Antimicrobial approach of abdominal post-surgical infections.

Abdominal surgical site infections (SSIs) are infections that occur after abdominal surgery. They can be superficial, involving the skin tissue only, or more profound, involving deeper skin tissues including organs and implanted materials. Currently, SSIs are large global health problem with an incidence that varies significantly depending on the United Nations' Human Development Index. The purpose of this review is to provide a practical update on the latest available literature on SSIs, focusing on causative pathogens and treatment with an overview of the ongoing studies of new therapeutic strategies.

MALDI-TOF Mass Spectrometry Analysis and Human Post-Mortem Microbial Community: A Pilot Study.

Introduction: The human post-mortem microbiome (HPM) plays a major role in the decomposition process. Successional changes in post-mortem bacterial communities have been recently demonstrated using high throughput metagenomic sequencing techniques, showing great potential as a post-mortem interval (PMI) predictor. The aim of this study is to verify the application of the mass spectrometry technique, better known as MALDI-TOF MS (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry), as a cheap and quick method for microbe taxonomic identification and for studying the PM microbiome. Methods: The study was carried out on 18 human bodies, ranging from 4 months to 82 years old and with a PMI range from 24 h up to 15 days. The storage time interval in the coolers was included in the final PMI estimates. Using the PMI, the sample study was divided into three main groups: seven cases with a PMI < 72 h; six cases with a PMI of 72−168 h and five cases with a PMI > 168 h. For each body, microbiological swabs were sampled from five external anatomical sites (eyes, ears, nose, mouth, and rectum) and four internal organs (brain, spleen, liver, and heart). Results: The HPM became increasingly different from the starting communities over time in the internal organs as well as at skin sites; the HPM microbiome was mostly dominated by Firmicutes and Proteobacteria phyla; and a PM microbial turnover existed during decomposition, evolving with the PMI. Conclusions: MALDI-TOF is a promising method for PMI estimation, given its sample handling, good reproducibility, and high speed and throughput. Although several intrinsic and extrinsic factors can affect the structure of the HPM, MALDI-TOF can detect the overall microbial community turnover of most prevalent phyla during decomposition. Limitations are mainly related to its sensitivity due to the culture-dependent method and bias in the identification of new isolates.

Postmortem interval assessment by MALDI-TOF mass spectrometry analysis in murine cadavers.

AIMS: This study assessed the use of matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry as an alternative method to identify species associated with the thanatomicrobiota and epinecrotic communities. METHODS AND RESULTS: The study was conducted on 10 murine cadavers, and microbiological swabs were collected from five external anatomical sites (eyes, ears, nose, mouth and rectum) and four internal organs (brain, spleen, liver, heart), during 16 and 30 days, for the thanatomicrobiota and epinecrotic communities, respectively. Our results revealed that the postmortem microbiota associated with the external cavities showed changes over time and reduced taxonomic diversity. The internal organs, initially sterile, showed signs of microbial invasion at 3 and 10 days postmortem for the liver-spleen and heart-brain, respectively. The postmortem microbiota was mainly dominated by Firmicutes and Proteobacteria. CONCLUSIONS: MALDI-TOF is a promising method for estimating postmortem interval (PMI), associated with rapid sample handling, good reproducibility and high productivity. SIGNIFICANCE AND IMPACT OF THE STUDY: This study investigated microbial changes during the decomposition process and proposed a simple strategy for PMI estimation. Results introducing the application of the MALDI-TOF method in the field of forensic.

Outer Membrane Vesicles Derived from Klebsiella pneumoniae Influence the miRNA Expression Profile in Human Bronchial Epithelial BEAS-2B Cells.

Klebsiella pneumoniae is an opportunistic pathogen that causes nosocomial and community-acquired infections. The spread of resistant strains of K. pneumoniae represents a growing threat to human health, due to the exhaustion of effective treatments. K. pneumoniae releases outer membrane vesicles (OMVs). OMVs are a vehicle for the transport of virulence factors to host cells, causing cell injury. Previous studies have shown changes of gene expression in human bronchial epithelial cells after treatment with K. pneumoniae OMVs. These variations in gene expression could be regulated through microRNAs (miRNAs), which participate in several biological mechanisms. Thereafter, miRNA expression profiles in human bronchial epithelial cells were evaluated during infection with standard and clinical K. pneumoniae strains. Microarray analysis and RT-qPCR identified the dysregulation of miR-223, hsa-miR-21, hsa-miR-25 and hsa-let-7g miRNA sequences. Target gene prediction revealed the essential role of these miRNAs in the regulation of host immune responses involving NF-ĸB (miR-223), TLR4 (hsa-miR-21), cytokine (hsa-miR-25) and IL-6 (hsa-let-7g miRNA) signalling pathways. The current study provides the first large scale expression profile of miRNAs from lung cells and predicted gene targets, following exposure to K. pneumoniae OMVs. Our results suggest the importance of OMVs in the inflammatory response.

Campylobacter jejuni bacteremia in Italian pediatric patients with acute lymphoblastic leukemia: Report of two cases.

Infections caused by Campylobacter jejuni are rarely associated with extraintestinal complications. C. jejuni bacteremia is difficult to detect in patients with hematological malignancies undergoing chemotherapy where the choice of appropriate antibiotic treatment is extremely important. We report two cases of C. jejuni bacteremia in Italian pediatric patients affected by acute lymphoblastic leukemia (ALL). Agreeing with the most recent epidemiological data, both clinical isolates showed a typical phenotypic antimicrobial resistance patterns with combined resistance to ciprofloxacin and tetracycline. To our knowledge, this is the first report of C. jejuni isolation from the blood of ALL pediatric patients in Italy, and it provides important epidemiological information on this rare infection.

Spontaneous bacterial peritonitis due to carbapenemase-producing Enterobacteriaceae: Etiology and antibiotic treatment.

Carbapenem antibiotics were first introduced in the 1980s and have long been considered the most active agents for the treatment of multidrug-resistant gram-negative bacteria. Over the last decade, carbapenem-resistant Enterobacteriaceae (CRE) have emerged as organisms causing spontaneous bacterial peritonitis. Infections caused by CRE have shown a higher mortality rate than those caused by bacteria sensitive to carbapenem antibiotics. Current antibiotic guidelines for the treatment of spontaneous bacterial peritonitis are insufficient, and rapid de-escalation of empiric antibiotic treatment is not widely recognized. This review summarizes the molecular characteristics, epidemiology and possible treatment of spontaneous bacterial peritonitis caused by CRE.

Seven years prevalence and distribution of high and low risk HPV genotypes in women living in the metropolitan area of Naples.

BACKGROUND: The objective of this work was to provide an update information on HPV age/genotype distribution by retrospectively analyzing a cohort of women living in the metropolitan area of Naples. METHODS: From January 2011 to December 2017, cervical scrape specimens from 1265 women, with abnormal cytological indication, were tested for HPV DNA. The presence and the viral genotypes were assessed by the Linear Array HPV genotyping test for the detection of 37 anogenital HPV-DNA genotypes. RESULTS: The overall prevalence of HPV infections was of 44.5% (95% CI 41.77-47.24). Among HR-HPV types, HPV-16 was the most common identified genotype, followed by HPV-31, -66, -59 and -51. As concern LR-HPV, HPV-53 resulted the most prevalent. Stratifying the study population by age, the total HPV infections showed a peak in younger women aged <23 years (58.5%), with a significative decrease by age (23-29 years, 54%; ≥ 30 years, 38.2%) (p < 0.001). CONCLUSION: We provided an HPV epidemiological analysis, highlighting the need to implement vaccination programmes and preventative screening strategies.

Isolation, characterization and analysis of pro-inflammatory potential of Klebsiella pneumoniae outer membrane vesicles.

Outer membrane vesicles (OMVs) are potent virulence factors, naturally secreted by gram-negative bacteria. Since Klebsiella pneumoniae has emerged as an important nosocomial pathogen, because of resistance to a wide spectrum of antibiotics, it is crucial to investigate its pathogenetic mechanism microorganism secretes outer membrane vesicles (OMVs), but the pathogenesis of Klebsiella pneumoniae as it relates to OMVs has not been well elucidated. In this study we focused on the isolation, characterization and evaluation of the virulence potential of OMVs obtained from Klebsiella pneumoniae. Our data demonstrate that Klebsiella pneumoniae OMVs are important secretory nanocomplexes that elicit a potent inflammatory response. Since OMVs are clearly involved in the pathogenesis of this bacterium during infection, further studies are required to determine whether they could be future targets for novel therapy and potential vaccine against Klebsiella pneumoniae.

Enrichment of semen culture in the diagnosis of bacterial prostatitis.

The objective was to investigate the diagnostic accuracy of our microbiological protocol to simplify the evaluation of bacterial prostatitis in the clinical practice. Our findings show the possibility to apply our alternative enrichment semen culture method to detect prostatic bacterial infection with higher sensitivity than the gold standard M&S technique.

A new enrichment diagnostic platform for semen culture.

Urogenital bacterial infections have been described in literature as a potential cause of infertility. For the consequences that a failure in diagnosis could have on the evolution of male urogenital infectious disease, an accurate microbiological procedure to investigate the bacterial species composition of seminal fluid plays a crucial role to better understand the eventual correlation with infertility. In order to improve the quality of semen culture investigations, we have developed a new enrichment diagnostic platform. Semen samples of 540 infertile men were simultaneously analyzed using the standard microbiological semen culture method and an alternative new experimental technique (Brain Heart Infusion broth, BHI, enrichment). Our results established the possibility to apply BHI enrichment to detect bacteria from semen samples with higher sensitivity (100%) and negative predictive value (100%) than the standard technique.

Distribution of Hepatitis C Virus Genotypes and Subtypes in the Metropolitan Area of Naples, Italy, in the Era of Interferon-Free Regimens.

AIMS: The objective of this work was to study the distribution of hepatitis C virus (HCV) genotypes and subtypes from 2010 to 2015 in 1,221 anti-HCV/HCV-RNA-positive specimens from patients living in the metropolitan area of Naples, since HCV genotypes and subtypes remain cornerstones in the management of chronic HCV infection even in the directly acting antivirals era. METHODS: The study was carried out on 1,221 anti-HCV/HCV-RNA-positive plasma samples collected between April 2010 and December 2015. RESULTS: Of the 1,221 patients enrolled, 633 (51.9%) were males and 588 (48.1%) were females, with a mean age of 60 ± 13 (SD) years. The most frequent HCV genotype observed was genotype 1 (68.1%; 1b in 55.3% and 1a in 9.5%); HCV genotype 2 was found in 289 samples (23.67%), genotype 3 in 6.47%, genotype 4 in only 19 samples, and only 2 samples were classified as genotype 5. The mean age of the patients with genotype 1a or 3 was lower (51 ± 12 and 49 ± 12 years, respectively) than those with genotype 1b (62 ± 11, p < 0.0001 for both) or 2 (62 ± 14, p < 0.0001 for both). CONCLUSIONS: The data from the present study suggest that HCV genotype 1b remains the most prevalent in this area, followed by genotype 2, 1a, and 3a.

Glycine-replaced derivatives of [Pro3,DLeu9]TL, a temporin L analogue: Evaluation of antimicrobial, cytotoxic and hemolytic activities.

In this study we designed and synthesized a new library of antimicrobial peptides correlated to [Pro3,DLeu9]TL 1, a temporin L derivative devoid of cytolytic effects in vitro, and investigated the correlation between the α-helical content of the compounds and their antibacterial, cytotoxic and hemolytic activities. We systematically replaced Gly in position 10 of reference peptide with several amino acids. Structure-activity relationship studies of these analogues were performed by means of antimicrobial and cytotoxicity assays along with CD spectroscopy analyses. NMR analysis was also accomplished for compound 10. As well, the most promising peptides were additionally evaluated for their activity against some clinical strains isolated from human skin and for their mechanism of action by studying the kinetics of membrane perturbation of some representative microbial strains. We identified novel analogues with interesting properties that make them attractive lead compounds for potential topical applications.

Infectivity inhibition by overlapping synthetic peptides derived from the gH/gL heterodimer of herpes simplex virus type 1.

Herpes simplex virus (HSV) is a human pathogen that infects epithelial cells. The cutaneous lesions, caused by the virus, spread to the nervous system creating several complications. Fusion of host membranes with the viral envelope is mandatory and mediated by a group of glycoproteins conserved in all Herpesviridae subfamilies, such as the glycoproteins B (gB), H (gH), L (gL) and D (gD). We investigated the inhibitory activity mediated by synthetic overlapping peptides spanning the entire ectodomains of gH and gL glycoproteins. We have performed a brute analysis of the complete gH/gL heterodimer in order to explore the inhibitory activity of peptides modelled on these glycoproteins against HSV-1 infection. Twenty-four of the gH peptides at a concentration of 150 µM reached the 50% of inhibition cut-off. Interestingly, they are mainly located in the gH carboxy-terminal domain. None of the gL peptides had a clear inhibiting effect. No peptide toxicity was observed by lactate dehydrogenase assay at the concentrations used in our experimental conditions. HSV-1 therapy is based on acyclovir treatment, but some resistant strains are emerging. In this scenario, innovative approaches for HSV-1 treatment are necessary. Our data support the direct involvement of the described domains in the process of virus penetration; therefore, these results are of relevance to the potential development of novel therapeutic compounds to prevent HSV-1 infections. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.

Non-dermatophytic onychomycosis diagnostic criteria: an unresolved question.

Non-dermatophytic moulds (NDMs) have been increasingly recognised as causative agents of onychomycosis. The diagnosis of onychomycosis is most often obtained by microscopic observation of nail specimens where fungal elements can be detected and cultured by standard mycological techniques. Direct microscopic examination does not always result positive in NDM onychomycosis; therefore to perform a correct diagnosis, a proper mycological culture is often required. The purpose of our study was to evaluate the role of direct microscopic examination in the NDM onychomycosis diagnosis. The results show that only 57.2% of the specimens from onychomycosis patients could be properly diagnosed showing positivity to both direct microscopic examination and NDMs culture isolation in two or more subsequent inoculations, while 42.8% of analysed specimens with a negative direct microscopic examination, showed NDMs growth after three or more subsequent inoculations. The large proportion of false negatives (more than 42%) could be related to the duration of the infection and/or to the experience and skills of the personnel dedicated to specimen collection. We point out the need for thoroughly evaluating all specimens showing cultural growth in at least three subsequent medium inoculations, whatever the result of the microscopic examination, in order to reduce false-negative rates. This strategy would allow for more accurate diagnosis of this mycosis.

Self-assembled or mixed peptide amphiphile micelles from Herpes simplex virus glycoproteins as potential immunomodulatory treatment.

The use of micelle aggregates formed from peptide amphiphiles (PAs) as potential synthetic self-adjuvant vaccines to treat Herpes simplex virus (HSV) infection are reported here. The PAs were based on epitopes gB409-505 and gD301-309, selected from HSV envelope glycoprotein B (gB) and glycoprotein D (gD), that had their N-terminus modified with hydrophobic moieties containing two C18 hydrocarbon chains. Pure and mixed micelles of gB and/or gD peptide epitopes were easily prepared after starting with the synthesis of corresponding PAs by solid phase methods. Structural characterization of the aggregates confirmed that they were sufficiently stable and compatible with in vivo use: critical micelle concentration values around 4.0 ⋅ 10(-7) mol ⋅ Kg(-1); hydrodynamic radii (RH) between 50-80 nm, and a zeta potential (ζ) around - 40 mV were found for all aggregates. The in vitro results indicate that both peptide epitopes and micelles, at 10 µM, triggered U937 and RAW 264.7 cells to release appreciable levels of cytokines. In particular, interleukin (IL)-23-, IL-6-, IL-8- or macrophage inflammatory protein (MIP)-2-, and tumor necrosis factor (TNF)-α-release increased considerably when cells were treated with the gB-micelles or gD-micelles compared with the production of the same cytokines when the stimulus was the single gB or gD peptide.

Biological activity of lactoferrin-functionalized biomimetic hydroxyapatite nanocrystals.

The emergence of bacterial strains resistant to antibiotics is a general public health problem. Progress in developing new molecules with antimicrobial properties has been made. In this study, we evaluated the biological activity of a hybrid nanocomposite composed of synthetic biomimetic hydroxyapatite surface-functionalized by lactoferrin (LF-HA). We evaluated the antimicrobial, anti-inflammatory, and antioxidant properties of LF-HA and found that the composite was active against both Gram-positive and Gram-negative bacteria, and that it modulated proinflammatory and anti-inflammatory responses and enhanced antioxidant properties as compared with LF alone. These results indicate the possibility of using LF-HA as an antimicrobial system and biomimetic hydroxyapatite as a candidate for innovative biomedical applications.