FBXO11 governs macrophage cell death and inflammation in response to bacterial toxins.

Staphylococcus aureus causes severe infections such as pneumonia and sepsis depending on the pore-forming toxin Panton-Valentine leukocidin (PVL). PVL kills and induces inflammation in macrophages and other myeloid cells by interacting with the human cell surface receptor, complement 5a receptor 1 (C5aR1). C5aR1 expression is tighly regulated and may thus modulate PVL activity, although the mechanisms involved remain incompletely understood. Here, we used a genome-wide CRISPR/Cas9 screen and identified F-box protein 11 (FBXO11), an E3 ubiquitin ligase complex member, to promote PVL toxicity. Genetic deletion of FBXO11 reduced the expression of C5aR1 at the mRNA level, whereas ectopic expression of C5aR1 in FBXO11-/- macrophages, or priming with LPS, restored C5aR1 expression and thereby PVL toxicity. In addition to promoting PVL-mediated killing, FBXO11 dampens secretion of IL-1ß after NLRP3 activation in response to bacterial toxins by reducing mRNA levels in a BCL-6-dependent and BCL-6-independent manner. Overall, these findings highlight that FBXO11 regulates C5aR1 and IL-1ß expression and controls macrophage cell death and inflammation following PVL exposure.

Skin Infections Due to Panton-Valentine Leukocidin-Producing S. Aureus.

BACKGROUND: Panton-Valentine leukocidin (PVL)-producing Staphylococcus aureus (PVL-SA) strains are frequently associated with large, recurring abscesses in otherwise healthy young individuals. The typical clinical presentation and the recommended diagnostic evaluation and treatment are not widely known. METHODS: This review is based on pertinent publications retrieved by a selective search in PubMed, with special attention to international recommendations. RESULTS: PVL-SA can cause leukocytolysis and dermatonecrosis through specific cell-wall pore formation. Unlike other types of pyoderma, such conditions caused by PVL-SA have no particular site of predilection. In Germany, the PVL gene can be detected in 61.3% (252/411) of skin and soft tissue infections with S. aureus. Skin and soft tissue infections with PVL-SA recur three times as frequently as those due to PVL-negative S. aureus. They are diagnosed by S. aureus culture from wound swabs and combined nasal/pharyngeal swabs, along with PCR for gene detection. The acute treatment of the skin abscesses consists of drainage, followed by antimicrobial therapy if needed. Important secondary preventive measures include topical cleansing with mupirocin nasal ointment and whole-body washing with chlorhexidine or octenidine. The limited evidence (level IIb) concerning PVL-SA is mainly derived from nonrandomized cohort studies and experimental analyses. CONCLUSION: PVL-SA skin infections are easily distinguished from other skin diseases with targeted history-taking and diagnostic evaluation.

LukS-PV inhibits hepatocellular carcinoma cells migration by downregulating HDAC6 expression.

BACKGROUND: Hepatocellular carcinoma (HCC) is a clinically common malignant tumor worldwide. LukS-PV is the S component of Panton-Valentine leukocidin secreted by Staphylococcus aureus, which has shown anti-cancer activity. Based on previous findings, this study investigated the effects of LukS-PV on HCC migration and the potential molecular mechanisms involving acetylation pathways. METHODS: After treating HCC cells with different concentrations of LukS-PV, we used scratch assays to determine the mobility of the cancer cells. Western blots were used to determine the expression levels of migration-related proteins. Quantitative proteomic sequencing was used to evaluate proteomic changes in target proteins. Immunoprecipitation and liquid chromatography coupled with tandem mass spectrometry analyses were used to validate the binding of related target proteins. RESULTS: LukS-PV inhibited HCC cell migration in a concentration-dependent manner. In addition, LukS-PV attenuated the expression of histone deacetylase (HDAC)6, which is highly expressed in HCC cells. Further studies showed that LukS-PV increased the acetylation level of α-tubulin by down-regulating HDAC6, which resulted in the inhibition of HCC cell migration. CONCLUSION: Taken together, our data revealed a vital role of LukS-PV in suppressing HCC cell migration by down-regulating HDAC6 and increasing the acetylation level of α-tubulin.

Regulatory relationship between macrophage autophagy and PVL-positive methicillin-resistant Staphylococcus aureus.

The present study intends to clarify the hypothesis that PVL-positive Methicillin-resistant S. aureus strain (PVL+-MRSA)-infected macrophages regulate autophagy and thus in turn inhibit phagocytosis through the in vitro and in vivo experiments. The autophagy of mouse macrophage cell line RAW264.7 was observed by fluorescence microscopy, and counted based on the number of each cell dot-like structure GFP-LC3. The protein levels of the phagocytic factors associated with autophagy were determined by western blotting. The phagocytosis of RAW264.7 on MRSA was determined by counting the colony. The clinically isolated and identified PVL+-MRSA strain was used to infect BALB/c mice (left nasal drip) to establish a mouse pneumonia model. PVL+-MRSA mice were then treated with 3-MA or linezolid. Bronchoalveolar lavage fluid (BALF) from mice was collected for macrophage counting by Flow cytometry assay. The right lung was aseptically isolated for counting the amount of bacteria. The results showed that PVL+-MRSA could induced the autophagy of macrophages, which in turn reduced the damage from macrophages, which were respectively alleviated by 3-MA and aggravated by rapamycin. Exogenous rPVL administrated into PVL--MRSA-infected macrophages caused the autophagy of macrophage. Exogenous rPVL, particularly A-Luk S-PV, administrated into macrophages also caused the autophagy of macrophage, which was reversed by PMX53, a C5aR antagonist. In a mouse pneumonia model, PVL+-MRSA could induced the autophagy of macrophages, which in turn reduced the damage from macrophages, which were respectively alleviated by 3-MA or linezolid. In conclusion, this study indicated PVL+-MRSA regulated macrophage autophagy, which in turns inhibit the phagocytosis of S. aureus by macrophage. This study may provide a potential target against S. aureus infection.

LukS-PV inhibits the proliferation of hepatocellular carcinoma cells by maintaining FOXO3 stability via the PI3K/AKT signaling pathway.

Hepatocellular carcinoma(HCC) is one of the common malignant tumors. LukS-PV is the S component of Panton-Valetine leukocidin(PVL) secreted by Staphylococcus aureus. Forkhead box O3 (FOXO3) is a member of the FOXO subfamily of transcription factors that acts as a tumor suppressor. In this study, we investigated the role of LukS-PV on the proliferation of HCC and explored possible mechanisms. We treated HCC cells with various concentrations of LukS-PV and evaluated the effect of LukS-PV on cell viability using the cell counting kit-8 and colony formation assays. Real-time PCR and western blot assays were used to analyze mRNA and protein expression levels, respectively. Immunofluorescence staining was performed to examine the intracellular localization of FOXO3. The expression of FOXO3 and its downstream target genes were analyzed by immunohistochemical staining. The protein synthesis inhibitor cycloheximide and the proteosome inhibitor MG132 were used to explore the potential mechanisms by which LukS-PV regulated FOXO3. We demonstrated that LukS-PV inhibited the proliferation of HCC cells in a concentration dependent manner. LukS-PV upregulated FOXO3 expression both in vitro and in vivo. Moreover, LukS-PV facilitated the entry of FOXO3 into the nucleus and, subsequently, regulated the transcription of downstream target genes. In addition, we discovered that LukS-PV decreased the expression of phosphorylated FOXO3 through the PI3K/AKT signaling pathway and maintained FOXO3 protein stability via the ubiquitin-proteasome pathway. Taken together, our data indicated that LukS-PV exert anticancer activities through FOXO3. LukS-PV may be a promising candidate for HCC treatment.

Paediatric osteoarticular infections caused by staphylococcus aureus producing panton-valentine leucocidin in morocco: Risk factors and clinical features.

OBJECTIVE: We aimed to estimate the prevalence of Staphylococcus aureus producing Panton-Valentine leucocidin (PVL) isolated from children diagnosed with osteoarticular infections (OAIs), and to examine risk factors and clinical features. METHODS: This prospective study was conducted from January 2017 to December 2018. All hospitalised children diagnosed with S. aureus OAI are included. Blood cultures, articular fluids, synovial tissues and/or bone fragments were collected for bacteriological culture. Antimicrobial susceptibility tests were determined by disk diffusion method. Genes encoding methicillin resistance (mecA) and PVL virulence factors (luk-S-PV and luk-F-PV) were detected by multiplex polymerase chain reaction. The demographic, clinical, laboratory, radiographic and clinical features were reviewed prospectively from medical records. RESULTS: A total of 37 children with S. aureus OAIs were included, 46% of them have PVL-positive infection and 70.6% were male. The mean age was 8.12 years (±4.57), and almost were from rural settings (76.5%). Children with Staphylococcus aureus producing Panton-Valentine leucocidin (SA-PVL) were significantly associated with type of infection (P = 0.005), location of infection (P = 0.037) and abnormal X-ray (P = 0.029). All strains SA-PVL+ are sensitive to methicillin, but one strain SA-PVL negative was methicillin-resistant S. aureus, confirmed by gene mecA positive. CONCLUSION: The prevalence of S. aureus infections producing PVL toxin was high in OAIs amongst Moroccan children, mainly due to methicillin-susceptible S. aureus. Type and location of infections and abnormal X-ray were significantly associated with SA-PVL. Routine diagnostic testing of PVL-SA, continuous epidemiological surveillance and multidisciplinary management of OAI is essential to prevent serious complications.

A case report: septic shock due to (tropical) pyomyositis and multiple metastatic embolisms caused by Panton Valentine Leukocidin-positive methicillin-sensitive staphylococcus aureus in a 12-year-old boy.

CASE REPORT: A 12-year-old boy, of Congolese roots and without medical history, first presented to our Emergency Department 3 days after blunt trauma of the left ankle. The boy represented on two more occasions in the next 3 days due to ongoing pain. On the last occasion he presented with severe hypoglycaemia. He was diagnosed with severe septic shock, secondary to subperiosteal abscess formation / osteomyelitis of the ankle. The patient was transferred to the paediatric intensive care unit where appropriate medical care was provided, including broad-spectrum antibiotic therapy, high dose vasopressor / inotropic support, surgical debridement of abscesses and below-knee amputation. PANTON VALENTINE LEUKOCIDIN TOXIN AND PYOMYOSITIS TROPICALIS: The causative organism was a methicillin-susceptible S. aureus, which upon further identification was a carrier of the PVL (Panton Valentine leukocidin) toxin. This pathogen is responsible for severe musculoskeletal infections. In children these infections are often associated with more severe clinical course requiring a higher need for surgical intervention and longer hospital stay.Tropical pyomyositis is a disease caused by Staphylococcus aureus, often seen in tropical countries, and classically presented with muscle abscesses. Young males between the ages of 10-40 years old are the most susceptible, and often present with a history of blunt trauma. Treatment generally requires a combination of an anti-staphylococcal agent, and an anti-toxic agent blocking bacterial protein-synthesis of PVL. Source control by surgical debridement also plays a major role in the treatment of PVL-infection. Despite agressive treatment, mortality still varies from 0.5% to 2%.

Molecular analysis of an increase in trimethoprim/sulfamethoxazole-resistant MRSA reveals multiple introductions into a tertiary care hospital, Germany 2012-19.

OBJECTIVES: Increasing spread of resistance could jeopardize the use of antifolates against MRSA infections. METHODS: We compared the prevalence of phenotypic trimethoprim/sulfamethoxazole resistance in 20 534 clinical Staphylococcus aureus isolates (19 096 MSSA and 1438 MRSA) of non-redundant patients at Heidelberg University Hospital over 8 years and performed WGS on trimethoprim/sulfamethoxazole-resistant MRSA. RESULTS: From 2012 to 2019, trimethoprim/sulfamethoxazole resistance in MSSA (674/19 096; 3.5%) ranged between 1.5% and 7.2% and in MRSA (135/1438; 9.4%) between 0.5% and 20.2%, reaching a peak in 2016 and 2018, respectively (Ptrend < 0.001). Trimethoprim/sulfamethoxazole resistance was more likely in outpatients than inpatients (P = 0.005), younger patients (P < 0.001), skin and soft tissue infections (SSTIs) (MRSA only, P = 0.05), submissions from pulmonology (MRSA only, P = 0.001), the upper respiratory tract (MSSA only, P < 0.001) and general surgery (MSSA only, P = 0.001). WGS of 76 trimethoprim/sulfamethoxazole-resistant MRSA revealed that 59% belonged to major pandemic CA-MRSA clones (ST22, ST8, ST398, ST772, ST30), 47% harboured Panton-Valentine leucocidin (PVL), 97% SCCmec IV/V, 71% dfrG and 28% dfrA. SNP-based phylogeny of trimethoprim/sulfamethoxazole-resistant MRSA core genomes favoured independent introduction over clonal expansion as the source, most prominently of dfrA+ trimethoprim/sulfamethoxazole-resistant ST22 MRSA from the Gaza Strip. CONCLUSIONS: The presented results support that trimethoprim/sulfamethoxazole-resistant S. aureus, formerly associated with SSTI from outpatients and S. aureus in the (sub)tropics, is on the rise in the temperate zone, potentially due to migration. Closer monitoring of trimethoprim/sulfamethoxazole resistance in S. aureus is recommended to safeguard the effectiveness of antifolate compounds.

Changing characteristics of S. aureus bacteremia caused by PVL-negative, MRSA strain over 11 years.

Community-acquired methicillin-resistant Staphylococcus aureus (MRSA) has emerged as an important cause of infection. We conducted a longitudinal study to evaluate changes in clinical and microbiological characteristics as well as outcomes of sequence type (ST) 72 MRSA bacteremia. We reviewed adult patients enrolled in a prospective cohort with ST72 MRSA bacteremia from August 2008 to December 2018 at Asan Medical Center, Seoul, South Korea. Changes in clinical characteristics, outcomes, and microbiological characteristics of patients over time were evaluated. Generalized linear and linear regression models were used to evaluate changes. Of the 1,760 isolates, 915 (62%) were MRSA bacteremia and 292 (31.9%) were ST72 MRSA. During the study period, the relative risk (RR) of MRSA bacteremia decreased annually by 3.7%; however, among MRSA bacteremia, RR of ST72 MRSA increased annually by 8.5%. Vancomycin minimum inhibitory concentration (MIC) decreased over the study period. Metastatic infection, persistent bacteremia, and recurrence of bacteremia within 12 weeks decreased significantly. There were no significant changes in 30-d and 12-week mortality. Antibiotic susceptibility of ST72 MRSA was evaluated, and the resistance rate to erythromycin decreased significantly. ST72 MRSA incidence increased annually; its vancomycin MIC and erythromycin resistance rate decreased over the 11 years.

Molecular characteristics of Staphylococcus aureus strains isolated from nasal samples of sixth year medical students during their pediatric services practices.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) strains are prevalent in healthcare services. Medical students are at risk for MRSA carriage, subsequent infection and potential transmission of nosocomial infection.Few studies have examined MRSA carriage among medical students. METHODS: In this prospective cohort study, between July 2016 and June 2017, two nasal swab samples were taken per student 4 weeks apart during their pediatric internship. MRSA typing was performed by staphylococcal cassette chromosome mec (SCCmec) types, Panton Valentine leukocidin (PVL) encoding genes. RESULTS: A total of 239 sixth year medical students, 164 (68.6%) male (M/F:2.1),with median age 25 years (min-max; 23-65 years) were included in this prospective cohort study. Among 239 students, 17 students (7.1%) were found to be colonized with methicillin-sensitive S. aureus (MSSA) at the beginning of pediatric internship. After 4 weeks, at the end of pediatric internship totally 52 students were found to be S. aureus colonized (21.8%). Three of 52 S. aureus isolates were MRSA (1.3%) and the rest was MSSA (20.5%), all were PVL gen negative. Two of three MRSA isolates were characterized as SCCmec type IV, one isolate was untypeable SCCmec. Nasal carriage of S. aureus increased from 7.1% to 21.5% (p < 0.001). Nasal S. aures colonization ratio was higher in students working in pediatric infectious disease service (p = 0.046). Smoking was found to be associated with a 2.37-fold [95% CI (1.12-5.00); p = 0.023] and number of patients in pediatric services was 2.66-fold [95% CI (1.13-6.27); p = 0.024] increase the risk of nasal S. aureus colonization. Gender was not found to increase risk of MRSA carriage. CONCLUSION: MSSA nasal carriage increased at the end of pediatric internship and significantly high in students working in pediatric infectious diseases services. Smoking and high number of patients in pediatric services significantly increase S.aureus colonization.

Staphylococcus aureus sensible a meticilina, productor de leucocidina de Panton Valentine: a propósito de dos casos pediátricos de infección osteoarticular / Panton-Valentine leukocidin toxin associated methicillin-susceptible Staphylococcus aureus infection: a report of two pediatric cases

Resumen Staphylococcus aureus coloniza la nasofaringe en un tercio de los individuos sanos y además es causante de infecciones graves en pediatría, como endocarditis, neumonía e infecciones osteoarticulares. Posee varios mecanismos de virulencia, siendo la leucocidina de Panton Valentine (LPV) uno de ellos, una exotoxina que causa muerte celular. Su producción está comúnmente relacionada con Staphylococcus aureus resistente a meticilina (SARM) e infecciones pulmonares y musculo-esqueléticas graves. Sin embargo, la producción de LPV no es exclusiva de SARM. Se presentan dos casos clínicos de pacientes con infección por Staphylococcus aureus sensible a meticilina productora de esta exotoxina.

Abstract Staphylococcus aureus colonizes the nasopharynx in one third of healthy individuals and is also responsible for several infections in pediatrics such as endocarditis, pneumonia and osteoarticular infections. It has several virulence mechanisms, such as Panton Valentine leukocidin (PVL), which is an exotoxin that causes cell death. It is commonly related to methicillin-resistant Staphylococcus aureus (MRSA) and more serious pulmonary and musculoskeletal infections. However, PVL is not exclusive to MRSA. Two clinical cases of patients with infection by methicillin-sensitive Staphylococcus aureus producing this exotoxin are presented.

Panton-Valentine Leukocidin Induces Cytokine Release and Cytotoxicity Mediated by the C5a Receptor on Rabbit Alveolar Macrophages.

Necrotizing pneumonia caused by Panton-Valentine leukocidin (PVL)-positive community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) has high mortality rates and is currently a serious clinical issue. PVL is a two-component toxin (LukS-PV and LukF-PV). It can cause necrosis in target cells by forming pores consisting of an octamer comprised of LukS-PV and LukF-PV. However, considering the specificity of PVL towards several target cells and species, the specific effect of PVL remains controversial. Therefore, we focused on necrotizing pneumonia caused by PVL-positive S. aureus and clarified the effect of PVL on alveolar macrophages, which play a central role in innate immunity in the alveolar space. We constructed recombinant PVL (rPVL) components and stimulated alveolar macrophages isolated from rabbits to evaluate cytotoxicity and pro-inflammatory cytokine release. Recombinant LukS-PV (rLukS-PV), but not recombinant LukF-PV (rLukF-PV), induced pro-inflammatory cytokine release. Specifically, tumor necrosis factor (TNF)-α release was mediated by the C5a receptor (C5aR) expressed on rabbit alveolar macrophages, and the toxicity of rPVL, consisting of rLukS-PV and rLukF-PV, towards rabbit alveolar macrophages was mediated by the same receptor. Overall, our findings shed light on the C5aR-mediated cytotoxic effect of PVL on alveolar macrophages, which may be useful for understanding the mechanism of necrotizing pneumonia caused by PVL.

Single-Chain Fragment Variables Targeting Leukocidin ED Can Alleviate the Inflammation of Staphylococcus aureus-Induced Mastitis in Mice.

Staphylococcus aureus is a vital bovine mastitis pathogen causing huge economic losses to the dairy industry worldwide. In our previous studies, leukotoxin ED (LukED) was detected in most S. aureus strains isolated from bovine mastitis. Here, four single-chain fragment variables (scFvs) (ZL8 and ZL42 targeting LukE, ZL22 and ZL23 targeting LukD) were obtained using purified LukE and LukD proteins as the antigens after five rounds of bio-panning. The complementarity-determining region 3 (CDR3) of the VH domain of these scFvs exhibited significant diversities. In vitro, the scFvs significantly decreased LukED-induced cell killing by inhibiting the binding of LukED to chemokine receptors (CCR5 and CXCR2) and reduced the death rates of bovine neutrophils and MAC-T cells caused by LukED and S. aureus (p < 0.05). In an S. aureus-induced mouse mastitis model, histopathology and MPO results revealed that scFvs ameliorated the histopathological damages and reduced the infiltration of inflammatory cells (p < 0.05). The ELISA and qPCR assays showed that scFvs reduced the transcription and expression levels of Tumor Necrosis Factor-alpha (TNF-α), interleukin-1ß (IL-1ß), IL-6, IL-8 and IL-18 (p < 0.05). The overall results demonstrated the protective anti-inflammatory effect of scFvs in vitro and in vivo, enlightening the potential role of scFvs in the prevention and treatment of S. aureus-induced mastitis.

Staphylococcus aureus and its Effects on the Prognosis of Bronchiectasis.

Bronchiectasis, which is an abnormal and irreversible dilation of one or several bronchial segments, causes significant morbidity and impaired quality of life to patients, mainly as the result of recurrent and chronic respiratory infections. Staphylococcus aureus is a microorganism known for its high infectious potential related to the production of molecules with great pathogenic power, such as enzymes, toxins, adhesins, and biofilm, which determine the degree of severity of systemic symptoms and can induce exacerbated immune response. This review highlighted the clinical significance of S. aureus colonization/infection in bronchiectasis patients, since little is known about it, despite its increasing frequency of isolation and potential serious morbidity.

Frequency Of Panton Valentine Leucocidin Gene In Staphylococcus Aureus From Skin And Soft Tissue Infections.

BACKGROUND: Staphylococcus aureus harbouring Panton Valentine Leucocidin gene are emerging and spreading worldwide. PVL gene was first identified by Noel Panton and Francis Valentine in 1932 who explained its ability to lyse leucocytes and its main relationship with skin and soft tissue infections. In Pakistan only limited data is available on the frequency and molecular analysis of PVL gene positive Staph aureus. Therefore, this study was conducted to understand the clinical epidemiology of PVL positive Staph aureus in our setup. Objectives of the study was aimed to determine the frequency of PVL gene in Staph aureus obtained from pus samples from skin and soft tissue infections from various departments; indoor and outdoor of a tertiary care hospital of Lahore. METHODS: 384 Staph aureus isolates from skin and soft tissue infections were selected from both indoor and outdoor departments of hospital. After identification by phenotypic methods, they were processed by PCR using luk-F and luk-S primers for the detection of PVL gene. RESULTS: 186 out of 384 Staph aureus isolates were positive for PVL gene. Overall frequency of PVL gene was 49%. Frequency of PVL gene in Staph aureus was 44.9% in males and 53.5% in females. The highest frequency of PVL gene was detected in paediatric age group. A large majority of positive isolates were from pus samples other than swabs and from the general surgery department. They mostly belong to indoor with indoor outdoor ratio of approximately 2:1. Frequencies of PVL gene in MRSA and MSSA were 51% and 44% respectively. Frequency of PVL gene was found to be high in Ciprofloxacin sensitive, Gentamicin sensitive, Erythromycin resistant and Fusidic acid resistant isolates. CONCLUSION: Almost half of Staph aureus isolates were found PVL positive. They were mostly multidrug resistant came from indoor setup. This situation is very alarming so, there is a need to adopt strict infection control policies in the hospitals to limit the widespread and injudicious use of antibiotics. There is also a need to apply PVL positive Staph aureus treatment to the effected individuals which involve not only antibiotics but also the decolonization of effected individuals and their close contacts.

Staphylococcal Panton-Valentine Leucocidin and Gamma Haemolysin Target and Lyse Mature Bone Marrow Leucocytes.

Staphylococcus aureus is a major human pathogen, inducing several infections ranging from the benign to the life-threatening, such as necrotising pneumonia. S. aureus is capable of producing a great variety of virulence factors, such as bicomponent pore-forming leucocidin, which take part in the physiopathology of staphylococcal infection. In necrotising pneumonia, Panton-Valentine leucocidin (PVL) induces not only lung injury and necrosis, but also leukopenia, regarded as a major factor of a poor prognosis. The aim of the present study was to evaluate the effect of bicomponent pore-forming leucocidin, PVL and gamma haemolysin on bone marrow leucocytes, to better understand the origin of leukopenia. Using multi-parameter cytometry, the expression of leucocidin receptors (C5aR, CXCR1, CXCR2, and CCR2) was assessed and toxin-induced lysis was measured for each bone marrow leucocyte population. We observed that PVL resulted in myeloid-derived cells lysis according to their maturation and their C5aR expression; it also induced monocytes lysis according to host susceptibility. Haemolysin gamma A, B, and C (HlgABC) displayed cytotoxicity to monocytes and natural killer cells, hypothetically through CXCR2 and CXCR1 receptors, respectively. Taken together, the data suggest that PVL and HlgABC can lyse bone marrow leucocytes. Nevertheless, the origin of leukopenia in severe staphylococcal infection is predominantly peripheral, since immature cells stay insensitive to leucocidins.

Staphylococcus aureus Infection and Persistence in Chronic Rhinosinusitis: Focus on Leukocidin ED.

Chronic rhinosinusitis (CRS) is thought to be a multifactorial disease that includes a direct involvement of bacteria that trigger inflammation and contribute to CRS pathogenesis. Staphylococcus aureus infection and persistence is associated with chronic rhinosinusitis (CRS), and it may be particularly relevant in the form with nasal polyps (CRSwNP). The large array of exotoxins deployed by S. aureus is instrumental for the bacterium to warrant its infection and dissemination in different human body districts. Here, we analyze the common Th2 environment in CRSwNP and prospect a possible dynamic role played by S. aureus leukocidins in promoting this chronic inflammation, considering leukocidin ED (LukED) as a strong prototype candidate worth of therapeutic investigation. CCR5 is an essential target for LukED to exert its cytotoxicity towards T cells, macrophages and dendritic cells. Therefore, CCR5 blockade might be an interesting therapeutic option for CRS and, more specifically, persistent and relapsing CRSwNP. In this perspective, the arsenal of CCR5 antagonists being developed to inhibit HIV-1 entry (CCR5 being the major HIV-1 co-receptor) could be easily repurposed for CRS therapeutic investigation. Finally, direct targeting of LukED by neutralizing antibodies could represent an important additional solution to S. aureus infection.

Inflammasome activation by Pseudomonas aeruginosa's ExlA pore-forming toxin is detrimental for the host.

During acute Pseudomonas aeruginosa infection, the inflammatory response is essential for bacterial clearance. Neutrophil recruitment can be initiated following the assembly of an inflammasome within sentinel macrophages, leading to activation of caspase-1, which in turn triggers macrophage pyroptosis and IL-1ß/IL-18 maturation. Inflammasome formation can be induced by a number of bacterial determinants, including Type III secretion systems (T3SSs) or pore-forming toxins, or, alternatively, by lipopolysaccharide (LPS) via caspase-11 activation. Surprisingly, previous studies indicated that a T3SS-induced inflammasome increased pathogenicity in mouse models of P. aeruginosa infection. Here, we investigated the immune reaction of mice infected with a T3SS-negative P. aeruginosa strain (IHMA879472). Virulence of this strain relies on ExlA, a secreted pore-forming toxin. IHMA879472 promoted massive neutrophil infiltration in infected lungs, owing to efficient priming of toll-like receptors, and thus enhanced the expression of inflammatory proteins including pro-IL-1ß and TNF-α. However, mature-IL-1ß and IL-18 were undetectable in wild-type mice, suggesting that ExlA failed to effectively activate caspase-1. Nevertheless, caspase-1/11 deficiency improved survival following infection with IHMA879472, as previously described for T3SS+ bacteria. We conclude that the detrimental effect associated with the ExlA-induced inflammasome is probably not due to hyperinflammation, rather it stems from another inflammasome-dependent process.

Tracking the evolution of the two successful CC59 methicillin-resistant Staphylococcus aureus clones in Taiwan: the divergence time of the two clades is estimated to be the 1980s.

Clonal complex 59 (CC59) is the dominant community-associated methicillin-resistant Staphylococcus aureus (MRSA) strain in Taiwan and includes the Asian-Pacific clone with Panton-Valentine leukocidin (PVL)-negative/staphylococcal cassette chromosome mec (SCCmec) IVg and the Taiwan clone characterised as PVL-positive/SCCmec V (5C2&5). Nevertheless, data on the evolutionary history of the two dominant CC59 MRSA clones in Taiwan are scarce. In this study, a total of 258 CC59 S. aureus strains from Taiwan were classified by multiple-locus variable-number tandem repeat analysis (MLVA), which revealed two major clusters (MT1 and MT2) with distinct mobile genetic elements (MGEs). However, sequencing and PCR mapping of the ß-lactamase-producing plasmid revealed no difference among all CC59 S. aureus strains. Bayesian evolutionary analysis of 18 of the CC59 S. aureus strains based on core genome alignment revealed two clades: (i) Clade A, which shared the samples with MT1, had the features of mainly harbouring gentamicin-resistant MES6272-2 or MES4578, φSA3 translocation in νSaß and SCCmec IVg; and (ii) Clade B, which shared the samples with MT2, had the features of mainly harbouring streptomycin-resistant MESPM1, PVL phage and SCCmec V (5C2&5). Based on the time-calibrated phylogenetic tree, the estimated time of divergence of the two clades was in the 1980s. These results suggest that the CC59 S. aureus progenitor acquired a ß-lactamase-producing plasmid and then developed the varied genetic backgrounds, which were associated with the acquisition and maintenance of distinct MGEs, leading to differences in antimicrobial susceptibility profiles and molecular virulence determinants.

Targeting NLRP3 and Staphylococcal pore-forming toxin receptors in human-induced pluripotent stem cell-derived macrophages.

Staphylococcus aureus causes necrotizing pneumonia by secreting toxins such as leukocidins that target front-line immune cells. The mechanism by which leukocidins kill innate immune cells and trigger inflammation during S. aureus lung infection, however, remains unresolved. Here, we explored human-induced pluripotent stem cell-derived macrophages (hiPSC-dMs) to study the interaction of the leukocidins Panton-Valentine leukocidin (PVL) and LukAB with lung macrophages, which are the initial leukocidin targets during S. aureus lung invasion. hiPSC-dMs were susceptible to the leukocidins PVL and LukAB and both leukocidins triggered NLPR3 inflammasome activation resulting in IL-1ß secretion. hiPSC-dM cell death after LukAB exposure, however, was only temporarily dependent of NLRP3, although NLRP3 triggered marked cell death after PVL treatment. CRISPR/Cas9-mediated deletion of the PVL receptor, C5aR1, protected hiPSC-dMs from PVL cytotoxicity, despite the expression of other leukocidin receptors, such as CD45. PVL-deficient S. aureus had reduced ability to induce lung IL-1ß levels in human C5aR1 knock-in mice. Unexpectedly, inhibiting NLRP3 activity resulted in increased wild-type S. aureus lung burdens. Our findings suggest that NLRP3 induces macrophage death and IL-1ß secretion after PVL exposure and controls S. aureus lung burdens.