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To evaluate theThe activity of antibiotics against planktonic cells was assessed by performing MIC, MBC, and time-kill assays. The activity against mature biofilms was evaluated, in a microtiter plate, both in terms of dispersion (crystal violet assay) and residual viability (viable cell count). The effect of drug exposure on selectedApramycin MICOur results warrant future pharmacokinetic and pharmacodynamic studies for supporting the rationale to repurpose apramycin, a veterinary aminoglycoside, for CF lung infections.

As a consequence of the current developments in the automotive industry and especially in electric vehicle manufacturing, new legitimate aspects regarding the safety of people, systems and infrastructures must be addressed. The need for higher powertrain power density is driving battery voltages up to 1200 Vdc, which poses significant safety challenges. Fatigue and instantaneous loads on battery and high voltage components in the form of historical data can support OEMs' efforts to improve safety and production effectiveness. Therefore, it is essential to monitor vehicle batteries individually and acknowledge them when necessary. This paper describes a wireless monitoring system that collects battery data and transmits it wirelessly (LoRa/Wi-Fi/4G) to end users to inform them about battery status while also generating historical data about battery profile. The purpose of this application is to ensure safer battery practices (charging/storage/maintenance) by informing the end-user of any abnormal condition, as well as to store historical data that can be used for battery behavior analysis under different operating conditions. Finally, we present a practical example of this system applied by Energica Motor Company in the FIM Enel MotoE World Cup.

Bloodstream infections (BSIs) remain a potentially life-threatening condition. The gold standard for the diagnosis of BSI is still blood cultures (BCs), and the diagnostic yield depends on clinical and technical factors that have an impact on collection and transportation. Hence, monitoring of the entire pre-analytical process from blood collection to transportation to the microbiology laboratory is critical. To optimize the clinical impact of the diagnostic and therapeutic procedures, a multidisciplinary approach and univocal protocols are mandatory. A board of specialists discussed the available evidence on the pre-analytical process and produced the present document to guide physicians and nurses on the ideal execution of BC: (1) timing and preparation for blood collection; (2) skin antisepsis; (3) blood volume; (4) sampling method and safety; (5) medium to be used; (6) time to BC transportation; and (7) quality assurance and quality management.

The in vitro activity of ceftolozane-tazobactam (C-T) was evaluated comparatively to other antibiotics against 188 Pseudomonas aeruginosa isolates collected from cystic fibrosis (CF) patients. Overall, the activity of C-T was comparable to colistin (susceptibility rate: 85.1% vs. 89.4%) but significantly higher than other antimicrobials. Particularly, C-T was active against 70% of meropenem nonsusceptible isolates and 64.1% of those nonsusceptible to beta-lactams. C-T was active against 70%, 58.1%, and 100% of multidrug-resistant, extensively drug-resistant (XDR), and pandrug-resistant isolates, respectively. No differences in C-T activity were found between isolates from children and adult patients, except for XDR ones significantly more susceptible in older patients. C-T and colistin exhibited comparable susceptibility rate (91.1% vs. 86.7%) also against 68 isolates collected during pulmonary exacerbations. Activity of C-T towards mucoid isolates was less than colistin (82.9% vs. 97.6%) but higher compared with other antibiotics. C-T represents a promising agent for treating CF lung infections.

Staphylococcus aureus represents a major human pathogen able to cause a number of infections, especially bloodstream infections (BSI). Clinical use of methicillin has led to the emergence of methicillin-resistant S. aureus (MRSA) and MRSA-BSI have been reported to be associated with high morbidity and mortality. Clinical diagnosis of BSI is based on the results from blood culture that, although considered the gold standard method, is time-consuming. For this reason, rapid diagnostic tests to identify the presence of methicillin-susceptible S. aureus (MSSA) and MRSA isolates directly in blood cultures are being used with increasing frequency to rapidly commence targeted antimicrobial therapy, also in the light of antimicrobial stewardship efforts. Here, we review and report the most common rapid non-molecular and molecular methods currently available to detect the presence of MRSA directly from blood.

Aim: Carbapenemase-resistant Enterobacteriaceae represents a major concern in hospital setting. Materials & methods: The evolutionary history of carbapenem-resistant Klebsiella pneumonia strains was analyzed by core genome multilocus sequence typing and Bayesian phylogenesis by whole genomes sequencing. Results: A great increase carbapenem-resistant K. pneumoniae causing blood stream infection was observed in the years 2015–2016. At multilocus sequence typing (MLST), they were prevalently ST512 and ST101. ST512 were core genome (cg)MLST 53, while ST101 mainly cgMLST453. The minimum-spanning tree, based on cgMLST, showed strains clustering based on the different STs. By Bayesian phylogenetic analysis, maximum clade credibility tree showed that strains were introduced in the year 2005 with the most probable location in the ICU ward. Two outbreaks by ST101 and ST512 strains with Tower T8 as the probable location were evidenced. Conclusion: Molecular epidemiology is a powerful tool to track the way of transmission of resistant bacteria within the hospital setting.

The intrinsic antibiotic resistance of Stenotrophomonas maltophilia, along with its ability to form biofilm both on abiotic surfaces and host tissues, dramatically affects the efficacy of the antibiotic therapy. In this work, 85 S. maltophilia strains isolated in several hospital of central Italy and from several clinical settings were evaluated for their genetic relatedness (by pulsed-field gel electrophoresis, PFGE), biofilm formation (by microtiter plate assay), and planktonic antibiotic resistance (by Kirby&ndash, Bauer disk diffusion technique). The S. maltophilia population showed a high genetic heterogeneity: 64 different PFGE types were identified, equally distributed in cystic fibrosis (CF) and non-CF strains, and some consisted of multiple strains. Most of the strains (88.2%) were able to form biofilm, although non-CF strains were significantly more efficient than CF strains. CF strains produced lower biofilm amounts than non-CF strains, both those from respiratory tracts and blood. Non-CF PFGE types 3 and 27 consisted of strong-producers only. Cotrimoxazole and levofloxacin were the most effective antibiotics, being active respectively against 81.2% and 72.9% of strains. CF strains were significantly more resistant to piperacillin/tazobactam compared to non-CF strains (90% vs. 53.3%), regardless of sample type. Among respiratory strains, cotrimoxazole was more active against non-CF than CF strains (susceptibility rates: 86.7% versus 75%). The multidrug resistant phenotype was significantly more prevalent in CF than non-CF strains (90% versus 66.7%). Overall, the multidrug-resistance level was negatively associated with efficiency in biofilm formation. Our results showed, for the first time, that in S. maltophilia both classical planktonic drug resistance and the ability of biofilm formation might favor its dissemination in the hospital setting. Biofilm formation might act as a survival mechanism for susceptible bacteria, suggesting that clinical isolates should be routinely assayed for biofilm formation in diagnostic laboratories.

The use of PCV7 for children immunization was gradually implemented in the Italian regions starting from 2006 and was replaced by PCV13 in 2010-2011. In this study we aimed to assess the PCV impact on invasive pneumococcal diseases (IPD) incidence, serotype distribution and antibiotic resistance in Italian children under 5years old.All IPD cases in children from 5 Italian regions (Emilia-Romagna, Lombardia, A. P. Bolzano, A. P. Trento, and Piemonte) reported through the nationwide surveillance system during 2008-2014 were included in this study. Pneumococcal isolates were subjected to serotyping, antibiotic susceptibility testing, and clonal analysis according to standard methods.During the study period overall IPD incidence decreased from 7.8 cases/100,000 inhabitants in 2008 to 3.0 cases/100,000 in 2014 (61% decrease, P0.001). In particular, from 2008 to 2014, PCV7-type IPD decreased from 2.92 to 0.13 cases/100,000 inhabitants (95% decrease, P0.001) while PCV13-non-PCV7 type IPD decreased from 3.2 to 0.89 cases/100,000 inhabitants (72% decrease, P=0.008). Conversely, non-vaccine serotype (NVS) IPD increased overtime, becoming more common than PCV13 serotype IPD in 2013-2014. Emergent NVS 24F and 12F were the most prevalent in 2014. Antibiotic resistance testing revealed an overall increasing trend in penicillin resistance, from 14% in 2008 to 23% in 2014. Erythromycin resistance showed a downward trend, from 38% in 2008 to 27% in 2014. While in 2008 PCV13 serotypes were the major responsible for antibiotic resistance, during the following years antimicrobial resistance due to NVS increased, mainly as a result of expansion of pre-existing clones.Both PCVs led to a substantial decrease in vaccine-related IPD incidence in the children population. However NVS-related IPD increased, becoming the most prevalent in the last two-years period. Continuous surveillance is an essential tool to monitor evolution of pneumococcal population causing IPD in children.

The intrinsic antibiotic resistance of Stenotrophomonas maltophilia, along with its ability to form biofilm both on abiotic surfaces and host tissues, dramatically affects the efficacy of the antibiotic therapy. In this work, 85 S. maltophilia strains isolated in several hospital of central Italy and from several clinical settings were evaluated for their genetic relatedness (by pulsed-field gel electrophoresis, PFGE), biofilm formation (by microtiter plate assay), and planktonic antibiotic resistance (by Kirby–Bauer disk diffusion technique). The S. maltophilia population showed a high genetic heterogeneity: 64 different PFGE types were identified, equally distributed in cystic fibrosis (CF) and non-CF strains, and some consisted of multiple strains. Most of the strains (88.2%) were able to form biofilm, although non-CF strains were significantly more efficient than CF strains. CF strains produced lower biofilm amounts than non-CF strains, both those from respiratory tracts and blood. Non-CF PFGE types 3 and 27 consisted of strong-producers only. Cotrimoxazole and levofloxacin were the most effective antibiotics, being active respectively against 81.2% and 72.9% of strains. CF strains were significantly more resistant to piperacillin/tazobactam compared to non-CF strains (90% versus 53.3%), regardless of sample type. Among respiratory strains, cotrimoxazole was more active against non-CF than CF strains (susceptibility rates: 86.7% versus 75%). The multidrug resistant phenotype was significantly more prevalent in CF than non-CF strains (90% versus 66.7%). Overall, the multidrug-resistance level was negatively associated with efficiency in biofilm formation. Our results showed, for the first time, that in S. maltophilia both classical planktonic drug resistance and the ability of biofilm formation might favor its dissemination in the hospital setting. Biofilm formation might in fact act as a survival mechanism for susceptible bacteria, suggesting that clinical isolates should be routinely assayed for biofilm formation in diagnostic laboratories.

As the cybersecurity risks in automotives grow due to vehicles increasing connectivity, enhancing the cybersecurity cognition of automotive networks has become an urgent requirement in industry. While modern microcontrollers offer ARM’s based Trustzone feature to secure critical applications, secure intra-vehicular protocols are required to design next generation vehicles. With standard security solutions not being practical to vehicles due to resource constraints and compatibility issues, in this work we present a lightweight technique to enable both a virtual trusted channel and a normal untrusted channel over the same physical CAN-bus network, which we call TrustNet. The goal of TrustNet is to secure CAN-bus sensitive communications by protecting against masquerade and replay attacks with minimum overhead and full legacy support, and at the same time to provide normal-world communications.

Streptococcus bovis/Streptococcus equinus complex (SBSEC), a non-enterococcal group D Streptococcus spp. complex, has been described as commensal bacteria in humans and animals, with a fecal carriage rate in humans varying from 5% to over 60%. Among streptococci, SBSEC isolates represent the most antibiotic-resistant species—with variable resistance rates reported for clindamycin, erythromycin, tetracycline, and levofloxacin—and might act as a reservoir of multiple acquired genes. Moreover, reduced susceptibility to penicillin and vancomycin associated with mobile genetic elements have also been detected, although rarely. Since the association of SBSEC bacteremia and colon lesions, infective endocarditis and hepatobiliary diseases has been established, particularly in elderly individuals, an accurate identification of SBSEC isolates to the species and subspecies level, as well as the evaluation of antibiotic resistance, are needed. In this paper, we reviewed the major methods used to identify SBSEC isolates and the antimicrobial resistance rates reported in the scientific literature among SBSEC species.

Purpose. Staphylococcus aureus isolates, collected from various clinical samples, were analysed to evaluate the contribution of the genetic background of both erythromycin-resistant (ERSA) and -susceptible (ESSA) S. aureus strains to biofilm formation. Methods. A total of 66 ESSA and 43 ERSA clinical isolates were studied for adhesiveness and biofilm formation under different atmospheres. All isolates were evaluated for phenotypic and genotypic macrolide resistance, and for clonal relatedness by pulsed-field gel electrophoresis (PFGE), and by spa typing on representative isolates. Results. A high genetic heterogeneity was encountered, although 10 major PFGE types accounted for 86 % with a few small spatially and temporally related clusters. Overall, biofilm formation under anoxia was significantly lower than under oxic and micro-aerophilic atmospheres. Biofilm formation by ESSA was significantly higher compared to ERSA under oxic and micro-aerophilic conditions. Adhesiveness to plastic was significantly higher among respiratory tract infection isolates under micro-aerophilic conditions, while surgical site infection isolates formed significantly higher biomass of biofilm under oxic and micro-aerophilic atmospheres compared to anoxia. Pulsotype 2 and 4 strains formed significantly higher biofilm biomass than pulsotype 1, with strains belonging to CC8 forming significantly more compared to those belonging to CC5, under both oxic and micro-aerophilic atmospheres. Conclusions. S. aureus biofilm formation appears to be more efficient in ESSA than ERSA, associated with specific S. aureus lineages, mainly CC8 and CC15, and affected by atmosphere. Further studies investigating the relationship between antibiotic resistance and biofilm formation could prove useful in the development of new strategies for the management of S. aureus infections.

With the increasing connectivity in modern vehicle infrastructure, solutions are required to harden the vehicle’s electronic architecture against potential attacks. One of the most important concerns is to ensure vehicle systems take appropriate and safe actions to diminish the probability of cyber-attacks success and mitigate the ramifications of potential unauthorized access. In this work, we present a layered systematic approach to achieve these goals along with a real implementation on an electric motorcycle. The developed secure infrastructure involves secure interconnection mechanisms to ensure trustworthy communications for electronic control nodes; hardware firewall to prevent interference and unauthorized access from untrusted applications/firmware; separated OS instances for different execution environments to provide best possible support for apps with different requirements and to further isolate apps from each other; and finally secure deployment of apps/firmware to ensure that apps are deployed unaltered to the automotive platform. In particular, considering both in-vehicle’s units resource-constraints, in terms of cost and size, and security needs that emerge in open bus-based networks, we introduce a secure technology that helps to prevent cyber-attacks in automotive Controller Area Network (CAN) protocol which is often used in in-vehicle networks. This technology, which we call secure CAN (sCAN), respects standard CAN-bus and realizes security mechanisms implemented in software or hardware, while adding less than 1 ms latency on the communication. sCAN in synergy with on-chip hardware firewalling which controls authorization level of on-chip accesses, and finally, together with securing the deployment of firmware updating over the air, build a fully-layered approach for connected vehicles to protect system assets, such as application and device authenticity, integrity, confidentiality, and availability.

Background Streptococcus pyogenes or group A streptococcus (GAS) is an important human pathogen responsible for a broad range of infections, from uncomplicated to more severe and invasive diseases with high mortality and morbidity. Epidemiological surveillance has been crucial to detect changes in the geographical and temporal variation of the disease pattern; for this purpose the M protein gene (emm) gene typing is the most widely used genotyping method, with more than 200 emm types recognized. Molecular epidemiological data have been also used for the development of GAS M protein-based vaccines. Methods The aim of this paper was to provide an updated scenario of the most prevalent GAS emm types responsible for invasive infections in developed countries as Europe and North America (US and Canada), from 1st January 2000 to 31st May 2017. The search, performed in PubMed by the combined use of the terms (“emm”) and (“invasive”) retrieved 264 articles, of which 38 articles (31 from Europe and 7 from North America) met the inclusion criteria and were selected for this study. Additional five papers cited in the European articles but not retrieved by the search were included. Results emm1 represented the dominant type in both Europe and North America, replaced by other emm types in only few occasions. The seven major emm types identified (emm1, emm28, emm89, emm3, emm12, emm4, and emm6) accounted for approximately 50–70% of the total isolates; less common emm types accounted for the remaining 30–50% of the cases. Most of the common emm types are included in either one or both the 26-valent and 30-valent vaccines, though some well-represented emm types found in Europe are not. Conclusion This study provided a picture of the prevalent emm types among invasive GAS (iGAS) in Europe and North America since the year 2000 onward. Continuous surveillance on the emm-type distribution among iGAS infections is strongly encouraged also to determine the potential coverage of the developing multivalent vaccines.

Corynebacterium species are increasingly recognized as opportunistic pathogens. A growing number of taxonomic studies has yielded a description of numerous new Corynebacterium species, such as those related to the urogenital tract, with Corynebacterium glucuronolyticum found to be rarely involved in genitourinary tract infections, particularly in male individuals.In this report, we describe a urethritis case caused by C. glucuronolyticum in a 37-year-old, apparently healthy male, who complained mild pain in the lower abdomen, with several urinary symptoms. While urethral and semen specimens did not yield positive results for microbiological evaluation, cultures of urine samples revealed the monomicrobial growth on blood-containing media of tiny colonies after 24h of incubation, clearly evident only after 48h of incubation under CO2-enriched atmosphere. Colonies were identified as C. glucuronolyticum both by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) and 16S rRNA gene sequencing. Oral ciprofloxacin gradually led to clinical improvement and, finally, to a complete recovery, in accordance with microbiological findings. In spite of its infrequent detection, C. glucuronolyticum might be a potential urogenital pathogen in males more commonly that what believed, perhaps due to slow growth leading to underrecognition; we suggest therefore to consider the organism in the differential diagnostics of bacterial diseases of the urinary tract.

International audience; A newly developed MLVA seven-loci scheme for Streptococcus pyogenes is described. The method can be successfully applied by using both agarose gel with visual inspections of bands and Lab on Chip technology. The potential of the present MLVA has been tested on a collection of 100 clinical GAS strains representing the most common emm types found in high-income countries plus 18 published gap-free genomes, in comparison to PFGE and MLST. The MLVA analysis defined 30 MLVA types with ten out of the considered 15 emm types exhibiting multiple and specific MLVA types. In only one occasion the same MLVA profile was shared between isolates belonging to two different emm types. A robust congruency between the methods was observed, with MLVA discriminating within clonal complexes as defined by PFGE or MLST. This new MLVA scheme can be adopted as a quick, low-cost and reliable typing method to track the short-term diffusion of GAS clones in inter-laboratory-based surveillance.

This study aimed to retrospectively identify 22 Streptococcus bovis clinical strains based on the new taxonomy, as well as to investigate their antibiotic-resistance and clonality. Strains were identified by Phoenix100 system, 16S rRNA sequencing, and two MALDI-TOF MS platforms (Bruker Biotyper, Vitek MS). Antibiotic resistance was determined both phenotypically and genotypically, and clonality was assessed by PFGE. Most of strains (63.6%) were isolated from urine, and diabetes was the most common underlying disease (31.8%). Phoenix100 system revealed all strains belonged to biotype II, and 16S rRNA sequencing identified all strains as S. gallolyticus subsp pasteurianus (SGSP). Although both MALDI-TOF MS systems correctly identified isolates to the species level, only Bruker Biotyper accurately identified to the subspecies level. Erythromycin-resistant strains (31.8%) were also clindamycin-resistant and positive for erm (B). Strains resistant to tetracycline (68.2%) were also resistant to erythromycin. PFGE showed high genetic variability identifying 17 different pulsotypes, most of which single.