Developing Disinfection Strategies for Controlling Human Norovirus, Sars-Cov-2, and Clostridioides difficile Endospores In Long-term Care Facilities

Long-term care facilities (LTCFs) provide an environment favorable for the transmission of three critical human pathogens: human norovirus (HuNoV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and Clostridioides difficile. Given residents in LTCFs are susceptible to infections due to their advanced ages and compromised immune systems, effective environmental surface disinfection plays a crucial role in controlling the spread of human pathogens within these settings and, therefore, mitigates the risk of infections caused by these pathogens. This dissertation aimed to assess the efficacy of various types of disinfectants against two HuNoV surrogates [feline calicivirus (FCV) and Tulane virus (TuV)], two SARS-CoV-2 surrogates [bovine coronavirus (BCoV) and human coronavirus (HCoV) OC43], and C. difficile endospores. The research encompasses surfaces commonly encountered in healthcare settings and public spaces, with a particular emphasis on the disinfection of these pathogens on soft porous surfaces in LTCFs. First, nine chemical disinfectants on EPA’s List G were selected using four criteria: 1) ready-to-use, 2) nonchlorine-based active ingredient, 3) commercially available, and 4) limited known health risks. Active ingredients of the products included hydrogen peroxide (H2O2), peracetic acid, quaternary ammonium compounds, or alcohols. The efficacy of the products against FCV, TuV and C. difficile spores was first screened using the American Society for Testing and Materials (ASTM) suspension test, and then the carrier test on stainless steel coupons for 1, 5 and 10 min (FCV, TuV) and 10 min (C. difficile spores). On stainless steel carriers, 8 of 9 products could reduce \u3e3 log10 PFU of FCV within 5 min. One most efficacious product containing H2O2 as key active ingredient, reduced \u3e5.1 log10 PFU of FCV and \u3e3.1 log10 TCID50 of TuV after 5 min, and \u3e6.0 log10 CFU of C. difficile endospores after 10 min. Of the five products containing H2O2,no strong correlation (R2=0.25, p=0.03) was observed between disinfection efficacy and H2O2 concentration. The addition of 0.025% ferrous sulfate to 1% H2O2 solution improved efficacy against all FCV, TuV, and C. difficile. Our results confirmed that both product formulation and the active ingredient concentration influence the efficacy. Additionally, TuV proved to be a more conservative surrogate for HuNoV than FCV. Next, this dissertation evaluated the efficacy of chemical disinfectants (products A, B, and C) and steam vapor against HuNoV on nylon carpet with two different backings. Carpet coupons (5×5 cm2) inoculated with a mixture of FCV and TuV were allowed to dry at room temperature under 30-50% relative humidity. The virus-inoculated carpet coupons were applied with three chemical disinfectants or steam vapor for different contact times. The viruses on the treated carpets were subsequently recovered and titrated. Additionally, the color and tensile strength of carpets were assessed after repeated disinfection 30 times to simulate long-term use for 1.5 years. Results suggested the efficacy of disinfectants was affected by the type of carpet backing. For carpet with a water-permeable backing (Color Accent®), products A, B, and C reduced 0.8, 3.1, and 0.9 log10 PFU of FCV, and 0.3, 2.5, and 0.4 log10 TCID50 of TuV after a 30-min contact time, respectively. For the carpet with a waterproof backing (Highlight®), only product B exhibited a substantial reduction of 5.0 log10 PFU for FCV and \u3e3.0 log10 TCID50 for TuV, while products A and C reduced 2.4 and 1.6 log10 PFU of FCV, and 1.2 and 1.2 log10 TCID50 of TuV, respectively. Impressively, steam vapor achieved a ≥5.2 log10 PFU reduction of FCV and \u3e3.2 log10 TCID50 reduction of TuV in just 15 s on both types of carpet. Additionally, two H2O2-based disinfectants significantly impacted the tensile strength of carpet backings after repeated disinfection, with only product B causing cracks on nylon carpet fibers. The overall results highlighted the potential efficacy of steam vapor against HuNoV on both carpet types. Furthermore, one H2O2-based product (product B) exhibited efficacy on waterproof carpet, though the repeated use of disinfectants did affect some properties of the carpet. To understand the transmission potential of SARS-CoV-2 via carpet, the persistence of SARS-CoV-2 surrogates, BCoV and HCoV OC43 on polyethylene terephthalate (PET) and nylon carpet was evaluated using both infectivity and RT-qPCR assays, and the efficacy of steam vapor treatment against BCoV and HCoV OC43 on nylon carpet was determined. After inoculation, the immediate recoveries were only 3.87% of HCoV OC43 from PET and 24.37% from nylon carpet. In contrast, the recovery rates of BCoV were 32.50% from PET and 34.86% from nylon carpet. Following a 1-h incubation at room temperature, BCoV and HCoV OC43 were reduced by 3.6 and \u3e2.8 log10 TCID50 on PET carpet but 0.6 and 1.8 log10 TCID50 on nylon carpet, respectively. The reduction of total genomic RNA of BCoV and HCoV OC43 was also less on nylon carpet than on PET carpet, with first-order decay rates (k values) at 0.86 and 0.27 h-1 for nylon and 1.19 and 0.67 h-1 for PET carpet, respectively. These findings suggest that both surrogates were more stable on nylon than on PET carpet. For carpet disinfection, steam vapor was demonstrated as an effective method for inactivating both surrogates on nylon carpet, by reducing \u3e3.0 log10 TCID50 of BCoV and \u3e3.2 log10 TCID50 of HCoV OC43 within 15 s. In response to the absence of a validated disinfection method against C. difficile endospores on carpet, this dissertation undertook a two-step approach. First, the recovery method for C. difficile endospores from the carpet was optimized by experimenting with three concentrations of Tween-80 and different stomaching durations. Subsequently, the efficacy of three EPA-registered disinfectants (two H2O2-based and one chlorine-based) and steam vapor against C. difficile endospores was evaluated on nylon carpet with two types of backing. The incorporation of 0.2% Tween-80, followed by a 3-min stomaching and subsequent sonication, substantially enhanced the recovery rate of C. difficile endospores, exceeding 60%. Product B was the most efficacious of the three disinfectants tested, which achieved reductions of 5.8 and 4.9 log10 CFU of C. difficile endospores within a 30-min contact time on carpet Highlight® and Color Accent®, respectively. Additionally, steam vapor treatment for 120 s exhibited strong efficacy, reducing \u3e6.0 and 4.9 log10 CFUof C. difficile endospores on carpet Highlight® and Color Accent®, respectively. Additionally, combining a 120-s steam vapor treatment with a less effective product A resulted in a 6.1 log10 CFU reduction of sensitized C. difficile endospores on carpet Highlight®. Furthermore, the efficacy of an aqueous photocatalytic disinfection system, known as photoClO2, was evaluated against HuNoV surrogates and C. difficile endospores on stainless steel and nylon carpet. The process was optimized by utilizing 1% NaClO2 and 10 ppm Eosin Y, which yielded a production rate of 60.64 ppm min-1 of ClO2 within a 4.5×4.5 cm2 area. Subsequently, the efficacy of the system was evaluated against FCV, TuV, and C. difficile endospores on both stainless steel and nylon carpet with two distinct backings under optimal lighting conditions. PhotoClO2 was efficacious in reducing\u3e5 log10 PFU of FCV in 45 min of contact time and \u3e3 log10 TCID50 of TuV in 60 min, but only 1.3 log10 CFU of C. difficile endospores in 120 min. On carpet Highlight®, photoClO2 achieved a 2.9 log10 PFU reduction of FCV and 2.5 log10 TCID50 reduction of TuV in 60 min, respectively, showcasing higher efficacy than carpet Color Accent® (a 1.3 log10 PFU reduction of FCV and 1.1 log10 TCID50 reduction of TuV, respectively). Under indoor lighting conditions, photoClO2 further exhibited its efficacy by inactivating 4.3 log10 PFU of FCV and 1.4 log10 TCID50 of TuV on stainless steel after 120 min. While photoClO2 proved highly effective against HuNoV surrogates, its efficacy against C. difficile endospores was limited regardless of surface material. This dissertation provides some insights into surface disinfection strategies, with a particular focus on soft porous carpet commonly found in LTCFs and various public areas. As the alternatives to bleach, H2O2-based disinfectants exhibited efficacy against HuNoV and C. difficile endospores. The research outcomes emphasize the critical consideration of the active ingredient and product formulation when selecting disinfectants to effectively inactivate pathogens. However, the repeated use of chemical disinfectants may adversely impact carpet properties such as fiber strength and backing integrity. In comparison to chemical disinfectants, steam vapor performed well with short contact time and can be used as a more suitable option for spot treatment or routine disinfection of HuNoV, SARS-CoV-2 and C. difficile endospores for carpet. It should be noted that the efficacy of these disinfection methods can be significantly affected by the carpet backing. This suggests the significance of considering carpet construction alongside materials when selecting disinfection approaches. While photoClO2 has exhibited efficacy against HuNoV surrogates, it\u27s necessary to emphasize that this process involves a slow yet long-lasting reaction. This specific characteristic could prove desirable in preventing the spread of pathogens between disinfection cycles. While this dissertation presented the efficacy of three types of disinfectants (chemical disinfectants, steam vapor, and photoClO2) against HuNoV and SARS-CoV-2 surrogates on carpet surfaces, the efficacy of these disinfectants should be validated using pathogenic HuNoV and SARS-CoV-2 to ensure their practical applicability

An Efficient Process for Co-Production of γ-Aminobutyric Acid and Probiotic \u3ci\u3eBacillus subtilis\u3c/i\u3e Cells

This study was to establish an integrated process for the co-production of γ-aminobutyric acid (GABA) and live probiotics. Six probiotic bacteria were screened and Bacillus subtilis ATCC 6051 showed the highest GABA-producing capacity. The optimal temperature and initial pH value for GABA production in B. subtilis were found to be 30 °C and 8.0, respectively. A variety of carbon and nitrogen sources were tested, and potato starch and peptone were the preferred carbon and nitrogen sources for GABA production, respectively. The concentrations of carbon source, nitrogen source and substrate (sodium L-glutamate) were then optimized using the response surface methodology. The GABA titer and concentration of viable cells of B. subtilis reached 19.74 g/L and 6.0 × 108 cfu/mL at 120 h. The GABA titer represents the highest production of GABA in B. subtilis. This work thus demonstrates a highly efficient co-production process for GABA and probiotic B. subtilis cells

Macrophage M1 polarization mediated via the IL-6/STAT3 pathway contributes to apical periodontitis induced by Porphyromonas gingivalis

Objective: To investigate the involvement of IL-6/STAT3 signaling pathway activation in macrophage polarization and bone destruction related to apical periodontitis (AP) stimulated by Porphyromonas gingivalis. Methodology: Macrophage polarization, IL-6/STAT3 expression, and the presence of P. gingivalis were detected in human AP tissues via RT-qPCR, western blotting, and immunohistochemistry staining. Murine bone marrow derived macrophages were isolated and cultured with P. gingivalis W83 in vitro, and levels of macrophage IL-6 expression, STAT3 phosphorylation, and macrophage polarization with or without the selective STAT3 phosphorylation inhibitor Stattic (5 μM) were detected via ELISA, western blotting, RT-qPCR, and flow cytometry, respectively. P. gingivalis-induced murine AP models were constructed, and bone destruction and macrophage polarization in the apical region were evaluated. Transwell co-culture systems were used to investigate the effects of macrophages infected with P. gingivalis on osteogenesis and osteoclastogenesis. Results: P. gingivalis was detected in human AP tissues that highly expressed IL-6/STAT3, and the M1 subtype of macrophages was more abundant in these tissues. P. gingivalis infection induced IL-6 expression, STAT3 phosphorylation, and M1 polarization of macrophages, while 5 μM of Stattic partially abolished these activation effects. Systemic STAT3 blockade via oral administration of Stattic at a dose of 25 mg kg-1 alleviated murine periapical bone resorption and apical infiltration of M1 macrophages induced by P. gingivalis infection in vivo. Furthermore, macrophages infected with P. gingivalis promoted bone destruction via secretion of IL-6, TNF-α, and RANKL, which hinder pre-osteoblast expression of Runx2 and accelerate pre-osteoclast expression of NFAT2. Conclusions:The activation of IL-6/STAT3 signaling pathway is involved in mediating macrophages M1 polarization in the P. gingivalis induced apical inflammatory context and may also be intimately involved in the bone loss caused by P. gingivalis infection, directing the M1 macrophage infiltration during the progression of AP.&nbsp

Identification and Heterologous Reconstitution of a 5-Alk(en)ylresorcinol Synthase from Endophytic Fungus \u3ci\u3eShiraia\u3c/i\u3e sp. Slf14

A new type III polyketide synthase gene (Ssars) was discovered from the genome of Shiraia sp. Slf14, an endophytic fungal strain from Huperzia serrata. The intron-free gene was cloned from the cDNA and ligated to two expression vectors pET28a and YEpADH2p-URA3 for expression in Escherichia coli BL21(DE3) and Saccharomyces cerevisiae BJ5464, respectively. SsARS was efficiently expressed in E. coli BL21(DE3), leading to the synthesis of a series of polyketide products. Six major products were isolated from the engineered E. coli and characterized as 1,3-dihydroxyphenyl-5-undecane, 1,3-dihydroxyphenyl-5-cis-6\u27-tridecene,1,3-dihydroxyphenyl-5-tridecane, 1,3-dihydroxyphenyl-5-cis-8\u27-pentadecene, 1,3-dihydroxyphenyl-5-pentadecane and 1,3-dihydroxyphenyl-5-cis-10\u27-heptadecene, respectively, based on the spectral data and biosynthetic origin. Expression of SsARS in the yeast also led to the synthesis of the same polyketide products, indicating that this enzyme can be reconstituted in both heterologous hosts. Supplementation of soybean oil into the culture of E. coli BL21(DE3)/SsARS increased the production titers of 1-6 and led to the synthesis of an additional product, which was identified as 5-(8\u27Z,11\u27Z-heptadecadienyl)resorcinol. This work thus allowed the identification of SsARS as a 5-alk(en)ylresorcinol synthase with flexible substrate specificity toward endogenous and exogenous fatty acids. Desired resorcinol derivatives may be synthesized by supplying corresponding fatty acids into the culture medium

Milk Oligopeptide Inhibition of (α)-Tocopherol Fortified Linoleic Acid Oxidation

This study investigated the effect of milk oligopeptides and (α)-tocopherol on inhibition of linoleic acid oxidation using Fe2+-vitamin C induced linoleic acid oxidation model through analysis of malondialdehyde, peroxide value, and conjugated diene and triene in the model. The alteration of milk oligopeptides maximal absorption wavelength, fluorescent feature, and secondary structure were further investigated to elucidate the interactions between milk oligopeptide and (α)-tocopherol that altered the inhibitory effect of linoleic acid oxidation. Results showed that Pro-Tyr-Tyr-Ala-Lys (PYYAK) and Ile-Pro-Ile-Gln-Tyr (IPIQY) with (α)-tocopherol significantly inhibited the oxidation of linoleic acid and reduced the formation of malondialdehyde by 38% and 41%, respectively. Additionally, Ile-Pro-Ile-Gln-Tyr-Val (IPIQYV) and (α)-tocopherol synergistically reduced the peroxide value in the model by 36.8%. Milk oligopeptides exhibited a blue shift on its maximal absorption wavelength, and their absorbance value decreased with the increase of the (α)-tocopherol concentration. The fluorescent intensity of milk oligopeptides was reduced with the addition of (α)-tocopherol and such fluorescent intensity reductions resulted from the static quenching process through the formation of milk oligopeptide-(α)-tocopherol complex. Fourier transform infrared spectroscopy analysis revealed that (α)-tocopherol significantly altered the secondary structure of milk oligopeptides and the percentage of β-turn obviously increased in milk oligopeptide-(α)-tocopherol complex. These indicated that the inhibition of linoleic acid oxidation might result from complex formed between milk oligopeptide and (α)-tocopherol through inter-molecular interactions

Perspectives on Using a Competitive Exclusion Approach to Control <i>Listeria monocytogenes</i> in Biological Soil Amendments of Animal Origin (BSAAO): A Review

Biological soil amendments of animal origin (BSAAO), such as animal waste or animal-waste-based composts, may contain foodborne pathogens such as Listeria monocytogenes. Due to the ubiquitous nature of Listeria, it is essential to understand the behavior of L. monocytogenes in BSAAO in order to develop preharvest prevention strategies to reduce pathogen contamination. As biological control agents, competitive exclusion (CE) microorganisms have been widely utilized in agriculture to control plant- or foodborne pathogens. Due to the diverse microbial community, animal wastes and composts are the potential sources for isolating CE strains for pathogen control. To explore the potential of using CE to control L. monocytogenes in BSAAO, we thoroughly reviewed the studies on the fate of L. monocytogenes in the agriculture field, and in the isolation and identification of CE from different matrices, and the applications of CE as a biological control method. Future studies using a next-generation sequencing approach to identify and characterize CE strains in complex microbial communities can provide a comprehensive picture of the microbial interactions between invading pathogens and the indigenous microbiota in BSAAO. This comprehensive review will provide insight into the development of effective biological control measures for preventing L. monocytogenes contamination in the agricultural field and enhancing food safety

Autophagy Proteins and clinical data reveal the prognosis of polycystic ovary syndrome

Abstract Objective We aimed to investigate the significance of autophagy proteins and their association with clinical data on pregnancy loss in polycystic ovary syndrome (PCOS), while also constructing predictive models. Methods This study was a secondary analysis. we collected endometrial samples from 33 patients with polycystic ovary syndrome (PCOS) and 7 patients with successful pregnancy control women at the Reproductive Center of the Second Hospital of Lanzhou University between September 2019 and September 2020. Liquid chromatography tandem mass spectrometry was employed to identify expressed proteins in the endometrium of 40 patients. R was use to identify differential expression proteins(DEPs). Subsequently, Metascape was utilized for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Multivariate Cox analysis was performed to analyze autophagy proteins associated with reproductive outcomes, while logistic regression was used for analyzing clinical data. Linear correlation analysis was conducted to examine the relationship between autophagy proteins and clinical data. We established prognostic models and constructed the nomograms based on proteome data and clinical data respectively. The performance of the prognostic model was evaluated by the receiver operating characteristic curve (ROC) and decision curve analysis (DCA). Results A total of 5331 proteins were identified, with 450 proteins exhibiting significant differential expression between the PCOS and control groups. A prognostic model for autophagy protein was developed based on three autophagy proteins (ARSA, ITGB1, and GABARAPL2). Additionally, another prognostic model for clinical data was established using insulin, TSH, TPOAB, and VD3. Our findings revealed a significant positive correlation between insulin and ARSA (R = 0.49), as well as ITGB1 (R = 0.3). Conversely, TSH exhibited a negative correlation with both ARSA (-0.33) and ITGB1 (R = -0.26). Conclusion Our research could effectively predict the occurrence of pregnancy loss in PCOS patients and provide a basis for subsequent research

Evaluation of the industrial cooperation of the Guangdong Hong Kong Macao Greater Bay Area: Based on the Origin-Destination pairs of industrial parks and coupling model efficiency

The industrial cooperation of Guangdong Hong Kong Macao Greater Bay Area (GBA) is one of the leading regional development strategies of this world-class urban agglomeration. This study constructed the industrial cooperation network based on the travel Origin-Destination (OD) connections among industrial parks. A multi-dimensional industrial cooperation and industrial development calculation index system were also set up to measure the nonlinear interaction relationship between them. The research found that an industrial collaboration network has been basically formed in the GBA, particularly presented by major cities. Some undeveloped cities may receive more benefits in the industrial collaboration network. The Covid-19 pandemic has had an impact in terms of within city connection instead of cross-city industrial cooperation. In addition, the degree of coupling between urban industrial coordination and urban industrial development has improved significantly over one decade and taking industrial collaboration as the input variables, industrial collaboration efficiently leads to industrial development outputs in almost every city in the GBA. Practically, decision makers should encourage and support intercity industrial collaboration, particularly between cities with closer geographic proximity, as it has been found to result in stronger cooperation and better economic enhancement. In addition, although industrial collaboration does not guarantee industrial development, when the collaboration systems and policies are enhanced, the synergy and coordination between them gradually improve. This highlights the potential benefits of continued investment in industrial collaboration for economic development

The Influence of Converting Food Crops to Forage Crops Policy Implementation on Herbivorous Livestock Husbandry Development&mdash;Based on Policy Pilot Counties in Hebei, China

In the context of increasing consumption of herbivorous livestock products, competition between humans and animals for food, and increasing environmental constraints, it is necessary to solve the problem of sustainable development of China&rsquo;s livestock industry and increase the protection and development of the grassland livestock industry while making good use of production resources in agricultural areas in order to explore the development potential of the herbivorous livestock industry in agricultural areas. The Converting Food Crops to Forage Crops Policy (CFFP), as an important measure of agricultural supply-side structural reform, aims to develop a high-quality forage industry and a high-quality herbivorous livestock industry. However, over the years of policy implementation, few studies have examined the impact effects of the policy on the development of the regional herbivorous livestock industry. To fill this research gap and provide theoretical support for subsequent policy implementation, the study used the synthetic control method to examine the impact of policy implementation on the development of herbivorous livestock production in the pilot counties in Hebei Province from 2010 to 2020. The study discovered that the policy&rsquo;s implementation encouraged the expansion of herbivorous livestock production in the pilot counties, but the policy&rsquo;s effects on various regions and livestock species varied due to the influence of local production bases and resource endowments