Molecular binding of different classes of organophosphates to methyl parathion hydrolase from Ochrobactrum species.

Methyl parathion hydrolase (MPH) is an enzyme of the metallo-ß-lactamase superfamily, which hydrolyses a wide range of organophosphates (OPs). Recently, MPH has attracted attention as a promising enzymatic bioremediator. The crystal structure of MPH enzyme shows a dimeric form, with each subunit containing a binuclear metal ion center. MPH also demonstrates metal ion-dependent selectivity patterns. The origins of these patterns remain unclear but are linked to open questions about the more general role of metal ions in functional evolution and divergence within enzyme superfamilies. We aimed to investigate and compare the binding of different OP pesticides to MPH with cobalt(II) metal ions. In this study, MPH was modeled from Ochrobactrum sp. with different OP pesticides bound, including methyl paraoxon and dichlorvos and profenofos. The docked structures for each substrate optimized by DFT calculation were selected and subjected to atomistic molecular dynamics simulations for 500 ns. It was found that alpha metal ions did not coordinate with all the pesticides. Rather, the pesticides coordinated with less buried beta metal ions. It was also observed that the coordination of beta metal ions was perturbed to accommodate the pesticides. The binding free energy calculations and structure-based pharmacophore model revealed that all the three substrates could bind well at the active site. However, profenofos exhibit a stronger binding affinity to MPH in comparison to the other two substrates. Therefore, our findings provide molecular insight on the binding of different OP pesticides which could help us design the enzyme for OP pesticides degradation.

Harnessing nature's arsenal: Ochrobactrum bacteria metabolites in the battle against root- knot nematode - Insights from in vitro and molecular docking studies.

Agricultural Productivity and plant health are threatened by the root-knot nematode. The use of biocontrol agents reduces the need for chemical nematicides and improves the general health of agricultural ecosystems by offering a more environmentally friendly and sustainable method of managing nematode infestations. Plant-parasitic nematodes can be efficiently managed with the use of entomopathogenic nematodes (EPNs), which are widely used biocontrol agents. This study focused on the nematicidal activity of the secondary metabolites present in the bacteria Ochrobactrum sp. identified in the EPN, Heterorhabditisindica against Root-Knot Nematode (Meloidogyne incognita). Its effect on egg hatching and survival of juveniles of root- knot nematode (RKN) was examined. The ethyl acetate component of the cell-free culture (CFC) filtrate of the Ochrobactrum sp. bacteria was tested at four different concentrations (25 %, 50 %, 75 % and 100 %) along with broth and distilled water as control. The bioactive compounds of Ochrobactrum sp. bacteria showed the highest suppression of M. incognita egg hatching (100 %) and juvenile mortality (100 %) at 100 % concentration within 24 h of incubation. In this study, unique metabolite compounds were identified through the Gas Chromatography- Mass Spectrometry (GC-MS) analysis, which were found to have anti- nematicidal activity. In light of this, molecular docking studies were conducted to determine the impact of biomolecules from Ochrobactrum sp. using significant proteins of M. incognita, such as calreticulin, sterol carrier protein 2, flavin-containing monooxygenase, pectate lyase, candidate secreted effector, oesophageal gland cell secretory protein and venom allergen-like protein. The results also showed that the biomolecules from Ochrobactrum sp. had a significant inhibitory effect on the different protein targets of M. incognita. 3-Epimacronine and Heraclenin were found to inhibit most of the chosen target protein. Among the targets, the docking analysis revealed that Heraclenin exhibited the highest binding affinity of -8.6 Kcal/mol with the target flavin- containing monooxygenase. Further, the in vitro evaluation of 3- Epimacronine confirmed their nematicidal activity against M. incognita at different concentrations. In light of this, the present study has raised awareness of the unique biomolecules of the bacterial symbiont Ochrobactrum sp. isolated from H. indica that have nematicidal properties.

Structure, Physicochemical Properties and Biological Activity of Lipopolysaccharide from the Rhizospheric Bacterium Ochrobactrum quorumnocens T1Kr02, Containing d-Fucose Residues.

Lipopolysaccharides (LPSs) are major components of the outer membranes of Gram-negative bacteria. In this work, the structure of the O-polysaccharide of Ochrobactrum quorumnocens T1Kr02 was identified by nuclear magnetic resonance (NMR), and the physical-chemical properties and biological activity of LPS were also investigated. The NMR analysis showed that the O-polysaccharide has the following structure: →2)-ß-d-Fucf-(1→3)-ß-d-Fucp-(1→. The structure of the periplasmic glucan coextracted with LPS was established by NMR spectroscopy and chemical methods: →2)-ß-d-Glcp-(1→. Non-stoichiometric modifications were identified in both polysaccharides: 50% of d-fucofuranose residues at position 3 were O-acetylated, and 15% of d-Glcp residues at position 6 were linked with succinate. This is the first report of a polysaccharide containing both d-fucopyranose and d-fucofuranose residues. The fatty acid analysis of the LPS showed the prevalence of 3-hydroxytetradecanoic, hexadecenoic, octadecenoic, lactobacillic, and 27-hydroxyoctacosanoic acids. The dynamic light scattering demonstrated that LPS (in an aqueous solution) formed supramolecular particles with a size of 72.2 nm and a zeta-potential of -21.5 mV. The LPS solution (10 mkg/mL) promoted the growth of potato microplants under in vitro conditions. Thus, LPS of O. quorumnocens T1Kr02 can be recommended as a promoter for plants and as a source of biotechnological production of d-fucose.

Detoxification of ars genotypes by arsenite-oxidizing bacteria through arsenic biotransformation.

The detoxification process of transforming arsenite (As(III)) to arsenate (As(V)) through bacterial oxidation presents a potent approach for bioremediation of arsenic-polluted soils in abandoned mines. In this study, twelve indigenous arsenic-oxidizing bacteria (AOB) were isolated from arsenic-contaminated soils. Among these, Paenibacillus xylanexedens EBC-SK As2 (MF928871) and Ochrobactrum anthropi EBC-SK As11 (MF928880) were identified as the most effective arsenic-oxidizing isolates. Evaluations for bacterial arsenic resistance demonstrated that P. xylanexedens EBC-SK As2 (MF928871) could resist As(III) up to 40 mM, while O. anthropi EBC-SK As11 (MF928880) could resist As(III) up to 25 mM. From these bacterial strains, genotypes of arsenic resistance system (ars) were detected, encompassing ars leader genes (arsR and arsD), membrane genes (arsB and arsJ), and aox genes known to be crucial for arsenic detoxification. These ars genotypes in the isolated AOBs might play an instrumental role in arsenic-contaminated soils with potential to reduce arsenic contamination.

If You're Not Confused, You're Not Paying Attention: Ochrobactrum Is Not Brucella.

Bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a severe animal and human disease. Recently, a group of taxonomists merged the brucellae with the primarily free-living, phylogenetically related Ochrobactrum spp. in the genus Brucella. This change, founded only on global genomic analysis and the fortuitous isolation of some opportunistic Ochrobactrum spp. from medically compromised patients, has been automatically included in culture collections and databases. We argue that clinical and environmental microbiologists should not accept this nomenclature, and we advise against its use because (i) it was presented without in-depth phylogenetic analyses and did not consider alternative taxonomic solutions; (ii) it was launched without the input of experts in brucellosis or Ochrobactrum; (iii) it applies a non-consensus genus concept that disregards taxonomically relevant differences in structure, physiology, population structure, core-pangenome assemblies, genome structure, genomic traits, clinical features, treatment, prevention, diagnosis, genus description rules, and, above all, pathogenicity; and (iv) placing these two bacterial groups in the same genus creates risks for veterinarians, medical doctors, clinical laboratories, health authorities, and legislators who deal with brucellosis, a disease that is particularly relevant in low- and middle-income countries. Based on all this information, we urge microbiologists, bacterial collections, genomic databases, journals, and public health boards to keep the Brucella and Ochrobactrum genera separate to avoid further bewilderment and harm.

Efficient gene targeting in soybean using Ochrobactrum haywardense-mediated delivery of a marker-free donor template.

Gene targeting (GT) for precise gene insertion or swap into pre-defined genomic location has been a bottleneck for expedited soybean precision breeding. We report a robust selectable marker-free GT system in soybean, one of the most economically important crops. An efficient Oh H1-8 (Ochrobactrum haywardense H1-8)-mediated embryonic axis transformation method was used for the delivery of CRISPR-Cas9 components and donor template to regenerate T0 plants 6-8 weeks after transformation. This approach generated up to 3.4% targeted insertion of the donor sequence into the target locus in T0 plants, with ∼ 90% mutation rate observed at the genomic target site. The GT was demonstrated in two genomic sites using two different donor DNA templates without the need for a selectable marker within the template. High-resolution Southern-by-Sequencing analysis identified T1 plants with precise targeted insertion and without unintended plasmid DNA. Unlike previous low-frequency GT reports in soybean that involved particle bombardment-mediated delivery and extensive selection, the method described here is fast, efficient, reproducible, does not require a selectable marker within the donor DNA, and generates nonchimeric plants with heritable GT.

Ochrobactrum sp. NBRISH6 Inoculation Enhances Zea mays Productivity, Mitigating Soil Alkalinity and Plant Immune Response.

Intensifying sodic land characterized by high alkaline pH is an incipient environmental hazard-limiting agricultural potential. In this study, we investigated the effects of plant growth-promoting bacteria Ochrobactrum sp. strain NBRISH6 on the growth and physiology of maize (Zea mays L.) grown under alkaline stress at two soil pH levels. Additionally, we also studied the effects of NBRISH6 on soil fertility parameters. A greenhouse experiment was designed using two live soils (pH 8.2 and 10.2) in earthen pots using maize as a host. Results revealed a significant increase in plant growth and a decrease in defense enzymes in both soil types due to NBRISH6 inoculation as compared to non-treated control. Furthermore, activities of all soil enzymes along with bacterial diversity increased in NBRISH6 treatment under normal as well as stressed conditions. In addition, field evaluation of NBRISH6 inoculation using maize was carried out under normal and alkaline conditions, which resulted in significant enhancement of all vegetative parameters as compared to respective controls. Therefore, the study suggested that Ochrobactrum sp. NBRISH6 can be used to develop a bioinoculant formulation to ameliorate abiotic stresses and enhanced crop productivity.

Ochrobactrum chromiisoli sp. nov., Isolated from Chromium-Contaminated Soil.

A Gram-stain-negative, non-spore-forming, flagellated, motile, aerobic, rod-shaped bacteria strain, designated YY2XT, was isolated from chromium-contaminated soil. Phylogenetic analysis based on 16S rRNA gene, recA gene, and whole genome indicated that the strain represented a new member of the genus Ochrobactrum, family Brucellaceae, class Alphaproteobacteria. The phylogenetic trees based on 16 s rRNA gene, revealed that Falsochrobactrum ovis DSM26720T (96.7%), Ochrobactrum gallinifaecis DSM15295T (96.2%), and Pseudochrobactrum asaccharolyticum DSM25619T (96.2%) are the most closely related phylogenetic neighbors of strain YY2XT. The draft genome of YY2XT was approximately 4,650,646 bp in size with a G + C content of 53.0 mol%. Average nucleotide identity and digital DNA-DNA hybridization values among strain YY2XT and the selected Brucellaceae species were 71.4-83.1% and 13.5-42.7%, which are below the recommended cut-off values for species delineation. Growth of strain YY2XT occurred within pH 5-10 (optimum, pH 7-8), 4 â„ƒ-42 °C (optimum, 30 °C), and NaCl concentrations of 0.0-6.0% (optimum, 1.0%). Major quinone system was ubiquinone 10, the major fatty acids were C16:0, C18:1ω7c, and C16:1ω7c and the major polyamines were spermidine and putrescine. Major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylmonomethylethanolamine, phosphatidylethanolamine, and four undefined lipids. On the basis of the phenotypic, genotypic and chemotaxonomic traits, strain YY2XT was considered to represent a novel species of the genus Ochrobactrum, for which the name Ochrobactrum chromiisoli sp. nov. is proposed. The type strain is YY2XT (= CCTCC AB 2023035T = JCM 36000T).

Development of an efficient marker-free soybean transformation method using the novel bacterium Ochrobactrum haywardense H1.

We have discovered a novel bacterium, Ochrobactrum haywardense H1 (Oh H1), which is capable of efficient plant transformation. Ochrobactrum is a new host for Agrobacterium-derived vir and T-DNA-mediated transformation. Oh H1 is a unique, non-phytopathogenic species, categorized as a BSL-1 organism. We engineered Oh H1 with repurposed Agrobacterium virulence machinery and demonstrated Oh H1 can transform numerous dicot species and at least one monocot, sorghum. We generated a cysteine auxotrophic Oh H1-8 strain containing a binary vector system. Oh H1-8 produced transgenic soybean plants with an efficiency 1.6 times that of Agrobacterium strain AGL1 and 2.9 times that of LBA4404Thy-. Oh H1-8 successfully transformed several elite Corteva soybean varieties with T0 transformation frequency up to 35%. In addition to higher transformation efficiencies, Oh H1-8 generated high-quality, transgenic events with single-copy, plasmid backbone-free insertion at frequencies higher than AGL1. The SpcN selectable marker gene is excised using a heat shock-inducible excision system resulting in marker-free transgenic events. Approximately, 24.5% of the regenerated plants contained only a single copy of the transgene and contained no vector backbone. There were no statistically significant differences in yield comparing T3 null-segregant lines to wild-type controls. We have demonstrated that Oh H1-8, combined with spectinomycin selection, is an efficient, rapid, marker-free and yield-neutral transformation system for elite soybean.

Biodegradation characteristics and mechanism of quinoline by Ochrobactrum sp. strain C2.

A quinoline-degrading strain, C2, which could completely degrade 250 mg/L of quinoline within 24 h, was isolated from coking wastewater. Strain C2 was identified as Ochrobactrum sp. on the basis of 16S rDNA sequence analysis According to 16S rDNA gene sequence analysis, Strain C2 was identified as Ochrobactrum sp. Strain C2 could utilize quinoline as the sole carbon sources and nitrogen sources to grow and degrade quinoline well under acidic conditions. The optimum inoculum concentration, temperature and shaking speed for quinoline degradation were 10%, 30 °C and 150 r/min, respectively. The degradation of quinoline at low concentration by the strain followed the first-order kinetic model. The growth process of strain C2 was more consistent with the Haldane model than the Monod model, and the kinetic parameters were: Vmax = 0.08 h-1, Ks = 131.5 mg/L, Ki = 183.1 mg/L. Compared with suspended strains, strain C2 immobilized by sodium alginate had better degradation efficiency of quinoline and COD. The metabolic pathway of quinoline by Strain C2 was tentatively proposed, quinoline was firstly converted into 2(1H) quinolone, then the benzene ring was opened with the action of catechol 1,2-dioxygenase and subsequently transformed into benzaldehyde, 2-pentanone, hydroxyphenyl propionic acid and others.

Bioaugmentation of quinoline-degrading bacteria for coking wastewater treatment: performance and microbial community analysis.

Ochrobactrum sp. XKL1, previously found to have the ability to efficiently degrade quinoline, was bioaugmented into a lab-scale A/O/O system to treat real coking wastewater. During the bioaugmentation stage, the removal of quinoline and pyridine of the O1 tank could be enhanced by 9.88% and 7.96%, respectively. High-throughput sequencing analysis indicated that the addition of XKL1 could significantly affect the alteration of microbial community structure in the sludge. In addition, the relative abundance of Ochrobactrum has demonstrated a trend of increasing first followed by decreasing with the highest abundance of 7.87% attained on the 94th day. The bioaugmentation effects lasted for about 14 days after the strains was inoculated into the reactor. Although a decrease in the relative abundance of XKL1 was observed for a rather short period of time, the bioaugmented A/O/O system has been proven to be more effective in the removal of organic pollutants than the control. Hence, the results of this study indicated that the bioaugmentation with XKL1 is a feasible operational strategy that would be able to enhance the removal of NHCs in the treatment of coking wastewater with complex composition and high organic concentrations.

A comparative study on chemical characterization and properties of surface active compounds from Gram-positive Bacillus and Gram-negative Ochrobactrum strains utilizing pure hydrocarbons and waste mineral lubricating oils.

Mineral lubricating oils are widely used in various industrial sectors for their applications in maintenance and functioning of machineries. However, indiscriminate dumping of these used oils have resulted in polluting the natural reservoirs which subsequently destroys ecological balance. Bacteria can emulsify or lower surface tension between phases of immiscible substrates and can acquire them as their carbon and energy sources. Such a phenomenon is mediated by production of extracellular polymers which can function as eminent surface active compounds based on their surfactant or emulsifying nature. The comparison between bacterial strains (Gram-positive Bacillus stratosphericus A15 and Gram-negative Ochrobactrum pseudintermedium C1) on utilization of pure straight chain hydrocarbons, waste mineral lubricating oils as sole carbon source and chemical characterization of the synthesized surface active compounds were studied. Characterization analysis by Ultraviolet Visible spectrophotometry, Fourier transform infrared spectroscopy, Nuclear Magnetic Resonance spectroscopy, Carbon-Hydrogen-Nitrogen analysis has given detailed structural elucidation of surface active compounds. The contrasting nature of bacterial strains in utilization of different hydrocarbons of waste mineral lubricating oils was observed in Gas Chromatography-Mass Spectroscopy analysis. The variation between both strains in utilization of hydrocarbons can be manifested in chemical structural differences and properties of the produced surface active compounds. Scanning Electron Microscopy has given detailed insight into the microstructural difference of the compounds. The utilization of lubricating oils can address waste disposal problem and offer an economical feasible approach for bacterial production of surface active compounds. Our results suggest that these surface active compounds can maneuver applications in environmental bioremediation and agriculture, pharmaceuticals and food as functional biomaterials.

Remediation of arsenic-contaminated paddy field by a new iron oxidizing strain (Ochrobactrum sp.) and iron-modified biochar.

Iron-oxidizing strain (FeOB) and iron modified biochars have been shown arsenic (As) remediation ability in the environment. However, due to the complicated soil environment, few field experiment has been conducted. The study was conducted to investigate the potential of iron modified biochar (BC-FeOS) and biomineralization by a new found FeOB to remediate As-contaminated paddy field. Compared with the control, the As contents of GB (BC-FeOS), GF (FeOB), GFN (FeOB and nitrogen fertilizer), GBF (BC-FeOS and FeOB) and GBFN (BC-FeOS, FeOB and nitrogen fertilizer) treatments in pore water decreased by 36.53%-80.03% and the microbial richness of iron-oxidizing bacteria in these treatments increased in soils at the rice maturation stage. The concentrations of available As of GB, GF, GFN, GBF and GBFN at the tillering stage were significantly decreased by 10.78%-55.48%. The concentrations of nonspecifically absorbed and specifically absorbed As fractions of GB, GF, GFN, GBF and GBFN in soils were decreased and the amorphous and poorly crystalline hydrated Fe and Al oxide-bound fraction was increased. Moreover, the As contents of GB, GF, GFN, GBF and GBFN in rice grains were significantly decreased (*P < 0.05) and the total As contents of GFN, GBF and GBFN were lower than the standard limit of the National Standard for Food Safety (GB 2762-2017). Compared with the other treatments, GBFN showed the greatest potential for the effective remediation of As-contaminated paddy fields.

Prospective bioremediation of toxic heavy metals in water by surfactant exopolysaccharide of Ochrobactrum pseudintermedium using cost-effective substrate.

Globally, the underlying peril of cumulative toxicity of heavy metals in water bodies contaminated by industrial effluents is a matter of great concern to the environmentalists. Heavy metals like lead, cadmium, and nickel are particularly liable for this. Such toxic water is not only hazardous to human health but also harmful to aquatic animals. Remedial measures are being taken by physico-chemical techniques, but most of them are neither eco-friendly nor cost-effective. Biological means like bioaccumulation of heavy metals by viable bacteria are often tedious. In the present study, biosorption of heavy metals is successfully expedited by surfactant exopolysaccharide (SEPS) of Ochrobactrum pseudintermedium C1 as a simple, safe, and economically sustainable option utilizing an easily available and cost-effective substrate like molasses extract. Its efficacy in bioremediation of toxic heavy metals like cadmium, nickel, and lead have been studied by UV-Vis spectrophotometry and verified by inductively coupled plasma-atomic emission spectroscopy (ICP-AES). FTIR and zeta potential studies have also been carried out to explore this novel biosorption potential. Results are conclusive and promising. Moreover, this particular SEPS alone can remediate all these three toxic heavy metals in water. For futuristic applications, it might be a prospective and cost-effective resource for bioremediation of toxic heavy metals in aqueous environment.

Enzyme-catalyzed biodegradation of penicillin fermentation residues by ß-lactamase OtLac from Ochrobactrum tritici.

BACKGROUND: Biodegradation of antibiotics is a promising method for the large-scale removal of antibiotic residues in the environment. However, the enzyme that is involved in the biodegradation process is the key information to be revealed. RESULTS: In this study, the beta-lactamase from Ochrobactrum tritici that mediates the biodegradation of penicillin V was identified and characterized. When searching the proteins of Ochrobactrum tritici, the ß-lactamase (OtLac) was identified. OtLac consists of 347 amino acids, and predicted isoelectric point is 7.0. It is a class C ß-lactamase according to BLAST analysis. The coding gene of OtLac was amplified from the genomic DNA of Ochrobactrum tritici. The OtLac was overexpressed in E. coli BL21 (DE3) and purified with Ni2+ column affinity chromatography. The biodegradation ability of penicillin V by OtLac was identified in an in vitro study and analyzed by HPLC. The optimal temperature for OtLac is 32 â„ƒ and the optimal pH is 7.0. Steady-state kinetics showed that OtLac was highly active against penicillin V with a Km value of 17.86 µM and a kcat value of 25.28 s-1 respectively. CONCLUSIONS: OtLac demonstrated biodegradation activity towards penicillin V potassium, indicating that OtLac is expected to degrade penicillin V in the future.

The first case of Ochrobactrum intermedium bacteremia in a pediatric patient with malignant tumor.

BACKGROUND: Ochrobactrum spp. are non-fermenting, Gram-negative bacilli that are regarded as emerging human pathogens of low virulence that can cause infections. The first identified case of Ochrobactrum intermedium was reported in 1998 in a liver transplantation patient with liver abcess. There are no reports of infections in pediatric patients. Here, we report the first case of O. intermedium bacteremia in a pediatric patient. CASE PRESENTATION: A two and a half years old male was admitted with fever, chills and nausea. He had been diagnosed as pineoblastoma and underwent surgical resection and chemotherapy. O. intermedium was isolated from his blood cultures and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), however, the Vitek II automated system failed to identify the organism. Then the pathogen was confirmed by 16S rDNA sequencing and average nucleotide identity result (ANI) confirmed the precise identification of O. intermedium at genomic level. In addition, the patient recovered well after antibiotic combined therapy. CONCLUSIONS: This, to our knowledge, is the first case of O. intermedium bacteremia in a pediatric patient with malignant tumor. Traditional biochemical identification methods such as API 20NE or VITEK2 system cannot differentiate O. anthropi and O. intermedium. MALDI-TOF may be a promising tool for rapid identification of microorganisms such as O. intermedium.

Identification and Assessment of a Biocontrol Agent, Ochrobactrum intermedium I-5, for Management of Alfalfa Root Rot Caused by Fusarium tricinctum.

Alfalfa root rot caused by Fusarium tricinctum is one of the most important soilborne diseases, resulting in significant losses to alfalfa agriculture worldwide. Fungicides used in management of the disease affect the environment and human health. In this study, a strain of Ochrobactrum intermedium (I-5), isolated from alfalfa rhizosphere soil, exhibited strong antifungal activity against a number of causative pathogens of alfalfa root rot and showed the strongest antagonistic activity against F. tricinctum (a longest radius/shortest radius ratio of 3.09). When applied at 10%, a filtrate of the strain liquid culture significantly reduced the spore production and germination and mycelial growth of F. tricinctum, and the inhibition rates were 76.67, 78.93, and 55.77%, respectively. Furthermore, a filtrate and suspension of the strain, when applied at 10%, reduced alfalfa root rot by >73% in repeated experiments. The strain clearly promoted the activities of invertase, urease, cellulose, and neutral phosphatase in alfalfa rhizosphere soil and significantly reduced the damage to rhizosphere soil quality attributable to alfalfa root rot. Moreover, the strain clearly promoted the growth of alfalfa without causing any evident damage to plants. The active substance produced by the strain was insensitive to heat and ultraviolet irradiation and displayed optimal efficacy at pH 8. To the best of our knowledge, this is the first study describing the use of O. intermedium for the biological control of alfalfa root rot. O. intermedium (I-5) has potential for application in the control of alfalfa root rot and improvement of the quality of cultivated alfalfa.

Functional analysis of the ocnE gene involved in nicotine-degradation pathways in Ochrobactrum intermedium SCUEC4 and its enzymatic properties.

The SCUEC4 strain of Ochrobactrum intermedium is a newly isolated bacterium that degrades nicotine can use nicotine as the sole carbon source via a series of enzymatic catalytic processes. The mechanisms underlying nicotine degradation in this bacterium and the corresponding functional genes remain unclear. Here, we analyzed the function and biological properties of the ocnE gene involved in the nicotine-degradation pathways in strain SCUEC4. The ocnE gene was cloned by PCR with total DNA of strain SCUEC4 and used to construct the recombinant plasmid pET28a-ocnE. The overexpression of the OcnE protein was detected by SDS-PAGE analysis, and study of the function of this protein was spectrophotometrically carried out by monitoring the changes of 2,5-dihydroxypyridine. Moreover, the effects of temperature, pH, and metal ions on the biological activities of the OcnE protein were analyzed. The optimal conditions for the biological activities of OcnE, a protein of approximately 37.6 kDa, were determined to be 25 °C, pH 7.0, and 25 µmol/L Fe2+, and the suitable storage conditions for the OcnE protein were 0 °C and pH 7.0. In conclusion, the ocnE gene is responsible for the ability of 2,5-dihydroxypyridine dioxygenase. These findings will be beneficial in clarifying the mechanisms of nicotine degradation in O. intermedium SCUEC4.

Malathion biodegradation by a psychrotolerant bacteria Ochrobactrum sp. M1D and metabolic pathway analysis.

An organophosphorus pesticide malathion biodegradation was investigated by using the bacteria Ochrobactrum sp. M1D isolated from a soil sample of peach orchards in Palampur, District Kangra, Himachal Pradesh (India). The bacterium was able to utilize malathion as the sole source of carbon and energy. The isolated bacterium was found psychrotolerant and could degrade 100% of 100 mg l-1 malathion in minimal salt medium at 20°C, pH 7·0 within 12 days with no major significant metabolites left at the end of the study. Through GCMS analysis, methyl phosphate, diethyl maleate, and diethyl 2-mercaptosuccinate were detected and identified as the major pathway metabolites. Based on the GCMS profile, three probable degradation pathways were interpreted. The present study is the first report of malathion biodegradation at both the psychrophilic and mesophilic conditions by any psychrotolerant strain and also through multiple degradation pathways. In the future, the strain can be explored to bio-remediate the malathion contaminated soil in the cold climatic region and to utilize the enzymatic systems for advanced biotechnology applications.

Pan-genomics of Ochrobactrum species from clinical and environmental origins reveals distinct populations and possible links.

Ochrobactrum genus is comprised of soil-dwelling Gram-negative bacteria mainly reported for bioremediation of toxic compounds. Since last few years, mainly two species of this genus, O. intermedium and O. anthropi were documented for causing infections mostly in the immunocompromised patients. Despite such ubiquitous presence, study of adaptation in various niches is still lacking. Thus, to gain insights into the niche adaptation strategies, pan-genome analysis was carried out by comparing 67 genome sequences belonging to Ochrobactrum species. Pan-genome analysis revealed it is an open pan-genome indicative of the continuously evolving nature of the genus. The presence/absence of gene clusters also illustrated the unique presence of antibiotic efflux transporter genes and type IV secretion system genes in the clinical strains while the genes of solvent resistance and exporter pumps in the environmental strains. A phylogenomic investigation based on 75 core genes depicted better and robust phylogenetic resolution and topology than the 16S rRNA gene. To support the pan-genome analysis, individual genomes were also investigated for the mobile genetic elements (MGE), antibiotic resistance genes (ARG), metal resistance genes (MRG) and virulence factors (VF). The analysis revealed the presence of MGE, ARG, and MRG in all the strains which play an important role in the species evolution which is in agreement with the pan-genome analysis. The average nucleotide identity (ANI) based on the genetic relatedness between the Ochrobactrum species indicated a distinction between individual species. Interestingly, the ANI tool was able to classify the Ochrobactrum genomes to the species level which were assigned till the genus level on the NCBI database.