is a significant health problem worldwide due to multi-drug resistance issues and absence of an effective vaccine. Patients infected with
have not shown a better immune response in successive ...infections. This might be explained by the fact that
possesses several mechanisms to evade the innate and adaptative immune responses at different levels. Macrophages are a key cellular component in the innate immune response against microorganisms. The current information suggests that gonococcus can hijack the host response by mechanisms that involve the control of macrophages activity. In this mini review, we intend to condense the recent knowledge on the macrophage-
interactions with a focus on strategies developed by gonococcus to evade or to exploit immune response to establish a successful infection. Finally, we discuss the opportunities and challenges of therapeutics for controlling immune manipulation by
.
Macrophages are critical cells in the innate immune response to microorganisms sensed in the tissues. During infections, the interaction between pathogens and macrophages leads to a macrophage ...response that includes cytokine production, antigen processing and presentation in the context of MHC molecules, expression of T cell costimulatory molecules and recruitment of innate defense effectors, which results in clearance of infection. However, Neisseria gonorrhoeae can suppress the protective immune response at this level, avoiding its detection and elimination. Studies addressed to develop the interactions between macrophages and Neisseria gonorrhoeae allow us to find potential targets to be exploited with vaccines and therapeutic drugs. In this chapter, we describe protocols to generate human monocyte-derived macrophages and assess their response to infection with Neisseria gonorrhoeae.
Current data suggest that Neisseria gonorrhoeae is able to suppress the protective immune response at different levels, such as B and T lymphocytes and antigen-presenting cells. The present report is ...focused on gonococcus evasion mechanism on macrophages (MФ) and its impact in the subsequent immune response. In response to various signals MФ may undergo classical-M1 (M1-MФ) or alternative-M2 (M2-MФ) activation. Until now there are no reports of the gonococcus effects on human MФ polarization. We assessed the phagocytic ability of monocyte-derived MФ (MDM) upon gonococcal infection by immunofluorescence and gentamicin protection experiments. Then, we evaluated cytokine profile and M1/M2 specific-surface markers on MФ challenged with N. gonorrhoeae and their proliferative effect on T cells. Our findings lead us to suggest N. gonorrhoeae stimulates a M2-MФ phenotype in which some of the M2b and none of the M1-MФ-associated markers are induced. Interestingly, N. gonorrhoeae exposure leads to upregulation of a Programmed Death Ligand 1 (PD-L1), widely known as an immunosuppressive molecule. Moreover, functional results showed that N. gonorrhoeae-treated MФ are unable to induce proliferation of human T-cells, suggesting a more likely regulatory phenotype. Taken together, our data show that N. gonorroheae interferes with MФ polarization. This study has important implications for understanding the mechanisms of clearance versus long-term persistence of N. gonorroheae infection and might be applicable for the development of new therapeutic strategies.
The convergence of hypervirulence and carbapenem resistance in the bacterial pathogen Klebsiella pneumoniae represents a critical global health concern. Hypervirulent K. pneumoniae (hvKp) strains, ...frequently from sequence type 23 (ST23) and having a K1 capsule, have been associated with severe community-acquired invasive infections. Although hvKp were initially restricted to Southeast Asia and primarily antibiotic-sensitive, carbapenem-resistant hvKp infections are reported worldwide. Here, within the carbapenemase production Enterobacterales surveillance system headed by the Chilean Public Health Institute, we describe the isolation in Chile of a high-risk ST23 dual-carbapenemase-producing hvKp strain, which carbapenemase genes are encoded in a single conjugative plasmid.
Phenotypic and molecular tests of this strain revealed an extensive resistance to at least 15 antibiotic classes and the production of KPC-2 and VIM-1 carbapenemases. Unexpectedly, this isolate lacked hypermucoviscosity, challenging this commonly used hvKp identification criteria. Complete genome sequencing and analysis confirmed the K1 capsular type, the KpVP-1 virulence plasmid, and the GIE492 and ICEKp10 genomic islands carrying virulence factors strongly associated with hvKp. Although this isolate belonged to the globally disseminated hvKp clonal group CG23-I, it is unique, as it formed a clade apart from a previously reported Chilean ST23 hvKp isolate and acquired an IncN KPC-2 plasmid highly disseminated in South America (absent in other hvKp genomes), but now including a class-I integron carrying bla
and other resistance genes. Notably, this isolate was able to conjugate the double carbapenemase plasmid to an E. coli recipient, conferring resistance to 1st -5th generation cephalosporins (including combinations with beta-lactamase inhibitors), penicillins, monobactams, and carbapenems.
We reported the isolation in Chile of high-risk carbapenem-resistant hvKp carrying a highly transmissible conjugative plasmid encoding KPC-2 and VIM-1 carbapenemases, conferring resistance to most beta-lactams. Furthermore, the lack of hypermucoviscosity argues against this trait as a reliable hvKp marker. These findings highlight the rapid evolution towards multi-drug resistance of hvKp in Chile and globally, as well as the importance of conjugative plasmids and other mobile genetic elements in this convergence. In this regard, genomic approaches provide valuable support to monitor and obtain essential information on these priority pathogens and mobile elements.
Drug delivery systems based on polymeric microparticles represent an interesting field of development for the treatment of several infectious diseases for humans and animals. In this work, we ...developed PLGA microparticles loaded with ceftiofur (PLGA-cef), a third- generation cephalosporin that is used exclusively used in animals. PLGA-cef was prepared by the double emulsion w/o/w method, and exhibited a diameter in the range of 1.5-2.2 μm, and a negative ζ potential in the range of -35 to -55 mV. The loading yield of PLGA-cef was ~7% and encapsulation efficiency was approximately 40%. The pharmacokinetic study demonstrated a sustained release profile of ceftiofur for 20 days. PLGA-cef administrated in a single dose was more effective than ceftiofur non-encapsulated in rats challenged with S. Typhimurium. The in vivo toxicological evaluation showed that PLGA-cef did not affect the blood biochemical, hematological and hemostasis parameters. Overall, the PLGA-cef showed slow in vivo release profile, high antibacterial efficacy, and low toxicity. The results obtained supports the safe application of PLGA-cef as sustained release platform in the veterinary industry.
Melanin is a pigment found in all biological kingdoms, and plays a key role in protection against ultraviolet radiation, oxidizing agents, and ionizing radiation damage. Melanin exerts an ...antimicrobial activity against bacteria, fungi, and parasites. We demonstrated an antifungal activity of synthetic and human melanin against
sp. The members of the
species complexes are capsulated yeasts, which cause cryptococcosis. For both species melanin is an important virulence factor. To evaluate if cryptococcal and human melanins have antifungal activity against
species they both were assayed for their antifungal properties and physico-chemical characters. Melanin extracts from human hair and different strains of
(
= 4) and
(
= 4) were investigated. The following minimum inhibitory concentrations were found for different melanins against
and
were (average/range): 13.7/(7.8-15.6) and 19.5/(15.6-31.2) μg/mL, respectively, for human melanin; 273.4/(125->500) and 367.2/(125.5->500) μg/mL for
melanin and 125/(62.5-250) and 156.2/(62-250) μg/mL for
melanin. Using Scanning Electron Microscopy we observed that human melanin showed a compact conformation and cryptococcal melanins exposed an amorphous conformation. Infrared spectroscopy (FTIR) showed some differences in the signals related to C-C bonds of the aromatic ring of the melanin monomers. High Performance Liquid Chromatography established differences in the chromatograms of fungal melanins extracts in comparison with human and synthetic melanin, particularly in the retention time of the main compound of fungal melanin extracts and also in the presence of minor unknown compounds. On the other hand, MALDI-TOF-MS analysis showed slight differences in the spectra, specifically the presence of a minor intensity ion in synthetic and human melanin, as well as in some fungal melanin extracts. We conclude that human melanin is more active than the two fungal melanins against Cryptococcus. Although some physico-chemical differences were found, they do not explain the differences in the antifungal activity against
of human and cryptococcal melanins. More detailed studies on the structure should be considered to associate structure and antifungal activity.
is the causative agent of the most frequently reported nosocomial diarrhea worldwide. The high incidence of recurrent infection is the main clinical challenge of
infections (CDI). Formation of
spores ...of the epidemic strain R20291 has been shown to be essential for recurrent infection and transmission of the disease in a mouse model. However, the underlying mechanisms of how these spores persist in the colonic environment remains unclear. In this work, we characterized the adherence properties of epidemic R20291 spores to components of the intestinal mucosa, and we assessed the role of the exosporium integrity in the adherence properties by using
mutant spores with a defective exosporium layer. Our results showed that spores and vegetative cells of the epidemic R20291 strain adhered at high levels to monolayers of Caco-2 cells and mucin. Transmission electron micrographs of Caco-2 cells demonstrated that the hair-like projections on the surface of R20291 spores are in close proximity with the plasma membrane and microvilli of undifferentiated and differentiated monolayers of Caco-2 cells. Competitive-binding assay in differentiated Caco-2 cells suggests that spore-adherence is mediated by specific binding sites. By using spores of a
mutant we demonstrated that the integrity of the exosporium layer determines the affinity of adherence of
spores to Caco-2 cells and mucin. Binding of fibronectin and vitronectin to the spore surface was concentration-dependent, and depending on the concentration, spore-adherence to Caco-2 cells was enhanced. In the presence of an aberrantly-assembled exosporium (
spores), binding of fibronectin, but not vitronectin, was increased. Notably, independent of the exosporium integrity, only a fraction of the spores had fibronectin and vitronectin molecules binding to their surface. Collectively, these results demonstrate that the integrity of the exosporium layer of strain R20291 contributes to selective spore adherence to components of the intestinal mucosa.
Base-pairing small RNAs (sRNAs) regulate gene expression commonly by direct interaction with cognate mRNAs. Nevertheless, recent studies have expanded this knowledge with the discovery of the RNA ...'sponges' which are able to interact and repress the functions of classical base-pairing sRNAs. In this work, we present evidence indicating that the sponge RNA SroC from Salmonella enterica serovar Typhimurium base pairs with the MgrR sRNA, thereby antagonizing its regulatory effects on both gene expression and resistance to the antimicrobial peptide polymyxin B (PMB). By a predictive algorithm, we determined putative SroC-MgrR base-pairing regions flanking the interaction area between MgrR and its target mRNA, eptB, encoding a LPS-modifying enzyme. With a two-plasmid system and compensatory mutations, we confirmed that SroC directly interacts and down-regulates the levels of MgrR, thus relieving the MgrR-mediated repression of eptB mRNA. Since it was previously shown that an Escherichia coli strain carrying an mgrR deletion is more resistant to PMB, we assessed the significance of SroC in the susceptibility of S. Typhimurium to PMB. Whereas the sroC deletion increased the sensitivity to PMB, as compared to the wild-type, the resistance phenotypes between the ΔmgrR and ΔsroCΔmgrR strains were comparable, evidencing that mgrR mutation is epistatic to the sroC mutation. Together, these results indicate that both SroC and MgrR sRNAs compose a coherent feed-forward loop controlling the eptB expression and hence the LPS modification in S. Typhimurium.
Objectives
We investigated the colonisation by Candida spp in patients using orthodontic fixed appliances by characterising the isolated Candida strains and by evaluating the host oral mucosa ...response through the measure of human β‐defensins 3 (HBD‐3) expression and Interleukin‐1ß/IL‐10.
Methods
Ninety patients were enrolled after signing an informed consent. Prevalence, susceptibility to fluconazole, genotyping and oral fungal burden of Candida sp. isolated were determined. Host responses were evaluated by measuring HBD‐3 expression as well as IL‐1ß and IL‐10 in saliva.
Results
The colonisation rate reached 6.7% (6/90), and 5 patients were colonised with C. albicans strains and one with one with C. tropicalis. The fluconazole MIC90/susceptibility of C. albicans strains ranged 1/0.25‐1 μg/mL. However, isolated strains did not present different genotype (SAB>0.9), C. albicans colonisation seems to be influenced by the duration of treatment and by level expression of HBD3 that were higher in colonised patients (not statistically different). A negative correlation between the fungal burden and IL‐1ß levels was found in colonised patients but not for IL‐10.
Conclusions
Our study revealed that patients with orthodontic fixed appliances were mainly colonised by C. albicans, which was related to a decrease in HBD‐3 expression and IL‐1ß levels.
(Ngo) is the etiological agent of gonorrhea, a sexually transmitted infection that initially infects the female lower genital tract. In untreated women, the bacteria can ascend to the upper genital ...reproductive tract and infect the fallopian tube (FTs), which is associated with salpingitis and can lead to impaired FT function and infertility. The extracellular matrix (ECM) plays an important role in cell migration and differentiation in the female genital tract, and some pathogens modify the ECM to establish successful infections. The ECM is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), their endogenous inhibitors; MMP deregulation causes pathological conditions in a variety of tissues.
The aim of this work was to analyze the expression and localization of MMP-3, MMP-8, MMP-9, and TIMP-1 in FT explants during Ngo infection using real-time PCR, immunohistochemistry, zymography and ELISA. No significant variations in MMP-3, MMP-9, and TIMP-1 transcript levels were observed. In contrast, a significant increase (
< 0.05) was observed for MMP-8 expression and was accompanied by stromal immunoreactivity in infected explants. ELISA results supported these findings and showed that MMP-8 release increased upon gonococcal infection.
Our results indicate that gonococcal infection induces increased MMP-8 expression, which might contribute to FT damage during infection.
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