Multiple Metal Resistance Research Articles

Multiple Metal Resistant Transferable Phenotypes in Bacteria as Indicators of Soil Contamination with Heavy Metals (6 pp)

Environmental contamination by heavy metals affects microbial communities. The number of single and multiple heavy metal resistant bacteria may be an indictor of the level of contamination. This paper details the isolation and characterisation of metal resistant microorganisms isolated from rhizosphere/soil samples obtained from an abandoned zinc, lead and copper mine and a local unaffected site. This data was compared to the level of heavy metal in the soils to establish the effect of metals on the microbial community and to determine the relationship between pollutant levels and resistant strains. This paper outlines the diversity of transferable resistance determinants between both sites and details the levels of heavy metal resistant bacteria and those expressing transferable multiple heavy metal tolerance. The sample sites were located in Co. Galway, Ireland. The first sample site (site A) was a former lead, zinc and copper mine, which was closed in 1961 due to exhaustion of ore. The second site (site B) was located two and a half kilometres from the mining site and was not affected by the mining operations. Composite soil samples were characterised for general soil matrix composition, organic content, pH and general chemical parameters. The soil was also enumerated for the total viable heterotrophic counts and tested on Pseudomonas selective agar (PSA) for total Pseudomonas counts and Sucrose Asparagine (SA), which is semi-selective for fluorescent Pseudomonas. Samples from both site A and site B were analysed by atomic absorption spectrophotometry for the presence of heavy metals. In the case of copper, which has a Dutch list recommended minimum permissible level of 190 µg/Kg dry weight, the levels detected at site A were 1270 µg/Kg dry weight while site B was detected at 36 µg/Kg. The arsenic levels detected at site A were eight times the permissible level (416 µg/Kg) while only half the permissible level was found at site B (13 µg/Kg). Zinc concentrations were also high at site A (4460 µg/Kg) while at site B (553 µg/Kg) they were well below the Dutchlist guidelines (720 µg/Kg). A large number of heavy metal tolerant strains were isolated from both sites. 270 isolates (site A (170) and site B (100)) were screened against 8 metals to examine the extent of multiple resistance. 82% of the strains from site A were found to be resistant to 5 metals. A total of 18% showed resistance to all 8 metals and of those examined only 4% were resistant to only one metal. In contrast isolates from site B showed no multiple resistance to more than 5 metals, while 62% showed resistance to individual metals only. Site A had a higher level of multiple heavy metal resistance strains. Stains isolated from site A had 23 (14%) isolates resistant to zinc, copper, nickel, arsenic and cobalt and site B had no bacteria resistant to all five of these selected metals. The transferability of heavy metal resistance was investigated in the case of 60 multiple heavy metal resistant isolates taken from site A and 50 multiple resistance isolates from site B. Transfer was only detected in isolates from site A, 13% showed transfer and expression of copper, zinc and arsenic resistance determinants. In most cases the transconjugants only expressed resistance to copper, zinc and arsenic, which were the metals, used for selection. This co-transfer of all three determinants suggests a genetic link between these resistance determinants. Heavy metal resistant bacteria are present in both sites, however, the number and presence of multiple transferable resistance phenotypes are confined to the isolates from the heavy metal contaminated site. The presence of high levels of heavy metals selects these multiple resistance phenotypes. Within these communities there seems to be little diversity between the microorganisms, which provides a hugely preferable environment for gene transfer of such metal resistant determinants. The experiments have shown a microbes ability to mobilize heavy metal determinants and a relationship between heavy metal resistance and metal contamination has been identified These multiple heavy metal resistant bacteria could eventually be used for detection and qualification of the level of heavy metal-polluted soil/water environments.

Multiple metal resistance in the ciliate protozoan, Vorticella microstoma, isolated from industrial effluents and its potential in bioremediation of toxic wastes.

Multiple metal resistance in the ciliate protozoan, Vorticella microstoma, isolated from industrial effluents and its potential in bioremediation of toxic wastes.

Identification and characterization of Rhodopseudomonas spp., a purple, non-sulfur bacterium from microbial mats

A species of facultative photo-organotrophic, purple, non-sulfur bacterium was isolated from mixed-species microbial mats, characterized and examined for metal tolerance and bioremediation potential. Contributing mats were natural consortia of microbes, dominated by cyanobacteria and containing several species of bacteria arranged in a laminar structure, stabilized within a gel matrix. Constructed microbial mats were used for bioremediation of heavy metals and organic chemical pollutants. Purple, non-sulfur bacteria are characteristically found in lower strata of intact mats, but their contributing function in mats survival and function by mediating the chemical environment has not been explored. The gram-negative rod-shaped bacterium, reported here, produced a dark red culture under phototrophic conditions, reproduced by budding and formed a lamellar intracytoplasmic membrane (ICM) system parallel to cytoplasmic membrane, which contained bacteriochlorophyll a and carotenoids. This strain was found to have multiple metal resistances and to be effective in the reductive removal of Cr(VI) and the degradation of 2,4,6-trichlorophenol. Based on the results obtained from morphology, nutrient requirements, major bacteriochlorophyll content, GC content, random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) profile and 16S-rDNA phylogenetic analysis, this member of the microbial mats may be identified as a new strain of the genus Rhodopseudomonas.

Metal and antibiotic resistance traits in Bradyrhizobium sp. (cajanus) isolated from soil receiving oil refinery wastewater

Forty isolates of Bradyrhizobium sp. (cajanus) were isolated from the nodules of pigeon pea plants grown in fields receiving petrochemical industrial wastewater for the past 12 years and characterized using standard methods. The heavy metal analysis of field soil and treated wastewater showed their presence in varying concentrations. All isolates showed resistance to one or more metals at concentrations >200 μg/ml. Multiple metal resistance was a common phenomenon in these isolates. There was no correlation between extractable soil metal concentration and the ability of the isolates to tolerate metal salts in their growth medium as evidenced from their minimum inhibitory concentration (MIC). However, high incidence of metal resistance and the multiple nature of resistance might have been the result of continuous exposure of these strains to heavy metals in the treated wastewater of Mathura Oil Refinery. These strains were also found to be resistant to one or more of the 13 antibacterial drugs tested.

Studies on mercury-detoxicating enzymes from a broad-spectrum mercury-resistant strain of Flavobacterium rigense.

Flavobacterium rigense strain PR2, a broad-spectrum mercury-resistant bacterium abundantly present in soil exhibited multiple metal resistance properties. Mercury resistance was due to the sequential action of two mercury-detoxicating enzymes, organomercurial lyase and mercuric reductase. The levels of these enzyme activities were determined using different mercury compounds as inducers and substrates. Mercuric reductase was partially purified from the bacterium and the physicochemical properties of the enzyme were studied. The effect of several enzyme inhibitors and heavy metal ions on the enzyme activity was also studied.

Differential sensitivity of pigmented and non-pigmented marine bacteria to metals and antibiotics

The response of pigmented and non-pigmented marine bacteria to metals and antibiotics was investigated. The two groups responded differently to heavy metals and antibiotics. Pigmented bacteria were more resistant to metals. Among the metals, Zn and Hg were less toxic to pigmented bacteria than Cd. Pigmented strains were resistant to antibiotics, particularly at higher concentrations. All the strains, irrespective of their pigment, showed multiple metal and drug resistance.