Shigellosis, also known as bacillary dysentery, is a bacterial disease caused by one of four species of genus ShigelIa. These include ShigelIa dysenteriae, Sh. sonnei, Sh. flexneri, and Sh. boydii. It is a common enteric disease, often acute and self-limiting, in almost all parts of the globe. It is characterized. by diarrhoea, lower abdominal cramps, and tenesmus.. The diarrheal stool often is mixed with blood and mucus.
CELLULAR MORPHOLOGY
The shigellae are short straight rod-shaped bacteria occurring singly. They are nonmotile, nonsporing, and noncapsulated. They are Gram-negative.
CULTURAL CHARACTERISTICS
The shigellae are facultatively anaerobes. They can grow in a temperature range of 25 to 42°C, optimum being 37°C. The growth occurs at slightly alkaline pH. As far as nutritional requirements are concerned, the shigellae can grow on simple media. They are chemoorganotrophic, having both respiratory and a fermentative type of metabolism. Several culture media, both differential and selective, are available for the cultivation of these bacteria, such as EMB Agar, Salmonella-Shigella agar, MacConkey agar, brilliant green agar, xylose lysine deoxicholate (XLD) agar, Hektoen enteric agar, bromothymol blue lactose agar, etc. Gram. negative (GN) broth is highly recommended as an enrichment medium for primary isolation of Shigella.
On nutrient agar the shigellae produce small, round, smooth, entire, raised, and translucent colonies.
BIOCHEMICAL CHARACTERISTICS
The shigellae are anaerogenic bacteria, except biotype 6 of Sh. flexneri. The four major species can be differentiated from each other on the basis of biochemical activities.They can be classified on the basis of mannitol fermentation into two
groups as shown in the following.
* Mannitol fermenting shigellae This group Includes Sh. flexeneri (excluding biotype 6) , Sh. sonnei, and Sh. boydii.
* Mannitol nonfermenting shigellae. This group includes all the biotypes of Sh. dysenteriae.
Though the shigellae are lactose nonfermentors, Sh. sonnei characteristically terments lactose when incubated for 48 hours.
ANTIGENIC CHARACTERISTICS
Since the members of the genus ShigelIa are nonmotile bacteria, thus differing from those of the genus Salmonella, H antigens are absent. In accord with the other members of the family Enterobacteriaceae, the complex lipopolysaccharide of the cell wall is responsible for the characterization of the O antigens of various members. The specificity of this antigen depends on the polysaccharide side chains of the complex. On the basis of differences in O antigens, one can classify the genus into four serogroups, namely A, B, C, and D. The serogroup A includes Sh. dysenteriae, B includes Sh. flexneri, C includes Sh.boydii, andCLINICAL MANIFESTATIONS
The onset of shigellosis is acute, with fever, malaise, abdominal pain, and watery diarrhea. As the disease progresses, bloody diarrhea with mucus, tenesemus, fecal urgency, and severe cramping abdominal pain becomes more prominent. Disruption of the normal motor function of the intestine is main cause of tenesemus and cramps. Nausea, vomiting, headache, and convulsions (in children) may occur. Fever is developed as a consequence of the action of the pyrogens released into the blood from the inflammatory reactions. The patients become weaker and more dehydrated.
Shigellosis caused by Sh. dysenteriae is more serious than those produced by other ShigelIa species. The fever is much higher, cramps are more severe, and diarrhea is more frequent and voluminous.
COMPLICATIONS
A patient of shigellosis may get involved in certain complications. These complications may be mild, such as arthritis, conjunctivitis, and morbilliform rash, or life-threatening, such as colonic perforation, shock, septicemia, vascular collapse, and hemolytic uremic syndrome.
LABORATORY DIAGNOSIS
The diagnosis of shigellosis depends on the isolation of the etiological agent from the stool of the patient and its identification. The rectal swabs can also be used for this purpose.
ISOLATION
Although the preferred method for maximal recovery of Shigella from clinical specimens is direct plating on agar media, enrichment of stool specimens in Gram-negative (GN) broth has been reported effective. Salenite broth may also be used in isolating Sh.sonnei. XLD agar is a selective and differential medium for ShigelIa. It contains xylose as differentiating carbohydrate, and since most of the Shigella strains do not ferment xylose, they appear as colorless colonies on the plate. However, Shigella isolates, which ferment xylose rapidly, can be missed on this medium. Therefore, additional plating media containing lactose as differentiating agent (such as EMB and MacConkey agar) should be used in conjunction with XLD agar.
IDENTIFICATION
The isolated colonies typical of Shigella should be identified on the basis of biochemical reactions. For this purpose the colonies may be transferred to TSI agar. Isolates whose TSI reaction is aikaline/acid with no H2S or gas should be screened serologically with antisera to ShigelIa 0 groups A, B, C, and D and inoculated to urea agar and motility medium. Urea-positive or motile cultures can then be discarded. lsolates that are nonmotile, anaerogenic, and urea-negative with other characteristic biochemical reactions may be considered presumptive ShigeIla species. Presumptive positive culture should be tested further biochemically.
THERAPY
The first consideration in the therapy of shigellosis is correction of the patient’s hydration abnormalities. For this oral fluid replacement is necessary. The rapidly rising fever that often results in convulsion in the young must be treated symptomatically. Appropriate application of antibiotics appears to be effective at reducing both the clinical symptoms and the duration of excretion of the organism. Treatment may be withheld in mild cases but is definitely indicated in severe cases or when the risk of transmission is high. Ampicillin, trimethoprim-suifamethoxazole, ciprofloxacin, and tetracycline are effective for sensitive strains.
CONTROL
The most effective method of controlling shigellosis, as in salmonellosis and many other enteric diseases, is to develop a safe water supply and an effective means of feces disposal. A successful control is possible by measures within the household and the local community. The patients, particularly children, in acute stage of dysentery must be isolated. lnfected cases should be restricted from handling food. All the objects used by the patients, such as thermometer, toilet seats, bed sheets, clothing, etc., must be appropriately disinfected. Because of the low numbers of the organisms needed to initiate an infection, asymptomatic carriers also contribute in the transmission of the disease.
PREVENTION
Shigellosis can be prevented by attention to hand washing to minimize person to person transmission and by protection of the water supply from human fecal contamination. Good individual and collective hygienic measures are proved fruitful in preventing shigellosis.lnsects, including flies and cockroaches, should also be controlled by the use of insecticides as they play very important role in the transmission of the disease.
SHIGELLA VACCINES
The severe clinical syndrome caused by Shigella species (particularly Sh. dysenteriae), their propensity to pandemic spread, and their resistance to most relevant antibiotics make a vaccine to prevent shigellosis a high priority. WHO has also placed Shigella vaccine development first in line of its priorities. Regretably, no Shigella vaccines seem to be ready for clinical trials.
The most important ShigelIa strains to be targeted for vaccine development are Sh. flexneri 2a, Sh. dysenteriae 1, and Sh. sonnei. However, the possible emergence of new serotypes has been emphasized. The emergence of Sh. flexneri serotypes 1, 3, and 6 has been observed in many countries. ln 1981, Formal prepared a Sh. sonnei vaccine by inserting into attenuated S. typhi. Ty21a the plasmid of Sh. sonnei that contains genes for the synthesis of the O antigens. The resultant hybrid strain, 5076-1C, expresses both S. typhi and Sh. sonnei O antigens. However, because of lot-to-lot variation in protective efficacy, controlled field trials were not undertaken. Another hybrid strain was prepared in 1990 by Format that expressed O antigens of Sh. flexneri. Linderberg prepared a vaccine comprising of a strain of Sh. flexneri with deletion of two genes that are necessary for the bacterium to proliferate in man. But all these efforts did not prove successful in preparing an effective vaccine that can immunize against different virulence factors necessary to produce disease.
Recently, two prototype attenuated vaccine strains of Sh. dysenteriae1 and Sh. flexneri 2a have been developed in the United States. The two strains are safe, highly immunogenic in stimulating lgA antibodies. This vaccine is in the preliminary stages of clinical trials. Another attenuated vaccine comprising of mutant Sh. flexneri has been developed in France. Clinical trials to determine its efficacy initiated in 1996 are currently ongoing. A vaccine has been developed in Sweden that utilizes auxotrophic mutant of Sh. flexneri Y strain. This vaccine was shown to be safe and immunologically potent enough to initiate antibody production against lipopolysaccharide (LPS) of the bacterium. Protection elicited by this vaccine remains to be tested in humans.
Another approach, based on expression of LPS in a live attenuated vector, is under development in Mexico. A similar approach has been taken in Switzerland. ln this case, a hybrid strain of cholera bacilli expresses both host-encoded lnaba and Sh. dysenteriae type 1 O antigens. The results of human trials are still awaited.
A novel approach has been taken in the United States. Parenteral vaccination with the non-covalent complexes of O-polysaccharide and ribosomal particles from Shigella induces a very powerful immune response in experimental animals. After successful clinical trials in animals, an extensive pian for human trials is now under consideration.
D includes Sh. sonnei. The serogroups A, B, and C are further subdivided into 31 individual serotypes, but serogroup D contains only one serotype. Sh. dysenteriae serotype 1 produces more serious disease than those produced by other serotypes.
SHIGELLA TOXIN
Sh. dysenteriae is known to produce a heat-labile exotoxin, called Shiga toxin, in addition of its heat-stable endotoxin. In experimental animals, such as mice or rabbits, this toxin causes peripheral paralysis and death. This is the reason that this toxin is called neurotoxin. However, Shigella neurotoxin differs from other classical neurotoxins such as tetanus and diphtheria toxins in that it targets the vasculature of the gray matter of the nervous system instead of nerve cells. Shiga toxin also acts as enterotoxins and contributes to the capillary destruction and focal hemorrhages that are associated with bloody stools. Other species of the genus ShigelIa produce Shiga-like toxins that have same clinical effects as that of Shiga toxin but-their quantity is very low as compare to the quantity of Shiga toxin produced by Sh. dysenteriae. Sh. flexneri and Sh. sonnei have been reported to produce cytotoxins. This toxin is stated to be associated with fever and occult stool blood. Their exact pathological role in the disease is yet to be determined.
RESISTANCE
The shigellae are highly susceptible to various physical and chemical agents. They are killed when exposed to 55°C for 1 hour. They can survive for many days in dried conditions especially when protected from sunlight in hypertonic solutions and seawater these bacteria show tolerance for 3 days. They are highly sensitive to radiation. Different disinfectants have bactericidal effects against the shigellae, such as phenol, acids, alkalies, and chlorine.
PATHOGENESIS
Shigellosis is a disease of man involving male and female of all ages. The source
of infection is the fecal material of infected persons or convalescent carriers. individuals are infected orally with the pathogens. The contaminated foods, flies, fomites, and water can serve as vectors. Direct spread is also possible especially in institutional environment, such as children’s daycare centers, hospitals, and prisons, and in individuals involved in homosexuality. The disease is very common in Pakistan and other nonindustriaiized countries where domestic water supply and waste water disposal is not adequately safe.
The incubation period of shigeliosis is usually less than 4 days. The shigillae are very effective pathogens in that the ingestion of 10-500 organisms routinely causes the disease. The organisms first attach to the mucosal epithelium of the ileum and colon and then invade in these layers. The organisms grow in the enterocytes and induce cytopathic changes in the colon including inflammatory reaction and micro-or- gross ulceration. The organisms cause death of enterocytes and spread to adjacent cells where they continue their lethal action. This leads to the erosion of the epithelium and ultimately formation of ulcers. Lesions of the stomach have also been reported in humans and monkeys, but their significance is not well established. The shigellae rarely have the ability to migrate the mesenteric lymph nodes and to cause bacteremia. Sh. dysenteriae is more likely to produce this condition than other species.
CLINICAL MANIFESTATIONS
The onset of shigellosis is acute, with fever, malaise, abdominal pain, and watery diarrhea. As the disease progresses, bloody diarrhea with mucus, tenesemus, fecal urgency, and severe cramping abdominal pain becomes more prominent. Disruption of the normal motor function of the intestine is main cause of tenesemus and cramps. Nausea, vomiting, headache, and convulsions (in children) may occur. Fever is developed as a consequence of the action of the pyrogens released into the blood from the inflammatory reactions. The patients become weaker and more dehydrated.
Shigellosis caused by Sh. dysenteriae is more serious than those produced by other ShigelIa species. The fever is much higher, cramps are more severe, and diarrhea is more frequent and voluminous.
COMPLICATIONS
A patient of shigellosis may get involved in certain complications. These complications may be mild, such as arthritis, conjunctivitis, and morbilliform rash, or life-threatening, such as colonic perforation, shock, septicemia, vascular collapse, and hemolytic uremic syndrome.
LABORATORY DIAGNOSIS
The diagnosis of shigellosis depends on the isolation of the etiological agent from the stool of the patient and its identification. The rectal swabs can also be used for this purpose.
ISOLATION
Although the preferred method for maximal recovery of Shigella from clinical specimens is direct plating on agar media, enrichment of stool specimens in Gram-negative (GN) broth has been reported effective. Salenite broth may also be used in isolating Sh.sonnei. XLD agar is a selective and differential medium for ShigelIa. It contains xylose as differentiating carbohydrate, and since most of the Shigella strains do not ferment xylose, they appear as colorless colonies on the plate. However, Shigella isolates, which ferment xylose rapidly, can be missed on this medium. Therefore, additional plating media containing lactose as differentiating agent (such as EMB and MacConkey agar) should be used in conjunction with XLD agar.
IDENTIFICATION
The isolated colonies typical of Shigella should be identified on the basis of biochemical reactions. For this purpose the colonies may be transferred to TSI agar. Isolates whose TSI reaction is aikaline/acid with no H2S or gas should be screened serologically with antisera to ShigelIa 0 groups A, B, C, and D and inoculated to urea agar and motility medium. Urea-positive or motile cultures can then be discarded. lsolates that are nonmotile, anaerogenic, and urea-negative with other characteristic biochemical reactions may be considered presumptive ShigeIla species. Presumptive positive culture should be tested further biochemically.
THERAPY
The first consideration in the therapy of shigellosis is correction of the patient’s hydration abnormalities. For this oral fluid replacement is necessary. The rapidly rising fever that often results in convulsion in the young must be treated symptomatically. Appropriate application of antibiotics appears to be effective at reducing both the clinical symptoms and the duration of excretion of the organism. Treatment may be withheld in mild cases but is definitely indicated in severe cases or when the risk of transmission is high. Ampicillin, trimethoprim-suifamethoxazole, ciprofloxacin, and tetracycline are effective for sensitive strains.
CONTROL
The most effective method of controlling shigellosis, as in salmonellosis and many other enteric diseases, is to develop a safe water supply and an effective means of feces disposal. A successful control is possible by measures within the household and the local community. The patients, particularly children, in acute stage of dysentery must be isolated. lnfected cases should be restricted from handling food. All the objects used by the patients, such as thermometer, toilet seats, bed sheets, clothing, etc., must be appropriately disinfected. Because of the low numbers of the organisms needed to initiate an infection, asymptomatic carriers also contribute in the transmission of the disease.
PREVENTION
Shigellosis can be prevented by attention to hand washing to minimize person to person transmission and by protection of the water supply from human fecal contamination. Good individual and collective hygienic measures are proved fruitful in preventing shigellosis.lnsects, including flies and cockroaches, should also be controlled by the use of insecticides as they play very important role in the transmission of the disease.
SHIGELLA VACCINES
The severe clinical syndrome caused by Shigella species (particularly Sh. dysenteriae), their propensity to pandemic spread, and their resistance to most relevant antibiotics make a vaccine to prevent shigellosis a high priority. WHO has also placed Shigella vaccine development first in line of its priorities. Regretably, no Shigella vaccines seem to be ready for clinical trials.
The most important ShigelIa strains to be targeted for vaccine development are Sh. flexneri 2a, Sh. dysenteriae 1, and Sh. sonnei. However, the possible emergence of new serotypes has been emphasized. The emergence of Sh. flexneri serotypes 1, 3, and 6 has been observed in many countries. ln 1981, Formal prepared a Sh. sonnei vaccine by inserting into attenuated S. typhi Ty21a the plasmid of Sh. sonnei that contains genes for the synthesis of the O antigens. The resultant hybrid strain, 5076-1C, expresses both S. typhi and Sh. sonnei O antigens. However, because of lot-to-lot variation in protective efficacy, controlled field trials were not undertaken. Another hybrid strain was prepared in 1990 by Format that expressed O antigens of Sh. flexneri. Linderberg prepared a vaccine comprising of a strain of Sh. flexneri with deletion of two genes that are necessary for the bacterium to proliferate in man. But all these efforts did not prove successful in preparing an effective vaccine that can immunize against different virulence factors necessary to produce disease.
Recently, two prototype attenuated vaccine strains of Sh. dysenteriae1 and Sh. flexneri 2a have been developed in the United States. The two strains are safe, highly immunogenic in stimulating lgA antibodies. This vaccine is in the preliminary stages of clinical trials. Another attenuated vaccine comprising of mutant Sh. flexneri has been developed in France. Clinical trials to determine its efficacy initiated in 1996 are currently ongoing. A vaccine has been developed in Sweden that utilizes auxotrophic mutant of Sh. flexneri Y strain. This vaccine was shown to be safe and immunologically potent enough to initiate antibody production against lipopolysaccharide (LPS) of the bacterium. Protection elicited by this vaccine remains to be tested in humans.
Another approach, based on expression of LPS in a live attenuated vector, is under development in Mexico. A similar approach has been taken in Switzerland. ln this case, a hybrid strain of cholera bacilli expresses both host-encoded lnaba and Sh. dysenteriae type 1 O antigens. The results of human trials are still awaited.
A novel approach has been taken in the United States. Parenteral vaccination with the non-covalent complexes of O-polysaccharide and ribosomal particles from Shigella induces a very powerful immune response in experimental animals. After successful clinical trials in animals, an extensive pian for human trials is now under consideration.
