Staphylococcus Research Articles

Putative staphylococcal enterotoxin possesses two common structural motifs for MHC-II binding

Staphylococcus aureus has become a significant cause of health risks in humankind. Staphylococcal superantigens (SAgs) or enterotoxins are the key virulent factors that can exhibit acute diseases to severe life-threatening conditions. Recent literature reports S. aureus has steadily gained new enterotoxin genes over the past few decades. In spite of current knowledge of the established SAgs, several questions on putative enterotoxins are still remaining unanswered. Keeping that in mind, this study sheds light on a putative enterotoxin SEl26 to characterize its structural and functional properties. In-silico analyses indicate its close relation with the conventional SAgs, especially the zinc-binding SAgs. Additionally, important residues that are vital for the T-cell receptor (TcR) and major histocompatibility complex class II (MHC-II) interaction were predicted and compared with established SAgs. Besides, our biochemical analyses exhibited the binding of this putative enterotoxin with MHC-II, followed by regulating pro-inflammatory and anti-inflammatory cytokines.

Evolution of Invasive and Colonizing Pediatric Staphylococcus aureus Isolates

Abstract Background Staphylococcus aureus is the most common bacterial pathogen isolated in children with invasive bloodstream and musculoskeletal infections. A major barrier to vaccine and novel therapeutic development against S. aureus is its continuous evolution. Many recent and current vaccine candidates target antigens that may have appeared crucial to its pathogenesis, only to see these factors recede from circulating invasive strains. Characterization of clinically relevant S. aureus isolates over time is necessary to find targets for novel therapeutics. Efforts toward new interventions should target factors that remain prevalent in invasive isolates over time. Methods We obtained 131 invasive S. aureus isolates from children admitted to the Monroe Carell Jr. Children's Hospital at Vanderbilt from 2010 to 2022, and 246 colonizing isolates. Whole genome sequences were obtained using the Illumina sequencing platform. Virulence factors and multi-locus sequence type (MLST) were determined using Geneious software and through PubMLST, an open-access curated database. Results There were 10 unique clonal complexes (CC) identified among 320 isolates. CC8 remains the most common invasive clonal complex, though CC8 is significantly less frequent now than in 2010-2012. Diversity of invasive strains has increased substantially, with the emergence of CC5 and CC121 lineages causing invasive disease in children. CC1 was found exclusively in colonization isolates. Virulence factor prevalence has changed significantly over time. Panton-Valentine leucocidin (PVL), enterotoxins K and Q (SEK/SEQ), and the pathogenic arginine catabolic mobile element (ACME), have decreased significantly in invasive S. aureus isolates since 2010, from 53-70% to 21-35% of current isolates. Leukocidin ED (LukED) and surface protein staphylokinase (Sak) are strongly associated with invasive isolates compared to colonization isolates. Conversely, toxic shock toxin (TST) and Staphylococcal enterotoxin B (SEB) were seen more commonly in colonization isolates (12-15%) compared to invasive isolates (5-7%). The genes encoding for gamma hemolysins (HlgA-C), leukocidin AB (LukAB), iron-regulated surface proteins (IsdA, IsdB), staphylococcal binding immunoglobulin protein (sbi), extracellular fibrinogen-binding protein (Efb), and clumping factors A and B (ClfA, ClfB) were identified in all isolates tested. Figure 1 Figure 2 Conclusions Significant shifts have occurred over the past decade in the predominant circulating S. aureus strains and their virulence factor repertoires. The once-dominant CC8 (USA300 clone) has receded, and with that has come a significant reduction in some factors once thought crucial for pathogenesis. These findings also have implications for infection prevention and control practices, as eradication of colonizing strains with low potential for pathogenicity may inadvertently allow for replacement by more virulent strains.

Ready-to-Eat Foods: A Potential Vehicle for the Spread of Coagulase-Positive Staphylococci and Antimicrobial-Resistant Staphylococcus aureus in Buea Municipality, South West Cameroon.

The consumption of ready-to-eat (RTE) foods contaminated with coagulase-positive staphylococci (CoPS) and especially Staphylococcus aureus puts consumers at a potential risk of food-borne disease or colonization and subsequent infection. This cross-sectional study determined the levels of CoPS and the presence of S. aureus in RTE foods sold in Buea municipality. A total of 420 RTE food samples, comprising 70 each of cake, bread, fruit salad, meat hot-pot, suya, and boiled rice were randomly purchased from February to August 2020. The CoPS counts were determined by culturing on Baird-Parker agar, and S. aureus was identified by amplification of the nuc gene using the polymerase chain reaction. All S. aureus isolates were screened for the presence of classical staphylococcal enterotoxin genes. To determine antimicrobial resistance profiles, each isolate was tested against 11 antimicrobials. Oxacillin-resistant S. aureus strains were analyzed for the presence of the mecA gene. Overall, 161 (38.3%) samples had detectable levels of CoPS ranging from 2.0 to 5.81 log10 CFU/g. Based on CoPS levels, 37 (8.81%) of the 420 RTE food samples-only fruit salad and meat hot-pot, had unsatisfactory microbiological quality. A total of 72 S. aureus isolates, comprising 52.78% from fruit salad, 16.67% from meat hot-pot, 12.5% from boiled rice, 9.72% from suya, 5.56% from bread, and 4.17% from cake, were recovered. None of the S. aureus isolates possessed any of the classical enterotoxin genes. All the isolates were susceptible to vancomycin and ofloxacin, while 68 (94.44%) and 66 (91.67%) were susceptible to oxacillin and ciprofloxacin, respectively. Resistance to penicillin (93.06%) was highest, followed by amoxicillin (91.67%) and erythromycin (79.17%). Four isolates were identified as methicillin-resistant S. aureus, all of which carried the mecA gene. A total of 24 antibiotypes were identified. Our findings showed that RTE foods sold in the Buea municipality are likely vehicles for the transmission of CoPS and antimicrobial-resistant S. aureus.

Bacterial cGAS senses a viral RNA to initiate immunity

Cyclic oligonucleotide-based antiphage signalling systems (CBASS) protect prokaryotes from viral (phage) attack through the production of cyclic oligonucleotides, which activate effector proteins that trigger the death of the infected host1,2. How bacterial cyclases recognize phage infection is not known. Here we show that staphylococcal phages produce a structured RNA transcribed from the terminase subunit genes, termed CBASS-activating bacteriophage RNA (cabRNA), which binds to a positively charged surface of the CdnE03 cyclase and promotes the synthesis of the cyclic dinucleotide cGAMP to activate the CBASS immune response. Phages that escape the CBASS defence harbour mutations that lead to the generation of a longer form of the cabRNA that cannot activate CdnE03. Since mammalian oligoadenylate synthetases also bind viral double-stranded RNA during the interferon response, our results reveal a conserved mechanism for the activation of innate antiviral defence pathways.

The Enterotoxin Gene Profiles and Enterotoxin Production of Staphylococcus aureus Strains Isolated from Artisanal Cheeses in Belgium.

A Staphyloccoccus aureus is one of the leading causes of food poisoning outbreaks (FPOs) worldwide. Staphylococcal food poisoning (SFP) is induced by the ingestion of food containing sufficient levels of staphylococcal enterotoxins (SEs). Currently, 33 SEs and SE-like toxins (SEls) have been described in the literature, but only five named "classical" enterotoxins are commonly investigated in FPOs due to lack of specific routine analytical techniques. The aims of this study were to (i) establish the genetic profile of strains in a variety of artisanal cheeses (n = 30) in Belgium, (ii) analyze the expression of the SE(l)s by these strains and (iii) compare the output derived from the different analytical tools. Forty-nine isolates of S. aureus were isolated from ten Belgian artisanal cheeses and were analyzed via microbiological, immunological, liquid chromatography mass spectrometry, molecular typing and genetic methods. The results indicated that classical SEs were not the dominant SEs in the Belgian artisanal cheeses that were analyzed in this study, and that all S. aureus isolates harbored at least one gene encoding a new SE(l). Among the new SE(l)s genes found, some of them code for enterotoxins with demonstrated emetic activity and ecg-enterotoxins. It is worth noting that the involvement of some of these new SEs has been demonstrated in SFP outbreaks. Thus, this study highlighted the importance of the development of specific techniques for the proper investigation of SFP outbreaks.

Epigallocatechin Gallate Alleviates Staphylococcal Enterotoxin A-Induced Intestinal Barrier Damage by Regulating Gut Microbiota and Inhibiting the TLR4-NF-κB/MAPKs-NLRP3 Inflammatory Cascade.

Natural phytochemicals have attracted increasing attention because of their promising ability to tackle bacteriotoxin-induced public safety concerns. However, it is unclear how natural phytochemicals regulate the intestinal barrier dysfunction caused by bacteriotoxin, such as staphylococcal enterotoxin A (SEA). This study aims to illustrate the in vitro and in vivo protective mechanism of epigallocatechin gallate (EGCG) on SEA-triggered intestinal barrier damage and inflammation. Results show that EGCG alleviates intestinal barrier damage by effectively inhibiting SEA-induced intestinal permeability increase, tight junction protein and mucin loss, and intestinal cell apoptosis. EGCG also reduces intestinal inflammation by suppressing the TLR4-NF-κB/MAPKs-NLRP3 pathway. Importantly, EGCG reverses gut microbiota dysbiosis and short-chain fatty acid (SCFA) content decrease induced by SEA. It is worth noting that this study also detects the direct interaction between the phytochemical and virulence factors and finds that EGCG effectively not only inhibits the secretion of SEA but also binds with the secreted SEA to attenuate its toxicity. Taken together, EGCG mitigates SEA-induced intestinal barrier dysfunction via gut microbiota SCFA-mediated TLR4-NF-κB/MAPKs-NLRP3 inflammatory cascade inhibition. Overall, this research provides enlightening insight into the application of bacteriotoxin-targeting natural compounds in the field of food safety and human wellness.

Investigation into the prevalence of enterotoxin genes and genetic background of Staphylococcus aureus isolates from retain foods in Hangzhou, China

BackgroundStaphylococcus aureus expresses numerous toxins, many of which are strongly believed to be responsible for specific symptoms and even diseases, making it significant in the pathogenesis of human health. Enterotoxins, which are vital toxins, are associated with foodborne illnesses that manifest through symptoms like vomiting and diarrhea. In the present study, 264 S. aureus isolates obtained from various retail foods in Hangzhou, China were further investigated the profiles of enterotoxin genes and genetic backgrounds.ResultsApproximately, 64.02% of the isolates from diverse sources contained at least one Staphylococcal Enterotoxin (SE) genes, displaying a total of 36 distinct combinations. Enterotoxin gene cluster (egc) encoded enterotoxin genes, normally designated by seg, sei, sem, sen, seo and selu, plus with sep were more frequently detected (33.73%, each). In contrast, see, ses and set were absent in any of the isolates tested. A total of 44 sequence types (STs), 20 clonal complexes (CCs) and 66 different staphylococcal protein A (spa) types (including six novel types) were identified among those 169 SE-positive isolates. Moreover, nineteen methicillin-resistant Staphylococcus aureus (MRSA) isolates were identified. The majority of those isolates belonged to the CC59-Sccmec IVa cluster and carried the seb-sek-seq gene cluster. The egc cluster, either coexisting with or without other enterotoxin genes, was observed in all isolates allocated into CC5, CC9, CC20, CC25, CC72 and ST672. Irrespective of the spa types and origins of the food, it appeared that seh was a distinct genetic element present in isolates belonging to the CC1 clonal lineage.ConclusionsThe results not only proposed a suspected relationship between distribution of enterotoxigenic strains and genetic backgrounds, but also attributed the presence of novel enterotoxins to potential hazards in food safety.

Genotypic diversity of staphylococcal enterotoxin B gene (seb) and its association with molecular characterization and antimicrobial resistance of Staphylococcus aureus from retail food

To investigate the expression pattern of staphylococcal enterotoxin B (SEB) in food and the genotypic diversity of SEB-encoding gene in association with molecular characteristics and antimicrobial resistance of S. aureus, 498 isolates from retail food were screened for seb gene and detected for SEB production in S. aureus. In addition, the seb nucleotide sequences, virulence genes, resistance genes, antimicrobial susceptibility and molecular characteristics of S. aureus were examined. A total of 45 (9.0 %) seb-positive S. aureus strains were identified, all of which expressed SEB. The detection rate of SEB-production strains was significantly higher from dairy-related sources than those from other sources (P < 0.05). In vitro simulations showed that S. aureus could grow and express SEB in both milk and pork, with SEB expression exceeding 20 ng/g after 1 day of storage at room temperature. There were 2 distinct SEB genotyping (SEB1 and SEB2) in the SEB amino acid sequences of the 45 isolates, including 4 amino acid differences (Ala-13Val, Ser14Ala, Asn192Ser, and Met222Leu). There was no significant difference (P > 0.05) in SEB production between SEB1 and SEB2 genotyping strains. Based on MLST clustering analysis, the same molecular type strains were found to have the same SEB genotyping, virulence gene profile, resistance gene profile and drug resistance profile. Among them, the dominant molecular types of SEB1 and SEB2 strains were CC1-ST188-t189 and CC59-ST59-t437, respectively. Compared to the CC1-ST188-t189 clonal strain, the CC59-ST59-t437 clonal strain carried a higher number of virulence and resistance genes and exhibited a broader resistance profile. Therefore, understanding the characteristics of the strains and their expression patterns in food can be effective in preventing food poisoning incidents.

Prevalence and Characteristics of Invasive Staphylococcus argenteus among Patients with Bacteremia in Hong Kong.

Staphylococcus argenteus is a novel Staphylococcus species derived from Staphylococcus aureus. Information on the prevalence and genetic characteristics of invasive S. argenteus in Asia is limited. In this study, 275 invasive S. aureus complex strains were retrieved from blood culture specimens in Hong Kong and re-analyzed using MALDI-TOF mass spectrometry and an in-house multiplex real-time PCR for S. argenteus. The prevalence of invasive S. argenteus in Hong Kong was found to be 4.0% (11/275). These strains were primarily susceptible to commonly used antibiotics, except penicillin. Whole-genome sequencing revealed the circulation of three S. argenteus genotypes (ST-2250, ST-1223, and ST-2854) in Hong Kong, with ST-2250 and ST-1223 being the predominant genotypes. The local ST-2250 and ST-1223 strains showed close phylogenetic relationships with isolates from mainland China. Antimicrobial-resistant genes (fosB, tet-38, mepA, blaI, blaZ) could be found in nearly all local S. argenteus strains. The ST-1223 and ST-2250 genotypes carried multiple staphylococcal enterotoxin genes that could cause food poisoning and toxic shock syndrome. The CRISPR/Cas locus was observed only in the ST-2250 strains. This study provides the first report on the molecular epidemiology of invasive S. argenteus in Hong Kong, and further analysis is needed to understand its transmission reservoir.

B-216 Development of a Point-of-care Multiplexed Room Temperature Stable Environmental Toxin Test

Abstract Background Hememics™ Biosensor System (HBS) is a platform technology enabling antigen and molecular testing in the field. The system consists of a reader, HemBox™, and a biosensing chip, HemChip™. The HemBox™ is a battery-operated, cell phone size with Bluetooth and WIFI module that can be integrated into the Health Management System. The biosensing chips can be programed with bio-receptors to detect pathogens in less than 5 min. This study presents performance characteristics of HBS as a rapid POC to detect toxins from environmental samples. Methods The testing protocol consists of insert a HemChip™ into the HemBox™ and mixing the sample with the buffer and apply a drop of this mixture to the chip. In less than 5 min, results are displayed on the screen of HemBox™ or transmitted to user’s cellphone via Bluetooth. Samples were prepared by spiking solutions of mud, serum, vegetable wash, and milk with Staphylococcal enterotoxin B (SEB) and ricin. Results HemChip™ was functionalized to recognize Ricin and SEB targets. For testing, these targets were mixed in various matrix. To achieve statistical significance, each sample tested represented a minimum of 10 data points. Conclusion Hememics™ Biosensor System (HBS) is designed to be the ultimate Portable Lab Diagnostics for field use. The ability to test for environmental toxins in the field is significant because it allows for quick and on-site detection of potentially harmful substances. Our biosensor system also eliminates the need for sample processing, collection and transportation, which can take significant time and resources. The results of field testing can be available almost immediately, allowing for prompt action to be taken in the case of high toxin levels.

Brain circuits for retching-like behavior.

Nausea and vomiting are important defensive responses to cope with pathogens and toxins that invade the body. The nucleus of the solitary tract (NTS) is important for initiating these responses. However, the molecular heterogeneities and cellular diversities of the NTS occlude a better understanding of these defensive responses. Here, we constructed the single-nucleus transcriptomic atlas of NTS cells and found multiple populations of NTS neurons that may be involved in these defensive responses. Among these, we identified Calbindin1-positive (Calb1+) NTS neurons that are molecularly distinct from Tac1+ neurons. These Calb1+ neurons are critical for nausea and retching induced by cereulide; an emetic toxin secreted by Bacillus Cereus. Strikingly, we found that cereulide can directly modulate vagal sensory neurons that innervate Calb1+ NTS neurons, a novel mechanism distinct from that for nausea and retching induced by Staphylococcal enterotoxin A. Together, our transcriptomic atlas of NTS neurons and the functional analyses revealed the neural mechanism for cereulide-induced retching-like behavior. These results demonstrate the molecular and cellular complexities in the brain that underlie defensive responses to the diversities of pathogens and toxins.

Effect of environmental factors on expression of staphylococcal enterotoxin genes.

Staphylococcal enterotoxins (SEs) secreted by Staphylococcus aureus (S. aureus) can cause foodborne disease, nausea, vomiting and diarrhea, and even death. Regulation of SE expression is related to accessory gene regulators (Agr). It is important to reveal which environmental factors influence regulation of SE expression to prevent SE food poisoning outbreak. Hence, natural environmental factors which may have an impact on SE expression were selected, such as temperature, food types, strains, and competing strains. Seven strains of S. aureus carrying different SE genes were collected from the Chinese Academy of Inspection and Quarantine (CAIQ) strain bank for study. Strains were cultured with different conditions. Temperature was 8°C, 22°C, and 30°C. Food type was milk powder and nutrient broth. Competing strains were Vibrio parahaemolyticus (V. parahaemolyticus), Escherichia coli (E. coli), and Bacillus cereus (B. cereus). The expression culture solution was pretreated by centrifugation, then determined by using SDS-PAGE, and distinguished SEs apart from each other by HPLC-ESI-TOF. There are 168 samples collected from SE expression culture; the result of SDS-PAGE suggests 23 samples were positive for SEs, and the other 145 samples were negative for SEs. The result of HPLC-ESI-TOF suggests that SEs with similar molecular weight can be distinguished in terms of m/z. The most important factor contributing to regulate expression of SEs was estimated by logistic regressive analysis. The result shows that McFadden R2 is 0.213; p value is 0.000 (p < 0.05); this result illustrates that the model is valid and meaningful. Strains, food types, temperature, and competing strands can explain the 21% change in SE expression. Temperature (z = 3.029, p = 0.002 < 0.01), strains (z = - 3.132, p = 0.002 < 0.01), and food types (z = - 2.415, p = 0.016 < 0.05) have significant impact on SE expression, and the competing strains (z = 1.230, p = 0.219 > 0.05) have no impact on the SE expression. More important impact on SE expression was estimated by OR value; the result shows that strength of temperature influencing on SE expression is bigger than strains and food types in terms of values of OR, temperature (OR = 2.862), strains (OR = 0.641), and food types (OR = 0.561); consequently, temperature is a key factor for stimulating SE expression and had high expression at 30°C. Therefore, food easily contaminated with S. aureus should be monitored intensively at early and late summer, when proper temperature for expressing SEs may result in S. aureus food poisoning prevalence.

Potential of Natural Phenolic Compounds as Antimicrobial Agents against Multidrug-Resistant Staphylococcus aureus in Chicken Meat.

Staphylococcus aureus is one of the most widespread foodborne bacteria that cause high morbidity, mortality, and economic loss, primarily if foodborne diseases are caused by pathogenic and multidrug-resistant (MDR) strains. This study aimed to determine the prevalence of S. aureus in chicken meat in Egyptian markets. Thus, this study might be the first to assess the efficiency of different natural phenolic compounds as novel antibacterial agents against MDR S. aureus pathogens isolated from raw chicken meat in the Egyptian market. The incidence and quantification of pathogenic S. aureus were detected in retail raw chicken meat parts (breast, thigh, fillet, and giblets). In total, 73 out of 80 (91.3%) of the chicken meat parts were contaminated, with S. aureus as the only species isolated. Of the 192 identified S. aureus isolates, 143 were coagulase-positive S. aureus and 117 isolates were MDR (81.8%, 117/143). Twenty-two antibiotic resistance profile patterns were detected. One strain was randomly selected from each pattern to further analyze virulence and resistance genes. Extracted DNA was assessed for the presence of antibiotic-resistance genes, i.e., vancomycin-resistance (vanA), aminoglycosides-resistance (aacA-aphD), apramycin-resistance (apmA), and methicillin-resistance (mecA), penicillin-resistance (blaZ), and virulence genes staphylococcal enterotoxins (sea and seb), Panton-Valentine leucocidin (pvl), clumping factor A (clfA), and toxic shock syndrome toxin (tst). Clustering analyses revealed that six S. aureus strains harbored the most virulence and resistance genes. The activity of hydroquinone was significantly higher than thymol, carvacrol, eugenol, and protocatechuic acid. Therefore, phenolic compounds, particularly hydroquinone, could potentially alternate with conventional antibiotics against the pathogenic MDR S. aureus inhabiting raw chicken meat. Hence, this study indicates that urgent interventions are necessary to improve hygiene for safer meat in Egyptian markets. Moreover, hydroquinone could be a natural phenolic compound for inhibiting foodborne pathogens.

Staphylococcus aureus enterotoxin coding genes identified from patients with atopic dermatitis in Iraq by molecular analysis

The present study determined the frequency of Staphylococcus aureus enterotoxins (B, C, Luk-pv) of atopic dermatitis isolated from (AD) cases in Laboratories of Baghdad. 54 Staphylococcus aureus isolates were subjected to primary identification tests using various methods (cultural characteristics, gram staining, biochemical tests, and vitek2 system). This study used antibiotic disc diffusion in fifty-four S. aureus isolates. Multidrug resistance (MDR) against different antimicrobial agents applied to polymerase chain reaction to amplify different genes coding for Staphylococcal enterotoxins, including 3 types (seb, sec and luk-pv), To ensure that the sequences of these genes match NCBI, DNA sequencing was performed for isolate No. (3). As a result of this study, 25 isolates had a multidrug resistance (MDR) percent (46.2%) against different antimicrobial agents—the results of DNA extraction and polymerase chain reaction directed to amplify the specific enterotoxin coding genes. This study showed that the (seb) gene is present in isolates of staph aureus bacteria isolated from patients with atopic dermatitis 12 /25 at a percentage (48 %). Furthermore, the absence of the sec gene in all Staphylococcus aureus isolates isolated from patients with atopic dermatitis—the results of the detection of the luk-pv 23\25 (92%) gene encoded for lukucidin.Polymerase chain reaction using different primers successfully identified Staphylococcus aureus enterotoxins (B, C)and luk-pv, luk-pv gene, which was the most frequent. Keywords: Staphylococcus aureus enterotoxin genes, polymerase chain reaction, Staphylococcus aureus, atopic dermatitis

Development of nanobodies against Staphylococcus enterotoxin B through yeast surface display

Staphylococcus enterotoxin B (SEB) is one of the primary virulence factors of Staphylococcus aureus but there is still a lack of targeted drugs. SEB activates immune cells via interacting with MHC-II on antigen-presenting cells, leading to the production of large amounts of pro-inflammatory cytokines. Blocking the interaction between SEB and MHC-II can avert the overactivation of immune cells. Nanobodies are the smallest functional antibodies that can bind stably to antigens. In this study, an ideal approach to obtain specific nanobodies without immunizing camelids was introduced. We constructed a library containing up to 5 × 108 nanobodies, and then screened those targeting SEB by using yeast surface display (YSD) technique and fluorescence-activated cell sorting (FACS). A total of 8 nanobodies with divergent complementarity-determining regions (CDRs) sequences were identified and one candidate Nb8 with high affinity to SEB was isolated. In vitro study demonstrated that Nb8 significantly inhibited SEB-induced inflammatory response. Molecular docking simulation indicated that the unique CDR3 sequence contributed to the binding of Nb8 to the MHC-II binding domain of SEB and accordingly cut off the connection between SEB and MHC-II. Our efforts contributed to the development of specific nanobodies for eliminating the threats of SEB.

The stability of Staphylococcal bacteriophage in presence of local vancomycin concentrations used in clinical practice.

To evaluate the stability of a clinically used Staphylococcal bacteriophage with doses of vancomycin that are encountered with local administration of vancomycin for musculoskeletal infections. A Staphylococcal bacteriophage was evaluated for stability in different pH ranges. Then that same bacteriophage was evaluated for stability with different concentrations of vancomycin and with vancomycin biodegradable antibiotic beads. The bacteriophage had stability within a pH range of 4-10. There was a statistically significant (P < 0.05) decrease in the amount of bacteriophage over 24h for vancomycin concentrations of 10mg/mL and 100mg/mL compared to lower vancomycin concentrations (1mg/mL, 0.1mg/mL and normal saline). However, no statistically significant decrease in the amount of bacteriophage was seen with biodegradable vancomycin beads over 24h. These findings have important clinical ramifications in that they show local administration of bacteriophages with concomitant local vancomycin powder therapy should be avoided. Moreover, these findings should spearhead further research into bacteriophage stability in in vivo environments.

Designing a novel chimeric multi-epitope vaccine subunit against Staphylococcus argenteus through artificial intelligence approach integrating pan-genome analysis, in vitro identification, and immunogenicity profiling

Staphylococcus argenteus is a newly identified pathogen that causes respiratory tract infections, skin infections, such as cellulitis, abscesses, and impetigo, and currently, there is no licensed vaccine available against it. To develop a vaccine against S. argenteus, a bacterial pan-genome analysis was applied to identify potential vaccine candidates. A total of 4908 core proteins were retrieved and utilized for identifying four proteins, including SG38 Panton-Valentine leukocidin LukS-PV protein, SG62 staphylococcal enterotoxin type A protein, SG39 enterotoxin B protein, and SG43 enterotoxin type C3 protein as potential vaccine candidates. Epitopes were predicted for these proteins using different types of B and T-cell epitope prediction tools, and only those with a non-toxic profile, antigenic, non-allergenic, and immunogenic were selected. The selected epitopes were linked to each other to form a multi-epitope vaccine construct, which was further linked to the PADRE sequence (AKFVAAWTLKAAA) and 50s ribosomal L7/L12 protein to enhance the vaccine’s antigenicity. The three-dimensional structure of the vaccine construct was assessed to determine its binding affinity with key Toll-like receptor 9 (TLR-9) and Toll-like receptor 5 (TLR-5) immune cell receptors. Our findings demonstrate that the vaccine exhibits favorable binding interactions with these immune cell receptors, indicating its potential efficacy. Molecular dynamic simulations further confirmed the accessibility of vaccine epitopes to the host immune system, substantiating its ability to elicit protective immune responses. Taken together, this study highlights the promising candidacy of the modeled vaccine construct for future in vivo and in vitro experimental investigations. Communicated by Ramaswamy H. Sarma

Enterotoxins A and B produced by Staphylococcus aureus increase cell proliferation, invasion and cytarabine resistance in acute myeloid leukemia cell lines

As in the case of cancer, the risk of infection increases when the host's immune system is not working properly. It has been shown that toxins produced by the bacteria responsible for bacterial infections can alter the properties of cancer cells as well as their sensitivity to chemotherapy agents. Staphylococcus aureus (S. aureus) is one of the most prevalent pathogens in acute myeloid leukemia (AML) patients and it produces several virulence factors, including Staphylococcal enterotoxin A (SEA) and Staphylococcal enterotoxin B (SEB). Cytotoxicity, transwell migration, invasion assays, and various transcriptomic and gene set enrichment (GSE) analyses were used to determine how SEA and SEB alter cell proliferation, migration, invasion, and Cytarabine (Cyt) resistance in AML cell lines. The treatment of AML cell lines with SEA/SEB caused an increase in cell proliferation and Cyt resistance. Toxins enhanced the proclivity of cells to migrate and invade, with around 50% of cells in the presence of SEA and SEB. Transcriptomic and gene set enrichment analyses, and subsequent PCR validations showed dysregulation of immune related genes and genesets. Apparently, this allows AML cells to escape and survive the undesirable environment created by toxins, possibly via the ER stress signaling pathway. Therefore, SEA and SEB can significantly alter the characteristics of AML cancer cells and evaluation of alterations in responsible immune genes and pathways may be crucial for controlling the progression of cancer. In addition, our results suggest that there may be a strong interaction between the immune related pathways and the ER signaling pathway.

Photoactivated Gallium Porphyrin Reduces Staphylococcus aureus Colonization on the Skin and Suppresses Its Ability to Produce Enterotoxin C and TSST-1.

Staphylococcus aureus is a key pathogen in atopic dermatitis (AD) pathogenicity. Over half of AD patients are carriers of S. aureus. Clinical isolates derived from AD patients produce various staphylococcal enterotoxins, such as staphylococcal enterotoxin C or toxic shock syndrome toxin. The production of these virulence factors is correlated with more severe AD. In this study, we propose cationic heme-mimetic gallium porphyrin (Ga3+CHP), a novel gallium metalloporphyrin, as an anti-staphylococcal agent that functions through dual mechanisms: a light-dependent mechanism (antimicrobial photodynamic inactivation, aPDI) and a light-independent mechanism (suppressing iron metabolism). Ga3+CHP has two additive quaternary ammonium groups that increase its water solubility. Furthermore, Ga3+CHP is an efficient generator of singlet oxygen and can be recognized by heme-target systems such as Isd, which improves the intracellular accumulation of this compound. Ga3+CHP activated with green light effectively reduced the survival of clinical S. aureus isolates derived from AD patients (>5 log10 CFU/mL) and affected their enterotoxin gene expression. Additionally, there was a decrease in the biological functionality of studied toxins regarding their superantigenicity. In aPDI conditions, there was no pronounced toxicity in HaCaT keratinocytes with both normal and suppressed filaggrin gene expression, which occurs in ∼50% of AD patients. Additionally, no mutagenic activity was observed. Green light-activated gallium metalloporphyrins may be a promising chemotherapeutic to reduce S. aureus colonization on the skin of AD patients.

Выявление стафилококкового энтеротоксина типа в в молочной продукции иммунохроматографией с визуальной и видеоцифровой детекцией

Detection of staphylococcal enterotoxins in food products is an important task of food poisoning prevention. The study was aimed to develop immunochromatography tests (ICTs) for detection of staphylococcal enterotoxins А (SEA) and be В (SEB), as well as to improve sensitivity of immunochromatography detection of staphylococcal enterotoxins (by the example of SEB) in dairy products relative to visual assessment by recording the analysis results with digital video recorders (DVR) using the principle of processing digital immunochromatogram images acquired using illumination in various spectral ranges. ICTs for detection of enterotoxins were designed as sandwich tests based on highly specific monoclonal antibodies (MABs) against staphylococcal enterotoxins. Milk, cream, sour cream, cheese artificially contaminated with SEB were analyzed. The analysis results were recorded visually or by DVR. DVR of immunochromatograms of the enterotoxin-containing dairy products acquired using illumination with white light in the wavelength range of 400–800 nm ensures a 4-fold increase in the SEB detection sensitivity, while that involving illumination with green light in the wavelength range having its maximum at 525 nm ensures a 4–8-fold increase relative to visual recording. The use of the “Reflecom” and “Zondazh” digital video immunochromatogram analyzers multiplies sensitivity of SEB detection by immunochromatography when assessing dairy products relative to visual recording.