Adaptation to a Commercial Quaternary Ammonium Compound Sanitizer Leads to Cross-Resistance to Select Antibiotics in Listeria monocytogenes Isolated From Fresh Produce Environments

The effective elimination of Listeria monocytogenes through cleaning and sanitation is of great importance to the food processing industry. Specifically in fresh produce operations, the lack of a kill step requires effective cleaning and sanitation to mitigate the risk of cross-contamination from the environment. As facilities rely on sanitizers to control L. monocytogenes, reports of the development of tolerance to sanitizers and other antimicrobials through cross-resistance is of particular concern. We investigated the potential for six L. monocytogenes isolates from fresh produce handling and processing facilities and packinghouses to develop cross-resistance between a commercial sanitizer and antibiotics. Experimental adaptation of isolates belonging to hypervirulent clonal complexes (CC2, CC4, and CC6) to a commercial quaternary ammonium compound sanitizer (cQAC) resulted in elevated minimum inhibitory concentrations (2–3 ppm) and minimum bactericidal concentrations (3–4 ppm). Susceptibility to cQAC was restored for all adapted (qAD) isolates in the presence of reserpine, a known efflux pump inhibitor. Reduced sensitivity to 7/17 tested antibiotics (chloramphenicol, ciprofloxacin, clindamycin, kanamycin, novobiocin, penicillin, and streptomycin) was observed in all tested isolates. qAD isolates remained susceptible to antibiotics commonly used in the treatment of listeriosis (i.e., ampicillin and gentamicin). The whole genome sequencing of qAD strains, followed by comparative genomic analysis, revealed several mutations in fepR, the regulator for FepA fluoroquinolone efflux pump. The results suggest that mutations in fepR play a role in the reduction in antibiotic susceptibility following low level adaptation to cQAC. Further investigation into the cross-resistance mechanisms and pressures leading to the development of this phenomenon among L. monocytogenes isolates recovered from different sources is needed to better understand the likelihood of cross-resistance development in food chain isolates and the implications for the food industry.

In vitro cytotoxicity and virucidal efficacy of potassium hydrogen peroxymonosulfate compared to quaternary ammonium compound under various concentrations, exposure times and temperatures against African swine fever virus

Background and Aim: The selection and proper application of disinfectants are crucial to the prevention of many diseases, so disinfectants must be evaluated before being used for the prevention of African swine fever (ASF). Three disinfectant products belonging to the group of potassium hydrogen peroxymonosulfates, product A and product B, and a quaternary ammonium compound called product C, were examined in vitro for host cell cytotoxicity and the efficacy of ASF virus inactivation. The study parameters included various concentrations, exposure times, temperatures, and degrees of cytotoxicity. Materials and Methods: Three disinfectant products were evaluated for cytotoxicity using primary porcine alveolar macrophage (PAM) cells at dilutions from 1:200 to 1:51,200. Disinfectants in concentrations of 1:200, 1:400, and 1:800 were prepared, the pH and the virucidal activity were tested. An equal volume of each dilution was mixed with the ASF virus and incubated at room temperature (20°C) or on ice (4°C) for 1 min, 5 min, or 30 min. Hemadsorption (HAD) or rosette formation was observed using an inverted microscope for 5 days after inoculation, and the virus titer was calculated as HAD50/mL. Each treatment and virus control were tested in triplicate, and the titers were reported as means and standard deviations. The reduction factor was used to measure inactivation. Results: Products A, B, and C at 1:400, 1:800, and 1:25,600 of dilution, respectively, did not show significant cytotoxic effects on PAM cells. Products A and B could inactivate ASF virus at 1:200 dilution within 5 min after exposure at 4°C. However, at 20°C, the exposure time had to be extended to 30 min to inactivate the virus. Product C could inactivate the virus at 1:400 dilution within 5 min under both temperature conditions, whereas at 1:800 dilution, the exposure time had to be extended to 30 min to completely inactivate the virus at 20°C. Conclusion: All disinfectants could inactivate ASF virus in various concentrations, under appropriate exposure times and reaction temperatures, and there was no evidence of host cell cytotoxicity. For the control of ASF in pig farms, the appropriate concentration, ambient temperature, and contact time of these disinfectants should be taken into account.

784. Efficiency of Quaternary Ammonium Disinfectants against SARS-CoV-2 and Coronavirus 229E

Background The recent pandemic of CoVid19 has increased our need to assess the impact of disinfectants on the inactivation of human coronaviruses. The goals of this study were 1) quantify the disinfection of SARS-CoV-2 and human coronavirus 229 inactivations by various quaternary ammonium formulations, and 2) demonstrate the impact of disinfectants on preventing fomite-to-finger transfer of coronaviruses. Methods We compared the inactivation of both SARS-Covid -2 and coronavirus 229E suspended in 5% fetal calf sera and dried on both metal and plastic surfaces. In addition, studies were conducted with a silinated quaternary ammonium compound that left a residual on the surface. Studies were also conducted on the finger transfer of coronavirus from various surfaces. The virus was allowed to dry on the surface for 30 minutes, then a transfer was conducted by placing the finger pad directly onto the contaminated surface. The finger was tested for the virus. The study was then repeated with virus-contaminated porcelain surfaces that were sprayed with a quaternary product or placed on a surface with a quaternary ammonium compound that left a residual. Results Several readily available quaternary ammonium formulations were evaluated and proved to be effective with greater than a 99.9% reduction in titer after drying on both metal and plastic surfaces. In addition, a silinated quaternary ammonium compound that left a residual on the surface was capable of inactivating SARS-CoV-2 for at least seven days after application. Studies on the finger transfer of coronavirus from various surfaces showed that the amount of virus transfer to the finger varied from 0.46 to 49.0% depending upon the surface. Little or no virus transfer occurred from treated surfaces compared to the untreated controls. In addition, coronavirus 229E appears to be a good model for use in disinfection assessments for SARS-CoV-2. Conclusion Our results demonstrate that various quaternary ammonium disinfectant formulations are effective against human coronaviruses. Finger transfer tests showed that transmission of coronavirus from surfaces can be prevented, reducing the risk of fomite transmission. Coronavirus 229E appears to be a good model for use in disinfection assessments for SARS-CoV-2. Disclosures Charles P. Gerba, Ph.D., Allied Biosciences (Grant/Research Support)Behr (Grant/Research Support)Corning Inc. (Grant/Research Support)PPG (Grant/Research Support)Procter and Gamble (Other Financial or Material Support, donation)Rickett and Coleman (Grant/Research Support)

STUDY OF PHARMACO-TOXICOLOGICAL PROPERTIES OF A QUATERNARY AMMONIUM COMPOUND THAT IS PART OF A DISINFECTANT

The study of the toxicological properties of a new disinfecting quaternary ammonium compound was carried out on laboratory animals of the vivarium of the Kazan State Medical University. Toxicological properties were evaluated on white mice, white rats, guinea pigs and rabbits by forming experimental and control groups. The value of acute oral toxicity, the indicator of the average lethal dose was experimentally determined, allergic properties were determined and the local irritant effect was evaluated. During the tests, it was found that the new quaternary ammonium compound belongs to the third hazard class.

Immobilizing Bactericides on Dental Resins via Electron Beam Irradiation

Polymerizable bactericides, such as quaternary ammonium compound–based monomers, have been intensively studied as candidates for immobilizing antibacterial components on dental resin. However, they predominantly exhibit a bacteriostatic behavior, rather than bactericidal, as the immobilized components are left with insufficient molecular movement to disrupt the bacterial surface structure through contact-mediated action. In this study, we developed a novel strategy to increase the density of the immobilized bactericide and enhance its antibacterial/antibiofilm properties by combining a surface-grafting technique with electron beam irradiation. A solution of the quaternary ammonium compound–based monomer, 12-methacryloyloxydodecylpyridinium bromide (MDPB), was coated on polymethyl methacrylate (PMMA) resin specimens at the concentrations of 30, 50, and 80 wt%. The coated resins were subsequently exposed to 10 MeV of electron beam irradiation at 50 and 100 kGy, followed by thermal stabilization at 60 °C. The antibacterial effect was evaluated by inoculating a Streptococcus mutans suspension on the coated PMMA resin samples, which exhibited bactericidal effects even after 28 d of aging ( P < 0.05, Tukey’s honestly significant difference test). Transmission electron microscopy and bacteriolytic activity evaluation revealed that the S. mutans cells had sustained membrane depolarization. Furthermore, the antibiofilm effects against S. mutans and bacteria collected from human saliva were assessed. The thickness and the percentage of membrane-intact cells of the S. mutans and multispecies biofilms formed on the MDPB-immobilized surfaces were significantly lower than the uncoated PMMA specimens, even after 28-d aging ( P < 0.05, Tukey’s honestly significant difference test). Thus, the immobilization of antibacterial MDPB via electron beam irradiation induced rapid membrane depolarization, increasing membrane permeability and eventually causing cell death. Our strategy substantially enhances the antibacterial properties of the resinous materials and inhibits biofilm formation, therefore demonstrating significant potential for preventing infectious diseases in the oral environment.

Characterization of proteobacterial plasmid integron-encoded qac efflux pump sequence diversity and quaternary ammonium compound antiseptic selection in E. coli grown planktonically and as biofilms

Qac efflux pumps from proteobacterial multidrug-resistant plasmids are integron-encoded and confer resistance to quaternary ammonium compound (QAC) antiseptics, however, many are uncharacterized and misannotated. A survey of >2000 plasmid-encoded qac identified 37 unique qac sequences that correspond to one of five representative motifs: QacE, QacEΔ1, QacF/L, QacH/I, and QacG. Antimicrobial susceptibility testing of each cloned qac member in Escherichia coli , highlighted distinctive antiseptic susceptibility patterns that were most prominent when cells grew as biofilms.

Evaluation of sensitivity to chemical disinfectants of mycobacterium tuberculosis strains isolated from HIV-infected individuals and patients with monotuberculosis infection compared to the reference strain

Objective. To study the sensitivity to chemical disinfectants against Mycobacterium tuberculosis isolated from HIV-infected individuals and patients with monotuberculosis in comparison to the reference strain of Mycobacterium terrae. Materials and methods. The sensitivity of 12 M. tuberculosis strains isolated from patients with HIV-associated tuberculosis (n = 6) and monotuberculosis (n = 6) to chlorine-containing and quaternary ammonium compound (QAC) disinfectants was studied. The M. terrae strain was used as a reference. Seedings of microbial suspensions treated with disinfectants on the culture medium Levenshtein-Jensen were carried out and were monitored for 3 months. Results. One, two, three months following the treatment of microorganisms with QAC-containing disinfectant in the absence of growth of the control strain, there was noted growth of M. tuberculosis, particularly intense in the strains isolated from HIV-infected persons. Conclusions. Thus, the resistance of tuberculosis pathogens to disinfectants can be acquired. M. tuberculosis strains isolated from patients with HIV-associated tuberculosis are detected to be more resistant to disinfectants than the strains isolated from patients with monotubercular infection.

Cetrimonium bromide promotes lipid clearance via TFEB-mediated autophagy-lysosome activation in hepatic cells

Autophagy plays a key role in the metabolism of macromolecules by the lysosomal degradative machinery. The transcription factor EB (TFEB) regulates autophagosome biogenesis and lysosome function, and promoting TFEB activity has emerged as a potential strategy for the treatment of metabolic disorders. Here, we describe that cetrimonium bromide (CTAB), a quaternary ammonium compound, promotes autophagy and lysosomal biogenesis by inducing the nuclear translocation of TFEB in hepatic cells. shRNA-mediated TFEB knockdown inhibits CTAB-induced autophagy and lysosomal biogenesis. Mechanistically, CTAB treatment inhibits the Akt-mTORC1 signaling pathway. Moreover, CTAB treatment markedly promotes lipid metabolism in both palmitate and oleate-treated HepG2 cells, and this promotion was attenuated by the depletion of TFEB. Altogether, our results indicate that CTAB activates the autophagy-lysosome pathway by inducing the nuclear translocation of TFEB via the inhibition of mTORC1 signaling. These results deepen our understanding of TFEB function and provide new insights into CTAB-mediated lipid metabolism.