Journal of Applied Bacteriology 1992,73, 3740
Survival of bacterial enteric pathogens in traditional fermented foods C. Simango and Grace Rukure' Department of Medical Laboratory Technology, University of Zimbabwe Medical School, Avondale and 'Blair Research Laboratory, Ministry of Health, Causeway, Zimbabwe 3777/09/91:accepted 3 February 1992
c . s I M A NG o A N D G . R u K u R E. 1992.The survival of strains of bacterial enteric pathogens was investigated in two traditional fermented foods (mahewu and sour porridge) and in unfermented porridge. The foods were inoculated with cell suspensions of Salmonella, Shigella, Campylobacter, Aeromonas species and pathogenic Escherichia coli which had a final concentration of 106-107 cfu/ml of food. None of the strains of Aeromonas and Campylobacter were detected in mahewu and sour porridge 20 min after inoculation. The salmonellas were not found 4 h after inoculation in either fermented foods but the shigellas and pathogenic E. coli strains were more tolerant to the low pH of the fermented foods. Some of the shigellas and pathogenic E. coli strains survived for 24 h after inoculation but showed a sharp decrease in numbers. All the strains of the enteric pathogens survived for 24 h in the unfermented porridge and increased in the numbers except for campylobacters, the numbers of which declined. These results suggest that the traditional fermented foods have bacteriostatic and bactericidal properties and are unlikely to play a major role in the transmission of bacterial enteric pathogens.
INTRODUCTION
Diarrhoea is a major cause of morbidity and mortality in children in developing countries. A number of bacterial enteric pathogens which include Shigella, Salmonella, Campylobacrer, Aeromonas species and pathogenic strains of Escherichia coli have been associated with childhood diarrhoea. Standards of personal hygiene and public sanitation are low in many communities in these countries and it has been shown that indigenous weaning foods are a significant source of childhood diarrhoea (Rowland et al. 1978; Barrel & Rowland 1979; Black et al. 1982). There is growing interest in the use of traditional fermented foods for weaning as it is thought that bacterial pathogens likely to cause diarrhoea could not survive for long periods in them because organic acids, e.g. lactate, propionate and acetate, are produced during the fermentation process and may be inhibitory. However, not much research has been carried out in this field. One study by Simango & Rukure (1991) on mahewu, a traditional fermented sour, non-alcoholic beverage in Zimbabwe, showed that none of the test strains of C. jejuni were detected 30 Correspondence to: D r C. Simango, Department of Medical Laboratory Technology, University of Zimbabwe Medical School, PO Box A1 78, Avondale, Zimbabwe.
min after inoculation in mahewu. All the strains of pathogenic E. coli were detected 24 h after inoculation but did not increase in numbers. This study was performed to investigate the survival of bacterial pathogens, which have been associated with childhood diarrhoea, in mahewu and sour porridge (fermented foods) and in unfermented porridge.
MATERIALS AND METHODS
Preparation of mahewu, sour and unfermented porrldge Mahewu was prepared by boiling a broth mixture of 200 g of maize meal flour and 2 1 of unchlorinated water for 10 min to gelatinize the starch. The cooked broth was cooled to 40"C, 100 g of sorghum malt flour was added and the mahewu broth was mixed thoroughly. Some of the unfermented mahewu broth was filtered through a 100 pm nylon bolting cloth. The pH of the filtrate was measured. The percentage of acidity, expressed as lactic acid, was determined by titrating the filtrate with sodium hydroxide with phenolphthalein as indicator (Anon. 1984). The unfiltered mahewu was left to ferment at room temperature (20"-
38 C . SIMANGO AND GRACE RUKURE
30°C) for 18 h. The fermented mahewu was then filtered and the pH and percentage of lactic acid of the filtrate determined as before. T h e mahewu broth and the fermented mahewu were not boiled before determination of acidity and p H because mahewu is not boiled before it is consumed. Sour porridge was prepared by mixing 300 g of maize meal flour with 2 I of water and allowing it to ferment at room temperature for 48 h. Before fermentation some of the broth mixture was filtered and boiled for 10 rnin to gelatinize the starch. When it was cool, the p H and the percentage of lactic acid in the filtrate were determined. After fermentation, the sour porridge broth was filtered and the filtrate was boiled for 10 min. After cooling the p H and percentage of lactic acid were determined. Unfermented porridge was prepared by adding 300 g of maize meal flour to 2 1 of water and filtering the broth mixture. The filtrate was boiled for 10 min and when cool the p H and percentage of lactic acid were determined. Fifty ml quantities of each type of food were distributed into 100 ml bottles with screw caps and were sterilized at 121°C for 15 min to stop the fermentation process. Survival of bacteria in mahewu, sour and unfermentedporridges
Shigella sonnei, Sh. flexneri, Salmonella species (Groups B and C), enteropathogenic E. coli (EPEC), enterotoxigenic (LT) E. coli (ETEC) and two strains of C. jejuni and Aeromonas species, which were isolated from stool specimens of children with diarrhoea, were used to inoculate the mahewu and porridge samples. The bacteria were subcultured twice on appropriate media (Oxoid): Mueller Hinton agar with 5% lysed sheep blood for Campylobacter, MacConkey agar for E. coli, Salmonella and Shigella, blood agar supplemented with ampicillin (10 pg/ml) for Aeromonas, before testing their survival in mahewu and the two types of porridge. A 24 h culture was used to prepare bacterial cell suspensions in 0.1% peptone water. The turbidity of the suspensions was standardized to match that of a barium chloride turbidity standard (prepared by adding 0.5 ml of 1.175% (w/v) barium chloride solution to 99.5 ml of 1% sulphuric acid). One ml of the cell suspension was added to 50 ml of sterile mahewu, sour and unfermented porridges in 100 ml bottles and the contents were mixed thoroughly. T h e inoculated samples, which had a final inoculum of 106-107 cfu/ml of food, were kept at 25°C. Surviving bacterial cells were enumerated by plating 10-fold serial dilutions of the inoculated samples in 0.1% peptone water at various periods of time (0, 10 and 20 min for Campylobacter and Aeromonas; 0, 1, 2, 3 and 4 h for Salmonella; 0, 3, 6 and 24 h for Shigella and E. coli) for the fermented foods. Serial dilutions of all the bacterial species
inoculated in the unfermented porridge were tested at 0, 3, 6 and 24 h after inoculation. A glass spreader was used to surface-plate 0.1 ml of each serial dilution in duplicate on appropriate media. T h e levels of inocula were determined by adding 1 ml of the standardized cell suspensions to 50 ml of 0.1% peptone water and surface-plating 10-fold serial dilutions in duplicate on the appropriate media. The minimum level of sensitivity of the counting method was 100 cfu/rnl of food sample. Preston Selective medium (Oxoid) with 5% lysed sheep blood was used for the enumeration of Campylobacter. T h e inoculated plates were incubated at 42'43°C for 48 h under micro-aerobic conditions and colonies typical of Campylobacter were counted. Representative colonies were confirmed as Campylobacter by a positive oxidase test and characteristic morphology of the bacterial cells. MacConkey agar medium was used for the enumeration of Salmonella, Shigella and E. coli. T h e inoculated plates were incubated at 37°C for 24 h. Colonies typical of Salmonella, Shigella and E. coli were counted. Representative colonies were confirmed as Salmonella, Shigella and E. coli by the API 20E system (API System, France). Aeromonas was enumerated on starch-ampicillin agar following the method of Palumbo et al. (1985). T h e cultures were incubated at room temperature for 48 h. After incubation, the plates which had growth were flooded with 5 ml of Lugol's iodine solution and amylase-positive colonies (those having clear zones surrounding the colonies) were counted as Aeromonas. Representative colonies were subcultured before the addition of iodine and were confirmed as Aeromonas by the API 20E system. RESULTS
T h e p H and percentage of total acidity, expressed as lactic acid, of mahewu, sour and unfermented porridges are shown in Table 1. Before the fermentation, the mahewu broth had a slightly lower p H than that of sour porridge. T h e pH of mahewu and sour porridge after fermentation
Table 1 The pH of mahewu, sour and unfermented porridges and percentages of lactic acid in these products
pH before fermentation pH after fermentation Percentage of lactic acid before fermentation Percentage of lactic acid after fermentation NA, Not applicable.
Mahew
Sour porridge
Unfermented porridge
5.5 2.74 0.07
5.95 2.91 0.02
5.96 NA
0.5
0.44
NA
0.04
BACTERIAL ENTERIC PATHOGENS IN FERMENTED FOODS
39
10
10
E
\
c ._ C 3
:.
m
-
10
b
ZI
0 V 0
10
10 Time (hl
Fig. 1 Survival of bacterial enteric pathogens in mahewu (pH 2.74) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); a, enterotoxigenic E. coli (ETEC); A, Shigella 1;A,Shigella 2; 0, Salmonella 1 ; 0 ,Salmonella 2
.,
Time (hl
Fig. 2 Survival of bacterial enteric pathogens in sour porridge (pH 2.91) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); enterotoxigenic E. coli (ETEC); A, Shigella 1 ;A,Shigella 2; 0,Salmonella 1 ; 0 ,Salmonella 2
"
I
l
l
0 1 2 3 4
.,
I
I
6
24 Time (h)
Fig. 3 Survival of bacterial enteric pathogens in unfermented porridge (pH 5.96) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); enterotoxigenic E. coli (ETEC); A, Shigella 1;A, Shigella 2; 0,Salmonella 1; 0 ,Salmonella 2; +, Campylobacter 1; x ,Campylobacter 2; - - - -, Aeromonas 1; -.-.-. ,Aeromonas 2
dropped to about half of that before fermentation. There was an increase in the percentage of lactic acid in both mahewu and sour porridge after fermentation. None of the strains of Aeromonas and Campylobacter tested survived for 10 and 20 min respectively in mahewu (pH 2.74) although heavy inocula of 106-107 cfu bacterial cells/ml of mahewu were used. The two strains of Salmonella could not be detected 4 h after inoculation in the mahewu while the two strains of Shigella and enterotoxigenic E. coli strain were not detected after 24 h (Fig. 1). Only the enteropathogenic E. coli strain survived for 24 h in mahewu but decreased by 3 log,, cfu. None of the strains of Aeromonas and Campylobacter tested survived for 10 min in sour porridge. The Salmonella strains could not be detected 3 h after inoculation in sour porridge while one strain of Shigella was detected after 24 h (Fig. 2). The two E. coli and one Shigella strains survived for 24 h in sour porridge but decreased in number by nearly 3 log,, cfu. All the bacterial pathogens survived in the unfermented porridge for 24 h. Aeromonas, Salmonella, Shigella and E.
40 C . SIMANGO AND GRACE RUKURE
coli strains increased from 1-5 to 2.5 loglo cfu (Fig. 3). The two Campylobacter strains decreased in the unfermented
porridge. DISCUSSION
There was variation in the survival of the various species of bacterial enteric pathogens in the fermented foods. Aeromonas and Campylobacter were the most sensitive to both mahewu and sour porridge, as they did not survive for 20 min. Salmonella could not be detected 4 h after inoculation. Shigella and E. colt were less affected by the mahewu and sour porridge but showed a marked decrease in numbers that survived for 24 h. These observations show that mahewu and sour porridge have metabolites, produced during the fermentation process, which are bactericidal to organisms such as Aeromonas, Campylobacter, and Salmonella and bacteriostatic to others such as Shigella and E. coli. This suggests that mahewu and sour porridge are unlikely sources of diarrhoea caused by Aeromonas, Campylobacter and Salmonella since these organisms did not survive for long periods after inoculation in the foods. These foods may also play only a minor role in the transmission of diarrhoea caused by Shigella and pathogenic E. coli as these bacteria do not grow in the fermented foods. A study by Mensah et al. (1988) on the survival of S h . jexneri in fermented Ghanaian maize dough showed that most of the strains were not detectable 6 h after inoculation in the maize dough that had been fermented for 3 d. This suggested that the fermented maize dough is an unlikely source of S h . jexneri. A few studies have shown that E. coli is tolerant to acidic conditions in fermented foods where the organism survived for at least 24 h. In a study by Feresu & Nyati (1990), E. colt strains were detected 24 h after inoculation in both traditional and commercial fermented milk, but there was a decrease in numbers. Simango & Rukure (1991) also observed the survival of pathogenic strains of E. coli 24 h after inoculation in mahewu which was prepared by women in a rural community. The observation that Aeromonas, Salmonella, Shigella and pathogenic E. coli survive and multiply in the unfermented porridge suggest that the unfermented porridge can be a vehicle for the transmission of bacterial enteric pathogens once it is contaminated with these organisms.
The results of the present study suggest that mahewu and sour porridge may be of considerable value in reducing the transmission of bacterial enteric pathogens through contaminated foods in developing countries if their use as weaning foods is promoted.
ACKNOWLEDGEMENTS
We thank the Secretary for Health for permission to publish and Christopher Nhandara for technical assistance.
REFERENCES ANON. (1984) Official Methods of Analysis of the Association of Oflcial Analytical Chemists, 14th edn. Washington: Association of Official Analytical Chemists. B A R R E L LR, . A . E . & R O W L A N DM , . G . M . (1979) Infant foods as a potential source of diarrhoea1 illness in rural West Africa. Transactions of the Royal Society of Tropical Medicine and Hygiene 73, 85-90. B L A C KR, . E . , B R O W NK , . H . , B E C K E RS, . , ABDULA L I M , A . R . M . & M E R S O N ,M . H . (1982) Contamination of weaning foods and transmission of enterotoxigenic Escherichia coli diarrhoea in children in rural Bangladesh. Transactions of the Royal Society of Tropical Medicine and Hygiene, 76, 259264. F E R E S US, . & N Y A T I H , . (1990) Fate of pathogenic and nonpathogenic Escherichia coli strains in two fermented milk products. Journal of Applied Bacteriology 69, 814-821. M E N S A H ,P . P . A . , T O M K I N SA , . M . , D R A S A RB, . S . & H A R R I S O NT, . J . (1988) Effect of fermentation of Ghanaian maize dough on the survival and proliferation of four strains of Shigclla jlesneri. Transactions of the Royal Society of Tropical Medicine and Hygiene 82, 635-636. P A L U M B O ,S . A . , M A X I N O , F . , W I L L I A M S ,A . C . , B U C H A N A NR, . L . & T H A Y E R D,. W . (1985) Starchampicillin agar for the quantitative detection of Aeromonas hydrophila. Applied and Environmental Microbiology 50, 10271030. ROWLAND, M . G . M . , B A R R E L LR, . A . E . & W H I T E H E A D , R . G . (1978) Bacterial contamination in traditional Gambian weaning foods. Lancet i, 136-138. S I M A N G OC., & R U K U R EG, . (1991) Survival of Campylobacter jejuni and pathogenic Escherichia coli in mahewu, a fermented cereal gruel. Transactions of the Royal Society of Tropical Medicine and Hygiene 85, 399-400.
Survival of bacterial enteric pathogens in traditional fermented foods C. Simango and Grace Rukure' Department of Medical Laboratory Technology, University of Zimbabwe Medical School, Avondale and 'Blair Research Laboratory, Ministry of Health, Causeway, Zimbabwe 3777/09/91:accepted 3 February 1992
c . s I M A NG o A N D G . R u K u R E. 1992.The survival of strains of bacterial enteric pathogens was investigated in two traditional fermented foods (mahewu and sour porridge) and in unfermented porridge. The foods were inoculated with cell suspensions of Salmonella, Shigella, Campylobacter, Aeromonas species and pathogenic Escherichia coli which had a final concentration of 106-107 cfu/ml of food. None of the strains of Aeromonas and Campylobacter were detected in mahewu and sour porridge 20 min after inoculation. The salmonellas were not found 4 h after inoculation in either fermented foods but the shigellas and pathogenic E. coli strains were more tolerant to the low pH of the fermented foods. Some of the shigellas and pathogenic E. coli strains survived for 24 h after inoculation but showed a sharp decrease in numbers. All the strains of the enteric pathogens survived for 24 h in the unfermented porridge and increased in the numbers except for campylobacters, the numbers of which declined. These results suggest that the traditional fermented foods have bacteriostatic and bactericidal properties and are unlikely to play a major role in the transmission of bacterial enteric pathogens.
INTRODUCTION
Diarrhoea is a major cause of morbidity and mortality in children in developing countries. A number of bacterial enteric pathogens which include Shigella, Salmonella, Campylobacrer, Aeromonas species and pathogenic strains of Escherichia coli have been associated with childhood diarrhoea. Standards of personal hygiene and public sanitation are low in many communities in these countries and it has been shown that indigenous weaning foods are a significant source of childhood diarrhoea (Rowland et al. 1978; Barrel & Rowland 1979; Black et al. 1982). There is growing interest in the use of traditional fermented foods for weaning as it is thought that bacterial pathogens likely to cause diarrhoea could not survive for long periods in them because organic acids, e.g. lactate, propionate and acetate, are produced during the fermentation process and may be inhibitory. However, not much research has been carried out in this field. One study by Simango & Rukure (1991) on mahewu, a traditional fermented sour, non-alcoholic beverage in Zimbabwe, showed that none of the test strains of C. jejuni were detected 30 Correspondence to: D r C. Simango, Department of Medical Laboratory Technology, University of Zimbabwe Medical School, PO Box A1 78, Avondale, Zimbabwe.
min after inoculation in mahewu. All the strains of pathogenic E. coli were detected 24 h after inoculation but did not increase in numbers. This study was performed to investigate the survival of bacterial pathogens, which have been associated with childhood diarrhoea, in mahewu and sour porridge (fermented foods) and in unfermented porridge.
MATERIALS AND METHODS
Preparation of mahewu, sour and unfermented porrldge Mahewu was prepared by boiling a broth mixture of 200 g of maize meal flour and 2 1 of unchlorinated water for 10 min to gelatinize the starch. The cooked broth was cooled to 40"C, 100 g of sorghum malt flour was added and the mahewu broth was mixed thoroughly. Some of the unfermented mahewu broth was filtered through a 100 pm nylon bolting cloth. The pH of the filtrate was measured. The percentage of acidity, expressed as lactic acid, was determined by titrating the filtrate with sodium hydroxide with phenolphthalein as indicator (Anon. 1984). The unfiltered mahewu was left to ferment at room temperature (20"-
38 C . SIMANGO AND GRACE RUKURE
30°C) for 18 h. The fermented mahewu was then filtered and the pH and percentage of lactic acid of the filtrate determined as before. T h e mahewu broth and the fermented mahewu were not boiled before determination of acidity and p H because mahewu is not boiled before it is consumed. Sour porridge was prepared by mixing 300 g of maize meal flour with 2 I of water and allowing it to ferment at room temperature for 48 h. Before fermentation some of the broth mixture was filtered and boiled for 10 rnin to gelatinize the starch. When it was cool, the p H and the percentage of lactic acid in the filtrate were determined. After fermentation, the sour porridge broth was filtered and the filtrate was boiled for 10 min. After cooling the p H and percentage of lactic acid were determined. Unfermented porridge was prepared by adding 300 g of maize meal flour to 2 1 of water and filtering the broth mixture. The filtrate was boiled for 10 min and when cool the p H and percentage of lactic acid were determined. Fifty ml quantities of each type of food were distributed into 100 ml bottles with screw caps and were sterilized at 121°C for 15 min to stop the fermentation process. Survival of bacteria in mahewu, sour and unfermentedporridges
Shigella sonnei, Sh. flexneri, Salmonella species (Groups B and C), enteropathogenic E. coli (EPEC), enterotoxigenic (LT) E. coli (ETEC) and two strains of C. jejuni and Aeromonas species, which were isolated from stool specimens of children with diarrhoea, were used to inoculate the mahewu and porridge samples. The bacteria were subcultured twice on appropriate media (Oxoid): Mueller Hinton agar with 5% lysed sheep blood for Campylobacter, MacConkey agar for E. coli, Salmonella and Shigella, blood agar supplemented with ampicillin (10 pg/ml) for Aeromonas, before testing their survival in mahewu and the two types of porridge. A 24 h culture was used to prepare bacterial cell suspensions in 0.1% peptone water. The turbidity of the suspensions was standardized to match that of a barium chloride turbidity standard (prepared by adding 0.5 ml of 1.175% (w/v) barium chloride solution to 99.5 ml of 1% sulphuric acid). One ml of the cell suspension was added to 50 ml of sterile mahewu, sour and unfermented porridges in 100 ml bottles and the contents were mixed thoroughly. T h e inoculated samples, which had a final inoculum of 106-107 cfu/ml of food, were kept at 25°C. Surviving bacterial cells were enumerated by plating 10-fold serial dilutions of the inoculated samples in 0.1% peptone water at various periods of time (0, 10 and 20 min for Campylobacter and Aeromonas; 0, 1, 2, 3 and 4 h for Salmonella; 0, 3, 6 and 24 h for Shigella and E. coli) for the fermented foods. Serial dilutions of all the bacterial species
inoculated in the unfermented porridge were tested at 0, 3, 6 and 24 h after inoculation. A glass spreader was used to surface-plate 0.1 ml of each serial dilution in duplicate on appropriate media. T h e levels of inocula were determined by adding 1 ml of the standardized cell suspensions to 50 ml of 0.1% peptone water and surface-plating 10-fold serial dilutions in duplicate on the appropriate media. The minimum level of sensitivity of the counting method was 100 cfu/rnl of food sample. Preston Selective medium (Oxoid) with 5% lysed sheep blood was used for the enumeration of Campylobacter. T h e inoculated plates were incubated at 42'43°C for 48 h under micro-aerobic conditions and colonies typical of Campylobacter were counted. Representative colonies were confirmed as Campylobacter by a positive oxidase test and characteristic morphology of the bacterial cells. MacConkey agar medium was used for the enumeration of Salmonella, Shigella and E. coli. T h e inoculated plates were incubated at 37°C for 24 h. Colonies typical of Salmonella, Shigella and E. coli were counted. Representative colonies were confirmed as Salmonella, Shigella and E. coli by the API 20E system (API System, France). Aeromonas was enumerated on starch-ampicillin agar following the method of Palumbo et al. (1985). T h e cultures were incubated at room temperature for 48 h. After incubation, the plates which had growth were flooded with 5 ml of Lugol's iodine solution and amylase-positive colonies (those having clear zones surrounding the colonies) were counted as Aeromonas. Representative colonies were subcultured before the addition of iodine and were confirmed as Aeromonas by the API 20E system. RESULTS
T h e p H and percentage of total acidity, expressed as lactic acid, of mahewu, sour and unfermented porridges are shown in Table 1. Before the fermentation, the mahewu broth had a slightly lower p H than that of sour porridge. T h e pH of mahewu and sour porridge after fermentation
Table 1 The pH of mahewu, sour and unfermented porridges and percentages of lactic acid in these products
pH before fermentation pH after fermentation Percentage of lactic acid before fermentation Percentage of lactic acid after fermentation NA, Not applicable.
Mahew
Sour porridge
Unfermented porridge
5.5 2.74 0.07
5.95 2.91 0.02
5.96 NA
0.5
0.44
NA
0.04
BACTERIAL ENTERIC PATHOGENS IN FERMENTED FOODS
39
10
10
E
\
c ._ C 3
:.
m
-
10
b
ZI
0 V 0
10
10 Time (hl
Fig. 1 Survival of bacterial enteric pathogens in mahewu (pH 2.74) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); a, enterotoxigenic E. coli (ETEC); A, Shigella 1;A,Shigella 2; 0, Salmonella 1 ; 0 ,Salmonella 2
.,
Time (hl
Fig. 2 Survival of bacterial enteric pathogens in sour porridge (pH 2.91) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); enterotoxigenic E. coli (ETEC); A, Shigella 1 ;A,Shigella 2; 0,Salmonella 1 ; 0 ,Salmonella 2
"
I
l
l
0 1 2 3 4
.,
I
I
6
24 Time (h)
Fig. 3 Survival of bacterial enteric pathogens in unfermented porridge (pH 5.96) at 25°C. 0,Enteropathogenic Escherichia coli (EPEC); enterotoxigenic E. coli (ETEC); A, Shigella 1;A, Shigella 2; 0,Salmonella 1; 0 ,Salmonella 2; +, Campylobacter 1; x ,Campylobacter 2; - - - -, Aeromonas 1; -.-.-. ,Aeromonas 2
dropped to about half of that before fermentation. There was an increase in the percentage of lactic acid in both mahewu and sour porridge after fermentation. None of the strains of Aeromonas and Campylobacter tested survived for 10 and 20 min respectively in mahewu (pH 2.74) although heavy inocula of 106-107 cfu bacterial cells/ml of mahewu were used. The two strains of Salmonella could not be detected 4 h after inoculation in the mahewu while the two strains of Shigella and enterotoxigenic E. coli strain were not detected after 24 h (Fig. 1). Only the enteropathogenic E. coli strain survived for 24 h in mahewu but decreased by 3 log,, cfu. None of the strains of Aeromonas and Campylobacter tested survived for 10 min in sour porridge. The Salmonella strains could not be detected 3 h after inoculation in sour porridge while one strain of Shigella was detected after 24 h (Fig. 2). The two E. coli and one Shigella strains survived for 24 h in sour porridge but decreased in number by nearly 3 log,, cfu. All the bacterial pathogens survived in the unfermented porridge for 24 h. Aeromonas, Salmonella, Shigella and E.
40 C . SIMANGO AND GRACE RUKURE
coli strains increased from 1-5 to 2.5 loglo cfu (Fig. 3). The two Campylobacter strains decreased in the unfermented
porridge. DISCUSSION
There was variation in the survival of the various species of bacterial enteric pathogens in the fermented foods. Aeromonas and Campylobacter were the most sensitive to both mahewu and sour porridge, as they did not survive for 20 min. Salmonella could not be detected 4 h after inoculation. Shigella and E. colt were less affected by the mahewu and sour porridge but showed a marked decrease in numbers that survived for 24 h. These observations show that mahewu and sour porridge have metabolites, produced during the fermentation process, which are bactericidal to organisms such as Aeromonas, Campylobacter, and Salmonella and bacteriostatic to others such as Shigella and E. coli. This suggests that mahewu and sour porridge are unlikely sources of diarrhoea caused by Aeromonas, Campylobacter and Salmonella since these organisms did not survive for long periods after inoculation in the foods. These foods may also play only a minor role in the transmission of diarrhoea caused by Shigella and pathogenic E. coli as these bacteria do not grow in the fermented foods. A study by Mensah et al. (1988) on the survival of S h . jexneri in fermented Ghanaian maize dough showed that most of the strains were not detectable 6 h after inoculation in the maize dough that had been fermented for 3 d. This suggested that the fermented maize dough is an unlikely source of S h . jexneri. A few studies have shown that E. coli is tolerant to acidic conditions in fermented foods where the organism survived for at least 24 h. In a study by Feresu & Nyati (1990), E. colt strains were detected 24 h after inoculation in both traditional and commercial fermented milk, but there was a decrease in numbers. Simango & Rukure (1991) also observed the survival of pathogenic strains of E. coli 24 h after inoculation in mahewu which was prepared by women in a rural community. The observation that Aeromonas, Salmonella, Shigella and pathogenic E. coli survive and multiply in the unfermented porridge suggest that the unfermented porridge can be a vehicle for the transmission of bacterial enteric pathogens once it is contaminated with these organisms.
The results of the present study suggest that mahewu and sour porridge may be of considerable value in reducing the transmission of bacterial enteric pathogens through contaminated foods in developing countries if their use as weaning foods is promoted.
ACKNOWLEDGEMENTS
We thank the Secretary for Health for permission to publish and Christopher Nhandara for technical assistance.
REFERENCES ANON. (1984) Official Methods of Analysis of the Association of Oflcial Analytical Chemists, 14th edn. Washington: Association of Official Analytical Chemists. B A R R E L LR, . A . E . & R O W L A N DM , . G . M . (1979) Infant foods as a potential source of diarrhoea1 illness in rural West Africa. Transactions of the Royal Society of Tropical Medicine and Hygiene 73, 85-90. B L A C KR, . E . , B R O W NK , . H . , B E C K E RS, . , ABDULA L I M , A . R . M . & M E R S O N ,M . H . (1982) Contamination of weaning foods and transmission of enterotoxigenic Escherichia coli diarrhoea in children in rural Bangladesh. Transactions of the Royal Society of Tropical Medicine and Hygiene, 76, 259264. F E R E S US, . & N Y A T I H , . (1990) Fate of pathogenic and nonpathogenic Escherichia coli strains in two fermented milk products. Journal of Applied Bacteriology 69, 814-821. M E N S A H ,P . P . A . , T O M K I N SA , . M . , D R A S A RB, . S . & H A R R I S O NT, . J . (1988) Effect of fermentation of Ghanaian maize dough on the survival and proliferation of four strains of Shigclla jlesneri. Transactions of the Royal Society of Tropical Medicine and Hygiene 82, 635-636. P A L U M B O ,S . A . , M A X I N O , F . , W I L L I A M S ,A . C . , B U C H A N A NR, . L . & T H A Y E R D,. W . (1985) Starchampicillin agar for the quantitative detection of Aeromonas hydrophila. Applied and Environmental Microbiology 50, 10271030. ROWLAND, M . G . M . , B A R R E L LR, . A . E . & W H I T E H E A D , R . G . (1978) Bacterial contamination in traditional Gambian weaning foods. Lancet i, 136-138. S I M A N G OC., & R U K U R EG, . (1991) Survival of Campylobacter jejuni and pathogenic Escherichia coli in mahewu, a fermented cereal gruel. Transactions of the Royal Society of Tropical Medicine and Hygiene 85, 399-400.
