African Health Sciences Makerere University Medical School ISSN: 1680-6905 EISSN: 1729-0503 Vol. 8, Num. 2, 2008, pp. 85-89

African Health Sciences, Vol. 8, No. 2, June, 2008, pp. 85-89

Bacterial indicators of pollution of the Douala lagoon, Cameroon: Public health implications

Jane-Francis T K Akoachere¹, Pius M Oben², Beryl S Mbivnjo¹, Lucy M Ndip¹, Gerald Nkwelang¹, Roland N Ndip^1,3.

¹Departments of Biochemistry and Microbiology, and ²Geology and Environmental Science, Faculty of Science, University of Buea, Box 63, Buea, Cameroon; ³Department of Biochemistry and Microbiology, Faculty of Science and Agriculture, University of Fort Hare, P/Bag X1314, Alice 5700, South Africa.
Author for correspondence: Roland N. Ndip^1,3.¹Departments of Biochemistry and Microbiology, ³Department of Biochemistry and Microbiology, Faculty of Science and Agriculture, University of Fort Hare, P/Bag X1314, Alice 5700, South Africa. Email: rndip@ufh.ac.za

Code Number: hs08020

Abstract

Background: Indiscriminate disposal of untreated wastes which are often heavily laden with sewage microorganisms some of which are pathogenic to humans into aquatic environments near cities could serve as potential dangers to human health.
Objective: A prospective study was undertaken to investigate the scope of potential bacterial pathogens and to assess the extent of pollution of the Douala lagoon.
Methods: A total of eighty water samples were collected fortnightly from the lagoon at five stations from March to October 2005 and analysed for heterotrophic bacterial densities, coliform counts, faecal coliform and faecal streptococcal counts. Bacteria were isolated and identified using standard microbiology and biochemical techniques.
Results: High heterotrophic bacterial counts (33 × 10⁵ 161 ×10⁵ CFU/ mL), total coliform counts (1.8 ×10² - 2.4 ×10² CFU/ 100 mL), faecal coliform counts (2.2 × 10² 2.4 × 10² CFU/ 100 mL) and faecal streptococcal counts (2.1 × 10² 2.3 x 10² CFU/ 100 mL) were observed in all sampling stations. Eleven species of bacteria: Bacteroides fragilis, Proteus vulgaris, Klebsiella pneumoniae, E. coli, Enterococcus faecalis, Enterobacter aerogenes, Citrobacter freundii, Aeromonas hydrophila, Pseudomonas aeruginosa, Bacillus mycoides and Serratia marcesens, were frequently isolated.
Conclusion: The presence of potential bacterial agents such as Bacteroides fragilis, Pseudomonas aeruginosa, Aeromonas hydrophila, Klebsiella pneumoniae and E. coli in the lagoon may pose a serious threat to the health and well being of users of the Lagoon and calls for urgent intervention.

Key words: Bacteria; Pathogenic; Coliform; Heterotrophic; Lagoon; Cameroon.

Introduction

Aquatic environments near cities are usually prone to overloading with a variety of pollutants either through direct or indirect discharges. This situation may be worsened by the indiscriminate disposal of untreated wastes, which are often heavily laden with sewage into actively used waters. Sewage polluted waters carry sewage microorganisms, some of which are pathogenic to humans^1-4.

Several researchers ^5-11 have documented the public health significance of faecal pollution of natural waters. Apart from health concerns, the indiscriminate dumping of untreated wastes into aquatic environments brings about physical, chemical and biological deteriorations of such water bodies when these discharges are beyond their self-purifying capacity^12-14. This will no doubt endanger the resident aquatic organisms as well as impair the beneficial uses of the water ^{6, 15, 16}.

The Douala lagoon, like many coastal lagoons, serves as a seaport, centre for recreational sailing and a sink for disposal of domestic and industrial wastes. A paucity of information exists on the extent of pollution of the lagoon and hence its public health implication. Ekane and Oben¹⁷ investigated the biochemical characteristics of the Douala lagoon and Limbe estuary and reported a great impact of human activities on these parameters. Oben and Oben¹⁸ in a plankton survey of the lagoon reported a high rate of mortality of some of these organisms and associated their findings to the influence of discharges from food and chemical industry. It is apparent that with the high level of waste disposal and other forms of contamination, the lagoon would likely represent a reservoir of potential bacterial and other pathogens. It is against this background that the present study was initiated since we are not aware of any investigation on the bacteriology of the lagoon. This study therefore appraises the level of bacterial contamination, the sanitary quality and potential health risk of the lagoon to humans.

Materials and methods

Study Site

The Douala lagoon is part of the coastal plain of Cameroon, which stretches from Rio-Del-Rey through Victoria, Tiko, Douala and Kribi down to Equatorial Guinea. The area experiences an ever-increasing population and industries (chemical, food processing, textile and petroleum) flank the lagoon.

Surface water samples (n= 80) were collected fortnightly for a period of eight months (March to October, 2005) using standard methods¹⁹. Quadruplet composite samples were collected into sterile 300 mL plastic bottles from the following points receiving either domestic or industrial wastes: station 1, behind an engineering company; station 2, after an effluent discharge point from the petroleum depot; station 3, an open space with no trees with recreational activities predominating; station 4, receives waste from a chemical company, also serves as a dump site for domestic wastes and, station 5, after the Wouri bridge, serves as dump site for domestic wastes (Fig.1) .

Samples were serially diluted. Total viable heterotrophic bacterial counts were obtained by the standard plate count (SPC) method on nutrient agar. Volumes of 0.1mL were plated in triplicate and incubated at 35^oC for 48 hours¹⁹.

Enumeration of Indicator Organisms

Standard methods were employed for the isolation and enumeration of indicator organisms¹⁹. Three-tube, 10-fold dilutions of McConkey broth were used to determine the most probable number (MPN) of total coliforms. For total coliforms, fermentation tubes (containing Durham tubes) in a 3-3-3-tube regimen were incubated at 37^oC for 24 hours. For faecal coliforms, incubation was at 44.5^oC for 24 hours. Faecal streptococci counts were obtained by inoculating samples into Hannay and Norton's sodium azide broth in similar regimen as for total and faecal coliforms and incubating at 37 ^oC for 72 hours². Counts (MPN) were estimated from probability tables¹⁹.

Identification of Isolates

Bacteria were identified based on colonial characteristics on nutrient and McConkey agar, Gram coloration, motility and biochemical reactions using previously established schemes^20-22. Isolates were confirmed using the API 20E (Biomerieux SA, France) kit according to the manufacturer's instruction.

Statistical analysis

Variations in the parameters analyzed between the sampling stations and sampling period were compared using the two-way analysis of variance test. P values <0.05 were considered significant.

Results

Heterotrophic bacterial counts, ranging from 33×10 ⁵ CFU/mL in station 2 to 161×10⁵ CFU/mL in station 3 were recorded in the lagoon (Fig. 2)_. The counts in station 3 were significantly higher (P<0.05) than in other stations. With respect to sampling period, counts were significantly higher (P<0.05) in the wet months (May to October) than in the dry months (March to April).

Also, very high coliform counts (1.8×10² 2.4×10² CFU/100mL), faecal coliform counts (2.2 ×10² — 2.4×10² CFU/100mL) and faecal streptococcal counts (2.1×10² 2.3×10² CFU/100mL (Fig.3) were observed throughout the study period. Counts though high reflected no significant differences (P>0.05) between the stations.

Eleven bacterial species were isolated from the lagoon (Table 1). The majority of these were enteric organisms: E.coli (100%), Klebsiella pneumoniae (100%), Proteus vulgaris (100%) and Enterobacter aerogenes (95%) predominating. Other gram-negative rods included Bacteroides fragilis (100%), Aeromonas hydrophila (50%), Citrobacter freundii (50%), Pseudomonas aeruginosa (45%) and Serratia marcescens (15%). Prominent among the gram-positive genera were Enterococcus faecalis (100%) and Bacillus mycoides (35%). Of these isolates, B. fragilis, P. vulgaris, K. pneumoniae, E. coli and E. faecalis were the most predominant, being isolated from all samples.

Discussion

Heterotrophic bacterial counts ranging from 33×10⁵ CFU/mL to 161×10⁵ CFU/mL were recorded in the lagoon. These counts were however significantly higher (P<0.05) in the wet months (May to October) than in the dry months (March to April). These high counts could be due to high levels of organic matter present in the lagoon as a result of indiscriminate dumping of wastes, and are in agreement with the findings of Tatah and Ikenebomeh²³ who in a similar study in Nigeria attributed such high counts to the high organic matter content of Ikpoba River. Fleisher et al ⁷ and Sequel et al²⁴ equally reported similar findings. During a rainfall, run off water washes organic matter as well as bacteria¹². We therefore speculate that this may also explain the significantly higher counts obtained in the wet months during which period run off get into the lagoon. Bacteria form a link between primary producers and consumers; it would therefore appear that pollution affects the aquatic food chain. The high bacterial counts are indicative of the sanitary status of the lagoon. It was also of interest to note the very high total coliform counts (1.8x10² 2.4x10² CFU/ 100mL), faecal coliform counts (2.2 x10² - 2.4x10² CFU/ 100mL) and faecal streptococcal counts (2.1x10² 2.3x10² CFU/ 100mL throughout the study period. Such counts are highly undesirable and indicate severe faecal pollution of the lagoon, hence the possible presence of human pathogens.

Eleven bacterial species were isolated from the lagoon. The majority of these were enteric organisms, with members of the Enterobacteriaceae (E. coli, Enterobacter aerogenes, Klebsiella pneumoniae, Proteus ulgaris) predominating. Of these isolates, B. fragilis, P. vulgaris, K. pneumoniae, E. coli and E. faecalis were the most predominant, being isolated from all samples. This is consistent with the findings of other investigators ^{1, 2, 12, 25, 26} who isolated these organisms from water bodies heavily contaminated with faecal material. The majority of our isolates are human pathogens. The lagoon is fed by polluted swamps and drainage ditches carrying faecal pollution from neighbouring densely crowded residential areas which have inadequate sanitary facilities. The uncontrolled urbanization around the swampy areas surrounding the lagoon harbours mostly the poor city dwellers who do not have access to clean water or basic sanitary

facilities. As waste disposal facilities are grossly inadequate to provide complete evacuation of both solid and liquid wastes, these inhabitants resort to use of the lagoon as an "infinite sink" for their wastes. The high incidence of human pathogenic bacteria in the lagoon may indicate their possible presence in fish and other foods obtained therein. This lays credence to the fact that other researchers^{26, 27} have isolated human pathogenic bacteria from aquatic foods from polluted waters. Though the lagoon is not used for drinking, it is of value for fishing and recreation thereby rendering it dangerous to the health of the population. Although cholera is endemic in Douala with some of the contributing factors to its endemicity being the location of the city of Douala at the mouth of the Wouri delta, presence of vast expanses of swamp, streams/drainage ditches infested with algae²⁸, we did not isolate Vibrio cholerae from samples during this study. Recent studies^29-31 have identified conditionally viable environmental cells of pathogenic Vibrio cholerae that resist cultivation by conventional techniques existing in surface waters as biofilms of partially dormant cells. This may explain the absence of this organism from our samples.

Conclusion

Based on our findings, it is likely that the lagoon could pose a serious threat to the health and well being of users and calls for urgent intervention. We therefore recommend that adequate waste disposal facilities should be provided to inhabitants of the Douala Municipality to prevent indiscriminate dumping of wastes into the lagoon; the Environmental Protection Agency should ensure that industries treat their wastes adequately before disposal into the lagoon and; the public should be educated on the health risks of indiscriminate disposal of wastes.

Acknowledgement

We thank the University of Buea for providing the equipment and some reagents used in this study under the Staff Development Grant Scheme.

References

1. Lipp EK, Farrah SA, and Rose JB. Assessment and impact of microbial fecal pollution and human enteric pathogen in a coastal community. Maritime Pollut Bull 2001; 42(4): 286 293.
2. Melo MT, Vieira RH, Saker-Sampaio S. and Hofer I.. Coliforms and Salmonella in Seawater near to domestic sewage sources in Fortaliza Cearca, Brazil. Microbiologia. 1997; 13(4): 463 470.
3. Olayemi AB. Bacteriological water assessment of an urban river in Nigeria. Intl J Environ Health Res. 1994; 4: 156-164.
4. Ogbodeminu FS. Effects of domestic and industrial wastes on the bacteriological quality of Kaduna River, Nigeria. Federal Ministry of Science and Technology. Lake Kanji Research institute Annual Report. 1986; ISSN 0331-9296.
5. Ekundayo JA. Environmental consequences of the pollution of Lagos Lagoon. Bull Sci Asso Nig. 1977; 3: 290-299.
6. Fleisher JM, Kay D, Wyer MD. and Godfree AF. Estimates of the severity of illnesses associated with bathing in marine recreational waters contaminated with domestic sewage. Intl J Epidemiol. 1998; 27(4): 722 726.
7. Fleisher JM, Kay D, Salmon RL, Jones F, Wyer MD, and Godfree AF. Marine waters contaminated with domestic sewage: non enteric illnesses associated with bather exposure in the UK. Amer J Pub Health. 1996; 86(9): 1228-1234.
8. George I, Anzil A, and Servais P. Quantification of fecal coliform inputs to aquatic systems through soil leaching. Water Res. 2004; 38(3): 611-618.
9. Obi CL, Potgieter N, Bessong PO, and Matsuang G. Scope of potential bacterial agents of diarrhea and microbial assessment of quality of river water sources in rural venda communities of SA. Water Sci Technol. 2003; 47(3):59-64.
10. Okafo CN, Umoh VJ, and Galadima M. Occurrence of pathogens on vegetables harvested from soils irrigated with contaminated streams. Sci Total Environ. 2003; 311(1-3):49-56.
11. Olyphant GA, Thomas J, Whitman RL, and Harper D. Characterization and statistical modeling of bacterial (Escherichia coli) outflows from watersheds that discharge into Southern Lake Michigari. Environ Monit Assess 2003; 81(1-3): 289-300.
12. Fujioka RS. Monitoring coastal marine waters for spore forming bacteria of fecal and soil origin to determine point from non-point source pollution. Water Sci Technol. 2001; 44(7): 181-188.
13. Horvat M, Covelli S, Faganeli J, Logar M, Mardic V, Rajar R, Sirca A, and Zagar D. Mercury in contaminated coastal environments, a case study: the Gulf of Trieste. Sci Total. Environ 1999; 237-238:43-56.
14. Sweeney A, and Sanudo Wilhelmy SA. Dissolved metal contamination in the East River-long Island Sound System: potential biological effects. Marine Pollut Bull. 2004; 48(7-8): 663-670.
15. Daby D, Turner J, and Jago C. Microbial and nutrient pollution of coastal bathing waters in Mauritius. Environ Intl. 2002; 27(7): 555 566.
16. Parsons EC. Trace metal pollution in Hong Kong: Implications for the Health of Hong Kong's Indo-pacific hump-backed dolphins (Souza Chinensis). Sci Total Environ. 1998; 214: 175-184.
17. Ekane ND, and Oben PM. Biochemical indicators of marine pollution of the Douala lagoon and Limbe estuary. In: Environmental Issues: Problems and Prospects. Lambi, C.M. (ed). Environmental issues: problems and prospects. Unique, Bamenda. 2001; p 119 132.
18. Oben PM, and Oben BO. Plankton Survey in National Waters of Cameroon Large Marine Ecosystem of Gulf of Guinea (LME-GOG). Prepared for the United Nations Development Organisation (UNIDO). 1999; pp. 20
19. American Public Health Association (APHA), American Water Works Association (AWWA) and Water Pollution Control Federation (WPCF). Standard Methods for the Examination of Water and Wastewater. 1989; pp.1193
20. Buchanan RE, Gibbons NE. Gram-negative faculttively anaerobic rods. In:Bergey's Manual of Determinative Bacteriology.8^th edition. Williams and Wilkins Co; Baltimore, USA. . 1974; p. 125 - 142
21. Cheesbrough M. Biochemical tests to identify bacteria. In: District Laboratory Practice in Tropical Countries Part II. Cambridge University Press. 2000; p. 206 - 224
22. Cowan ST, and Steel KJ Manual for the Identification of Medical Bacteria. 4^th edition. Cambridge University Press, London.1985; pp. 265
23. Tatah JFK and Ikenebomeh MJ. Influence of brewery effluent on heterotrophic counts and some physical parameters of Ikpoba Tropical River, Nigeria. Nig J Microbiol. 1999; 13: 55-58.
24. Sequel CG, Mudge SM, Salgado C, and Toledo M. Tracing sewage in the marine environment: altered signatures in Conception Bay, Chile. Water Res 2001; 35(17): 4166-474.
25. Akpata TVI, Oyenekan JA, and Nwanko DI. Impacts of organic pollution on bacterial, plankton and benthic populations of Lagos Lagoon, Nigeria. International J Ecol Environ Sci 1993; 19: 73-82.
26. Olayemi AB, Adedayo O, and Ojo AO. Microbiological studies on freshwater fishes from the Asa River, Ilorin, Nigeria. J Aquacul Trop. 1990; 5: 135-139.
27. Ndip RN, Akoachere J-FTK, Mokosso DK, Ndip LM, and Anyangwe IAN. Carriage of Vibrio species by shrimps harvested from the coastal waters of South West Cameroon. East Afri Med J. 2002; 79:146-149.
28. Guevart E, Noeske J, Essomba JM, Edjenguele M, Bita A, Mouangue A, and Manga B. Factors contributing to endemic cholera in Douala, Cameroon. Med Trop 2006; 66(3): 283-91.
29. Alam M, Sultana M, Nair GB, Sack RB, Sack DA, Siddique AK, Ali A, Huq A, and Colwell RR. Toxigenic Vibrio cholerae in the aquatic environment of Mathbaria, Bangladesh. Appl Environ Microbiol. 2006; 72(4):2849-55.
30. Binsztein N, Costagliola MC, Pichel M, Jurquiza V, Ramirez FC, Akselman R, Vacchino M, Huq, A, and Colwell R. Viable but nonculturable Vibrio cholerae O1 in the environment of Argentina. Appl Environ Microbiol. 2004; 70(12): 7481-6.
31. Faruque SM, Biswas K, Udden SMN, Ahmad QS, Sack DA, Nair GB, and Mekalanos JJ. Transmissibility of cholera: In vivo-formed bioflms and their relationship to infectivity and persistence in the environment. P N A S, USA. 2006; 103(16):6350-6355.

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