A Comparative Study on Antimicrobial Resistance in Escherichia coli Isolated from Channel Catfish and Related Freshwater Fish Species.

Antimicrobial resistance (AMR) trends in 114 generic Escherichia coli isolated from channel catfish and related fish species were investigated in this study. Of these, 45 isolates were from commercial-sized channel catfish harvested from fishponds in Alabama, while 69 isolates were from Siluriformes products, accessed from the U.S. Department of Agriculture Food Safety and Inspection Service' (FSIS) National Antimicrobial Resistance Monitoring System (NARMS) program. Antibiotic susceptibility testing and whole genome sequencing were performed using the GenomeTrakr protocol. Upon analysis, the fishpond isolates showed resistance to ampicillin (44%), meropenem (7%) and azithromycin (4%). The FSIS NARMS isolates showed resistance to tetracycline (31.9%), chloramphenicol (20.3%), sulfisoxazole (17.4%), ampicillin (5.8%) and trimethoprim-sulfamethoxazole, nalidixic acid, amoxicillin-clavulanic acid, azithromycin and cefoxitin below 5% each. There was no correlation between genotypic and phenotypic resistance in the fishpond isolates, however, there was in NARMS isolates for folate pathway antagonists: Sulfisoxazole vs. sul1 and sul2 (p = 0.0042 and p < 0.0001, respectively) and trimethoprim-sulfamethoxazole vs. dfrA16 and sul1 (p = 0.0290 and p = 0.013, respectively). Furthermore, correlations were found for tetracyclines: Tetracycline vs. tet(A) and tet(B) (p < 0.0001 each), macrolides: Azithromycin vs. mph(E) and msr(E) (p = 0.0145 each), phenicols: Chloramphenicol vs. mdtM (p < 0.0001), quinolones: Nalidixic acid vs. gyrA_S83L=POINT (p = 0.0004), and ß-lactams: Ampicillin vs. blaTEM-1 (p < 0.0001). Overall, we recorded differences in antimicrobial susceptibility testing profiles, phenotypic-genotypic concordance, and resistance to critically important antimicrobials, which may be a public health concern.

Low-dose Oral Thimerosal for the Treatment of Oral Herpes: Clinical Trial Results and Improved Outcome After Post-hoc Analysis.

BACKGROUND: Thimerosal (TML) is an organomercury antimicrobial. Low doses (1/250th of the amount in a typical vaccine dose) may promote an antiviral immune response. Low-dose TML (BTL-TML) was evaluated for safety and efficacy against herpes labialis in two FDA-approved, randomized, double blind, placebo-controlled clinical trials. METHODS: BTL-TML was evaluated in a Phase IIa trial for its ability to block progression to lesion in subjects with recurrent oral herpes caused by dental trauma. Subjects were administered BTL-TML or a saline control over a 7-day period. In a Phase IIb trial, BTL-TML was evaluated for its ability to block progression to lesion over a 7-day period in subjects with herpes lip infections induced by exposure to ultraviolet (UV) radiation. RESULTS: Progression to lesion post-dental procedure was prevented in 54.5% (12/22) TML subjects versus 22.2% (2/9) control subjects (p = 0.106). Progression to lesion post-UV irradiation was blocked in 47.8% (11/23) BTL-TML treatment subjects and 42.8% (6/14) control subjects. A post-hoc analysis yielded 52.2% (12/23) BTL-TML subjects with no progression to lesion versus 28.6% (6/21) control subjects with no progression (p = 0.099). There were no significant differences in adverse effects between treatment and control groups in either trial. CONCLUSIONS: Neither clinical trial showed a statistically significant effect of BTL-TML on progression to lesion. However, the post-hoc analysis suggested there is a 48-hour period following UV radiation exposure during which the anti-herpes activity of antivirals such as BTL-TML is reduced. Accordingly, BTL-TML may have promise in subsequent, properly designed and powered clinical trials.

Effects of cannabis oil extract on immune response gene expression in human small airway epithelial cells (HSAEpC): implications for chronic obstructive pulmonary disease (COPD).

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is commonly associated with both a pro-inflammatory and a T-helper 1 (Th1) immune response. It was hypothesized that cannabis oil extract can alleviate COPD symptoms by eliciting an anti-inflammatory Th2 immune response. Accordingly, the effects of cannabis oil extract on the expression of 84 Th2 and related immune response genes in human small airways epithelial cells (HSAEpC) were investigated. METHODS: HSAEpC from a single donor were treated with three dilutions of a standardized cannabis oil extract (1:400, 1:800 and 1:1600) along with a solvent control (0.25% [2.5 ul/ml] ethanol) for 24 h. There were four replicates per treatment dilution, and six for the control. RNA isolated from cells were employed in pathway-focused quantitative polymerase chain reaction (qPCR) microarray assays. RESULTS: The extract induced significant (P < 0.05) changes in expression of 37 tested genes. Six genes (CSF2, IL1RL1, IL4, IL13RA2, IL17A and PPARG) were up-regulated at all three dilutions. Another two (CCL22 and TSLP) were up-regulated while six (CLCA1, CMA1, EPX, LTB4R, MAF and PMCH) were down-regulated at the 1:400 and 1:800 dilutions. The relationship of differentially-expressed genes of interest to biologic pathways was explored using the Database for Annotation, Visualization and Integrated Discovery (DAVID). CONCLUSIONS: This exploratory investigation indicates that cannabis oil extract may affect expression of specific airway epithelial cell genes that could modulate pro-inflammatory or Th1 processes in COPD. These results provide a basis for further investigations and have prompted in vivo studies of the effects of cannabis oil extract on pulmonary function. TRIAL REGISTRATION: NONE (all in vitro experiments).

Can Unconventional Immunomodulatory Agents Help Alleviate COVID-19 Symptoms and Severity?

Severe acute respiratory syndrome coronavirus 2 (SARS coronavirus 2, or SARS-CoV-2) is the cause of the respiratory infection known as COVID-19. From an immunopathological standpoint, coronaviruses such as SARS-CoV-2 induce increased levels of a variety of T-helper 1 (Th1) and inflammatory cytokines and chemokines, including interleukin-1 (IL-1), IL-6, CCL2 protein, and CXCL10 protein. In the absence of proven antiviral agents or an effective vaccine, substances with immunomodulatory activity may be able to inhibit inflammatory and Th1 cytokines and/or yield an anti-inflammatory and/or Th2 immune response to counteract COVID-19 symptoms and severity. This report briefly describes the following four unconventional but commercially accessible immunomodulatory agents that can be employed in clinical trials to evaluate their effectiveness at alleviating disease symptoms and severity: low-dose oral interferon alpha, microdose DNA, low-dose thimerosal, and phytocannabinoids.

Occurrence of Listeria monocytogenes in Ready-to-Eat Meat and Poultry Product Verification Testing Samples from U.S. Department of Agriculture-Regulated Producing Establishments, 2005 through 2017.

ABSTRACT: Ready-to-eat (RTE) meat and poultry product samples collected between 2005 and 2017 from RTE-producing establishments for the U.S. Department of Agriculture, Food Safety and Inspection Service (FSIS) ALLRTE/RTEPROD_RAND (random) and RTE001/RTEPROD_RISK (risk-based) sampling projects were tested for Listeria monocytogenes (Lm). Data for 45,897 ALLRTE/RTEPROD_RAND samples collected from 3,607 distinct establishments and 112,347 RTE001/RTEPROD_RISK samples collected from 3,283 distinct establishments were analyzed for the presence of Lm. These data were also analyzed based upon the percentages of establishments with positive samples, annual production volume, sanitation control alternatives, geographic location, and season or month of sample collection. Results revealed low occurrence of Lm-positive samples from the random and risk-based sampling projects, with 152 (0.33%) positive samples for ALLRTE/RTEPROD_RAND and 403 (0.36%) positive samples for RTE001/RTEPROD_RISK. The percentage of positive samples significantly decreased over time, from about 0.7% in 2005 and 2006 to about 0.2% in 2017 (P < 0.05). From 2005 to 2017, 3.9% of establishments sampled under the ALLRTE/RTEPROD_RAND sampling project had at least one Lm-positive sample. Similarly, 10.0% of establishments sampled under the RTE001/RTEPROD_RISK sampling project had at least one positive sample. Samples positive for Lm were found in all geographic regions in all months. Thus, in 13 years of RTE product sampling in FSIS-regulated establishments (2005 through 2017), <0.4% of samples were positive for Lm in both risk-based and random sampling projects. The low prevalence of Lm in these products suggests that the combination of FSIS policies and industry practices may be effective for controlling Lm contamination. Information obtained from these sampling projects is relevant to the ongoing prevention of foodborne Lm illnesses from RTE meat and poultry products.

Occurrence of Salmonella in Ready-to-Eat Meat and Poultry Product Samples from U.S. Department of Agriculture-Regulated Producing Establishments. I. Results from the ALLRTE and RTE001 Random and Risk-Based Sampling Projects, from 2005 to 2012.

Ready-to-eat (RTE) meat and poultry product samples collected from RTE-producing establishments for the ALLRTE (random) and RTE001 (risk-based) sampling projects of the Food Safety and Inspection Service (FSIS) were tested for both Salmonella and Listeria monocytogenes. The FSIS analyzed Salmonella results for RTE meat and poultry product samples collected for the two sampling projects from 2005 to 2012. Data for 24,385 ALLRTE samples collected from 3,023 establishments and 66,653 RTE001 samples collected from 2,784 establishments were evaluated for the percentages of Salmonella-positive samples, product types of positive samples, and Salmonella serotypes. There also were descriptive summaries with respect to establishment hazard analysis and critical control point (HACCP) size, production volumes, L. monocytogenes control alternatives, geographic location, and season or month of sample collection. Results showed low occurrences of Salmonella-positive samples from the ALLRTE and RTE001 sampling projects, with 14 positive samples (0.06%) for ALLRTE and 33 positive samples (0.05%) for RTE001. Percentages of establishments with at least one Salmonella-positive sample averaged 0.46% for ALLRTE and 1.11% for RTE001. Three product types-sausage products, pork barbecue, and head cheese-accounted for 62% of all positive samples. There were 27 distinct serotypes from 48 Salmonella isolates, with serotypes Infantis and Typhimurium being the most common (5 isolates each). All but one of the Salmonella-positive samples were obtained from establishments with HACCP sizes of small or very small. More than half of the positive samples were obtained from establishments using L. monocytogenes control alternative 3 (sanitation only, highest-risk category). Positive Salmonella samples were found in all geographic regions at all times of the year. Information obtained from these sampling projects is relevant to the prevention of foodborne Salmonella illnesses from RTE meat and poultry products.

Occurrence of Salmonella in Ready-to-Eat Meat and Poultry Product Samples from U.S. Department of Agriculture-Regulated Producing Establishments. II. Salmonella in Ready-to-Eat Pork Barbecue Products, from 2005 to 2012.

Ready-to-eat (RTE) meat and poultry product samples from the random ALLRTE and risk-based RTE001 sampling projects of the Food Safety and Inspection Service (FSIS) were tested for both Listeria monocytogenes and Salmonella. In the course of analyzing Salmonella data for calendar years 2005 to 2012, it was observed that 8 (17.0%) of 47 positive samples were from pork barbecue. The eight Salmonella-positive samples, from seven establishments in a single state, were from 1,085 pork barbecue samples tested nationwide (0.74% positive) and from 296 samples tested from that one state (2.7% positive). The seven establishments represented 30.4% of 23 federal establishments in that state that had pork barbecue samples tested for Salmonella. A follow-up sample from intensified verification testing at one of the seven establishments also was positive for Salmonella. Upon further examination, contamination appeared to be influenced by regional differences in production methods. Notably, the style of pork barbecue that tested positive for Salmonella used a vinegar- and pepper-based sauce in which the ingredients were mixed without cooking. All the establishments with Salmonella-positive samples followed the practice of first cooking the pork and then adding the barbecue sauce ingredients (vinegar, pepper, other spices, etc.) after cooking (postlethality exposure). In addition to the sauce ingredients, other possible sources of contamination included employee hygiene and food handling practices and cross-contamination from other Salmonella-contaminated products and from commonly used equipment. Based on these findings, the FSIS issued guidelines recommending changes in production methods that would minimize or eliminate pork barbecue as a potential source of foodborne Salmonella infections.

Low-dose oral interferon modulates expression of inflammatory and autoimmune genes in cattle.

While the safety and efficacy profiles of orally administered bovine interferon (IFN) alpha have been documented, the mechanism(s) that result in clinical benefits remain elusive. One approach to delineating the molecular pathways of IFN efficacy is through the use of gene expression profiling technologies. In this proof-of-concept study, different (0, 50, 200 and 800 units) oral doses of natural bovine IFN (type I) were tested in cattle to determine if oral IFN altered the expression of genes that may be pivotal to the development of systemic resistance to viral infections such as foot-and-mouth disease (FMD). Oral IFN was administered twice: Time 0 and 8h later. Blood was collected at 0, 8 and 24h after the first IFN administration, and DNA isolated from peripheral blood mononuclear cells (PBMCs) was employed in quantitative polymerase chain reaction (qPCR) microarray assays. Within 8h, 50 and 200 units of oral IFN induced significant (P<0.05) changes in expression of 41 of 92 tested autoimmune and inflammatory response-associated genes. These data suggest that orally administered IFN is a viable approach for providing short-term antiviral immunity to livestock exposed to viruses such as FMD virus (FMDV) until such a time that an effective vaccine can be produced and distributed to producers.

Effects of streptolysin o on extracellular matrix gene expression in normal human epidermal keratinocytes.

ML-05 is a non-hemolytic form of streptolysin O, the membrane-damaging extracellular toxin produced by certain streptococci. ML-05 stimulates keratinocyte migration and proliferation in wound-healing scratch assays and promotes wound healing in a human skin organ culture wound model. Pathway-focused DNA microarrays were used to elucidate ML-05's mechanism of action in wound healing processes. Normal human epidermal keratinocytes (NHEK) were treated with varying concentrations of ML-05 for 24 hours, followed by RNA extraction and cRNA production. Gene expression profiling utilized microarrays containing nucleic acid probes for 113 extracellular matrix (ECM) genes. Microarrays yielded 6 upregulated and 4 downregulated genes with ≥2-fold changes and p<0.05 in t-tests. Quantitative real-time polymerase chain reactions (qPCR) were used to verify gene regulation. Upregulated genes of interest were VCAN (formerly CSPG2, encoding versican), CD44 (encoding hyaluronan receptor), ICAM1 (encoding intercellular adhesion molecule-1) and CTGF (encoding connective tissue growth factor). All four upregulated genes encode proteins involved in promoting keratinocyte migration and proliferation. Downregulated genes of interest were MMP9 (encoding matrix metalloproteinase 9) and SPP1 (encoding osteopontin). ML-05 may enhance wound healing through the expression of specific genes encoding proteins capable of promoting keratinocyte migration, proliferation, and other activities related to maintaining ECM structure and function.

Cytotoxic xanthones from Psorospermum molluscum from the Madagascar rain forest.

Two new cytotoxic xanthones were isolated from extracts of the Madagascar rain forest plant Psorospermum cf. molluscum using bioassay-guided fractionation with the Escherichia coli SOS chromotest. The structures of the new dihydrofuranoxanthones, designated 3',4'-deoxy-4'-chloropsoroxanthin-(3',5'-diol) ( 1) and psoroxanthin ( 4), were determined on the basis of 2D-NMR, MS, and UV spectroscopic data and are structurally related to the psorospermins, a known class of plant antitumor agents. A new hydroxyprenylated xanthone ( 5) is also described. Xanthones 1 and 4 showed selective in vitro cytotoxicity against ABAE cells (bovine endothelial cell line).

Streptolysin O enhances keratinocyte migration and proliferation and promotes skin organ culture wound healing in vitro.

ML-05, a modified form of the hemolytic and cytotoxic bacterial toxin, streptolysin O, is currently being investigated as a treatment for collagen-related disorders such as scleroderma and fibrosis. Furthermore, ML-05 may be effective in promoting wound healing and alleviating the formation of hypertrophic scars and keloids. To investigate the effects of ML-05 on wound-healing processes, in vitro wound-healing scratch assays (using human primary epidermal keratinocytes and dermal fibroblasts) and a human skin organ culture wound model were utilized. ML-05 markedly enhanced keratinocyte migration and proliferation in wound scratch assays. ML-05 did not affect either proliferation or migration of dermal fibroblasts, indicating that ML-05's effects on cell migration/proliferation may be keratinocyte-specific. ML-05 was tested in a dose-dependent manner in a skin organ culture wound model using two different application methods: Through the culture media (dermal exposure) or direct topical treatment of the wound surface. ML-05 was found to accelerate wound healing as measured by reepithelialization, particularly after topical application. Therefore, ML-05 may have potential as a wound-healing agent that promotes reepithelialization through stimulation of keratinocyte migration and proliferation.

The use of streptolysin o for the treatment of scars, adhesions and fibrosis: initial investigations using murine models of scleroderma.

Diseases and conditions involving the deposition of excessive amounts of collagen include scleroderma, fibrosis, and scar and surgical adhesion formation. Diseases such as scleroderma may result from acute and chronic inflammation, disturbances in the normal parenchymal area, and activation of fibroblasts. ML-05, a modified form of the hemolytic and cytotoxic bacterial toxin, streptolysin O, is being developed for the treatment of such collagen-related disorders. At sublytic concentrations in vitro, ML-05 was shown to activate CD44 expression. This may modulate production of collagen, hyaluronate, and their associated enzymes to allow a restoration of normal extracellular matrices within tissues. More importantly, ML-05 appeared to decrease skin collagen levels in two in vivo models of collagen disorders, the tight skin mouse (Tsk) model of scleroderma, and the bleomycin-induced mouse skin fibrosis model. In the Tsk model, levels of hydroxyproline (a measure of total collagen) decreased by 25% in the Tsk+ML-05 treatment group relative to the Tsk+saline control group over a 3-month period. In the bleomycin-induced skin fibrosis study, hydroxyproline levels decreased from 15-22% over a 6-week period in a bleomycin-induced ML-05 treatment group (relative to levels in a bleomycin-induced, untreated control group). Hydroxyproline levels in samples from this treatment group were only slightly greater than levels in an uninduced control group at 8 weeks. Thus, ML-05 treatment appeared to reduce collagen levels in two separate mouse skin fibrosis models, one genetically based and the other chemically induced.

Bioactive saponins from Acacia tenuifolia from the suriname rainforest.

Bioassay-guided fractionation of the MeOH extract of Acacia tenuifolia using the engineered yeast strains 1138, 1140, 1353, and Sc7 as the bioassay tool resulted in the isolation of the three new saponins 3, 5, and 6 and the three known saponins 1, 2, and 4. The structures of the new compounds were established on the basis of HRMS, 1D and 2D NMR spectral data on the intact saponins, and GC-MS analyses of the sugars. Compounds 1,2 and 5,6 showed cytotoxicity against mammalian cell lines.