In the food manufacturing processing there are a lot of problems that affect safety and health food, drink then human. The most important problems food poisoning and spoilage that includes any changes in physical, chemical, sensory and appearance of food because affect the validity of food and nutritional value as well as human health. However, these changes occur by contamination and raise the microorganisms in food.
Food poisoning and foodborne diseases outbreak widespread causes human illnesses and sometimes gets lethal disease all over the world. It is occurs by consuming contaminated water or food by spoilage and pathogenic microorganisms particularly bacteria because they are widespread and anywhere. Also, need simple requirements for growth. Although there are huge numbers of bacteria species, some of them are causative whether food poisoning or foodborne diseases. The most important types of bacteria responsible for foodborne diseases Shigella spp.
Shigella spp are genus of bacteria Gram negative, rod shape, non motile, facultative aerobes and non spore forming, members of Enterobacteriaceae family. Classification in four main species: S. boydii, S. dysenteriae, S. flexneri and S. sonnei. These species found in the human intestine, cause foodborne disease and proliferate whether through food or by human. The most important of these species that cause severe symptoms is S. dysenteriae.
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The disease characteristics associated with Shigella spp
'Bacterial foodborne diseases can cause illness by ingestion of small numbers of microbes usually incubation period is long in range 1-25 days, frequently transmitted and cause illness by contaminated water. Whereas, bacterial food poisoning can cause illness either by a huge numbers of microbes or by production of toxin in food, incubation period is short in range 2-36 hours, food poisoning does not occur by contaminated water'. (Trickett, 39)
'Few microbes of Shigella spp (approximately 10-100 bacterial cells) can cause illness compared to other types of microorganisms. Also, transmission by food is less common compared with other microbes' (Navia). 'Shigella spp is a markedly contagious microbes and one of the most important human pathogen that causes outbreaks of bacillary dysentery particularly in the developing countries' (Bing). So, have been recorded a huge number of Shigellosis waterborne outbreaks all over the world. 'Roughly 165 million cases and over one million deaths are reported annually all over the world by bacillary dysentery' (Sharma), '91% from this incidences occurring in developing countries' (Navia). 'This endemic is a severe ulcerative infection in human intestine and becomes more serious' (Bing) 'among infants and young children less than five years' (Vijaya). 'The most prevalent species are S. flexneri and S. sonnei that causing about 60% and 15% of incidences, respectively'. (Osorio)
'Dysentery occurs either via Shigella (bacillary dysentery) or by amoeba (amoebic dysentery). Amoebic dysentery is' (Bing) 'infrequent in Europe and caused by a protozoa. Although, all species usually can produce acute illnesses. Shigella dysentery occurs less than other organisms but causes a more acute illnesses and epidemic' (Osorio) 'incubation period 1-7 days. Clinical characteristics include severe abdominal pains, fever, inflammatory, lethargy, convulsion, nausea, colic, vomiting, watery diarrhea and dehydration. Characterized by mucus and bloody feces. Can be transmitted via wastewater or by consuming contaminated food, often directly by person poor personal hygiene (fecal oral transmission) and insufficient sanitation. Also by vegetables or fruit harvested from a field irrigated with contaminated water, can also be transmitted by insects breed in human sewage. So, it is presence in water indicates contamination with human feces. Infectiousness acquired by contaminated food, drinking or swimming in contaminated water' (Bing). 'Once ingested the organisms invade the gastrointestinal epithelial cells then multiply and spread to uninfect gastrointestinal cells' (Peng). 'Shigella spp can survive in environmental water'. (Sharma)
Isolation and confirmation of the presence of Shigella spp
There are several media have been designed for the growth of different microorganisms. These media include:
Eosin Methylene Blue Agar (EMB) designed for detection, isolation and confirmation of Gram negative coliforms, contains sucrose and lactose with two dyes.
MacConkey agar is a culture media use for growth and isolation of Gram negative bacteria by lactose fermentation, contains bile salts and crystal violet dye (to prevent Gram positive bacteria), neutral red dye, peptone and lactose. Use peptone instead lactose to raise the rate of pH and increase formation of colorless colonies.
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Hektoen Enteric (HE) Agar is a culture media use to growth and isolation of Gram negative microorganisms and recovers fecal microbes particularly in Shigella and Salmonella. Contains dyes, bile salts, H2S and indicators of lactose fermentation.
SS Agar. Salmonella Shigella Agar
Steps to isolate microbe:
'Suspend the powder 75 g in one liter purified water and mix thoroughly
Heat to dissolve, but do not overheat or autoclave
Cool 45-50Ù’ C and use
Test samples of the final product
Incubate plates at 35Ù’ C for 18-24 hours, protected from light
Shigella appear fair to good growth and color green colonies'
The recovery of Shigella spp
'This study developed approach for rapid detection of Shigella spp and other pathogens by combining immunocapturing of the microbe and a universal primer PCR for enhance test sensitivity. In this study used S. dysenteriae1, S. sonnei, S. boydii and S. flexneri 1a,2a,3a,4,5 and Y . Were grown separately at 37Ù’ C in L-broth media. Then put it in polystyrene 96-well plates and use carbonic acid buffer 0.05 (pH9.6) at 4Ù’ C for 18 hour. Then washed with phosphate-buffered saline (0.05% Tween 20) and incubated at 37Ù’ C for 1 hour. Then washed and wells were incubated with distilled H2O and heated at 100Ù’C for 5 min to denature the bacterial DNA. Then centrifuged for 20 min. then washed with sterile 0.85% NaCL. Then dissolved in sterile distilled H2O and heated at 100Ù’C\5 min to denature the bacterial DNA. Each UPPCR mixture consisted 2.5 PCR buffer, 3mM MgCl2 and subjected to 40 cycles of 94Ù’C for 1 min (denaturation), 51Ù’C for 1 min (annealing), and 70Ù’C for 2.5 min (extension).
Neutralization and coating antibody tests were used to check the specificity of UPPCR. In the neutralization test cell containing 20CFU were mixed with equal volumes of monoclonal antibodies against S. dysenteriae prior to incubation at 37Ù’C for 1 hour and added to wells. Coating test monoclonal antibodies against S. dysenteriae were replaced with monoclonal antibodies against hepatitis B surface antigen as a coating antibody. The new approach described increased sensitivity the epidemiological and diagnosis of S. dysenteriae. Also, can be used directly to detect microbe in environment and can be obtained the results within 5 hours'.