Evaluation of assays for screening polycyclic aromatic hydrocarbon-degrading potential of bacteria

Abstract

Within a 30-day incubation laboratory study, the polycyclic aromatic hydrocarbon (PAH) degradation profile of two bacteria, Planomicrobium sp. RNP01 and Rhodococcus sp. RNP05 were studied by three microtiter plate assays to reveal the combination of certain biological and biochemical characteristics which are reliable indicators in evaluation of bacterial biodegradation abilities. The three assays, which are hydrocarbon growth assay, 2,6-DCPIP assay and dehydrogenase activity assay revealed that Rhodococcus sp. RNP05 exhibited better potential for PAH degradation than Planomicrobium sp. RNP01. Differences between initial and final optical density and specific growth rate constants were significantly higher (r = 0.995, P < 0.05) in case of Rhodococcus sp. RNP05 on all tested substrates, as compared to Planomicrobium sp. RNP01. This was confirmed by GC-FID analyses. Dehydrogenase activity of Rhodococcus sp. RNP05 was higher (r = 0.9995, P < 0.05) than Planomicrobium sp. RNP01 and correlated positively with the hydrocarbon growth assay (r = 0.999, P < 0.05, for Rhodococcus sp. RNP05, r = 0.986, P < 0.05 for Planomicrobium sp. RNP01). This study has shown that the combination of these assays could be used for determining the bioremediation potential of PAHs in petroleum contaminated soil with the ability of screening a large number of bacterial strains.

Degradacija policikličnih aromatičnih ugljovodonika (polycyclic aromatic hydrocarbons - PAH) ispitivana je testovima u mikrotitar pločama pomoću dva bakterijska soja Planomicrobium sp. RNP01 i Rhodococcus sp. RNP05, u periodu od 30 dana. Rezultati ispitivanja su ukazali da se kombinacijom određenih bioloških i biohemijskih karakteristika mogu kreirati dobri indikatori u proceni bakterijskog degradacionog potencijala. Tri testa, 2,6-DCPIP test, test dehidrogenazne aktivnosti i test rasta na ugljovodonicima pokazali su da Rhodococcus sp. RNP05 ima veći potencijal za degradaciju PAH jednjenja u odnosu na Planomicrobium sp. RNP01. Razlike u početnoj i krajnjoj optičkoj gustini i specifične konstante rasta bile su značajno više (r = 0,995, P < 0,05) u testu sa Rhodococcus sp. RNP05 na svim testiranim supstratima, u poređenju sa Planomicrobium sp. RNP01, a dobijeni rezultati su potvrđeni gasno-hromatogafskom-FID analizom. Dehidrogenazna aktivnost soja Rhodococcus sp. RNP05 bila je viša u odnosu na Planomicrobium sp. RNP01 (r = 0,9995, P < 0,05) i u pozitivnoj korelaciji sa testom rasta na ugljovodonicima (r = 0,999, P < 0,05, za Rhodococcus sp. RNP05, r = 0,986, P < 0,05 za Planomicrobium sp. RNP01). Rezultati prikazanog istraživanja ukazuju na to da se kombinacija ovih testova može koristiti za određivanje bioremedijacionog potencijala za razgradnju PAH jedinjenja u zemljištu zagađenim naftom, pri čemu se dobija i mogućnost testiranja velikog broja bakterijskih sojeva.