IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L.22 (3) 2009 The Impact of Ascending Levels of Crude Oil Pollution on Growth of Olive ( Olea europaea Linn) Seedlings B. A. A . Ali , H . H. Ali and G . A. Shaker Iraq natural History Museum and Research Centre , Unive rsity of Baghdad Abstract A st udy on the imp act of ascending lev els of crude oil on the growt h of transp lanted seedlings ( M arch2005 ) of Olive (Ol ea europaea Linn) was carried out at t he exp erimental area of Iraq Natural Hist ory M useum and Research centre / Baghd ad University (Bab-Al-M adham –Baghd ad) grown under field condition and continued till Ap ril 2008.The exp eriment was laid out in comp lete rando mized desi gn ( CR D ) with five levels of p ollution (0.0 , 0.5 , 1.0 , 2.0 and 3.0 liter / seedlin g ) p oured at the soil surface , each seedling represented one replicate and was replicated four times . Data collected from the exp eriment were visual sy mptoms , p ercents of seedlings death, p lant height and total dry weight of harvest ed p lants. T he results indicated that crude oil p ollution caused an adverse effect on the Olive (Olea europaea Linn). The crude oil led to leaf chlorosis , dry ness and death of seedlings. The adverse effects were p rop ortional with the levels of p ollution. Plant heights and total dry weights were sign ificantly reduced (p < 0.05) as a consequence of p ollution. Results of the st udy indicated that the effect of p ollution clearly app eared after three weeks at the high levels then the sy mptoms extended to lower levels during the first six months. The st udy showed that treated plants with low levels of crude oil p ollution were less growt h but survived and st ay ed alive, while the high er levels led to seedlings de ath. It was obvious t hat the adversely affected p lants could not be able to recov er even after 27 months after p ollution, the time of the exp eriment. Introduction Olive tree (Olea europaea Linn) , is a small tree , mostly not exceeding 6 t0 9 m in height , and bear ing small lan ceolate leaves and axillaries flowers oblong. While the p roduction of oil is the main aim of p lanting this sp ecies ; a sp ecific orchard man agement should be app lied to maintain acceptable levels of p roductivity , oil quality , and cost of p roduction (1). It was introduced to Iraq since a long time ago and extensively cultivated in different p arts of Iraq( 2 ) esp ecially in the up p er parts of t he country . The tree is considered as t oler ant for drou ght, and for nitrogen deficiency (3 ).Recently the Iraqi M inist ry of Agriculture considers the tree as one of the important economic p lants and adop ted many p rojects to exp and planting areas and encouraged farmers to grow the tree in their private plantations. Soil p rotection and remediation of oil p olluted soil sites became an imp ortant p art of the environmen tal p rotection p olicy in many oil p roducin g countries. Resear ch in p reventing and recuperating oil p ollution became an inte gral p art of p etroleum industry activities because the crude o il h as consider ed as the most last ing and persistent environmental pollutants affecting the soil ecosy st ems. Crude oil pollution is t he most frequent one in Iraq than in any other producing countries due to t he wars , fighting and unn atural security conditions , where p ipe lines , oil transp orting trailers and tanks were mostly exp osed to attack by exp losions causing an intensive crude oil - soil pollution . IBN AL- HAITHAM J . FO R PURE & APPL. SC I VO L.22 (3) 2009 Plants resp ond differently to p ollution dep ending on many factors such as typ e and level of pollutant , a ge and size of the plant and Its inherent characterist ic, in addition to environmental conditions under which the p lant is grown. Oil p ollution has adversely effects on different p lant sp ecies ( 4) ,( 5) , (6) and (7) referr ed to crude oil as p hy totoxic for many p lants and consider ed the p oor growt h of oil t reated plants as attributed to suffocation of p lants caused by exclusion of air by the oil or exh austion of oxy gen by the increased microbial activity and toxicity from some oil comp ounds . In sp ite of the adverse effects of oil p ollution on the plants, no much works were done in Iraq and due to the shortage of the available literatures related to this subject current study had been done and aimed to invest igate the p erformance of Olive tree (Olea europaea ) growt h under different levels of crude oil pollution . Materials and Methods Fifty seedlings of Olive (Olea europaea ) n early 30-40 cm tall and fr ee of insects or dieses infections obtained from a local nursery and were transp lanted in a clay soil with distances of one by one meter between seed lin gs at the exp erimental ar ea of the Iraq Natural History Research Centre & M useum / Baghd ad University on M ay of 2005 . Twenty healthy seedlings and near ly equally height of them w ere selected on De cember 2005 . Five levels of Basrah light crude o il obtain ed from Iraqi Oil M inist ry used ( 0.0 (L0) , 0.5 ( L1), 1.0 (L2) , 2.0 (L3) and 3.0 (L4) liter of oil / seedlin gs) were distributed randomly on the selected seedlin gs and each level was repeated four times . Soil Crude o il p ollution was simulated by p ouring the crude oil over the surface soil and inorder to ensure that the amounts in all treatment were distributed over the same ar ea, a ring of a plast ic tube with diameter of 30 cm was made , and was p ut on the soil to surround the seedling, then the crude oil was distributed equally and evenly inside the ring. Resulted trees were h arvest ed on Ap ril 2008. The imp act of the seedlin g as affected by crude o il was st udied depending on the visual sy mptoms and observations app eared on the p lants, p ercents of death seedlings( p ercent of death seedling to the 4 replications of each treatment) were collected during the exp eriment t ime and on the measurements of the heights and total dry weight of harvest ed trees. Complete randomized design (CRD) was used as an exp erimental d esign and the results were were st atist ically analyzed by using Statist ica program (99 edition)(8). Results and Discussion The treated surface soils were turned immediately to oily with brown color after adding the crude oil. The resulted color was p rop ortional with oil level used. The surface soil of L1 , L2 and L3 was able to absorb the crude oil amounts directly after the addition , while soil of L4 took more time to absorb and became saturated with it. T he intensity of the brown color of the surface soils started to the reduce gradually with in time and this could be due to t he degradation of the oil or the penetration within the soil. The treated surface soils were able to recover to the original color during the first six months of the exp eriment . The consumed time to recover the original soil color was also p rop ortional with t he levels of crude oil used . All the seedlings of Olea europaea exp osed to crude oil resp onded negatively to p ollution in comp arison with control treatment ( L0) Fig (1). Chlorosis (y ellowing of p lant leaves) was the early distinguished sy mptoms observed on all of the crude oil treated seedlings . Although the chlorosis was a common feature for all of the treate seedlings, their y ellow color intensity was different and ranging between light to intensive depending on levels of p ollution being used . The chlorosis was first ly observed in some of the old leaves and then develop ed with in time to sp eared all over the p lant and eventually led to the dryness and death of seedlings in some cases esp ecially in the higher levels of p ollution ( L3 and IBN AL- HAITHAM J . FO R PURE & APPL. SC I VO L.22 (3) 2009 L4). In sp ite of no obvious cases of infection were observed during the time of the exp eriment, it was p ossible to ensure that t he dryness and death followed the chlorosis were due to p ollution and not for other reasons and therefore, p ut the dead seedlings under a close visual and microscopic examination . First case of seedling dry ness and death was recorded on L4 treatment after three weeks, and then the p henomena app eared on other treatments within the first six months after p ollution. Crude oil p ollution caused death to some rep lications of the treated seedlings and the p ercent of death was directly p rop ortional to the level of p ollution and ranged between 75 to 100 % Fig( 2 ). The increase of the death p ercentage of Olea europaea with the increase of the levels of crude oil p ollution was clearly indicated to an adverse effect of oil on this sp ecies. It is well known that the visible sy mptoms have often p rovided the first indication of other p roblems which might be resulted from the impaired metabolism within the plant. Crude oil p ollution led to different sy mptoms on seedling during the first six months after exp osing to crude oil. Old leaves of treated seedling became p ale in comp arison t o control and this could be due to the breakdown in chlorop hy ll p roduction. Such sy mptom had similar asp ects of nitrogen deficiency , where sp ecific sy mptoms of nitrogen deficiency first app ear on the oldest leaves. Plant suffering from nitrogen deficiency is p ale in color in comp arison to healthy one due to the breakdown in chlorop hy ll p roduction. Such observations were noticed by (9) where they referred that t he sp ecific sy mptoms of nitrogen deficiency first app eared ( as with deficiencies of p hosp horus and p otassium) on the oldest leaves, and these become p aler then turned to chlorosis ( M arked y ellowing) . It is well known t hat crude oil pollution leads t o increase soil organic carbon as well as the p op ulations of total heterotrop hic microorganisms and also, a reduction in nitrate-nitrogen and available phosp horus (10).The shortage of nutrients availability ,esp ecially the nitrogen as a result of the increase of the total soil heterotrop hic could be exp ected according to the current st udy since no nitrogen fertilizer was added. For t his reason, the nitrogen deficiency could be as one cause of the chlorosis and can not be rolled out. The adverse effects of crude on Olea europaea trees continued for more than 27 months after p ollution . No more death happ ened for trees after the first six months but the survived trees from crude oil p ollution p ossessed weaker growt h in comp arison with control as reflected by trees height and total dry weight .The mean height for control trees was 203.7 cm , but due to p ollution it was reduced to 79.5 , 36.5 , 14.7 , 0.0 cm for L1, L2, L3 and L4 resp ectively table (1). Total dry weigh also was reduced from 675.5 to 195.87, 30.62 , 12.25 , 0.0 gm for the same three levels Table (2). Current results clearly refer to the adverse effects of crude oil on p lants growt h and were in an agreement with to the results of (4),(5),(7), (11), (12), and (13) . Since up take of water and nutrients is carried out by the root sy st em, untreated seedlings of Olea europaea with sound, healthy roots grew normally while the treated seedlings could suffer from anatomical aberration that may caused by p resence of oil films in a sp ecific parts of p lant leading to a negative effect on the growt h. In anatomical study (14) noticed the p resence of oil films in the epidermal and cortical regions of root, st em and leaves of the plant . The increase of death p ercent of Olea europaea seedlings with increasing of p ollution as occurred in the current st udy , could be due to one or a combination of following factors; disrup tion of root-soil-water relationship , direct imp act on p lant metabolic p rocesses , toxicity of living cells, reduced oxy gen exchange between the atmosp here and soil , nutrients immobilization and an alteration in the p hy sical, chemical and biological p rop erties of soil. Such factors could have observed visual sy mptoms , reduction in biomass or deaths of seedlings. From the foregoing, it is clear that crude oil may affect p lant by causing direct death due to coating the roots and soil p articles by it or t hrough the affect of toxic comp onents, where the oil or its comp onents interfere with cellular p rocesses, resulting the plant death . Conclusion Some conclusions could be mentioned according to this exp eriment ; (a): Crude oil app lication to soil IBN AL- HAITHAM J . FO R PURE & APPL. SC I VO L.22 (3) 2009 has adverse effects on the growt h of Olea europaea seedlings ,(b): The effect being p rop ortional to the concentration of crude oil is app lied, (c): Oil may affect the p lants by causing direct death in high levels of p ollution , (d): In some levels of p ollution olive p lants may undergo a weakness in growt h and reduction of biomass p roducing a st unted plants, and (e) the effect of p ollution by crude oil on Olea europaea may last more than 27 months (t he period of the exp eriment). Acknowledgeme nts The authors are grateful to Iraqi ministry of oil and the management of Dorra refinery for p roviding the Basroh crude oil that was used in the current st udy . References 1-M ichelakis,N.(2002).ISHS Acta Hort iculturae. 586 :239-24 http://www.actahort.org/books/586/586_45.htm 2 - Chakravarty , H.I. 1976. Plant wealth of Iraq. Vol (1), Botany Directorate, M inist ry of Agricu lture and Agr arian Reform, Baghd ad, Iraq. 3 - Fernández-Escobar ,R., Sÿ nchez-Zamora ,M .A., Uceda, M . and Beltr ,G. (2002).ISHS Acta Hort iculturae 586: 158-163. http://www.actahort.org/members/showp df?booknrarnr=586_88 4 - Agbo gidi, O.M , Onosode,A.T and Ofuoku,A.U. (2006). Europ ean. J. of Scientific Research, 13 (2). :165-171. 5 - Akaninwor, J.O. , Ay eleso, A.O. and M onago , C.C ( 2007). Scientific Research and Essay. 2(4): 127-131. 6 - Ogbonna,D.N , Iwegbu e,C.M .A , T.G. Sokari and Akoko, I.O.( 2007). Earth and Environmental Science. 27( 2) :303-309. 7 -Udo,E.J. and Fey emi,A.A.A. (1975). 4:537-540. 8-Statist ica for windows "99 Edition" , (1999) . Kernel release 5.5 A , StatSoft,Inc . 9 - Snowball, K. and Robson , A.D (1983) .Symptoms of nutrient deficiency . Dep artment of soil science and plant nutrition, Institute of Agriculture, University of West ern Australia. 10- Okolo, J.C. , Amadi, E.N and Odu, C.T .I (2005). App lied ecolo gy and environmental r esearch 3 (1) : 47-53. 11 - Issoufi, I , Rhykerd, R.L and Smiciklas, K.D (2006). J Agron Crop Sci, 192 ( 4 ):310-317. 12-Baek, Ky ung-Hwa , Kim, Hee-Sik , Hee-M ock, Oh , Yoon, By ung-Dae,Yoon , Ki m,Jaisoo and Lee, In- Sook. (2004). J.of Environ sci and Health. A39 (9 ): 2465-2472. 13 - Gill,L.S. , Ny awuame,H.G.K. and Ehihametelor, A.G. (1992). Newsletter 5:46-50. 14 - Anoliefo, G.O. (1991). Forcados blend oil effect in resp iration,metabolism, elemental comp osition and growt h of Citrullus vulgaris (Schard).Ph.D. T hesis Benn.pp. 6-30. In Gill,L.S. , Ny awuame,H.G.K. and Ehihametelor, A.G. (1992). IBN AL- HAITHAM J . FO R PURE & APPL. SC I VO L.22 (3) 2009 Table (1): Plant height (cm) at the end of the experiment as influenced by five levels of crude oil 0.0 refer to dead plant . Means with different superscripts are significantly different (LSD) at p< 0 Table( 2): Plant dry weight ( gm) at the end of the experiment as influenced by Five levels of crude oil in soil Mean Dry W eight (gm) Replications T reatments (Levels o f soil pollution) 4th 3d 2nd 1St 675.5 a 195.87 b 30.62 b 12.25 b 0.00 b 661 0.0 0.0 49.0 0.00 686 0.0 61.25 0.00 0.00 980 416.5 61.25 0.00 0.00 735.0 367.0 0.00 0.00 0.00 L0 L1 L2 L3 L4 0.0 refer to dead plant . Means with different sup erscrip ts are significan tly different (LSD) at p< 0.05 Plant Height (cm) Replications T reatments Mean 4th 3d 2nd 1St Levels of soil pollution 203.7a 79. 5b 36. 5 b 14.7 b 0.0 b 186.0 0.0 0.0 59.0 0.0 192.0 0.0 52.0 0.0 0.0 228.0 166.0 94.0 0.0 0.0 0 .209 152.0 0. 0 0.0 0.0 L0 L1 L2 L3 L4 IBN AL- HAITHAM J. FO R PURE & APPL. SC I VO L.22 (3) 2009 Fig (1): Eff ect of five levels of crud e oil on the growth of Olive seedling af ter six month from p ollution Fig (2): Effect of five levels of crude oil pollution on the percent of death of olive plan L0 L1 L2 L3 L4 100 75 50 50 Levels of crud e oil P ercent of Death (%) 2009) 3( 22المجلد للعلوم الصرفة والتطبیقیة مجلة ابن الھیثم Olea europaea نمو شتالت في النفط الخام ب التلوث من متزایدةتأثیر مستویات الزیتون شاكر عبد الوهاب حسن حسین علي و كونر،باسم عباس عبد علي جامعة بغداد ،مركز بحوث و متحف التاریخ الطبیعي الخالصة في المساحة التجربیة في 2005في اذار نمو شتالت الزیتون المزروعة في مختلفة من النفط الخام مستویاتتأثیر درس الى 2005من آذار المدة تحت الظروف الحقلیة) بغداد –باب المعظم (جامعة بغداد /مركز بحوث و متحف التاریخ الطبیعي ط الخام وهي فصممت التجربة الحقلیة على التصمیم العشوائي الكامل باستخدام خمسة مستویات من الن. 2008نیسان كل شتلة اذ ،أربعة مكررات عمالباست و اضیفت على سطح التربة )الشتلة / لتر نفط 3.0و 2.0، 1.0، 0.5 ، 0.0( ة الو ،عراض المرئیة التي ظهرت على النباتاتاأل ت تسجیل من التجربة شملالتي جمعت البیانات . مثلت مكرر نسبة المئوی كلي للنباتات ،و أطوال النباتات ،لموت النباتات أشارت النتائج إلى أن التلوث النفطي أدى إلى . بعد الحصاد و الوزن الجاف ال االصفرار على أعراضطي إلى ظهور فالتلوث الن أدى .ر سلبیة على نباتات الزیتون تناسبت مع مستویات التلوثاثآظهور نتیجة 0.05على مستوى احتمال ائیااحص الجافة هاأوزان النباتات و أطوال انخفضت . احیانا و الجفاف ثم موت النبات األوراق المستوى االعلى من التلوث ثم فياثر التلوث ظهر واضحا بعد ثالثة اسابیع الدراسة الى اننتائج اشارت . لتلوث النفطيل ت البحث ت نتائجكما أظهر .المستویات االدنى وذلك خالل االشهر الستة االولى امتد ظهور االعراض الىبعدها ان النباتا وث ها استطاعت ان تنجو ولكن و اً نمو كانت اقل القلیلة من النفط الخام مستویات الالمعاملة ب تبقى حیة في حین ادى التل كان واضحا ان النباتات التي تأثرت سلبیا بالتلوث استمر معها التأثیر السلبي ولم دوق. الى موت الشتالتبالمستویات العالیة .استمرار التجربةمدة ، شهرا من بدء التلوث النفطي 27تتعاف حتى بعد مرور