Ochrobactrum Intermedium Research Articles

Reduction of hexavalent chromium by Ochrobactrum intermedium BCR400 isolated from a chromium-contaminated soil

Hexavalent chromium-resistant Ochrobactrum intermedium BCR400 was isolated from chromium contaminated soil collected from Vadodara, Gujarat. It reduced 100 mg Cr(VI)/L completely in 52 h with initial Cr(VI) reduction rate of 1.98 mg/L/h. The Cr(VI) reduction rate decreased with increase in Cr(VI) concentration from 100 to 500 mg/L. The addition of anthraquinone-2-sulphonic acid (AQS) to culture O. intermedium BCR400 significantly enhanced its chromium reduction rate. The activation energy of AQS-mediated Cr(VI) reduction (120.69 KJ/mol) was 1.1-fold lower than non-mediated Cr(VI) reduction. An increase in the activities of quinone reductase and chromate reductase in cells grown in presence of AQS/AQS + Cr(VI) suggests their role in reduction of Cr(VI) by O. intermedium. Both chromate reductase and quinone reductase activities were FAD independent, required NADH as reductant, displayed maximum activity at pH (7.0) and temperature (30 °C). Thus Cr(VI) bioremediation potential of O. intermedium can be enhanced by augmentation of system with AQS as redox mediator.

Isolation and characterization of lead-tolerant Ochrobactrum intermedium and its role in enhancing lead accumulation by Eucalyptus camaldulensis

In this study, the potential of rhizospheric bacteria in promoting the growth and Pb accumulation by the woody plant Eucalyptus camaldulensis under hydroponic conditions was investigated for the first time. Three Pb-tolerant bacteria were isolated from the rhizosphere of E. camaldulensis grown in Pb-contaminated soils in the Bo Ngam Pb mine, Thailand. Based on analysis of partial 16S rRNA gene sequence, the three isolates were identified as Microbacterium paraoxydans BN-2, Ochrobactrum intermedium BN-3, and Bacillus fusiformis BN-4. Among these strains, O. intermedium BN-3 showed the highest tolerance to not only Pb but also Cd and Zn. After growth in the presence of Pb, the membranes of O. intermedium BN-3 cells exhibited an increase in unsaturated fatty acid levels but a decrease in fluidity. In hydroponic studies, inoculation of O. intermedium BN-3 significantly increased the biomass and Pb accumulation by E. camaldulensis compared to the uninoculated control. The results suggested the role of the natural rhizospheric bacteria localized to the root surface of E. camaldulensis in promoting Pb accumulation and plant growth. Our results indicate that O. intermedium BN-3 and other indigenous rhizospheric bacteria have the potential to improve the efficiency of phytoremediation of Pb-contaminated sites.

Application of a continuously stirred tank bioreactor (CSTR) for bioremediation of hydrocarbon-rich industrial wastewater effluents

A continuously stirred tank bioreactor (CSTR) was used to optimize feasible and reliable bioprocess system in order to treat hydrocarbon-rich industrial wastewaters. A successful bioremediation was developed by an efficient acclimatized microbial consortium. After an experimental period of 225 days, the process was shown to be highly efficient in decontaminating the wastewater. The performance of the bioaugmented reactor was demonstrated by the reduction of COD rates up to 95%. The residual total petroleum hydrocarbon (TPH) decreased from 320 mg TPH l −1 to 8 mg TPH l −1. Analysis using gas chromatography–mass spectrometry (GC–MS) identified 26 hydrocarbons. The use of the mixed cultures demonstrated high degradation performance for hydrocarbons range n-alkanes (C10–C35). Six microbial isolates from the CSTR were characterized and species identification was confirmed by sequencing the 16S rRNA genes. The partial 16S rRNA gene sequences demonstrated that 5 strains were closely related to Aeromonas punctata ( Aeromonas caviae), Bacillus cereus, Ochrobactrum intermedium, Stenotrophomonas maltophilia and Rhodococcus sp. The 6th isolate was affiliated to genera Achromobacter. Besides, the treated wastewater could be considered as non toxic according to the phytotoxicity test since the germination index of Lepidium sativum ranged between 57 and 95%. The treatment provided satisfactory results and presents a feasible technology for the treatment of hydrocarbon-rich wastewater from petrochemical industries and petroleum refineries.

SEASONAL EFFECTS OF DOMESTIC WASTEWATERS ON THE CR (VI) REDUCTION POTENTIAL OF <i>BACILLUS CEREUS</i> S-6 AND <i>OCHROBACTRUM INTERMEDIUM</i> CRT-1

Present study deals with the seasonal effects of domestic wastewater on the chromium reduction potential of strains Bacillus cereus S-6 and Ochrobactrum intermedium CrT-1. The presence of domestic wastewater from various localities caused a 20%—30% decrease in the Cr (VI) reduction potential of these bacteria. However, Cr (VI) reduction potential was 5%—20% higher in the presence of domestic wastewater collected in the summer season as compared to the winter season. The optimum temperature for reduction was observed at 37°C in case of B. cereus (96.9% Cr (VI) reduction) and 45°C in O. intermedium (96% Cr (VI) reduction). The pH preferences for optimum Cr (VI) reduction in these strains was pH 7 and pH 9 in case of B. cereus (93% Cr (VI) reduction) and O. intermedium (97% Cr (VI) reduction), respectively. The rate ofCr (VI) reduction was more in both strains when initially supplied with 100|ig ml-1 of K2Cr04as compared to 500|ag ml-1 and 1000(xg ml1. In the presence of three domestic wastewater samples, the rate of Cr(VI) reduction was slightly less where the effluent was used in sterilised form.

SEASONAL EFFECTS OF DOMESTIC WASTEWATERS ON THE CR (VI) REDUCTION POTENTIAL OF BACILLUS CEREUS S-6 AND OCHROBACTRUM INTERMEDIUM CRT-1

Present study deals with the seasonal effects of domestic wastewater on the chromium reduction potential of strains Bacillus cereus S-6 and Ochrobactrum intermedium CrT-1. The presence of domestic wastewater from various localities caused a 20%—30% decrease in the Cr (VI) reduction potential of these bacteria. However, Cr (VI) reduction potential was 5%—20% higher in the presence of domestic wastewater collected in the summer season as compared to the winter season. The optimum temperature for reduction was observed at 37°C in case of B. cereus (96.9% Cr (VI) reduction) and 45°C in O. intermedium (96% Cr (VI) reduction). The pH preferences for optimum Cr (VI) reduction in these strains was pH 7 and pH 9 in case of B. cereus (93% Cr (VI) reduction) and O. intermedium (97% Cr (VI) reduction), respectively. The rate of Cr (VI) reduction was more in both strains when initially supplied with 100μg ml -1 of K 2 CrO 4 as compared to 500μg ml -1 and 1000μg ml -1 . In the presence of three domestic wastewater samples, the rate of Cr(VI) reduction was slightly less where the effluent was used in sterilised form.

Ochrobactrum pecoris sp. nov., isolated from farm animals

Two Gram-negative, rod-shaped, non-spore-forming strains, designated 08RB2639(T) and 08RB2781-1, were isolated from a sheep (Ovis aries) and a domestic boar (Sus scrofa domestica), respectively. By 16S rRNA gene sequencing, the isolates revealed identical sequences and were shown to belong to the Alphaproteobacteria. They exhibited 97.8 % 16S rRNA gene sequence similarity with Ochrobactrum rhizosphaerae PR17(T), O. pituitosum CCUG 50899(T), O. tritici SCII24(T) and O. haematophilum CCUG 38531(T) and 97.4 % sequence similarity with O. cytisi ESC1(T), O. anthropi LMG 3331(T) and O. lupini LUP21(T). The recA gene sequences of the two isolates showed only minor differences (99.5 % recA sequence similarity), and strain 08RB2639(T) exhibited the highest recA sequence similarity with Ochrobactrum intermedium CCUG 24694(T) (91.3 %). The quinone system was ubiquinone Q-10, with minor amounts of Q-9 and Q-11, the major polyamines were spermidine, putrescine and sym-homospermidine and the major lipids were phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and phosphatidylcholine, with moderate amounts of the Ochrobactrum-specific unidentified aminolipid AL2. The major fatty acids (>20 %) were C₁₈:₁ω7c and C₁₉:₀ cyclo ω8c. These traits were in excellent agreement with the assignment of the isolates to the genus Ochrobactrum. DNA-DNA relatedness and physiological and biochemical tests allowed genotypic and phenotypic differentiation from other members of the genus Ochrobactrum. Hence, it is concluded that the isolates represent a novel species, for which the name Ochrobactrum pecoris sp. nov. is proposed (type strain 08RB2639(T) = DSM 23868(T) = CCUG 60088(T) = CCM 7822(T)).

Cicer arietinum growth promotion by Ochrobactrum intermedium and Bacillus cereus in the presence of CrCl3 and K2CrO4

The present study deals with the effect of two chromium-resistant bacteria Ochrobactrum intermedium CrT-1 and Bacillus cereus S-6 on chickpea (Cicer arietinum var NM-88) plants in the presence of two chromium salts CrCl3 and K2CrO4 (300 μg ml−1). Overall, the chromate salt severely affected various growth, yield and biochemical parameters of the chickpea plants. In general, bacterial inoculations enhanced various growth and biochemical parameters by alleviating the toxic effects of chromate salt. A reduction of 43% in seedling length was recorded when the plants were supplemented with 300 μg ml−1 of K2CrO4 in comparison to the chromium-free control. The toxicity of chromate salt was also obvious on the fresh biomass (40%), dry weight (116%), and auxin content (143%). Higher concentrations of chromium contents were detected in root (1.912 mg Cr g−1 dry weight) and smaller quantities were transferred into shoots (0.086 mg Cr g−1 dry weight) and leaves (0.074 mg Cr g−1 dry weight).

血清検査においてブルセラ病と診断された輸入牛から分離されたOchrobactrum intermedium の同定事例

輸入検疫では，牛の監視伝染病であるブルセラ病をこれまで多頭数摘発している． しかし，これらの摘発牛は血清反応検査に基づく診断によるもので，摘発牛からのブルセラ菌分離事例はない． 今般，動物検疫所で輸入検疫を実施したオーストラリア産肥育用素牛において，試験管凝集反応でブルセラ病患畜と診断された牛からBrucella 属菌と生化学的性状および抗原性が近似している菌を分離した． 分離菌はPCR検査でブルセラ属およびブルセラアボルタスと同一部位にバンドが確認されこれらと識別できなかった． しかし，遺伝子解析によりBrucella 属菌は否定され，Ochrobactrum intermedium と同定した．

Ochrobactrum ciceri sp. nov., isolated from nodules of Cicer arietinum

A Gram-staining-negative, aerobic, rod-shaped, non-spore-forming bacterial strain, Ca-34(T), was isolated from nodules of chickpea (Cicer arietinum) in Pakistan and studied for its taxonomic affiliation. The almost full-length 16S rRNA gene sequence showed highest similarities to those of strains of the genus Ochrobactrum. Based on results of MALDI-TOF MS and 16S rRNA gene sequence similarity (98.6 %), strain Ca-34(T) and Ochrobactrum intermedium LMG 3301(T) are phylogenetic neighbours; the two strains shared DNA-DNA relatedness of 64 %. The fatty acid profile [predominantly C(18 : 1)omega7c (67.7 %) and C(19 : 0) cyclo omega8c (19.6 %)] also supported the genus affiliation. Metabolically, strain Ca-34(T) differed from other type strains of Ochrobactrum in many reactions and from all type strains in testing positive for gelatin hydrolysis and in testing negative for assimilation of alaninamide and l-threonine. Based on phenotypic and genotypic data, we conclude that strain Ca-34(T) represents a novel species, for which we propose the name Ochrobactrum ciceri sp. nov. (type strain Ca-34(T) =DSM 22292(T) =CCUG 57879(T)).

Effects of a reputed immunostimulant on the innate immune system of goat kids

Forty newborn Majorera goats were randomly assigned to one of two groups. On the first day of life, the kids were fed atomized colostrum. The kids were subsequently fed a milk replacer (16% w/v) twice a day. The experimental group received two intramuscular doses of a reputed immunostimulant composed of a Propionibacterium acnes extract, Ochrobactrum intermedium lipopolysaccharides, and Proclin ® at 10 and 40 days of age, whereas, the control group received two doses of saline solution. Blood samples were collected weekly from kids age 10 to 66 days postpartum to measure the total and differential white blood cell counts, the plasma IgG, IgA, and IgM levels, and the activities of the total and the alternative complement systems. The white blood cell and the differential cell counts were the same for both groups. No statistical differences were found for the plasma IgG, IgA, and IgM concentrations. Complement system activity was undetectable in both groups. Summarizing, the reputed immunostimulant did not affect the immune system of goats. Thus, veterinarians should carefully consider whether it is prudent to recommended this kind of product, because while it increases farmers’ expenses it apparently does not improve the state of the innate immune system of the neonatal goat.

Phylogenetic profiling of bacterial community from two intimately located sites in Balramgari, North-East coast of India

Microbial communities in coastal subsurface sediments play an important role in biogeochemical cycles. In this study microbial communities in tidal subsurface sediments of Balramgari in the state of Orissa, India were investigated using a culture independent approach. Two 16S rDNA cloned libraries were prepared from the closely located (100 m along the coast) subsurface sediment samples. Library I sediment samples had higher organic carbon content but lower sand percentage in comparison to Library II. A total of 310 clone sequences were used for DOTUR analysis which revealed 51 unique phylotypes or operational taxonomic units (OTUs) for both libraries. The OTUs were affiliated with 13 major lineages of domain bacteria including Proteobacteria (α, β, δ and λ), Acidobacteria, Actinobacteria, Cyanobacteria, Chloroflexi, Firmicutes, Verrucomicrobia, Bacteroidetes, Gemmatimonadetes and TM7. We encountered few pathogenic bacteria such as Aeromonas hydrophila and Ochrobactrum intermedium, in sediment from Library I. ∫-LIBSHUFF comparison depicts that the two libraries were significantly different communities. Most of the OTUs from both libraries possessed ≥85% to <97% similarity to RDP database sequences depicting the putative presence of new species, genera and phylum. This work revealed the complex and unique bacterial diversity from coastal habitat of Balramgari and shows that, in coastal habitat a variability of physical and chemical parameter has a prominent impact on the microbial community structure.

Degradation of polychlorinated biphenyl (PCB) by a consortium obtained from a contaminated soil composed of Brevibacterium, Pandoraea and Ochrobactrum

An indigenous polychlorinated biphenyl (PCB)-degrading bacterial consortium was obtained from soils contaminated by transformer oil with a high content of PCBs. The PCB degrader strains were isolated and identified as Brevibacterium antarcticum, Pandoraea pnomenusa, and Ochrobactrum intermedium by 16S rRNA gene sequence phylogenetic analysis. The PCB-degrading ability of the consortium and of individual strains was determined by using GC/MS. The PCB-degrading capacities of the consortium were evaluated for three concentrations of transfomer oil ranging from 55 to 152 μM supplemented with 0.001% biphenyl and 0.1% of Tween 80 surfactant. PCB biodegradation by the consortium was favored in the presence of both additives and the greatest extent of biodegradation (67.5%) was obtained at a PCB concentration of 55 μM. Each bacterial species exhibited a particular pattern of degradation relating to specific PCB congeners. Isolated strains showed a moderate degradation capability towards tetra-, hepta-, and octa-chlorobiphenyls; although no effect on penta-, hexa-, and nona-chlorobiphenyls was observed. Recently, PCB degradation capacity was recognized in a Pandorea member; however, this is the first study that describes the ability of Brevibacterium and Ochrobactrum species to degrade PCBs.

Genetic diversity and phylogenetic relationships of bacteria belonging to the Ochrobactrum– Brucella group by recA and 16S rRNA gene-based comparative sequence analysis

The genetic diversity and phylogenetic interrelationships among 106 Ochrobactrum strains ( O. anthropi: 72, O. intermedium: 22, O. tritici: 5, O. oryzae: 2, O. grignonense: 2, O. gallinifaecis: 1, O. lupini: 2), the type strains of the eight Brucella species and other closely related taxa were studied by recA and rrs gene (16S rRNA) comparative sequence analysis. Both markers correctly delineated the various Ochrobactrum species; however, resolution at the subspecies level was considerably higher in the recA gene-based approach. Phylogenetic analyses using neighbor-joining, parsimony, and maximum likelihood algorithms generated trees with similar topologies but the overall branching order, and also the order of the subclades, were not stable in either assay, which could be explained by generally high recA and rrs sequence similarities. Ochrobactrum and Pseudochrobactrum formed separate clades distinct from other Alphaproteobacteria with Bartonella, Agrobacterium, and Rhizobium as the closest relatives. O. gallinifaecis was the most distinct member, when compared to the type species O. anthropi, with rrs and recA similarities of 96.2% and 81.4%. Brucella species were indistinguishable, exhibiting high rrs and recA gene similarities of 98.6% and 85.5% compared with Ochrobactrum intermedium. At the protein level, all RecA sequences among the various Ochrobactrum species and between Ochrobactrum and Brucella were highly similar with only a few amino acid substitutions. O. anthropi and O. tritici were indistinguishable by means of their RecA proteins. A set of initially biochemically classified strains did not cluster within their assigned species and they either grouped within other known species or grouped as potential novel Ochrobactrum species. In further investigations, these strains were reclassified and described as novel species. In summary, Ochrobactrum is a highly diverse genus comprising several novel species. We recommend recA- in addition to rrs gene-analysis for correct species allocation and subtyping of novel Ochrobactrum isolates.

Specific detection and differentiation of Ochrobactrum anthropi, Ochrobactrum intermedium and Brucella spp. by a multi-primer PCR that targets the recA gene

Ochrobactrum anthropi, Ochrobactrum intermedium and Brucella spp. are phenotypically and genetically closely related pathogens that may cause disease with similar clinical presentation. Consequently, difficulties in their identification and differentiation have been reported. In this study, a sensitive recA gene-based multi-primer single-target PCR (MP-ST-PCR) was developed that allowed the specific detection and differentiation of these clinically relevant pathogens. The specificity of the assay was evaluated using a representative panel of 50 O. anthropi and 16 O. intermedium strains and the type strains of all Brucella spp. Detection limits for purified DNA from O. anthropi, O. intermedium and Brucella melitensis were 100, 10 and 100 fg, respectively. Brucella DNA was also successfully detected in various clinical specimens from a human patient with culture-proven brucellosis and from a Brucella-infected sheep and its aborted fetuses. The sensitivity of the MP-ST-PCR was comparable to that of an evaluated in-house Brucella real-time PCR assay. The developed assay closes a diagnostic gap and provides a simple but robust tool for the sensitive detection and correct identification of O. anthropi, O. intermedium and Brucella spp.

Screening and characterization of antifungal clusterbean (Cyamopsis tetragonoloba) rhizobacteria

About 377 guar (Cyamopsis tetragonoloba) rhizobacteria were isolated from cultivated soils of north-west India (Thar Desert) and their antifungal activity against Macrophomina phaseolina (strains of groundnut, mungbean and guar) and Fusarium oxysporum (strains of chickpea and cumin) was examined. Isolates were characterised for generic types and physiological/functional diversity. About 19% isolates representing 24% locations were inhibitory to fungal growth. Isolates 009071, 009073, 009078 and 102354 recorded maximum inhibition of pathogenic fungi on plates. Isolate 034206 gave highest %RI, 009073 showed maximum protease activity and 102354 gave highest salt tolerance. Net house and field screening results revealed that isolates 004052, 009071, 009073, 001001, 094340 and 102354 had potential for biocontrol of disease. Partial sequencing of 16S rRNA gene of 61 isolates showed that 85% of isolates belonged to genus Bacillus. Phylogenetically, however, there were four clusters in the Bacillus group comprising of Bacillus subtilis, B. cereus, B. pumilus and B. sphaericus. One isolate was identified as B. flexus, while six isolates were Bacillus spp. Four isolates were identified as Achromobacter xylosoxidans, two as Bacterium (unclassified bacteria), and one each as Ochrobactrum intermedium, Pseudomonas aeruginosa and Ralstonia sp.

Brucellosis of the Common Vole (Microtus arvalis)

A systemic disease occurred in a wild population of the common vole Microtus arvalis in South Moravia (Czech Republic) during the years 1999-2003. Acute infections were characterized by edema of extremities, occasionally with colliquating abscesses, arthritis, lymphadenitis, perforations of the skin resulting from colliquated abscesses, orchitis, and peritoneal granulomas. From the clinical samples, small Gram-negative coccobacilli were isolated and identified as Ochrobactrum intermedium by API 20NE and colistin sensitivity profiles. However, subsequent rrs (16S rRNA) and recA (recombinase A) gene sequencing analysis of two isolates (CCM 4915=CAPM 6434; CCM 4916=CAPM 6435) identified them as Brucella sp. with sequence identities of 100% to other Brucella spp. Analysis of the omp2a/b genes confirmed the two isolates as Brucella. In AMOS polymerase chain reaction (PCR), a 2000-bp fragment was generated that was not seen in other brucellae. Experimental infection of outbred ICR mice with these isolates resulted in a mortality rate of 50%. Based on the results of the molecular investigations and the mortality observed in experimentally infected mice we conclude that the epizootic was caused by Brucella sp. and not by Ochrobactrum intermedium. The study demonstrates the limitations of commercial biochemical test systems in accurately differentiating among Ochrobactrum and Brucella.

Ochrobactrum intermedium ANKI, a nitrogen-fixing bacterium able to decolorize azobenzene

Morphological and biochemical properties of the nitrogen-fixing strain Ochrobactrum intermedium ANKI, intensely growing on media with azo compounds, and its resistance to various common xenobiotics were investigated. The kinetics of azobenzene transformation by O. intermedium ANKI was studied. Under cometabolism conditions, up to 40 mg of azobenzene per liter of medium were decolorized within one week. It was shown that the strain possessed molybdenum-dependent nitrogenase activity, and its nitrogenase system was sensitive to oxygen and fixed nitrogen in the medium.

Ochrobactrum pseudintermedium sp. nov., a novel member of the family Brucellaceae, isolated from human clinical samples

Three novel Gram-negative, non-fermenting aerobic bacilli were isolated from human clinical samples. They shared more than 99.8 % of the 16S rRNA gene nucleotide positions. The strains were related to Ochrobactrum intermedium with about 97.48 % 16S rRNA gene sequence similarity. In 16S rRNA gene-, dnaK- and rpoB-based phylogenies, the strains were grouped in a lineage that was distinct from other Ochrobactrum species in the family Brucellaceae. Fatty acid composition, polar lipids, quinone system, DNA-DNA relatedness, genome organization, and physiological and biochemical data differentiated these isolates from recognized species of the genus Ochrobactrum. The three clinical strains therefore represent a novel species within the genus Ochrobactrum, for which the name Ochrobactrum pseudintermedium sp. nov., is proposed. The type strain is ADV31(T) (=CIP 109116(T)=DSM 17490(T)). The DNA G+C content of strain ADV31(T) was 54.5 mol%.

Biodegradation of nicotine from tobacco waste extract by Ochrobactrum intermedium DN2

Ochrobactrum intermedium DN2 was used to degrade nicotine in tobacco waste extracts. The optimal temperature and pH of nicotine degradation by strain DN2 was 30-37 degrees C and 7.0, respectively. Under these optimal conditions, the average degradation rate of nicotine in a 30L fed-batch culture was 140.5 mg 1(-1) h(-1). The results of this study indicate that strain DN2 may be useful for reducing the nicotine content of reconstituted tobacco.

Growth stimulatory effect of Ochrobactrum intermedium and Bacillus cereus on Vigna radiata plants

This study assessed the plant growth-promoting ability of the bacterial strains Ochrobactrum intermedium (isolate CrT-1) and Bacillus cereus (isolate S-6). Two chromium resistant bacterial strains isolated from chromium-contaminated wastewater and soils were identified as O. intermedium CrT-1 and B. cereus S-6. These strains were inoculated on seeds of mungbean Vigna radiata var NM-92, which were germinated and grown under chromate salts (300 microg ml(-1) of CrCl(3)or K(2)CrO(4)). The data show that Cr(VI) was more toxic because of its better availability to plants roots when compared with Cr(III). The major part of Cr(VI) supplied to the seedlings was reduced to Cr(III) in the rhizosphere by the bacterial strains, thus lowering the toxicity of chromium to seedlings. Strains have significant Cr(VI) resistance and reduction potential and have ability to enhance mungbean plant growth under chromium stress. These strains could be utilized for the growth of economically important cash crops as well as for the bioremediation of chromium-polluted soils.