We report on the genomic characterization of 47 multi-drug resistant, carbapenem resistant and ESBL-producing K. pneumoniae isolates from the influent (I) and effluent (E) of three wastewater treatment plants (WWTPs) and from Romanian hospital units which are discharging the wastewater in the sampled WWTPs. The K. pneumoniae whole genome sequences were analyzed for antibiotic resistance genes (ARGs), virulence genes and sequence types (STs) in order to compare their distribution in C, I and E samples. Both clinical and environmental samples harbored prevalent and widely distributed ESBL genes, i.e. bla_SHV, bla_OXA, bla_TEM and bla_{CTX M}. The most prevalent carbapenemase genes were bla_NDM-1, bla_OXA-48 and bla_KPC-2. They were found in all types of isolates, while bla_OXA-162, a rare bla_OXA-48 variant, was found exclusively in water samples. A higher diversity of carbapenemases genes was seen in wastewater isolates. The aminoglycoside modifying enzymes (AME) genes found in all types of samples were aac(6’), ant(2'')Ia, aph(3'), aaD, aac(3) and aph(6). Quinolone resistance gene qnrS1 and the multi-drug resistance oqxA/B pump gene were found in all samples, while qnrD and qnrB were associated to aquatic isolates. The antiseptics resistance gene qacEdelta1 was found in all samples, while qacE was detected exclusively in the clinical ones. Trimethroprim-sulfamethoxazole (dfrA, sul1 and sul2), tetracyclines (tetA and tetD) and fosfomycin (fosA6, known to be located on a transpozon) resistance genes were found in all samples, while for choramphenicol and macrolides some ARGs were detected in all samples (catA1 and catB3 / mphA), while other (catA2, cmIA5 and aac(6’)Ib / mphE and msrE) only in wastewater samples. The rifampin resistance genes arr2 and 3 (both carried by class I integrons) were detected only in water samples. The highly prevalent ARGs preferentially associating with aquatic versus clinical samples could ascribe potential markers for the aquatic (bla_SHV-145, qacEdelta1, sul1, aadA1, aadA2) and clinical (bla_OXA-1, bla_SHV-106,bla_TEM-150, aac(3)Iia, dfrA14, oqxA10; oqxB17,catB3, tetD) reservoirs of AR. Moreover, some ARGs (oqxA10; bla_SHV-145; bla_SHV-100, aac(6')Il, aph(3')VI, armA, arr2, cmlA5, bla_CMY-4, mphE, msrE, oqxB13, bla_OXA-10) showing decreased prevalence in influent versus effluent wastewater samples could be used as markers for the efficiency of the WWTPs in eliminating AR bacteria and ARGs. The highest number of virulence genes (75) was recorded for the I samples, while for E and C samples it was reduced to half. The most prevalent belong to three functional groups: adherence (fim genes), iron acquisition (ent, fep, fyu, irp and ybt genes) and the secretion system (omp genes). However, none of the genes associated with hypervirulent K. pneumoniae have been found. A total of 14 STs were identified. The most prevalent clones were ST101, ST219 in clinical samples and ST258, ST395 in aquatic isolates. These STs were also the most frequently associated with integrons. ST45 and ST485 were exclusively associated with I samples, ST11, ST35, ST364 with E and ST1564 with C samples. The less frequent ST17 and ST307 aquatic isolates harbored bla_OXA-162, which was co-expressed in our strains with bla_CTX-M-15 and bla_OXA-1.

The expression of bioactivity depends on the assimilation of different classes of natural substances (e.g., phenolic compounds) in vivo. Six functional extracts (Aspalathus linearis, leaves; Paullinia cupana, seeds; Aristotelia chilensis, berries; Ilex paraguariensis, leaves; Syzygium aromaticum, cloves, and wild berries) were analyzed in vitro and in vivo as an alternative to alleviating pathologies associated with oxidative stress (proliferation of cancer cells). The purpose of this research was to evaluate the in vitro and in vivo antioxidant and cytotoxic potential of hydroalcoholic solutions, in addition to the assimilation capacity of bioactive components in Saccharomyces boulardii cells. In vivo antioxidant capacity (critical point value) was correlated with the assimilation ratio of functional compounds. The results of in vitro antioxidant activities were correlated with the presence of quercetin (4.67 ± 0.27 mg/100 mL) and chlorogenic acid (14.38 ± 0.29 mg/100 mL) in I. paraguariensis. Bioassimilation of the main nutraceutical components depended on the individual sample. Phenolic acid levels revealed the poor assimilation of the main components, which could be associated with cell viability to oxidative stress.

The ability to orally administer silver nanoparticles (AgNPs) in enteric capsules implies a direct interaction with the colon microbiota. The in vitro effect provides a portrayal of the functional properties under in vivo conditions. The purpose of this study was to describe a green AgNP synthesis process, using aqueous extract from Lactarius piperatus mushroom, and to characterize the nanomaterial. We determined its antimicrobial and antioxidant effects in vitro in the microbiota of healthy individuals via the GIS1 system—a colon transit simulator. Per the quantitative polymerase chain reaction (qPCR) results, the antimicrobial properties of the AgNPs affected the initial share of different enteric species by decreasing the Bacteroides, Enterobacteriaceae, and Lactobacillus populations and favoring the Bifidobacterium group. The association between AgNPs and wild mushroom L. piperatus extract had a synergistic antibacterial activity against various pathogenic microorganisms while the mushroom extract reduced biofilm formation. Administration of AgNP maintained its constant antioxidant status, and it was correlated with a reduction in ammonium compounds. The physicochemical characterization of these NPs complemented their biochemical characterization. The maximum ultraviolet-visible spectroscopy (UV-VIS) absorbance was observed at 440 nm, while the Fourier transform infrared (FT-IR) spectrum reached a peak at 3296 cm^–1, which was correlated with the high-performance liquid chromatographic analysis (HPLC). The major phenolic compound was homogentisic acid. The size (49 ± 16 nm in diameter) and spherical shape of the NPs were correlated with their biological effects in vitro.

The multiple causes of cardiovascular diseases signify a major incidence and developmental risk of this pathology. One of the processes accountable for this pathologic development is the instauration of dysbiosis and its connection with an inflammatory process. Low antioxidant colonic protection encourages the progression of inflammation, with cardiovascular dysfunctions being a secondary consequence of the dysbiosis. Curcumin is one of the bioactive compounds displaying promising results for the reduction of an inflammatory process. The present study aims at demonstrating the capacity of three extracts drawn from Curcuma (C.) longa through an in vitro simulation process, for microbiota modulation in patients with hypertension. The acidic pH in the extraction process determined a high curcumin content in the extracts. The major phenolic compound identified was curcumin III, 622 ± 6.88 µg/mL for the ethanol/water/acetic acid extract. Low EC50 values were associated (0.2 µg/mL for DPPH scavenging activity) with the presence of curcumin isomers. A metabolic pattern became evident because the relationship between the short-chain fatty acids acted as a clinical biomarker. The curcumin present stimulated the formation of butyric and propionic acids. Microbiota activity control included a high degree of curcumin degradation and biotransformation in the other phenolic compounds. This developmental process was supported by the progression in the enterobacteria with a corresponding escalation in the pH level. The metabolomic pattern demonstrated a performance similar to the administration of dietary fibre, with the positive effects being dose-dependent.

Non-nutritive sweeteners represent an ingredient class that directly affects human health, via the development of inflammatory processes that promote chronic diseases related to microbiota dysbiosis. Several in vitro tests were conducted in the static GIS1 simulator. The aim of the study was to highlight the effect of sweeteners on the microbiota pattern of healthy individuals, associated with any alteration in the metabolomic response, through the production of organic acids and ammonium. The immediate effect of the in vitro treatment and the influence of the specific sweetener type on the occurrence of dysbiosis were evaluated by determining the biomarkers of the microbiota response. The presence of the steviol reduced the ammonium level (minimum of 410 mg/L), while the addition of cyclamate and saccharin caused a decrease in the number of microorganisms, in addition to lowering the total quantity of synthesized short-chain fatty acids (SCFAs). The bifidobacteria appeared to decrease below 10² genomes/mL in all the analyzed samples at the end of the in vitro simulation period. Barring the in vitro treatment of steviol, all the sweeteners tested exerted a negative influence on the fermentative profile, resulting in a decline in the fermentative processes, a rise in the colonic pH, and uniformity of the SCFA ratio.

Acinetobacter baumannii has emerged worldwide as a dominant pathogen in a broad range of severe infections, raising an acute need for efficient antibacterials. This is the first report on the resistome and virulome of 33 extended drug-resistant and carbapenem-resistant A. baumannii (XDR CRAB) strains isolated from hospitalized and ambulatory patients in Bucharest, Romania. A total of 33 isolates were collected and analyzed using phenotypic antibiotic susceptibility and conjugation assays, PCR, whole-genome sequencing (WGS), pulsed-field gel electrophoresis (PFGE) and MultiLocus Sequence Typing (MLST). All isolates were extensively drug-resistant (XDR), being susceptible only to colistin. The carbapenem resistance was attributed by PCR mainly to bla_OXA-24 and bla_OXA-23 genes. PFGE followed by MLST analysis demonstrated the presence of nine pulsotypes and six sequence types. WGS of seven XDR CRAB isolates from healthcare-associated infections demonstrated the high diversity of resistance genes repertoire, as well as of mobile genetic elements, carrying ARGs for aminoglycosides, sulphonamides and macrolides. Our data will facilitate the understanding of resistance, virulence and transmission features of XDR AB isolates from Romanian patients and might be able to contribute to the implementation of appropriate infection control measures and to develop new molecules with innovative mechanisms of action, able to fight effectively against these bugs, for limiting the spread and decreasing the infection rate and mortality.