Distribution and activity of petroleum hydrocarbon degrading bacteria in the North Sea and Baltic Sea

Distribution and Activity of Petroleum Hydrocarbon Degrading Bacteria in the North Sea and Baltic Sea

Kai B r u n s , Gerhard D a h l m a n n andWilfried G u n k e l

UDC 547.912:576.8.095.32; North Sea; Baltic Sea

Summary

Data were collected in 1988 and 1989 on the distribution and activity of petroleum hydrocarbon degrading bacteria in the North Sea and Baltic Sea. Crude oil degrading bacteria and the number of bacteria which in particular degrade naphthalene were quantified using a modified dilution method (MPN). Crude oil degrading bacteria were present in all of about 100 water samples, with as many as 103 ml -~ in some samples. Numbers of naphthalene degrading bacteria were at least tenfold lower. There is obviously a greater connection between this bacteria group and petroleum hydrocarbon (PHC) contamination than between the more nonspecific group of crude oil degrading bacteria and PHC contamination. Data from the North Sea show an extremely high abundance of hydrocarbon degrading bacteria, even in winter, while in the southern Baltic Sea low numbers of bacteria were found and slower crude oil degradation was observed.

Verteilung und Aktivit~it von erdiilkohlenwasserstoffabbauenden Bakterien in Nord- und Ostsee (Zusammenfassung)

1988 und 1989 wurden Daten tiber Verteilung und Aktivit~it von erd61kohlenwas- serstoffabbanenden Bakterien in Nord- und Ostsee gesammelt. Roht~labbauende Bakte- rien und die Zahl yon Bakterien, die insbesondere Naphthalen abbauen, wurden mit einer modifizierten L6sungsmethode (MPN) bestimmt. Roh61abbanende Bakterien wurden in allen ca. 100 Wasserproben nachgewiesen; in einigen Proben mit nicht weniger als 103/ml. Die Zahl der naphthalenabbauenden Bakterien war mindestens um den Faktor 10 ldeiner. Ein st~kerer Zusammenhang zwischen dieser Bakteriengmppe und der Belastung des Meerwassers dutch Erd61-Kohlenwasserstoffe als zwischen der eher unspezifischen Gruppe der roh61abbauenden Bakterien und dieser Belastung ist wahrscheinlich. Daten aus der Nordsee zeigen, selbst im Winter, eine sehr hohe Zahl yon kohlenwasserstoffabbauenden Bakterien, w~arend in der sttdlichen Ostsee eine niedri- gere Anzahl von Bakterien bestimmt und ein langsamerer Roh61abbau beobachtet wurde.

1 Introduction

According to estimates by the National Research Council [1985], around 3.3 mio tons of petroleum hydrocarbons (PHC) enter the world's seas every year. As the North Sea and the Baltic Sea are relatively enclosed, marginal seas surrounded by densely populated and highly industrialized states, they are particularly affected. Annual input into the North Sea is estimated to be between 128 000 and 198 000 tons, while that for the Baltic Sea is estimated at between 20 300 and 66 200 tons (Tab. 1). The main sources are land runoff (33.6 to 88.7 %); shipping (1.8 to 23.5 %); and atmospheric input (4.9 to 15.1%).

360 Dr. Hydrogr. Z. 45, 1993, H. 6. B r u n s e t al., Petroleum hydrocarbon degrading bacteria

T a b l e 1 Estimated input of petroleum hydrocarbons into the North Sea and Baltic Sea (x 103 �9 t �9 a-l), modified according to 1)Institute of Offshore Engineering, 1984,

loc. cit. S i d e et al., 1985; z) E n c k e I l , 1987.

SOURCE N O R T H S E A 1) BALTIC SEA 2)

x 103 - t- a -1 x 103 - t- a 1

Naturalseeps 0 .3- 0.8 - Atmospheric 19 1 - 10

Rivers, land runoff (incl. inland municipal waste) 40 - 80 18 - 39 Coastal sewage discharges 3 - 14 no statement

Coastal refineries 6 0.2 Off terminal operations (incl. ballast water treatment) 0.8 0 .1- 0.2

Other coastal industrial effluent 9 0 .7 - 1.3 Accidental losses from tankers at sea 5 - 12 0 .2- 9

Operational discharges form tankers at sea and losses from general shipping 21.8- 33.8 0.16-6.5 Offshore production 23 < 0.005

Total: 127.9-198.4 20.3-66.2

According to these results, PHC concentrations in the North Sea and Baltic Sea should show a mean annual increase of 2.56 - 3.96 gg �9 1-1 and 1 - 3.3 I.tg. 1-1 respectively. Such an increase, however, is not detected because of the. degradation of PHCs in sea water by physical, chemical and biochemical processes (see summaries in National Research Council [1985]), A t l a s [1984], S a l o m o n s etal. [1988], G i b s o n [1984]).

The bacteria which play an essential role in biochemical degradation are restricted in their activity in the North Sea and Baltic Sea by relatively low temperatures, limited oxygen supply and limited availability of inorganic nitrogen and phosphorus. Recent results suggest that photooxidation is the more significant cause of PHC-degradation in the sea rather than bacteria (Tj e s s e m e t al., 1984), but this is yet to be proved. Examination of sea water has hitherto focused on the determination of PHC concentrations on the one hand and bacteria on the other (DHI [1980 to 1989], G u n k e l [1973]). Only few results are available from joint investigations ( G u n k e l et al. [19801).

The aim of our investigations in the North Sea and Baltic Sea was to ascertain the extent to which PHC concentrations can be correlated with the number of bacteria in the same water body in different areas. In addition, we also studied the degradative activity of naturally occurring bacterial communities at selected stations.

In January 1988, during a cruise with the RV Gauss around the UK, near-surface water samples were examined for PHC and PHC-degrading bacteria. A similar cruise aboard RV Valdivia was undertaken in the central Baltic Sea in April 1989 to collect near-surface and near-bottom samples. Samples taken at a station off the Gulf of Riga were incubated over several weeks so as to determine the extent of microbial degradation. Also in April 1989, the RV Friedrich Heincke left for the German Bight. The aim of this cruise was t o collect microbiological data and to investigate mineral oil degradation in nutrient gradients between the Elbe estuary and the open sea.

Dt. Hydrogr. Z. 45, 1993, H. 6. B r u n s e t al., Petroleum hydrocarbon degrading bacteria 361

2 Materials and Methods

2.1 P e t r o l e u m H y d r o c a r b o n s ( P H C )

Water samples were taken at a depth of 1 m using a 10 1 glass f a s k sampler rinsed with hexane (S t a d 1 e r and S c h o m a k e r [1977]) and extracted using n-hexane after the addition of an internal standard to the glass flask sampler. The extract was then dried with Na2SO 4 and concentrated in a rotary evaporator. The PHC concentration was determined using ultraviolet fluoresensce spectroscopy (Perkin-Elmer 650-10 S) as recommended by IGOSS (310nm excitation/360 nm emission). The PHC concentration is expressed in topped Ekofisk crude oil equivalents as g g . 1-1 (IOC [1984], D a h l m a n n and L a n g e [1981]). This method is used to screen affected areas and their degree of pollution (Anon. [1982]).

Biodegraded crude oils were separated by solid-phase extraction (Bond Elut Cl8-cartridges, Analytichem, Harbor City, USA) eluted with n-hexane/dichloromethane 1 : 1 (T h e o b a 1 d [1988]). The dried and concentrated extracts were analysed using glass capillary gas chromato- graphy (Sichromat 1, Siemens) with a flame ionisation detector and a fused silica column (25 m, coating: 5 % phenyl-methyl-silicone) as described by B r u n s e t al. [1989]). For the method used, deviations of individual measurements ranged from between 6 and a maximum of 15%.

2.2 B a c t e r i a

Water samples for the microbiological investigations were taken using 1 or 2 1 sterile bottles attached to modified Zobell samplers. In water > 100 m, sterile plastic bags and a bacteriological Niskin sampler were used (General Oceanics, Miami, USA). During the cruise around the UK, sub-samples were mainly taken from the glass flask sampler before the addition of the extraction medium and immediately examined.

The heterotrophic, saprophytic bacteria as colony-forming units (CFU) were ascertained using the pure-plate method in triplicate. 2216 E-M-Agar was used as the culture medium ( G u n k e l [1964]; R h e i n h e i m e r [1977]). Incubation in the dark at a temperature of 18 ~ lasted 21 days.

The number of PHC-degrading bacteria were quantified using the Most Probable Number method (MPN) in triplicate. The mineral salt medium according to G u n k e 1 and T r e k e 1 [1967] was modified by the addition of 0.5 ml �9 1-1 trace-metal solution according to C o h e n - B a z i r e et al. [1957]. This intensifies naphthalene decomposition (Bruns [1986]). 1 ml topped Ekofisk crude and diesel oil were added to 10 ml of liquid medium as the sole carbon source. Naphthalene-degrading bacteria were enriched in a medium containing naphthalene (70 mg in 10 mL medium) washed in A. dest. (B r u n s [1986]. The samples were incubated in the dark at a temperature of 18 ~ and their MPN was calculated using their turbidity values after 6 weeks (De M a n [1975]).

2.3 D e g r a d a t i v e E x p e r i m e n t s

Biodegradation experiments were carried out in 1 1 bottles in which 50 mg �9 1-1 topped Ekofisk crude were incubated with 0.5 1 of water sample. At the same time, samples were incubated after adding inorganic nitrogen (5 mg NHnC1 �9 1,31 mg �9 1-1 N), phosphorus (25 mg Na2PO 4 x 12 H20 and 5 mg KzHPO 4 �9 3,05 mg �9 1-1 P) and trace minerals to them (see above). The samples were lightly shaken or incubated in a vertical rotating apparatus at 18 ~ in the dark.

After a specified incubation period, a non-supplemented water sample and one with added inorganic nutrients were examined rnicrobiologically. The degradative activity in two doubled samples was halted by adding 10 ml �9 1-1 HC1 (10%) to each. These samples were then cooled and stored in the dark until processing in the laboratory.

362 Dt. Hydrogr. Z. 45, 1993, H. 6. B r u n s e t al., Petroleum hydrocarbon degrading bacteria

3 Results and Discussion

3.1 P H C c o n c e n t r a t i o n s a n d P H C - d e g r a d i n g b a c t e r i a in t h e N o r t h S e a a n d B a l t i c S e a

Figure 1 shows the location of sampling stations on the cruise around the UK in January/ February 1988. Colony-forming bacteria (2.3 to 2.2 x 104 CFU. m1-1) and mineral oil degrading bacteria (0.03 -4.3 x 102 N . ml -a) were found in' all the water samples. The number of naphthalene-degrading bacteria amounted to a maximum of 2.3 x 102 N . ml -l. This bacteria group could not be found at 15 from a total of 45 stations.

Both high PHC concentrations (1.40 to 5.39 gg �9 1-1) and high numbers of bacteria, partly consisting of a large proportion of PHC-degrading bacteria, were found in the German Bight. Similarly high values were measured as far as the Straight of Dover although the greatest PHC concentrations and maximum values for the bacteria group under investigation were found off the Rhine estuary. Bacteria nmnbers and PHC concentrations decreased in the English Channel towards the Atlantic except at coastal stations.

On entering St George's Channel, it was found that CFU and PHC concentrations increased due to the influence of the coast. Only in the Irish Sea were PHC concentrations and the proportion of PHC degrading bacteria considerably higher, with values similar to those in the German Bight: petroleum hydrocarbon-degrading bacteria amounted to a maximum of 7.03 x 103 CP-M, 9.3 x 101 per mL and naphthalene-degrading bacteria measured 9.3 with a maximum PHC concentration of 6.6 ~tg - 1 1.

Measurements taken in the Mersey estuary (Irish Sea) clearly showed the connection between PHC input and the number of bacteria. At the offshore station, a reduction of about 50 % was observed in PHC concentration (4.82 compared with 2.36 gg �9 1 -~) and the bacteria groups were diminished by a factor of 10 (CFU: 9.57 x 102 compared with 2.47 x 101; oil-MPN: 1.50 x 102 compared with 2.10 x 101; naphthalene-MPN: 2.3 compared with 0.23 N . ml-1). PHC concentration and the proportion of PHC-degrading bacteria also decreased in the North Channel towards the Atlantic (4.82 compared with 2.36 gg �9 1-1).

PHC concentrations and the number of bacteria measured in the German Bight, the Rhine and Thames estuaries and the Irish Sea show these to be more seriously contaminated areas. Data on the composition of bacterial communities indicate a) great degrading activity and b) chronic pollution. It was possible to prove the presence of e. g. naphthalene-degrading bacteria mostly only in water bodies with a PHC concentration no less than 1 ~tg �9 1-1. The less specific group of oil degraders was even found in water samples with a hydrocarbon concentration below 0.4 gg �9 1-1. The presence of naphthalene-degrading bacteria might therefore indicate a higher level of PHC pollution.

It even seems feasible to correlate the seasonal fluctuations in PHCs with the number of naphthalene-degrading bacteria present: in the German Bight, PHC concentrations in the summer, lower by a factor of 2 to 3 than in the winter, were found regularly (DHI [1980 to 89]). Near Helgoland, for instance, the number of naphthalene-degrading bacteria in the summer was 2.5 N . mk 1 ( B r u n s [1986]). On this winter cruise, however, it was found to be 230 N �9 m 1 - 1 .

Compared with the PHC concentrations in the North Sea, those found in the surface water (5 m) of the central Baltic in April 1989 were low (Fig. 2). They were evenly distributed and ranged from 1.07 - 2.95 gg - 1-1 in the surface water. This can be attributed to the relatively high atmospheric input of PHCs into the North Sea. Higher PHC concentrations, 0.63 - 5.88 gg �9 k I were measured 10 m above the sea floor.

CFU and mineral-oil-degrading bacteria were detected in all the water samples, but in considerably fewer numbers than in the North Sea. Values [N �9 m1-1] for saprophytic bacteria (CFU) were between 7 and 4.9 x 102; for oil-degrading bacteria they were between 0.091 and 2.3 x 10~; and for naphthalene-degrading bacteria they lay between 0.023 and 2.3 x 1011 (Fig. 2).

Saprophytic and oil-degrading bacteria were found in all water samples. Slight increases in both bacteria groups were observed in the Gulf of Gdansk, along the west coast to the Gulf of

Dt. Hydrogr+ Z. 45, 1993, H. 6. B r u n s e t al., Pe t ro leum hydrocarbon degrading bacteria 363

~ +7+,7,71 ~ �9 " r+ +.,~ N

_ _ o "6 i l ~I+"o

++ "~ W{ ~+ 1,0 o

N + + ~2

++ I+ I - . . ~ ~ ~ .

o

o

~ ~ 0

o ~ ,4m

0

ff~ 113 ~ I/3 u3 i/~ ,~- ~

.:.+ ,_:

~ o +.., : D < u o ~,=,~

u m

~ ~ ...-

" ~ O0 , . 0

- ~ ~ ~

; N


1 5 ~ 1 6 o 1 7 ~ 1 8 o 1 9 ~ 2 0 ~ 2 1 o 2 2 ~ 2 3 ~ 2 4 ~

5 6 ~

5 3 0 . . . . . . . . . . . . . . . . . . . . . . . . . . . , , , , ~ . . . . . . . , . . . . . . . . . . ..-.~=-.~ ..=..~.. 5 3 0

1 5 ~ 1 6 ~ 1 7 ~ 1 8 o 1 9 o 2 0 0 2 1 ~ 2 2 ~ 2 3 ~ 2 4 ~

Fig. 2 Distribution of CFU (left bar: [~) , crude oil (intermediate bar: [ ] ) and naphthalene-degrading bacteria (right bar: �9 ) as [lg N - mF 1] as well as PHC concentrations [~tg - 12] (below the plotted bars) in the central Baltic (RV "Valdivia", 30. 3. 1989 - 15. 4. 1989) at a depth of 5 m. The arrow indicates the station where samples were taken for biodegradation experiments at a depth

o f l m.

Dt. Hydrogr. Z. 45, 1993, H. 6. B r u n s et al., Petroleum hydrocarbon degrading bacteria 365

Finland as well as to the west of Oland mad Bornholm. Where conditions differed from those in the North Sea was in the small proportion of bacteria solely able to degrade polycyclic naphthalene. This group was traceable in only 20 out of 24 samples. In addition, their number was mostly at the lower detection limit (0.023 N �9 m1-1) except for a station to the west of Bornholm and two stations in the northern Baltic Proper (23 N �9 ml 1).

3.2 D e g r a d a t i v e A c t i v i t y in t h e N o r t h S e a a n d B a l t i c S e a

Bacteria numbers alone are not sufficient to evaluate the degradative activity of a bacterial community. Features of bacteria are their rate of increase and effective selection mechanisms which cause a bacterial community to adapt to local conditions.

Over a period of several weeks, oil-degrading experiments were carried out to determine the extent of bacterial oil degradation; 0.5 1 water samples were incubated with 50 mg �9 1-1 Ekofisk crude. Samples from one station at the latitude of the Gulf of Riga (see arrow in Fig. 2) were incubated for 84 days, for comparison. The number of bacteria of all the groups examined increased in three days by a factor of up to 10 000 (Fig. 3), as had been repeatedly observed in the case of North Sea bacteria ( B r u n s et al. [1989], G u n k e l and D a h l m a n n [1987]). Within 14 days of incubation, each of the bacteria groups reached or had already exceeded maximum values. As in the case of North Sea bacteria, the addition of inorganic nitrogen, phosphorus and trace elements resulted in faster and increased growth of CFU and mineral-oil- degrading bacteria without the number of naphthalene-degrading bacteria being greatly influenced.

Bacteria [lg N mL-1] 11 lO 9 8 7 6 ~ . . . . . . . . . . . , -':Y-Y2.-"l . . . . . . . . . . . .

3 2 1

0 i I I

11

w i t h o u t i n o r g a n i c s 10 9 8 7 6 5 4 3 2 1 0

10 w i t h i n o r g a n i c sa l t s - lo 9 9 8 8 6 y -,~ - 6

- . . . . . . �9 . . . . . . . . . . . . . . . . . . " . . . . . . . . . A. 5 4 r "4 3 3 2 2 1 -1 0 I t I I 0

0 10 20 30 40 Incubation period [d]

Fig. 3 Microbial crude oil degradation in the Baltic Sea growth of bacterial community [lg N �9 ml 1] (CFU: - 0 - ; oil-MPN: -am-; naphthalene-MPN: - i ' ) taken at a depth of 1 m during incubation with 50 mg �9 1-1 Ekofisk oil in pure seawater (left) and with additionally available

inorganic nitrogen and phosphorus (right).


The extent of mineral oil degradation was different, however. The natural nitrogen and phosphorus concentrations in the sample water from the Baltic amounted to 8.21 gmol �9 1-1 and 0.55 /.tmol �9 k 1. It was therefore to be expected that the mineral oil degradation in the non-supplemented incubation samples would be slight. Indeed, the biodegradation led merely to the incomplete degradation of alkanes with a chain-length < C17. Unlike previous experiments off Helgoland, what was surprising here was that at first hardly any intensification in degradative activity was observed when nitrogen and phosphorus were additionally available. The concentration of the alkanes with a carbon chain length > C15 decreased only after 28 days' incubation; and the branched chain alkane pristane remained unaffected even after 84 days' incubation.

Nevertheless, not all the findings were consistent: the results of the mineral oil incubation tests are subject to deviations that are due to the incubation method used. For each incubation period, separate water samples containing mineral oil were incubated and examined in microbiological and chemical analyses. Deviations up to a maximum of 25 % can thus be attributed to water sample inhomogeneities such as, for instance, differences in the numbers of bacteria or protozoa in the samples at the start. The "grazing" of protozoa has a crucial influence on the selection of highly degradative bacteria (F e n c h e 1 [1982]).

In the same month, the extent of mineral oil degradation was investigated at 10 stations in the German Bight. See Fig. 4 for the positions of the stations and the composition of the bacterial communities at a depth of lm. Values for saprophytic bacteria were between 2.3 and 1.46 x 1 0 4 C F U �9 m l - 1 ; for crude-oil-degrading bacteria they were between 0.36 and 1.5 x 1011; for naphthalene-degrading bacteria they were 0.36 and 2.3 N . ml -~. The influence of the Elbe is clearly seen in the large number of CFU and mineral-oil-degrading bacteria. There was a general decrease in the size of bacterial communities as the distance from eutrophic water increased. The proportion of bacteria specializing in alternative food sources (oil MPN) increased in relation to heterotrophic bacteria. Polycyclic PHCs and similar substances did not presumably play an essential part in the modified food supply as the number of naphthalene-degrading bacteria in the open sea was mostly at the lower detection limit.

Like the Baltic Sea bacteria, these bacterial communities were incubated with 50 mg�9 1-1 Ekofisk crude. Two weeks later, chemical analysis of the degraded oils showed that mainly alkanes had been degraded. In the non-supplemented incubations from stations 1 to 7, a correlation between the extent of alkane degradation and nitrogen and phosphorus concentrations in the sea water could clearly be identified (Fig. 5). Two weeks later, the alkanes in the immediate spheres of influence of the rivers Elbe and Weser (stations 1 and 2 and station 4 respectively) had been reduced by between 70 and 90 %. Degradation off Helgoland (station 3) only reached 30 to 50%, while in the open sea (station 7), the degradation was observed of only Clo and Cll alkanes. In contrast to the results of the Baltic Sea investigations, a more intensive degradative process was detected when nitrogen and phosphorus were added. Pristane, known to be more resistant to degradation, was also affected.

The amount of the added phosphorus and nitrogen was greater than the natural concentration by a factor of 1000. Despite this, the extent of degradation of mineral oil at offshore stations was considerably smaller. It is now believed that microbial mineral oil degradation is not only limited by the availability of nitrogen and phosphorus, as has been assumed until now, but that other, previously unknown factors also play a role. Which factors these are is still a matter of conjecture.

Earlier investigations showed that the order of magnitude and composition of the number of bacteria are almost the same within 3 to 7 days' incubation. This is why little importance should be attached to the size and composition of bacterial communities. The possible preadaptation of bacterial communities in chronically polluted near-coastal areas should be considered as a factor (H u d a k et al. [1988]) or the presence of protozoa which are responsible for selecting active and rapidly growing bacteria ("microbial loop", F e n c h e 1 [1982]).


5 6 "

5 5 "

54. =

3 ~ 4 .~ 5 ~ 6 ~ 7 " B* 9 " O ~

BACTERIA

[Ig N mL-1] 2

5 6 "

55*

54-"

5.3" �9 " . . . . . 5,.3* 3* 4 ~ 5 ~ 6* 7 ~ B* 9 ~ 10 ~

Fig. 4 Distribution of CFU (left bar: [ ] ) , crude oil (intermediate bar: [ ] ) and naphthalene-degrading bacteria (right bar: �9 ) as [lg N �9 ml 1] in the German Bight (RV "Friedrich Heincke",

24. 4. - 28.4. 1989) at a depth of 1 m.

N [umol 1--1] P [umol L - l ]

100 3

80 J l ' ~ N ( t o t a l ~ L ~ ~' 2 2,5 6O

1,5 )

40 ~'.,.. ~ 1 20 P04- P ~ " - - " " ~ ..~x~.

0,5 " ' - 'h ' ' ' ' -~ . . . . . . . . . .k

0 , , I l , , l , l T 0

1 2 3 4 5 6 7 8 9 10

Stat ion

Fig. 5 Concen t r a t i on o f n i t rogen (left b a r : - ~ - ) and p h o s p h o r u s ( in te rmedia te b a r : - ' ~ - ) [pmol �9 1 1] in the G e r m a n B i gh t in Apr i l 1989 (for the pos i t ion o f the s a m p l i n g s ta t ions cf. Fig. 5).

368 Dr. Hydrogr. Z. 45, 1993, H. 6. B r u n s e t al., Petroleum hydrocarbon degrading bacteria

4 Conclusions

Simultaneous estimates of PHC concentrations and degrading bacteria highlight the ecological stress and the biological reaction of the system. This does not mean, however, that a direct connection can be expected to exist between bacteria numbers and PHC concentration, as comments on the physiological characteristics of a bacterial community cannot be based on high numbers of bacteria alone (compare H u d a k et al. [1988]). Comparison of PHC concentrations and bacteria numbers is further limited by the use of fluorescence spectroscopy in that the method can only indicate polluted areas. Despite this limitation, oil degraders are present in all the water samples, but the ability to degrade naphthalene in unpolluted water does not seem to be widespread. Naphthalene-degrading bacteria are, however, always present in more polluted water. The riverine influx of PHC results in the continuous presence of naphthalene-degrading bacteria in estuaries and causes greater degradative activity which is due to sufficient concentrations of phosphorus and nitrogen.

The negligible degradative activity found in the Baltic is based on only one investigation whose results may not be applied to the whole Baltic Sea area. While our results agree with those by T s y b a n and I z r a e l , as shown in a review by G o c k e et al. [1990], more investigations are necessary in areas directly influenced by the coast to enable comparison of microbial degradative potential in the North Sea and Baltic Sea.

It was found that intensive microbial oil degradation in the German Bight only occurs very near the coast because of the nitrogen and phosphorus levels there. Bacteria in the open sea are also able to degrade PHCs, given sufficient nitrogen and phosphorus. Their level of activity is not as high as bacteria in coastal waters, however. This indicates that other, as yet unknown factors influence microbial mineral oil degradation in the open sea.

Our results show that the percentage of microorganisms involved in PHC degradation in the open sea is very small. Photooxidation as discussed by T j e s s e m et al. [1984] may indeed be more important in the degradation of PHCs than microbial degradative processes.

A c k n o w l e d g e m e n t s

This paper was originally presented at the 4th European marine microbiology symposium in Kiel, FRG in 1990. It forms part of the Ph.D. thesis on ,,Mikrobieller Abbau von Mineralt len im Meer, Abbauaktivitat und Ver~inderungen in der Zusammensetzung der Rest t le" at Justus- Liebig-University, Giessen, FRG. Work on the thesis was undertaken at the Bundesamt for Seeschiffahrt und Hydrographic in Hamburg and at the Biologische Anstalt Helgoland on Helgoland.

The oceanographic data were supplied by Mr. Mangelsdorf and Mr. Treutner of the Biologische Anstalt Helgoland, to whom sincere thanks are due.

References

Anonymus, 1982: Proceedings of a workshop on the analysis of hydrocarbons in seawater. March 23 - April 3, 1981. Baltic Sea Envi- ronment Proceedings No. 6. Helsinki Com- mission, Helsinki.

A t l a s , R. M. (ed.), 1984: Petroleum Micro- biology. New York: Macmillian Publishing Company.

B r u n s, K, 1986: Einfltisse von Naphthalen auf marine Bakterien. Master Thesis, Justus- Liebig-Universit~it Giessen.

B r u n s , K. ,W. G u n k e l , G. D a h l m a n n und N. T h e o b al d, 1989: Biologischer Ab- bau von Erdtlkohlenwasserstoffen in der

Nordsee unter besonderer Berficksichtigung yon schwer abbaubaren Komponenten, die als Leitsubstanzen fur die chemisch analytische U-berwachung der Meeresumwelt dienen sol- len. Wiss. Techn. Ber. 1989-3. Deutsches Hy- drographisches Institut, Hamburg.

C o h e n - B a z i r e , G . ,W.R . S i s t r o m and R. Y. S t a n i e r , 1957: Kinetics studies of pigment synthesis by non-sulphur purple bacteria. J. Cell. Comp. Physiol. 49, 25-68.

D a h l m a n n , G. andW. L a n g e , 1981: In- vestigation of the distribution of petroleum hydrocarbons in the German Bight by means of fluorescence spectroscopy. Dt. hydrogr. Z. 34, 150-161.


DHI, 1982-1991: l)-berwachung des Meeres. Bericht fiir das Jahr 1980-1989. Deutsches Hydrographisches Institut, Hamburg.

E n c k e l l , E., 1987: Oil pollution load on the Baltic Sea; a compilation of measured and estimated load. Baltic Sea Envrion. Proc. No. 22, 2-9.

F e n c h e 1, T., 1982: Ecology of heterotrophic microflagellates IV. Quantitative occurence and importance as bacterial consumers. Mar. Ecol. Prog. Ser. 9, 35-42.

G i b s o n , D. T. (ed.), 1984: Microbial degradation of organic compounds. New York: Marcel Dekker.

G o c k e , K., et al., 1990: Second periodic assessment of the state of the marine environment of the Baltic Sea, 1984-1988; General conclusions., 7. Microorganisms. In: Baltic Marine Environment Protection Commission - Helsinki Commission - (ed.), Baltic Sea Environment Proceedings No. 35 A.

G u n k e l , W., 1964: Einwirkungen des kalten Winters 1962/63 auf die Bakterienpopulatio- nen vor Helgoland. Helgolander wiss. Mee- resunters. 10, 246-256.

G u n k e l , W. and G. D a h l m a n n , 1 9 8 7 : Bacterial degradation of heavy fuel oil in seawater. Baltic Sea Environ. Proc. 22, 68-80.

G u n k e l , W. 1973: Distribution and abundance of oil-oxidizing bacteria in the North Sea, p. 127-139. In: D. G. Adhearn and S. E Meyers (ed.), The microbial degradation of oil pollutants. Louisiana State University, Ba- ton Rouge.

G u n k e l , W. undH. H. T r e k e l , 1967: Zur Methodik der quantitativen Erfassung 61ab- banender Bakterien in verrlten Sedimenten und Brden, Ol-Wassergemischen, Olen und teerartigen Substanzen. Helgol~inder. wiss. Meeresunters. 16, 336-348.

G u n k e 1, W., et al., 1980: Preliminary results of baseline studies of hydrocarbons and bacteria in the North Sea: 1975, 1976 and 1977, p. 223-247 Ponencias del simposio internatio-

nalen Resistencia a los antibioticos y micro- biologia marina. Santiago de Compostela.

H u d a k , J. R und J. A. F u h r m a n , 1988: Effects of four organic pollutants on the growth of natural marine bacterioplankton populations. Mar. Ecol. Prog. Ser. 47, 185-194.

Intergovernmental Oceanographic Commission, 1984: Manual for monitoring oil and dissol- ved/dispersed petroleum hydrocarbons in marine waters and on beaches. Manuals and Guides No. 13. IOC/UNER Paris.

de M a n , J. C., 1975: The probability of most probable number. Europ. J. Appl. Microbiol. 1, No. 67-78.

National Research Council (ed.), 1985: Oil in the sea, inputs, fates, and effects. Washington D. C.: National Academy Press.

R h e i n h e i m e r , G., 1977: Microbial ecology of a brackish water environment. Berlin: Springer.

S a l o m o n s , W. , B. L. B a y n e , E. K. D u u r s m a und U. F r r s t e r (eds.), 1988: Pollution of the North Sea - An assessment. Berlin: Springer.

S i d e , J . , Ch . H e r d u n d W. G r o g a n , 1985: Oil spill pollution: The North Sea expe- rience of cooperative measures, p. 615-620. In: American Petroleum Institute (ed.), Proc. 1985 Oil spill conference. American Petro- leum Institute, Washington D.C.

S t a d l e r , D. a n d K. S c h o m a k e r , 1977: Ein Glaskugelsch6pfer zur kontaminations- freien Entnahme von Seewasser unter der Oberfl~iche ftir die Analyse von Kohlenwas- serstoffen und halogenierten Kohlenwasser- stoffen. Dt. Hydrogr. Z. 30, 20-25.

T h e o b a l d , N., 1988: Rapid preliminary se- paration of petroleum hydrocarbons by solid-phase extraction cartridges. Anal. Chim. Acta 204, 135-144.

T j e s s e m , K. andK. H. P a l m o r k , 1984: An overview of auto/photooxidation of petroleum in the marine environment. ICES, Ma- rine Chemistry Working Group, p. 1-25.

Submitted: 20. 6. 1993 Accepted: 17. 12. 1993

Authors' addresses:

Kai Bruns* Mozartstrage 3 35463 Fernwald 1

Dr. G. Dahlmann Bundesamt far Seeschiffahrt und Hydrographie Postfach 30 12 20 20305 Hamburg

* Author to whom correspondence should be addressed

Prof. Dr. W. Gunkel Biologische Anstalt Helgoland NotkestraBe 31 22607 Hamburg

Documents

Distribution and activity of petroleum hydrocarbon degrading bacteria in the North Sea and Baltic Sea