Role In Antimicrobial Resistance Research Articles

Deciphering the Coevolutionary Dynamics of L2 β-Lactamases via Deep Learning.

L2 β-lactamases, serine-based class A β-lactamases expressed by Stenotrophomonas maltophilia, play a pivotal role in antimicrobial resistance (AMR). However, limited studies have been conducted on these important enzymes. To understand the coevolutionary dynamics of L2 β-lactamase, innovative computational methodologies, including adaptive sampling molecular dynamics simulations, and deep learning methods (convolutional variational autoencoders and BindSiteS-CNN) explored conformational changes and correlations within the L2 β-lactamase family together with other representative class A enzymes including SME-1 and KPC-2. This work also investigated the potential role of hydrophobic nodes and binding site residues in facilitating the functional mechanisms. The convergence of analytical approaches utilized in this effort yielded comprehensive insights into the dynamic behavior of the β-lactamases, specifically from an evolutionary standpoint. In addition, this analysis presents a promising approach for understanding how the class A β-lactamases evolve in response to environmental pressure and establishes a theoretical foundation for forthcoming endeavors in drug development aimed at combating AMR.

P-type ATPase zinc transporter Rv3270 of Mycobacterium tuberculosis enhances multi-drug efflux activity.

Metal homeostasis is maintained by the uptake, storage and efflux of metal ions that are necessary for the survival of the bacterium. Homeostasis is mostly regulated by a group of transporters categorized as ABC transporters and P-type ATPases. On the other hand, efflux pumps often play a role in drug-metal cross-resistance. Here, with the help of antibiotic sensitivity, antibiotic/dye accumulation and semi-quantitative biofilm formation assessments we report the ability of Rv3270, a P-type ATPase known for its role in combating Mn2+ and Zn2+ metal ion toxicity in Mycobacterium tuberculosis, in influencing the extrusion of multiple structurally unrelated drugs and enhancing the biofilm formation of Escherichia coli and Mycobacterium smegmatis. Overexpression of Rv3270 increased the tolerance of host cells to norfloxacin, ofloxacin, sparfloxacin, ampicillin, oxacillin, amikacin and isoniazid. A significantly lower accumulation of norfloxacin, ethidium bromide, bocillin FL and levofloxacin in cells harbouring Rv3270 as compared to host cells indicated its role in enhancing efflux activity. Although over-expression of Rv3270 did not alter the susceptibility levels of levofloxacin, rifampicin and apramycin, the presence of a sub-inhibitory concentration of Zn2+ resulted in low-level tolerance towards these drugs. Of note, the expression of Rv3270 enhanced the biofilm-forming ability of the host cells strengthening its role in antimicrobial resistance. Therefore, the study indicated that the over-expression of Rv3270 enhances the drug efflux activity of the micro-organism where zinc might facilitate drug-metal cross-resistance for some antibiotics.

Colistin the last resort drug in 21st century antibiotics to combat Multidrug resistance superbugs

Polymyxin' E' (Colistin) is considered the last resort therapy against Multidrug resistance (MDR) bacteria, mainly Klebsiella peumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Escherichia coli and play a critical role in causing life-threatening infection, and their prevalence is increasing as a big concern globally. Apart from immunological adaptation, chromosomal mutations and plasmid-mediated genes are mostly associated with this resistance at the molecular level. Therefore, the current review extensively focused on Colistin as a drug in 21st-century antibiotics, the activities spectrum with diverse resistance mechanisms of bacteria against Colistin, and emerging approaches of Colistin from discovery to tackling MDR. In the study, we got to know about the challenges and new developments with old weapons like phage therapy as well as new approaches like Phage display and drug repurposing, in addition to the chromosomal and plasmid-mediated genes that play a role in antimicrobial resistance (AMR). The present study would provide insight into the prognostic aspect of combating MDR.

Efflux Pumps In Antimicrobial Resistance: Mechanism, Regulation And Therapeutic Implications

Efflux pumps play a crucial role in antimicrobial resistance, enabling bacteria to extrude a wide range of antibiotics and other antimicrobial compounds, thereby reducing their intracellular concentration and rendering them ineffective. Understanding the mechanisms underlying efflux pump-mediated resistance is essential for the development of effective strategies to combat this growing threat. This review paper provides an overview of the various efflux pump systems, their regulation mechanisms, and their impact on antimicrobial resistance. Additionally, we discuss the potential therapeutic interventions that target efflux pumps to restore the efficacy of antimicrobial agents.

Bacteria exposed to antiviral drugs develop antibiotic cross-resistance and unique resistance profiles

Antiviral drugs are used globally as treatment and prophylaxis for long-term and acute viral infections. Even though antivirals also have been shown to have off-target effects on bacterial growth, the potential contributions of antivirals to antimicrobial resistance remains unknown. Herein we explored the ability of different classes of antiviral drugs to induce antimicrobial resistance. Our results establish the previously unrecognized capacity of antivirals to broadly alter the phenotypic antimicrobial resistance profiles of both gram-negative and gram-positive bacteria Escherichia coli and Bacillus cereus. Bacteria exposed to antivirals including zidovudine, dolutegravir and raltegravir developed cross-resistance to commonly used antibiotics including trimethoprim, tetracycline, clarithromycin, erythromycin, and amoxicillin. Whole genome sequencing of antiviral-resistant E. coli isolates revealed numerous unique single base pair mutations, as well as multi-base pair insertions and deletions, in genes with known and suspected roles in antimicrobial resistance including those coding for multidrug efflux pumps, carbohydrate transport, and cellular metabolism. The observed phenotypic changes coupled with genotypic results indicate that bacteria exposed to antiviral drugs with antibacterial properties in vitro can develop multiple resistance mutations that confer cross-resistance to antibiotics. Our findings underscore the potential contribution of wide scale usage of antiviral drugs to the development and spread of antimicrobial resistance in humans and the environment.

Detection of resistance and virulence plasmids in Campylobacter coli and Campylobacter jejuni isolated from North Carolina food animal production, 2018–2019

Campylobacter remains the leading cause of bacterial foodborne illness in the U.S. and worldwide. Campylobacter plasmids may play a significant role in antimicrobial resistance (AMR) and virulence factor distribution, and potentially drive rapid adaptation. C. coli (n = 345) and C. jejuni (n = 199) isolates collected from live cattle, swine, turkey, and chickens, poultry carcasses at production, and retail meat in N.C. were analyzed to determine plasmid prevalence, extrachromosomal virulence and AMR genes, and the phylogeny of assembled plasmids. Putative plasmids ranging from <2 kb to 237kb were identified with virulence factors present in 66.1% (228/345) C. coli and 88.4% (176/199) C. jejuni plasmids (promoting adherence, invasion, exotoxin production, immune modulation, chemotaxis, mobility, and the type IV secretion system). AMR genes were identified in 21.2% (73/345) C. coli and 28.1% C. jejuni plasmids (conferring resistance to tetracyclines, aminoglycosides, beta-lactams, nucleosides, and lincosamides). Megaplasmids (>100 kb) were present in 25.7% (140/544) of the isolates and carried genes previously recognized to be involved with interspecies recombination. Our study highlights the extensive distribution and diversity of Campylobacter plasmids in food animal production and their role in the dissemination of biomedically important genes. Characterizing Campylobacter plasmids within the food animal production niche is important to understanding the epidemiology of potential emerging strains.

Role of Antimicrobial Resistance in Outcomes of Acute Endophthalmitis.

This study explores local trends in antimicrobial resistance and its influence on long-term visual outcomes following treatment with broad-spectrum empiric intravitreal antibiotics. All patients undergoing intraocular sampling for endophthalmitis from Auckland between January 2006-May 2023 were included. The impact of antimicrobial resistance on the final visual outcome was analysed using logistic regression models. 389 cases of endophthalmitis were included, and 207 eyes (53.2%) were culture positive. When tested, all Gram-positive microorganisms were fully susceptible to Vancomycin, and all Gram-negative microorganisms demonstrated full or intermediate susceptibility to Ceftazidime. Resistance to at least one antimicrobial agent was present in 89 culture results (43.0%), and multidrug resistance (resistant to ≥3 antimicrobials) in 23 results (11.1%). No increase in resistance was observed over time. The primary procedure was a tap and inject in 251 eyes (64.5%), and early vitrectomy was performed in 196 eyes (50.3%). Severe vision loss (≤20/200) occurred in 167 eyes (42.9%). Antimicrobial resistance was associated with an increased risk of retinal detachment (OR 2.455 p = 0.048) but not vision loss (p = 0.288). High sensitivity to Vancomycin and Ceftazidime was present in our population, reinforcing their role as first-line empiric treatments. Resistant microorganisms were associated with an increased risk of retinal detachment but no alteration in final visual outcome.

Hospital and urban wastewaters shape the matrix and active resistome of environmental biofilms

Understanding the dynamics of antibiotic resistance gene (ARG) transfer and dissemination in natural environments remains challenging. Biofilms play a crucial role in bacterial survival and antimicrobial resistance (AMR) dissemination in natural environments, particularly in aquatic systems. This study focused on hospital and urban wastewater (WW) biofilms to investigate the potential for ARG dissemination through mobile genetic elements (MGEs). The analysis included assessing the biofilm extracellular polymeric substances (EPS), microbiota composition as well as metatranscriptomic profiling of the resistome and mobilome. We produced both in vitro and in situ biofilms and performed phenotypic and genomic analyses. In the in vitro setup, untreated urban and hospital WW was used to establish biofilm reactors, with ciprofloxacin added as a selective agent at minimal selective concentration. In the in situ setup, biofilms were developed directly in hospital and urban WW pipes.We first showed that a) the composition of EPS differed depending on the growth environment (in situ and in vitro) and the sampling origin (hospital vs urban WW) and that b) ciprofloxacin impacted the composition of the EPS. The metatranscriptomic approach showed that a) expression of several ARGs and MGEs increased upon adding ciprofloxacin for biofilms from hospital WW only and b) that the abundance and type of plasmids that carried individual or multiple ARGs varied depending on the WW origins of the biofilms. When the same plasmids were present in both, urban and hospital WW biofilms, they carried different ARGs. We showed that hospital and urban wastewaters shaped the structure and active resistome of environmental biofilms, and we confirmed that hospital WW is an important hot spot for the dissemination and selection of antimicrobial resistance. Our study provides a comprehensive assessment of WW biofilms as crucial hotspots for ARG transfer. Hospital WW biofilms exhibited distinct characteristics, including higher eDNA abundance and expression levels of ARGs and MGEs, highlighting their role in antimicrobial resistance dissemination. These findings emphasize the importance of understanding the structural, ecological, functional, and genetic organization of biofilms in anthropized environments and their contribution to antibiotic resistance dynamics.

Evaluation of the Effect of Plectranthus amboinicus L. Leaf Extracts on the Bacterial Antioxidant System and Cell Membrane Integrity of Pseudomonas aeruginosa PA01 and Staphylococcus aureus NCTC8325.

Plectranthus amboinicus (Indian borage) has been extensively studied for its medicinal properties, which can be exploited to develop new antimicrobial therapeutics. The current study investigated the effect of Plectranthus amboinicus leaf extracts on the catalase activity, reactive oxygen species, lipid peroxidation, cytoplasmic membrane permeability, and efflux pump activity in S. aureus NCTC8325 and P. aeruginosa PA01. As the enzyme catalase protects bacteria against oxidative stress, disruption of its activity creates an imbalance in reactive oxygen species (ROS) levels, which subsequently oxidizes lipid chains, leading to lipid peroxidation. In addition, bacterial cell membranes are a potential target for new antibacterial agents, as efflux pump systems play a crucial role in antimicrobial resistance. Upon exposure of the microorganisms to Indian borage leaf extracts, the observed catalase activity decreased by 60% and 20% in P. aeruginosa and S. aureus, respectively. The generation of ROS can cause oxidation reactions to occur within the polyunsaturated fatty acids of the lipid membranes and induce lipid peroxidation. To investigate these phenomena, the increase in ROS activity in P. aeruginosa and S. aureus was studied using H2DCFDA, which is oxidized to 2',7'-dichlorofluorescein (DCF) by ROS. Furthermore, the concentration of lipid peroxidation product (malondialdehyde) was assessed using the Thiobarbituric acid assay and was shown to increase by 42.4% and 42.5% in P. aeruginosa and S. aureus, respectively. The effect of the extracts on the cell membrane permeability was monitored using diSC3-5 dye and it was observed that the cell membrane permeability of P. aeruginosa increased by 58% and of S. aureus by 83%. The effect on efflux pump activity was investigated using Rhodamine-6-uptake assay, which displayed a decrease in efflux activity of 25.5% in P. aeruginosa and 24.2% in S. aureus after treatment with the extracts. This combination of different methods to study various bacterial virulence factors provides a more robust, mechanistic understanding of the effect of P. amboinicus extracts on P. aeruginosa and S. aureus. This study thus represents the first report of the assessment of the effect of Indian borage leaf extracts on bacterial antioxidant systems and bacterial cell membranes, and can facilitate the future development of bacterial resistance modifying agents derived from P. amboinicus.

Antimicrobial resistance and mechanisms of epigenetic regulation.

The rampant use of antibiotics in animal husbandry, farming and clinical disease treatment has led to a significant issue with pathogen resistance worldwide over the past decades. The classical mechanisms of resistance typically investigate antimicrobial resistance resulting from natural resistance, mutation, gene transfer and other processes. However, the emergence and development of bacterial resistance cannot be fully explained from a genetic and biochemical standpoint. Evolution necessitates phenotypic variation, selection, and inheritance. There are indications that epigenetic modifications also play a role in antimicrobial resistance. This review will specifically focus on the effects of DNA modification, histone modification, rRNA methylation and the regulation of non-coding RNAs expression on antimicrobial resistance. In particular, we highlight critical work that how DNA methyltransferases and non-coding RNAs act as transcriptional regulators that allow bacteria to rapidly adapt to environmental changes and control their gene expressions to resist antibiotic stress. Additionally, it will delve into how Nucleolar-associated proteins in bacteria perform histone functions akin to eukaryotes. Epigenetics, a non-classical regulatory mechanism of bacterial resistance, may offer new avenues for antibiotic target selection and the development of novel antibiotics.

Quaternary Ammonium Compounds: A Chemical Class of Emerging Concern.

Quaternary ammonium compounds (QACs), a large class of chemicals that includes high production volume substances, have been used for decades as antimicrobials, preservatives, and antistatic agents and for other functions in cleaning, disinfecting, personal care products, and durable consumer goods. QAC use has accelerated in response to the COVID-19 pandemic and the banning of 19 antimicrobials from several personal care products by the US Food and Drug Administration in 2016. Studies conducted before and after the onset of the pandemic indicate increased human exposure to QACs. Environmental releases of these chemicals have also increased. Emerging information on adverse environmental and human health impacts of QACs is motivating a reconsideration of the risks and benefits across the life cycle of their production, use, and disposal. This work presents a critical review of the literature and scientific perspective developed by a multidisciplinary, multi-institutional team of authors from academia, governmental, and nonprofit organizations. The review evaluates currently available information on the ecological and human health profile of QACs and identifies multiple areas of potential concern. Adverse ecological effects include acute and chronic toxicity to susceptible aquatic organisms, with concentrations of some QACs approaching levels of concern. Suspected or known adverse health outcomes include dermal and respiratory effects, developmental and reproductive toxicity, disruption of metabolic function such as lipid homeostasis, and impairment of mitochondrial function. QACs' role in antimicrobial resistance has also been demonstrated. In the US regulatory system, how a QAC is managed depends on how it is used, for example in pesticides or personal care products. This can result in the same QACs receiving different degrees of scrutiny depending on the use and the agency regulating it. Further, the US Environmental Protection Agency's current method of grouping QACs based on structure, first proposed in 1988, is insufficient to address the wide range of QAC chemistries, potential toxicities, and exposure scenarios. Consequently, exposures to common mixtures of QACs and from multiple sources remain largely unassessed. Some restrictions on the use of QACs have been implemented in the US and elsewhere, primarily focused on personal care products. Assessing the risks posed by QACs is hampered by their vast structural diversity and a lack of quantitative data on exposure and toxicity for the majority of these compounds. This review identifies important data gaps and provides research and policy recommendations for preserving the utility of QAC chemistries while also seeking to limit adverse environmental and human health effects.

The Capsule Increases Susceptibility to Last-Resort Polymyxins, but Not to Other Antibiotics, in Klebsiella pneumoniae.

The extracellular capsule is a virulence factor present in many facultative pathogens, but its role in antimicrobial resistance remains controversial. To shed light on this debate, we tested six antibiotics on four Klebsiella pneumoniae species complex strains. Noncapsulated strains exhibited increased tolerance to polymyxins, but not to other antibiotics, as measured using the MIC. Our results urge caution on the use of therapeutic agents that target the capsule and may result in selection for its inactivation.

A Systematic Quantitative Determination of the Antimicrobial Efficacy of Grape Seed Extract against Foodborne Bacterial Pathogens

Concerns regarding the role of antimicrobial resistance (AMR) in disease outbreaks are growing due to the excessive use of antibiotics. Moreover, consumers are demanding food products that are minimally processed and produced in a sustainable way, without the use of chemical preservatives or antibiotics. Grape seed extract (GSE) is isolated from wine industry waste and is an interesting source of natural antimicrobials, especially when aiming to increase sustainable processing. The aim of this study was to obtain a systematic understanding of the microbial inactivation efficacy/potential of GSE against Listeria monocytogenes (Gram-positive), Escherichia coli and Salmonella Typhimurium (Gram-negative) in an in vitro model system. More specifically, for L. monocytogenes, the effects of the initial inoculum concentration, bacterial growth phase and absence of the environmental stress response regulon (SigB) on the GSE microbial inactivation potential were investigated. In general, GSE was found to be highly effective at inactivating L. monocytogenes, with higher inactivation achieved for higher GSE concentrations and lower initial inoculum levels. Generally, stationary phase cells were more resistant/tolerant to GSE as compared to exponential phase cells (for the same inoculum level). Additionally, SigB appears to play an important role in the resistance of L. monocytogenes to GSE. The Gram-negative bacteria under study (E. coli and S. Typhimurium) were less susceptible to GSE as compared to L. monocytogenes. Our findings provide a quantitative and mechanistic understanding of the impact of GSE on the microbial dynamics of foodborne pathogens, assisting in the more systematic design of natural antimicrobial-based strategies for sustainable food safety.

Biofilm formation in vitro by Leptospira interrogans strains isolated from naturally infected dogs and their role in antimicrobial resistance

Leptospira interrogans is a biofilm-forming pathogen, however, there are few data involving Brazilian strains isolated from dogs and their antimicrobial sensitivity in planktonic and biofilm forms. The potential for biofilm formation and antimicrobial resistance in naturally infected dogs is a fundamental approach towards disease epidemiology and the establishment of consistent prophylaxis and control measures. The objective of this study was to evaluate in vitro biofilm formation of a reference strain (L. interrogans, sv. Copenhageni L1 130 - L20) and of L. interrogans isolated from dogs (C20, C29, C51, C82), with subsequent evaluation of antimicrobial susceptibility in planktonic and biofilm forms. The semi quantification of biofilm production revealed a dynamic process of development over time, with mature biofilm formation early on the seventh day of incubation. All strains were efficient for in vitro biofilm formation and, in this form, they were considerably more resistant compared to their planktonic form, with MIC90 of 1600 μg/mL for amoxicillin, 800 μg/mL for ampicillin, and >1600 μg/mL for doxycycline and ciprofloxacin. The strains studies were isolated on naturally infected dogs that might act as reservoirs and sentinels for human infections. The potential to antimicrobial resistance together with the close relation between dogs and humans indicates the need for greater actions on disease control and surveillance. Moreover, biofilm formation may contribute to the persistence of Leptospira interrogans in the host and these animals can act as chronic carriers, disseminating the agent in the environment.

Functional metagenomic libraries generated from anthropogenically impacted environments reveal importance of metabolic genes in biocide and antibiotic resistance

Anthropogenic activities result in the release of antimicrobial resistant bacteria and a cocktail of antimicrobial compounds into the environment that may directly select or indirectly co-select for antimicrobial resistance (AMR). Many studies use metagenome sequencing or qPCR-based approaches to study the environmental resistome but these methods are limited by a priori knowledge. In this study, a functional metagenomic approach was used to explore biocide resistance mechanisms in two contaminated environments and a pristine site, and to identify whether potentially novel genes conferring biocide resistance also conferred resistance or reduced susceptibility to antibiotics. Resistance was predominately mediated through novel mechanisms exclusive of the well-known qac efflux genes. UDP-galactose 4-epimerase (galE) –like genes were identified in both contaminated environments and were shown to confer cross-resistance to biocides and clinically important antibiotics for the first time (to our knowledge), compared to knockout mutants. GalE -like genes were also co-located with transposons, suggesting mobilisation potential. These results show that housekeeping genes may play a significant yet underappreciated role in AMR in environmental microbiomes.

The Role of Outer Membrane Proteins in UPEC Antimicrobial Resistance: A Systematic Review.

Uropathogenic Escherichia coli (UPEC) are one of the most common agents of urinary tract infection. In the last decade, several UPEC strains have acquired antibiotic resistance mechanisms and some have become resistant to all classes of antibiotics. UPEC outer membrane proteins (OMPs) seem to have a decisive role not only in the processes of invasion and colonization of the bladder mucosa, but also in mechanisms of drug resistance, by which bacteria avoid killing by antimicrobial molecules. This systematic review was performed according to the PRISMA guidelines, aiming to characterize UPEC OMPs and identify their potential role in antimicrobial resistance. The search was limited to studies in English published during the last decade. Twenty-nine studies were included for revision and, among the 76 proteins identified, seven were associated with antibiotic resistance. Indeed, OmpC was associated with β-lactams resistance and OmpF with β-lactams and fluoroquinolone resistance. In turn, TolC, OmpX, YddB, TosA and murein lipoprotein (Lpp) were associated with fluoroquinolones, enrofloxacin, novobiocin, β-lactams and globomycin resistances, respectively. The clinical implications of UPEC resistance to antimicrobial agents in both veterinary and human medicine must propel the implementation of new strategies of administration of antimicrobial agents, while also promoting the development of improved antimicrobials, protective vaccines and specific inhibitors of virulence and resistance factors.

The Role of Antimicrobial Resistance in Refractory and Recurrent Bacterial Vaginosis and Current Recommendations for Treatment.

Bacterial vaginosis (BV), the most common cause of vaginal discharge, is characterized by a shift in the vaginal microbiota from Lactobacillus species dominance to a diverse array of facultative and strict anaerobic bacteria which form a multi-species biofilm on vaginal epithelial cells. The rate of BV recurrence after therapy is high, often >60%. The BV biofilm itself likely contributes to recurrent and refractory disease after treatment by reducing antimicrobial penetration. However, antimicrobial resistance in BV-associated bacteria, including those both within the biofilm and the vaginal canal, may be the result of independent, unrelated bacterial properties. In the absence of new, more potent antimicrobial agents to eradicate drug-resistant pathogenic vaginal microbiota, treatment advances in refractory and recurrent BV have employed new strategies incorporating combination therapy. Such strategies include the use of combination antimicrobial regimens as well as alternative approaches such as probiotics and vaginal fluid transfer. Our current recommendations for the treatment of refractory and recurrent BV are provided.

Membrane Efflux Pumps of Pathogenic Vibrio Species: Role in Antimicrobial Resistance and Virulence

Infectious diseases caused by bacterial species of the Vibrio genus have had considerable significance upon human health for centuries. V. cholerae is the causative microbial agent of cholera, a severe ailment characterized by profuse watery diarrhea, a condition associated with epidemics, and seven great historical pandemics. V. parahaemolyticus causes wound infection and watery diarrhea, while V. vulnificus can cause wound infections and septicemia. Species of the Vibrio genus with resistance to multiple antimicrobials have been a significant health concern for several decades. Mechanisms of antimicrobial resistance machinery in Vibrio spp. include biofilm formation, drug inactivation, target protection, antimicrobial permeability reduction, and active antimicrobial efflux. Integral membrane-bound active antimicrobial efflux pump systems include primary and secondary transporters, members of which belong to closely related protein superfamilies. The RND (resistance-nodulation-division) pumps, the MFS (major facilitator superfamily) transporters, and the ABC superfamily of efflux pumps constitute significant drug transporters for investigation. In this review, we explore these antimicrobial transport systems in the context of Vibrio spp. pathogenesis and virulence.

Mammaliicoccus spp. from German Dairy Farms Exhibit a Wide Range of Antimicrobial Resistance Genes and Non-Wildtype Phenotypes to Several Antibiotic Classes.

Simple SummaryWorldwide, antimicrobial resistance (AMR) is of major concern for human and animal health since infections with multidrug-resistant bacteria are often more challenging and costly. In the family Staphyloccocaceae, the species Staphylococcus aureus in particular was reported to cause severe infections. Although most of the other Staphylococcaceae members were not shown to cause severe illnesses, the transmission of AMR genes to harmful species might take place. Therefore, the monitoring of AMR potential in different environments is of high relevance. Mammaliicocci on dairy farms might represent such an AMR gene reservoir. Thus, in this study, the AMR potential of mammaliicocci isolates from German dairy farms was investigated. Whole-genome sequencing (WGS) of the isolates was conducted to evaluate the phylogenetic relationship of the isolates and analyze AMR genes. In addition, antimicrobial susceptibility testing was performed to compare the AMR genotype with the phenotype. It turned out that mammaliicocci may harbor large numbers of different AMR genes and exhibit phenotypic resistance to various antibiotics. Since some AMR genes are likely located on mobile genetic elements, such as plasmids, AMR gene transmission between members of the Staphylococcaceae family might occur.Mammaliicocci might play a major role in antimicrobial resistance (AMR) gene transmission between organisms of the family Staphylococcaceae, such as the potentially pathogenic species Staphylococcus aureus. The interest of this study was to analyze AMR profiles of mammaliicocci from German dairy farms to evaluate the AMR transmission potential. In total, 65 mammaliicocci isolates from 17 dairy farms with a history of MRSA detection were analyzed for AMR genotypes and phenotypes using whole genome sequencing and antimicrobial susceptibility testing against 19 antibiotics. The various genotypic and phenotypic AMR profiles of mammaliicocci from German dairy farms indicated the simultaneous occurrence of several different strains on the farms. The isolates exhibited a non-wildtype phenotype to penicillin (58/64), cefoxitin (25/64), chloramphenicol (26/64), ciprofloxacin (25/64), clindamycin (49/64), erythromycin (17/64), fusidic acid (61/64), gentamicin (8/64), kanamycin (9/64), linezolid (1/64), mupirocin (4/64), rifampicin (1/64), sulfamethoxazol (1/64), streptomycin (20/64), quinupristin/dalfopristin (26/64), tetracycline (37/64), tiamulin (59/64), and trimethoprim (30/64). Corresponding AMR genes against several antimicrobial classes were detected. Linezolid resistance was associated with the cfr gene in the respective isolate. However, discrepancies between genotypic prediction and phenotypic resistance profiles, such as for fusidic acid and tiamulin, were also observed. In conclusion, mammaliicocci from dairy farms may carry a broad variety of antimicrobial resistance genes and exhibit non-wildtype phenotypes to several antimicrobial classes; therefore, they may represent an important source for horizontal gene transfer of AMR genes to pathogenic Staphylococcaceae.

Community Pharmacists' Knowledge, Attitude, and Nonprescription Dispensing Practices of Antibiotics: An Explorative Study in a Selected City of South India.

This study aimed to assess the community pharmacists' knowledge of antibiotics, their attitude toward antibiotic usage and antibiotic resistance, and their nonprescription dispensing practices of antibiotics. A cross sectional-questionnaire-based study was conducted among 75 community pharmacists practicing in a selected city of South India. Data on their age, years of experience, and educational qualifications were obtained. A modified, 33-items, prevalidated structured questionnaire was used to assess the community pharmacists knowledge, attitude, and nonprescription antibiotic dispensing practices knowledge, attitudes and practices (KAP). The responses obtained were expressed in descriptive statistics. The association between years of experience and their KAP was assessed using Pearson's correlation. Most pharmacists (60%) agreed that antibiotics are used for bacterial infections, and 35% believed that antibiotics could be given for pain and inflammation. Fourty-one percentage of pharmacists agreed that dispensing antibiotics without prescription increases the risk of antibiotic resistance. Seventy-two percentage agreed that they are responsible for taking a prominent role in antimicrobial resistance and infection-control programs in healthcare. Only 46% of pharmacists stated that they always dispensed antibiotics only with a prescription, and 56% dispensed antibiotics for longer than the doctor prescribed. Amoxicillin, metronidazole, and cephalexin were the most commonly dispensed antibiotics without a prescription. The most common reason for dispensing antibiotics without a prescription was the fear of losing customers. The study identified an average KAP interquartile range 1 among community pharmacists, indicating a lack of awareness of antibiotic resistance and dispensing antibiotics without a prescription.