Recognition of environmental contaminant and pathogenic bacteria by means of redox potential methodology.

The time-consuming nature of culturing methods has urged the exploration of rapid modern technologies. One promising alternative utilizes redox potential, which describes the oxidative changes within complex media, indicating oxygen and nutrient consumption, as well as the production of reduced substances in the investigated biological system. Redox potential measurement can detect microbial activity within 16 h, what is significantly faster than the minimum 24 h incubation time of the reference plate counting technique. The redox potential based method can be specific with selective media, but bacterial strains have unique kinetic pattern as well. The proposed method suggests evaluation of the curve shape for the differentiation of environmental contaminant and pathogenic microbial strains. Six bacterial species were used in validation (Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Listeria innocua, Listeria monocytogenes, and Listeria ivanovii). Descriptive parameters reached 98.2 % accuracy and Gompertz model achieved 91.6 % accuracy in classification of the selected 6 bacteria species.•Mathematical model (Gompertz function) and first order descriptive parameters are suggested to describe the specific shape of redox potential curves, while Support Vector Machine (SVM) is recommended for classification.•Due to the concentration dependent time to detection (TTD), pre-processing applies standardization according to the inflection point time.

New bacterial strains isolated from Tunisian biotopes: A sustainable enzymatic approach for decolorization and detoxification of Congo Red and Malachite Green.

Seven bacterial strains, isolated from various Tunisian biotopes, were investigated for Congo Red (CR) and Malachite Green (MG) decolorization. The isolated strains underwent morphological and biochemical tests, including assessments for antibiotic sensitivity as well as biofilm formation. One selected strain, ST11, was partially identified as Paenibacillus sp. strain ST11. The newly isolated crude bacterial filtrates (NICBFs) effectively decolorized CR and MG. Specifically, six and seven NICBFs were found to be effective for degrading CR (150 mg l-1) and MG (50 mg l-1), respectively. Under non-optimized conditions, CR and MG could be decolorized up to 80% within 6-12 h. The degradation products of CR and MG, characterized by UV-visible and FT-IR techniques, demonstrated both decolorization and transformation, highlighting the role of enzymes in dye degradation. Phytotoxicity and cytotoxicity studies evaluated the impact of treated and untreated CR and MG. Some NICBFs showed promise as powerful biological tools, reducing and sometimes detoxifying CR and MG, commonly used as fertilizers. The potential applications of these NICBFs in decolorization and bioremediation of dye-rich textile effluents were explored. The screening also identified environmentally friendly, cost-effective bacterial strains adaptable to various conditions through phytotoxicity and cytotoxicity studies.

Response of Arugula to Integrated Use of Biological, Inorganic, and Organic Fertilization.

This study evaluated the effects of solely and integrated application of inorganic (NPK), commercial organic (NC), and biological (MIX, mixed strains Ensifer meliloti and Azotobacter chroococcum) fertilizers on the chemical characteristics of arugula biomass and its yield, as well as changes in soil microbiological parameters after the experiment in relation to the control treatment (Ø). The experiment was performed in semi-controlled greenhouse conditions, in pots, from the 4th decade of March to the 2nd decade of September, in 2023, at three cutting times/swaths, during one agricultural season, with Vertisol soil. For soil characterization, the following parameters were analysed: granulometric composition using sieving and sedimentation procedure; soil acidity-potentiometrically; SOM-soil organic matter by Kotzmann method; total N using CNS analyser; available P-spectrophotometrically; available K-flame photometrically; total number of microorganisms on an agarized soil extract medium; fungi on a solid Czapek agar; actinomycetes on a solid Krasiljnikov agar with saccharose; Azotobacter spp. on a liquid Fyodorov medium with mannitol; ammonifiers on a liquid medium with asparagine; and dehydrogenase activity-spectrophotometrically. For plant characterization, the following parameters were determined: N and C, both on CNS analyser; P on spectrophotometer; K on flame photometer; air-dried yield biomass. A stimulative effect on all microbiological parameters was found in the treatment with integrated use of organic and biological fertilizer, except for fungi, which grew better in the treatments with separate inorganic and organic fertilizers. Generally, the stimulative impact on plant chemical parameters manifested in combined inorganic and biological, organic and biological, and inorganic and organic fertilization treatments, and was inhibited in treatment without fertilization, in all three swaths, which could also be stated for the plant yield. Positive influence of all fertilization treatments on chemical parameters was observed for the second swath in relation to the first and the third. The total yield in the NPK+MIX treatment was 121%, and in the NC+MIX treatment, it was 87% higher compared to the control (Ø). In general, integrated use of inorganic and biological, organic and biological, and inorganic and organic fertilizers, respectively, could be proposed as an optimal fertilization treatment in arugula cultivation.

A Ranking Tool for "Category Killer" Microbial Biobanks.

Microbial biobanks preserve and provide microbial bioresources for research, training, and quality control purposes. They ensure the conservation of biodiversity, contribute to taxonomical research, and support scientific advancements. Microbial biobanks can cover a wide range of phylogenetic and metabolic diversity ("category killers") or focus on specific taxonomic, thematic, or disease areas. The strategic decisions about strain selection for certain applications or for the biobank culling necessitate a method to support prioritization and selection. Here, we propose an unbiased scoring approach based on objective parameters to assess, categorize, and assign priorities among samples in stock in a microbial biobank. We describe the concept of this ranking tool and its application to identify high-priority strains for whole genome sequencing with two main goals: (i) genomic characterization of quality control, reference, and type strains; (ii) genome mining for the discovery of natural products, bioactive and antimicrobial molecules, with focus on human diseases. The general concept of the tool can be useful to any biobank and for any ranking or culling needs.

Discovery of disease-adapted bacterial lineages in inflammatory bowel diseases.

Gut bacteria are implicated in inflammatory bowel disease (IBD), but the strains driving these associations are unknown. Large-scale studies of microbiome evolution could reveal the imprint of disease on gut bacteria, thus pinpointing the strains and genes that may underlie inflammation. Here, we use stool metagenomes of thousands of IBD patients and healthy controls to reconstruct 140,000 strain genotypes, revealing hundreds of lineages enriched in IBD. We demonstrate that these strains are ancient, taxonomically diverse, and ubiquitous in humans. Moreover, disease-associated strains outcompete their healthy counterparts during inflammation, implying long-term adaptation to disease. Strain genetic differences map onto known axes of inflammation, including oxidative stress, nutrient biosynthesis, and immune evasion. Lastly, the loss of health-associated strains of Eggerthella lenta was predictive of fecal calprotectin, a biomarker of disease severity. Our work identifies reservoirs of strain diversity that may impact inflammatory disease and can be extended to other microbiome-associated diseases.

Molecular Characterization of Escherichia coli Causing Urinary Tract Infections Through Next-Generation Sequencing: A Comprehensive Analysis of Serotypes, Sequence Types, and Antimicrobial and Virulence Genes.

Introduction An enormous increase in antimicrobial resistance (AMR) among bacteria isolated from human clinical specimens contributed to treatment failures. Increased surveillance through next-generation sequencing (NGS) or whole genome sequencing (WGS) could facilitate the study of the epidemiology of drug-resistant bacterial strains, resistance genes, and other virulence determinants they are potentially carrying. Methods This study included 30 Escherichia coli (E. coli) isolates obtained from patients suffering from urinary tract infections (UTIs) attending Prathima Institute of Medical Sciences, Karimnagar, India. All bacterial isolates were identified, and antimicrobial susceptibility patterns were determined through conventional microbiological techniques and confirmed by automated systems. All the isolates were investigated using NGS to identify genes coding for resistance, such as extended-spectrum beta-lactamases (ESBLs), metallo-beta-lactamases, and virulence genes. Multilocus sequence typing (MLST) was used to understand the prevalent strain types, and serotyping was carried out to evaluate the type of O (cell wall antigen) and H (flagellar antigen) serotypes carried by the isolates. Results The conventional antimicrobial susceptibility testing revealed that 15 (50%) isolates were resistant to imipenem (IPM), 10 (33.33%) were resistant to amikacin (AK), 13 (43.33%) were resistant to piperacillin-tazobactam (PTZ), 17 (56.66%) were resistant to cephalosporins, and 14 (46.66%) were resistant to nitrofurantoin (NIT). Among the isolates, 26 (86.66%) had revealed the presence of multiple antibiotic-resistant genes with evidence of at least one gene coding for beta-lactamase resistance. There was a high prevalence of blaCTX-M (19/30, 63.33%) genes, followed by blaTEM and blaOXA-1. The blaNDM-5 gene was found in three isolates (3/30, 10%). The virulence genes identified in the present study were iutA, sat, iss, and papC, among others. The E. coli serotype found predominantly belonged to O25:H4 (5, 16.66%), followed by O102:H6 (4, 13.33%). A total of 16 MLST variants were identified among the examined samples. Of the MLST-based sequence types (STs) identified, ST-131 (7, 23.33%) was the predominant one, followed by ST-167 (3, 10%) and ST-12 (3, 10%). Conclusions The study results demonstrated that the E. coli strains isolated from patients suffering from UTIs potentially carried antimicrobial resistance and virulence genes and belonged to different strain types based on MLST. Careful evaluation of bacterial strains using molecular analyses such as NGS could facilitate an improved understanding of bacterial antibiotic resistance and its virulence potential. This could enable physicians to choose appropriate antimicrobial agents and contribute to better patient management, thereby preventing the emergence and spread of drug-resistant bacteria.

Exploring the potential of earthworm gut bacteria for plastic degradation.

The use of plastic mulch films in agriculture leads to the inevitable accumulation of plastic debris in soils. Here, we explored the potential of earthworm gut-inhabiting bacterial strains (Mycobacterium vanbaalenii (MV), Rhodococcus jostii (RJ), Streptomyces fulvissimus (SF), Bacillus simplex (BS), and Sporosarcina globispora (SG) to degrade plastic films (⌀ = 15 mm) made from commonly used polymers: low-density polyethylene film (LDPE-f), polylactic acid (PLA-f), polybutylene adipate terephthalate film (PBAT-f), and a commercial biodegradable mulch film, Bionov-B® (composed of Mater-Bi, a feedstock with PBAT, PLA and other chemical compounds). A 180-day experiment was conducted at room temperature (x̄ =19.4 °C) for different strain-plastic combinations under a low carbon media (0.1× tryptic soy broth). Results showed that the tested strain-plastic combinations did not facilitate the degradation of LDPE-f (treated with RJ and SF), PBAT-f (treated with BS and SG), and Bionov-B (treated with BS, MV, and SG). However, incubating PLA-f with SF triggered a reduction in the molecular weights and an increase in crystallinity. Therefore, we used PLA-f as model plastic to study the influence of temperature ("room temperature" & "30 °C"), carbon source ("carbon-free" & "low carbon supply"), and strain interactions ("single strains" & "strain mixtures") on PLA degradation. SF and SF + RJ treatments significantly fostered PLA degradation under 30 °C in a low-carbon media. PLA-f did not show any degradation in carbon-free media treatments. The competition between different strains in the same system likely hindered the performance of PLA-degrading strains. A positive correlation between the final pH of culture media and PLA-f weight loss was observed, which might reflect the pH-dependent hydrolysis mechanism of PLA. Our results situate SF and its co-culture with RJ strains as possible accelerators of PLA degradation in temperatures below PLA glass transition temperature (Tg). Further studies are needed to test the bioremediation feasibility in soils.

An Insight into Fluorinated Imines and Hydrazones as Antibacterial Agents.

Fluorinated imines (Schiff bases) and fluorinated hydrazones are of particular interest in medicinal chemistry due to their potential usefulness in treating opportunistic strains of bacteria that are resistant to commonly used antibacterial agents. The present review paper is focused on these fluorinated molecules revealing strong, moderate or weak in vitro antibacterial activities, which have been reported in the scientific papers during the last fifteen years. Fluorinated building blocks and reaction conditions used for the synthesis of imines and hydrazones are mentioned. The structural modifications, which have an influence on the antibacterial activity in all the reported classes of fluorinated small molecules, are highlighted, focusing mainly on the importance of specific substitutions. Advanced research techniques and innovations for the synthesis, design and development of fluorinated imines and hydrazones are also summarized.

Evaluation of Osteoconductive and Antimicrobial Properties of Novel Graphene on Dental Implants: An In Vitro Study.

Introduction Graphene oxide (GO) has emerged as a promising material in dentistry, leveraging its exceptional properties. This study evaluates the physicochemical attributes of GO and elucidates its derived biological properties. These encompass biocompatibility, antibacterial efficacy, as well as its influence on osteogenic and odontogenic differentiation processes. Understanding the intricate interplay between the physicochemical and biological aspects of GO provides valuable insights into its potential applications in various dental contexts. Materials and methods The study group (so; titanium discs surface coated with GO) and the control group (co; plain/uncoated machined titanium discs) were divided based on cell attachment and cell proliferation assays (n=60). These groups were further divided into subgroups (n=30) based on the tested time intervals, specifically 24 hours, 48 hours, and 72 hours. The study and controlgroups were further subdivided into three subgroups (n=10) based on the microorganisms tested i.e Porphyromonas gingivalis, Prevotella intermedia and Fusobacteria nucleatum. Results The results of this in vitro study suggest that GO-coated titanium dental implants have both increased osteogenic potential and antimicrobial efficacy. Graphene has good potential as a promising alternative to traditional surface treatments, and a graphene-coated implant can be used for enhanced osseointegration.  Conclusion The osteogenic potential and the cell attachment were higher on titanium surfaces coated with GO nanoparticles when compared to plain titanium discs at 24, 48 and 72 hours respectively.

Green Synthesis of Silver Nanoparticles Using Drymaria cordata and Their Biocompatibility with Hemoglobin: A Therapeutic Potential Approach.

In this study, we present the synthesis and characterization of AgNPs using Drymaria cordata along with an assessment of their antioxidant, antibacterial, and antidiabetic activities. Antibacterial activities using four bacterial strains, free radical scavenging assays (DPPH and ABTS), and carbohydrate hydrolyzing enzyme inhibition assays were done to examine the therapeutic efficacy of AgNPs. Additionally, herein, we also evaluated the biocompatibility of the AgNPs using hemoglobin (Hb) as a model protein. A comprehensive analysis of Hb and AgNP interactions was carried out by using various spectroscopic, imaging, and size determination studies. Spectroscopic results showed that the secondary structure of Hb was not altered after its interaction with AgNPs. Furthermore, the thermal stability was also well maintained at different concentrations of nanoparticles. This study demonstrated a low-cost, quick, and eco-friendly method for developing AgNPs using D. cordata, and the biocompatible nature of AgNPs was also established. D. cordata-mediated AgNPs have potential applications against bacteria and diabetes and may be utilized for targeted drug delivery.

Biocontrol of almond canker diseases caused by Botryosphaeriaceae fungi.

BACKGROUND: Botryosphaeria dieback is a canker disease caused by fungal species of the Botryosphaeriaceae family that threatens almond productivity. The most common control measure to prevent canker development is the application of fungicides which are being phased out by European Union regulations. In the present study, two sets of bacterial strains were evaluated for their antifungal activity against pathogenic Botryosphaeriaceae species through in vitro and in vivo antagonism assays. RESULTS: The rhizospheric bacteria Pseudomonas aeruginosa AC17 and Bacillus velezensis ACH16, as well as the endophytic bacteria Bacillus mobilis Sol 1-2, respectively inhibited 87, 95, and 63% of the mycelial growth of Neofusicoccum parvum, Botryosphaeria dothidea, Diplodia seriata, and Macrophomina phaseolina. Additionally, they significantly reduced the length of lesions caused by N. parvum and B. dothidea in artificially inoculated detached almond twigs. All these bacterial strains produce hydrolytic enzymes that are able to degrade the fungal cell wall. P. aeruginosa AC17 also produces toxic volatile compounds, such as hydrogen cyanide. This strain was the most effective in controlling Botryosphaeria dieback in planta under controlled conditions at a level similar to the biocontrol agent Trichoderma atroviride and standard chemical fungicide treatments. CONCLUSION: Pseudomonas aeruginosa AC17 is the best candidate to be considered as a potential biocontrol agent against Botryosphaeriaceae fungi affecting almond. © 2023 Society of Chemical Industry.

Identification and specificity validation of unique and antimicrobial resistance genes to trace suspected pathogenic AMR bacteria and to monitor the development of AMR in non-AMR strains in the environment and clinical settings.

The detection of developing antimicrobial resistance (AMR) has become a global issue. The detection of developing antimicrobial resistance has become a global issue. The growing number of AMR bacteria poses a new threat to public health. Therefore, a less laborious and quick confirmatory test becomes important for further investigations into developing AMR in the environment and in clinical settings. This study aims to present a comprehensive analysis and validation of unique and antimicrobial-resistant strains from the WHO priority list of antimicrobial-resistant bacteria and previously reported AMR strains such as Acinetobacter baumannii, Aeromonas spp., Anaeromonas frigoriresistens, Anaeromonas gelatinfytica, Bacillus spp., Campylobacter jejuni subsp. jejuni, Enterococcus faecalis, Escherichia coli, Haemophilus influenzae, Helicobacter pylori, Klebsiella pneumonia subsp. pneumoniae, Pseudomonas aeruginosa, Salmonella enterica subsp. enterica serovar Typhimurium, Thermanaeromonas toyohensis, and Vibrio proteolyticus. Using in-house designed gene-specific primers, 18 different antibiotic resistance genes (algJ, alpB, AQU-1, CEPH-A3, ciaB, CMY-1-MOX-7, CMY-1-MOX-9, CMY-1/MOX, cphA2, cphA5, cphA7, ebpA, ECP_4655, fliC, OXA-51, RfbU, ThiU2, and tolB) from 46 strains were selected and validated. Hence, this study provides insight into the identification of strain-specific, unique antimicrobial resistance genes. Targeted amplification and verification using selected unique marker genes have been reported. Thus, the present detection and validation use a robust method for the entire experiment. Results also highlight the presence of another set of 18 antibiotic-resistant and unique genes (Aqu1, cphA2, cphA3, cphA5, cphA7, cmy1/mox7, cmy1/mox9, asaI, ascV, asoB, oxa-12, acr-2, pepA, uo65, pliI, dr0274, tapY2, and cpeT). Of these sets of genes, 15 were found to be suitable for the detection of pathogenic strains belonging to the genera Aeromonas, Pseudomonas, Helicobacter, Campylobacter, Enterococcus, Klebsiella, Acinetobacter, Salmonella, Haemophilus, and Bacillus. Thus, we have detected and verified sets of unique and antimicrobial resistance genes in bacteria on the WHO Priority List and from published reports on AMR bacteria. This study offers advantages for confirming antimicrobial resistance in all suspected AMR bacteria and monitoring the development of AMR in non-AMR bacteria, in the environment, and in clinical settings.

Isolation, screening, and identification of chitinase-producing bacterial strains from riverbank soils at Ambo, Western Ethiopia.

Chitinases are hydrolytic enzymes that dissolve the glycosidic linkages in chitin. Chitin is a cell wall component of fungi and fund in exoskeleten of worms and arthropods. Chitinase has been applied in agriculture, as a biopesticide for the control of plant fungal infections, in medicine, and in waste management. This research aimed to isolate, screen, and identification of chitinase-producing bacteria from riverbank soils. Twenty nine chitinolytic bacteria were isolated from the river bank soil samples, from which 9 of them had strong chitinolytic properties. Chitinase production was determined by zones of hydrolysis produced after 96 h of incubation at 37 °C. The different bacterial isolates were characterized morphologically, microscopically, and biochemically and finally eight strain were identified at species level by Matrix Assisted Laser Desorption Ionization - Time of Flight Mass Spectrometry (MALDI-TOF MS). From the eight, bacterial isolates investigated in this study Stenotrophomonas maltophilia showed the highest chitinase enzyme activity (625 µg/mL) followed by Pseudomonas putida with the enzyme activity of (553 µg/mL) and the least enzyme activity was recorded for Lilliottia amnigena (80 µg/mL). An incubation temperature of 45 °C, neutral pH and an incubation period of 96 h are found to be the optimum condition for the chitinase enzyme production from Stenotrophomonas maltophilia. The results of this study indicated the possibility of the production of chitinase from the chitinolytic bacterial isolates, which was highly useful for a variety of applications, including biocontrol of harmful insects and pathogenic fungi as well as in the biochemical, pharmaceutical, and medical sectors.

Microbial consortia degrade several widely used organic UV filters, but a number of hydrophobic filters remain recalcitrant to biodegradation.

Organic UV filters are important ingredients in many personal care products, including sunscreens. Evaluating the biodegradability of organic UV filters is key to estimate their recalcitrance and environmental fate and thus central to their overall environmental risk assessment. In order to further understand the degradation process, the aim was to investigate whether specific consortia could degrade certain UV filters. Several bacterial strains were isolated from enrichment cultures actively degrading octocrylene (OC), butyl methoxydibenzoylmethane (BM), homosalate (HS), and 2-ethylhexyl salicylate (ES) and were utilized to construct an in-house consortium. This synthetic consortium contained 27 bacterial strains and degraded OC, BM, HS, and ES 60-80% after 12 days, but not benzophenone-3 (BP3), methoxyphenyl triazine (BEMT), methylene bis-benzotriazolyl tetramethylbutylphenol (MBBT), diethylhexyl butamido triazone (DBT), ethylhexyl triazone (EHT), or diethylamino hydroxybenzoyl hexyl benzoate (DHHB). Furthermore, several commercial microbial mixtures from Greencell were tested to assess their degradation activity toward the same organic UV filters. ES and HS were degraded by some of the commercial consortia, but to a lesser extent. The rest of the tested UV filters were not degraded by any of the commercial bacterial mixes. These results confirm that some organic UV filters are recalcitrant to biodegradation, while others are degraded by a specific set of microorganisms.

Molecular Characterization and Antibiotic Susceptibility Pattern of Bacterial Strains Isolated From Wound of Patients With Diabetes.

BACKGROUND:  Diabetic wound infections are susceptible to various pathogens, particularly bacteria, due to the immunocompromised state of diabetic patients. Staphylococcus aureus is frequently implicated in diabetic wounds. To ascertain the presence of multiple antibiotic resistance in bacterial pathogens derived from diabetic wound infections, a comprehensive analysis is required. MATERIALS AND METHODS: The present cross-sectional investigation was carried out at a tertiary care facility. The samples were collected in aseptic conditions from the Endocrinology unit, specifically from local in-hospital patients (n=140). These samples were then assessed for their susceptibility to the commonly used antibacterial medications within the study area. The specimens were obtained from the lesions of individuals diagnosed with diabetes. The subjects were subjected to inoculation using various media and cultures. RESULTS: The findings of this study revealed that a collective sum of 122 bacterial isolates was acquired. The conclusions of the antibiotic susceptibility analysis revealed that the gram-positive isolates had a higher level of resistance to penicillin G (93.18%). However, they demonstrated sensitivity to vancomycin (100%) and linezolid (LZD) (95%). The gram-negative isolates exhibited complete resistance, at a rate of 100%, to penicillin, specifically amoxicillin (AMC), as well as to sulfonamides, such as sulfamethoxazole/trimethoprim (SXT), which belong to the antibiotic classes mentioned. CONCLUSION: In conclusion, there has been a notable rise in antibiotic resistance.

Analysis of antibacterial and cytotoxic potential of medicinal plants from Cholistan desert, Pakistan.

This study aimed to investigate the antibacterial and cytotoxic activity of 03 medicinal plants, Calligonum polygonides, Farsetia hamiltonii, and Pulcaria crispa, from Cholistan desert, Pakistan. The active constituents of plants species were extracted in 05 different solvents and the extracts were tested against various bacterial strains and brine shrimps. Although all Calligonum polygonides's extracts except chloroform were active against Staphylococcus aureus the most active was the acetone extract (21 ± 0.00 mm at 200 µg/disc) and activity was better than Caricef (p-value 0.03). While its water extract was more potent (18 ± 1.45 mm at 200 µg/disc) than Augmentin and Caricef (p-value < 0.005). The methanol extract's activity (15 ± 0.39 mm in 200 µg/disc) was comparable to Fucidin against Proteus vulgaris (p-value > 0.99) and activity of diethyl ether extract against Escherichia coli (10 ± 1.16 mm in 200 µg/disc) was same as of Urixin (p-value 0.91). Farsetia hamiltonii's acetone extract against Pseudomonas aeruginosa (10 ± 0.15 mm in 1 µg/disc) was more active than Augmentin Caricef and Cefotax (p-value < 0.02) and against Staphylococcus aureus (15 ± 1.15 mm in 200 µg/disc) activity was higher than Caricef (p-value 0.03). All Pulicaria crispa's extracts except water extract were found active against Staphylococcus aureus. However, the diethyl ether extract was most effective (25 + 0.00 mm at 150 µg /disc) and activity was more than Augmentin, Oxy-tetracycline, Fucidin, Urixin, Ceftriaxone (p-value < 0.05). Although all extracts were exhibited cytotoxic activity, the Calligonum polygonides's acetone extract (100%), Farsetia hamiltonii's diethyl ether extract (90%) and Pulicaria crispa's methanol extract (100%) were most active at 1000 µg/ml concentration. This study validated the medicinal significance of the studied plants and thus opens the way for their therapeutic applications.

Protocol for the High-quality Plasmid Isolation from Different Recalcitrant Bacterial Species: Agrobacterium spp., Rhizobium sp., and Bacillus thuringiensis.

High yield of good quality plasmid DNA from gram -ve bacteria (Agrobacterium tumefaciens, A. rhizogenes, and Rhizobium sp.) and gram +ve bacterium (Bacillus thuringiensis) is difficult. The widely used plasmid extraction kits for Escherichia coli yield a low quantity of poor-quality plasmid DNA from these species. We have optimized an in-house modification of the QIAprep Spin Miniprep kit protocol of Qiagen, consisting of two extraction steps. In the first, the centrifugation after adding neutralization buffer is followed by ethanol (absolute) precipitation of plasmid DNA. In the second extraction step, the precipitated DNA is dissolved in Tris-EDTA (TE) buffer, followed by an addition of 0.5 volumes of 5 M sodium chloride and 0.1 volumes of 20% (w/v) sodium dodecyl sulfate. After incubation at 65 °C for 15 min, the plasmid DNA is extracted with an equal volume of chloroform:isoamyl alcohol (CIA). RNase (20 mg/mL) is added to the upper phase retrieved after centrifugation and is incubated at 37 °C for 15 min. The extraction of the plasmid DNA with an equal volume of CIA is followed by centrifugation and is precipitated from the retrieved upper phase by adding an equal volume of absolute ethanol. The pellet obtained after centrifugation is washed twice with 70% (v/v) ethanol, air dried, dissolved in TE buffer, and quantified. This easy-to-perform protocol is free from phenol extraction, density gradient steps, and DNA binding columns, and yields high-quality plasmid DNA. The protocol opens an easy scale up to yield a large amount of high-quality plasmid DNA, useful for high-throughput downstream applications. Key features The protocol is free from density gradient steps and use of phenol. The protocol is an extension of the QIAprep Spin Miniprep kit (Qiagen) and is applicable for plasmid DNA isolation from difficult-to-extract bacterial species. The protocol facilitates the direct transformation of the ligation product into Agrobacterium by skipping the step of E. coli transformation. The plasmids isolated are of sequencing grade and the method is useful for extracting plasmids for metagenomic studies. Graphical overview Overview of the plasmid isolation protocol (modified QIAprep Spin Miniprep kit) of the present study.

The Plant Growth-Promoting Potential of Halotolerant Bacteria Is Not Phylogenetically Determined: Evidence from Two Bacillus megaterium Strains Isolated from Saline Soils Used to Grow Wheat.

(1) Background: Increasing salinity, further potentiated by climate change and soil degradation, will jeopardize food security even more. Therefore, there is an urgent need for sustainable agricultural practices capable of maintaining high crop yields despite adverse conditions. Here, we tested if wheat, a salt-sensitive crop, could be a good reservoir for halotolerant bacteria with plant growth-promoting (PGP) capabilities. (2) Methods: We used two agricultural soils from Algeria, which differ in salinity but are both used to grow wheat. Soil halotolerant bacterial strains were isolated and screened for 12 PGP traits related to phytohormone production, improved nitrogen and phosphorus availability, nutrient cycling, and plant defence. The four 'most promising' halotolerant PGPB strains were tested hydroponically on wheat by measuring their effect on germination, survival, and biomass along a salinity gradient. (3) Results: Two halotolerant bacterial strains with PGP traits were isolated from the non-saline soil and were identified as Bacillus subtilis and Pseudomonas fluorescens, and another two halotolerant bacterial strains with PGP traits were isolated from the saline soil and identified as B. megaterium. When grown under 250 mM of NaCl, only the inoculated wheat seedlings survived. The halotolerant bacterial strain that displayed all 12 PGP traits and promoted seed germination and plant growth the most was one of the B. megaterium strains isolated from the saline soil. Although they both belonged to the B. megaterium clade and displayed a remarkable halotolerance, the two bacterial strains isolated from the saline soil differed in two PGP traits and had different effects on plant performance, which clearly shows that PGP potential is not phylogenetically determined. (4) Conclusions: Our data highlight that salt-sensitive plants and non-saline soils can be reservoirs for halotolerant microbes with the potential to become effective and sustainable strategies to improve plant tolerance to salinity. However, these strains need to be tested under field conditions and with more crops before being considered biofertilizer candidates.

Self-assembled and Zn(II)-coordinated dipeptide nanoparticles with membrane-rupturing action on bacteria.

Metal ion-coordinated self-assembled short-chain amino acid peptide molecules with multi-photon excitation wavelengths and their photoluminescence properties are advantageous for fluorescence-based diagnostics and treatments of biological diseases based on their extra features of antibacterial agents. We have designed a novel strategy based on tryptophan molecule coordinated with Zn(II) ions in the form of biocompatible spherical nanoparticles of diameter 30-80 nm which have been used for antibacterial treatments against different kinds of pathogenic bacteria (Escherichia coli, Salmonella typhimurium, and Pseudomonas). Preferably, we have used tryptophan-phenylalanine (Trp-Phe), a dipeptide molecule having tryptophan as principal material against E. coli strains as antimicrobial agents for surface rupturing and killing purposes. Furthermore, based on single amino acid, tryptophan, self-assembled and Zn(II)-coordinated dipeptide nanoparticles (Zn-DPNPs) were studied against three types of multi-drug-resistant bacteria as an active antimicrobial agent. These antibacterial efficient nanoparticles may have best alternative of antibiotic drugs for clinical applications. The capability of self-assembled fluorescence behavior of Zn-coordinated dipeptide molecules and higher hydrophobicity against bacterial cell wall will perform as antimicrobial fluorescent agents. KEY POINTS: • Zn(II) and Cu(II) better coordinated into self-assembled NPs. • Fluorescence signals showed interaction of NPs with gram -ve cell wall. • Significant surface-damaging effects were observed in the case of Cu-DPNPs and Zn-DPNPs.

Prevalence of Multidrug-Resistant Pathogens Causing Neonatal Early and Late Onset Sepsis, a Retrospective Study from the Tertiary Referral Children's Hospital.

Introduction: Sepsis is the most severe infectious disease with the highest mortality rate, particularly among neonates admitted to the neonatal intensive care unit (NICU). This study retrospectively analyzed the epidemiology, antibiotic resistance profiles, and prevalence of multidrug-resistant (MDR) bacteria isolated from blood or cerebrospinal fluid (CSF) cultures in order to evaluate the appropriateness of initial empirical therapy for neonatal sepsis. Methods: A retrospective study was conducted in the NICU from January 1, 2015, to December 31, 2022. Microbiological data from patients admitted to the NICU were anonymously extracted from the Laboratory of Microbiology database. Neonatal sepsis was classified into two types: early-onset sepsis (EOS), which occurs within the first 72 hours of life, and late-onset sepsis (LOS) for those begins later. Results: A total of 679 bacterial strains, 543 from blood and 136 from CSF, were detected in 631 neonates. Among these, 378 isolates (55.67%) were Gram-positive bacteria, and 301 isolates (44.33%) were Gram-negative bacteria. The most frequently isolated pathogens were Coagulase-negative staphylococci (CoNS) (36.52%), followed by Klebsiella pneumoniae (20.47%) and Escherichia coli (13.84%). In EOS, 121 strains were found, CoNS represented the majority (33.88%), followed by Klebsiella pneumoniae (23.97%) and Escherichia coli (8.26%). Early-onset septicemia exhibited 67 (55.37%) MDR bacteria. In LOS, 558 strains were isolated, CoNS represented the majority of pathogens (37.10%), followed by Klebsiella pneumoniae (19.71%) and Escherichia coli (15.05%). Late-onset septicemia showed 332 (59.50%) MDR bacteria. High rates of MDR were found in CoNS (76.21%), carbapenem-resistant Klebsiella pneumoniae (66.91%), and MRSA (33.33%). Conclusion: The study revealed an alarming prevalence of MDR strains isolated from neonatal sepsis, emphasizing the necessity of finding effective prevention and treatment measures. Colistin can be used for MDR Gram-negative bacteria, while vancomycin and teicoplanin can be considered treatment therapies for staphylococcal infections.