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The murine immune response to Escherichia coli O6:K-alone or pre-exposed to 0.1 x MIC of aztreonam was investigated. Relative to mice immunized with untreated bacteria, mice immunized with antibiotic-treated microorganisms presented a significantly enhanced protection towards a challenge of 100 x LD50 of viable E. coli O6:K-. Previous injection of 0.1 mL of serum drawn from mice immunized with treated and untreated bacteria protected non-immunized mice towards a challenge of 10 x LD50 of viable E. coli O6:K--. Serum from mice immunized with treated bacteria also protected non-immunized mice towards a lethal challenge of E. coli O111. The antiserum contained high titre of IgG antibodies that cross-reacted with lipopolysaccharide isolated from smooth and rough Gram-negative bacteria. Immunoblotting showed additional bands of reactivity to the untreated E. coli O6:K-. Immunization with antibiotic-treated bacteria led to the production of type specific and cross reactive antibodies that protected animals against viable homologous and heterologous lethal challenges. [ABSTRACT FROM AUTHOR]

OSPAR, Rake and Flood accumulation zone methods were applied at 29 beaches of the German and Lithuanian Baltic Sea coast (2011-2018) to monitor cigarette butt pollution. Also, butt pollution prevention measure - ballot bin, was tested. The number of cigarette butts was significantly higher in Germany than in Lithuania, but the percent of butts from total litter items was similar. Rake method turned out to be suitable for cigarette butt assessment, while the OSPAR method underestimates the butt pollution. The visitor number on the beach had a significant effect on the cigarette butt number: in remote beaches, without visitors, usually, no butts were observed, while the highest number and the percent of butts were observed in beaches with the highest number of visitors. The ballot bin campaign may have increased the public awareness about pollution, but turned out to be in-efficient in reducing it., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

HHV-6A and HHV-6B are ubiquitous human betaherpesviruses sharing more than 80% homology. HHV-6B is the most common cause of encephalitis in transplant patients and its primary infection may cause the exanthema subitum and febrile seizures in infants. HHV-6A and HHV-6B are able to infect several immune cell types such as T cells, monocytes and dendritic cells (DCs). In this study we found that HHV-6 B derived from patients affected by exanthema subitum impaired monocyte differentiation into DCs, as the infected cells acquired less CD1a DC marker and retained more CD14 monocyte marker. In agreement with the previous finding that HHV-6B dysregulated autophagy and induced endoplasmic reticulum (ER) stress in cells in which it replicated, here we found that these effects occurred also in differentiating monocytes and that ER stress relief, by using the chemical chaperone sodium 4-phenylbutirate (PBA), partially restored DC formation. This suggests that the induction of ER stress, likely exacerbated by autophagy inhibition, could contribute to the immune suppression induced by HHV-6B derived from exanthema subitem patients., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The Tyr705 STAT3 constitutive activation, besides promoting PEL cell survival, contributes to the maintenance of viral latency. We found indeed that its de-phosphorylation by AG490 induced KSHV lytic cycle. Moreover, Tyr705 STAT3 de-phosphorylation, mediated by the activation of tyrosine phosphatases, together with the increase of Ser727 STAT3 phosphorylation contributed to KSHV lytic cycle induction by TPA. We then observed that p53-p21 axis, essential for the induction of KSHV replication, was activated by the inhibition of Tyr705 and by the increase of Ser727 STAT3 phosphorylation. As a possible link between STAT3, p53-p21 and KSHV lytic cycle, we found that TPA and AG490 reduced the expression of KAP-1, promoting p53 stability, p21 transcription and KSHV lytic cycle activation in PEL cells., (Copyright © 2018. Published by Elsevier Inc.)

Herpesviruses are known to manipulate autophagy to optimize their replication, counteract immune response and probably to promote tumourigenesis. This study explored, for the first time, the impact of human herpesvirus (HHV)-6 lytic infection on autophagy and demonstrated that HHV-6A and B (viruses sharing more than 80 % homology) differently affected this cellular process. Indeed, while HHV-6A (GS) infection of HSB2 cells promoted autophagy, HHV-6B (Z29) or the virus isolated from the serum of roseola infantum-affected patient-inhibited autophagy in Molt-3 cells or in PBMCs, respectively. Interestingly, the different behaviour of HHV-6A and B on the autophagic process was accompanied by different effects on endoplasmic reticulum stress, unfolded protein response and cell survival that was more strongly reduced by HHV-6B infection. We hypothesize that the ability to inhibit autophagy displayed by HHV-6B could be due to the fact that it contains gene homologues of those encoding for TRS1; the protein responsible for the block of autophagy by human cytomegalovirus. Understanding how HHV-6A/B infection regulates autophagy could be of particular interest, as it has been recently shown that this virus may be involved in Alzheimer's disease in which a dysregulation of autophagy may also play a role.

In cancer patients, the immune system is often altered with an excess of inhibitory factors, such as immunosuppressive cytokines, produced by regulatory T cells (Treg) or myeloid-derived suppressor cells (MDSC). The manipulation of the immune system has emerged as one of new promising therapies for cancer treatment, and also represents an attractive strategy to control prostate cancer (PCa). Therapeutic cancer vaccines and immune checkpoint inhibitors have been the most investigated in clinical trials. Many trials are ongoing to define the effects of immune therapy with established treatments: androgen deprivation therapy (ADT) and chemotherapy (CT) or radiotherapy (RT). This article discusses some of these approaches in the context of future treatments for PCa.

Human herpesviruses (HHVs) cause many serious acute and persistent central nervous system (CNS) disorders. Because these infections manifest with various, often non-specific, symptoms and signs, and because specific therapy is often available, accurate diagnosis is essential. Cerebrospinal fluid (CSF) from 146 patients with acute meningitis or meningoencephalitis and 9 with "other neurological disorders" were analyzed by using an automatic system for nucleic acid extraction and quantitative real-time polymerase chain reaction (PCR) for herpes simplex 1 and 2 (HSV-1, HSV-2), Epstein-Barr virus (EBV), human cytomegalovirus (HCMV), herpesvirus-6 (HHV-6), and varicella-zoster virus (VZV). HHVs DNA was detected in 52 of 155 (33.5%) analyzed samples. In 39 CSF samples from patients with meningoencephalitis we found: VZV in 13, HSV-1 in 12, EBV in 6, HHV-6 in 4, and HSV-2 in 4. Co-infections of EBV and HSV-2, HSV-1 and HSV-2, HSV-1 and VZV were also disclosed in four cases. In addition, two patients with Guillain-Barré syndrome had HCMV and one showed HHV6 positivity, two patients with myelitis / polymyeloradiculitis had VZV and HCMV respectively, HHV-6 DNA was found in one patient with lateral amyotrophic sclerosis. Three CSF specimens from HIV-infected patients with CNS complications had HHV-6 or EBV DNA. Moreover quantitative data were also correlated to clinical conditions to obtain more information on the virus aetiopathogenic role.

Transplant patients are at high risk of post-transplant lymphoproliferative disorder (PTLD). A strong correlation between Epstein-Barr virus (EBV) and PTLD is observed in pediatric patients with primary infection after transplant. Because many patients have responded to reversal of immunosuppressive therapy, an early identification of EBV is essential for the reduction of immunosuppression and/or introduction of antiviral therapy to prevent PTLD. Polymerase chain reaction (PCR) is a specific and sensitive method to identify EBV DNA in blood. The aim of our study was to establish a protocol for monitoring EBV infection in transplanted patients for early identification those at high risk of PTLD. Viral presence in peripheral blood leukocytes (PBL) and serum samples was revealed by Nested PCR; positive specimens were quantified with Real Time PCR (RT-PCR). DNA in PBL was observed in 12 cases and 6 showed EBV in sera. Quantitative analysis showed a wide range of EBV DNA copies in leukocytes that were higher than in sera. Two patients displayed high viral load values in both PBL and sera associated with clinical evidence of PTLD. Our data suggest that the study of the EBV load represents an essential approach in the diagnosis of PTLD and the analysis of serum samples could provide useful information in the post-transplant monitoring of high-risk patients.

Immunosuppressive status in solid organ transplant recipients is often related to the reactivation of Human Cytomegalovirus (HCMV) infection that remains one of the major causes of morbidity and mortality. Therefore, the early detection of HCMV followed by infection monitoring is important to institute prompt and appropriate treatment. In recent years good results have been obtained by HCMV DNA amplification methods; qualitative and quantitative approaches have shown good sensitivity and specificity, but they often require post-PCR manipulation that adds time to the analysis and may lead to contamination problems. Recently, Real Time PCR (RT-PCR) has been proposed in HCMV DNA analysis as a valid method for its good sensitivity and rapidity. In the present study, twenty-five solid organ transplant recipients were analyzed for HCMV diagnosis; 60 peripheral blood leukocytes and 120 plasma samples were tested by RT-PCR and the results compared to those obtained by a qualitative Nested PCR and a quantitative DNA enzyme immunoassay.

Because of the considerable impact of human cytomegalovirus (HCMV) infection, sensitive, specific, and standardized methods are required for rapid and accurate evaluation of viral load in monitoring transplant recipients. The aim of the present study was to evaluate the usefulness of a multiplex polymerase chain reaction (PCR) for the coamplification of HCMV-DNA and beta-globin genomic sequence in polymorphonuclear leukocytes (PMNL). Analysis and quantification of PCR products were carried out by a DNA enzyme immunoassay (DEIA), which is based on the hybridization of amplified DNA with a single-stranded DNA probe, which coats microtitre wells. Colorimetric detection of the DNA-antibody complex was carried out and optical density (O.D.) was recorded at 450/630 nm. To quantify HCMV/DNA load, a standard curve to which samples O.D. refer was obtained by amplifying serial dilutions of recombinant PGEM-3Z plasmid DNA containing a genomic fragment of glycoprotein B. 340 PMNL specimens from 102 solid organ recipients were tested for the detection of pp65 antigen and HCMV-DNA. The results showed a good correlation between viral load and clinical symptoms of HCMV infection; high specificity and predictive values for HCMV disease were found by PCR, using a cut-off limit of 10(3) genomic copies per 2 x 10(5) PMNL. These findings indicate that the system described is an efficient and reproducible diagnostic method easy to apply for routine diagnosis and therapeutic monitoring of transplanted patients., (Copyright 2000 Wiley-Liss, Inc.)

The use of sensitive and specific methods for rapid and reliable diagnosis is required due to the considerable impact of human cytomegalovirus (HCMV) in organ transplant recipients. For this purpose the demonstration of the presence of viral antigens in peripheral blood leukocytes (PMNLs) and of viral nucleic acids in the same cells or in sera would seem to be of valid support. The present study was designed to test pp65 antigen, HCMV DNA and HCMV late mRNA in order to provide clinical information for the management of the infection. Fifty solid organ recipients were monitored for six months after transplant. The data obtained from the various tests were analysed from the first evidence of HCMV infection revealed by positive antigenaemia and/or DNA-polymerase chain reaction (PCR). In 3 asymptomatic and in 7 symptomatic patients, PCR became positive 1-2 weeks before antigenaemia but PCR did not discriminate the clinical evolution of HCMV infection. The antigenaemia test well correlated to the development of viral infection being positive in all symptomatics and in 31, 2% of asymptomatics. The antigenic load > 100/2 x 10(5) positive cells was always associated with clinical signs of illness. The detection of late mRNA was more indicative of the virus replicative status in the follow-up of patients treated with ganciclovir. In some cases there was evidence, prior to the other two tests, the block of viral replication due to the antiviral therapy and in others the onset of HCMV infection relapse.

The capacity of human and murine polyclonal and monoclonal antibodies to inhibit lipopolysaccharide (LPS)-induced tumor necrosis factor-alpha (TNF-alpha) release from human monocytes was investigated. Human pooled immunoglobulin G (IVIG), human IgM monoclonal antibody (HA-1A) directed against the lipid A moiety of LPS, and murine IgG monoclonal antibody (MT-1F) raised in mice against antibiotic-treated Escherichia coli O6:K- were either added simultaneously with LPS to monocytes or preincubated for 1 h at 37 degrees C before being added to monocytes. TNF-alpha content in the monocyte supernatants was then tested. Simultaneous addition of increasing concentrations of IVIG (from 0.3 to 2.5 mg/ml) and 10 micrograms/ml of LPS to monocytes induced an enhanced release of TNF-alpha by monocytes in a dose dependent fashion. Preincubation of IVIG with LPS abolished the additive effect, but did not inhibit LPS-induced TNF-alpha release by monocytes. The simultaneous addition of LPS and HA-1A to monocytes had no additive effect nor did it inhibit TNF-alpha release. On the other hand, inhibition of TNF-alpha release was observed when HA-1A was preincubated with LPS before being added to monocytes. In all instances MT-1F inhibited TNF-alpha release when the monocytes were stimulated with smooth type LPS, but not with LPS isolated from rough mutants.

In a randomized double blind study, we analyzed the efficacy of IVIG in the infectious complications in patients at high risk of developing sepsis syndrome. Two groups of twenty patients were enrolled, one receiving 250 mg/Kg of IVIG on the first and seventh day after admission and the other receiving sterile saline as placebo. Serum samples were drawn before IVIG administration and 24, 48 and 72 hours afterwards. The same schedule was used for patients treated with placebo. Sera pooled from healthy donors served as controls. On all the samples, opsonic and bactericidal activity as well as C3, total IgG and serum TNF content were tested. IVIG did not significantly affect total IgG and C3 content. Similarly, opsonic and bactericidal activity tested against E. coli 06 :K-, E. coli 0111 and SAC I was not modified ranging within HPS values. Furthermore, IVIG administration did not change the TNF level. A lower incidence of bacteremia in IVIG treated patients was observed.

The activation and differentiation of resting B cells into Ig secreting cells are regulated by T cells, macrophages and their secreted factors. The present study evaluated the effect of cyclosporin A (CsA) on this process. Peripheral blood lymphomonocytes (PBMC) drawn from healthy donors were stimulated with protein A (PA) or with lipopolysaccharides plus pokeweed (LPS+PWM) in either the presence or the absence of CsA. Phenotypic B cell changes and immunoglobulin production was then analyzed. The data revealed that CsA decreased the expression of B cell surface receptors of the activation phase, and enhanced the resting phase receptors. Different effects of CsA were found on B cell differentiation, depending on its induction by PA or LPS+PWM. In the first system, CsA decreased the expression of differentiation phase receptors and the secretion of free Ig. In cultures stimulated with LPS+PWM, CsA increased the differentiated phase receptors and Ig secretion. Thus, CsA seemed to act as a blocking agent of the activation phase and as a modulator of the differentiation phase and of IgG secretion, depending upon the antigen used for stimulation.

Pseudomonas aeruginosa produces several extracellular substances such as enzymes and toxins which seem to contribute to its pathogenicity. In particular, alkaline protease and elastase production seems to affect bacterial adherence. Aim of this study was to isolate an elastase deficient mutant of P. aeruginosa and to demonstrate a possible correlation between enzyme production and adherence to WEHI cells. Mutant strain showed a significant reduction of elastase and protease alkaline activity, as the decrease of absorbance values demonstrate. Furthermore the adherence to WEHI cells of mutant strain was strongly reduced with respect to the wild strain. Our results prove that proteolytic enzymes play an important role in adherence, probably modifying the cell surfaces and so enhancing adherence.

Since infections represent one of the greatest complications in immunocompromised hosts, our group, concerned with bacteriological monitoring, has applied its attention to microorganisms such as Pseudomonas sp, Staphylococcus sp and Candida sp. Pseudomonas sp strains have been identified using conventional techniques and the API system 20 NE. According to Varaldo's scheme, Staphylococci sp have been divided into six "lyogroup" and then, the typing of Candida sp has been checked by cultural and biochemical tests. The results obtained have demonstrated that the Pseudomonas aeruginosa species is prevalent not only in the urinary tract but also in the respiratory tract. As for the Staphylococci sp, the VI lyogroup showed the greatest percentage of strains and than, as regards Candida sp it has been possible to observe that, over a given period, the percentage of pluricontamination has been almost constant.

A total of 274 recently isolated Gram-positive e Gram-negative strains have been tested. The method used for the determination of the minimal inhibitory concentration of the antibiotics was by agar dilution and the minimal bactericidal concentrations were determined by "replica plating" system. All strains demonstrated a good sensitivity, above all, to Norfloxacin which inhibited Gram-positive bacteria at the concentration of 3.12 micrograms/ml and, concerning Gram-negative, at the concentration of 0.10 microgram/ml for K.E.S. group, 0.19 microgram/ml for Escherichia coli, 6.25 micrograms/ml for Proteus sp and 1.56 microgram/ml for Pseudomonas aeruginosa. Also with the other antibiotics we obtained good results especially with Nitrofurantoin concerning Enterococcus and Ceftazidime concerning all the strains. On the basis of these results we can conclude that Norfloxacin is one of the most active antibiotics among those used against strains implicated in urinary infections.

Recent clinical isolates were tested for production of some extracellular factors such as alkaline protease and elastase. They were also assayed for adhesiveness to WEHI cells. It is well known that extracellular production of substances other than toxins is related to virulence and may increase adherence. The present investigation aimed to evaluate the role of extracellular proteins in adherence. Alkaline protease production was assayed using a test performed with casein as substrate while elastase activity was investigated with the elastin-congored method. Our results demonstrated that P. aeruginosa strains which are good alkaline protease and elastase producers adher better than those showing no or low protease and elastase activity.