Heba A. R. Abdelhaleem

Whole-genome sequencing (WGS) of microbial isolates is a valuable tool for mapping the genomes of novel organisms and is helpful for understanding plant-bacteria interactions. The close relationships between bacteria and plants are essential for maintaining healthy ecosystems, whether the bacteria reside the plant or in the rhizosphere surrounding its roots. In this study, isolation, characterization, and WGS were performed to identify promising plant growth-promoting rhizobacteria (PGPR) using the rhizospheric soil sample of jatropha tree roots. Out of 100 isolates, six (FACU 2024, 2, 3, 4, 5, and 6) exhibited phosphate-solubilizing bacteria (PSB) traits, including solubilizing phosphate and producing indole acetic acid (IAA), and the ability of other plant growth-promoting (PGP) traits was tested. Isolate FACU 2024 exhibited the highest values for IAA production (12.1 µg/ml), soluble phosphate release (300 µg/ml), and phosphate solubilization index (6.7). Therefore, FACU 2024 was molecularly identified as Escherichia coli. The WGS analysis revealed that E. coli FACU 2024 possesses one chromosome and one plasmid with a total length of 4.8 Mb and were submitted on GenBank under accession numbers CP147009 and CP147010. The bacterial genome contained about 142 PGP genes, ranging from 258 to 3744 bp and associated with phosphate solubilization, siderophore production, indole acetic acid (IAA) production, nitrogen metabolism, nitrogen fixation, and nitrite/nitrate reduction. Moreover, genomic islands (GIs) were enriched with genes associated with horizontal gene transfer (HGT), stress response, and environmental adaptation, and prophage analyses were carried out. In addition, 15 heavy metal resistance genes were annotated, such as those for As, Cd, Zn, Pb, Cu, Fe, and Co, ranging from 426 to 2505 bp. This study provides the first comprehensive genetic evidence linking E. coli to key PGPR traits alongside genes conferring resistance to multiple heavy metals. This strain demonstrates potential as a PGPR in addition to heavy metal bioremediation.

Industrial workplaces, particularly those involved in ore processing or smelting, pose a high risk of exposure to cadmium, a highly toxic metal. This study isolated and identified eight cadmium-resistant strains from industrial wastewater for their ability to resist cadmium. Enterobacter roggenkampii FACU2 demonstrated exceptional cadmium removal capabilities during our analysis, successfully eliminating 62% of the cadmium. Additionally, transmission electron microscopy (TEM) was utilized to examine the morphological change between the most and least efficient strains that responded to cadmium stress at the cellular level. Compared to the control bacteria, the treated bacteria exhibited notably higher levels of cadmium adsorption and accumulation within their cells. A complete genome analysis revealed that E. roggenkampii FACU2 has one chromosome and one plasmid with a size of 4,856,454 bp and 80,926 bp, respectively, in addition to harboring numerous heavy metal-resistant genes related to cadmium and other heavy metals. Moreover, the gene expression of four cadmium-resistant genes (czcA, cadA, czcC and czcD) showed that the high cadmium concentration led to a significant increase in czcA and cadA mRNA levels, thus indicating the activation of cadmium-resistant genes in the E. roggenkampii FACU2 compared to Enterobacter sp. strain FACU. Due to its ability to remove cadmium and other heavy metals, this strain holds promise as a source of genes for biological treatment methods. This application could contribute to environmental purification, ultimately benefiting human health.

Many organisms have adapted to survive in environments with high levels of arsenic (As), a naturally occurring metalloid with various oxidation states and a common element in human activities. These organisms employ diverse mechanisms to resist the harmful effects of arsenic compounds. Ten arsenic-resistant bacteria were isolated from contaminated wastewater in this study. The most efficient bacterial isolate able to resist 15,000 ppm Na₂HAsO₄·7H₂O was identified using the 16S rRNA gene and whole genome analysis as Enterobacter cloacae FACU. The arsenic E. cloacae FACU biosorption capability was analyzed. To further unravel the genetic determinants of As stress resistance, the whole genome sequence of E. cloacae FACU was performed. The FACU complete genome sequence consists of one chromosome (5.7 Mb) and two plasmids, pENCL 1 and pENCL 2 (755,058 and 1155666 bp, respectively). 7152 CDSs were identified in the E. cloacae FACU genome. The genome consists of 130 genes for tRNA and 21 for rRNAs. The average G + C content was found to be 54%. Sequencing analysis annotated 58 genes related to resistance to many heavy metals, including 16 genes involved in arsenic efflux transporter and arsenic reduction (five arsRDABC genes) and 42 genes related to lead, zinc, mercury, nickel, silver, copper, cadmium and chromium in FACU. Scanning electron microscopy (SEM) confirmed the difference between the morphological responses of the As-treated FACU compared to the control strain. The study highlights the genes involved in the mechanism of As stress resistance, metabolic pathways, and potential activity of E. cloacae FACU at the genetic level.

The phenylurea herbicides are persistent in soil and water, necessitating the creation of methods for removing them from the environment. This study aimed to examine the soil microbial diversity, searching for local bacterial isolates able to efficiently degrade the phenylurea herbicide isoproturon, 3-(4-isopropylphenyl)-1, 1-dimethylurea (IPU). The best isolates able to effectively degrade IPU were selected, characterized, and identified as Pseudomonas putida and Acinetobacter johnsonii. The catechol 1, 2-dioxygenase enzyme's catA gene was amplified, cloned, and expressed in E. coli M15. The Expressed E. coli showed high degradation efficiency (44.80%) as analyzed by HPLC after 15 days of inoculation in comparison to P. putida (21.60%). The expression of the catA gene in P. putida and expressed E. coli was measured using quantitative polymerase chain reaction (qPCR). The results displayed a significant increase in the mRNA levels of the catA gene by increasing the incubation time with IPU. Hydrophilic interaction chromatography (HILIC) mass spectrometry analysis revealed that three intermediate metabolites, 1-(4-isopropylphenyl)-3-methylurea (MDIPU), 4-Isopropylaniline (4-IA) and 1-(4-isopropylphenyl) urea (DDIPU) were generated by both P. putida and expressed E. coli. In addition, IPU-induced catA activity was detected in both P. putida and expressed E. coli. The supernatant of both P. putida and expressed E. coli had a significant influence on weed growth. The study clearly exhibited that P. putida and expressed E. coli were capable of metabolizing IPU influentially and thus could be utilized for bioremediation and biodegradation technology development.