, 70/139 (50.4%) were CRAb containing carbapenemase blaOXA-23 and 69/139 (49.6%) belonged to CSAb (Table 1). Figure 1. shows the prevalence of antimicrobial resistance (AMR) genes in CRAb and CSAb isolates with qacE genes. For CRAb isolates, 56% had qacE genes while 39% of CSAb had qacE genes. About 42% of CRAb isolates were multi-drug resistant (MDR), indicating a prevalence of qacE genes among the MDR isolates. Table 2 shows the presence of qacE genes by sequence type (ST). In the ST2 (Pasteur) and ST406 (Pasteur) strains, qacE gene was mainly present. Interestingly, most of the Acinetobacter isolates that belong to ST2 (ST195 and ST281, Oxford) are CRAb with qacE gene.Figure 1:Different AMR genes and qacE in both CRAb and CSAb isolates Conclusion Clinical isolates in our study had an even distribution of both CRAb and CSAb. The presence of qacE gene was more prevalent in the CRAb isolates compared to the CSAb isolates. While different STs harbored the qacE gene, the prevalence of qacE gene was highest in ST2 among all isolates. Future studies are needed to determine the reduced susceptibility to other commonly used hospital disinfectants. Disclosures Keith S. Kaye, MD, MPH, Abbvie: Advisor/Consultant|Abbvie: Honoraria|Entasis: Advisor/Consultant|Entasis: Honoraria|GSK: Advisor/Consultant|GSK: Honoraria|Merck: Advisor/Consultant|Merck: Honoraria|Shionogi: Advisor/Consultant|Shionogi: Honoraria|VenatoRx: Advisor/Consultant|VenatoRx: Honoraria

Impact of clonal lineages on susceptibility of Staphylococcus lugdunensis to chlorhexidine digluconate and chloride benzalkonium.

Little is known about susceptibility of Staphylococcus lugdunensis to antiseptics. The objective of this study was to evaluate, at the molecular and phenotypic level, the susceptibility of 49 clinical S. lugdunensis strains (belonging to the seven clonal complexes [CCs] defined by multilocus sequence typing) to two antiseptics frequently used in healthcare settings (chlorhexidine digluconate [CHX] and chloride benzalkonium [BAC]). The minimum inhibitory concentrations (MICs), by broth microdilution method, varied for BAC from 0.25mg/L to 8mg/L (MIC50 = 1mg/L, MIC90 = 2mg/L) and for CHX from 0.5mg/L to 2mg/L (MIC50 = 1mg/L, MIC90 = 2mg/L). The BAC and CHX minimum bactericidal concentrations (MBCs) varied from 2mg/L to 8mg/L (MBC50 = 4mg/L, MBC90 = 8mg/L) and from 2mg/L to 4mg/L (MBC50 and MBC90 = 4mg/L), respectively. A reduced susceptibility to CHX (MIC = 2mg/L) was observed for 12.2% of the strains and that to BAC (MIC ≥ 4mg/L) for 4.1%. The norA resistance gene was detected in all the 49 isolates, whereas the qacA gene was rarely encountered (two strains; 4.1%). The qacC, qacG, qacH, and qacJ genes were not detected. The two strains harboring the qacA gene had reduced susceptibility to both antiseptics and belonged to CC3. The norA gene was detected in all the strains, suggesting that it could belong to the core genome of S. lugdunensis. S. lugdunensis is highly susceptible to both antiseptics tested. Reduced susceptibility to BAC and CHX was a rare phenomenon. Of note, a tendency to higher MICs of BAC was detected for CC3 isolates. These results should be confirmed on a larger collection of strains.

Profile and resistance levels of 136 integron resistance genes

Integrons have played a major role in the rise and spread of multidrug resistance in Gram-negative pathogens and are nowadays commonplace among clinical isolates. These platforms capture, stockpile, and modulate the expression of more than 170 antimicrobial resistance cassettes (ARCs) against most clinically-relevant antibiotics. Despite their importance, our knowledge on their profile and resistance levels is patchy, because data is scattered in the literature, often reported in different genetic backgrounds and sometimes extrapolated from sequence similarity alone. Here we have generated a collection of 136 ARCs against 8 antibiotic families and disinfectants. Cassettes are cloned in a vector designed to mimic the genetic environment of a class 1 integron, and transformed in Escherichia coli. We have measured the minimal inhibitory concentration (MIC) to the most relevant molecules from each antibiotic family. With more than 500 MIC values, we provide an exhaustive and comparable quantitation of resistance conferred by ARCs. Our data confirm known resistance trends and profiles while revealing important differences among closely related genes. We have also detected genes that do not confer the expected resistance, to the point of challenging the role of the whole family of qac genes in resistance against disinfectants. Our work provides a detailed characterization of integron resistance genes at-a-glance.

Detection of The Quaternary Ammonium Compound Antiseptic Resistance Gene (qac A/B qac C and qac E gene) In Pseudomonas Aeruginosa Recovered from Topical Wound Infection

Background: Quaternary ammonium compounds (QACs) have represented one of the most visible and effective classes of antiseptics and disinfectants substances that used in different parts of life including food production, water treatment and healthcare such as hospitals to prevent infections and intoxications. The increased use of disinfectants containing quaternary ammonium compounds (QACs) has raised concerns about the development of resistance. Aim of the study: this study pursues to determination frequency of antiseptic resistant Quaternary Ammonium Compound (qacA/B, qacC and qacE gene) in Pseudomonas aeruginosa, recovered from topical wound infection. Methods: Two swab samples were collected from 200 patients complain from burn, traumatic injury and wound after surgery, Suspected isolates of P. aeruginosa which showed Beta hemolysis on Blood agar media while occur colorless colonies (non-lactose fermenter) on MacConkey. Furthermore, identified using VITEK-2 System. The efficacy of antiseptics chlorhexidine (CLX), benzalkonium (BZK) and cetrimide (CET) was tested against P. aeruginosa by Well diffusion assay test and MIC procedure. The confirmed P. aeruginosa isolates have been used for Genomic DNA extraction from a fresh overnight culture, after that DNA templates was used to target (qacA/B, qacC and qacE gene) by using specific Primers sequences. Results: Out of 56 P. aeruginosa a considerable number of isolates 48 (85.7%) of them were resistance to chlorhexidine and 8 (14.3%) were sensitive, moreover substantial number of isolates 42 (75.0%) of them were resistance to cetrimide and 14 (25.0%) were sensitive; No sensitive rate to benzalkonium were observed in all isolates, qacA/B gene were identified in 32(57.1%) were positive for this gene while qacC gene were detected in 46(82.1%) were positive for this gene, Furthermore, qacE gene were detected in 50 (89.2%) were positive for this gene in 56 P. aeruginosa isolates. Conclusion: the current study, shows that the qacA/B, qacC and qacE genes which harbored resistance to quaternary ammonium compound antiseptics are widespread in Pseudomonas aeruginosa isolates in wound and burn patients.

Effect of chlorine disinfectant influx on biological sewage treatment process under the COVID-19 pandemic: Performance, mechanisms and implications

Since the onset of the COVID-19 Pandemic, large amounts of chlorine-containing disinfectants have been used to interrupt the spread of SARS-CoV-2 and residual chlorine eventually entered the hospital or municipal sewage treatment facilities. However, little is known about the effect of chlorine influx on the biological sewage treatment process. Here we investigated the effect of chlorine on the microbiome and the mechanism of microbial chlorine resistance in the activated sludge of the aerobic treatment process, using metagenomic and metatranscriptomic sequencing. We found that chlorine could negatively impact the aerobic treatment performance regarding nitrogen/COD removal with a dose-dependent effect, and the dual effects of chlorine dose and interaction time differentiated the microbial community in activated sludge. The decline of nitrogen/COD removal was attributed to the compressed activity of functional microorganisms, such as the ammonia oxidation bacteria, under chlorinated conditions, and the damage cannot be recovered in a short term. In addition, some microorganisms could survive in chlorinated conditions by up-regulating the chlorine resistance genes (CRGs) expression (approximately 1.5 times) and stimulating new CRGs expression. In particular, species Acinetobacter johnsonii could resist high concentrations of chlorine through various mechanisms, especially the overexpression of efflux pump function encoded by qac genes play a key role. Based on these results, considering the persistence of the epidemic and extensive use of chlorine disinfectants, it cannot be ignored that large amounts of residual chlorine are entering the biological treatment facility, and strictly de-chlorination measures or microbial chlorine resistance regulations before entering should be implemented.

Resistance profiles and genotyping of extended-spectrum beta-lactamase (ESBL) -producing and non-ESBL-producing E. coli and Klebsiella from retail market fishes.

Studies on antimicrobial resistance (AMR) profiles and epidemiological affirmation for AMR transmission are limited in fisheries and aquaculture settings. Since 2015, based on Global Action Plan on AMR by World Health Organization (WHO) and World Organization for Animal Health (OIE), several initiatives have been under taken to enhance the knowledge, skills and capacity to establish AMR trends through surveillance and strengthening of epidemiological evidence. The focus of this study was to determine the prevalence of antimicrobial resistance (AMR), its resistance profiles and molecular characterization with respect to phylogroups, antimicrobial resistance genes (ARGs), virulence genes (VGs), quaternary ammonium compounds resistance (QAC) genes and plasmid typing in retail market fishes. Pulse field gel electrophoresis (PFGE) to understand the genetic lineage of the two most important Enterobacteriaceae members, E. coli and Klebsiella sp. was performed. 94 fish samples were collected from three different sites viz., Silagrant (S1), Garchuk (S2) and North Guwahati Town Committee (NGTC) Region (S3) in Guwahati, Assam. Out of the 113 microbial isolates from the fish samples, 45 (39.82%) were E. coli; 23 (20.35%) belonged to Klebsiella genus. Among E. coli, 48.88% (n=22) of the isolates were alerted by the BD Phoenix M50 instrument as ESBL, 15.55% (n=7) as PCP and 35.55% (n=16) as non-ESBL. E. coli (39.82%) was the most prevalent pathogen among the Enterobacteriaceae members screened and showed resistance to ampicillin (69%) followed by cefazoline (64%), cefotaxime (49%) and piperacillin (49%). In the present study, 66.66% of E. coli and 30.43% of Klebsiella sp. were categorized as multi drug resistance (MDR) bacteria. CTX-M-gp-1, with CTX-M-15 variant (47%), was the most widely circulating beta-lactamase gene, while other ESBL genes blaTEM (7%), blaSHV (2%) and blaOXA-1-like (2%) were also identified in E. coli. Out of the 23 isolates of Klebsiella, 14(60.86%) were ampicillin (AM)-resistant (11(47.82%) K. oxytoca, 3(13.04%) K. aerogenes), whereas 8(34.78%) isolates of K. oxytoca showed intermediate resistance to AM. All Klebsiella isolates were susceptible to AN, SCP, MEM and TZP, although two K. aerogenes were resistant to imipenem. DHA and LAT genes were detected, respectively, in 7(16%) and 1(2%) of the E. coli strains while a single K. oxytoca (4.34%) isolate carried MOX, DHA and blaCMY-2 genes. The fluoroquinolone resistance genes identified in E. coli included qnrB (71%), qnrS (84%), oqxB (73%) and aac(6)-Ib-cr (27%); however, in Klebsiella, these genes, respectively, had a prevalence of 87%, 26%, 74% and 9%. The E. coli isolates belonged to phylogroup A(47%), B1(33%) and D(14%). All of the 22(100%) ESBL E. coli had chromosome-mediated disinfectant resistance genes viz., ydgE, ydgF, sugE(c), mdfA while 82% of ESBL E. coli had emrE. Among the non-ESBL E. coli isolates, 87% of them showed the presence of ydgE, ydgF and sugE(c) genes, while 78% of the isolates had mdfA and 39% had emrE genes respectively. 59% of the ESBL and 26% of the non-ESBL E. coli had showed the presence of qacEΔ1. The sugE(p) was present in 27% of the ESBL-producing E. coli and in 9% of non-ESBL isolates. Out of the 3 ESBL-producing Klebsiella isolates, 2(66.66%) K. oxytoca isolates were found harboring plasmid-mediated qacEΔ1 gene while one (33.33%) K. oxytoca isolate had sugE(p) gene. IncFI was the most prevalent plasmid type detected in the isolates studied, with A/C (18%), P (14%), X, Y (9% each) and I1-Iγ (14%, 4%). 50% (n=11) of the ESBL and 17% (n=4) of the non-ESBL E. coli isolates harboured IncFIB and 45% (n=10) ESBL and one (4.34%) non-ESBL E. coli isolates harboured IncFIA. Dominance of E. coli over other Enterobacterales and diverse phylogenetic profiles of E. coli and Klebsiella sp. suggests the possibility of contamination and this may be due to compromised hygienic practices along the supply chain and contamination of aquatic ecosystem. Continuous surveillance in domestic markets must be a priority in addressing antimicrobial resistance in fishery settings and to identify any unwarranted epidemic clones of E. coli and Klebsiella that can challenge public health sector.

Class 1 integron carrying qacEΔ1 gene confers resistance to disinfectant and antibiotics in Salmonella

Salmonella has presented increasingly alarming rates of antimicrobial resistance believed to be a result of a high prevalence of integrons. It is speculated that disinfectant-resistant isolates are due to the expression of qacEΔ1, an efflux pump located in the 3′ conserved sequence (3′CS) of class 1 integrons. With this concern, we tested the antibiotic and disinfectant resistance of 581 Salmonella strains collected from different sources, and characterized their integron structures. Gene expression and induction experiments were also performed. Results showed that Salmonella have high resistance to antimicrobials, especially to sulfonamides (SAs, 78.83 %), tetracyclines (TCs, 75.04 %) and benzalkonium chloride (BC, 87.26 %). The multi-drug resistance (MDR) frequency reached up to 63.17 %, and the prevalence of intI1 was 45.78 %. Molecular characterization of class 1 integrons exhibited nine different gene cassette arrays, of these, dfrA12-orf-aadA2 (n = 75), EstX (n = 25) and aadA2 (n = 14) were the most frequent. Importantly, 74.06 % of intI1-positive isolates were carrying qacEΔ1-sul1 genes in the 3′CS. This study also demonstrated that phenotypic resistance to both antibiotics and disinfectants was significantly correlated with the emergence of intI1 (p < 0.05). 91.37 % of qacEΔ1-sul1 positive Salmonella were found with disinfectant resistance. Additionally, expression of qacEΔ1 gene in Escherichia coli confirmed qacEΔ1 is predominantly involved in conferring disinfectant resistance. Disinfectant induction experiments further implicated qacEΔ1 in disinfectant resistance. RT-qPCR revealed a disinfectant-mediated increase in the relative expression of antibiotic-resistant genes (ARGs), aadA2 and dfrA12 on the integron, and efflux pump genes (mdtH and acrD) indicating that disinfectant could trigger co or cross-resistance. Therefore, our study confirmed that using disinfectant could provide selection pressure for strains with acquired resistance to antibiotics, providing new insights into the public health impact of Salmonella and guide continued efforts in antimicrobial stewardship and prevention of antibiotic resistance.

A comparative study of intestinal Pseudomonas aeruginosa in healthy individuals and ICU inpatients

The human intestinal tract is considered the most important reservoir of the opportunistic pathogens, including Pseudomonas aeruginosa, which is often overlooked but critical due to its antimicrobial resistance and virulence. Public health interventions to control this pathogen require a comprehensive understanding of its epidemiology and genomics. In the current study, we identified P. aeruginosa strains from 2,605 fecal samples collected between 2021 to 2022. Among these samples, 574 were from ICU inpatients in Zhejiang province, while 2,031 were obtained from healthy individuals residing in ten different provinces in China. The prevalence of P. aeruginosa intestinal carriage was found to be higher in ICU inpatients (10.28%, 95% CI: 7.79%–12.76%) than that in healthy individuals (3.99%, 81/2,031, 95% CI: 3.14%–4.84%). Similarly, the prevalence of carbapenem-resistant P. aeruginosa (CRPA) was higher in ICU inpatients (32.2%) compared to healthy individuals (7.41%). The population structure analysis of our isolates revealed a predominantly non-clonal distribution, with 41 distinct sequence types identified among 59 P. aeruginosa isolates from ICU inpatients and 38 different STs among 81 P. aeruginosa isolates from healthy individuals. These findings suggest that the individual acquisition of P. aeruginosa is more frequent than patient-to-patient transmission, as evidenced by the polyclonal population structure. Antimicrobial susceptibility testing and genome analysis indicated that P. aeruginosa strains from ICU inpatients exhibited significantly higher resistance rates to most antimicrobials and harbored a greater number of acquired resistance genes compared to strains from healthy individuals. Notably, in ICU inpatients, we identified three isolates of ST463, all of which shared the conserved Tn3-TnpR-ISKpn8-blaKPC-ISKpn6 genetic context. Additionally, five isolates carrying the qacE gene were also identified, these findings suggest that small-scale transmission events may still occur within the ICU setting, posing significant challenges for clinical management. With regard to virulence factors, we observed similar profiles between the two groups, except for phzA2, phzB2, and pilA, which were statistically higher in isolates from healthy individuals. This may be because the accumulating resistance mutations in ICU-derived P. aeruginosa are linked to a decrease in virulence.