Colicin Typing of Shigella sonnei

The relative amount of extracellular polymer which remains about Azotobacter vinelandii, Zoogloea ramigera, Klebsiella pneumoniae, and Diplococcus pneumoniae after critical-point drying was studied by electron microscopy. The results obtained with this technique are compared to those obtained with methods that illustrate extracellular polymer, such as freeze-etching and ruthenium red staining. Comparative results indicate critical-point drying to be a rapid, reliable method for the determination of capsule-like polymer surrounding bacterial cells. In addition, critical-point drying can be used to observe morphogenetic changes, such as vesicle production.     

Colicine Typing of Shigella sonnei

The colicine typing method of Shigella sonnei is described with experimental evidence supporting it as well as the manner of selection of eight indicator strains. The disparity of principle between this method and that of Abbott and Shannon's method is (1) selection of the indicators only from wild strains existing in this country, (2) employment of heart infusion broth for colicine production, (3) performance of the typing within 48 hours, and (4) determination of types and subtypes of test-strains by combining their colicinogenic activity against the indicators and their sensitivity to colicines produced by the indicators. A modification of the method is advocated which requires three days to extract colicine by cultivation and one day for sensitivity tests and which uses peptone as the sole nutriment in media. The efficiency of the technique of Abbott and Shannon, McGeachie and McCormick, and the authors' two methods was compared using the selected indicators. Only the technique of McGeachie and McCormick showed some discrepancies.     

Colicine Typing of Shigella sonnei

Colicine typing of Shigella sonnei using the methods of Naito et al. and Abbott and Shannon were performed in parallel on a large number of epidemic strains, the indicator strains used in both methods, and on stock strains used to test the accuracy of the indicators. The following results were obtained after typing 462 epidemic strains: by Naito's method, 18 strains were in A₁, 87 in A₂, 33 in B₁, 253 in B₂, 38 in C₁, 3 in C₂, 2 in D, and 3 in E, and 25 strains were unclassifiable; while by Abbott and Shannon's method, using the present authors' simplified designation, 189 strains were in type 6/11, 124 in type 4/14, 85 in type O, and 41 in type 8, and 23 strains were either in other types or found unclassifiable. Naito's type A was in large part, equivalent to Abbott and Shannon's type O, but included some type 6/11 and type 4/14 strains. Naito's type B consisted of types 6/11, 4/14, 3A and 13, and type E consisted of 1A, IB, 3, 5, 9, and 10. Type C coincided with type 8 and type D with type 12. In addition, there were strains determined to be Abbott and Shannon's types 6/11, 4/14, 2, 7 and 9. The Abbott and Shannon's method revealed the possibilities of these strains to be classified in further detail. This is considered to be attributable not merely to the lack of indicators in the Naito's method corresponding to those of Abbott and Shannon's, but also to insufficient production of certain colicines in the Naito's method. Because of the type distribution found in Japan at present, further investigation should be done so that types 6 and 11, and types 4 and 14 can be separated.     

Epidemiologic Study of Shigella sonnei from Sequential Outbreaks and Sporadic Cases Using Different Typing Techniques

We noted that eight outbreaks of Shigella sonnei from an unknown source occurred sequentially in Aichi Prefecture, Japan, between October 1992-June 1993. For comparative purposes we analyzed 53 outbreak-related isolates of Shigella sonnei using different subtyping methods and studied the epidemiology of the outbreaks. It appeared from our study that DNA-based techniques such as plasmid typing and pulsed-field gel electrophoresis (PFGE) were more useful tools for subtyping Shigella sonnei than colicin typing and the antimicrobial susceptibility test. Moreover, according to PFGE analysis, four genetically related isolates of Shigella sonnei were responsible for the eight sequential outbreaks. To further investigate the epidemiology of outbreaks, 58 sporadic isolates of Shigella sonnei from overseas travelers with shigellosis during the same period were also examined. We found that some sporadic isolates from travelers in Asia were genetically related to those of the outbreak-related isolates, indicating that genetically related isolates prevailed in Asia during this period, probably because of the extensive movement of people or food.     

Bacteriophage Typing as an Epidemiological Tool for Urinary Escherichia coli

Phage typing was used to identify strains of Escherichia coli isolated from urinary and nonurinary sources. When eight phages isolated in Pennsylvania were used to type 717 cultures from Missouri, 50.3% of 624 urinary isolates and 34.4% of 93 nonurinary isolates were typable. Strains from nonurinary sources were not found commonly in urine. When five additional phages isolated in Missouri were added to the original set of eight phages, 80.4% of 331 urinary isolates were typable. When this set of phages was used to type 552 urinary cultures isolated in California, Minnesota, Ohio, Pennsylvania, Virginia, and West Virginia, 82.0% of the cultures were typable. Some common phage types were found in high incidence among cultures from the different regions. No correlation was found between phage type and the pattern of resistance to antibiotics. Phage typing data were presented also on the number of strains in individual urine specimens and the recurrences of strains in patients with chronic bacteriuria. Of 97 fecal isolates, 75.2% of the cultures were typable, and the most common phage type was observed in high incidence among the urinary isolates from this region. When 75 cultures from nine other genera of enteric bacteria were typed, only the shigellae were lysed. In view of the information obtained by phage typing and the ease and speed with which it can be done, it is suggested that phage typing be considered a new tool in epidemiological studies of urinary tract infections by E. coli.     

Lipopolysaccharides of Shigella sonnei

Immunobiological properties of native lipopolysaccharides (LPS) from virulent and avirulent strains of Shigella sonnei bacteria (LPS-V and LPS-A, respectively) were studied. In avirulent bacteria, LPS-V induced immunosuppressive activity specific of the virulent strain. LPS of the avirulent strain, whereas LPS-A lacked this property. Native LPS-V with immunosuppressive activity were isolated from the virulent strain by and immune affinity method. Treatment of LPS-V with phenol or TCA abolished its activity and converted it into the LPS-A form. The data showed that LPS-A can be converted back to the LPS-V form by redox treatment. This approach seems to be promising for activating LPS extracted from cells with TCA or a water-phenol mixture.     

Acid tolerance of Shigella sonnei and Shigella flexneri

AIMS: Determination of the behaviour of Shigella sonnei and Sh. flexneri under acid conditions. METHODS AND RESULTS: The growth and survival of Shigella spp. (9 isolates) in acidified Brain Heart Infusion (BHI) (pH 5.0-3.25 with pH intervals of 0.25) was determined after 6, 24 and 30 h incubation at 37 degrees C. Subsequently, survival of shigellae was studied in apple juice and tomato juice stored at 7 degrees C and 22 degrees C for up to 14 days and in strawberries and a fresh fruit salad, kept at 4 degrees C for 4 and 48 h. CONCLUSIONS: The minimum pH for growth in acidified BHI for Sh. flexneri and Sh. sonnei was, respectively, pH 4.75 and pH 4.50. Survival in fruit juices and fresh fruits depended upon their pH, the type of strain and the incubation temperature. Shigella spp. Survived for up to 14 days in tomato juice and apple juice stored at 7 degrees C. The shortest survival time (2-8 d) was observed in apple juice at 22 degrees C. Sh. sonnei but not Sh. flexneri was recovered after 48 h from strawberries and fruit salad kept at 4 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: Acid foods, especially if kept at refrigeration temperatures, support survival of Shigella spp. and may cause Shigella food poisoning.     

O-specific L-hapten in the composition of Shigella sonnei

Along with classical lipopolysaccharide (LPS), O-specific material not precipitated by ultracentrifugation has been isolated from the water-phenol extract of S. sonnei avirulent strain 9090 possessing complete antigenic properties. The purification of O-antigen contained in the supernatant fluid has been carried out by the gel filtration of the fluid, previously treated with ribonuclease, in a column packed with Sephadex G-100. The polysaccharide nature of O-antigen thus obtained, the absence of lipid A and KDO and the low content of hexoses, or core-specific saccharides of S. sonnei LPS, in this antigen make it possible to classify this material with O-components of microbial cells, described by different authors as "native protoplasmic polysaccharide" or "L-hapten" and formed by polymers of LPS O-side chains. The content of this component in S. sonnei strains under study is, on the average, 2.5% of the weight of dry microbial substance. L-hapten preparations obtained in the course of our investigations have been found to contain two O-specific antigens detected by immunoelectrophoresis and immunodiffusion, as well as by sedimentation in saccharose gradient, where they form peaks corresponding to 4.3 S and 10.8 S. This polysaccharide O-antigen is supposed to be capable of interaction with ribosomal particles and suitable for use as a component of ribosomal dysentery vaccines.     

Detection of immunosuppression factors in Shigella sonnei

A factor, making noninvasive shigellae and other bacteria capable of suppressing immunological memory and secondary immune response manifested as delayed hypersensitivity, has been detected in the germ-free filtrates of the broth cultures of invasive S. sonnei.