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Microflora of Minimally Processed Frozen Vegetables Sold in Gaborone, Botswana

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Microflora of Minimally Processed Frozen Vegetables Sold in Gaborone, Botswana 2581 Journal of Food Protection, Vol. 69, No. 11, 2006, Pages 2581–2586 Copyright ᮊ, International Association for Food Protection Microflora of Minimally Processed Frozen Vegetables Sold in Gaborone, Botswana TINNA A. MANANI,1 ERNEST K. COLLISON,2 AND SISAI MPUCHANE2* 1University of Malawi, Lilongwe, Malawi; and 2University of Botswana, Gaborone, Botswana MS 06-181: Received 28 March 2006/Accepted 13 June 2006 ABSTRACT Downloaded from http://meridian.allenpress.com/jfp/article-pdf/69/11/2581/1679939/0362-028x-69_11_2581.pdf by guest on 23 September 2021 Two hundred samples of minimally processed, frozen, and prepacked potato chips, peas, corn, and a variety of combined vegetables from supermarkets in Gaborone, Botswana, were examined microbiologically. Determination of aerobic mesophilic plate count, aerobic psychrotrophic plate count, lactic acid bacteria, yeasts and molds, coliforms, Listeria spp., and Staphy- lococcus aureus were done. Chips had the lowest mean log values for all of the microorganisms enumerated except yeasts and molds. The mean log values for single vegetables ranged from 3.6 to 9.1, 3.4 to 8.9, 2.9 to 5.6, and 2.1 to 6.5 log CFU/ g aerobic mesophilic plate count, aerobic psychrotrophic plate count, lactic acid bacteria, and yeasts and molds, respectively. The microbial profiles of peas and corn were almost similar (P Ͻ 0.001). The mean values for combined vegetables were clustered within 4.6 and 5.4 and 4.2 and 5.2 log CFU/g aerobic mesophilic plate count and aerobic psychrotrophic plate count, respectively. All of the vegetables had a coliform population distribution ranging from 0 to Ͻ104 most probable number per g. The predominant gram-negative bacteria isolated included members of Enterobacteriaceae and Pseudomonaceae (86.2%). Escherichia coli was not detected in all of the samples. The organisms isolated included those responsible for spoilage in frozen vegetables, namely Pseudomonas, Klebsiella, Corynebacterium, lactic acid bacteria, and Flavobacterium. The predom- inant lactic acid bacteria were Lactobacillus spp. (55.9%). Other spoilage organisms were yeasts, and Cryptococcus spp. (55.4%) was predominant. Pathogens, namely Listeria monocytogenes, were also isolated at a rate of 2 to 10%, of which 4% was from corn, 2% each from peas and country crop, and 10% from stir-fry. Bacillus cereus was also isolated and accounted for 7.7% of the microorganisms from corn. S. aureus was isolated from all of the vegetables. Enterotoxigenic strains were from corn, peas, mixed vegetables, and stir-fry, and all of them produced enterotoxin A. In addition, the isolates from stir-fry vegetables also produced enterotoxins B and C. The study reveals the presence of pathogens and emerging opportunistic pathogens in the ready-to-use or ready-to-eat vegetables. If E. coli is the only indicator for safety and acceptability, consumers may be exposed to foodborne diseases. Inclusion of other groups as indicator organisms is suggested. Retailers are urged to invest in standby generators to maintain the cold chain. The consumer demand for high-quality foods requiring MP vegetables is from industrialized countries, and infor- only a minimal amount of effort and time for preparation mation is lacking regarding microbial profiles, ecology, has led to the introduction of minimally processed (ready- quality, and safety of MP vegetables in developing coun- to-use or ready-to-eat) convenient foods preserved by mild tries where the use of such food is on the increase. Thus, methods (1). Minimally processed (MP) vegetables consist this study was undertaken to determine the microbial qual- of raw fresh cut produce, which has undergone minimal ity of a variety of MP vegetables sold in retail supermarkets processing such as trimming, peeling, slicing, shredding, in Botswana. cutting, washing, and blanching before packaging in sealed Most of these products are imported into the country pouches or in plastic trays. and transported by road. The efficiency of the maintenance The potential for microbial contamination is high be- of the cold chain to wholesalers and retailers and conditions cause of the wide variety of conditions to which the pro- during sale would have a bearing on microbiological qual- duce is exposed during growth, harvest, processing, and ity of the consumed vegetables. distribution (15). Slicing and shredding procedures as well as improper refrigeration during storage have been associ- MATERIALS AND METHODS ated with an increase in the number of mesophilic and psy- Sample collection. Two hundred MP vegetables were pur- chrotrophic aerobic microorganisms (2). One of the psy- chased in frozen state from various supermarkets in Gaborone. chrotrophic bacterial pathogens is Listeria monocytogenes, These vegetables are packed in sealed plastic bags, which are which is capable of growing at chill temperatures that are usually displayed in open freezer units. The samples comprised heavily relied upon by the food industry to maintain prod- 50 each of combined vegetables, corn, peas, and potato chips. The uct quality (23) and to extend product shelf life (1). samples were purchased from five supermarkets, and from each Most published information on the microbial quality of supermarket, 10 samples of corn, peas, and chips were bought. The numbers of combined vegetables were determined by avail- * Author for correspondence. Tel: ϩ267 3552598; Fax: ϩ267 3909476; ability from different supermarkets. The samples were transferred E-mail: [email protected]. to the laboratory in ice coolers within an hour of collection and 2582 MANANI ET AL. J. Food Prot., Vol. 69, No. 11 sampling was done within 24 h of collection. Combined vegetable bation at 35ЊC for 24 and 48 h. Tubes showing growth and gas samples refer to a mixture of different types of vegetables in one were considered positive for total coliforms. Fecal coliforms were pack. The packaged vegetables used were 10 samples of stir-fry also enumerated by the three-tube MPN method using lauryl tryp- (cabbage, beans, carrots, and onions); 10 samples of mixed veg- tose broth, incubated at 44ЊC for 24 and 48 h. A loopful of culture etables (carrots, onions, baby marrows, beans, peppers, and cab- showing growth and gas was streaked on eosin methylene blue bage); 16 samples of country crop (cauliflower florets, carrot roun- agar (Oxoid). Typical E. coli colonies produced a green metallic dels, cut green beans, and broccoli cuts); 5 samples of vegetable sheen color on eosin methylene blue agar. curry (cauliflower, carrots, peas, corn, beans, potatoes, and curry E. coli. The samples were cultured using a standard plate spices); 9 samples of Mexican mix (broccoli, cauliflower, corn, technique on sorbitol MacConkey agar (HiMedia, Mumbai, India) red peppers, and spices); and 50 samples each of single vegeta- for the isolation of E. coli. Purified isolates were subjected to bles, which refers to those packaged as single entities and include various morphological and biochemical tests including API 20E potato chips, corn, and peas. galleries. Identification was carried out using the ATB reader (bioMe´rieux) Sample preparation. The packaged contents of the vegeta- Downloaded from http://meridian.allenpress.com/jfp/article-pdf/69/11/2581/1679939/0362-028x-69_11_2581.pdf by guest on 23 September 2021 bles were mixed, and a total of 50 g was aseptically transferred LAB. Lactic acid bacteria (LAB) were enumerated on MRS into a sterile stomacher bag, and 450 ml of sterile 0.1% peptone (Oxoid) agar (14). The samples were cultured using the pour plate water was added. The sample-diluent mixture was blended for 4 method with an overlay (11) and were incubated at 37ЊC for 24 min in a Stomacher 400 laboratory blender (Seward Medical, Lon- to 48 h, after which representative colonies were isolated, purified, don, UK). Serial, 10-fold dilutions 10Ϫ1 to 10Ϫ6 were made using and maintained on MRS agar slants at 4ЊC for identification. the blended sample and 90-ml aliquots of 0.1% peptone water for Gram positives, which were catalase negative and were iso- analysis of aerobic (mesophiles and psychrotrophs) bacterial, co- lated from MRS agar, were treated as presumptive LAB. These liform, yeast and mold, and lactic acid bacterial counts. Samples isolates were characterized and identified using API 50CH and were also analyzed for the presence of Escherichia coli, Staphy- API 50CHL medium (bioMe´rieux), according to the manufactur- lococcus aureus, and Listeria. er’s specifications. Enumeration and detection of microorganisms. Aerobic S. aureus. Triplicate spread plate technique was used on mesophilic plate counts and aerobic psychrophilic plate counts Baird-Parker (Merck, Whitehouse Station, N.J.) supplemented were done as follows: Six replicate plates were prepared with plate with egg yolk tellurite emulsion FD 046 (HiMedia) and the plates count agar (Oxoid, Basingstoke, UK) using the standard pour plate were incubated at 37ЊC for 24 h. Typical representative colonies Њ method. One set of triplicate plates was incubated at 37 C for 24 from each plate were picked and subcultured on nutrient agar Њ h for mesophilic counts and the other set at 7 C for 10 days for slants (Oxoid) for further tests. psychrotrophic counts (24). After incubation, colonies were count- Gram-positive cocci isolated as presumptive S. aureus from ed, and representative colonies were isolated, purified, and sub- Baird-Parker medium were tested for coagulase activity using rab- Њ cultured on nutrient agar slants and were maintained at 4 C until bit coagulase plasma PL850 (Pro-Lab, Austin, Tex.). Organisms further characterization. that showed a positive result for coagulase tests were subjected to Biochemical characterization, identification, and confir- API STAPH (bioMe´rieux) for confirmation. The isolates con- mation. A total of 727 colonies were isolated (348 were from firmed as S. aureus were tested for toxin production using SET- plate count agar, 167 from Baird-Parker agar, 111 from deMan RPLA staphylococcal enterotoxin test kit TD900 (Oxoid) accord- Rogosa Sharpe [MRS], and 101 from dichloran rose bengal chlor- ing to the manufacturer’s specifications.
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