VOSKOBOYNIKOV G.M., ILYINSKY V.V., LOPUSHANSKAYA E.M., PUGOVKIN D.V.

Murmansk Marine Biological Institute KSC RAS, Russia, Lomonosov Moscow State University, Russia, FSUE Mendeleev Russian Federal Research Institute of meteorology, Russia

Fucus algae High resistance to oil contamination (Voskoboynikov, et al., 2004; Stepanyan, Voskoboynikov, 2006) was detected earlier.

It was registered that Fucus vesiculosus keeps ability to photosynthesis, growth, polysaccharides synthesis even after long-term petroleum influence. We set up a hypothesis of macrophytes role in water surface purifying of oil pollution not only due to oil products retention by analogy with slick bar, and of inclusion them in metabolism, and also of oil neutralizing due to formation of algae and hydrocarbon oxidizing bacteria symbiotic association.

During experiments at sea and basins we detected high ability of fucus algae to stop oil slick spread provided that they are properly positioned on substrates of surface water layer.

Analyses have shown, that oil mass concentration in algae from zone of permanent contamination (30 days) oil content in water constitutes 6956 mg/kg, 3238 mg/kg of them are removed from surface and 3718 mg/kg are extracted from algae tissues. In a week after algae taking from polluted place and putting them in moist room at the temperature analogous to habitat, oil mass concentration amounted 1084 mg/kg, 832 mg/kg of them were from surface and 252 mg/kg were extracted from algae tissues.

In two weeks oil mass concentration in polluted algae amounted to 807 mg/kg.

Thus, the role of marine macrophytes in water surface purifying of oil pollution not only due to sorption and mechanic retention of oil slicks, and also due to this contamination destruction is confirmed. Apparently, that oil hydrocarbon destruction takes place as a result of their inclusion in fucus metabolism or due to biodegradation by hydrocarbon oxidizing bacteria or due to these processes together.

We have experimentally verified whether epiphytic hydrocarbon oxidizing bacteria (EHB) living on focus thallomes surface are really able to biodegradation of oil hydrocarbon (OH). For this purpose we subjected fragments of focus thallome to ultrasonic processing for removal of located on them bacteria. Preliminarily focus thallomes were washed with sterile sodium chloride solution (3 %) and then put in a sterile 100 ml glass containing 50 ml of sterile mineral medium MMS. Conditions of ultrasonic processing are the following: processing time is 1 minute, current is 0,4А, ultrasonic processing frequency is 22 kHz.

After ultrasonic processing fucus was taken from a glass with medium, and the medium containing desorbed from focus EHB were put in 0,5 l tilted flasks containing 150 ml of MMS medium. Then in these inoculated flasks with medium 1 ml of sterile diesel oil (DO) was brought in as the only carbon and energy source. Flasks were put in rocking-bar and incubated for 3 weeks under room temperature of 20 degree centigrade (200С).
DO extraction from flasks was carried out with 100 ml of perchloromethane in a separatory funnel during 30 minutes. We precipitated extracts during one day, then poured out low layer of extragent, run it through the column with alumina and analysed on spectrophotometer “SP-1100” (Pye Unicam, England) over the range 2500 - 3500 sm1.

We found out the following:

1. EHB living on fucus thallomes are able to destruct oil under conditions of laboratory experiment. Maximum DO consumption by bacteria during 3 weeks of exposition amounted to 49 % of control.
2. EHB living on fucus thallomes taken from oil contaminated waters consumed 7-9% of DO more than bacteria living on focus from extracted of pure waters. It testifies their higher potential of hydrocarbon oxidizing activity.

Thus, we have experimentally shown that epiphytic bacteria living on fucus thallomes are able to destruct OH, and EHB desorbed from surface of fucus taken from oil contaminated waters (Murmansk port) are able to destroy more OH than EHB, desorbed from fucus from oil uncontaminated waters (Belokamenka settlement coast).

At the second stage we determined quantity of cultivated (able to grow on nutrient medium) EHB living on fucus thallomes taken from oil-contaminated and from pure waters. First, we have chosen optimum method of bacteria removal (desorption) from fucus thallomes. We compared three different methods: ultrasonic processing of thallomes on device UZDN-1, processing of thallomes on mixer-dispenser IKA ULTRA-TURRAX Tube

Drive with attachment-dispenser DT-20 (Germany) and also manual removal of bacterial coating from fucus thallomes surface with sterile cotton wad. The last method appeared to be the most efficient and it was used in further activities on definition of EHB quantity on fucus thallomes.

Fucus taken from oil-contaminated waters and from waters free of the given type of contamination were subjected to removal with a wad. It was found that EHB quantity on fucus from contaminated waters considerably exceeds EHB quantity on fucus from pure waters and this difference can make 2 - 4 orders.

Thus, by now there is no doubt that EHB living on fucus surface able to destroy OH and their quantity on fucus from oil-contaminated waters is higher, than on fucus from places free of these pollutants. It allows us to assume that EHB take part directly in processes of natural waters purification of oil contamination in fucus plantations.

Ecological investigations were carried out at financial support of RAS Presidium “Innovations and developments program” and Global fund of ecological researches.

OIL AND GAS OF ARCTIC SHELF 2008