Pseudomonas Research Articles

Isolation and Characterization of a Novel Jumbo Phage HPP-Temi Infecting Pseudomonas aeruginosa Pa9 and Increasing Host Sensitivity to Ciprofloxacin

Pseudomonas aeruginosa is a bacteria responsible for many hospital-acquired infections. Phages are promising alternatives for treating P. aeruginosa infections, which are often intrinsically resistant. The combination of phage and antibiotics in clearing bacterial infection holds promise due to increasing reports of enhanced effectiveness when both are used together. The aim of the study is to isolate and characterize a novel P. aeruginosa phage and determine its effectiveness in in vitro combination with antibiotics in controlling P. aeruginosa. In this study, a novel jumbo myophage HPP-Temi infecting P. aeruginosa Pa9 (PP334386) was isolated from household sewage. Electron micrographs of the phage were obtained to determine the morphological features of HPP-Temi virions. Complete genome analysis and a combination of Pseudomonas phage HPP-Temi with antibiotics were examined. The phage HPP-Temi was able to productively infect P. aeruginosa ATCC 9027 but was unable to infect a closely related genus. The phage was stable at 4–37 °C, 0.5% NaCl, and pH 8 for at least one hour. The HPP-Temi genome is a 302,719-bp-long dsDNA molecule with a GC content of 46.46%. The genome was predicted to have 436 ORFs and 7 tRNA genes. No virulence factor-related genes, antimicrobial resistance, or temperate lifestyle-associated genes were found in the phage HPP-Temi genome. Phage HPP-Temi is most closely related to the known or tentative representatives of the Pawinskivirus genus and can be proposed as a representative for the creation of a novel phage species in that genus. The phage and antibiotics (Ciprofloxacin) combination at varying phage titers (103, 106, 109) were used against P. aeruginosa Pa9 (PP334386) at 3.0 × 108 CFU/mL, which was carried out in triplicate. The result showed that combining antibiotics with phage significantly reduced the bacteria count at 103 and 106 titers, while no growth was observed at 109 PFU/mL. This suggests that the effect of phage HPP-Temi in combination with antibiotics is a potential and promising agent for the control of P. aeruginosa infections.

Occurrence of Multi-Drug-Resistant Bacteria in Rainbow Trout–Lettuce Bio-Integrated Culture Systems in Chile

In an aquaponic system, fish and plants are cultivated together in a symbiotic environment where they mutually benefit, using significantly less water than traditional farming methods. The main aim of this study was to investigate the occurrence of antimicrobial resistance in two aquaponic systems implemented in two Chilean high schools using rainbow trout and lettuce cultures. When water samples (fish tank, biofilter, and plant raft) were analyzed over a three-month period, no resistance to oxytetracycline was detected, whereas the occurrence of resistance to florfenicol was rather small, ranging from 0.01% to 3.1% of bacterial culturable counts. Eighteen isolates were recovered from various sources as representatives of the florfenicol-resistant population, and all of them belonged to the Pseudomonas genus, showing a multi-drug-resistance phenotype and exhibiting simultaneous resistance to 7–13 antimicrobials. All isolates exhibited resistance to amoxicillin, chloramphenicol, florfenicol, and furazolidone and susceptibility to meropenem, oxytetracycline, oxolinic acid, flumequine, ciprofloxacin, and enrofloxacin. Five and two isolates carried the amphenicol-resistance-encoding genes floR and cmlA, respectively, whereas no carriage of integrons or the fexA, fexB, pexA, optrA, and cfr genes encoding for florfenicol resistance was detected. Eleven isolates carried plasmids, but only two of them were able to transfer their plasmid content by conjugation. The knowledge of the microbiome associated with aquaponic systems is still scarce, and their role as potential reservoirs of antimicrobial-resistant bacteria and related genes of these systems must be elucidated.

Molecular identification and antimicrobial resistance patterns of enterobacterales in community urinary tract infections among indigenous women in Ecuador: addressing microbiological misidentification

BackgroundAntibiotic resistance of Enterobacterales poses a major challenge in the treatment of urinary tract infections (UTIs). In low- and middle-income countries (LMICs), standard microbiological (i.e. urine culture and simple disk diffusion test) methods are considered the “gold standard” for bacterial identification and drug susceptibility testing, while PCR and DNA sequencing are less commonly used. In this study, we aimed to re-identifying Enterobacterales as the primary bacterial agents responsible for urinary tract infections (UTIs) by comparing the sensitivity and specificity of traditional microbiological methods with advanced molecular techniques for the detection of uropathogens in indigenous women from Otavalo, Ecuador.MethodsA facility-based cross-sectional study was conducted from October 2021 to February 2022 among Kichwa-Otavalo women. Pathogens from urine samples were identified using culture and biochemical typing. Morphological identification was doble-checked through PCR and DNA sequencing of 16S, recA, and rpoB molecular barcodes. The isolates were subjected to antimicrobial susceptibility-testing using disk diffusion test.ResultsThis study highlighted a 32% misidentification rate between biochemical and molecular identification. Using traditional methods, E. coli was 26.19% underrepresented meanwhile Klebsiella oxytoca was overrepresented by 92.86%. Furthermore, the genera Pseudomonas, Proteus, and Serratia were confirmed to be E. coli and Klebsiella spp. by molecular method, and one Klebsiella spp. was reidentified as Enterobacter spp. The susceptibility profile showed that 59% of the isolates were multidrug resistant strains and 31% produced extended spectrum beta-lactamases (ESBLs). Co-trimoxazole was the least effective antibiotic with 61% of the isolates resistant. Compared to previous reports, resistance to nitrofurantoin and fosfomycin showed an increase in resistance by 25% and 15%, respectively.ConclusionsCommunity-acquired UTIs in indigenous women in Otavalo were primarily caused by E. coli and Klebsiella spp. Molecular identification (16S/rpoB/recA) revealed a high rate of misidentification by standard biochemical and microbiological techniques, which could lead to incorrect antibiotic prescriptions. UTI isolates in this population displayed higher levels of resistance to commonly used antibiotics compared with non-indigenous groups. Accurate identification of pathogens causing UTIs and their antibiotic susceptibility in local populations is important for local antibiotic prescribing guidelines.

Marine Pseudomonas: diving into the waves of blue biotechnology.

From marine to terrestrial environments, Pseudomonas spp. exhibit a remarkable ability not only to adapt but also thrive even amidst adverse conditions. This fact turns Pseudomonas spp. into one of the most prominent candidates for novel biotechnological solutions. Even though terrestrial isolates have been extensively studied, there is still an almost untapped source to be explored in marine Pseudomonas. Harnessing such strains offers an opportunity to discover novel bioactive compounds that could address current global challenges in healthcare and sustainable development. Therefore, this minireview aimed to provide an overview of the main recent discoveries regarding antimicrobials, antifouling, enzymes, pigments, and bioremediation strategies derived from marine isolates of Pseudomonas spp. Future research perspectives will also be discussed to foster forthcoming endeavors to explore the marine counterparts of such a prolific bacterial genus.

Evaluation of the Robustness Under Alkanol Stress and Adaptability of Members of the New Genus Halopseudomonas

Many species of the genus Pseudomonas are known to be highly tolerant to solvents and other environmental stressors. Based on phylogenomic and comparative genomic analyses, several Pseudomonas species were recently transferred to a new genus named Halopseudomonas. Because of their unique enzymatic machinery, these strains are being discussed as novel biocatalysts in biotechnology. In order to test their growth parameters and stress tolerance, five Halopseudomonas strains were assessed regarding their tolerance toward different n-alkanols (1-butanol, 1-hexanol, 1-octanol, 1-decanol), as well as to salt stress and elevated temperatures. The toxicity of the solvents was investigated by their effects on bacterial growth rates and presented as EC50 concentrations. Hereby, all Halopseudomonas strains showed EC50 values up to two-fold lower than those previously detected for Pseudomonas putida. In addition, the activity of the cis-trans isomerase of unsaturated fatty acids (Cti), which is an urgent stress response mechanism known to be present in all Pseudomonas species, was monitored in the five Halopseudomonas strains. Although several of the tested species were known to contain the cti gene, no significant phenotypic activity could be detected in the presence of the assayed stressors. A bioinformatic analysis of eight cti-carrying Halopseudomonas strains examining promotor binding sites, binding motifs and signal peptides showed that most of the cti genes have a lipoprotein signal peptide and promotor regions and binding motifs that do not coincide with those of Pseudomonas. These insights represent putative reasons for the absence of the expected Cti activity in Halopseudomonas, which in turn has always been observed in cti-carrying Pseudomonas. The lack of Cti activity under membrane stress conditions when the cti gene is present has never been documented, and this could represent potential negative implications on the utility of the genus Halopseudomonas for some biotechnological applications.

Identification of receptor-binding protein and host receptor of non-lytic dsRNA phage phiNY.

To date, complete genome sequences of 14 double-stranded RNA (dsRNA) phages are available, and studies have shown that the host range of dsRNA phages is limited. The hosts of most dsRNA phages belong to the genus Pseudomonas. However, the dsRNA phage phiNY, which has a non-lytic life cycle, was isolated from Microvirgula aerodenitrificans. Currently, the interaction between dsRNA phage phiNY and its host bacteria is unclear, which is not beneficial to a comprehensive understanding of dsRNA phage biology and the exploitation of dsRNA phage with non-lytic life cycle for biomedical applications and others. Phage adsorption is a crucial step through the interactions between receptor-binding protein (RBP) of the phage and its receptors to initiate the infection process, which dictates host range specificity. Thus, we identified the RBP and host receptor of phiNY. Through homology alignment, amino acid sequence similarity analysis, and the phylogenetic tree analysis, orf11, located in the M-segment of dsRNA phage phiNY, encodes a putative RBP. We further performed the whole-cell enzyme-linked immunosorbent assay (ELISA), western blotting assay, and indirect immunofluorescence assay and demonstrated that this orf11 is an RBP. Finally, using affinity chromatography, ELISA, and dynamic light scattering, we identified lipopolysaccharides (LPSs) on the surface of the host M. aerodenitrificans strain LH9 as host receptors involved in the adsorption of the dsRNA bacteriophage phiNY and observed the state of phiNY RBP after combining with LPS by atomic force microscopy. These results will guide future studies on phage-host interaction in a dsRNA phage with a non-lytic life cycle.IMPORTANCEThe interactions between the lytic dsRNA phages and their host receptors have been clarified in previous studies. However, the interaction between the dsRNA phage phiNY (which has a non-lytic life cycle) and its host receptors during the dsRNA phage adsorption process was unknown. Here, we found that phiNY uses the orf11 protein as a receptor-binding protein (RBP). In addition, we found that this orf11 recognizes lipopolysaccharide from the host bacterium Microvirgula aerodenitrificans strain LH9 as a specific receptor. These results suggest that phiNY, like lytic dsRNA phages, uses an RBP to bind to a similar host receptor (i.e., lipopolysaccharide). Determining the interaction between the dsRNA phage phiNY and its host receptors will help to elucidate the mechanisms underlying the phiNY non-lytic life cycle and enhance our understanding of its infection mechanism.

Development of bacterial bioformulations using response surface methodology.

Bacterial consortia exhibiting plant growth promoting properties have emerged as a sustainable approach for crop improvement. As the main challenge associated with them is viability loss and performance under natural conditions, a robust approach for designing bioformulation is needed. In this study, an efficient bioformulation was developed using spontaneous mutants of three bacterial strains for growth promotion of Cajanus cajan. Optimization of additives for solid [carboxymethylcellulose (CMC), and glycerol) and liquid [polysorbate, CMC, and polyvinyl pyrrolidone (PVP)] bioformulations was done by Response Surface Methodology using Central Composite Design. The stability of each bioinoculant in the formulation was assessed at 30°C and 4°C. The efficiency of the liquid bioformulation was checked in planta in sterile, and subsequently in non-sterile, soil. The maximum cell count was observed in solid bioformulation with 0.1g L-1 CMC and 50% glycerol (8.10×108, 3.69×108, and 7.39×108 for Priestia megaterium, Azotobacter chroococcum, and Pseudomonas sp. SK3, respectively) and in liquid bioformulation comprising 1% PVP, 0.1g L-1 CMC, and 0.025% polysorbate (8×109, 3.8×109, and 6.82×109 for Priestia megaterium, Azotobacter chroococcum, and Pseudomonas sp. SK3, respectively). The bioinoculants showed a higher viability (6 months) at 4°C compared to 30°C. Triple culture consortia enhanced plant growth in comparison to the control. The strains could be detected in soil till 45 days after sowing. The study established a systematic process for developing a potent bioformulation to promote agricultural sustainability. Using mutant strains, the bioinoculants could be tracked. In planta assays revealed that the triple culture consortium out-performed mono and dual cultures in terms of impact on plant growth.

Draft genome sequence of Pseudomonas sp. ZS001 isolated from peanut plantation soil.

Pseudomonas sp. ZS001 is a bacterium potentially degrading soluble starch isolated from peanut plantation soil sprayed with starch-based material in Dasi Town, Tianjin. Reporting the genome sequence of strain ZS001 can help us understand the genome composition, potential functional characteristics, and application value of Pseudomonas members at the genetic level.

Pseudomonas koreensis AB36 from a gold mine: genomic insights into adaptation to extreme conditions and potential plant growth promotion

AbstractThe genus Pseudomonas is one of the most varied and widespread bacterial genera, with species found in most environment. They are known to degrade organic and inorganic compounds, produce secondary metabolites, and enhance plant growth. The genome of Pseudomonas koreensis AB36, a heavy metal resistant organism isolated from a gold mine was sequenced to unveil the versatile metabolic potential of the organism. The genome is a single circular chromosome of 5,902,614 bp, with G + C content of 60.1%. There are 4154 similar orthologous gene clusters shared among strain AB36 and other sequenced P. koreensis strains with 7 clusters found alone in the genome of strain AB36. Genome mining of the organism predicted 8 biosynthetic gene clusters using antiSMASH including three non-ribosomal peptide synthethase (NRPS) clusters, arylpolyene and bacteriocin. The genome contains putative genes for heavy metal transport/resistance. These results show the heavy metal resistance ability and degradation of xenobiotic compounds of strain AB36 as well as its potential to adapt to various environments.

Effect of Bacillus subtilis – based microbiological preparations on the numerical composition of cultured forms of endophytic bacteria, content of mineral nutrition elements, photosynthetic pigments, as well as on chlorophyll fluorescence indices in leaves of columnar apple trees

The results of agrochemical field experiments conducted in an orchard of columnar apple tree (Malus domestica Borkh.) in the Moscow Oblast during the period of 2021–2022 are presented. The research objects were the ‘Valuta’, ‘Triumph’, and ‘President’ varieties. The effect of foliar treatments of plants with Bacillus subtilis (strain V167) and Bacillus subtilis (strain V417) strains, as well as Bacillus subtilis – based microbiological preparations “Extrasol” (strain Ch13) and “Phytosporin-M” (strain 26D) on such indicators as the microbiological and chemical composition of leaves, the content of photosynthetic pigments in leaves, and chlorophyll fluorescence indices was studied. Plant samples were analyzed using standard methodology. When applying strains and microbiological preparations based on Bacillus subtilis, the taxonomic composition of endophytic bacterial forms cultured on dense nutrient media and isolated from leaves was represented by the Bacillus genus. For comparison, when treating plants with Bacillus subtilis strain V417 and “Phytosporin-M”, the Pseudomonas genus was also detected. The number of cultivated forms of endophytic bacteria in the treated plants was consistently growing, having reached the maximum value of 406800 CFU/g in the “Extrasol” variant. Notably, the content of photosynthetic pigments in leaves depended on both the plant variety and the strain used. High values of maximal efficiency of PSII photochemistry (Fv/Fm) were observed both in experimental and control variants. A consistent increase in leaf N content was detected when applying “Extrasol”. In this respect, the ‘President’ variety showed the greatest response, i.e., a 16.1 % increase compared to the control. The ‘President’ and ‘Triumph’ varieties demonstrated an increase in leaf K content under the influence of foliar treatment of plants with the studied strains and microbiological preparations. The greatest effect of 17.2 % above the control was observed in the ‘Triumph’ variety when treated with “Extrasol”. Foliar treatments of ‘President’ and ‘Valuta’ plants with microbiological preparations contributed to increased, compared to the control, Mg contents in their leaves.

Metagenome-based microbial community analysis of urine-derived fertilizer.

Phosphorus is essential for food production and its supply is limited. Urine is an excellent source of phosphorus and one way to produce fertilizer is through conversion of urine to struvite (MgNH3PO4.6H2O). The present study aimed to understand the bacterial portion of the microbial community composition and dynamics of plasmid-mediated antimicrobial resistant genes during the optimized process of struvite production from composite human urine. Samples for DNA extraction was collected from fresh urine, stored urine and struvite during the process of struvite production. Shotgun metagenomic analysis was employed to understand the bacterial community. The most dominant phyla in the fresh and stored urine samples were Pseudomonadata, which comprised of 60% and 43% respectively, followed by Bacillota, comprised of 25% and 39% respectively. The struvite sample was dominated by the phylum Bacilliota (61%), Pseudomonadota (18%) and bacteroidota (12%). Members of the above phyla persisted in dominating each sample accordingly. Member of the family Morganellaceae was dominant in the fresh sample while the stored urine and struvite samples were dominated by the family Clostridiaceae. A decrease of members of the class Gammaproteobacteria was observed from the fresh to the struvite sample though not statistically significant. The genus Pseudomonas remained to be the most dominant member of Gammaproteobacteria in the fresh and stored urine sample with OTU count of 12,116 and 6,155 with a marked decrease by half in the stored sample. On the other hand, members of the genera Clostridium, Enterococcus, Bacteroides in the stored samples and Clostridium, Alkaliphilus and Pseudomonas in the struvite samples were dominant. 96% of the identified genera were shared in all the samples and the antimicrobial resistance genes (ARGs) identified in the fresh urine were shared by the struvite but not by the stored urine (e.g. sul, cat, aph and aac members). The presence of high abundance of ARGs in struvite needs attention in the persistence and transmissibility of the ARGs before application for agriculture.

Open and closed structures of L-arginine oxidase by cryo-electron microscopy and X-ray crystallography.

L-arginine oxidase (AROD, EC 1.4.3.25) is an oxidoreductase that catalyzes the deamination of L-arginine, with flavin adenine dinucleotide (FAD) as a cofactor. Recently identified AROD from Pseudomonas sp. TPU 7192 (PT-AROD) demonstrates high selectivity for L-arginine. This enzyme is useful for accurate assays of L-arginine in biological samples. The structural characteristics of the FAD-dependent AROD, however, remain unknown. Here, we report the structure of PT-AROD at a resolution of 2.3 Å by cryo-electron microscopy. PT-AROD adopts an octameric structure with D4 symmetry, which is consistent with its molecular weight in solution, estimated by mass photometry. Comparative analysis of this structure with that determined using X-ray crystallography reveals open and closed forms of the lid-like loop at the entrance to the substrate pocket. Furthermore, mutation of Glu493, located at the substrate binding site, diminishes substrate selectivity, suggesting that this residue contributes significantly to the high selectivity of PT-AROD.

Characterization of Plant Growth Promoting Rhizobacteria and Their Benefits on Soybean Growth

Background: Soybean is the most important legume in the world and its seeds contains 40% protein and 20% oil. In recent years, more attention has been paid to the use of plant growth-promoting (PGP) rhizobacteria as a biofertilizer alternative to chemical fertilizers, which may pose risks to the environment. The rhizobia inoculation of soybean is a sustainable practice to induce atmospheric nitrogen fixation and subsequently improve crop productivity and soil fertility. Methods: In this study, 19 indigenous rhizobia isolated from soybean were tested in vitro for their plant growth-promoting properties (PGPR), while genotypic characterization included sequencing of the 16S rRNA gene. Result: Morphological characterisation has shown that all of the isolates were rod-shaped gram negative bacteria. The sequencing has shown that 12 out of 19 isolates belong to the genus Pseudomonas, four of which belong to P. fluorescens species. Isolates which belong to the Pseudomonas genus have shown the highest ability of indole-3-acetic acid synthesis, phosphorous solubilization and along with isolates SGN6, SGN7 and SGS4 (Sphingomonas sanguinis) potassium solubilisation too. All the isolates from Pseudomonas genus as well as those belonging to B. japonicum species have shown the protease synthesis abilities while amylase synthesis abilities was observed only in the isolate SAK2 (P. chlororaphis). The most efficient strains in in vitro biological nitrogen fixation assay belong to Pseudomonas and Rhizobium genera. Most of the isolates were positive to the organic acid production while all of them have shown the ability of the exopolysaccharide production.

Distribution and characterization of endophytic and rhizosphere bacteriome of below-ground tissues in Chinese fir plantation.

None declared.Conflict of interestPlantation of Chinese fir, a popular woody tree species, faces sustainable issues such as nutrient deficiency and increasing disease threat. Rhizosphere and endophytic bacteria play important roles in plant' nutrient absorption and stress alleviation. Our understanding on the microbiome structure and functions are proceeding rapidly in model plants and some crop species. Yet, the spatial distribution and functional patterns of the bacteriome for the woody trees remain largely unexplored. In this study, we collected rhizosphere soil, non-rhizosphere soil, fine root, thick root and primary root samples of Chinese fir, and investigated the structure and distribution of bacteriome, as well as the beneficial effects of endophytic bacterial isolates. We discovered that Burkholderia and Paraburkholderia genera were overwhelmingly enriched in rhizosphere soil and the abundance of Pseudomonas genus was significantly enhanced in fine root. By isolating and testing the nutrient absorption and pathogen antagonism functions of representative endophytic bacteria species in Pseudomonas and Burkholderia, we noticed that P-solubilising functional isolates was enriched in fine root, while pathogen antagonism isolates was enriched in thick root. As a conclusion, our study revealed that the endophytic and rhizosphere environments of Chinese fir hold distinct structure and abundance of bacteriome, with potential specific functional enrichment of some bacterial clades. These findings assist us to further study the potential regulation mechanism of endophytic functional bacteria by the host tree, which will contribute to beneficial microbe application in forestry plantation and sustainable development.

Understanding the biosynthesis, metabolic regulation, and anti-phytopathogen activity of 3,7-dihydroxytropolone in Pseudomonas spp.

The genus Pseudomonas is a prolific source of specialized metabolites with significant biological activities, including siderophores, antibiotics, and plant hormones. These molecules play pivotal roles in environmental interactions, influencing pathogenicity, inhibiting microorganisms, responding to nutrient limitation and abiotic challenges, and regulating plant growth. These properties mean that pseudomonads are suitable candidates as biological control agents against plant pathogens. Multiple transposon-based screens have identified a Pseudomonas biosynthetic gene cluster (BGC) associated with potent antibacterial and antifungal activities, which produces 7-hydroxytropolone (7-HT). In this study, we show that this BGC also makes 3,7-dihydroxytropolone (3,7-dHT), which has strong antimicrobial activity toward Streptomyces scabies, a potato pathogen. Through metabolomics and reporter assays, we unveil the involvement of cluster-situated genes in generating phenylacetyl-coenzyme A, a key precursor for tropolone biosynthesis via the phenylacetic acid catabolon. The clustering of these phenylacetic acid genes within tropolone BGCs is unusual in other Gram-negative bacteria. Our findings support the interception of phenylacetic acid catabolism via an enoyl-CoA dehydratase encoded in the BGC, as well as highlighting an essential role for a conserved thioesterase in biosynthesis. Biochemical assays were used to show that this thioesterase functions after a dehydrogenation-epoxidation step catalyzed by a flavoprotein. We use this information to identify diverse uncharacterized BGCs that encode proteins with homology to flavoproteins and thioesterases involved in tropolone biosynthesis. This study provides insights into tropolone biosynthesis in Pseudomonas, laying the foundation for further investigations into the ecological role of tropolone production.IMPORTANCEPseudomonas bacteria produce various potent chemicals that influence interactions in nature, such as metal-binding molecules, antibiotics, or plant hormones. This ability to synthesize bioactive molecules means that Pseudomonas bacteria may be useful as biological control agents to protect plants from agricultural pathogens, as well as a source of antibiotic candidates. We have identified a plant-associated Pseudomonas strain that can produce 3,7-dihydroxytropolone, which has broad biological activity and can inhibit the growth of Streptomyces scabies, a bacterium that causes potato scab. Following the identification of this molecule, we used a combination of genetic, chemical, and biochemical experiments to identify key steps in the production of tropolones in Pseudomonas species. Understanding this biosynthetic process led to the discovery of an array of diverse pathways that we predict will produce new tropolone-like molecules. This work should also help us shed light on the natural function of antibiotics in nature.

Decomposition of Polythene by Bacteria Isolated from Waste Disposal Sites in Gusau City

The accumulation of polythene can be hazardous and can cause various environmental problems. This work was aimed at studying the biodegradation of polythene by bacteria isolated from waste disposal areas in North West, (Zamfara state) Nigeria. A total of twenty soil samples were collected from four different locations, such as waste disposal areas, hospitals, roadsides, and petrol pumps station. Spread plate method was carried out on Nutrient agar for the growth of bacteria. A total of 13 bacteria were isolated from different areas. The total bacteria count in hospital ranged from 80 × 10-4 - 248 × 10-4 cfu/mL, roadsides ranged from 116 × 10-4 - 208 × 10-4 cfu/mL, petrol pump ranged from 88 × 10-4 - 184 × 10-4 cfu/mL and waste disposal areas ranged from 116 × 10-4 - 264 × 10-4 cfu/mL. The microbial species associated with the polythene materials were identified as Pseudomonas sp, Bacillus sp, Staphylococcus aureus, Providencia stuartii, Clostridium perfringes, Corynebacterium diphtheria, Listeria monocytogen. The bacteria isolates were screened for polyethylene degradation using mineral salt medium containing polyethylene as sole source of carbon and nitrogen for their ability to degrade polyethylene for the period of 2 months. The efficacy of the microbes in the degradation of polythene were analyzed using liquid culture method, among the bacteria Pseudomonas aeruginosa degraded 75% of polythene, Bacillus subtilis 71.4% in a period of 2 months.

In-depth characterization of food and environmental microbiomes across different meat processing plants

BackgroundProcessing environments can be an important source of pathogenic and spoilage microorganisms that cross contaminate meat and meat products. The aim of this study was to characterize the microbiome of raw materials, processing environments and end products from 19 facilities producing different meat products.ResultsThe taxonomic profiles of the microbial communities evolved along processing, from raw materials to end products, suggesting that food contact (FC) surfaces play an important role in modulating the microbiome of final products. Some species persisted with the highest relative abundance in raw materials, food processing environments and/or in the final product, including species from the genera Pseudomonas, Staphylococcus, Brochothrix, Acinetobacter and Psychrobacter. Processing environments showed a very diverse core microbiota, partially shared with the products. Pseudomonas fragi and Pseudomonas sp. Lz4W (in all sample and facility types) and Brochothrix thermosphacta, Psychrobacter sp. and Psychrobacter sp. P11F6 (in raw materials, FC surfaces and end products) were prominent members of the core microbiota for all facilities, while Latilactobacillus sakei was found as a dominant species exclusively in end products from the facilities producing fermented sausages. Processing environments showed a higher amount of antimicrobial resistance genes and virulence factors than raw materials and end products. One thousand four hundred twenty-one medium/high-quality metagenome-assembled genomes (MAGs) were reconstructed. Of these, 274 high-quality MAGs (completeness > 90%) corresponded to 210 putative new species, mostly found in processing environments. For two relevant taxa in meat curing and fermentation processes (S. equorum and L. sakei, respectively), phylogenetic variation was observed associated with the specific processing facility under study, which suggests that specific strains of these taxa may be selected in different meat processing plants, likely contributing to the peculiar sensorial traits of the end products produced in them.ConclusionsOverall, our findings provide the most detailed metagenomics-based perspective up to now of the microbes that thrive in meat, meat products and associated environments and open avenues for future research activities to better understand the microbiome functionality and potential contribution to meat quality and safety.BW5-yJYgBWXLb4-yN7W47nVideo

Plant-Based Ingredients Utilized as Fat Replacers and Natural Antimicrobial Agents in Beef Burgers.

The present study aimed to find solutions based on the use of plant-based ingredients that would improve the nutritional quality of meat products as well as ensure sensory and microbiological quality. Two fat replacers, lemon albedo (Citrus lemon) and carob seed gum (Ceratonia siliqua), were investigated by chemical analysis and panel testing to evaluate their effect on the nutritional and sensory quality of beef burgers. The antimicrobial activity of two plant extracts, from nettle (Urtica dioica) leaves and medlar (Eriobotrya japonica) seeds, was studied, evaluating the intensity of inhibitory action and the minimum inhibitory concentration against Pseudomonas spp. and Listeria innocua strains by plate test. In addition, the antioxidant activity of both extracts was evaluated. Based on the results, lemon albedo and medlar seed extracts were validated in a food model (beef burger) by a storage test and a challenge test. The storage test results highlight that medlar seed extract prevents the formation of thiobarbituric acid reactive substances (TBARSs) and ensures microbiological quality, inhibiting Enterobacteriaceae and Pseudomonas spp. Anti-Listeria efficacy was confirmed in situ by challenge test results. In conclusion, although fat replacers ensure nutritional and sensory quality, they do not satisfy microbiological quality. This study clearly demonstrates that the safety of low-fat burgers can only be achieved through the combination of appropriate fat replacers with well-selected natural antimicrobial extracts.

Unraveling the Genomic Diversity of the Pseudomonas putida Group: Exploring Taxonomy, Core Pangenome, and Antibiotic Resistance Mechanisms.

The genus Pseudomonas is characterized by its rich genetic diversity, with over 300 species been validly recognized. This reflects significant progress made through sequencing and computational methods. Pseudomonas putida group comprises highly adaptable species that thrive in diverse environments and play various ecological roles, from promoting plant growth to being pathogenic in immunocompromised individuals. By leveraging the GRUMPS computational pipeline, we scrutinized 26363 genomes labeled as Pseudomonas in NCBI GenBank, categorizing all Pseudomonas spp. genomes into 435 distinct species-level clusters or cliques. We identified 224 strains deposited under the taxonomic identifier "Pseudomonas putida" distributed within 31 of these species-level clusters, challenging prior classifications. Nine of these 31 cliques contained at least six genomes labeled as "Pseudomonas putida" and were analyzed in depth, particularly clique_1 (P. alloputida) and clique_2 (P. putida). Pangenomic analysis of a set of 413 P. putida group strains revealed over 2.2 million proteins and more than 77000 distinct protein families. The core genome of these 413 strains includes 2226 protein families involved in essential biological processes. Intraspecific genetic homogeneity was observed within each clique, each possessing a distinct genomic identity. These cliques exhibit distinct core genes and diverse subgroups, reflecting adaptation to specific environments. Contrary to traditional views, nosocomial infections by P. alloputida, P. putida, and P. monteilii have been reported, with strains showing varied antibiotic resistance profiles due to diverse mechanisms. This review enhances the taxonomic understanding of key P. putida group species using advanced population genomics approaches and provides a comprehensive understanding of their genetic diversity, ecological roles, interactions, and potential applications.

New insights into uranium biomineralization mediated by Pseudomonas sp. WG2-6 in the presence of organic phosphorus: Promoting effect of extracellular polymeric substance and formation of U-P nanominerals

Microbial biomineralization significantly affects the uranium (U) behavior in the environment. However, the mechanism of microbial biomineralization of U is still not fully understood. In this study, a dominant bacterium (Pseudomonas sp. WG2–6) was isolated from U tail mining area. Abiotic precipitation tests demonstrated that U biomineralization was entirely attributed to the mediation of Pseudomonas sp. WG2–6 when the concentration ratio of exogenous β-glycerophosphate (SGP) to U was 10:1. Pseudomonas sp. WG2–6 exhibited strong immobilization ability towards U (97.59 %) according to batch experiments, and acylamide, carbonyls, and phosphate groups were the main functional groups that interacted with U. Besides, U mainly existed in the form of amorphous U-P complexes after biomineralization by Pseudomonas WG2–6, which could be converted into crystalline nano-minerals H2(UO2)2(PO4)2·8H2O in the presence of SGP. In particular, the formation and structural composition changes of extracellular polymeric substance (EPS) as well as the decrease in U4f binding energy were observed during the U biomineralization process of Pseudomonas sp. WG2–6 in the presence of SGP, indicating that EPS provided the nucleation site for the formation of stable biomineralized products. This work provides new insight into the mechanism of U microbial biomineralization and a theoretical basis for the remediation of U contaminated environments through microbial biomineralization.