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Antimicrobial-resistant Klebsiella pneumoniae is one of the predominant pathogens in healthcare settings. However, the prevalence and resistome of this organism within residential aged care facilities (RACFs), which are potential hotspots for antimicrobial resistance, remain unexplored. Here, we provide a phenotypic and molecular characterization of antimicrobial-resistant K. pneumoniae isolated from RACFs. K. pneumoniae was isolated from urine, faecal and wastewater samples and facility swabs. The antimicrobial susceptibility profiles of all the isolates were determined and the genomic basis for resistance was explored with whole-genome sequencing on a subset of isolates. A total of 147 K. pneumoniae were isolated, displaying resistance against multiple antimicrobials. Genotypic analysis revealed the presence of beta-lactamases and the ciprofloxacin-resistance determinant QnrB4 but failed to confirm the basis for the observed cephalosporin resistance. Clonal spread of the multidrug-resistant, widely disseminated sequence types 323 and 661 was observed. This study was the first to examine the resistome of K. pneumoniae isolates from RACFs and demonstrated a complexity between genotypic and phenotypic antimicrobial resistance. The intra-facility dissemination and persistence of multidrug-resistant clones is concerning, given that residents are particularly vulnerable to antimicrobial resistant infections, and it highlights the need for continued surveillance and interventions to reduce the risk of outbreaks. [ABSTRACT FROM AUTHOR]

Understanding the role of foods in the emergence and spread of antimicrobial resistance necessitates the initial documentation of antibiotic resistance genes within bacterial species found in foods. Here, the NCBI Pathogen Detection database was used to query antimicrobial resistance gene prevalence in foodborne and human clinical bacterial isolates. Of the 1,843,630 sequence entries, 639,087 (34.7%) were assigned to foodborne or human clinical sources with 147,788 (23.14%) from food and 427,614 (76.88%) from humans. The majority of foodborne isolates were either Salmonella (47.88%), Campylobacter (23.03%), Escherichia (11.79%), or Listeria (11.3%), and the remaining 6% belonged to 20 other genera. Most foodborne isolates were from meat/poultry (95,251 or 64.45%), followed by multi-product mixed food sources (29,892 or 20.23%) and fish/seafood (6503 or 4.4%); however, the most prominent isolation source varied depending on the genus/species. Resistance gene carriage also varied depending on isolation source and genus/species. Of note, Klebsiella pneumoniae and Enterobacter spp. carried larger proportions of the quinolone resistance gene qnrS and some clinically relevant beta-lactam resistance genes in comparison to Salmonella and Escherichia coli. The prevalence of mec in S. aureus did not significantly differ between meat/poultry and multi-product sources relative to clinical sources, whereas this resistance was rare in isolates from dairy sources. The proportion of biocide resistance in Bacillus and Escherichia was significantly higher in clinical isolates compared to many foodborne sources but significantly lower in clinical Listeria compared to foodborne Listeria. This work exposes the gaps in current publicly available sequence data repositories, which are largely composed of clinical isolates and are biased towards specific highly abundant pathogenic species. We also highlight the importance of requiring and curating metadata on sequence submission to not only ensure correct information and data interpretation but also foster efficient analysis, sharing, and collaboration. To effectively monitor resistance carriage in food production, additional work on sequencing and characterizing AMR carriage in common commensal foodborne bacteria is critical. [ABSTRACT FROM AUTHOR]

Summary: Klebsiella pneumoniae is among the World Health Organization's list of priority pathogens, notorious for its role in causing healthcare-associated infections and neonatal sepsis globally. Containment of K. pneumoniae transmission depends on the continued effectiveness of antimicrobials and of biocides used for topical antisepsis and surface disinfection. Klebsiella pneumoniae is known to disseminate antimicrobial resistance (AMR) through a large auxiliary genome made up of plasmids, transposons and integrons, enabling it to evade antimicrobial killing through the use of efflux systems and biofilm development. Because AMR mechanisms are also known to impart tolerance to biocides, AMR is frequently linked with biocide resistance (BR). However, despite extensive research on AMR, there is a gap in knowledge about BR and the extent to which AMR and BR mechanisms overlap remains debatable. The aim of this paper is to review and summarise the current knowledge on the determinants of BR in K. pneumoniae and highlight content areas that require further inquiry.

Benzalkonium chloride is a quaternary ammonium compound with bactericidal activity, which is widely used in domestic and healthcare settings as well as in food processing environments. It plays an important role in the prevention and control of foodborne pathogens. However, the presence of biofilms or improper use of benzalkonium chloride may enable foodborne pathogens to adapt to bactericides, affecting the tolerance to homologous or heterologous biocidal agents. The review summarizes the relationship between the adaptation of foodborne pathogens to benzalkonium chloride and their tolerance to this biocide. The reasons for the increased tolerance to benzalkonium chloride are explained from the perspectives of the expression of efflux pump genes, cell membrane modification, and the formation of the viable but non-cultivable (VBNC) state. Furthermore, the influence of benzalkonium chloride adaptation on the antibiotic resistance of foodborne pathogens is discussed, and the reasons for the increased antibiotic resistance are summarized from the perspectives of biofilm formation, spontaneous mutations, horizontal gene transfer, and the role of efflux pumps in antibiotic resistance. This review could help understand the mechanism of the adaption of foodborne pathogens to benzalkonium chloride and provide a reference for the rational use of biocides and the prevention and control of foodborne pathogens.

Antimicrobial-resistant Klebsiella pneumoniae is one of the predominant pathogens in healthcare settings. However, the prevalence and resistome of this organism within residential aged care facilities (RACFs), which are potential hotspots for antimicrobial resistance, remain unexplored. Here, we provide a phenotypic and molecular characterization of antimicrobial-resistant K. pneumoniae isolated from RACFs. K. pneumoniae was isolated from urine, faecal and wastewater samples and facility swabs. The antimicrobial susceptibility profiles of all the isolates were determined and the genomic basis for resistance was explored with whole-genome sequencing on a subset of isolates. A total of 147 K. pneumoniae were isolated, displaying resistance against multiple antimicrobials. Genotypic analysis revealed the presence of beta-lactamases and the ciprofloxacin-resistance determinant QnrB4 but failed to confirm the basis for the observed cephalosporin resistance. Clonal spread of the multidrug-resistant, widely disseminated sequence types 323 and 661 was observed. This study was the first to examine the resistome of K. pneumoniae isolates from RACFs and demonstrated a complexity between genotypic and phenotypic antimicrobial resistance. The intra-facility dissemination and persistence of multidrug-resistant clones is concerning, given that residents are particularly vulnerable to antimicrobial resistant infections, and it highlights the need for continued surveillance and interventions to reduce the risk of outbreaks.

Acinetobacter baumannii, a pathogenic bacterium acquired in hospitals, causes diverse infections in humans. Previous studies have reported resistance among A. baumannii strains, potentially selecting multi-drug-resistant variants. In Pakistan, research has primarily focused on carbapenem-resistant A. baumannii (CRAB) strains, overlooking the investigation of efflux pumps (EPs) and biocide resistance. This study aims to assess A. baumannii strains from five hospitals in Pakistan, focusing on antibiotic and biocide susceptibility, the impact of EP inhibitors on antimicrobial susceptibility, and the distribution of ARGs and STs. A total of 130 non-repeated Acinetobacter baumannii isolates were collected from five tertiary care hospitals in Pakistan and identified using API 20NE and multiplex PCR. Antimicrobial susceptibility testing utilized disc diffusion and broth microdilution assays, while biocide susceptibility was assessed with various agents. The impact of an efflux pump inhibitor (NMP) on antibiotic susceptibility was evaluated. PCR screening for ARGs and EPGs was followed by DNA sequencing validation. MLST was performed using the Pasteur scheme. Most isolates demonstrated resistance to tested antibiotics, with varying levels of susceptibility to biocides. All isolates exhibited the intrinsic class D β-lactamase blaOXA-51, while acquired blaOXA-23 was present in all CRAB isolates. Among EPs, adeJ, abeD, amvA, and aceI were prevalent in almost all isolates, with adeB found in 93% of isolates and adeG, adeT1, adeT2, and qacEΔ1 displaying lower prevalence ranging from 65% to 79%. The most common STs were ST589 and ST2, accounting for 28.46% and 25.38% of isolates, respectively, followed by ST642 at 12.6%. These findings indicate that A. baumannii strains in Pakistan are resistant to antibiotics (excluding colistin and tigecycline) and may be developing biocide resistance, which could contribute to the selection and dissemination of multi-drug-resistant strains. [ABSTRACT FROM AUTHOR]

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Chlorhexidine (CHD) is a cationic biocide used ubiquitously in healthcare settings. Proteus mirabilis, an important pathogen of the catheterized urinary tract, and isolates of this species are often described as "resistant" to CHD-containing products used for catheter infection control. To identify the mechanisms underlying reduced CHD susceptibility in P. mirabilis, we subjected the CHD tolerant clinical isolate RS47 to random transposon mutagenesis and screened for mutants with reduced CHD minimum inhibitory concentrations (MICs). One mutant recovered from these screens (designated RS47-2) exhibited ~8-fold reduction in CHD MIC. Complete genome sequencing of RS47-2 showed a single mini-Tn5 insert in the waaC gene involved in lipopolysaccharide (LPS) inner core biosynthesis. Phenotypic screening of RS47-2 revealed a significant increase in cell surface hydrophobicity and serum susceptibility compared to the wildtype, and confirmed defects in LPS production congruent with waaC inactivation. Disruption of waaC was also associated with increased susceptibility to a range of other cationic biocides but did not affect susceptibility to antibiotics tested. Complementation studies showed that repression of smvA efflux activity in RS47-2 further increased susceptibility to CHD and other cationic biocides, reducing CHD MICs to values comparable with the most CHD susceptible isolates characterized. The formation of crystalline biofilms and blockage of urethral catheters was also significantly attenuated in RS47-2. Taken together, these data show that aspects of LPS structure and upregulation of the smvA efflux system function in synergy to modulate susceptibility to CHD and other cationic biocides, and that LPS structure is also an important factor in P. mirabilis crystalline biofilm formation. [ABSTRACT FROM AUTHOR]

Understanding the role of foods in the emergence and spread of antimicrobial resistance necessitates the initial documentation of antibiotic resistance genes within bacterial species found in foods. Here, the NCBI Pathogen Detection database was used to query antimicrobial resistance gene prevalence in foodborne and human clinical bacterial isolates. Of the 1,843,630 sequence entries, 639,087 (34.7%) were assigned to foodborne or human clinical sources with 147,788 (23.14%) from food and 427,614 (76.88%) from humans. The majority of foodborne isolates were either Salmonella (47.88%), Campylobacter (23.03%), Escherichia (11.79%), or Listeria (11.3%), and the remaining 6% belonged to 20 other genera. Most foodborne isolates were from meat/poultry (95,251 or 64.45%), followed by multi-product mixed food sources (29,892 or 20.23%) and fish/seafood (6503 or 4.4%); however, the most prominent isolation source varied depending on the genus/species. Resistance gene carriage also varied depending on isolation source and genus/species. Of note, Klebsiella pneumoniae and Enterobacter spp. carried larger proportions of the quinolone resistance gene qnrS and some clinically relevant beta-lactam resistance genes in comparison to Salmonella and Escherichia coli. The prevalence of mec in S. aureus did not significantly differ between meat/poultry and multi-product sources relative to clinical sources, whereas this resistance was rare in isolates from dairy sources. The proportion of biocide resistance in Bacillus and Escherichia was significantly higher in clinical isolates compared to many foodborne sources but significantly lower in clinical Listeria compared to foodborne Listeria. This work exposes the gaps in current publicly available sequence data repositories, which are largely composed of clinical isolates and are biased towards specific highly abundant pathogenic species. We also highlight the importance of requiring and curating metadata on sequence submission to not only ensure correct information and data interpretation but also foster efficient analysis, sharing, and collaboration. To effectively monitor resistance carriage in food production, additional work on sequencing and characterizing AMR carriage in common commensal foodborne bacteria is critical.

Chlorhexidine (CHD) is a cationic biocide used ubiquitously in healthcare settings. Proteus mirabilis, an important pathogen of the catheterized urinary tract, and isolates of this species are often described as “resistant” to CHD-containing products used for catheter infection control. To identify the mechanisms underlying reduced CHD susceptibility in P. mirabilis, we subjected the CHD tolerant clinical isolate RS47 to random transposon mutagenesis and screened for mutants with reduced CHD minimum inhibitory concentrations (MICs). One mutant recovered from these screens (designated RS47-2) exhibited ~ 8-fold reduction in CHD MIC. Complete genome sequencing of RS47-2 showed a single mini-Tn5 insert in the waaC gene involved in lipopolysaccharide (LPS) inner core biosynthesis. Phenotypic screening of RS47-2 revealed a significant increase in cell surface hydrophobicity and serum susceptibility compared to the wildtype, and confirmed defects in LPS production congruent with waaC inactivation. Disruption of waaC was also associated with increased susceptibility to a range of other cationic biocides but did not affect susceptibility to antibiotics tested. Complementation studies showed that repression of smvA efflux activity in RS47-2 further increased susceptibility to CHD and other cationic biocides, reducing CHD MICs to values comparable with the most CHD susceptible isolates characterized. The formation of crystalline biofilms and blockage of urethral catheters was also significantly attenuated in RS47-2. Taken together, these data show that aspects of LPS structure and upregulation of the smvA efflux system function in synergy to modulate susceptibility to CHD and other cationic biocides, and that LPS structure is also an important factor in P. mirabilis crystalline biofilm formation.

Staphylococcus aureus biofilms are resistant to both antibiotics and disinfectants. As Staphylococci cell walls are an important defence mechanism, we sought to examine changes to the bacterial cell wall under different growth conditions. Cell walls of S. aureus grown as 3-day hydrated biofilm, 12-day hydrated biofilm, and 12-day dry surface biofilm (DSB) were compared to cell walls of planktonic organisms. Additionally, proteomic analysis using high-throughput tandem mass tag-based mass spectrometry was performed. Proteins involved in cell wall synthesis in biofilms were upregulated in comparison to planktonic growth. Bacterial cell wall width (measured by transmission electron microscopy) and peptidoglycan production (detected using a silkworm larva plasma system) increased with biofilm culture duration (p < 0.001) and dehydration (p = 0.002). Similarly, disinfectant tolerance was greatest in DSB, followed by 12-day hydrated biofilm and then 3-day biofilm, and it was least in the planktonic bacteria––suggesting that changes to the cell wall may be a key factor for S. aureus biofilm biocide resistance. Our findings shed light on possible new targets to combat biofilm-related infections and hospital dry surface biofilms. [ABSTRACT FROM AUTHOR]

Acinetobacter baumannii is especially known as a cause of nosocomial infections worldwide. It shows intrinsic and acquired resistances to numerous antimicrobial agents, which can render the treatment difficult. In contrast to the situation in human medicine, there are only few studies focusing on A. baumannii among livestock. In this study, we have examined 643 samples from turkeys reared for meat production, including 250 environmental and 393 diagnostic samples, for the presence of A. baumannii. In total, 99 isolates were identified, confirmed to species level via MALDI-TOF-MS and characterised with pulsed-field gel electrophoresis. Antimicrobial and biocide susceptibility was tested by broth microdilution methods. Based on the results, 26 representative isolates were selected and subjected to whole-genome sequencing (WGS). In general, A. baumannii was detected at a very low prevalence, except for a high prevalence of 79.7% in chick-box-papers (n = 118) of one-day-old turkey chicks. The distributions of the minimal inhibitory concentration values were unimodal for the four biocides and for most of the antimicrobial agents tested. WGS revealed 16 Pasteur and 18 Oxford sequence types, including new ones. Core genome MLST highlighted the diversity of most isolates. In conclusion, the isolates detected were highly diverse and still susceptible to many antimicrobial agents. [ABSTRACT FROM AUTHOR]

Molecular insights into the mechanisms of resistance to disinfectants are severely limited, together with the roles of various mobile genetic elements. Genomic islands are a well-characterised molecular resistance element in antibiotic resistance, but it is unknown whether genomic islands play a role in disinfectant resistance. Through whole-genome sequencing and the bioinformatic analysis of Serratia sp. HRI, an isolate with high disinfectant resistance capabilities, nine resistance islands were predicted and annotated within the genome. Resistance genes active against several antimicrobials were annotated in these islands, most of which are multidrug efflux pumps belonging to the MFS, ABC and DMT efflux families. Antibiotic resistance islands containing genes encoding for multidrug resistance proteins ErmB (macrolide and erythromycin resistance) and biclomycin were also found. A metal fitness island harbouring 13 resistance and response genes to copper, silver, lead, cadmium, zinc, and mercury was identified. In the search for disinfectant resistance islands, two genomic islands were identified to harbour smr genes, notorious for conferring disinfectant resistance. This suggests that genomic islands are capable of conferring disinfectant resistance, a phenomenon that has not yet been observed in the study of biocide resistance and tolerance. [ABSTRACT FROM AUTHOR]

Environmental reservoir of antibiotic and other resistance genes are an emerging public health concern, and especially so in food production sites. This study aimed to explore the co-occurrence of antibiotic resistance genes (ARG) and resistance genes to other compounds, including heavy metals and biocides, in urban vegetable garden soils. Total soil DNA was extracted and subjected to metagenomic analysis on ARG, metal resistance genes (MRG), biocide resistance genes (BRG), multi-compound resistance genes (MCRG), and mobile genetic elements (MGE). The relative abundance of ARGs and MRGs were 2.00–2.90 × 10−1 and 4.22–6.00 × 10−2 copies per 16S rRNA gene, respectively. The most abundant ARGs included multidrug, vancomycin, bacitracin, fosmidomycin, and macrolide-lincosamide-streptogramin (MLS) resistance genes. Common MRGs consisted of genes with resistance to copper (dnaK, copA, cueA), arsenic (arsM), multi-metal (recG_BAC0356, ruvB, cueA), iron (acn), and chromium (recG). BRGs identified were those conferring resistance to multi-biocide (smrA, mexW, tbtB, and emhB), peroxide (sodA, sodB), and phenolic compound (triC). MCRGs were found with resistance to biocide-metal (dpsA, ruvB, actP, ideR), drug-biocide-metal (mdtB), or drug-biocide (mexF, ceoB, sme, mexK, sdeB). Multidrug resistance genes (mexF, mexW, multidrug transporter gene), MLS resistance gene (macB), and MCRG (ruvB) showed most correlations with ARGs and resistance genes to other compounds. The data demonstrate extensive co-occurrence of ARGs, MRGs, BRGs, and MCRGs in urban agricultural environments and suggest the great potential of soil contaminants as co-selection agents for antibiotic resistance.

Disinfection is a key strategy to reduce the burden of infections. The contact of bacteria to biocides—the active substances of disinfectants—has been linked to bacterial adaptation and the development of antimicrobial resistance. Currently, there is no scientific consensus on whether the excessive use of biocides contributes to the emergence and spread of multidrug resistant bacteria. The comprehensive analysis of available data remains a challenge because neither uniform test procedures nor standardized interpretive criteria nor harmonized terms are available to describe altered bacterial susceptibility to biocides. In our review, we investigated the variety of criteria and the diversity of terms applied to interpret findings in original studies performing biocide susceptibility testing (BST) of field isolates. An additional analysis of reviews summarizing the knowledge of individual studies on altered biocide susceptibility provided insights into currently available broader concepts for data interpretation. Both approaches pointed out the urgent need for standardization. We, therefore, propose that the well-established and approved concepts for interpretation of antimicrobial susceptibility testing data should serve as a role model to evaluate biocide resistance mechanisms on a single cell level. Furthermore, we emphasize the adaptations necessary to acknowledge the specific needs for the evaluation of BST data. Our approach might help to increase scientific awareness and acceptance. [ABSTRACT FROM AUTHOR]

Microbes frequently encounter heavy metals and other toxic compounds generated from natural biogeochemical processes and anthropogenic activities. Here, we analyzed the prevalence and association of genes conferring resistance to heavy metals, biocides, and antimicrobial compounds in 394 genome sequences of clinical human-derived S. enterica from New Hampshire, USA. The most prevalent was the gold operon (gesABC-golTSB), which was present in 99.2% of the genomes. In contrast, the other five heavy metal operons (arsenic, copper, mercury, silver, tellurite) were present in 0.76% (3/394)-5.58% (22/394) of the total population. The heavy metal operons and three biocide resistance genes were differentially distributed across 15 sequence types (STs) and 16 serotypes. The number of heavy metal operons and biocide resistance genes per genome was significantly associated with high number of antimicrobial resistance (AMR) genes per genome. Notable is the mercury operon which exhibited significant association with genes conferring resistance to aminoglycosides, cephalosporins, diaminopyrimidine, sulfonamide, and fosfomycin. The mercury operon was co-located with the AMR genes aac(3)-IV, ant(3")-IIa, aph(3')-Ia, and aph(4)-Ia, CTX-M-65, dfrA14, sul1, and fosA3 genes within the same plasmid types. Lastly, we found evidence for negative selection of individual genes of each heavy metal operon and the biocide resistance genes (dN/dS < 1). Our study highlights the need for continued surveillance of S. enterica serotypes that carry those genes that confer resistance to heavy metals and biocides that are often associated with mobile AMR genes. The selective pressures imposed by heavy metals and biocides on S. enterica may contribute to the co-selection and spread of AMR in human infections. [ABSTRACT FROM AUTHOR]

Microbes frequently encounter heavy metals and other toxic compounds generated from natural biogeochemical processes and anthropogenic activities. Here, we analyzed the prevalence and association of genes conferring resistance to heavy metals, biocides, and antimicrobial compounds in 394 genome sequences of clinical human-derived S. enterica from New Hampshire, USA. The most prevalent was the gold operon (gesABC-golTSB), which was present in 99.2% of the genomes. In contrast, the other five heavy metal operons (arsenic, copper, mercury, silver, tellurite) were present in 0.76% (3/394)–5.58% (22/394) of the total population. The heavy metal operons and three biocide resistance genes were differentially distributed across 15 sequence types (STs) and 16 serotypes. The number of heavy metal operons and biocide resistance genes per genome was significantly associated with high number of antimicrobial resistance (AMR) genes per genome. Notable is the mercury operon which exhibited significant association with genes conferring resistance to aminoglycosides, cephalosporins, diaminopyrimidine, sulfonamide, and fosfomycin. The mercury operon was co-located with the AMR genes aac(3)-IV, ant(3”)-IIa, aph(3’)-Ia, and aph(4)-Ia, CTX-M-65, dfrA14, sul1, and fosA3 genes within the same plasmid types. Lastly, we found evidence for negative selection of individual genes of each heavy metal operon and the biocide resistance genes (dN/dS < 1). Our study highlights the need for continued surveillance of S. enterica serotypes that carry those genes that confer resistance to heavy metals and biocides that are often associated with mobile AMR genes. The selective pressures imposed by heavy metals and biocides on S. enterica may contribute to the co-selection and spread of AMR in human infections.

Enterococcus are opportunistic pathogens that have been gaining importance in the clinical setting, especially in terms of hospital-acquired infections. This problem has mainly been associated with the fact that these bacteria are able to present intrinsic and extrinsic resistance to different classes of antibiotics, with a great deal of importance being attributed to vancomycin-resistant enterococci. However, other aspects, such as the expression of different virulence factors including biofilm-forming ability, and its capacity of trading genetic information, makes this bacterial genus more capable of surviving harsh environmental conditions. All these characteristics, associated with some reports of decreased susceptibility to some biocides, all described in this literary review, allow enterococci to present a longer survival ability in the hospital environment, consequently giving them more opportunities to disseminate in these settings and be responsible for difficult-to-treat infections. [ABSTRACT FROM AUTHOR]

ABSTRACT Development of extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli is one the greatest threats faced by mankind. Among animals, chickens, pigs, and cattle are reservoirs of these pathogens worldwide. Nevertheless, there is a knowledge gap on ESBL-producing E. coli from small ruminants (i.e., sheep and goats) in China. The aim of this study was to identify and characterize the resistance profiles, resistomes, and sequence features of 67 ESBL-producing E. coli isolates from sheep in northwest China. The findings showed that blaCTX-M and blaTEM were the most prevalent. Interestingly, we found that the resistance gene mcr-1 was widespread in sheep merely from Shaanxi areas, accounting for 19.2% (5/26). The highly prevalent serotypes and FumC-FimH (CH) typing isolates were O8 and C4H32, respectively. High-risk E. coli clones, such as sequence type 10 (ST10), ST23, ST44, and ST58, were also found in China’s sheep population. A total of 67 ESBL-producing isolates were divided into five phylogenetic groups, namely, B1 (n = 47, 70.1%), B2 (n = 1, 1.5%), C (n = 14, 20.9%), E (n = 1, 1.5%), and F (n = 1, 1.5%), with the phylogenetic groups for 3 isolates (4.5%) remaining unknown. Moreover, ESBL-producing E. coli isolates were also characterized by the abundance and diversity of biocide/metal resistance genes and insert sequences. We found that in ESBL-producing E. coli isolates, there were two different types of isolates, those containing ESBL genes or not, which led to large discrepancies between resistance phenotypes and resistomes. In summary, our study provides a comprehensive overview of resistance profiles and genome sequence features in ESBL-producing E. coli and highlights the possible role of sheep as antibiotic resistance gene disseminators into humans. IMPORTANCE Antimicrobial resistance (AMR), especially the simultaneous resistance to several antibiotics (multidrug resistance [MDR]), is one of the greatest threats to global public health in the 21st century. Among animals, chickens, pigs, and cattle are reservoirs of these pathogens worldwide. Nevertheless, there is a knowledge gap on ESBL-producing E. coli from small ruminants in China. This study is the largest and most comprehensive analysis of ESBL-producing E. coli isolates from sheep, including antibiotic resistance profiles, phylogenetic groups, serotypes, multilocus sequence types (MLST), insert sequences (IS), antibiotic resistance genes, disinfectant resistance genes, and heavy metal resistance genes. We recommend extending the surveillance of AMR of sheep-origin E. coli to prevent future public health risks.

Acinetobacter baumannii, a pathogenic bacterium acquired in hospitals, causes diverse infections in humans. Previous studies have reported resistance among A. baumannii strains, potentially selecting multi-drug-resistant variants. In Pakistan, research has primarily focused on carbapenem-resistant A. baumannii (CRAB) strains, overlooking the investigation of efflux pumps (EPs) and biocide resistance. This study aims to assess A. baumannii strains from five hospitals in Pakistan, focusing on antibiotic and biocide susceptibility, the impact of EP inhibitors on antimicrobial susceptibility, and the distribution of ARGs and STs. A total of 130 non-repeated Acinetobacter baumannii isolates were collected from five tertiary care hospitals in Pakistan and identified using API 20NE and multiplex PCR. Antimicrobial susceptibility testing utilized disc diffusion and broth microdilution assays, while biocide susceptibility was assessed with various agents. The impact of an efflux pump inhibitor (NMP) on antibiotic susceptibility was evaluated. PCR screening for ARGs and EPGs was followed by DNA sequencing validation. MLST was performed using the Pasteur scheme. Most isolates demonstrated resistance to tested antibiotics, with varying levels of susceptibility to biocides. All isolates exhibited the intrinsic class D β-lactamase blaOXA-51, while acquired blaOXA-23 was present in all CRAB isolates. Among EPs, adeJ, abeD, amvA, and aceI were prevalent in almost all isolates, with adeB found in 93% of isolates and adeG, adeT1, adeT2, and qacEΔ1 displaying lower prevalence ranging from 65% to 79%. The most common STs were ST589 and ST2, accounting for 28.46% and 25.38% of isolates, respectively, followed by ST642 at 12.6%. These findings indicate that A. baumannii strains in Pakistan are resistant to antibiotics (excluding colistin and tigecycline) and may be developing biocide resistance, which could contribute to the selection and dissemination of multi-drug-resistant strains.

Acinetobacter baumannii is especially known as a cause of nosocomial infections worldwide. It shows intrinsic and acquired resistances to numerous antimicrobial agents, which can render the treatment difficult. In contrast to the situation in human medicine, there are only few studies focusing on A. baumannii among livestock. In this study, we have examined 643 samples from turkeys reared for meat production, including 250 environmental and 393 diagnostic samples, for the presence of A. baumannii. In total, 99 isolates were identified, confirmed to species level via MALDI-TOF-MS and characterised with pulsed-field gel electrophoresis. Antimicrobial and biocide susceptibility was tested by broth microdilution methods. Based on the results, 26 representative isolates were selected and subjected to whole-genome sequencing (WGS). In general, A. baumannii was detected at a very low prevalence, except for a high prevalence of 79.7% in chick-box-papers (n = 118) of one-day-old turkey chicks. The distributions of the minimal inhibitory concentration values were unimodal for the four biocides and for most of the antimicrobial agents tested. WGS revealed 16 Pasteur and 18 Oxford sequence types, including new ones. Core genome MLST highlighted the diversity of most isolates. In conclusion, the isolates detected were highly diverse and still susceptible to many antimicrobial agents.

Molecular insights into the mechanisms of resistance to disinfectants are severely limited, together with the roles of various mobile genetic elements. Genomic islands are a well-characterised molecular resistance element in antibiotic resistance, but it is unknown whether genomic islands play a role in disinfectant resistance. Through whole-genome sequencing and the bioinformatic analysis of Serratia sp. HRI, an isolate with high disinfectant resistance capabilities, nine resistance islands were predicted and annotated within the genome. Resistance genes active against several antimicrobials were annotated in these islands, most of which are multidrug efflux pumps belonging to the MFS, ABC and DMT efflux families. Antibiotic resistance islands containing genes encoding for multidrug resistance proteins ErmB (macrolide and erythromycin resistance) and biclomycin were also found. A metal fitness island harbouring 13 resistance and response genes to copper, silver, lead, cadmium, zinc, and mercury was identified. In the search for disinfectant resistance islands, two genomic islands were identified to harbour smr genes, notorious for conferring disinfectant resistance. This suggests that genomic islands are capable of conferring disinfectant resistance, a phenomenon that has not yet been observed in the study of biocide resistance and tolerance.

Microorganisms are capable of colonizing extreme environments like deep biosphere and oil reservoirs. The prokaryotes diversity in exploited oil reservoirs is composed of indigenous microbial communities and artificially introduced microbes. In the present work, high throughput sequencing techniques were applied to analyze the microbial community from the injected and produced water in a neotropical hyper-thermophile oil reservoir located in the Orinoquia region of Colombia, South America. Tepidiphilus is the dominant bacteria found in both injection and produced waters. The produced water has a higher microbial richness and exhibits a Tepidiphilus microdiversity. The reservoir injected water is recycled and treated with the biocides glutaraldehyde and tetrakis-hydroxymethyl-phosphonium sulfate (THPS) to reduce microbial load. This process reduces microbial richness and selects a single Tepidiphilus genome (T. sp. UDEAICP_D1) as the dominant isolate. Thermus and Hydrogenobacter were subdominants in both water systems. Phylogenomic analysis of the injection water dominant Tepidiphilus positioned it as an independent branch outside T. succinatimandens and T. thermophilus lineage. Comparative analysis of the Tepidiphilus genomes revealed several genes that might be related to the biocide-resistant phenotype and the tolerance to the stress conditions imposed inside the oil well, like RND efflux pumps and type II toxin-antitoxin systems. Comparing the abundance of Tepidiphilus protein-coding genes in both water systems shows that the biocide selected Tepidiphilus sp. UDEAICP_D1 genome has enriched genes annotated as ABC-2 type transporter, ABC transporter, Methionine biosynthesis protein MetW, Glycosyltransferases, and two-component system NarL. [ABSTRACT FROM AUTHOR]

Microorganisms are capable of colonizing extreme environments like deep biosphere and oil reservoirs. The prokaryotes diversity in exploited oil reservoirs is composed of indigenous microbial communities and artificially introduced microbes. In the present work, high throughput sequencing techniques were applied to analyze the microbial community from the injected and produced water in a neotropical hyper-thermophile oil reservoir located in the Orinoquia region of Colombia, South America. Tepidiphilus is the dominant bacteria found in both injection and produced waters. The produced water has a higher microbial richness and exhibits a Tepidiphilus microdiversity. The reservoir injected water is recycled and treated with the biocides glutaraldehyde and tetrakis-hydroxymethyl-phosphonium sulfate (THPS) to reduce microbial load. This process reduces microbial richness and selects a single Tepidiphilus genome (T. sp. UDEAICP_D1) as the dominant isolate. Thermus and Hydrogenobacter were subdominants in both water systems. Phylogenomic analysis of the injection water dominant Tepidiphilus positioned it as an independent branch outside T. succinatimandens and T. thermophilus lineage. Comparative analysis of the Tepidiphilus genomes revealed several genes that might be related to the biocide-resistant phenotype and the tolerance to the stress conditions imposed inside the oil well, like RND efflux pumps and type II toxin-antitoxin systems. Comparing the abundance of Tepidiphilus protein-coding genes in both water systems shows that the biocide selected Tepidiphilus sp. UDEAICP_D1 genome has enriched genes annotated as ABC-2 type transporter, ABC transporter, Methionine biosynthesis protein MetW, Glycosyltransferases, and two-component system NarL.

Disinfection is a key strategy to reduce the burden of infections. The contact of bacteria to biocides—the active substances of disinfectants—has been linked to bacterial adaptation and the development of antimicrobial resistance. Currently, there is no scientific consensus on whether the excessive use of biocides contributes to the emergence and spread of multidrug resistant bacteria. The comprehensive analysis of available data remains a challenge because neither uniform test procedures nor standardized interpretive criteria nor harmonized terms are available to describe altered bacterial susceptibility to biocides. In our review, we investigated the variety of criteria and the diversity of terms applied to interpret findings in original studies performing biocide susceptibility testing (BST) of field isolates. An additional analysis of reviews summarizing the knowledge of individual studies on altered biocide susceptibility provided insights into currently available broader concepts for data interpretation. Both approaches pointed out the urgent need for standardization. We, therefore, propose that the well-established and approved concepts for interpretation of antimicrobial susceptibility testing data should serve as a role model to evaluate biocide resistance mechanisms on a single cell level. Furthermore, we emphasize the adaptations necessary to acknowledge the specific needs for the evaluation of BST data. Our approach might help to increase scientific awareness and acceptance.

Enterococcus are opportunistic pathogens that have been gaining importance in the clinical setting, especially in terms of hospital-acquired infections. This problem has mainly been associated with the fact that these bacteria are able to present intrinsic and extrinsic resistance to different classes of antibiotics, with a great deal of importance being attributed to vancomycin-resistant enterococci. However, other aspects, such as the expression of different virulence factors including biofilm-forming ability, and its capacity of trading genetic information, makes this bacterial genus more capable of surviving harsh environmental conditions. All these characteristics, associated with some reports of decreased susceptibility to some biocides, all described in this literary review, allow enterococci to present a longer survival ability in the hospital environment, consequently giving them more opportunities to disseminate in these settings and be responsible for difficult-to-treat infections.

Listeria monocytogenes can survive in food production facilities and can be transmitted via contamination of food during the various stages of food production. This study was conducted to compile the results of three independent previous studies on the genetic diversity of L. monocytogenes in a poultry production company in Spain and to determine the potential virulence and sanitizer resistance of the strains by using both genotype and phenotype analyses. L. monocytogenes was detected at three production stages: a broiler abattoir, a processing plant, and retail stores marketing fresh poultry products from the same company. These three stages spanned three locations in three provinces of Spain. A set of 347 L. monocytogenes isolates representing 39 subtypes was obtained using pulsed-field gel electrophoresis (PFGE). A total of 28 subtypes (68%) had a fulllength internalin A gene, and two subtypes had a phenotype with low potential for virulence because of a mutation in the prfA gene. A total of 32 subtypes (82%) were classified as benzalkonium chloride resistant (BAC-R) and contained the resistance determinant bcrABC (21 subtypes, 54%) or the resistance gene qacH (11 subtypes, 28%). A total of 13 persistent BAC-R subtypes (minimum of 3 months between the first and last sample from with the isolate was recovered) were identified at the abattoir and processing plant. The three production stages shared a unique subtype (PFGE type 1), which had the mutation in the prfA gene and the bcrABC resistance determinant. Whole genome sequencing revealed this subtype to be sequence type 31. Limited genetic diversity was noted in the isolates studied, including some subtypes that were persistent in the environment of the investigated facilities. Given the high prevalence of BAC-R subtypes, these results support the association between resistance to biocides and persistence of L. monocytogenes. [ABSTRACT FROM AUTHOR]

Listeria monocytogenes is the etiological agent of listeriosis, a foodborne illness associated with high hospitalizations and mortality rates. This bacterium can persist in food associated environments for years with isolates being increasingly linked to outbreaks. This review presents a discussion of genomes of Listeria monocytogenes which are commonly regarded as persisters within food production environments, as well as genes which are involved in mechanisms aiding this phenotype. Although criteria for the detection of persistence remain undefined, the advent of whole genome sequencing (WGS) and the development of bioinformatic tools have revolutionized the ability to find closely related strains. These advancements will facilitate the identification of mechanisms responsible for persistence among indistinguishable genomes. In turn, this will lead to improved assessments of the importance of biofilm formation, adaptation to stressful conditions and tolerance to sterilizers in relation to the persistence of this bacterium, all of which have been previously associated with this phenotype. Despite much research being published around the topic of persistence, more insights are required to further elucidate the nature of true persistence and its implications for public health.

Klebsiella pneumoniae is among the World Health Organization's list of priority pathogens, notorious for its role in causing healthcare-associated infections and neonatal sepsis globally. Containment of K. pneumoniae transmission depends on the continued effectiveness of antimicrobials and of biocides used for topical antisepsis and surface disinfection. Klebsiella pneumoniae is known to disseminate antimicrobial resistance (AMR) through a large auxiliary genome made up of plasmids, transposons and integrons, enabling it to evade antimicrobial killing through the use of efflux systems and biofilm development. Because AMR mechanisms are also known to impart tolerance to biocides, AMR is frequently linked with biocide resistance (BR). However, despite extensive research on AMR, there is a gap in knowledge about BR and the extent to which AMR and BR mechanisms overlap remains debatable. The aim of this paper is to review and summarise the current knowledge on the determinants of BR in K. pneumoniae and highlight content areas that require further inquiry., (© 2024 The Authors.)

The main goal of this work was to approach food industry conditions in the comparison of the susceptibility of biofilms of Listeria monocytogenes to the biocides benzalkonium chloride (BAC) and peracetic acid (PAA). Twelve isolates of L. monocytogenes , including nine well characterized BAC resistant strains were used. Biofilms were produced on stainless steel coupons (SSC), at 11 °C (refrigeration temperature) or at 25 °C (room temperature), in culture media simulating clean (nutrient limiting) or soiled (nutrient rich) growth conditions. Neither different nutrient availability nor growth temperature showed significant effect (p >.05) on biofilm formation. PAA confirmed to be more effective than BAC in biofilm elimination. Biofilms formed under nutritional stress tended to differentiate more the response to BAC of the resistant or sensitive strains, but the resistant or sensitive phenotype of the planktonic cells did not dictate biofilm susceptibility. Unlabelled Image • Resistant or sensitive strain phenotype did not dictate biofilm susceptibility. • Biofilm formation was not affected by nutrient availability nor growth temperature. • Biofilms grown under nutrient scarcity differentiate more their response to BAC. [ABSTRACT FROM AUTHOR]

ABSTRACT The Riverton City dump is Jamaica’s largest solid waste disposal site, but it lacks engineered protection for leachate containment and treatment. Shotgun metagenomics was used to survey the microbial communities in the Riverton City dump leachate and in surface waters of the Duhaney River, an urban waterway abutting the dump. The community within the leachate pond was taxonomically distinct from that found in the surface waters of the Duhaney River. Higher microbial diversity was observed within the dump leachate, with members of the Bacteroidetes, Firmicutes, Gammaproteobacteria, Deltaproteobacteria, and Tenericutes being the most abundant, while the river community was dominated by Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria. The microbial communities exhibit similar functional potential profiles, including chemoorganoheterotrophy as the dominant metabolism, and the potential to degrade aromatic compounds. From reconstruction of metagenome-assembled genomes (MAGs), organisms within both environments are predicted to survive in the presence of multiple antibiotics, antiseptics, biocides, and metals. Strong virulence potential coincided with the most diverse multiple resistance profiles in 1 of 5 leachate MAGs and 5 of 33 river MAGs. Unexpectedly, the microbial resistance profiles were more varied and widespread in the river populations, where we had expected the chemical composition of the leachate to select and enrich for resistance characteristics. This study provides valuable insights into the total functional potential of a landfill leachate microbial community and identifies possible human health hazards within the Duhaney River and Riverton City dump, urban environments with the potential to impact human populations. IMPORTANCE Landfill leachate is a persistent contamination threat for terrestrial waters. Microbial metabolism in landfills transforms contaminants and contributes to greenhouse gas emissions. A better understanding of landfill-associated microbial communities will inform bioremediation of solid waste environments and improve pathogen monitoring. We leveraged shotgun metagenomics to investigate the microbial communities of the Riverton City dump and the adjoining Duhaney River near Kingston City, Jamaica. We identified no overlap between the microbial communities inhabiting the Riverton City dump leachate and the Duhaney River. Both communities are predicted to degrade aromatic compounds, which are ubiquitous environmental pollutants. Adversely, microbes in both environments are predicted to withstand widely used antibiotics, antiseptics, and metal contamination. The absence of evidence for microbial transfer from the leachate to the river is encouraging; however, the Duhaney River contained several organisms with predicted pathogenic lifestyles, indicating that the river represents a human health risk regardless of impact from the dump.

Background: Most studies on biocide resistance and its genetic determinants arise from environmental or food-borne microbial isolates and only a few from clinically relevant isolates., Objectives: This study determines the proportion of biocide resistance against five commonly used biocides and detects biocide resistance genes among MDR bacterial isolates using PCR., Methods: Consecutive MDR isolates (n ​= ​180) were included (30 each of Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, Staphylococcus aureus, and Enterococcus species) from clinical specimens of various inpatient units at JIPMER. The isolates were challenged at 0.5,1 and 2 Macfarland (McF) inoculum with discrete dilutions of disinfectants. The minimum bactericidal concentrations (MBCs) for 70% Ethanol, 1.5% Cresol, 2% Glutaraldehyde, 1% Cetrimide, and 1% Chlorhexidine were determined for the isolates using ATCC reference strains as controls. PCR was performed targeting qac A/B, G; smr; and nfx B genes., Results: For all biocides, MDR isolates had MBCs less than the maximum MBCs of ATCC strains. For MDR K. pneumoniae, A. baumannii, and P. aeruginosa, the highest MBCs of chlorhexidine and cetrimide were ≥75 and ​≥ ​150 ​μg/ml respectively at 0.5 McF inoculum; whereas these organisms grew at higher inoculum (2McF) even at commercially recommended biocidal concentration (1%) corresponding to 750 and 1500 ​μg/ml of chlorhexidine and cetrimide respectively. Meanwhile, the highest MBCs of MDR E. coli were 75 and 150 ​μg/ml for chlorhexidine and cetrimide respectively. Interestingly, the Gram-positive cocci survived the action of up to 35% ethanol. The nfxB and qacG genes were detected in 87% and 6.67% of MDR P. aeruginosa isolates respectively with no biocide resistance genes detected among the other organisms., Conclusions: Biocide dilutions challenged with higher inoculum indicated a narrow margin of effectiveness for certain biocides. Although a significant proportion of clinical MDR isolates of P. aeruginosa harbored biocide resistance genes, this finding had no phenotypic correlation., (Copyright © 2023 Indian Association of Medical Microbiologists. Published by Elsevier B.V. All rights reserved.)

The major contamination sources, biofilm-forming ability and biocide resistance of Staphylococcus aureus in tilapia-processing plants were evaluated. Twenty-five processing control points were analysed twice in two factories, including whole tilapias, frozen fillets, water and food-contact surfaces. No final product was contaminated with S. aureus. However, high concentrations of S. aureus carrying enterotoxin (se) genes were found in several processing points of both factories due to the application of inadequate hygienic and handling procedures, which generate a high risk of cross-contamination of the tilapia fillets with staphylococcal enterotoxins. Nine S. aureus strains were characterized by RAPD-PCR using primers AP-7, ERIC-2 and S. A wide diversity of se gene profiles was detected, most strains being multi-se-carriers. All S. aureus strains showed high biofilm-forming ability on stainless steel and polystyrene. Biofilm-forming ability was correlated with the presence of fliC H7 and the type of origin surface (metallic or plastic). A marked resistance of S. aureus to peracetic acid and sodium hypochlorite was also observed, required doses being higher than those recommended by manufacturers to be eradicated. Case-by-case approaches are thus recommended to determine the sources and degree of contamination present in each factory, which would allow applying precise responses that avoid, or at least reduce, the presence of bacterial pathogens and the emergence of antimicrobial resistance. [ABSTRACT FROM AUTHOR]

Abstract: Although most Brazilian dairy products meet high technological standards, there are quality issues regarding milk production, which may reduce the final product quality. Several microbial species may contaminate milk during manufacture and handling. If antimicrobial usage remains uncontrolled in dairy cattle, the horizontal transfer of antimicrobial resistance genes in foodstuffs may be of particular concern for both food producers and dairy industry. This study focused on the evaluation of putative Gram positive cocci in Minas cheese and of antimicrobial and biocide resistance genes among the isolated bacteria. Representative samples of 7 different industrially trademarked Minas cheeses (n = 35) were processed for selective culture and isolation of Gram positive cocci. All isolated bacteria were identified by DNA sequencing of the 16S rRNA gene. Antimicrobial resistance genes were screened by PCR. Overall, 208 strains were isolated and identified as follows: Enterococcus faecalis (47.6%), Macrococcus caseolyticus (18.3%), Enterococcus faecium (11.5%), Enterococcus caseliflavus (7.7%), Staphylococcus haemolyticus (7.2%), Staphylococcus aureus (4.3%), Staphylococcus epidermidis (2.9%), and Enterococcus hirae (0.5%). The genetic markers mecA (78.0%) and smr (71.4%) were the most prevalent, but others were also detected, such as blaZ (65.2%), msrA (60.9%), msrB (46.6%), linA (54.7%), and aacA‐aphD (47.6%). The occurrence of opportunist pathogenic bacteria harboring antimicrobial resistance markers in the cheese samples are of special concern, since these bacteria are not considered harmful contaminating agents according to the Brazilian sanitary regulations. However, they are potentially pathogenic bacteria and the cheese may be considered a reservoir for antimicrobial resistance genes available for horizontal transfer through the food chain, manufacturing personnel and consumers. [ABSTRACT FROM AUTHOR]

The major contamination sources, serogroups, biofilm-forming ability and biocide resistance of Listeria monocytogenes persistent in tilapia-processing facilities were assessed. Twenty-five processing-control points were examined twice in two factories, including whole tilapias, frozen fillets, water and food-contact surfaces. L. monocytogenes were detected in 4 and 20% of points of Factory A and B respectively, but at low concentrations. Contamination was due to inadequate handling of tilapias in the slaughter room of Factory A and to the application of ineffective sanitizing procedures in Factory B. Seven strains were characterized by RAPD-PCR using primers HLWL85, OPM-01 and DAF4. Genotypic similarity allowed tracing the contamination source of tilapia fillets in Factory B and detecting a prevalent strain in Brazilian tilapia-processing facilities. The serogroup II (including the serotype 1/2c) was the most frequently found, followed by serogroup I (1/2a) and III (1/2b), whereas the serotype 4b was not detected. All strains showed high biofilm-forming ability on stainless steel and polystyrene, but biofilm formation was positively correlated with the type of origin surface. Biofilms were highly resistant to peracetic acid and sodium hypochlorite, being required doses higher than those recommended by manufacturers to be eradicated. Peracetic acid was more effective than sodium hypochlorite, but the use of disinfectants with similar mechanisms of action increases the risk of cross-resistance. Case-by-case approaches are thus recommended to determine the sources and degree of contamination present in each factory, which would allow applying precise responses to control the persistence of bacterial pathogens such as L. monocytogenes. [ABSTRACT FROM AUTHOR]

Bacteria attaching surfaces contacting water reproduce in n the matrix they produce and form a slime layer called biofilm. Increased antimicrobial resistance of bacteria forming biofilm is thought to result from quorum sensing (QS) systems' becoming active. To remove unwanted biofilms from industrial environments, the most commonly used agents are antimicrobial agents called biocides. In the present study, how 61 different bacteria species out of 84 bacteria isolated from the waste water treatment system formed biofilm, and their resistance to Chloramine T trihydrate (Merck) and Pen water BC8120 (Hidrokim), widely used commercial biocides, were investigated. In addition, the relationship between QS systems and biocide resistance of the bacteria identified to be resistant was determined. At the end of the study, it was found that the bacteria treated with ten different concentrations of the biocides (%1, %0,5, %0,2, %0,1, %0,01, %0,001, %0,0001, %0,00001, %0,000001, %0,0000001) for 24 hours, 48 hours and 72 hours developed resistance at various levels depending on the dosage and duration of the application. However, it was also determined that the resistance of numbered 73 to 0,1%, 0,2%, 0,5% and 1% concentrations of Chloramine T trihydrate and the resistance of bacteria numbered 84 to 0,01%, 0,0001%, 0,00001%, 0,2% and 1% concentrations of Chloramine T trihydrate were based on the QS systems. [ABSTRACT FROM AUTHOR]

Background: Biocides and antibiotics are used to eradicate or prevent the growth of microbial species on surfaces (occasionally on catheters), or infected sites, either in combination or sequentially, raising concerns about the development of co-resistance to both antimicrobial types. The effect of such compounds on Salmonella enterica, a major food-borne and zoonotic pathogen, has been analysed in different studies, but only few works evaluated its biological cost, and the overall effects at the genomic and transcriptomic levels associated with diverse phenotypes resulting from biocide exposure, which was the aim of this work. Results: Exposure to triclosan, clorhexidine, benzalkonium, (but not to hypochlorite) resulted in mutants with different phenotypes to a wide range of antimicrobials even unrelated to the selective agent. Most biocide-resistant mutants showed increased susceptibility to compounds acting on the cell wall (β-lactams) or the cell membranes (poly-L-lysine, polymyxin B, colistin or toxic anions). Mutations (SNPs) were found in three intergenic regions and nine genes, which have a role in energy production, amino acids, carbohydrates or lipids metabolism, some of them involved in membrane transport and pathogenicity. Comparative transcriptomics of biocide-resistant mutants showed over-expression of genes encoding efflux pumps (sugE), ribosomal and transcription-related proteins, cold-shock response (cpeE) and enzymes of microaerobic metabolism including those of the phosphotransferase system. Mainly ribosomal, metabolic and pathogenicity-related genes had affected expression in both in vitro-selected biocide mutants and field Salmonella isolates with reduced biocide susceptibility. Conclusions: Multiple pathways can be involved in the adaptation of Salmonella to biocides, mainly related with global stress, or involving metabolic and membrane alterations, and eventually causing "collateral sensitivity" to other antimicrobials. These changes might impact the bacterial-environment interaction, imposing significant bacterial fitness costs which may reduce the chances of fixation and spread of biocide resistant mutants. [ABSTRACT FROM AUTHOR]

Background: Antibacterial biocides and metals can co-select for antibiotic resistance when bacteria harbour resistance or tolerance genes towards both types of compounds. Despite numerous case studies, systematic and quantitative data on co-occurrence of such genes on plasmids and chromosomes is lacking, as is knowledge on environments and bacterial taxa that tend to carry resistance genes to such compounds. This effectively prevents identification of risk scenarios. Therefore, we aimed to identify general patterns for which biocide/metal resistance genes (BMRGs) and antibiotic resistance genes (ARGs) that tend to occur together. We also aimed to quantify co-occurrence of resistance genes in different environments and taxa, and investigate to what extent plasmids carrying both types of genes are conjugative and/or are carrying toxin-antitoxin systems. Results: Co-occurrence patterns of resistance genes were derived from publicly available, fully sequenced bacterial genomes (n = 2522) and plasmids (n = 4582). The only BMRGs commonly co-occurring with ARGs on plasmids were mercury resistance genes and the qacEΔ1 gene that provides low-level resistance to quaternary ammonium compounds. Novel connections between cadmium/zinc and macrolide/aminoglycoside resistance genes were also uncovered. Several clinically important bacterial taxa were particularly prone to carry both BMRGs and ARGs. Bacteria carrying BMRGs more often carried ARGs compared to bacteria without (p < 0.0001). BMRGs were found in 86 % of bacterial genomes, and co-occurred with ARGs in 17 % of the cases. In contrast, co-occurrences of BMRGs and ARGs were rare on plasmids from all external environments (<0.7 %) but more common on those of human and domestic animal origin (5 % and 7 %, respectively). Finally, plasmids with both BMRGs and ARGs were more likely to be conjugative (p < 0.0001) and carry toxin-antitoxin systems (p < 0.0001) than plasmids without resistance genes. Conclusions: This is the first large-scale identification of compounds, taxa and environments of particular concern for co-selection of resistance against antibiotics, biocides and metals. Genetic co-occurrences suggest that plasmids provide limited opportunities for biocides and metals to promote horizontal transfer of antibiotic resistance through co-selection, whereas ample possibilities exist for indirect selection via chromosomal BMRGs. Taken together, the derived patterns improve our understanding of co-selection potential between biocides, metals and antibiotics, and thereby provide guidance for risk-reducing actions. [ABSTRACT FROM AUTHOR]

Staphylococcus epidermidis is a versatile agent, being both a commensal and a nosocomial pathogen usually with an opportunistic role in association with implanted foreign body materials. Pre-operative antiseptic preparation is an important strategy for reducing the risk of complications such as surgical site infection ( SSI). Currently, the most widely used antiseptics are alcohols, quaternary ammonium compounds ( QACs), and the bisbiguanide chlorhexidine. Occurrence of resistance to the latter agent has drawn increasing attention. The aim of this study was to investigate if decreased susceptibility to chlorhexidine among S. epidermidis was present in our setting, a Swedish university hospital. Staphylococcus epidermidis (n = 143), retrospectively collected, were obtained from prosthetic joint infections ( PJI) (n = 61), post-operative infections after cardiac surgery (n = 31), and the skin of the chest after routine disinfection prior to cardiac surgery (n = 27). In addition, 24 commensal isolates were included. Minimum inhibitory concentration ( MIC) of chlorhexidine was determined on Mueller Hinton agar plates supplemented with serial dilutions of chlorhexidine. Five QAC resistance genes, qacA/B, smr, qacH, qacJ, and qacG, were detected using PCR. Decreased susceptibility to chlorhexidine was found in 54% of PJI isolates, 68% of cardiac isolates, 21% of commensal isolates, and 7% of skin isolates from cardiac patients, respectively. The qacA/B gene was present in 62/143 isolates (43%), smr in 8/143 (6%), and qacH in one isolate (0.7%). The qacA/B gene was found in 52% of PJI isolates, 61% of cardiac isolates, 25% of commensal isolates, and 19% of the skin isolates. In conclusion, decreased susceptibility to chlorhexidine, as well as QAC resistance genes, were prevalent among S. epidermidis isolates associated with deep SSIs. [ABSTRACT FROM AUTHOR]