Microbiological Examination of Decaying Vegetable (Pumpkin Leaves) Sold at Afia Nine Obiagu, Enugu.
CHAPTER ONE
Aim and objectives of the study
The main aim of this study is to ascertain the microbial load that cause decaying in pumpkin leaves.
The following are the specific objectives of the study:
- To ascertain microbial safety of decayed pumpkin leaves
- To isolate micro-organisms associated with decayed pumpkin leaves
- To Identify the microbial types
- To determine the microbial load of the sampled decayed pumpkin leaves
- To determine the microbial load, of each type
CHAPTER TWO
LITERATURE REVIEW
Introduction
Food plays a vital role in the rise and growth or the fall and decline of a nation. This is because of its effect on the health of the population. Food plays an important role in meeting immediate or urgent food need of people in many cities and towns. Thousands of people depend on food daily. The foods are cheap and easily accessible (Tambekar et al., 2008). Cooked food can be described as the state of food ready for immediate consumption at the point of sale (Eni et al., 2001). It is also defined as food consumed ordinarily in the same state as that in which it is sold by food sellers, hawkers or at local market and does not include nut in the shell and whole, raw fruits, vegetables that are intended for peeling or washing before consumption (Eni et al., 2001). It is a popular option for people ‘‘on the go’’. However, they often come with a lot of risk. This is because cooked foods can be consumed at the point of sale without further cooking or preparation (Castro-Rosas et al., 2007). Cooked food can be raw or cooked, hot or chilled and can be consumed without further heat treatment (Tsang, 2002). Different terms have been used to describe such cooked food. These include: ‘‘convenient’’, ‘‘ready’’, ‘‘instant’’ and ‘‘fast’’ foods.
A general observation of our society shows a social pattern characterised by increased mobility, large numbers of workers and less family-or-home centered activities. This situation has resulted in more cooked foods taken outside home; thus, there is increase in food seller services (Musa and Akande, 2002). The present economic condition has resulted in situation where food vending has become increasingly important in most countries. This contributes to income inflow for individuals involved in selling of these foods (Agwa et al., 2012). The lack of hygiene and sanitation during preparation and marketing of these foods provide ample opportunities for the increase in the number of food-borne pathogens (Desai and Varadaraj, 2009).
In the food matrix, microbes grow in compact ecosystems in which they can exploit metabolites produced by other microbes (Gram et al., 2002). Growth and metabolism of spoilage and pathogenic bacteria produce compounds such as secondary metabolites that can affect the quality of the foods, causing either chemical or microbiological changes in the products (Gram et al., 2002). There is a wide recognition of the role of food in spreading diseases (Marambio et al., 2001). Potential health risks are associated with contamination of foods by E. coli, Salmonella typhi, Pseudomonas spp., S. aureus and Proteus spp. during preparation, post cooking and other handling stages (Garode and Waghode, 2012; Ghosh et al., 2007; Hanoshiro et al., 2004).
Microorganisms are abundant and ubiquitous in nature and their role as causative agents of food-borne illnesses was recognized as far as the 19th century. It was discovered by the end of the 19th century that microorganisms were responsible for a variety of food-borne diseases. Microbial contamination of foods presents challenges that are different from those posed by toxins and physical hazards (Van Ermengen, 1998).
CHAPTER THREE
MATERIALS AND METHODS
Materials Collection of the decayed pumpkin leaves (Fluted pumpkin leaves samples)
Five samples of pumpkin leaves were collected from different food vendors in Afia Nine Obiagu, Enugu. The leaves samples were collected in sterile plates and covered. They were allowed to stay for 72 hours microbial growth.
Sterilization of Materials
All the glass wares used were wrapped with craft paper and fastened with a masking tape and sterilized using a hot air oven at 180ºC for 1 hour. Each culture medium after preparation was stoppered with cotton wool and rapped with an aluminium foil. The flasks were packed into the autoclave for sterilization at 121ºC for 15 minutes. The bench used was also sterilized using alcohol and a cotton wool to swab it so as to free it from contaminants.
Preparation of Media
MacConkey agar, blood agar and nutrient agar media were prepared according to the instruction of manufacturer. Meanwhile, 250ml of nutrient agar, 250ml of MacConkey agar and 100ml of blood agar were prepared.
Measuring of the vegetable leaves Sample for Culturing
One gram of the leaves sample was weighed out using a spatula, weighing balance and a beaker, 9 ml of distilled water was added to the leaves in the beaker and stirred with a glass rod for homogeneity. After mixing, serial dilution was done in other to reduce the bacterial load or concentration so as to obtain a discrete growth on the plate. The level of serial dilution in each was 105. Then, two test tubes were used in the culturing. They were cultured into different media that were prepared. The last two test tubes were used because their bacterial loads were less.
Inoculation and Incubation
From the last two test tubes, 0.1ml of the leaves suspension was plated or inoculated out into the different media plates, their levels of serial dilution was put down on the plates (i.e., the plates were labeled, B(102), C(103), D(104) and E(105). The suspension was spread on the solid surface of the agar medium using a bent glass rod. The plates were inverted when dried and incubated at (37º) for 24 hours in the incubator for growth.
CHAPTER FOUR
RESULTS AND DISCUSSION
Results
The results obtained were based on the examination of the colonial characteristics, gram staining reactions and biochemical test carried out.
Discussion
The important sources of Bacillus species are primary soil and water (Jay, 2003). Thus, these organisms are the predominant spoilage organism of decayed pumpkin leaves because at the onset of spoilage of the leaves, there were always heavy/profuse growth of pure colonies of Bacillus species on the various growth media used. Thus, this corresponds with the spoilage of the individual components of the leaves, as Bacillus species were among the major organism responsible for the spoilage of the leaves condiments.
In the process of inoculating the sour decayed pumpkin leaves on various solid growth media, particularly MacConkey agar and blood agar media,there were often growths of mixed colonies after 24 hours of incubation. Micro-organisms usually isolated were Streptococcus faecalis, Escherichia coli and Staphylococcus aureus. The growth of these organisms especially Streptococcus feacalis and E. coli suggests contamination may be from the water used in washing the leaves or water used in washing of the cooking utensils. The contamination might also result from the handlers, especially in the process of testing the plates used in conveying the leaves to the laboratory.
Faecal contamination was suggested to be the source of these organisms in the leaves since they are of water and faecalis origin. However, the idea of sterilization was introduced to ascertain if these organisms were intrinsic of the soured leaves. Thus, the plates used in conveying the leaves were sterilized by autoclaving at 121ºC for 15 minutes and after placing the leaves on the plates, they were heated to 80ºC to kill the contaminants. These leavess were allowed to spoil and later 24 hours of their inoculation, and incubation, there were neither growth of Escherichia coli, Streptococcus aureus, but a profuse growth of Bacillus species (aerobic spore formers).
CHAPTER FIVE
CONCLUSION AND RECOMMENDATION
In summary, this study has suggested that various organisms spoil decayed pumpkin leaves with Bacillus spp. as the predominant spoilage organism. Since most of the spoilage organisms are able to produce toxins, decayed pumpkin leaves when sour or spoiled is unfit for human consumption. Thus, it is therefore necessary to preserve pumpkin leaves from spoilage by proper refrigeration or freezing.
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