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Download the complete microbiology project topic and material (chapter 1-5) titled THE DIVERSITY OF THE ORGANISMS ASSOCIATED WITH GROUNDNUT PASTE (OKWA OSE). here on See below for the abstract, table of contents, list of figures, list of tables, list of appendices, list of abbreviations and chapter one. Click the DOWNLOAD NOW button to get the complete project work instantly.



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Groundnut is one of the leguminous crops grown worldwide. It contains about 25-30% protein, and 40-48% oil of high quality. Italso contains niacin, thiamine and other Vitamin-B componentsplus 11 of the 13 essential minerals like calcium (Maula, 1985).

In the Philippines, about 92.0% of its total groundnut production is consumed as food, 0.5% is used as seeds and 7.5% for non­food uses. Products prepared from groundnut are flour, protein

Isolate, cheese, and paste for shortening and defatted meal forsnack foods. The seed coat is a source of commercial tannin and thiamine. According to a survey conducted by (Galvez et al., 2002), groundnut paste was the most preferred over the other groundnut products like fried, roasted or boiled and produced and available in all the regions of the Philippines, the most common of which are the flowing types. However the oil separates during storage, and the product needs to be remixed for better eating quality. This separation is a problem due to higher tendency of the product to become rancid.The stabilizer keeps the oil from separating from the groundnut paste and improves texture, increases shelf life, and keeps the groundnut paste fresh which most consumers prefer (Malupangue, 2005).

Groundnut paste (Okwa-Ose) is a semi-perishable product that is subject to a number of microbial, chemical and physical deteriorative changes, which affect the final quality of the finished product. The shelf life is greatly dependent on the quality of groundnut used and the conditions of the groundnut used for making the groundnut paste. Deterioration of groundnut paste arises from putrefaction of protein fraction caused by bacterial metabolism; darkening, which results from an interaction between sugar and protein in the product and; oxidative rancidity that develops in the unsaturated portion of oil when it is exposed to air (Woodroof, 1983).






The scope of the study is embedded on enlightening the public on the need of preserving groundnut paste to avoid deterioration which may lead to the transmission of pathogenic organisms. In order words, if the groundnut paste has been affected by pathogenic organisms leading to its deterioration, such groundnut paste should be discarded and consumption of it avoided in other to sustain healthy living.


This study is aimed at studying the diversity of the organisms associated with groundnut paste (Okwa Ose).


  1. To isolate microorganisms associated with groundnut paste.
  2. To characterize the isolated organisms.


Groundnut paste is a paste made from dry roasted groundnuts. This paste is generally used as a spread on toast or sandwich, garden egg etc. Groundnut paste is a healthy food which is full of nutrients such as protein, carbohydrate, monounsaturated fatty acids, folate, niacin, pantothenic acid, pyridoxine, riboflavin, thiamine, vitamin E, vitamin C, vitamin A, sodium, magnesium, calcium, manganese, phosphorous, selenium, copper, iron and zinc  (Maula, 1985).

Groundnuts, as they are called belong to the family of Fabaceae and species of Arachis hypogea. The plant itself is a small annual plant in which the branch of the flower touches the ground and grows underneath the ground. The plant is thought to have originated in the American continent and through Spanish travelers it was spread all over the world.

Unlike costly nuts such as cashew nuts, pistachio nuts, almonds nut and walnuts, groundnuts are low cost nuts that offers similar benefits (Ritcher et al; 1992).






Shelled peanuts




First stage roasting (20 mins, 150°C)




Deskinning the kernels


Sorting out defective kernels


1st Grinding


Addition of ingredients (salt, ehuru, and stabilizer)


2nd grinding


Groundnut paste






Fig. 5.1 Process flow chart for groundnut paste.

(Sokari, 1991).








People who consume groundnut and groundnut products on a regular basis have an overall higher intake of protein fats (poly-saturated and mono-saturated) fiber, vitamins A and E and many other nutrients (Ritcher et al., 1992).

Groundnuts contain no amount of saturated fats. Apart from Its great taste, groundnut paste has vital nutrients that are essentialfor the body. Some of the healthy benefits of groundnut pasteare;

Rich Source of Protein

Groundnut pastecontains high amount of proteinto30g. Proteins that we eat are broken down into amino acids which are then utilized in each and every cell for repairing and building the body. A cell divides by using protein as its building materials(Ritcher et al; 1992).

Lower Cholesterol Level with Unsaturated Fats

The amount of fats contained in groundnut paste is almost equal to that of the fats found in olive oil (Ritcher et al., 1992).

Groundnut paste contains both polyunsaturated and monounsaturated fats. As these fats are not saturated, they are good to consume without putting the heart at risk. The unsaturated fats in groundnut pastes help lowering bad cholesterol or low density lipoprotein and promote the circulation of good cholesterol or high density lipoprotein.

Reduce Risk of Type II Diabetes

Consuming groundnut paste can be beneficial in reducing the risk of type II diabetes (Maula, 1985).

Unsaturated fats have been noted to improve insulin sensitivity. Research into groundnut paste consumption and diabetes showed that higher intake of groundnut paste and other nuts lowers the risk of type II diabetes.

Source of Vitamin

Groundnut paste contains many vitamins that are good for our body to function property. Vitamin a found in groundnut paste is helpful for eyesight, while vitamin (help to boost the immune system and heals simple ulcers faster. On the other hand, vitaminE found in groundnut paste is also a very important micronutrient needed by our body to dissolve complex fatty acid structures, fat blockage and arteries.

Anti-oxidant Properties

Groundnut paste contains antioxidant properties due to the presence of folate, niacin pantothenic acid, pyridoxine, riboflavin, and thiamine. One of the antioxidant found in groundnut paste is resveratrol.Resveratrol is a polyphenol antioxidant which has been found effective in controlling certain types of cancers, heart diseases, Alzheimer’s disease, viral and fungal infections and degenerative nerve diseases (Ritcher et al; 1992).

Anticancer Properties

Groundnut paste contains B-sitosterol, a polyphenol which has a great property of fighting against cancer, particularly colon, prostate and breast cancers.

Groundnut and its products such as groundnut oil, groundnut paste, is ideal sources of polytosterol (Ritcher et al; 1992).



Good Source of Magnesium

Magnesium is one of the most needed elements in our body. It presence in our body helps in more than 300 biochemical reactions which are extremely important for our survival-Groundnut paste is a good source of magnesium (170mg/100g). This makes up for 42% of the daily recommended value of magnesium. Magnesium has an important role to play in muscle,bone and immunity development in the body. Magnesium also helps in regulating the blood sugar levels and blood pressure (Ritcher et al; 1992).

High in Potassium

Groundnut paste contains potassium (70mg/100g) that acts as an electrolyte and is a fluid balancing element in the body. In companion to sodium which directly puts pressure on the cardiovascular system in the form of hypertension, potassium does not put any pressure either on the blood or on the cardiovascular system. In effect, potassium is a heart-friendly element which is found in high qualities in groundnut paste (Ritcher et al; 1992).

Reduces Risk of Gallstones

Gallstones is one of the major health risks in developed countries which is caused by overweight, crash diets, certain types of cholesterol drugs and birth control pills. A study on groundnuts and nut consumption and the risk of gallstones was taken up. The results of the study showed that over a period of 2 decades, women who consumed groundnut paste and nuts on regular basis had reduced their risk of developing gallstones (Ritcher et al; 1992).

Dietary Fiber

Groundnut and groundnut paste are high in dietary fiber. One cup or approximately 125g of groundnuts and groundnut paste has 12g and 20g of dietary fiber respectively.

Dietary fiber is one of the important parts in our diet as lack of dietary fiber can lead to several health problems and diseases such as constipation, diabetes, cholesterol and heart diseases (Ritcher et al; 1992).


Aflatoxins are natural occurring carcinogenic by-product of common fungi on grains and other crops, particularly maize andgroundnuts. They pose a significant public health risk in many tropical developing countries and are also a barrier to the growth of domestic and international commercial markets for food andfeed.It is also reported that domestic commodities most susceptible to afflatoxin are peanuts, corn, cotton-seed and tree nuts (almondspeacans, walnuts).Moreover, aflotoxin is a toxin class I carcinogenic by-product of fungi that colonize maize and groundnuts among other crops. However more than 4.5 billion people (64% of the world inhabitant) in developing countries may be chronically exposed to aflatoxin in their diets.Aflatoxin is also reported as a class I carcinogenic that contributes 28% of all new liver cancers. It also increases TB. The stunted children were found to have 30-40% more aflatoxin in their blood than those with normal body weight. The percentage of stunted children was reported to be 46% for those less than five years of age in Malawi.As climate stiffs, so do the complex communities of aflatoxin-producing fungi. This includes changes in the quality of aflatoxin-producers in the environment and alterations to fungal community structure (Carpenter; 1972).

Fluctuation in climate also influence predisposition of hosts to contamination by altering crop development and by affecting insets that create wounds on which aflatoxin-producers proliferate. Aflatoxin contamination is prevalent both in warm humid climates and irrigated hot deserts. However, rain and temperature influence the crop phases different with dry hot conditions favoring aflatoxin contamination during the crop development and warm, wet conditions favoring it after maturation.

Post-harvest contamination may take place due to a number of factors that include damage of kernels, crop stresses such as drought and insect infestation and inadequate drying and storing facilities and soaking of shells to simplify hand shelling of groundnuts which induces ideal conditions for Aspergillus infection.

The Association of Official Analytical Chemists (AOAC) member used a variety of methods in determining aflatoxins in groundnuts which include the Enzyme Linked Immunosorbent Assay (ELISA) Solid-phase Radio Immunoassay (RIA).

High pressure liquid chromatography, different thin layer chromatography and minicolumn detection methods (Carpenter; 1972).


There are different groups of microorganisms that may be found in groundnut paste.  The presence ofany particularspecie of organism is based on the source of contamination. Microorganisms that may   be found   in   groundnut   paste are of different groups which include fungi and bacteria (Carpenter; 1972).


The major group of fungi found to be responsible for the spoilage of groundnut and groundnut products to Aspergillus.




Aspergillus species are widely distributed and caused numerous kinds of spoilage.  They rot figs and dates decay tobacco andcigars spoil nuts and bread and grow on leather and clothing in humid weather, (Carpenter; 1972).

The specie of Aspergillus found in nuts and nut products is the Aspergiilus Fiarus. Aspergifius Flarus was found in the infected peanut meal, together with alcohol-extractable toxins termed aflatoxin (Prescott et al; 2008). This particular specie of fungi can be a contaminant to groundnut paste especially when the groundnut used for the preparation of the paste is already contaminated by these organisms. The Aspergilius Flarus is a facultative fungus. They invade plant tissues directly or attack tissues that have been predisposed by stresses such as dry weather or damages caused by insects, nematodes, natural cracking and harvest equipment (Pettit, 1984).

The yellow mold fungus, Aspergiilus Flarus is commonly found in the seed of both rotten and apparently healthy pods of groundnut. Many stains of this fungus are capable of producing aflatoxins that render the seed unacceptable due to high toxicity for human or animal consumption (Reddy and MacDonald, 1983). Aflatoxins are carcinogenic and produced by the Aspergiilus group of fungi that have been identified as B1, 82, G1 and G2 (Pettit, 1984).There have been many studies to improve groundnut paste as a food commodity. Most of these address such problems as

  1. The prevention of oil separation on the surface,
  2. Improvement of smoothness and spreadability,
  3. Improvement of the consistency and stickiness,
  4. Development of a type that can be blocked and sliced,
  5. Enhancement of flavor by the addition of optional ingredients,
  6. Effects of added fats, carbohydrates and stabilizers on the final quality, and;
  7. Prevention of rapid deterioration of groundnut paste during storage. All these problems define or set the limits of the shelf-life stability of groundnut paste. Color along with other quality, safety and nutritional factors have achieved a more preeminent position in the minds of the consumers. This has necessitated a greater concern on the   part of the food manufacturers in assessing the color of foods. Mulego et al. (1990) reported that groundnuts that are water blanched at 90°C   for   10   minutes   are   lighter   in   color.   The   color   of groundnut paste is basically affected by roasting time. The most serious   problem   of   natural   groundnut   paste   is   the tendency of the oil to separate. Oil is released during the grinding of peanuts. The improvement of emulsion stability in groundnut paste is characterized by the absence of twolayers of oil and meal phase during ordinary conditions of storage, and improved texture, consistency, spreadability, flavor, color as well as nutritional value. Without stabilizers, the groundnut meal settles at the bottom and forms a hard layer while the oil remains on top (Aryana et al, 2000). Several efforts have already been made to answer the problem. Among the solutions arrived at and researches done to address this particular problem were those cited by Gills and Resurreccion (2000) and Woodroof (1983), which include special grinding of roasted groundnuts, the heat treatment of paste after packaging, and the incorporation in groundnut paste of various substances, including water, honey, glycerin, mono- and di- glycerides, and vegetable oils hydrogenated to various degrees of hardness. Some groundnut pastes were stabilized by incorporating into them commercially hydrogenated groundnut oil (m.p. 14S°C) and iodine value of eight (Mitchell, 1950). Other commercial stabilizers incorporated in groundnut oil, are hydrogenated groundnut oil, and salt (Holman and Quimby, 1950).

Stabilizers used for groundnut paste butter are partially or fully hydrogenated vegetable of Is. Hydrogenated oils are usually suggested as stabilizing agents for groundnut paste because of their efficient homogenization and crystallization. The use of unhydrogenated palm oil has also been studied for its stabilizing action on groundnut paste (Gills and Resurreccion, 2000;Hinds etal,1994), Other known important factors contributing to the protection of the groundnut paste from oil separation that have been cited in literature are the storage temperature and the temperature at which the stabilizer was incorporated.

Groundnut (Arachis hypogaea L.) occupies an important position in the developing countries. The major groundnut producing countries are India, China and the United States, It was introduced in Nigeria in the 16th century and it has been estimated that 1.4 million hectare is cultivated for groundnut in Nigeria.

Groundnut paste is a food paste made primarily from ground dry roasted groundnut. Groundnut paste has been frequentlyassociated with food illness in which initial contamination is traceable to food handlers. Numerical epidemiological studies and reports have implicated foods of ready to eat origin as the major vehicle associated with food-borne pathogens. Person to person transmission has also been described (Sokari, 1991). Escherichia coil, Staphylococcus aureus, Yersinia enter-coilitid, Salmonella spp yeasts and moulds have been used to assess the microbiological safety and sanitation conditions during processing and keeping quality of groundnut paste and groundnut products (Consumer Report, 2009).

In Nigeria, groundnut paste is produced traditionally on a small scale and as such has been given little or no attention on the microbiological   quality   and   safety   of   traditionally   processed Nigeria groundnut paste is lacking in literature. Groundnut (Arachis Hypogaea L.) is an essential item in several confection any products and supplementary feeding programs. It is one of the most important sources of edible vegetable oils and cultivated   principally for this purpose. In Nigeria,   groundnut is widely consumed as boiled, roasted or dried nut as a complimentwith banana, garden eggs etc. It is high in colonies due to its highfats and protein contents.

Okwa-Ose,   as   locally   known   and   called   in   Eastern   Nigeria,   is   acohesive, comminuted food product prepared from clean, soundand  shelled  groundnuts   by grinding   roasted   mature kernels  towhich salt, pepper and ehuru spice (Mondora Myristica) are addedas a seasoning and flavoring agents. It is a common householdproduct that is conventionally eaten with garden eggs andkolanuts, especially in traditional ceremonial settings.

Okwa-Ose   has   a   long   shelf   life   at   room   temperature   and   isconventionally prepared only with groundnuts.


Mycotoxins are produced by various fungi and are consideredpoisonous contaminants in susceptible foods and feeds. Aflatoxinhas been identified as the most toxic mycotoxin associated withgroundnuts, and hence the toxicity, and measures to manage andprevent contamination by aflatoxin has been discussed in detail inthis review. In addition to aflatoxins, another commonly occurringnatural contaminant of groundnut is the mycotoxin cyclopiazonic acid,produced by several species of Penicillium and Aspergillus (Lansden& Davidson, 1983). Cyclopiazonic acid is a potent inhibitor of thereticular form of the Ca2þ ATPase pump (Plenge-Tellecheaet al., 1997). The compound is not considered to bea potent toxin in humans due to low oral LD50 values, in the range of30e70 mg/kg in rodents (Antony et al., 2003;Nishie et al., 1987). On peanuts, the natural level ofcontamination by this toxin is only 6.5 ppm (Lansden & Davidson,1983), thus, the compound is toxic to humans only when it isconsumed at levels that exceed the natural level of intake of thetoxin (Burdock & Flamm, 2000; Van Rensburg, 1984). Further, inmost cases, both cyclopiazonic acid and aflatoxin, both produced byAspergillus flavus, are present concurrently (Urano et al., 1992), andthis effectively disguises the presence of cyclopiazonic acid. A studyconducted by Van Rensburg (1984) concluded that in the presenceof usual hygiene and aflatoxin control practices in peanuts, nofurther control or screening measures for cyclopiazonic acid iswarranted (Pettit, 1984).

Bacteria Found in Groundnutn Paste

In today’s food community there is the possibility for more than 200 diseases to be transmitted via consumed food products.  The Centers for Disease Control and Prevention estimates that there are approximately 76 million cases of food borne illnesses and disease each year.  Of the reported cases 325,000 conclude as hospitalizations and there is an upwards of 5,000 deaths related to food borne illness each year with the occurrences appearing in the very old, very young or in those individuals who have a suppressed immune system (Pettit, 1984).

Salmonella spp. was the organism of concern during the Peter Pan recall.  Salmonellosis is one of the most common food borne diseases as it produces an enterotoxin that interrupts the natural balance of the secretion of fluid and electrolytes.  There are over 2000 species of Salmonella, all are considered to be pathogenic, and they have shown to be resistant to many antibiotics, freezing, and drying.  (Fung, 2007).

A hypothesis for the project addresses that organic and all-natural products would have higher counts of microorganisms as their processing procedures do not included additives or preservatives that would assist in reducing the number of organisms that could survive in the peanut butter environment.Similar to groundnuts, Salmonella can survive in groundnut paste despite its low aw. Burnett et al., (2000) found that with an initialinoculum of 5.68 log CFU/g, Salmonella was found at greater than3 log CFU/g after 1 week at 5 _C and 21 _C. Even after 24 weeks,approximately 2 and 1 log CFU/g Salmonella was detected at 5 _Cand 21 _C, respectively. A 3-strain cocktail of SalmonellaTennessee inoculated in peanut butter survived for 2 weeks at 4 and 22 _C(Park et al.,2008). Further, Kenney and Beuchat (2004) reportedthat another pathogen, Listeriamonocytogenes could alsosurvive for 24 weeks in groundnut paste at 20 _C with an initialinoculation level of 4 log CFU/g. Although no illness was reported,potential contamination by L.monocytogenes also promptednationwide recalls of certain brands of peanut butter, and peanutbutter & jelly sandwiches in 2010 and 2011 respectively (US FDA,2010; US FDA 2011a). Results of these studies suggest that anypathogen that contaminates peanut butter after heat processingcould remain in the product throughout its shelf life (Pettit, 1984).



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