Klebsiella pneumoniae is an opportunistic pathogen that frequently causes nosocomial and community-acquired (CA) infections. Until now, a limited number of studies has been focused on the analyses of changes affecting the virulence attributes. Genotypic and phenotypic methods were used to characterise the 39 clinical K. pneumoniae isolates; all belonged to the pan-drug resistant, widespread clone ST 15 and expressed the K24 capsule. PFGE has revealed that the isolates could be divided into three distinct genomic clusters. All isolates possessed allS and uge genes, known to contribute to the virulence of K. pneumoniae and 10.25% of the isolates showed hypermucoviscosity, 94.87% produced type 1 fimbriae, 92.3% produced type 3 fimbriae, and 92.3% were able to produce biofilm. In vivo persistence could be supported by serum resistance 46.15%, enterobactin (94.87%) and aerobactin (5.12%) production and invasion of the INT407 and T24 cell lines. Sequence analysis of the whole genomes of the four representative strains 11/3, 50/1, 53/2 and 53/3 has revealed high sequence homology to the reference K. pneumoniae strain HS11286. Our results represent the divergence of virulence attributes among the isolates derived from a common ancestor clone ST 15, in an evolutionary process that occurred both in the hospital and in the community. Full article

The development of new pharmaceutical solutions for treating various diseases results from a growing understanding of the benefits of using essential oils. One of the most often used volatile materials among essential oils is the oil of the citronella plant, termed citronella essential oil (CITEO), which has potential for use in food and medicine. Its wide use is limited due to lipophilicity, high volatility and poor physicochemical stability. With this background, the present study aims to evaluate the properties of CITEO-nanoemulsion (CITEO-NE) by analyzing its antimicrobial activities against Staphylococcus aureus (S. aureus) and Candida albicans (C. albicans) and its anticancer activity against, human skin adenocarcinoma cell line (A431). The CITEO-NE was prepared and evaluated for the size range of 130 ± 5 nm, polydispersity index (PDI) of 0.127 and zeta potential −12.6 mV. The percentage % of entrapment efficiency (%EE) of nanoemulsions loaded with CIT was very high at the beginning of the study, at 95.5 ± 4.775%. The MIC was observed to be 500 µg/mL for CITEO and 250 µg/mL for CITEO-NE against S. aureus and 250 µg/mL for CITEO and 125 µg/mL for CITEO-NE against C. albicans. The time-kill assay also suggests the effectiveness of CITEO-NE against the test pathogens as a novel alternative therapy. The IC₅₀ values of CITEO and CITEO-NE exhibited significant cytotoxic properties against the A431 cell line, with 41.20 μg/mL and 37.71 μg/mL, respectively. Hence, our findings revealed that encapsulation of CITEO increased the pharmacological properties. Full article

Medicinal plants are an essential source of traditional curatives for numerous skin diseases. Polyalthia longifolia (Sonn.) Thwaites (Annonaceae family) is a medicinal plant used to cure skin illnesses. P. longifolia is usually applied in folkloric therapeutical systems to treat skin diseases. The methicillin-resistant Staphylococcus aureus (MRSA) bacteria is among the essential bacteria contributing to skin diseases. Hence, to verify the traditional medicinal claim of P. longifolia usage in skin disease treatment, the current research was performed to study the synergistic antibacterial activity of standardized Polyalthia longifolia methanol leaf extract (MEPL) against MRSA bacteria. The synergistic antimicrobial activity result of ceftriaxone, when mixed with MEPL, against MRSA was investigated by the disc diffusion method, broth microdilution method, checkerboard dilution test, and modulation of mecA gene expression by multiplex polymerase chain reaction (multiplex PCR). The MEPL extract exhibited good synergistic antimicrobial activity against MRSA. Using the checkerboard method, we confirmed the synergistic effect of MEPL from P. longifolia and ceftriaxone (2:1) for MRSA with a marked reduction of the MIC value of the ceftriaxone from 8000 µg/mL to 1000 µg/mL. Moreover, the combination of MEPL with ceftriaxone significantly (p < 0.05) inhibited the presence of the resistant mecA gene in the tested strain. The LC–ESI–MS/MS analysis identified compounds that were reported to exhibit antimicrobial activity. Conclusively, the MEPL extract, an important etiological agent for skin diseases, showed worthy synergistic antimicrobial action against MRSA bacteria, thus supporting the traditional use of P. longifolia. Full article

Rare information is available on clinical Enterococcus faecium encountered in Sardinia, Italy. This study investigated the antimicrobial susceptibility profiles and genotypic characteristics of E. faecium isolated at the University Hospital of Sassari, Italy, using the Vitek2 system and PCR, MLST, or WGS. Vitek2 revealed two VanB-type vancomycin-resistant Enterococcus faecium (VREfm) isolates (MICs mg/L = 8 and ≥32) but failed to detect vancomycin resistance in one isolate (MIC mg/L ≤ 1) despite positive genotypic confirmation of vanB gene, which proved to be vancomycin resistant by additional phenotypic methods (MICs mg/L = 8). This vanB isolate was able to increase its vancomycin MIC after exposure to vancomycin, unlike the “classic” occult vanB-carrying E. faecium, becoming detectable by Vitek 2 (MICs mg/L ≥ 32). All three E. faecium had highly mutated vanB₂ operons, as part of a chromosomally integrated Tn1549 transposon, with common missense mutations in VanH and VanB₂ resistance proteins and specific missense mutations in the VanW accessory protein. There were additional missense mutations in VanS, VanH, and VanB proteins in the vanB₂-carrying VREfm isolates compared to Vitek2. The molecular typing revealed a polyclonal hospital-associated E. faecium population from Clade A1, and that vanB₂-VREfm, and nearly half of vancomycin-susceptible E. faecium (VSEfm) analyzed, belonged to ST117. Based on core genome-MLST, ST117 strains had different clonal types (CT), excluding nosocomial transmission of specific CT. Detecting vanB₂-carrying VREfm isolates by Vitek2 may be problematic, and alternative methods are needed to prevent therapeutic failure and spread. Full article

Critically ill patients suffering from severe infections are prone to pathophysiological pharmacokinetic changes that are frequently associated with inadequate antibiotic serum concentrations. Minimum inhibitory concentrations (MICs) of the causative pathogens tend to be higher in intensive care units. Both pharmacokinetic changes and high antibiotic resistance likely jeopardize the efficacy of treatment. The use of extracorporeal circulation devices to support hemodynamic, respiratory, or renal failure enables pharmacokinetic changes and makes it even more difficult to achieve an adequate antibiotic dose. Besides a clinical response, antibiotic pharmacokinetic optimization is important to reduce the selection of strains resistant to common antibiotics. In this review, we summarize the present knowledge regarding pharmacokinetic changes in critically ill patients and we discuss the effects of extra-corporeal devices on antibiotic treatment together with potential solutions. Full article

Antimicrobial resistance is a huge threat against the public health sphere and is a major cause of global mortality and morbidity. Antibiotic misuse and overuse have led to the development of many resistant bacterial strains. One particular bacterium of concern is methicillin-resistant Staphylococcus aureus (MRSA), which is the most common resistant bacteria in humans. Antibiotic development has been unable to keep up with the rapid evolution of antibiotic-resistant organisms, and there is an urgent need to identify alternative agents to combat this problem. The purpose of this systematic review is to explore the literature on the antibacterial properties of Eucalyptus globulus essential oil against MRSA. The articles used in this review were obtained through a systematic search of the literature across four databases, with the timeline being between 2002 and 2022. Twenty studies were included in this review, which used various methods to investigate the antibacterial properties of E. globulus essential oil, alone or in combination with other agents, against MRSA. The findings suggest that E. globulus essential oil has antibacterial properties against MRSA, which can be enhanced when used in combination with other agents, such as other essential oils and antibiotics. Full article

Rapid detection of extended-spectrum-β-lactamase (ESBL) is of paramount importance to accelerate clinical decision-making, optimize antibiotic treatment, and implement adequate infection control measures. This study was aimed at assessing the impact of direct detection of CTX-M ESBL-producers on antimicrobial management of Escherichia coli bloodstream infections over a 2-year period. This study included all E. coli bloodstream infection (BSI) events that were serially processed through a rapid workflow with communication to the clinicians of direct detection of CTX-M ESBL-producers and conventional culture-based workflow. Antimicrobial management was retrospectively analyzed to assess the contribution of the rapid test result. A total of 199 E. coli BSI events with a report of direct detection of CTX-M ESBL production results were included. Of these, 33.7% (n = 67) and 66.3% (n = 132) were reported as positive and negative CTX-M producers, respectively. Detection of CTX-M positive results induced more antibiotic therapy modifications (mainly towards carbapenem-containing regimens, p < 0.01), and antimicrobial susceptibility testing results of CTX-M ESBL-producing E. coli isolates induced more antibiotic escalations towards carbapenem-containing regimens (p < 0.01). Direct detection of CTX-M ESBL-producing E. coli resulted in a remarkable rate of antibiotic optimizations on the same day of blood culture processing. Observing antibiotic management following the availability of antimicrobial susceptibility testing results, additional early optimizations in escalation could probably have been made if the rapid test data had been used. Detection of CTX-M negative results resulted in few therapeutic changes, which could have probably been higher, integrating epidemiological and clinical data. Full article

Antimicrobial resistance is a major public health concern worldwide. Albeit to a lesser extent than bacteria, fungi are also becoming increasingly resistant to antifungal drugs. Moreover, due to the small number of antifungal classes, therapy options are limited, complicating the clinical management of mycoses. In this view, antimicrobial peptides (AMPs) are a potential alternative to conventional drugs. Among these, Proline-rich antimicrobial peptides (PrAMPs), almost exclusively of animal origins, are of particular interest due to their peculiar mode of action. In this study, a search for new arginine- and proline-rich peptides from plants has been carried out with a bioinformatic approach by sequence alignment and antimicrobial prediction tools. Two peptide candidates were tested against planktonic cells and biofilms of Candida albicans and Candida glabrata strains, including resistant isolates. These peptides showed similar potent activity, with half-maximal effective concentration values in the micromolar range. In addition, some structural and functional features, revealing peculiar mechanistic behaviors, were investigated. Full article

LF3872 was isolated from the milk of a healthy lactating and breastfeeding woman. Earlier, the genome of LF3872 was sequenced, and a gene encoding unique bacteriocin was discovered. We have shown here that the LF3872 strain produces a novel thermolabile class III bacteriolysin (BLF3872), exhibiting antimicrobial activity against antibiotic-resistant Staphylococcus aureus strains. Sequence analysis revealed the two-domain structural (lysozyme-like domain and peptidase M23 domain) organization of BLF3872. At least 25% residues of this protein are expected to be intrinsically disordered. Furthermore, BLF3872 is predicted to have a very high liquid-liquid phase separation. According to the electron microscopy data, the bacterial cells of LF3872 strain form co-aggregates with the S. aureus 8325-4 bacterial cells. LF3872 produced bacteriolysin BLF3872 that lyses the cells of the S. aureus 8325-4 mastitis-inducing strain. The sensitivity of the antibiotic-resistant S. aureus collection strains and freshly isolated antibiotic-resistant strains was tested using samples from women with lactation mastitis; the human nasopharynx and oral cavity; the oropharynx of pigs; and the cows with a diagnosis of clinical mastitis sensitive to the lytic action of the LF3872 strain producing BLF3872. The co-cultivation of LF3872 strain with various antibiotic-resistant S. aureus strains for 24 h reduced the level of living cells of these pathogens by six log. The LF3872 strain was found to be able to co-aggregate with all studied S. aureus strains. The cell-free culture supernatant of LF3872 (CSLF3872) induced S. aureus cell damage and ATP leakage. The effectiveness of the bacteriolytic action of LF3872 strain did not depend on the origin of the S. aureus strains. The results reported here are important for the creation of new effective drugs against antibiotic-resistant strains of S. aureus circulating in humans and animals. Full article

Antimicrobial resistance (AMR) is an important public health concern around the world. Limited information exists about AMR in grasslands-based systems where antibiotics are seldom used in beef cattle. The present study investigated the impacts of oxytetracycline (OTC) on the microbiome, antibiotic resistance genes (ARGs), and virulence factor genes (VFGs) in grazing steers with no previous exposure to antibiotic treatments. Four steers were injected with a single dose of OTC (TREAT), and four steers were kept as control (CONT). The effects of OTC on fecal microbiome, ARGs, and VFGs were assessed for 14 days using 16S rRNA sequencing and shotgun metagenomics. Alpha and beta microbiome diversities were significantly affected by OTC. Following treatment, less than 8% of bacterial genera had differential abundance between CONT and TREAT samples. Seven ARGs conferring resistance to tetracycline (tet32, tet40, tet44, tetO, tetQ, tetW, and tetW/N/W) increased their abundance in the post-TREAT samples compared to CONT samples. In addition, OTC use was associated with the enrichment of macrolide and lincosamide ARGs (mel and lnuC, respectively). The use of OTC had no significant effect on VFGs. In conclusion, OTC induced short-term alterations of the fecal microbiome and enrichment of ARGs in the feces of grazing beef cattle. Full article

No prospective evidence exists on the pharmacokinetic/pharmacodynamic (PK/PD) target attainment of ceftazidime in adult patients on general wards. We aimed to investigate whether the PK/PD target of ceftazidime (50% T > MIC) is attained in adult patients on general wards with adequate and impaired renal function receiving regular and guideline-recommended reduced doses of ceftazidime. In this observational, prospective, bicenter cohort study, adult patients admitted to a general ward receiving ceftazidime as part of standard care were included. Three blood samples per patient within 72 h after start of treatment were collected. Data were analyzed with nonlinear mixed effects modeling. The primary endpoint was target attainment of 50% T > MIC during the first 24 h of treatment (50% T_0–24 > MIC). Forty patients were included from whom 121 blood samples were obtained. All 25/25 patients with adequate renal function, 9/10 patients with moderately impaired renal function (eGFR 30–50 mL/min/1.73 m²) and 5/5 patients with severe impaired renal function (eGFR < 30 mL/min/1.73 m²) attained 50% T_0–24 > MIC when applying the clinical breakpoint MIC for Pseudomonas aeruginosa of 8 mg/L. The one patient not attaining the PK/PD target did not differ in any of the collected patients’ characteristics, except that this patient was the oldest in the study population. However, age was not statistically significantly associated with clearance or volume of distribution in the population pharmacokinetic model and, therefore, not likely the cause for this patient not attaining the PK/PD target. Our results suggest ≥90% probability of the PK/PD target attainment of ceftazidime in patients on general wards with adequate and impaired renal function receiving regular and guideline-recommended reduced doses of ceftazidime for treatment of infections with Pseudomonas aeruginosa and all bacteria with lower MIC-values. Full article

Periodontal disease is a relevant oral disease in dogs and nisin–biogel has been previously proposed to be used in its control. Enterococci, as inhabitants of the oral cavity with a high genetic versatility, are a reliable bacterial model for antimicrobial studies. Our goal was to evaluate the in vivo influence of the long-term dental application of the nisin–biogel on the virulence and antimicrobial signatures of canine oral enterococci. Twenty dogs were randomly allocated to one of two groups (treatment group—TG with nisin–biogel dental application, or control group—CG without treatment) and submitted to dental plaque sampling at day 0 and after 90 days (T90). Samples were processed for Enterococcus spp. isolation, quantification, identification, molecular typing and antimicrobial and virulence characterization. From a total of 140 enterococci, molecular typing allowed us to obtain 70 representative isolates, mostly identified as E. faecalis and E. faecium. No significant differences (p > 0.05) were observed in the virulence index of the isolates obtained from samples collected from the TG and CG at T90. At T90, a statistically significant difference (p = 0.0008) was observed in the antimicrobial resistance index between the isolates from the TC and CG. Oral enterococci were revealed to be reservoirs of high resistant and virulent phenotypes. Full article

Diabetic foot ulcers (DFU) are one of the most serious and devastating complications of diabetes and account for a significant decrease in quality of life and costly healthcare expenses worldwide. This condition affects around 15% of diabetic patients and is one of the leading causes of lower limb amputations. DFUs generally present poor clinical outcomes, mainly due to the impaired healing process and the elevated risk of microbial infections which leads to tissue damage. Nowadays, antimicrobial resistance poses a rising threat to global health, thus hampering DFU treatment and care. Faced with this reality, it is pivotal to find greener and less environmentally impactful alternatives for fighting these resistant microbes. Antimicrobial peptides are small molecules that play a crucial role in the innate immune system of the host and can be found in nature. Some of these molecules have shown broad-spectrum antimicrobial properties and wound-healing activity, making them good potential therapeutic compounds to treat DFUs. This review aims to describe antimicrobial peptides derived from green, eco-friendly processes that can be used as potential therapeutic compounds to treat DFUs, thereby granting a better quality of life to patients and their families while protecting our fundamental bio-resources. Full article

An antimicrobial consumption (AMC) study was performed in Trinidad and Tobago at the Eastern Regional Health Authority (ERHA). A retrospective, cross-sectional survey was conducted from 1 November 2021 to 30 March 2022. Dosage and package types of amoxicillin, azithromycin, co-amoxiclav, cefuroxime, ciprofloxacin, levofloxacin, moxifloxacin, nitrofurantoin and co-trimoxazole were investigated. Consumption was measured using the World Health Organization’s Antimicrobial Resistance and Consumption Surveillance System methodology version 1.0, as defined daily doses (DDD) per 1000 population per day (DID). They were also analyzed using the ‘Access’, ‘Watch’ and ‘Reserve’ classifications. In the ERHA, AMC ranged from 6.9 DID to 4.6 DID. With regards to intravenous formulations, the ‘Watch’ group displayed increased consumption, from 0.160 DID in 2017 to 0.238 DID in 2019, followed by a subsequent drop in consumption with the onset of the COVID-19 pandemic. Oral co-amoxiclav, oral cefuroxime, oral azithromycin and oral co-trimoxazole were the most highly consumed antibiotics. The hospital started off as the higher consumer of antibiotics, but this changed to the community. The consumption of ‘Watch’ group antibiotics increased from 2017 to 2021, with a drop in consumption of ‘Access’ antibiotics and at the onset of COVID-19. Consumption of oral azithromycin was higher in 2021 than 2020. Full article

Bacterial infection can occur in patients hospitalized with SARS-CoV-2 in various conditions, resulting in poorer outcomes, such as a higher death rate. This current systematic review was conducted in order to assess the efficiency of multiplex PCR in detecting bacterial infections in hospitalized COVID-19 patients, as well as to analyze the most common bacterial pathogens and other factors that interfere with this diagnosis. The research was conducted using four electronic databases (PubMed, Taylor&Francis, Web of Science, and Wiley Online Library). Out of 290 studies, nine were included in the systematic review. The results supported the use of multiplex PCR in detecting bacteria, considering its high sensitivity and specificity rates. The most common bacterial pathogens found were Klebsiella pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, and Haemophilus influenzae. The median age at admission was 61.5 years, and the majority of patients were men (70.3%), out of a total of 1553 patients. The proportion of ICU admission was very high, with a pooled proportion of 52.6% over the analyzed studies, and an average duration of hospitalization of 13 days. The mortality rate was proportionally high, as was the rate of ICU admission, with a pooled mortality of 24.9%. It was discovered that 65.2% of all patients used antibiotics before admission, with or without medical prescription. Antibiotic treatment should be considered consciously, considering the high risks of developing antibiotic resistance. Full article