Ice cream is a nutritionally enriched congealed dairy product consumed by all age groups particularly children, during summer (Sharif et al., 2005). The ingredients of Ice cream may be various combinations of milk, cream, evaporated or condensed milk, dried milk, colouring material, flavors, fruits, nuts, sweetening agents, eggs and eggs products stabilizer. Any of these may account for the various specific species of bacteria (Yaman et al., 2006).
Ice cream is a nutritious food for human and also an excellent medium for the growth of many microorganisms some of which may cause diseases in human beings e.g. Cholera, typhoid, bacillary dysentery.
Contaminated ice cream causes several outbreaks of gastrointestinal diseases in a number of countries in Asia, Europe and North America (Dijuretic et al., 1997; Dijuretic and Wall, 1996; Chug, 1997).
In England and Wales two outbreaks of S. enteritidis phage type 4 infections were reported in 1990 and 1995 due to consumption of ice cream (Hennessy et al., 1996).
Quality of ice cream depends on extrinsic factors that include manufacture procedure, as well as intrinsic factors that include the proportion of ingredients used. Primary sources of microbial contamination of ice cream include water and raw milk, whereas secondary sources include flavoring agents, utensils and handling.
Possible sources of these microorganisms in ice cream have been reported to include raw materials used for the composition of ice cream-mix, such as milk and milk powder, cream, flavouring and colouring substances and sanitizer (Verma, 1972; Bathla and Rao, 1973) and from contaminate air during processing (Gomez, 1969).
In Gilgit town ice cream is manufactured on small scale by using the dried and raw milk and sale in the city at retail outlets.
The aim of this study was to determine the bacteriological quality of commercially soled ice creams in Gilgit town and their potential risk to public health.
MATERIALS AND METHODS
Collection of samples: Twenty ice cream samples (cups) were randomly
collected from eight different areas/location of Gilgit town (Fig.
1). The collected samples were immediately shifted to the laboratory of
Department of Biological Sciences, Karakoram International University, Gilgit
in a cold box.
Microbiological analysis: Serial dilutions: In serial dilutions 1 gram of ice cream was aseptically transferred into 9 ml of distilled water and homogenized by vertex. Subsequent serial dilutions were made up to 105.
Culture of samples: For aerobic enumeration of colonies 1 ml sample was cultured on Nutrient agar (Oxoid) and incubated at 37oC for 24 h. The following day the total number of colonies were counted and read morphology, for their identification. Separated colonies were sub culture on freshly prepared Nutrient agar, MacConkey (Oxoid) and Salmonella Shigella agar (Oxoid) plates. Further identification of the Gram negative colonies was performed by biochemical tests such as production of urease, utilization of sugars, carbon and production of indole, H2S gas, oxidase and on the basis of motility of the bacteria (WHO Manual, 1987) and the gram positive bacteria colonies were identified by coagulase and catalase tests.
The total viable counts in samples of the eight locations of Gilgit town are
presented in Table 1. The samples of all the areas show heavy
contamination of bacteria ranging from > 2.2x103 CFU/g to 6.8x104
CFU/g. The mean results indicate that the highest contamination was found in
Punial Road ice cream samples 6.6x104 CFU/g followed by Park Hotel
link Road 3.6x104 CFU/g, Nasim Cinema Bazaar 2.5x104 CFU/g,
Hospital Road 8.5x103 CFU/g, Jamat Khana Bazar 7.7x103,
Yadgar chowk 4.5x103 CFU/g, Shaheed Milat road 3.4x103 CFU/g
and Khazana Road ice cream samples had 2.4x103 CFU/g.
collection area of Ice cream samples in Gilgit city
Table 2 show the occurrence of different bacteria in the
investigated ice cream samples. Escherichia coli is the commonest organism
and was isolated from all the samples of all the areas, Klebsiella sp.
was isolated from 17 samples except sample numbers PRIS2 PRIS3
of Punial road and sample no. NCBIS1 of Nasim Cinema Bazar.
Overall isolation of Proteus was from 09 samples i.e. in all the samples
of hospital road, in one sample of each of Punial road, Khazana road, Jamat
khana Bazar, Park hotel link road Yadgar Chowk and Shaheed Millat road. Bacillus
sp. were isolated from 4 samples, i.e. in one sample each of Punial road,
Khazana road, Jamat Khana Bazar and yadgar chowk. Salmonella was isolated from
3 sample i.e. one each from Punial road, Nasim Cinema road and Park hotel link
road. Staphylococcus sp. was in 10 samples i.e. in two samples of Punial
road, one sample each of Hospital road and Park hotel link road, in 02 samples
each of Jamat Khana bazar, Nasim Cinema bazar.
Table 3 shows incidence of different bacteria isolated during the investigation.
The infestation ratio were: Escherichia coli 20 (20) 100%, Klebsiella sp. 17 (20) 85%, Proteus 9 (20) 45%, Bacillus sp. 04 (20) 20%, Salmonella 03 (20) 15%, Staphylococcus sp. 10 (20) 50%.
Viable count (CFU/g) of various bacteria isolated from ice cream samples
The results obtained in this study represent the current status of microbiological quality of ice cream being sold in Gilgit town. All the analyzed ice cream samples (n = 20) showed heavy contamination of notable bacteria (E. coli, Klebsiella, Proteus, Salmonella and Staphylococcus) which indicates fecal contamination. The presence of this high level of fecal coliforms contamination represents a public health risk due to the possible presence and transmission of pathogens such as enteropathogenic Escherichia coli, Hepatitis A virus, poliomyelitis virus, while Entamoeba histolytica may also be present in the ice cream (Arias and Windrantz, 2000). The mode of transmission of all these bacteria is fecal-oral route and or via common house flies.
The results suggest negligence such as poor sanitation during the preparation and/or storage of these products. These include the observed dirty premises and utensils used, the use of bare hands in preparing the products (personal communication with the handlers).
In this study three most important genera Escherichia coli, Salmonella
sp. and Staphylococcus sp. were also isolated. Escherichia coli
strains EPEC, ETEC, EHEC and invasive strains are pathogenic to children
of <5 years. Salmonella is still the most important acute agent causing
food bone diseases (Tood, 1997). Consumption of ice cream contaminated with
enteropathogenic bacteria such as Salmonella has been the cause of several
outbreaks (Hennessy et al., 1996). In the Northern Areas of Pakistan
Gilgit Ahmed et al. (2005) isolated twenty one Salmonella typhi
from 585 suspected gastroenteritis patients. Staphylococcus, which is
commonly Staphylococcus aureus when transmitted from man and animal,
can lead to staphylococcal food poisoning as a result of growth of the organism
and release of enterotoxin into the food.
Species isolated from different ice cream samples from areas
and percentage infestation of bacteria isolated from Ice cream
Entrotoxin production and secretion occurs especially when ice cream products
are not hygienically prepared and stored. The presence of starch and protein
encourage enterotoxin production by the microorganisms (Wistreich and Lechtman,
1980). The possible source (s) of this organism in ice cream could be from human
nose where it is commonly found; hands, skin and clothing of handlers (Hobbs
and Golbert, 1982). Coughing, talking and sneezing produce droplets, which could
settle on, ice cream during transportation In Gilgit most of the population
produce homemade ice cream for domestic use as well as commercial purposes by
using the ingredients dry milk, sugar and water.
The presence of fecal coliform indicates post-treatment contamination which may either come from water, lack of personal hygiene of the ice cream manufacturer, utensils used for ice cream and distribution environment. A study conducted by Ahmed and Shakoori (2002) reported 640-683 E. coli colonies /100 of drinking water. The ice cream manufacturers use the same water for the preparation of ice cream as well as for washing of their hands and utensils. Once the ice cream become contaminated, freezing temperature later could not make the product safer (Jay, 1996).