Phenome-wide association study identifies new clinical phenotypes associated with Staphylococcus aureus infections.

BACKGROUND: Phenome-Wide Association study (PheWAS) is a powerful tool designed to systematically screen clinical observations derived from medical records (phenotypes) for association with a variable of interest. Despite their usefulness, no systematic screening of phenotypes associated with Staphylococcus aureus infections (SAIs) has been done leaving potential novel risk factors or complications undiscovered. METHOD AND COHORTS: We tailored the PheWAS approach into a two-stage screening procedure to identify novel phenotypes correlating with SAIs. The first stage screened for co-occurrence of SAIs with other phenotypes within medical records. In the second stage, significant findings were examined for the correlations between their age of onset with that of SAIs. The PheWAS was implemented using the medical records of 754,401 patients from the Marshfield Clinic Health System. Any novel associations discovered were subsequently validated using datasets from TriNetX and All of Us, encompassing 109,884,571 and 118,538 patients respectively. RESULTS: Forty-one phenotypes met the significance criteria of a p-value < 3.64e-5 and odds ratios of > 5. Out of these, we classified 23 associations either as risk factors or as complications of SAIs. Three novel associations were discovered and classified either as a risk (long-term use of aspirin) or complications (iron deficiency anemia and anemia of chronic disease). All novel associations were replicated in the TriNetX cohort. In the All of Us cohort, anemia of chronic disease was replicated according to our significance criteria. CONCLUSIONS: The PheWAS of SAIs expands our understanding of SAIs interacting phenotypes. Additionally, the novel two-stage PheWAS approach developed in this study can be applied to examine other disease-disease interactions of interest. Due to the possibility of bias inherent in observational data, the findings of this study require further investigation.

Transcriptional markers classifying Escherichia coli and Staphylococcus aureus induced sepsis in adults: A data-driven approach.

Sepsis is a life-threatening condition mainly caused by gram-negative and gram-positive bacteria. Understanding the type of causative agent in the early stages is essential for precise antibiotic therapy. This study sought to identify a host gene set capable of distinguishing between sepsis induced by gram-negative bacteria; Escherichia coli and gram-positive bacteria; Staphylococcus aureus in community-onset adult patients. In the present study, microarray expression information was used to apply the Least Absolute Shrinkage and Selection Operator (Lasso) technique to select the predictive gene set for classifying sepsis induced by E. coli or S. aureus pathogens. We identified 25 predictive genes, including LILRA5 and TNFAIP6, which had previously been associated with sepsis in other research. Using these genes, we trained a logistic regression classifier to distinguish whether a sample contains an E. coli or S. aureus infection or belongs to a healthy control group, and subsequently assessed its performance. The classifier achieved an Area Under the Curve (AUC) of 0.96 for E. coli and 0.98 for S. aureus-induced sepsis, and perfect discrimination (AUC of 1) for healthy controls from the other conditions in a 10-fold cross-validation. The genes demonstrated an AUC of 0.75 in distinguishing between sepsis patients with E. coli and S. aureus pathogens. These findings were further confirmed in two distinct independent validation datasets which gave high prediction AUC ranging from 0.72-0.87 and 0.62 in distinguishing three groups of participants and two groups of patients respectively. These genes were significantly enriched in the immune system, cytokine signaling in immune system, innate immune system, and interferon signaling. Transcriptional patterns in blood can differentiate patients with E. coli-induced sepsis from those with S. aureus-induced sepsis. These diagnostic markers, upon validation in larger trials, may serve as a foundation for a reliable differential diagnostics assay.

MicroRNA-939 amplifies Staphylococcus aureus-induced matrix metalloproteinase expression in atopic dermatitis.

Background: Atopic dermatitis (AD) is a common chronic inflammatory skin diseases that seriously affects life quality of the patients. Staphylococcus aureus (S. aureus) colonization on the skin plays an important role in the pathogenesis of AD; however, the mechanism of how it modulates skin immunity to exacerbate AD remains unclear. MicroRNAs are short non-coding RNAs that act as post-transcriptional regulators of genes. They are involved in the pathogenesis of various inflammatory skin diseases. Methods: In this study, we established miRNA expression profiles for keratinocytes stimulated with heat-killed S. aureus (HKSA). The expression of miR-939 in atopic dermatitis patients was analyzed by fluorescence in situ hybridization (FISH). miR-939 mimic was transfected to human primary keratinocyte to investigate its impact on the expression of matrix metalloproteinase genes (MMPs) in vitro. Subsequently, miR-939, along with Polyplus transfection reagent, was administered to MC903-induced atopic dermatitis skin to assess its function in vivo. Results: MiR-939 was highly upregulated in HKSA-stimulated keratinocytes and AD lesions. In vitro studies revealed that miR-939 increased the expression of matrix metalloproteinase genes, including MMP1, MMP3, and MMP9, as well as the cell adhesion molecule ICAM1 in human primary keratinocytes. In vivo studies indicated that miR-939 increased the expression of matrix metalloproteinases to promote the colonization of S. aureus and exacerbated S. aureus-induced AD-like skin inflammation. Conclusions: Our work reveals miR-939 is an important regulator of skin inflammation in AD that could be used as a potential therapeutic target for AD.

Integrated transcriptomic analysis reveals immune signatures distinguishing persistent versus resolving outcomes in MRSA bacteremia.

Introduction: Staphylococcus aureus bacteremia (SAB) is a life-threatening infection particularly involving methicillin-resistant S. aureus (MRSA). In contrast to resolving MRSA bacteremia (RB), persistent MRSA bacteremia (PB) blood cultures remain positive despite appropriate antibiotic treatment. Host immune responses distinguishing PB vs. RB outcomes are poorly understood. Here, integrated transcriptomic, IL-10 cytokine levels, and genomic analyses sought to identify signatures differentiating PB vs. RB outcomes. Methods: Whole-blood transcriptomes of propensity-matched PB (n=28) versus RB (n=30) patients treated with vancomycin were compared in one independent training patient cohort. Gene expression (GE) modules were analyzed and prioritized relative to host IL-10 cytokine levels and DNA methyltransferase-3A (DNMT3A) genotype. Results: Differential expression of T and B lymphocyte gene expression early in MRSA bacteremia discriminated RB from PB outcomes. Significant increases in effector T and B cell signaling pathways correlated with RB, lower IL-10 cytokine levels and DNMT3A heterozygous A/C genotype. Importantly, a second PB and RB patient cohort analyzed in a masked manner demonstrated high predictive accuracy of differential signatures. Discussion: Collectively, the present findings indicate that human PB involves dysregulated immunity characterized by impaired T and B cell responses associated with excessive IL-10 expression in context of the DNMT3A A/A genotype. These findings reveal distinct immunologic programs in PB vs. RB outcomes, enable future studies to define mechanisms by which host and/or pathogen drive differential signatures and may accelerate prediction of PB outcomes. Such prognostic assessment of host risk could significantly enhance early anti-infective interventions to avert PB and improve patient outcomes.

Long non-coding RNA (CMR) involved in autoprotection in S. aureus mastitis in dairy cows by regulating miR-877/FOXM1.

Long non-coding RNAs (LncRNAs) are dysregulated in a variety of human diseases and are highly involved in the development and progression of tumors. Studies on lncRNAs associated with cow mastitis have been lagging behind compared to humans or model animals, therefore, the aim of this study was to explore the mechanism of LncRNAs (CMR) involved in autoprotection against S. aureus mastitis in Bovine Mammary Epithelial Cells (BMECs). First, qRT-PCR was used to examine the relative expression of CMR in a S. aureus mastitis model of BMECs. Then, cell proliferation and apoptosis were detected by EdU and apoptosis assay. Finally, the targeting relationship between miRNAs and mRNA/LncRNAs was determined by dual luciferase reporter gene, qRT-PCR and western blotting techniques. The results showed that CMR was upregulated in the S. aureus mastitis model of BMECs and promoted the expression of inflammatory factors, and SiRNA-mediated CMR inhibited the proliferation of mammary epithelial cells and induced apoptosis. Mechanistically, CMR acts as a competitive endogenous RNA (ceRNA) sponge miR-877, leading to upregulation of FOXM1, a target of miR-877. Importantly, either miR-877 overexpression or FOXM1 inhibition abrogated CMR knockdown-induced apoptosis promoting cell proliferation and reducing inflammatory factor expression levels. In summary, CMR is involved in the regulation of autoprotection against S. aureus mastitis through the miR-877/FOXM1 axis in BMECs and induces immune responses in mammary tissues and cells of dairy cows, providing an important reference for subsequent prevention and control of cow mastitis and the development of targeted drugs.

Collaborative Cross mice have diverse phenotypic responses to infection with Methicillin-resistant Staphylococcus aureus USA300.

Staphylococcus aureus (S. aureus) is an opportunistic pathogen causing diseases ranging from mild skin infections to life threatening conditions, including endocarditis, pneumonia, and sepsis. To identify host genes modulating this host-pathogen interaction, we infected 25 Collaborative Cross (CC) mouse strains with methicillin-resistant S. aureus (MRSA) and monitored disease progression for seven days using a surgically implanted telemetry system. CC strains varied widely in their response to intravenous MRSA infection. We identified eight 'susceptible' CC strains with high bacterial load, tissue damage, and reduced survival. Among the surviving strains, six with minimal colonization were classified as 'resistant', while the remaining six tolerated higher organ colonization ('tolerant'). The kidney was the most heavily colonized organ, but liver, spleen and lung colonization were better correlated with reduced survival. Resistant strains had higher pre-infection circulating neutrophils and lower post-infection tissue damage compared to susceptible and tolerant strains. We identified four CC strains with sexual dimorphism: all females survived the study period while all males met our euthanasia criteria earlier. In these CC strains, males had more baseline circulating monocytes and red blood cells. We identified several CC strains that may be useful as new models for endocarditis, myocarditis, pneumonia, and resistance to MRSA infection. Quantitative Trait Locus (QTL) analysis identified two significant loci, on Chromosomes 18 and 3, involved in early susceptibility and late survival after infection. We prioritized Npc1 and Ifi44l genes as the strongest candidates influencing survival using variant analysis and mRNA expression data from kidneys within these intervals.

DNA methylation haplotype block signatures responding to Staphylococcus aureus subclinical mastitis and association with production and health traits.

BACKGROUND: DNA methylation has been documented to play vital roles in diseases and biological processes. In bovine, little is known about the regulatory roles of DNA methylation alterations on production and health traits, including mastitis. RESULTS: Here, we employed whole-genome DNA methylation sequencing to profile the DNA methylation patterns of milk somatic cells from sixteen cows with naturally occurring Staphylococcus aureus (S. aureus) subclinical mastitis and ten healthy control cows. We observed abundant DNA methylation alterations, including 3,356,456 differentially methylated cytosines and 153,783 differential methylation haplotype blocks (dMHBs). The DNA methylation in regulatory regions, including promoters, first exons and first introns, showed global significant negative correlations with gene expression status. We identified 6435 dMHBs located in the regulatory regions of differentially expressed genes and significantly correlated with their corresponding genes, revealing their potential effects on transcriptional activities. Genes harboring DNA methylation alterations were significantly enriched in multiple immune- and disease-related pathways, suggesting the involvement of DNA methylation in regulating host responses to S. aureus subclinical mastitis. In addition, we found nine discriminant signatures (differentiates cows with S. aureus subclinical mastitis from healthy cows) representing the majority of the DNA methylation variations related to S. aureus subclinical mastitis. Validation of seven dMHBs in 200 cows indicated significant associations with mammary gland health (SCC and SCS) and milk production performance (milk yield). CONCLUSIONS: In conclusion, our findings revealed abundant DNA methylation alterations in milk somatic cells that may be involved in regulating mammary gland defense against S. aureus infection. Particularly noteworthy is the identification of seven dMHBs showing significant associations with mammary gland health, underscoring their potential as promising epigenetic biomarkers. Overall, our findings on DNA methylation alterations offer novel insights into the regulatory mechanisms of bovine subclinical mastitis, providing further avenues for the development of effective control measures.

Genomic characteristics of cfr and fexA carrying Staphylococcus aureus isolated from pig carcasses in Korea.

The emergence of transferable linezolid resistance genes poses significant challenges to public health, as it does not only confer linezolid resistance but also reduces susceptibility to florfenicol, which is widely used in the veterinary field. This study evaluated the genetic characteristics of linezolid-resistant Staphylococcus aureus strains isolated from pig carcasses and further clarified potential resistance and virulence mechanisms in a newly identified sequence type. Of more than 2500 strains isolated in a prior study, 15 isolated from pig carcasses exhibited linezolid resistance (minimum inhibitory concentration ≥ 8 mg/L). The strains were characterized in detail by genomic analysis. Linezolid-resistant S. aureus strains exhibited a high degree of genetic lineage diversity, with one strain (LNZ_R_SAU_64) belonging to ST8004, which has not been reported previously. The 15 strains carried a total of 21 antibiotic resistance genes, and five carried mecA associated with methicillin resistance. All strains harbored cfr and fexA, which mediate resistance to linezolid, phenicol, and other antibiotics. Moreover, the strains carried enterotoxin gene clusters, including the hemolysin, leukotoxin, and protease genes, which are associated with humans or livestock. Some genes were predicted to be carried in plasmids or flanked by ISSau9 and the transposon Tn554, thus being transmittable between staphylococci. Strains carrying the plasmid replicon repUS5 displayed high sequence similarity (99%) to the previously reported strain pSA737 in human clinical samples in the United States. The results illustrate the need for continuous monitoring of the prevalence and transmission of linezolid-resistant S. aureus isolated from animals and their products.

Histidine transport is essential for the growth of Staphylococcus aureus at low pH.

Staphylococcus aureus is an opportunistic pathogen capable of causing many different human diseases. During colonization and infection, S. aureus will encounter a range of hostile environments, including acidic conditions such as those found on the skin and within macrophages. However, little is known about the mechanisms that S. aureus uses to detect and respond to low pH. Here, we employed a transposon sequencing approach to determine on a genome-wide level the genes required or detrimental for growth at low pH. We identified 31 genes that were essential for the growth of S. aureus at pH 4.5 and confirmed the importance of many of them through follow up experiments using mutant strains inactivated for individual genes. Most of the genes identified code for proteins with functions in cell wall assembly and maintenance. These data suggest that the cell wall has a more important role than previously appreciated in promoting bacterial survival when under acid stress. We also identified several novel processes previously not linked to the acid stress response in S. aureus. These include aerobic respiration and histidine transport, the latter by showing that one of the most important genes, SAUSA300_0846, codes for a previously uncharacterized histidine transporter. We further show that under acid stress, the expression of the histidine transporter gene is increased in WT S. aureus. In a S. aureus SAUSA300_0846 mutant strain expression of the histidine biosynthesis genes is induced under acid stress conditions allowing the bacteria to maintain cytosolic histidine levels. This strain is, however, unable to maintain its cytosolic pH to the same extent as a WT strain, revealing an important function specifically for histidine transport in the acid stress response of S. aureus.

High prevalence of heteroresistance in Staphylococcus aureus is caused by a multitude of mutations in core genes.

Heteroresistance (HR) is an enigmatic phenotype where, in a main population of susceptible cells, small subpopulations of resistant cells exist. This is a cause for concern, as this small subpopulation is difficult to detect by standard antibiotic susceptibility tests, and upon antibiotic exposure the resistant subpopulation may increase in frequency and potentially lead to treatment complications or failure. Here, we determined the prevalence and mechanisms of HR for 40 clinical Staphylococcus aureus isolates, against 6 clinically important antibiotics: daptomycin, gentamicin, linezolid, oxacillin, teicoplanin, and vancomycin. High frequencies of HR were observed for gentamicin (69.2%), oxacillin (27%), daptomycin (25.6%), and teicoplanin (15.4%) while none of the isolates showed HR toward linezolid or vancomycin. Point mutations in various chromosomal core genes, including those involved in membrane and peptidoglycan/teichoic acid biosynthesis and transport, tRNA charging, menaquinone and chorismite biosynthesis and cyclic-di-AMP biosynthesis, were the mechanisms responsible for generating the resistant subpopulations. This finding is in contrast to gram-negative bacteria, where increased copy number of bona fide resistance genes via tandem gene amplification is the most prevalent mechanism. This difference can be explained by the observation that S. aureus has a low content of resistance genes and absence of the repeat sequences that allow tandem gene amplification of these genes as compared to gram-negative species.

Enhanced Biofilm Formation by Tetracycline in a Staphylococcus aureus Naturally Lacking ica Operon and atl.

Staphylococcus aureus is a major, widespread pathogen, and its biofilm-forming characteristics make it even more difficult to eliminate by biocides. Tetracycline (TCY) is a major broad-spectrum antibiotic, the residues of which can cause deleterious health impacts, and subinhibitory concentrations of TCY have the potential to increase biofilm formation in S. aureus. In this study, we showed how the biofilm formation of S. aureus 123786 is enhanced in the presence of TCY at specific subinhibitory concentrations. S. aureus 123786 used in this study was identified as Staphylococcal Cassette Chromosome mec III, sequence type239 and naturally lacking ica operon and atl gene. Two assays were performed to quantify the formation of S. aureus biofilm. In the crystal violet (CV) assay, the absorbance values of biofilm stained with CV at optical density (OD)540 nm increased after 8 and 16 hr of incubation when the concentration of TCY was 1/2 minimum inhibitory concentration (MIC), whereas at the concentration of 1/16 MIC, the absorbance values increased after 16 and 24 hr of incubation. In tetrazolium salt reduction assay, the absorbance value at OD490 nm of S. aureus 123786 biofilms mixed with 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium solution increased after 8 hr when the concentration of TCY was 1/4 MIC, which may be correlated with the higher proliferation and maturation of biofilm. In conclusion, the biofilm formation of S. aureus 123786 could be enhanced in the presence of TCY at specific subinhibitory concentrations.

Relationship between biofilm-forming microorganisms (BFM: Staphylococcus aureus and Pseudomonas aeruginosa) and DEFB1 gene variants on ß-defensin levels in periprosthetic joint infection (PJI).

BACKGROUND: The purpose of this study was to investigate the relationship between biofilm-forming microorganisms (BFM) and DEFB1 gene variants on ß-defensin levels in patients with periprosthetic joint infection (PJI) of Mexican origin. METHODS AND RESULTS: One hundred and five clinical aspirates were obtained from patients with suspected PJI. After microbiologic culture, samples were classified as non-septic and septic; of the latter, only those positive for Staphylococcus aureus and Pseudomonas aeruginosa were selected. ß-Defensin levels were quantified by ELISA, DNA was extracted from total leukocytes of the samples, and - 20G > A (rs11362) and - 44 C > G (rs1800972) variants were genotyped using TaqMan probes. Forty-one clinical aspirates were non-septic, 18 were positive for S. aureus and 18 were positive for P. aeruginosa. It was observed that ß-defensin levels were higher in the P. aeruginosa group compared to S. aureus group (2339.0 pg/mL IQR = 1809.2 vs. 1821.3 pg/mL IQR = 1536.4) and non-septic group (2339.0 pg/mL IQR = 1809.2 vs. 1099.7 pg/mL IQR = 1744.5, P < 0.001). The CG genotype of the rs1800972 variant was associated with higher ß-defensin levels compared to the CC genotype for both P. aeruginosa and S. aureus (1905.8 vs. 421.7 pg/mL, P = 0.004; and 1878.2 vs. 256.4 pg/mL, P = 0.006, respectively). CONCLUSIONS: Our results show that ß-defensin levels are significantly elevated in patients with BFM-associated PJI compared to those without infection. Furthermore, carriers of the CG genotype of the rs1800972 variant have an increased risk of PJI. Further research is needed to replicate these findings in a larger population.

Searching for Virulence Factors among Staphylococcus lugdunensis Isolates from Orthopedic Infections: Correlation of ß-hemolysin, hemolysin III, and slush Genes with Hemolytic Activity and Synergistic Hemolytic Activity.

Staphylococcus lugdunensis is an emerging high-virulent pathogen. Here, the presence and expression of virulence genes (icaA, fbl, vwbl, fbpA, slush A, B and C, and genes of the putative ß-hemolysin and hemolysin III) and the ability to induce synergistic hemolytic activity and hemolysis after 24, 48 and 72 h were investigated in a collection of twenty-two S. lugdunensis clinical isolates. The collection of isolates, mainly from implant orthopedic infections, had previously been grouped by ribotyping/dendrogram analysis and studied for biofilm matrices, biomasses and antibiotic resistances. Two isolates, constituting a unique small ribogroup sharing the same cluster, exhibited an amplicon size of the slush operon (S. lugdunensis synergistic hemolysin) which was shorter than the expected 977 bp. This outcome can predict the genetic lineage of the S. lugdunensis strains. One isolate (cra1342) presented two deletions: one of 90 bp in slush A and the other of 91 bp in slush B. Another isolate (N860314) showed a single 193 bp deletion, which encompassed part of the slush B terminal sequence and most of slush C. The isolate N860314 was devoid of hemolytic activity after 24 h, and the first consideration was that the deleted region deals with the coding of the active enzymatic site of the slush hemolysin. On the other hand, cra1342 and N860314 isolates with different slush deletions and with hemolytic activity after 24 and 48 h, respectively, could have replaced the hemolytic phenotype through other processes.

Panton-Valentine Leukocidin (PVL) genes may not be a reliable marker for community-acquired MRSA in the Dakahlia Governorate, Egypt.

BACKGROUND: Methicillin-resistant Staphylococcus aureus is linked to both nosocomial and community infections. One of the key virulence factors of S. aureus is Panton-Valentine leukocidin (PVL). The PVL genes are mostly associated with community-acquired MRSA (CA-MRSA). This study evaluates the prevalence of PVL genes as a marker for CA-MRSA at tertiary hospitals in Mansoura, Dakahlia, Egypt. S. aureus was isolated from clinical specimens obtained from different departments of tertiary hospitals, outpatient clinics, and hospital healthcare workers (HCWs). PCR was used to detect the mecA, PVL, and SCCmec genes among the recovered isolates. Standard broth microdilution method was used to determine the minimum inhibitory concentrations (MIC) of nine antibiotics against S. aureus. RESULTS: Two hundred S. aureus isolates were recovered and identified out of the total isolates (n = 320). The mecA gene was detected in 103 S. aureus isolates (51.5%). Among the MRSA isolates, 46.60% were PVL-positive. The incidence of the PVL genes of MRSA in nosocomial (HA), outpatient clinics (CA), and HCWs was 46.66%, 56.52%, and 42%, respectively. All MRSA isolates showed resistance to cefoxitin. The percentage of resistance to most tested antibiotics was high, except for ciprofloxacin (6.85%). Both antibiotic resistance and multidrug resistance among MRSA isolates were generally higher in PVL-positive isolates than in PVL-negative isolates in HA- and CA-MRSA isolates. While SCCmec type V was the most prevalent in PVL-positive MRSA stains, type I was the most prevalent in PVL-negative isolates. CONCLUSION: This study revealed that PVL genes are generally highly prevalent among mecA-positive MRSA isolates, whether they are CA-MRSA, HA-MRSA, or HCW isolates. Therefore, PVL is not a valid marker for CA-MRSA in Mansoura, Dakahlia Governorate, Egypt, as has been reported in other countries. Further epidemiologic studies are required to track the incidence of PVL in HA-MRSA, CA-MRSA, and HCW isolates in other Egyptian governorates.

Inactivation of the Sts enzymes promotes resistance to lethal Staphylococcus aureus infection.

Staphylococcus aureus is a highly infective Gram-positive bacterial pathogen that causes a wide range of diseases in both healthy and immunocompromised individuals. It can evade host immune defenses by expressing numerous virulence factors and toxins. Coupled with the inability of the human host to develop protective immunity against S. aureus, the emergence of antibiotic-resistant strains complicates treatment options. The non-canonical Sts phosphatases negatively regulate signaling pathways in varied immune cell types. To determine the role of the Sts proteins in regulating host responses to a Gram-positive microorganism, we investigated the response of mice lacking Sts expression to S. aureus infection. Herein, we demonstrate that Sts -/- animals are significantly resistant to lethal intravenous doses of S. aureus strain USA300. Resistance is characterized by significantly enhanced survival and accelerated bacterial clearance in multiple peripheral organs. Infected Sts -/- animals do not display increased levels of cytokines TNFα, IFNγ, and IL-6 in the spleen, liver, and kidney during the early stages of the infection, suggesting that a heightened pro-inflammatory response does not underlie the resistance phenotype. In vivo ablation of mononuclear phagocytes compromises the Sts -/- enhanced CFU clearance phenotype. Additionally, Sts -/- bone marrow-derived macrophages demonstrate significantly enhanced restriction of intracellular S. aureus following ex vivo infection. These results reveal the Sts enzymes to be critical regulators of host immunity to a virulent Gram-positive pathogen and identify them as therapeutic targets for optimizing host anti-microbial responses.

Toxin exposure and HLA alleles determine serum antibody binding to toxic shock syndrome toxin 1 (TSST-1) of Staphylococcus aureus.

Life-threatening toxic shock syndrome is often caused by the superantigen toxic shock syndrome toxin-1 (TSST-1) produced by Staphylococcus aureus. A well-known risk factor is the lack of neutralizing antibodies. To identify determinants of the anti-TSST-1 antibody response, we examined 976 participants of the German population-based epidemiological Study of Health in Pomerania (SHIP-TREND-0). We measured anti-TSST-1 antibody levels, analyzed the colonization with TSST-1-encoding S. aureus strains, and performed a genome-wide association analysis of genetic risk factors. TSST-1-specific serum IgG levels varied over a range of 4.2 logs and were elevated by a factor of 12.3 upon nasal colonization with TSST-1-encoding S. aureus. Moreover, the anti-TSST-1 antibody levels were strongly associated with HLA class II gene loci. HLA-DRB1*03:01 and HLA-DQB1*02:01 were positively, and HLA-DRB1*01:01 as well as HLA-DQB1*05:01 negatively associated with the anti-TSST-1 antibody levels. Thus, both toxin exposure and HLA alleles affect the human antibody response to TSST-1.

A statistical genomics framework to trace bacterial genomic predictors of clinical outcomes in Staphylococcus aureus bacteremia.

Outcomes of severe bacterial infections are determined by the interplay between host, pathogen, and treatments. While human genomics has provided insights into host factors impacting Staphylococcus aureus infections, comparatively little is known about S. aureus genotypes and disease severity. Building on the hypothesis that bacterial pathoadaptation is a key outcome driver, we developed a genome-wide association study (GWAS) framework to identify adaptive mutations associated with treatment failure and mortality in S. aureus bacteremia (1,358 episodes). Our research highlights the potential of vancomycin-selected mutations and vancomycin minimum inhibitory concentration (MIC) as key explanatory variables to predict infection severity. The contribution of bacterial variation was much lower for clinical outcomes (heritability <5%); however, GWASs allowed us to identify additional, MIC-independent candidate pathogenesis loci. Using supervised machine learning, we were able to quantify the predictive potential of these adaptive signatures. Our statistical genomics framework provides a powerful means to capture adaptive mutations impacting severe bacterial infections.

Transcriptome analysis of Macrobrachium rosenbergii hemocytes in response to Staphylococcus aureus infection.

The aquaculture industry has suffered significant financial losses as a result of disease outbreaks. In particular, disease outbreaks have become a major problem that can seriously affect the sustainable development of the Macrobrachium rosenbergii aquaculture industry. It is crucial to determine the defense mechanism of the host after pathogenic invasion in order to provide effective defense measures after disease outbreaks. Shrimp, like other invertebrates, primarily depend on their innate immune systems to defend against pathogens, and recognize and resist pathogens through humoral and cellular immune responses. In this investigation, we used RNA-seq technology to investigate the transcriptome of hemocytes from M. rosenbergii induced by Staphylococcus aureus. Our main targets were immune pathways and genes related to innate immunity. RNA-seq identified 209,069 and 204,775 unigenes in the control and experimental groups, respectively. In addition, we identified 547 and 1734 differentially expressed genes (DEGs) following S. aureus challenge after 6 and 12 h (h), respectively. GO and KEGG enrichment analysis revealed that the DEGs were significantly enriched in several biological signalling pathways, including NOD-like receptor, PI3K-Akt, Toll and Imd, IL-17, TGF-beta, RIG-I-like receptor, cAMP, apoptosis, and C-type lectin receptor. Sixteen DEGs were chosen at random for qPCR verification; these results concurred with those from sequencing. Our findings revealed that immune-related genes play an important role in antibacterial activities and have specific functions for gram-positive bacteria. These results provide more data for the prevention of M. rosenbergii diseases and offer a basis for the better prevention of diseases.

Horizontal transfer of probable chicken-pathogenicity chromosomal islands between Staphylococcus aureus and Staphylococcus agnetis.

Staphylococcus agnetis is an emerging pathogen in chickens but has been most commonly isolated from sub-clinical mastitis in bovines. Previous whole-genome analyses for known virulence genes failed to identify determinants for the switch from mild ductal infections in cattle to severe infections in poultry. We now report identification of a family of 15 kbp, 17-19 gene mobile genetic elements (MGEs) specific to chicken osteomyelitis and dermatitis isolates of S. agnetis. These MGEs can be present in multiple copies per genome. The MGE has been vectored on a Staphylococcus phage that separately lysogenized two S. agnetis osteomyelitis strains. The S. agnetis genome from a broiler breeder case of ulcerative dermatitis contains 2 orthologs of this MGE, not associated with a prophage. BLASTn and phylogenetic analyses show that there are closely related intact MGEs found in genomes of S. aureus. The genome from a 1980s isolate from chickens in Ireland contains 3 copies of this MGE. More recent chicken isolates descended from that genome (Poland 2009, Oklahoma 2010, and Arkansas 2018) contain 2 to 4 related copies. Many of the genes of this MGE can be identified in disparate regions of the genomes of other chicken isolates of S. aureus. BLAST searches of the NCBI databases detect no similar MGEs outside of S. aureus and S. agnetis. These MGEs encode no proteins related to those produced by Staphylococcus aureus Pathogenicity Islands, which have been associated with the transition of S. aureus from human to chicken hosts. Other than mobilization functions, most of the genes in these new MGEs annotate as hypothetical proteins. The MGEs we describe appear to represent a new family of Chromosomal Islands (CIs) shared amongst S. agnetis and S. aureus. Further work is needed to understand the role of these CIs/MGEs in pathogenesis. Analysis of horizontal transfer of genetic elements between isolates and species of Staphylococci provides clues to evolution of host-pathogen interactions as well as revealing critical determinants for animal welfare and human diseases.

Whole-genome sequencing analysis of molecular epidemiology and silent transmissions causing meticillin-resistant Staphylococcus aureus bloodstream infections in a university hospital.

BACKGROUND: The emergence of novel genomic-type clones, such as community-associated meticillin-resistant Staphylococcus aureus (MRSA) and livestock-associated MRSA, and their invasion into hospitals have become major concerns worldwide; however, little information is available regarding the prevalence of MRSA in Japan. Whole-genome sequencing (WGS) has been conducted to analyse various pathogens worldwide. Therefore, it is important to establish a genome database of clinical MRSA isolates available in Japan. AIM: A molecular epidemiological analysis of MRSA strains isolated from bloodstream-infected patients in a Japanese university hospital was conducted using WGS and single-nucleotide polymorphism (SNP) analysis. Additionally, through a review of patients' clinical characteristics, the effectiveness of SNP analysis as a tool for detecting silent nosocomial transmission that may be missed by other methods was evaluated in diverse settings and various time points of detection. METHODS: Polymerase-chain-reaction-based staphylococcal cassette chromosome mec (SCCmec) typing was performed using 135 isolates obtained between 2014 and 2018, and WGS was performed using 88 isolates obtained between 2015 and 2017. FINDINGS: SCCmec type II strains, prevalent in 2014, became rare in 2018, whereas the prevalence of SCCmec type IV strains increased from 18.75% to 83.87% of the population, and became the dominant clones. Clonal complex (CC) 5 CC8 and CC1 were detected between 2015 and 2017, with CC1 being dominant. In 88 cases, SNP analyses revealed nosocomial transmissions among 20 patients which involved highly homologous strains. CONCLUSIONS: Routine monitoring of MRSA by whole-genome analysis is effective not only for gaining knowledge regarding molecular epidemiology, but also for detecting silent nosocomial transmission.