Severe Acute Respiratory Infections (SARI) surveillance in over-65-years-old patients: the experience of a University hospital (seasons 2017-2018 and 2018-2019).

Background: Influenza is a relevant public health problem, also due to the risk of complications. The most effective measure to prevent influenza is vaccination; therefore, at present, there is consensus among European countries, regarding the need for routine seasonal influenza vaccination of elderly and individuals at increased risk of severe influenza. At the same time, influenza surveillance is necessary to understand the viruses circulating and effectiveness of vaccination strategies. The present study reports the results of two seasons influenza surveillance (2017/2018 and 2018/2019) conduced in an University Hospital in Rome among hospitalized patients aged ≥65 years. Study design: A prospective cohort study. Methods: The study consisted of systematic daily screening of all admissions among patients aged ≥65 years meeting a syndromic SARI case definition during two consecutive influenza seasons: 2017/2018 and 2018/2019. Characteristics of patients and their risk factors were collected by a standardized questionnaire and nose-pharyngeal swabs were performed to each patient. Influenza vaccine effectiveness (IVE), rates of vaccinated subjects and case fatality rate were also evaluated. Results: Influenza was laboratory confirmed in 11 (9.9%) of the 111 and 11 (9.6%) of the 115 enrolled patients in seasons 2017/18 and 2018/19, respectively. Adjusted IVE against all influenza type, calculated for each season, was 88.5% (95% CI: 38.9 to 97.8) and 61.7% (95% CI: -59.9 to 90.9) for 2017/2018 and 2018/2019 seasons, respectively. Our analysis shows a Case Fatality Rate of 2.7% and 4.3% for the 2017/18 and 2018/19 seasons, respectively. Conclusions: The surveillance of SARI conduced in one hospital in Rome confirmed that influenza is an important cause of hospital admissions. Routine monitoring of infectious diseases and related aetiology associated with SARI, also at the local-level, is useful for targeting the right preventive measures.

Biodistribution and intracellular localization of hyaluronan and its nanogels. A strategy to target intracellular S. aureus in persistent skin infections.

Intracellular pathogens are a critical challenge for antimicrobial therapies. Staphylococcus aureus (S. aureus) causes approximately 85% of all skin and soft tissue infections in humans worldwide and more than 30% of patients develop chronic or recurrent infections within three months, even after appropriate antibacterial therapies. S. aureus is also one of the most common bacteria found in chronic wounds. Recent evidences suggest that S. aureus is able to persist within phagolysosomes of skin cells (i.e. keratinocytes, phagocytic cells), being protected from both the immune system and a number of antimicrobials. To overcome these limits, nano-formulations that enable targeted therapies against intracellular S. aureus might be developed. Herein, the biodistribution and intracellular localisation of hyaluronan (HA) and HA-based nanoparticles (nanogels, NHs) are investigated, both after intravenous (i.v.) injections (in mice) and topical administrations (in ex vivo human skin). Results indicate HA and NHs accumulate especially in skin and liver of mice after i.v. injection. After topical application on human skin explants, no penetration of both HA and NHs was detected in skin with intact stratum corneum. By contrast, in barrier-disrupted human skin (with partial removal and loosening of stratum corneum), HA and NHs penetrate to the viable epidermis and are taken up by keratinocytes. In mechanically produced wounds (skin without epidermis) they accumulate in wound tissue and are taken up by dermis cells, e.g. fibroblasts and phagocytic cells. Interestingly, in all cases, the cellular uptake is CD44-mediated. In vitro studies confirmed that after CD44-mediated uptake, both HA and NHs accumulate in lysosomes of dermal fibroblasts and macrophages, as previously reported for keratinocytes. Finally, the colocalisation between intracellular S. aureus and HA or NHs is demonstrated, in macrophages. Altogether, for the first time, these results strongly suggest that HA and HA-based NHs can provide a targeted therapy to intracellular S. aureus, in persistent skin or wound infections.

Surveillance of healthcare acquired infections by "alert microorganisms": preliminary results.

BACKGROUND: We evaluated the trend of four years (2015 - 2018) of "alert organisms" surveillance carried out at the 450 bed teaching hospital S. Andrea in Rome. METHODS: All patients with an "Alert organism" isolation were screened. In accordance with definitions used by the Centers for Disease Control patients with an "alert organism" isolation were evaluated for infection or colonization, by an infection control team (ICT). RESULTS: Between April 2015 and December 2018 a total 4,762 specimens with "Alert organism" isolation were screened and 1,601 patients were surveyed and included in the study. Overall 780 (48.8%) patients developed an healthcare acquired infection (HAI) at our institution, whereas 311 (19.4%) entered with a community acquired infection, 254 (15.8%) with an infection acquired in another healthcare setting and 256 (16.0%) resulted simply colonized. The 780 patients who developed an HAI at our institution presented 878 infectious episodes and the isolation of 931 microorganisms. C. difficile infections were the most common (27.2%), followed by 21.3% respiratory tract infections, 16.9% urinary tract infections, 15.5% surgical site infections, 12.5% bloodstream infections, 3.6% ulcers and 3.0% others. Among HAI group Gram negatives (54.1%) were more frequent than Gram positives (45.9%), whereas in patients entering in the hospital already with a community infection Gram positives overpassed Gram negatives (58.7% vs. 41.3%; p<0.001). Most common pathogens responsible for HAI were C. difficile (25.6%), Klebsiella spp. (25.5%), MRSA (19.6%) and Acinetobacter spp. (15.3%). Notably 30.0% HAI at other institutions were represented by C. difficile. Impressively, >40% of community acquired infections were related to MRSA. CONCLUSIONS: The present study provided some useful insight into the major multi-resistant pathogens epidemiology at our institution. The Authors succeeded in organizing a multidisciplinary ICT that created a partnership feeling with the hospital personnel.

Risk factors for recurrence in patients with Clostridium difficile infection due to 027 and non-027 ribotypes.

OBJECTIVES: Our objective was to evaluate factors associated with recurrence in patients with 027+ and 027- Clostridium difficile infection (CDI). METHODS: Patients with CDI observed between January and December 2014 in six hospitals were consecutively included in the study. The 027 ribotype was deduced by the presence of tcdB, tcdB, cdt genes and the deletion Δ117 in tcdC (Xpert® C. difficile/Epi). Recurrence was defined as a positive laboratory test result for C. difficile more than 14 days but within 8 weeks after the initial diagnosis date with reappearance of symptoms. To identify factors associated with recurrence in 027+ and 027- CDI, a multivariate analysis was performed in each patient group. Subdistributional hazard ratios (sHRs) and 95% confidence intervals (95%CIs) were calculated. RESULTS: Overall, 238 patients with 027+ CDI and 267 with 027- CDI were analysed. On multivariate analysis metronidazole monotherapy (sHR 2.380, 95%CI 1.549-3.60, p <0.001) and immunosuppressive treatment (sHR 3.116, 95%CI 1.906-5.090, p <0.001) were factors associated with recurrence in patients with 027+ CDI. In this patient group, metronidazole monotherapy was independently associated with recurrence in both mild/moderate (sHR 1.894, 95%CI 1.051-3.410, p 0.033) and severe CDI (sHR 2.476, 95%CI 1.281-4.790, p 0.007). Conversely, non-severe disease (sHR 3.704, 95%CI 1.437-9.524, p 0.007) and absence of chronic renal failure (sHR 16.129, 95%CI 2.155-125.000, p 0.007) were associated with recurrence in 027- CDI. CONCLUSIONS: Compared to vancomycin, metronidazole monotherapy appears less effective in curing CDI without relapse in the 027+ patient group, independently of disease severity.

Cervicovaginal bacteria and fungi in pregnant diabetic and non-diabetic women: a multicenter observational cohort study.

OBJECTIVE: We evaluated the prevalence of cervicovaginal Bacteria, group B Streptococcus (GBS), Gardnerella vaginalis (GV), Candida spp., Chlamydia trachomatis (CT), Mycoplasma hominis (MH) and Ureaplasma urealyticum (UU) in pregnant women with and without diabetes mellitus (DM). PATIENTS AND METHODS: Cervicovaginal swabs were gathered from 473 pregnant patients divided into 127 diabetic and 346 non-diabetic. The results were correlated to gestational age, parity and glycemic control. RESULTS: A higher prevalence of MH/UU (p=0.012) was found in the diabetic patients. After the 28th week of pregnancy, the prevalence for all investigated microorganisms appeared similar except for MH/UU (p=0.014). In multigravida, there were statistical differences between two groups in testing for Bacteria (p=0.015) and for MH/UU (p=0.037). The diabetic condition correlated to the state of multigravida in cases positive for Candida spp. (p=0.049) and in those testing positive for at least one microorganism (p=0.043). Pregnant with a blood glucose > 92 have twice the risk of being positive to a single microbiological test than those with better glycemic control. CONCLUSIONS: The higher prevalence of MH/UU after the 28th weeks can be explained with the physiologically reduced insulin tolerance characteristic of this gestational period. Among the diabetic testing positive to Candida spp. the statistically significant association was observed only in multigravida condition. These data suggest that diabetic multigravida women are at increased risk for Candida spp. infection in relation to the improper glycemic control.

Imaging bacteria with radiolabelled quinolones, cephalosporins and siderophores for imaging infection: a systematic review.

Bacterial infections are still one of the main causes of patient morbidity and mortality worldwide. Nowadays, many imaging techniques, like computed tomography or magnetic resonance imaging, are used to identify inflammatory processes, but, although they recognize anatomical modifications, they cannot easily distinguish bacterial infective foci from non bacterial infections. In nuclear medicine, many efforts have been made to develop specific radiopharmaceuticals to discriminate infection from sterile inflammation. Several compounds (antimicrobial peptides, leukocytes, cytokines, antibiotics…) have been radiolabelled and tested in vitro and in vivo, but none proved to be highly specific for bacteria. Indeed factors, including the number and strain of bacteria, the infection site, and the host condition may affect the specificity of tested radiopharmaceuticals. Ciprofloxacin has been proposed and intensively studied because of its easy radiolabelling method, broad spectrum, and low cost, but at the same time it presents some problems such as low stability or the risk of antibiotic resistance. Therefore, in the present review studies with ciprofloxacin and other radiolabelled antibiotics as possible substitutes of ciprofloxacin are reported. Among them we can distinguish different classes, such as cephalosporins, fluoroquinolones, inhibitors of nucleic acid synthesis, inhibitors of bacterial cell wall synthesis and inhibitors of protein synthesis; then also others, like siderophores or maltodextrin-based probes, have been discussed as bacterial infection imaging agents. A systematic analysis was performed to report the main characteristics and differences of each antibiotic to provide an overview about the state of the art of imaging infection with radiolabelled antibiotics.

Saccharomyces cerevisiae fungemia, a possible consequence of the treatment of Clostridium difficile colitis with a probioticum.

The yeast Saccharomyces boulardii is a biotherapeutic agent used for the prevention and treatment of several gastrointestinal diseases, such as diarrhoea caused by Clostridium difficile, in addition to the antibiotic therapy. In this study we report a case of Saccharomyces cerevisiae fungemia in a patient with Clostridium difficile-associated diarrhoea (CDAD) treated orally with S. boulardii in association with vancomycin. The identification of the S. cerevisiae was confirmed by molecular technique. Fungemia is a rare, but a serious complication to treatment with probiotics. We believe it is important to remind the clinicians of this risk when prescribing probiotics, especially to immunocompromised patients.

Microbiological and molecular characterization of extreme drug-resistant carbapenemase-producing Klebsiella pneumoniae isolates.

Fifteen Klebsiella pneumoniae clinical isolates showing non-susceptibility to carbapenems and resistant to colistin were collected in an Italian hospital. All isolates resulted negative to AmpC, MBL and ESBL production but positive to modified Hodge test, therefore were evaluated as KPC producers. The presence of blaKPC genes was verified by polymerase chain reaction (PCR) and sequencing. Furthermore, molecular typing was performed by multilocus sequence typing (MLST). PCR analysis and nucleotide sequencing revealed that all 15 isolates carried the blaKPC-3 gene. MLST analysis attributed the isolates from all patients to belong to the sequence type ST512. All isolates showed extensively drug-resistant (XDR) phenotype. The emergence of colistin-resistant K. pneumoniae underscores the implementation of strict control measures to prevent the dissemination of these organisms in hospitals.

Kodamaea ohmeri isolate from two immunocompromised patients: first report in Italy.

Kodamaea ohmeri is an unusual yeast-form fungus that has recently been identified as an important aetiological agent of fungaemia, endocarditis, cellulitis, funguria and peritonitis in immunocompromised patients. We present two new isolated of K. ohmeri. The microorganisms were identified by CHROMagar Candida medium, VitekII system and API ID32C. Biochemical identification of the two yeast isolates was confirmed by sequence analysis of the 26S ribosomal DNA. Antifungal susceptibility testing done by Sensititre YeastOne showed that the isolates were susceptible to amphotericin B, voriconazole and itraconazole. This work is the first report of isolation of K. ohmeri in immunocompromised patients in Italy.

Detection of different Borrelia burgdorferi genospecies in serum of people with different occupational risks: short report.

This study is aimed at applying a previously described PCR-based method to detect B. burgdorferi sensu lato and different Borrelia genospecies in total DNA preparations of serum samples collected from people with different occupational risks for tick bite and with serological evidence of borreliosis. Among the seropositive samples, the PCR for B. burgdorferi confirmed the positivity in 65 percent of the forestry workers and in 60 percent of the subjects living in the same area. None of the seronegative subjects belonging to the control group showed the presence of B. burgdorferi sensu lato DNA. Results on genospecies distribution show that B. afzelii was the predominant species, followed by B. garinii and finally by B. valaisiana.

Chlamydia pneumoniae and atherosclerosis: current state and future prospectives.

Chlamydia pneumoniae, an intracellular bacterial pathogen, is known as a leading cause of human respiratory tract infections worldwide. Over the last decade, several reports in the literature have suggested that infection with C. pneumoniae may contribute to the pathogenesis of atherosclerosis. In order to play a causative role in chronic disease, C. pneumoniae would need to persist within infected tissue for extended periods of time, thereby stimulating a chronic inflammatory response. C. pneumoniae has been shown to disseminate systemically from the lungs through infected peripheral blood mononuclear cells and to localize in arteries where it may infect endothelial cells, vascular smooth muscle cells, monocytes/macrophages and promote inflammatory atherogenous process. The involvement of C. pneumoniae in atherosclerosis was investigated by seroepidemiological and pathological studies, in vivo and in vitro studies, and in clinical antibiotic treatment trials. This review will provide an update on the role of C. pneumoniae in atherosclerosis focusing on the recent insights and suggesting areas for future research.

Evaluation by real-time PCR of the expression of S. flexneri virulence-associated genes ospB and phoN2 under different genetical backgrounds.

Under conditions of activated type III secretion Shigella flexneri up-regulates the expression of numerous genes, including the virulence plasmid (pINV)-encoded ospB and phoN2 genes. ospB and phoN2 are virulence-associated genes which are part of a bicistronic transcriptional unit encoding OspB, a protein (effector) of unknown function secreted by the type III secretion (TTS) apparatus, and PhoN2 (apyrase or ATP-diphosphohydrolase), a periplasmic protein involved in polar IcsA localization on the surface of S. flexneri. In this work we used real-time PCR to measure transcription of ospB and phoN2 of wild-type S. flexneri strain M90T as well as of derivative mutants impaired in definite virulence traits. The results obtained confirmed and extended previous reports indicating that the expression of ospB and phoN2 genes is modulated in a virB-dependent, mxiE-independent manner under conditions of non-activated secretion, while their expression is considerably induced in a mxiE-dependent manner under conditions of activated secretion. That the expression of the ospB-phoN2 operon is up-regulated in condition of activated secretion, indicates that probably the expression of these two genes might be important, especially during the later stages of infection of S. flexneri.

No evidence of involvement of Chlamydia pneumoniae in lung cancer by means of quantitative real-time polymerase chain reaction.

Chlamydia pneumoniae, an obligate intracellular pathogen, is well-known as etiological agent of acute respiratory infections; the repeated or prolonged exposure to chlamydial antigens may promote the persistence of C. pneumoniae in the respiratory tract leading to chronic diseases, such as chronic obstructive pulmonary disease and asthma. The predilection of C. pneumoniae to cause respiratory tract infections combined with its persistent nature suggest that it might play a role in lung cancer. The aim of our study is to evaluate the involvement of C. pneumoniae in pathogenesis of lung cancer. We therefore investigated the presence of C. pneumoniae DNA in tumor lung tissues by using real-time PCR assay. Simultaneously, tumor and healthy tissues from the same patient with primary carcinoma lung were analyzed. C. pneumoniae DNA was not detected in a single lung tumor tissue by means of an highly sensitive, and specific real-time PCR assay based on FRET hybridization probes. In conclusion, this study does not support the involvement of C. pneumoniae in the pathogenesis of lung cancer, suggesting that further investigations are needed to clarify other potential causative factors for the development of this malignancy.

Genotype and phenotype relationship in MLSb resistance in Streptococcus pyogenes and Streptococcus agalactiae isolated in central Italy.

The aim of this work is to study a correlation between phenotype and genotype in clinical isolates of erythromycin-resistant Streptococcus spp. Among the 25 erythromycin-resistant S. pyogenes, we detected six strains with iMLSB, nine with cMLSB and two with M phenotypes. Among 14 erythromycin-resistant S. agalactiae, we detected five strains with iMLSB, seven with cMLSB and none with an M phenotype. Moreover, 8 S. pyogenes and 2 S. agalactiae showed a phenotype not matching the known ones described in literature, defining an unknown phenotype. Upon examination, the genetic profiles, erm(A), erm(B) and mef(A), of the clinical isolates did not easily correlate with a specific phenotype. Our findings highlighted that the whole matter of phenotypic diversity in macrolide-resistant S. pyogenes and S. agalactiae strains and the correlation with their genetic profiles should be submitted to a more careful analysis of phenotypic and genotypic characterization.

In vitro susceptibility of isolates of Borrelia burgdorferi s.l. to antimicrobial agents.

In the present study, we investigate the in vitro antimicrobial activity of macrolides, beta-lactams and tetracycline against Borrelia burgdorferi s.l. clinical and tick isolates. Minimal inhibitory concentrations (MICs) were determined in normal growth condition and after pre-exposure of the strains to sub-MIC of the founder of each drug family. All the classes of tested antibiotics showed good antibacterial activity against all the borreliae isolates and there were no significant susceptibility differences among clinical and tick isolates. After pre-exposure of the strains to sub-MIC of erythromycin, cefoxitin and tetracycline, we observed that some strains of B. burgdorferi s.l. showed higher MIC values to both the pre-exposed drug and drugs of the same family. The less susceptibility of borreliae, in the last growth condition in vitro, could be one of the justifications of clinical results indicating the limited efficacy of these antibiotics in treatment of B. burgdoferi infections.

Chlamydia pneumoniae in asymptomatic carotid atherosclerosis.

We evaluated, in 415 patients with asymptomatic carotid atherosclerosis: (i) the prevalence of C. pneumoniae DNA in atherosclerotic carotid plaques and peripheral blood mononuclear cells (PBMC); (ii) the distribution of C. pneumoniae in atherosclerotic carotid plaques and PBMC from the same patients; (iii) the correlation between circulating anti-chlamydial antibodies and the presence of C. pneumoniae DNA. Overall, 160 atherosclerotic carotid plaques and 174 PBMC specimens from patients with asymptomatic carotid atherosclerosis were examined by ompA nested touchdown PCR for presence of C. pneumoniae. In addition, C. pneumoniae DNA was detected in 81 specimens of atherosclerotic carotid plaque and PBMC obtained from the same patients. C. pneumoniae DNA was found in 36.9% of atherosclerotic carotid plaques and in 40.2% of PBMC specimens examined (P=NS). With regard to 81 patients, C. pneumoniae DNA was detected in 27.2% of atherosclerotic carotid plaques and in 44.4% of PBMC specimens(P=0.05). In 18 patients, the presence of C. pneumoniae DNA in PBMC specimens and atherosclerotic carotid plaques coincided (P=0.005). No statistically significant association was found between anti-C. pneumoniae antibodies (IgG and IgA) and positive PCR results. In conclusion, our results suggest that the detection of C. pneumoniae DNA in PBMC specimens seems to be a first-choice method to identify the patients at risk for endovascular chlamydial infection.

Detection of Chlamydia pneumoniae in atherosclerotic coronary arteries.

Chlamydia pneumoniae has recently been associated with the development of coronary heart diseases by sero-epidemiological studies and by direct detection of the organism in atherosclerotic tissues. The aim of our study was to employ a semi-nested PCR approach to investigate the presence of C. pneumoniae in both normal and atherosclerotic coronary arteries of humans obtained at autopsy. Moreover, we have evaluated the role of infection with C. pneumoniae in relation to the extent of coronary atherosclerosis. One hundred and eighty coronary artery specimens were collected at autopsy from 60 consecutive subjects (three arterial segments from each subject). Atherosclerosis in each arterial segment was graded histologically by the Stary classification. Thirty normal coronary arteries were also taken at autopsy as control. PCR results evidenced the presence of C. pneumoniae DNA in atherosclerotic coronary arteries in 19 (31.7%) of 60 subjects examined, while none of the 30 subjects with non-atherosclerotic tissues was positive (p=0.001). Moreover, of the 180 atherosclerotic specimens examined, C. pneumoniae DNA was detected in 3.4% (2/59) of mild atherosclerotic lesions, and in 14.0% (17/121) of advanced atherosclerotic lesions (p=0.05). Our results demonstrate that the presence of C. pneumoniae DNA may be associated with the severity of coronary atherosclerosis.

Multicentric study of seroprevalence of Borrelia burgdorferi and Anaplasma phagocytophila in high-risk groups in regions of central and southern Italy.

The aim of this study was to evaluate the seroprevalence of B. burgdorferi and A. phagocytophila in populations of workers from 4 Italian regions, known to be exposed to tick bites. A total of 712 serum samples collected were divided as follows: 387 samples were obtained from workers at risk for tick bites and 325 from individuals that were not considered to be at risk of ticks bites and served as the control group. Antibodies against B. burgdorferi were found in 29 (7.5%) of the 387 risk workers and in 4 (1.2%) of the 325 control group. Antibodies reactive with the HGE agent were found in 22 (5.7%) of the 387 risk workers and in 3 (0.9%) of the 325 control group. Antibodies to both B. burgdorferi and A. phagocytophila were found in 1.6% of the forestry workers confirming the possibility of coinfection or concurrent infection. The present finding show significant differences between seroprevalence of the risk workers and that of the people with no risk for tick exposure.