Cent Afr J Med, 2000 Jun, 46(6), 150 - 3 An outbreak of dysentery in a rural district of Zimbabwe: the role of personal hygiene at public gatherings; Midzi SM et al.; OBJECTIVES: To characterize the nature and extent of the outbreak; to determine the risk factors associated with contracting shigellosis; and to institute disease control and preventive measures . DESIGN: Case control study . SETTING: Nyaure Ward, Goromonzi District, Mashonaland East Province, Zimbabwe . SUBJECTS: 52 cases and 52 controls . A case was defined as any resident of Nyaure Ward who presented with bloody diarrhoea (three or more loose stools/day) between 25 July and 25 October 1997 . MAIN OUTCOME MEASURES: Frequencies of symptoms, types of treatment received, risk factors for contracting the illness . RESULTS: The median age was 17 (Q1 = 8, Q3 = 30) years for cases and 19 (Q1 = 7, Q3 = 28) years for controls . Prominent symptoms were abdominal cramps (96.2%), malaise and fever (92.3%), nausea and vomiting (50.0%) . The median duration of diarrhoea was 13 (Q1 = 6, Q3 = 14) days . Eighteen (34.6%) cases were admitted and of these five were referred to a central hospital and two of them complicated with haemolytic uraemic syndrome . The case fatality rate was 1.6% . Twenty four(46.1%) of the cases had Shigella dysenteriae type I, sensitive to Nalidixic acid, Kanamycin, and Doxycycline but resistant to Metronidazole, isolated from the stool specimen . Water samples did not yield any pathogens . Significant risk factors associated with contracting dysentery were shared hand washing in the same standing water at gatherings{OR = 8.47, 95% CI: (2.43-31.33)} or within homes{OR = 60.43, 95% CI: (15.73 to 256.00)} . The use of Blair toilets was a protective factor{OR = 0.03, 95% CI: (0.01 to 0.11)} . CONCLUSION: The epidemiologic evidence implicated contamination of water used for shared hand washing before meals as the source of the infection . The common practice of shared hand washing in standing water should be discouraged at all gatherings and within households to avoid future outbreaks. Curr Biol, 2001 Jan 23, 11(2), R53 - 6 Pathogen evolution: How good bacteria go bad; Stephens C et al.; Recent findings suggest that dysentery-causing Shigella strains have arisen several times from Escherichia coli via plasmid acquisition and phenotypic convergence . Similarly, three Bacillus strains with distinct pathogenic properties are derivatives of a single species whose behavior is profoundly altered by acquired plasmids. J Clin Microbiol, 2001 Mar, 39(3), 1048 - 56 Molecular epidemiology of a Shigella flexneri outbreak in a mountainous township in Taiwan, Republic of China; Chiou CS et al.; An outbreak of shigellosis occurred in a township of Nantou Conuty in central Taiwan from August to October in 1996 . The infections extended to two neighboring townships and continued to the end of 1996 . Forty cases were confirmed during the period, in contrast to only one confirmed case in Nantou County in 1996 before the outbreak . All of these 41 cases in 1996 were identified as infections with Shigella flexneri serotype 2a . In order to trace the source of the infections, the 41 isolates recovered were analyzed by plasmid profile and pulsed-field gel electrophoresis (PFGE) . There was no correlation between the plasmid profile results and the PFGE results, and the latter were used for subtyping of the 41 isolates . Twenty-two isolates (53%) had the same NotI and XbaI PFGE patterns, and 4 isolates (10%) had an additional unstable plasmid band in their NotI patterns but otherwise had the same NotI and XbaI patterns as the 22 isolates . These 26 isolates were designated the outbreak strain, and of these, 24 appeared in eight villages in one township and 2 appeared in a neighboring township . Fourteen of the remaining 15 isolates, including the isolate recovered 7 months before the outbreak, had both NotI and XbaI PFGE patterns closely related to those of the outbreak strain, indicating that Shigella infections were endemic in the area . By tracing the first isolation dates of the outbreak strain in individual villages and the neighboring township, it was found that the strain spread along the major arterial road and its branch road as time passed . Our molecular typing results and epidemiological data demonstrated the endemic nature of the outbreak strain as well as a person-to-person mode of transmission for the widespread infections the strain caused. J Bacteriol, 2001 Mar, 183(6), 1928 - 37 Complete genomic sequence of the lytic bacteriophage phiYeO3-12 of Yersinia enterocolitica serotype O:3; Pajunen MI et al.; phiYeO3-12 is a T3-related lytic bacteriophage of Yersinia enterocolitica serotype O:3 . The nucleotide sequence of the 39,600-bp linear double-stranded DNA (dsDNA) genome was determined . The phage genome has direct terminal repeats of 232 bp, a GC content of 50.6%, and 54 putative genes, which are all transcribed from the same DNA strand . Functions were assigned to 30 genes based on the similarity of the predicted products to known proteins . A striking feature of the phiYeO3-12 genome is its extensive similarity to the coliphage T3 and T7 genomes; most of the predicted phiYeO3-12 gene products were >70% identical to those of T3, and the overall organizations of the genomes were similar . In addition to an identical promoter specificity, phiYeO3-12 shares several common features with T3, nonsubjectibility to F exclusion and growth on Shigella sonnei D(2)371-48 (M . Pajunen, S . Kiljunen, and M . Skurnik, J . Bacteriol . 182:5114-5120, 2000) . These findings indicate that phiYeO3-12 is a T3-like phage that has adapted to Y . enterocolitica O:3 or vice versa . This is the first dsDNA yersiniophage genome sequence to be reported. Epidemiol Infect, 2000 Dec, 125(3), 523 - 30 An epidemiological study of Plesiomonas shigelloides diarrhoea among Japanese travellers; Shigematsu M et al.; Plesiomonas shigelloides is often regarded as a non-pathogenic bacterial species that is occasionally isolated from patients with diarrhoea . However, a review of travellers returning to Japan with diarrhoeal illness through Kansai Airport revealed that the incidence of P . shigelloides from microbiologically confirmed cases increased from 23.2% in 1987 to 77.8% in 1999 . We carried out a descriptive epidemiological study to identify patterns associated with diarrhoea due to this organism . Selected P . shigelloides isolates from this patient group were compared by pulsed-field gel electrophoresis of SpeI total chromosomal DNA digests to determine their genetic heterogeneity . Over the study period (whole of 1996 and first 2 months of 1999), 1149 of 1659 (69.3%) patients with microbiologically confirmed gastroenteritis yielded P . shigelloides . Infection was characterized by watery diarrhoea five times per day that persisted for 3 days . No statistically significant association was found between factors such as age, gender, destination, length of trip, but multivariable logistic regression analysis revealed an association between additional symptoms (vomiting, fever, abdominal pain) age and gender . The molecular fingerprints of a selection of 39 isolates and 3 reference strains of P . shigelloides were highly variable and each had a unique profile . We conclude that although P . shigelloides infections are usually mild and self-limiting, this organism may contribute to a significant proportion of travellers' diarrhoea in the Orient . The species is characterized by great heterogeneity at the DNA level. Epidemiol Infect, 2000 Dec, 125(3), 499 - 503 A community waterborne outbreak of gastro-enteritis attributed to Shigella sonnei; Alamanos Y et al.; An outbreak of gastro-enteritis occurred in a community of 2213 persons located near the city of Ioannina, in North-western Greece . Two hundreds and eighty-eight inhabitants of the village of Eleoussa, suffered from gastro-enteritis between 11 and 22 October . The peak of the epidemic occurred during the first 3 days (11-13 October) . The highest risk of developing gastro-enteritis was observed in the age group 0-14 years (41.4%) and decreased significantly with age (P < 0.01) . Patients over 65 years were more frequently hospitalized than those in other age groups (P < 0.05) . Shigella sonnei was isolated from both, water samples and faeces of patients . Control measures were implemented on the second day of the outbreak . Environmental conditions suggest that contaminationof the water system occurred by groundwater. Zh Mikrobiol Epidemiol Immunobiol, 2000 Nov-Dec, (6), 74 - 6 {Bactericidal properties of hemo-cytolysin from Vibrio cholerae non O1 P-11702 strain in a panel of indicator cultures for detection of vibriocins}; Telesmanich NP et al.; The influence of the preparation of hemo-cytolysin, obtained from V . cholerae non O1 strain P-11702 and inducing lysis of both red blood cells and V . cholerae cultures using a panel of indicator cultures for the detection of vibriocins, was studied . The set of indicator cultures contained 2 Shigella flexneri strains, 1 S . dysenteriae strain, 3 S . sonnei strains, 3 Escherichia coli strains and 2 V . cholerae strains, one of them being atypical . Hemo-cytolysin exhibited lytic activity with respect to S . dysenteriae, S . sonnei strains and 1 V . cholerae strain . i.e . to 4 out of 11 indicator strains . V . cholerae atypical strain proved to be resistant to the preparation in contrast to 33 V . cholerae typical strains, studied previously. Zh Mikrobiol Epidemiol Immunobiol, 2000 Nov-Dec, (6), 38 - 41 {Impaired immune response associated with ABO antigen system in patients with purulent abscesses and dysentery}; Zemskov AM et al.; On the basis of a considerable number of facts--the results of the immunological survey of 197 patients with purulent infections of soft tissues and 103 shigellosis patients--the character and manifestation of immunological disturbances were found to depend on the genetic markers of blood (antigens of the AB0 system) which proved to differ in different type of pathology in patients. Zh Mikrobiol Epidemiol Immunobiol, 2000 Nov-Dec, (6), 25 - 7 {Effect of hydroxylamine hydrochloride on lipopolysaccharide fatty acid composition in Shigella dysenteriae 1}; Zakharova NE et al.; Changes in the fatty acid composition of S . dysenteriae 1 lipid A after the treatment of lipopolysaccharide (LPS) with hydrosylamine hydrochloride (HH) and 4 degrees C, 37 degrees C and 56 degrees C were studied with the use of gas-liquid chromatographicmass-spectrometry . The treatment with HH led to a decrease in the toxicity of LPS, but produced no changes in the content of the main fatty acid components of lipid A (lauric, myristic, oxymyristic and palmitic acids) . At the same time the total number of minor fatty acid derivatives decreased from 11 (in the original LPS) to 5 in LPS treated with HH at 56 degrees C. Bull World Health Organ, 2000, 78(12), 1466 - 73 Epub 2003 Nov 17. Safety of community drinking-water and outbreaks of waterborne enteric disease: Israel, 1976-97; Tulchinsky TH et al.; Waterborne disease remains a major public health problem in many countries . We report findings on nearly three decades of waterborne disease in Israel and the part these diseases play in the total national burden of enteric disease . During the 1970s and 1980s, Israel's community water supplies were frequently of poor quality according to the microbiological standards at that time, and the country experienced many outbreaks of waterborne enteric disease . New regulations raised water quality standards and made chlorination of community water supplies mandatory, as well as imposing more stringent guidelines on maintaining water sources and distribution systems for both surface water and groundwater . This was followed by improved compliance and water quality, and a marked decline in the number of outbreaks of waterborne disease; no outbreaks were detected between 1992 and 1997 . The incidence of waterborne salmonellosis, shigellosis, and typhoid declined markedly as proportions of the total burden of these diseases, but peaked during the time in which there were frequent outbreaks of waterborne disease (1980-85) . Long-term trends in the total incidence of reported infectious enteric diseases from all sources, including typhoid, shigellosis, and viral hepatitis (all types) declined, while the total incidence of salmonellosis increased . Mandatory chlorination has had an important impact on improving water quality, in reducing outbreaks of waterborne disease in Israel, and reducing the total burden of enteric disease in the country. Kansenshogaku Zasshi, 2000 Dec, 74(12), 1004 - 11 {A Shigella sonnei outbreak in Nagasaki}; Hamamoto A et al.; We have experienced an outbreak of dysentery in Nagasaki . Shigella sonnei were positively cultured from 467 patients out of suspected 821 cases, and 346 patients were admitted . 121 patients were treated with oral antimicrobials in the outpatient clinic . Five patients were diagnosed as secondary infection . We treated a total of 96 patients in Nagasaki Municipal Medical Center, and studied the clinical and bacterial features in these 96 patients . Chief complaints included fever, abdominal pain and diarrhea . Most diarrheal patients showed waterly diarrhea and only a few were bloody (3 of 47) . Treatment of levofloxacine 300 mg a day for 5 days successfully eliminated S . sonnei from all culture positive patients . An environmental surveillance revealed that water in a well at the university to which many patients were using was the origin of the infection with positive cultures of S . sonnei . No difference between the clinical and environmental isolates was observed in results on biochemical, serological and enzymatic tests . All isolates were susceptible to levofloxacin and to ofloxacin, but three isolates showed resistance of fosfomycin with MIC above 64 micrograms/ml . In analysis of pulsed-field gel electrophoresis, both clinical and environmental isolates were considered to be closely related. J Bacteriol, 2001 Feb, 183(3), 951 - 8 Periplasmic transit and disulfide bond formation of the autotransported Shigella protein IcsA; Brandon LD et al.; The Shigella outer membrane protein IcsA belongs to the family of type V secreted (autotransported) virulence factors . Members of this family mediate their own translocation across the bacterial outer membrane: the carboxy-terminal beta domain forms a beta barrel channel in the outer membrane through which the amino-terminal alpha domain passes . IcsA, which is localized at one pole of the bacterium, mediates actin assembly by Shigella, which is essential for bacterial intracellular movement and intercellular dissemination . Here, we characterize the transit of IcsA across the periplasm during its secretion . We show that an insertion in the dsbB gene, whose gene product mediates disulfide bond formation of many periplasmic intermediates, does not affect the surface expression or unipolar targeting of IcsA . However, IcsA forms one disulfide bond in the periplasm in a DsbA/DsbB-dependent fashion . Furthermore, cellular fractionation studies reveal that IcsA has a transient soluble periplasmic intermediate . Our data also suggest that IcsA is folded in a proteinase K-resistant state in the periplasm . From these data, we propose a novel model for the secretion of IcsA that may be applicable to other autotransported proteins. Cell Microbiol, 2000 Dec, 2(6), 601 - 16 Functional role for the class IX myosin myr5 in epithelial cell infection by Shigella flexneri; Graf B et al.; Efficient control of Shigella-induced, rho-dependent cytoskeletal rearrangements seems to be required to shape the delicate cellular structures associated with bacterial invasion of epithelial cells . We therefore studied a class IX myosin and rho antagonist, the GTPase-activating protein (GAP) myr5, for a potential role in the bacterial entry process . We show that myr5 is recruited into bacterial entry spots . The recruitment pattern resembled that of rhoC or ezrin, but not rhoA, rac or CDC42, while in vitro GAP activity of myr5 was similar for rhoA, B or C . Analysis of myr5 mutants suggested that GTPase- or ATP-binding activites are not required for Shigella-induced recruitment of this atypical myosin to the bacterial entry site . Functional studies revealed a potential dual role of the myosin functions and the GAP module of myr5 for bacterial internalization. Cell Microbiol, 2000 Dec, 2(6), 579 - 90 Targeting of an enteropathogenic Escherichia coli (EPEC) effector protein to host mitochondria; Kenny B et al.; Many Gram-negative pathogens use a type III secretion apparatus to deliver effector molecules into host cells to subvert cellular processes in favour of the pathogen . Enteropathogenic Escherichia coli (EPEC) uses such a system to deliver the Tir effector molecule into host cells . In this paper, we show that the gene upstream of tir, orf19, encodes an additional type III secreted effector protein . Orf19 is delivered into host cells by a mechanism independent of endocytosis, but dependent on EspB . Orf19 is targeted to host mitochondria, where it appears to interfere with the ability to maintain membrane potential . Although the precise role of Orf19 remains to be elucidated, its interaction with mitochondria suggests a possible role in the subversion of key functions of these organelles, such as energy production or control of cell death . This is the first example of a type III secreted protein targeted to mitochondria; it is probable that homologues (present in EPEC and Shigella species) and other bacterial effectors will also target this organelle. Cell Microbiol, 2000 Jun, 2(3), 195 - 205 Actin-based motility of pathogens: the Arp2/3 complex is a central player; Cossart P; Bacterial actin-based motility has provided cell biologists with tools that led to the recent discovery that, in many forms of actin-based motilities, a key player is a protein complex named the Arp2/3 complex . The Arp2/3 complex is evolutionally conserved and made up of seven polypeptides involved in both actin filament nucleation and organization . Interestingly, this complex is inactive by itself and recent work has highlighted the fact that its activation is achieved differently in the different types of actin-based motilities, including the well-known examples of Listeria and Shigella motilities . Proteins of the WASP family and small G-proteins are involved in most cases . It is interesting that bacteria bypass or mimic some of the events occurring in eukaryotic systems . The Shigella protein IcsA recruits N-WASP and activates it in a Cdc42-like fashion . This activation leads to Arp2/3 complex recruitment, activation of the complex and ultimately actin polymerization and movement . The Listeria ActA protein activates Arp2/3 directly and, thus, seems to mimic proteins of the WASP family . A breakthrough in the field is the recent reconstitution of the actin-based motilities of Listeria and N-WASP-coated E . coli (IcsA) using a restricted number of purified cellular proteins including F-actin, the Arp2/3 complex, actin depolymerizing factor (ADF or cofilin) and capping protein . The movement was more effective upon addition of profilin, alpha-actinin and VASP (for Listeria) . Bacterial actin-based motility is now one of the best-documented examples of the exploitation of mammalian cell machineries by bacterial pathogens. Cell Microbiol, 2000 Jun, 2(3), 187 - 93 Bacterial signals and cell responses during Shigella entry into epithelial cells; Tran Van Nhieu G et al.; Shigella invades epithelial cells by inducing cytoskeletal reorganization localized at the site of bacterial-host cell interaction . During entry, the Shigella type III secretion apparatus allows the insertion of a pore that contains the IpaB and IpaC proteins into cell membranes . Insertion of this complex is thought to allow translocation of the carboxy-terminus moiety of IpaC, but also of other Shigella effectors, such as IpaA, into the cell cytosol . IpaC triggers actin polymerization and the formation of filopodial and lamellipodial extensions dependent on the Cdc42 and Rac GTPases . IpaA, on the other hand, binds to the focal adhesion protein vinculin and induces depolymerization of actin filaments . IpaA and the GTPase Rho are not required for actin polymerization at the site of bacterial contact with the cell membrane, but allow the transformation of the IpaC-induced extensions into a structure that is productive for bacterial entry . Rho is required for the recruitment at entry foci of ezrin, a cytoskeletal linker required for Shigella entry, and also of the Src tyrosine kinase . The Src tyrosine kinase activity, which is required for Shigella-induced actin polymerization, also appears to be involved in a negative regulatory loop that downregulates Rho at the site of entry. Cell Microbiol, 2000 Feb, 2(1), 19 - 33 CD44 binds to the Shigella IpaB protein and participates in bacterial invasion of epithelial cells; Skoudy A et al.; Shigella entry into epithelial cells is characterized by a transient reorganization of the host cell cytoskeleton at the site of bacterial interaction with the cell membrane, which leads to bacterial engulfment in a macropinocytic process . Using affinity chromatography on HeLa cell extracts, we show here that the hyaluronan receptor CD44 associates with IpaB, a Shigella protein that is secreted upon cell contact . Overlay and solid-phase assays indicated that IpaB binds directly to the extracellular domain of CD44; binding is saturable and inhibitable, with a half-maximal inhibitory concentration of 175 nM . Immunoprecipitation experiments showed that IpaB associates with CD44 during Shigella entry . CD44 is recruited at bacterial entry sites and localizes at the plasma membrane of cellular extensions induced by Shigella . Pretreatment of cells with an anti-CD44 monoclonal antibody resulted in inhibition of Shigella-induced cytoskeletal reorganization, as well as inhibition of bacterial entry, whereas transfection of CD44 in cells that are deficient for CD44 results in increased bacterial binding to cells and internalization . The IpaB-CD44 interaction appears to be required for Shigella invasion by initiating the early steps of the entry process. Cell Microbiol, 1999 Sep, 1(2), 183 - 93 The secreted IpaB and IpaC invasins and their cytoplasmic chaperone IpgC are required for intercellular dissemination of Shigella flexneri; Page AL et al.; Invasion of epithelial cells by Shigella flexneri involves entry and dissemination . The main effectors of entry, IpaB and IpaC, are also required for contact haemolytic activity and escape from the phagosome in infected macrophages . These proteins are stored in the cytoplasm in association with the chaperone IpgC, before their secretion by a type III secretion apparatus is activated by host cells . We used a His-tagged IpgC protein to purify IpgC-containing complexes and showed that only IpaB and IpaC are associated with IpgC . Plasmids expressing His6-IpgC either alone or together with IpaB or IpaC under the control of an IPTG-inducible lac promoter were introduced into ipgC, ipaB or ipaC mutants . Induction of expression of the recombinant plasmid-encoded proteins by IPTG allowed bacteria to enter epithelial cells, and the role of these proteins in dissemination was investigated by incubating infected cells in either the absence or the presence of IPTG . The size of plaques produced by recombinant strains on cell monolayers was regulated by IPTG, indicating that IpgC, IpaB and IpaC were each required for efficient dissemination . Electron microscopy analysis of infected cells indicated that these proteins were necessary for lysis of the membrane of the protrusions during cell-to-cell spread. Bull Exp Biol Med, 2000 Nov, 130(11), 1093 - 4 Changes in agent variability and time course of disease incidence during a year (as exemplified by dysentery); Savilov ED et al.; The vulnerability of epidemic process during the period of minimum annual incidence of the disease is validated . Biological properties of Shigella sonnei are studied and their variability examined using the index for evaluation of the mean number of variations for a sign . Minimum agent heterogeneity coincides with minimum incidence of disease and maximum heterogeneity with its seasonal rises. J Clin Ultrasound, 2001 Jan, 29(1), 48 - 50 Sonographic findings in Shigella colitis; Fujii Y et al.; We present a case of colitis caused by Shigella sonnei in which abdominal sonography helped in the early diagnosis, leading to successful conservative management . Sonography showed diffuse wall thickening and layer stratification in the descending and sigmoid colon . Although Shigella colitis is relatively rare, the possibility must be considered when patients have diffuse wall thickening with distinct layer stratification in the left colon . Br J Nutr, 2000 Nov, 84(5), 775 - 9 Effect of an energy-dense diet on the clinical course of acute shigellosis in undernourished children; Mazumder RN et al.; To date there have been few reports on the impact of dietary intervention on the clinical course of acute shigellosis . Current management of acute shigellosis is primarily focused on antibiotic therapy with less emphasis on nutritional management . In a randomised clinical trial, we examined the role of an energy-dense diet on the clinical outcome in malnourished children with acute dysentery due to shigellosis . Seventy-five children aged 12--48 months with acute dysentery randomly received either a milk--cereal formula with an energy density of 4960 kJ/l (test group) or a milk-cereal formula with energy of 2480 kJ/l (control group) for 10 d in hospital . In both milk-cereal formulas, protein provided 11 % energy . In addition, the standard hospital diet was offered to all children and all children received an appropriate antibiotic for 5 d . The mean food intakes (g/kg per d) in the test and control groups were: 112 (SE 2.28) and 116 (SE 3.48) on day 1; 118 (SE 2.72) and 107 (SE 3.13) on day 5; 120 (SE 2.25) and 100 (SE 3.83) on day 10 . The mean energy intakes (kJ/kg per d) in the test and control groups respectively were: 622 (SE 13.2) and 315 (SE 11.3) on day 1; 655 (SE 15.1) and 311 (SE 7.98) on day 5; 672 (SE 14.7) and 294 (SE 11.1) on day 10 . The food and energy intakes were mostly from the milk-cereal diet . There was no difference between two groups in resolution of fever, dysenteric (bloody and or mucoid) stools, stool frequency and tenesmus . However, vomiting was more frequently observed among the test-group children during the first 5 d of intervention (67 % v . 41 %, There was an increase in the mean weight-for-age (%) in the test group compared with the control group after the 10 d of dietary intervention (6.2 (SE 0.6) v . 2.7 (SE 0.4), In addition, resolution of rectal prolapse was better (26 % v . 8 %, in the test group v . control group after 5 d, and 13 % v . 6 %, after 10 d of dietary intervention . Supplementation with a high-energy diet does not have any adverse effect on clinical course of acute shigellosis and reduces the incidence of rectal prolapse in malnourished children. Hong Kong Med J, 2000 Dec, 6(4), 375 - 80 Plesiomonas shigelloides infection in Hong Kong: retrospective study of 167 laboratory-confirmed cases; Wong TY et al.; OBJECTIVE: To study the epidemiological, clinical, and microbiological features of Plesiomonas shigelloides infection in Hong Kong . DESIGN: Retrospective study . SETTING: Infectious Disease Unit of a district hospital, Hong Kong . PATIENTS: Patients with laboratory-confirmed cases of Plesiomonas shigelloides infection between 1 January 1995 and 31 December 1998 . MAIN OUTCOME MEASURES: Epidemiological and clinical data, antibiotic sensitivity, and clinical outcome . RESULTS: There was an increasing trend in the number of isolates of Plesiomonas shigelloides obtained and the prevalence of the bacterium . A total of 197 isolates were obtained from 188 patients, and most isolates (172; 87.3%) were obtained during the summer . Clinical and epidemiological data were available for 167 patients (85 males, 82 females) . Patient age ranged from 1 month to 95 years; the mean and median ages of the patients older than 15 years were 51.0 and 40.5 years, respectively (n=132) . Only 35 (21.0%) of the 167 patients had a history of travel outside Hong Kong, whereas 21 (12.6%) had a history of consuming seafood or uncooked food; 39 (23.4%) had underlying medical conditions . Most patients (165; 98.8%) had symptoms of Plesiomonas shigelloides infection . Nine (5.4%) patients had had chronic diarrhoea for more than 2 weeks; watery and bloody diarrhoea was discharged by 122 (73.1%) and 42 (25.1%) of the patients, respectively . All 197 Plesiomonas shigelloides isolates were sensitive to ofloxacin, or levofloxacin and ceftriaxone . Resistance or partial resistance was recorded for ampicillin (72%), tetracycline (67%), co-trimoxazole (12%), and chloramphenicol (5%) . The majority of patients (142/167; 85.0%) had self-limiting cases of infection, but 25 patients were given antibiotics for more severe symptoms at the time of presentation; there were two deaths . CONCLUSIONS: The occurrence of Plesiomonas shigelloides infection in Hong Kong is increasing, although most cases of are self-limiting. Nat Med, 2001 Feb, 7(2), 180 - 5 Downregulation of bactericidal peptides in enteric infections: a novel immune escape mechanism with bacterial DNA as a potential regulator; Islam D et al.; Antibacterial peptides are active defense components of innate immunity . Several studies confirm their importance at epithelial surfaces as immediate barrier effectors in preventing infection . Here we report that early in Shigella spp . infections, expression of the antibacterial peptides LL-37 and human beta-defensin-1 is reduced or turned off . The downregulation is detected in biopsies from patients with bacillary dysenteries and in Shigella- infected cell cultures of epithelial and monocyte origin . This downregulation of immediate defense effectors might promote bacterial adherence and invasion into host epithelium and could be an important virulence parameter . Analyses of bacterial molecules causing the downregulation indicate Shigella plasmid DNA as one mediator. Am J Physiol Gastrointest Liver Physiol, 2001 Mar, 280(3), G319 - 23 Microbes and microbial toxins: paradigms for microbial-mucosal interactions III . Shigellosis: from symptoms to molecular pathogenesis; Sansonetti PJ; Interaction of Shigella flexneri with epithelial cells includes contact of bacteria with the cell surface and release of Ipa proteins through a specialized type III secreton . A complex signaling process involving activation of small GTPases of the Rho family and c-src causes major rearrangements of the subcortical cytoskeleton, thereby allowing bacterial entry by macropinocytosis . After entry, shigellae escape to the cell cytoplasm and initiate intracytoplasmic movement through polar nucleation and assembly of actin filaments caused by bacterial surface protein IcsA, which binds and activates neuronal Wiskoff-Aldrich syndrome protein (N-WASP), thus inducing actin nucleation in an Arp 2/3-dependent mechanism . Actin-driven motility promotes efficient colonization of the host cell cytoplasm and rapid cell-to-cell spread via protrusions that are engulfed by adjacent cells in a cadherin-dependent process . Bacterial invasion turns infected cells to strongly proinflammatory cells through sustained activation of nuclear factor-kappaB . A major consequence is interleukin (IL)-8 production, which attracts polymorphonuclear leukocytes (PMNs) . On transmigration, PMNs disrupt the permeability of this epithelium and promote its invasion by shigellae . At the early stage of infection, M cells of the follicle-associated epithelium allow bacterial translocation . Subsequent apoptotic killing of macrophages in a caspase 1-dependent process causes the release of IL-1beta and IL-18, which accounts for the initial steps of inflammation. Mol Microbiol, 2001 Feb, 39(3), 792 - 800 Secretion of predicted Inc proteins of Chlamydia pneumoniae by a heterologous type III machinery; Subtil A et al.; Chlamydia spp . are strictly intracellular pathogens that grow inside a vacuole, called an inclusion . They possess genes encoding proteins homologous to components of type III secretion machineries, which, in other bacterial pathogens, are involved in delivery of bacterial proteins within or through the membrane of eukaryotic host cells . Inc proteins are chlamydial proteins that are associated with the inclusion membrane and are characterized by the presence of a large hydrophobic domain in their amino acid sequence . To investigate whether Inc proteins and other proteins exhibiting a similar hydropathic profile might be secreted by a type III system, we used a heterologous secretion system . Chimeras were constructed by fusing the N-terminal part of these proteins with a reporter, the Cya protein of Bordetella pertussis, and these were expressed in various strains of Shigella flexneri . We demonstrate that these hybrid proteins are secreted by the type III secretion system of S . flexneri, thereby providing evidence that IncA, IncB and IncC are secreted by a type III mechanism in chlamydiae . Moreover, we show that three other proteins from Chlamydia pneumoniae, all of which have in common the presence of a large hydrophobic domain, are also secreted by S . flexneri type III secretion machinery. Mol Microbiol, 2001 Feb, 39(3), 652 - 63 Structure and composition of the Shigella flexneri "needle complex", a part of its type III secreton; Blocker A et al.; Type III secretion systems (TTSSs or secretons), essential virulence determinants of many Gram-negative bacteria, serve to translocate proteins directly from the bacteria into the host cytoplasm . Electron microscopy (EM) indicates that the TTSSs of Shigella flexneri are composed of: (1) an external needle; (2) a transmembrane domain; and (3) a cytoplasmic bulb . EM analysis of purified and negatively stained parts 1, 2 and a portion of 3 of the TTSS, together termed the "needle complex" (NC), produced an average image at 17 A resolution in which a base, an outer ring and a needle, inserted through the ring into the base, could be discerned . This analysis and cryoEM images of NCs indicated that the needle and base contain a central 2-3 nm canal . Five major NC components, MxiD, MxiG, MxiJ, MxiH and MxiI, were identified by N-terminal sequencing . MxiG and MxiJ are predicted to be inner membrane proteins and presumably form the base . MxiD is predicted to be an outer membrane protein and to form the outer ring . MxiH and MxiI are small hydrophilic proteins . Mutants lacking either of these proteins formed needleless secretons and were unable to secrete Ipa proteins . As MxiH was present in NCs in large molar excess, we propose that it is the major needle component . MxiI may cap at the external needle tip. J Appl Microbiol, 2001 Feb, 90(2), 229 - 36 A colony blot immunoassay to detect enteroinvasive Escherichia coli and Shigella in water samples; Szakal D et al.; AIMS: The aim of the study was to develop a colony blot immunoassay to detect Shigella and enteroinvasive Escherichia coli (EIEC) in water . METHODS AND RESULTS: Spiked samples were filtered through nitrocellulose membranes . Colony prints on the filters were tested with a monoclonal antibody specific to IpaC, an antigen coded by the invasion plasmid of Shigella and EIEC . Invasive pathogens could be successfully detected with the technique, even in the presence of a large number of non-pathogenic bacterial cells . The method was significantly more sensitive in identifying pathogen-containing samples then the traditional culture-based approach . CONCLUSION: The IpaC-specific colony blot immunoassay is an inexpensive method for identifying the aetiological agents of bacillary dysentery in water samples . SIGNIFICANCE AND IMPACT OF THE STUDY: The technique could be particularly useful in detecting enteroinvasive E . coli which often remains undetected by bio- and serotyping. Microb Pathog, 2001 Jan, 30(1), 1 - 8 Genetic organization of the she pathogenicity island in Shigella flexneri 2a; Al-Hasani K et al.; In this study we report the complete nucleotide sequence and genetic organization of the she pathogenicity island (PAI) of Shigella flexneri 2a strain YSH6000T . The 46 603 bp she PAI is situated adjacent to the 3' terminus of the pheV tRNA gene and includes an imperfect direct repeat of the 3'-terminal 22 bp of the pheV gene at the right boundary of the PAI . The she PAI carries a bacteriophage P4-like integrase gene within the pheV -proximal boundary of the PAI, intact and truncated mobile genetic elements, plasmid-related sequences, open reading frames exhibiting high sequence similarity to those found on the locus of enterocyte effacement (LEE) PAI of enterohemorrhagic Escherichia coli (EHEC), and the SHI-2 PAI of S . flexneri and several other open reading frames of unknown function . The she PAI also encodes two autotransporter proteins, including SigA, a cytopathic protease that contributes to intestinal fluid accumulation and Pic, a protease with mucinase, and hemagglutinin activities . In addition, an open reading frame (orf) termed sap, has high sequence similarity to the gene encoding Antigen 43, a surface-located autotransporter protein of E . coli . The ShET1 enterotoxin genes, associated predominantly with S . flexneri 2a strains, are also located on the she PAI . Infect Immun, 2001 Feb, 69(2), 1072 - 83 Parameters underlying successful protection with live attenuated mutants in experimental shigellosis; Bernardini ML et al.; Because the use of live attenuated mutants of Shigella spp . represents a promising approach to protection against bacillary dysentery (M . E . Etherridge, A . T . M . Shamsul Hoque, and D . A . Sack, Lab . Anim . Sci . 46:61-66, 1996), it becomes essential to rationalize this approach in animal models in order to optimize attenuation of virulence in the vaccine candidates, as well as their route and mode of administration, and to define the correlates of protection . In this study, we have compared three strains of Shigella flexneri 5--the wild-type M90T, an aroC mutant, and a double purE aroC mutant--for their pathogenicity, immunogenicity, and protective capacity . Protection against keratoconjunctivitis, induced by wild-type M90T, was used as the protection read out in guinea pigs that were inoculated either intranasally or intragastrically . Following intranasal immunization, the aroC mutant elicited weak nasal tissue destruction compared to M90T and achieved protection correlated with high levels of local anti-lipopolysaccharide immunoglobulin A (IgA), whereas the purE aroC double mutant, which also elicited weak tissue destruction, was not protective and elicited a low IgA response . Conversely, following intragastric immunization, only the M90T purE aroC double mutant elicited protection compared to both the aroC mutant and the wild-type strain . This mutant caused mild inflammatory destruction, particularly at the level of Peyer's patches, but it persisted much longer within the tissues . This could represent an essential parameter of the protective response that, in this case, did not clearly correlate with high anti-lipopolysaccharide IgA titers. J Clin Microbiol, 2001 Feb, 39(2), 670 - 4 Clonal relationships among Shigella serotypes suggested by cryptic flagellin gene polymorphism; Coimbra RS et al.; The presence of cryptic fliC alleles in the genomes of 120 strains representative of the four Shigella species was investigated . One fragment was obtained by PCR amplification of fliC, with a size varying from 1.2 to 3.2 kbp, depending on the species or serotype . After digestion with endonuclease HhaI, the number of fragments in patterns varied from three to nine, with sizes of between 115 and 1,020 bp . Patterns sharing most of their bands were grouped to constitute an F type . A total of 17 different F types were obtained from all strains included in this study . A unique pattern was observed for each the following serotypes: Shigella dysenteriae 1, 2, 8, and 10 and S . boydii 7, 13, 15, 16, and 17 . On the contrary, S . dysenteriae serotype 13 and S . sonnei biotype e were each subdivided into two different F types . S . flexneri serotypes 3a and X could be distinguished from the cluster containing S . flexneri serotypes 1 to 5 and Y . S . flexneri serotype 6 clustered with S . boydii serotypes 1, 2, 3, 4, 6, 8, 10, 11, 14, and 18 and S . dysenteriae serotypes 4, 5, 6, 7, 9, 11, and 12 . Two other clusters were outlined: one comprising S . dysenteriae serotypes 3, 12, 13 (strain CDC598-77), 14, and 15 and the other one joining S . boydii serotypes 5 and 9 . None of the 17 fliC patterns was found in the fliC HhaI pattern database previously described for Escherichia coli . Overall, this work supports the hypothesis that Shigella evolved from different ancestral strains of E . coli . Moreover, the method outlined here is a promising tool for the identification of some clinically important Shigella strains as well as for confirmation of atypical isolates as Shigella spp. J Bacteriol, 2001 Feb, 183(4), 1269 - 76 Structure-function analysis of the Shigella virulence factor IpaB; Guichon A et al.; Infection by the gram-negative bacterium Shigella flexneri results in dysentery, an acute inflammatory disease of the colon . Essential events in the pathogenesis of Shigella infections include bacterial invasion of epithelial cells, escape from the phagosome, and induction of apoptosis in macrophages . The Shigella virulence factor invasion plasmid antigen B (IpaB) is required for all of these processes . Induction of apoptosis is dependent on IpaB binding to the cysteine protease caspase-1 (Casp-1) . The activation of this enzyme triggers both apoptosis and release of the proinflammatory cytokine interleukin-1beta . Several IpaB mutants were generated to correlate function with protein subdomains . We determined that the N-terminal portion of IpaB is necessary for stable expression of IpaB . A putative amphipathic alpha-helical domain preserves the structure of IpaB . We found 10 consecutive residues within the amino terminus of the hydrophobic region that play a critical role in invasion, phagosomal escape, and cytotoxicity . An IpaB mutant carrying a mutation in this region binds to Casp-1 yet is not cytotoxic, even following direct delivery to the macrophage cytoplasm . These results indicate that the association between IpaB and Casp-1 is only a step in the activation of macrophage apoptosis. J Med Microbiol, 2000 Apr, 49(4), 355 - 60 Molecular analysis of Shigella sonnei isolated from three well-documented outbreaks in school children; Lee TM et al.; Fifty-eight isolates of Shigella sonnei from three outbreaks in school children and eight control isolates from epidemiologically unrelated sporadic clinical infections in Taiwan were compared by antibiotic susceptibility testing and molecular typing . Antibiotic susceptibility testing showed that all strains except one sporadic isolate were multi-resistant . Ribotyping after restriction endonuclease digestion with SalI, PvuII and HindII generated the same ribosomal pattern in 65 of the 66 isolates . Plasmid profile analysis and pulsed-field gel electrophoresis (PFGE) produced eight and nine distinct patterns, respectively, and were in agreement with the epidemiological relationship of the outbreak strains . Nevertheless, some of the sporadic isolates could be discriminated only by a combination of these two methods . This study showed that plasmid profiling in combination with PFGE may be superior to ribotyping in molecular epidemiological investigations of S . sonnei. Presse Med, 2000 Dec 2, 29(37), 2040 - 1 {Shigella: from the rupture to the invasion and to the destruction of the colonic epithelium}; Sansonetti P; CROSSING THE EPITHELIAL BARRIER: Shigella acts on the immune system in order to cross the intestinal epithelial barrier which is normally impermeable to intestinal flora . Among the effectors of the immune system of the colonic mucosa are translocation sites corresponding to lymphatic follicles carrying a follicular epithelium in the presence of M cells . These cells capture molecules, particles and microorganisms from the intestinal barrier that they translocate to immune competent cells present in the follicles (antigen presenting cells including resident macrophages and then B and T lymphocytes) . MACROPHAGE APTOTOSIS AND ITS CONSEQUENCES: The first cell infected by Shigella appears to be the macrophage once the translocation has been made via the M cells . This infection rapidly leads to apoptotic death of the macrophage allowing the Shigella to infect the laterobasal epithelial cells and the development of an inflammatory response . This response in turn facilitates the invasion, rupture and permeability of the intestinal epithelium and the initiation, by Shigella, of a cell-to-cell invasion pathway. Trends Cell Biol, 2001 Jan, 11(1), 30 - 38 Surfing pathogens and the lessons learned for actin polymerization; Frischknecht F et al.; A number of unrelated bacterial species as well as vaccinia virus (ab)use the process of actin polymerization to facilitate and enhance their infection cycle . Studies into the mechanism by which these pathogens hijack and control the actin cytoskeleton have provided many interesting insights into the regulation of actin polymerization in migrating cells . This review focuses on what we have learnt from the actin-based motilities of Listeria, Shigella and vaccinia and discusses what we would still like to learn from our nasty friends, including enteropathogenic Escherichia coli and Rickettsia Appl Environ Microbiol, 2001 Jan, 67(1), 142 - 7 Rapid detection, identification, and enumeration of Escherichia coli cells in municipal water by chemiluminescent in situ hybridization; Stender H et al.; A new chemiluminescent in situ hybridization (CISH) method provides simultaneous detection, identification, and enumeration of culturable Escherichia coli cells in 100 ml of municipal water within one working day . Following filtration and 5 h of growth on tryptic soy agar at 35 degrees C, individual microcolonies of E . coli were detected directly on a 47-mm-diameter membrane filter using soybean peroxidase-labeled peptide nucleic acid (PNA) probes targeting a species-specific sequence in E . coli 16S rRNA . Within each microcolony, hybridized, peroxidase-labeled PNA probe and chemiluminescent substrate generated light which was subsequently captured on film . Thus, each spot of light represented one microcolony of E . coli . Following probe selection based on 16S ribosomal DNA (rDNA) sequence alignments and sample matrix interference, the sensitivity and specificity of the probe Eco16S07C were determined by dot hybridization to RNA of eight bacterial species . Only the rRNA of E . coli and Pseudomonas aeruginosa were detected by Eco16S07C with the latter mismatch hybridization being eliminated by a PNA blocker probe targeting P . aeruginosa 16S rRNA . The sensitivity and specificity for the detection of E . coli by PNA CISH were then determined using 8 E . coli strains and 17 other bacterial species, including closely related species . No bacterial strains other than E . coli and Shigella spp . were detected, which is in accordance with 16S rDNA sequence information . Furthermore, the enumeration of microcolonies of E . coli represented by spots of light correlated 92 to 95% with visible colonies following overnight incubation . PNA CISH employs traditional membrane filtration and culturing techniques while providing the added sensitivity and specificity of PNA probes in order to yield faster and more definitive results. Comp Immunol Microbiol Infect Dis, 2001 Jan, 24(1), 39 - 55 Enteropathogenicity of Plesiomonas shigelloides and Aeromonas spp . in experimental mono- and coinfection with Cryptosporidium parvum in the intestine of neonatal BALB/c mice; Vitovec J et al.; Enteropathogenicity of Plesiomonas shigelloides, Aeromonas hydrophila, A . caviae and A . sobria was studied both in monoinfections and in coinfections with coccidium Cryptosporidium parvum in neonatal BALB/c mice . In monoinfection experiments, neonatal BALB/c mice were orally infected with 7 x 10(7) or 7 x 10(8) CFU, respectively, of a strain of P . shigelloides or a strain of an Aeromonas spp . In coinfection experiments, the neonatal mice were, in addition to being orally infected with one of the four bacterial species, orally infected with an inoculum containing 10(5) oocysts of C . parvum . Results from monoinfections with P . shigelloides revealed long-term colonisation of the neonatal mouse intestine by this pathogen, along with associated pathological lesions . The lesions varied in severity from atrophy to necrosis of the mucosal inner surface of the ileum and colon, with predilection to the colon and brush border of colonic enterocytes . The effects of coinfection of P . shigelloides with C . parvum were characterised by bacteremia and heavy colonisation of the intestine by P . shigelloides . In addition, extensive necrotising inflammatory changes in the ileum and colon were accompanied by diarrhoea and deaths of coinfected mice . In contrast, the results from monoinfections of neonatal mice with Aeromonas spp . showed only a short-term colonisation of the intestine by the pathogen . However, when mice were coinfected with A . hydrophila and C . parvum, then the growth of the bacterial species was prolonged, and occurred in both the spleen and intestine . However, no substantial clinical or histopathological changes were observed in mice, whether monoinfected with Aeromonas spp . or coinfected with C . parvum . Our study suggests that experimental monoinfections of neonatal BALB/c mice with P . shigellodes, Aeromonas spp . and C . parvum, together with coinfections (each bacterial species with the protozoan C . parvum), may serve as a useful model to study the initial steps of gastrointestinal colonisation and diarrhoeal disease syndromes caused by enteropathogenic bacteria and protozoa, individually and in combination. Pharmacotherapy, 2000 Dec, 20(12), 1417 - 22 Effect of antipyretic therapy on the duration of illness in experimental influenza A, Shigella sonnei, and Rickettsia rickettsii infections; Plaisance KI et al.; STUDY OBJECTIVES: To determine whether antipyretic therapy prolongs the course of experimental influenza A, Shigella sonnei, and Rickettsia rickettsii infections . DESIGN: Retrospective observational study . SETTING: University Center for Vaccine Development . SUBJECTS: Fifty-four volunteers with experimentally induced influenza A, 45 with S . sonnei, and 21 with R . rickettsii infections participated . INTERVENTIONS: Subjects from the six influenza A studies were challenged intranasally . If they met certain criteria, they were offered aspirin or acetaminophen for symptomatic relief . Subjects from the three Shigella studies were challenged with the bacteria and then given trimethoprimsulfamethoxazole . Acetaminophen also could be administered . In the one R . rickettsii trial, subjects were inoculated intradermally and treated with tetracycline . Again, acetaminophen was administered for symptomatic relief . MEASUREMENTS AND MAIN RESULTS: Data, excerpted from subjects' study records, were evaluated using Wilcoxon tests, Spearman's correlation coefficients, and multiple regression analysis . Two-tailed hypotheses with a p value of 0.05 were used for all of the analyses . There was a striking correlation between antipyretic therapy and duration of illness in subjects infected with influenza A and S . sonnei, but not R . rickettsii . CONCLUSIONS: Multivariate analysis suggested that antipyretic therapy prolonged illness in subjects infected with influenza A, but its use was the result of prolonged illness in those infected with S . sonnei . The precise nature of these relationships requires a prospective, randomized, placebo-controlled trial. Nature, 2000 Dec 7, 408(6813), 732 - 5 IRSp53 is an essential intermediate between Rac and WAVE in the regulation of membrane ruffling; Miki H et al.; Neural Wiskott-Aldrich syndrome protein (N-WASP) functions in several intracellular events including filopodium formation, vesicle transport and movement of Shigella frexneri and vaccinia virus, by stimulating rapid actin polymerization through the Arp2/3 complex . N-WASP is regulated by the direct binding of Cdc42 (refs 7, 8), which exposes the domain in N-WASP that activates the Arp2/3 complex . A WASP-related protein, WAVE/Scar, functions in Rac-induced membrane ruffling; however, Rac does not bind directly to WAVE, raising the question of how WAVE is regulated by Rac . Here we demonstrate that IRSp53, a substrate for insulin receptor with unknown function, is the 'missing link' between Rac and WAVE . Activated Rac binds to the amino terminus of IRSp53, and carboxy-terminal Src-homology-3 domain of IRSp53 binds to WAVE to form a trimolecular complex . From studies of ectopic expression, we found that IRSp53 is essential for Rac to induce membrane ruffling, probably because it recruits WAVE, which stimulates actin polymerization mediated by the Arp2/3 complex. Mol Microbiol, 2001 Jan, 39(1), 100 - 11 Identification of functional regions within invasion plasmid antigen C (IpaC) of Shigella flexneri; Picking WL et al.; Shigella flexneri causes bacillary dysentery with symptoms resulting from the inflammation that accompanies bacterial entry into the cells of the colonic epithelium . The effectors of S . flexneri invasion are the Ipa proteins, particularly IpaB and IpaC, which are secreted at the host-pathogen interface following bacterial contact with a host cell . Of the purified Ipa proteins, only IpaC has been shown to possess quantifiable in vitro activities that are related to cellular invasion . In this study, ipaC deletion mutants were generated to identify functional regions within the IpaC protein . From these data, we now know that the N-terminus and an immunogenic central region are not required for IpaC-dependent enhancement of cellular invasion by S . flexneri . However, to restore invasiveness to an ipaC null mutant of S . flexneri, the N-terminus is essential, because IpaC mutants lacking the N-terminus are not secreted by the bacterium . Deletion of the central hydrophobic region eliminates IpaC's ability to interact with phospholipid membranes, and fusion of this region to a modified form of green fluorescent protein converts it into an efficient membrane-associating protein . Meanwhile, deletion of the C-terminus eliminates the mutant protein's ability to establish protein-protein contacts with full-length IpaC . Interestingly, the mutant form of ipaC that restores partial invasiveness to the S . flexneri ipaC null mutant also restores full contact-mediated haemolysis activity to this bacterium . These data support a model in which IpaC possesses a distinct functional organization that is important for bacterial invasion . This information will be important in defining the precise role of IpaC in S . flexneri pathogenesis and in exploring the potential effects of purified IpaC at mucosal surfaces. Mol Microbiol, 2000 Nov, 38(4), 760 - 71 The virulence plasmid pWR100 and the repertoire of proteins secreted by the type III secretion apparatus of Shigella flexneri; Buchrieser C et al.; Bacteria of Shigella spp . are the causative agents of shigellosis . The virulence traits of these pathogens include their ability to enter into epithelial cells and induce apoptosis in macrophages . Expression of these functions requires the Mxi-Spa type III secretion apparatus and the secreted IpaA-D proteins, all of which are encoded by a virulence plasmid . In wild-type strains, the activity of the secretion apparatus is tightly regulated and induced upon contact of bacteria with epithelial cells . To investigate the repertoire of proteins secreted by Shigella flexneri in conditions of active secretion, we determined the N-terminal sequence of 14 proteins that are secreted by a mutant in which secretion was deregulated . Sequencing of the virulence plasmid pWR100 of the S . flexneri strain M90T (serotype 5) has allowed us to identify the genes encoding these secreted proteins and suggests that approximately 25 proteins are secreted by the type III secretion apparatus . Analysis of the G+C content and the relative positions of genes and open reading frames carried by the plasmid, together with information concerning the localization and function of encoded proteins, suggests that pWR100 contains blocks of genes of various origins, some of which were initially carried by four different plasmids. Kansenshogaku Zasshi, 2000 Oct, 74(10), 834 - 40 {Species and serovar-distribution, and drug-resistance of Shigella strains isolated from imported and domestic cases during 1995-1999 in Tokyo}; Matsushita S et al.; A total of 290 Shigella strains consisting of 180 imported strains and 110 domestic strains isolated during 1995-1999 in Tokyo were examined regarding their species and serovar-distribution and their drug-resistance . In both groups, S . sonnei (70.0% in the imported strains, 80.9% in the domestic strains) was found to be the most prevalent species, followed by S . flexneri (20.0% in the imported strains, 19.1% in the domestic strains) . S . dysenteriae and S . boydii were only isolated in the imported cases . Among the S . flexneri serovar, 1b, 2a, 6, 2b, and 3a were predominant in the imported strains, whereas 1b and 2a were predominant in the domestic strains . Provisional new serovar Shigella strains were isolated from 11 imported cases and 2 domestic cases . The drug-resistance test using 9 drugs (CP, TC, SM, KM, ABPC, ST, NA, FOM, and NFLX) showed that 92.2% of the imported strains and 94.5% of the domestic strains were resistant to some of the drugs tested . Drug-resistance patterns of the resistant strains varied in 25 types . Among those, a triple drug-resistance type with TC.SM.ST was found as the most frequent pattern in both groups . None of the strains were resistant to NFLX. Rev Cubana Med Trop, 2000 Jan-Apr, 52(1), 10 - 4 {Plesiomonas shigelloides, a Vibrionaceae to be taken into account}; Bravo L et al.; The antigenic structure and antimicrobial susceptibility were studied in 99 strains isolated from patients with acute diarrhea (6 strains from an outbreak of digestive transmission disease in Santiago de Cuba) and a strain isolated from a patient who died from infections neurological syndrome (INS, meningitis) . Four new serotypes (093, 994, 095, 096), which had not been described in the world classification, were identified from the Cuban isolated strains and were included in the International Serotyping Scheme by the International Reference Center located in Prague, Czech Republic . For the first time in Cuba, the circulation of serotypes 017:H11, 011: H2, 023 . H1alc, 057: H3 which show cross reaction to Shiguella species was proved . Those strains from the outbreak of digestive disease belonged to serotype 050: H11 and had a thermostable toxin . The first case of infectious neurologic syndrome with Plesionomas shigelloides etiology reported in Cuba was described; the strain corresponded to serotype 050: H11 . The worldwide reported pattern of antimicrobial resistance was demonstrated. Braz J Med Biol Res, 2000 Dec, 33(12), 1437 - 42 Studies of the small bowel surface by scanning electron microscopy in infants with persistent diarrhea; Fagundes-Neto U et al.; We describe the ultrastructural abnormalities of the small bowel surface in 16 infants with persistent diarrhea . The age range of the patients was 2 to 10 months, mean 4.8 months . All patients had diarrhea lasting 14 or more days . Bacterial overgrowth of the colonic microflora in the jejunal secretion, at concentrations above 10(4) colonies/ml, was present in 11 (68.7%) patients . The stool culture was positive for an enteropathogenic agent in 8 (50.0%) patients: for EPEC O111 in 2, EPEC O119 in 1, EAEC in 1, and Shigella flexneri in 1; mixed infections due to EPEC O111 and EAEC in 1 patient, EPEC O119 and EAEC in 1 and EPEC O55, EPEC O111, EAEC and Shigella sonnei in 1 . Morphological abnormalities in the small bowel mucosa were observed in all 16 patients, varying in intensity from moderate 9 (56.3%) to severe 7 (43.7%) . The scanning electron microscopic study of small bowel biopsies from these subjects showed several surface abnormalities . At low magnification (100X) most of the villi showed mild to moderate stunting, but on several occasions there was subtotal villus atrophy . At higher magnification (7,500X) photomicrographs showed derangement of the enterocytes; on several occasions the cell borders were not clearly defined and very often microvilli were decreased in number and height; in some areas there was a total disappearance of the microvilli . In half of the patients a mucus-fibrinoid pseudomembrane was seen partially coating the enterocytes, a finding that provides additional information on the pathophysiology of persistent diarrhea. Clin Infect Dis, 2000 Dec, 31(6), 1386 - 9 Epub 2000 Nov 29. Transient increase in diarrheal diseases after the devastating earthquake in Kocaeli, Turkey: results of an infectious disease surveillance study; Vahaboglu H et al.; Immediately after the devastating earthquake in Turkey in August 1999, an infectious disease surveillance system was established in Kocaeli Province (the biggest area affected) . This surveillance study was mainly focused on diarrheal diseases . During a 33-day period, 1,468 stool cultures were processed . Diarrheal diseases increased step-by-step and later decreased to the initial level by the end of this period . Cases were scattered throughout the entire region, and the identified causes were various, indicating a multifocal increase . Of the identified causes, Shigella species were the most common . Nevertheless, Shigella isolates also belonged to distinct serotypes and clones . This study indicated a multifocal, multiclonal increase in diarrheal diseases after this massive disaster, thus indicating the necessity to set up infectious disease surveillance systems after such events. Commun Dis Intell, 2000 Sep, 24(9), 272 - 5 An outbreak of multi-resistant Shigella sonnei in a long-stay geriatric nursing centre; McCall B et al.; An outbreak of Shigella sonnei infection in a long-stay nursing centre was detected during routine surveillance of notifications in July 1999 . Subsequent investigations identified 13 cases of multi-resistant S . sonnei infection affecting nine staff, three community members associated with the centre and one resident of the centre . Each isolate of S . sonnei was genetically indistinguishable . The outbreak investigation identified contact with residents with vomiting and diarrhoea as a significant risk factor for infection amongst staff providing nursing care . This association, and the duration of the outbreak over several months, suggests that transmission was most likely person-to-person . This outbreak demonstrates the importance of infection control policies and hygiene measures in long-stay nursing facilities. Infect Immun, 2000 Dec, 68(12), 6624 - 32 Isolation and characterization of a Shigella flexneri invasin complex subunit vaccine; Turbyfill KR et al.; The invasiveness and virulence of Shigella spp . are largely due to the expression of plasmid-encoded virulence factors, among which are the invasion plasmid antigens (Ipa proteins) . After infection, the host immune response is directed primarily against lipopolysaccharide (LPS) and the virulence proteins (IpaB, IpaC, and IpaD) . Recent observations have indicated that the Ipa proteins (IpaB, IpaC, and possibly IpaD) form a multiprotein complex capable of inducing the phagocytic event which internalizes the bacterium . We have isolated a complex of invasins and LPS from water-extractable antigens of virulent shigellae by ion-exchange chromatography . Western blot analysis of the complex indicates that all of the major virulence antigens of Shigella, including IpaB, IpaC, and IpaD, and LPS are components of this macromolecular complex . Mice or guinea pigs immunized intranasally with purified invasin complex (invaplex), without any additional adjuvant, mounted a significant immunoglobulin G (IgG) and IgA antibody response against the Shigella virulence antigens and LPS . The virulence-specific response was very similar to that previously noted in primates infected with shigellae . Guinea pigs (keratoconjunctivitis model) or mice (lethal lung model) immunized intranasally on days 0, 14, and 28 and challenged 3 weeks later with virulent shigellae were protected from disease (P<0.01 for both animal models). Plasmid, 2000 Nov, 44(3), 220 - 30 A novel IS-like element frequently inserted in a putative virulence regulator in bovine mastitis isolates of Streptococcus dysgalactiae; Vasi J et al.; Streptococcus dysgalactiae, a Lancefield group C streptococcus, is commonly isolated from bovine mastitis . We recently identified a putative regulon in two S . dysgalactiae strains, 8215 and Epi9, consisting of two consecutive genes, dmg and dem, coding for a possible regulatory protein and an M-like protein with fibrinogen- and IgG-binding-properties, respectively . During these studies a short sequence homologous to an IS element was found to be inserted in the dmg gene of strain 8215 . The present investigation describes the complete sequence of this IS-like element, named ISSdy1, which consists of 1218 bp and contains two ORFs, flanked by imperfect repeats . The nucleotide sequence of the IS-like element shows 82% identity to the previously reported sequence of IS199 from Streptococcus mutans V403 . The deduced amino acid sequences of the ORFs also revealed high homology to transposases from IS elements in Enterococcus faecium, Escherichia coli, and Shigella dysenteriae, all belonging to the IS3 family . We studied the distribution of ISSdy1 in 57 S . dysgalactiae isolates using PCR analysis with specific primers derived from the IS element . Ninety-eight percent of the isolates contained the ISSdy1 element . Surprisingly, in the majority of studied strains a copy of the IS-like element was found to be inserted in the dmg gene, a putative virulence regulator . J Infect Dis, 2000 Dec, 182(6), 1780 - 3 Epub 2000 Oct 17. Enteroaggregative escherichia coli virulence factors in traveler's diarrhea strains; Vila J et al.; Enteroaggregative Escherichia coli (EAEC) is associated with diarrhea in Spanish travelers to developing countries . In this study, the polymerase chain reaction was used to test EAEC isolates for genes encoding putative virulence factors, including EAEC adhesins, the plasmid-encoded toxin (Pet), a heat-stable enterotoxin (EAST), and Shigella enterotoxins 1 and 2 (ShET1 and ShET2) . Findings included a low prevalence of genes for Pet (4.3%), ShET2 (4.3%), and the adherence factor AAF/II (8.7%) . The overlapping genes encoding the ShET1 and the Pic mucinase were present in most EAEC strains tested (56.5%); however, some strains that carried this locus did not produce both proteins, as determined by Western immunoblot . Surprisingly, ShET1 and ShET2 genes were also found in other E . coli pathotypes, as was the EAST toxin locus . These findings underscore the heterogeneity of EAEC strains and suggest that the ShET1 may be an important virulence factor in traveler's diarrhea. Schweiz Rundsch Med Prax, 2000 Sep 28, 89(39), 1559 - 65 {Chronic diarrhea: value of microbiology in diagnosis}; Kist M; Chronic diarrhoea of the adult is defined as diarrhea during 30 days or longer . Frequent causes of chronic diarrhea in the immunocompetent adult without recent travel to developing countries are noninfectious processes, including laxatives misuse, diseases causing chronic maldigestion, osmotically active artificial sweeteners (i.e . sorbitol), hormonal disorders or drugs with intestinal side effects . Infectious agents as the cause of chronic diarrhea are important in two populations, namely in travelers returning from tropical countries bearing a significant risk of intestinal parasitic infections and in immunocompromised patients, especially AIDS patients with CD4 cell counts below 50 per microliter . Intestinal parasites and C . difficile, Y . enterocolitica, Shigellae and Cytomegalovirus are the most important causative agents of chronic diarrhea . Intestinal pathogens were identified in 46% of chronic, but only in 16.5% of acute diarrhea episodes of HIV-infected patients . An extensive medical history including recent travel as well as the detailed characteristics of onset of symptoms and of their time course is essential for the diagnosis . All patients should have a complete differential blood count, ESR, determination of electrolytes, liver enzymes, creatinine, blood glucose, and serum albumin . Tests to exclude hyperthyriodism, or pancreatic insufficiency as well as a d-xylose absorption test can be included, if appropriate . Microbiological-parasitological investigations are obligatory in patients with chronic diarrhea returning from countries with increased risk of traveler diarrhea, in cases of suspected immunodeficiency, if sudden onset of symptoms with fever is reported, after antibiotic treatment, and in children below six years of age . As a rule, stool specimens are appropriate, for the detection of cytomegalovirus colonic biopsies are necessary . In the latter case colonosigmoidoscopy has no diagnostic advantage . One single stool specimen is sufficient for the detection of bacteria or toxins, in contrast to parasitological investigations, where only three consecutive specimens provide sufficient diagnostic sensitivity. Prikl Biokhim Mikrobiol, 2000 Sep-Oct, 36(5), 597 - 602 {Lipopolysaccharides of Shigella sonnei}; Borisova EV et al.; 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. Infect Immun, 2000 Nov, 68(11), 6449 - 56 Key role for DsbA in cell-to-cell spread of Shigella flexneri, permitting secretion of Ipa proteins into interepithelial protrusions; Yu J et al.; DsbA, a disulfide bond catalyst, is necessary for realization of the pathogenic potential of Shigella flexneri . Sh42, a mutant strain differing from wild-type M90TS solely because it expresses nonfunctional DsbA33G (substitution for 33C at the active site), secreted less IpaB and IpaC than M90TS in response to various stimuli in vitro . A kinetic study demonstrated that Sh42 responded more slowly to Congo red than M90TS . By modulating relative concentrations of functional and nonfunctional DsbA within bacteria, functional enzyme has been shown to be necessary for intercellular spread . By confocal microscopy, M90TS dividing in protrusions was shown to secrete Ipa proteins from the septation furrow, anticipating lysis of protrusions, while Sh42 showed minimal Ipa secretion in this location . In the light of a previous demonstration that DsbA is not necessary for entry of epithelial cells, we conclude that a role in virulence of this disulfide bond catalyst lies in facilitating secretion of Ipa proteins specifically within epithelial protrusions, in turn allowing cell-to-cell spread of S . flexneri. Infect Immun, 2000 Nov, 68(11), 6329 - 36 TonB is required for intracellular growth and virulence of Shigella dysenteriae; Reeves SA et al.; To assess the importance of TonB-dependent iron transport systems to growth of Shigella in vivo, a tonB mutant of Shigella dysenteriae was isolated and tested in cultured cells . The tonB mutant invaded epithelial cells, but did not form plaques in confluent monolayers of Henle cells, indicating an inability of this mutant to spread from cell to cell . The rate of intracellular multiplication of the tonB mutant was reduced significantly compared to that of the wild type . The loss of virulence in the tonB mutant was not due to loss of either Shu or Ent, the TonB-dependent systems which allow for transport of heme and ferrienterobactin, respectively . A shuA mutant lacking the outer membrane receptor for heme, an entB mutant defective in enterobactin synthesis, and a shuA entB double mutant each were able to invade cultured cells, multiply intracellularly, and form wild-type plaques . The ability of S . dysenteriae to access iron during intracellular growth was assessed by flow cytometric analysis of an iron- and Fur-regulated shuA-gfp reporter construct . Low levels of green fluorescent protein expression in the intracellular environment were observed in all strains, indicating that iron is available to intracellular bacteria, even in the absence of TonB-dependent iron transport . The failure of the tonB mutant to grow well in an iron-replete intracellular environment suggests that TonB plays a role in addition to heme- and siderophore-mediated iron acquisition in vivo, and this function is required for the intracellular growth and intercellular spread of S . dysenteriae. Mol Microbiol, 2000 Oct, 38(1), 8 - 19 IpgD, a protein secreted by the type III secretion machinery of Shigella flexneri, is chaperoned by IpgE and implicated in entry focus formation; Niebuhr K et al.; Invasion of epithelial cells by Shigella flexneri involves entry and intercellular dissemination . Entry of bacteria into non-phagocytic cells requires the IpaA-D proteins that are secreted by the Mxi-Spa type III secretion machinery . Type III secretion systems are found in several Gram-negative pathogens and serve to inject bacterial effector proteins directly into the cytoplasm of host cells . In this study, we have analysed the IpgD protein of S . flexneri, the gene of which is located on the virulence plasmid at the 5' end of the mxi-spa locus . We have shown that IpgD (i) is stored in the bacterial cytoplasm in association with a specific chaperone, IpgE; (ii) is secreted by the Mxi-Spa type III secretion system in amounts similar to those of the IpaA-D proteins; (iii) is associated with IpaA in the extracellular medium; and (iv) is involved in the modulation of the host cell response after contact of the bacterium with epithelial cells . This suggests that IpgD is an effector that might be injected into host cells to manipulate cellular processes during infection. FEMS Immunol Med Microbiol, 2000 Oct, 29(2), 129 - 36 Role of porin of Shigella dysenteriae type 1 in modulation of lipopolysaccharide mediated nitric oxide and interleukin-1 release by murine peritoneal macrophages; Biswas T; The ability of Shigella dysenteriae type 1 porin to induce the release of nitric oxide (NO) and interleukin-1 (IL-1) from peritoneal macrophages of mouse and to regulate lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) mediated release of the two proinflammatory mediators was investigated . Porin released nitrite when added to macrophage cultures . A maximum of 3.2-fold nitrite release by macrophages was observed with 100 ng ml(-1) of porin . The nitrite release of LPS was enhanced significantly by lower concentrations of porin, whereas the effect of IFN-gamma was enhanced by porin at higher concentrations . Polysaccharide (PS) moiety of LPS stimulated the nitrite release of elicited macrophages by 1.6-fold compared to untreated control . It also enhanced the stimulatory effect of 1 and 10 ng ml(-1) of porin by 1.3-fold . Lipid A (LPA) moiety of LPS did not release nitrite, nor did it increase the porin mediated nitrite production . Porin treated 24 h old macrophage culture supernatants were applied for ConA activated thymocyte proliferation as a measure for determination of IL-1 release . Sixty percent depletion of thymocyte proliferation was observed when the porin treated macrophage supernatants were absorbed with anti-IL-1 antibody . A maximum of 5.5-fold increase of thymocyte proliferation over control was found with 1 and 10 ng ml(-1) of porin . One or 10 ng ml(-1) of porin and LPS augmented the thymocyte growth, 1.5-fold beyond that obtained by porin and 1.8-/1 . 7-fold more than that obtained by LPS, alone . Similarly, porin and IFN-gamma co-stimulated the cell growth also . PS enhanced the thymocyte proliferation by 5-fold . It also enhanced the thymocyte growth by co-stimulating 1.4-fold the effect observed by 1 or 10 ng ml(-1) of porin alone . LPA could not participate in the cell proliferating activity nor did it enhance the stimulatory effect of porin . Therefore, both nitrite release and thymocyte proliferation by LPS could be substituted by PS only . The tight association of the two bacterial outer membrane components, porin and LPS, could be a necessary co-signal for boosting the release of the two proinflammatory mediators, namely NO and IL-1, which may be associated with the inflammatory response of the colon during Shigella invasion. FEMS Immunol Med Microbiol, 2000 Oct, 29(2), 107 - 13 Specific detection of Plesiomonas shigelloides isolated from aquatic environments, animals and human diarrhoeal cases by PCR based on 23S rRNA gene; Gonzalez-Rey C et al.; Twenty-five strains of Plesiomonas shigelloides isolated from aquatic environment, 10 strains from human cases of diarrhoea and five strains from animals were identified by the polymerase chain reaction technique based on 23S rRNA gene . For this purpose, two primers targeted against part of the 5' half of the 23S rRNA gene of P . shigelloides (Escherichia coli number C-912, G-1195; Plesiomonas number C-906, G-1189) were designed . Results from our study indicated that this method might serve as a tool for a rapid and sensitive identification of P . shigelloides from different environmental and clinical sources. Clin Infect Dis, 2000 Sep, 31(3), 728 - 33 Epub 2000 Oct 04. Application of a geographic information system to the tracking and control of an outbreak of shigellosis; McKee Jr KT et al.; A personal computer-based commercial geographic information system (GIS) was applied to an outbreak of Shigella sonnei infection at Fort Bragg, North Carolina . We used a database consisting of demographic, temporal, and home-address information for all recognized cases of S . sonnei that occurred among health care beneficiaries from 23 May 1997 through 14 August 1997 . We imported this database into the GIS, which contained a digitized basemap of the local community . Through simultaneous examination of temporal and spatial distribution of the 59 identified cases of S . sonnei, a focus of infection in a single housing area was identified . Targeted education among residents of the neighborhood in which there was intense transmission was associated with prompt extinction of the epidemic . A GIS offers an efficient and practical way to directly visualize the dynamics of transmission of infectious diseases in the setting of a community outbreak. Bioorg Khim, 2000 Jul, 26(7), 539 - 47 {Isotyping of human C4 complement using differences in the functional activity of C4A and C4B isotypes}; Kozlov LV et al.; The difference in the functional activity of the isotypes A and B of component C4 of human complement was used to determine their ratio and to detect the inherited deficiency of the isotypes . ELISA methods were developed for the quantitative assay of component C4 (conventional sandwich method) and its functional activity . When determining the functional activity, the classic pathway of the complement and therefore of component C4 was activated by activators sorbed on ELISA microplates (immunoglobulin IgG3 or liposaccharide of the Shigella sonnei cell walls, which activates the complement by binding component C1) . The nascent fragment C4b is covalently bound to the target activator; C4Ab binds better to the target protein (immunoglobulin), and C4Bb to the target carbohydrate (liposaccharide) . Therefore, when immunoglobulin is a target activator, isotype C4A is bound and determined; and when the complement is activated by liposaccharide, isotype C4B is determined . The ratio of the activities determined by the two methods indicates a deficiency in the individual isotypes of component C4 or its absence . The rabbit polyclonal monospecific antibodies against the human component C4 and the conjugates of these antibodies with horseradish peroxidase were used in the methods described. Mol Microbiol, 2000 Sep, 37(6), 1293 - 305 Genetic analysis of Escherichia coli K1 gastrointestinal colonization; Martindale J et al.; Strains of Escherichia coli expressing the K1 polysaccharide capsule colonize the large intestine of newborn infants, and are the leading cause of Gram-negative septicaemia and meningitis in the neonatal period . We used signature-tagged mutagenesis (STM) to identify genes that E . coli K1 requires to colonize the gastrointestinal (GI) tract . A total of 2140 mTn5 mutants was screened for their capacity to colonize the GI tract of infant rats, and 16 colonization defective mutants were identified . The mutants have transposon insertions in genes affecting the synthesis of cell surface structures, membrane transporters, transcriptional regulators, enzymes in metabolic pathways, and in genes of unknown function, designated dgc (defective in GI colonization) . Three dgcs are absent from the whole genome sequence of E . coli K-12, although related sequences are found in other pathogenic strains of E . coli and in Shigella flexneri . Additionally, immunohistochemistry was used to define the nature of the colonization defect in five mutants including all dgc mutants . STM was successfully applied to examine the factors involved in E . coli K1 colonization, and the findings are relevant to the pathogenesis of other enteric infections. J Nat Toxins, 2000 Aug, 9(3), 299 - 313 Escherichia coli Shiga toxin; Nakao H et al.; The Stx family contains two types called Stx1 (verotoxin 1: VT1 or Shiga-like toxin: SLT1) and Stx2 (VT2, SLT2); both toxins are encoded by bacteriophages . Stx1 is identical to Shiga toxin produced by Shigella dysenteriae type I . Stx2 is heterogeneous and immunologically different from Stx1 . Although many variations are found in Stx family, all Stx has an A-B structure: the A subunit has N-glycosidase activity and the B subunit binds to a membrane glycolipid, globotriaosylceramide (Gb3) . The A subunit cleaves a single adenine residue from the 28S rRNA component of eukaryotic ribosomes, resulting in inhibition of protein synthesis . Stx-producing Escherichia coli (STEC) is known to cause hemorrhagic enterocolitis and hemolytic-uremic syndrome (HUS) . Stx plays a role in the occurrence of blood in the feces and in the HUS by their action on the endothelial cells of blood vessels in the intestinal submucosa and in the renal glomeruli . Epidemiologically, Stx2 seems to be more important than Stx1 in development of HUS . The action of Stx is not limited to inhibition of protein synthesis . Stx induces macrophages to express tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and interleukin-6 (IL-6) in vitro . These cytokines and lipopolysaccharide (LPS) are reported to increase the susceptibility of cells to Stx . A variety of cells such as tubular epithelial cells, may be targets for Stx-mediated apoptosis . Apoptosis is considered to contribute to the pathogenesis of HUS caused by STEC . In this review, recent progress in Stx-related research is summarized. Infect Immun, 2000 Oct, 68(10), 6056 - 61 Comparison of O-antigen gene clusters of Escherichia coli (Shigella) sonnei and Plesiomonas shigelloides O17: sonnei gained its current plasmid-borne O-antigen genes from P . shigelloides in a recent event; Shepherd JG et al.; Escherichia coli Sonnei has an O antigen identical to that of Plesiomonas shigelloides O17, and its O-antigen gene cluster is located on a plasmid . By sequencing the chromosomal O-antigen gene cluster of P . shigelloides O17 and comparing it with that of Sonnei, we showed that Sonnei gained its O-antigen genes recently. Infect Immun, 2000 Oct, 68(10), 5502 - 8 Tripeptidyl peptidase II promotes maturation of caspase-1 in Shigella flexneri-induced macrophage apoptosis; Hilbi H et al.; The invasive enteropathogenic bacterium Shigella flexneri activates apoptosis in macrophages . Shigella-induced apoptosis requires caspase-1 . We demonstrate here that tripeptidyl peptidase II (TPPII), a cytoplasmic, high-molecular-weight protease, participates in the apoptotic pathway triggered by Shigella . The TPPII inhibitor Ala-Ala-Phe-chloromethylketone (AAF-cmk) and clasto-lactacystin beta-lactone (lactacystin), an inhibitor of both TPPII and the proteasome, protected macrophages from Shigella-induced apoptosis . AAF-cmk was more potent than lactacystin and irreversibly blocked Shigella-induced apoptosis by 95% at a concentration of 1 microM . Conversely, peptide aldehyde and peptide vinylsulfone proteasome inhibitors had little effect on Shigella-mediated cytotoxicity . Both AAF-cmk and lactacystin prevented the maturation of pro-caspase-1 and its substrate pro-interleukin 1beta in Shigella-infected macrophages, indicating that TPPII is upstream of caspase-1 . Neither of these compounds directly inhibited caspase-1 . AAF-cmk and lactacystin did not impair macrophage phagocytosis or the ability of Shigella to escape the macrophage phagosome . TPPII was also found to be involved in apoptosis induced by ATP and the protein kinase inhibitor staurosporine . We propose that TPPII participates in apoptotic pathways. J Cell Sci, 2000 Oct, 113 Pt 19, 3375 - 86 Myosin light chain kinase plays an essential role in S . flexneri dissemination; Rathman M et al.; Shigella flexneri, the causitive agent of bacillary dysentery, has been shown to disseminate in colonic epithelial cells via protrusions that extend from infected cells and are endocytosed by adjacent cells . This phenomenon occurs in the region of the eukaryotic cell's adherens junctions and is inhibited by pharmacological reagents or host cell mutations that completely disrupt the junctional complex . In this study, inhibitors of the myosin light chain kinase (MLCK) were shown to dramatically decrease intercellular spread of S . flexneri but to have no inhibitory effect on bacterial entry, multiplication or actin-based motility within the host cell . Furthermore, cell-to-cell spread of Listeria monocytogenes, another bacterial pathogen that uses an actin-based mechanism to move within the eukaryotic cytoplasm and to spread from cell to cell, was not affected by the MLCK inhibitors, indicating that (1) the inhibition of S . flexneri cell-to-cell spread in treated cells is not due to a complete break down of cell-cell contacts, which was subsequently confirmed by confocal microscopy, and (2) MLCK plays a role in a S . flexneri-specific mechanism of dissemination . Myosin has been shown to play a role in a variety of membrane-based phenomena . The work presented here suggests that activation of this molecule via phosphorylation by MLCK, at the very least participates in the formation of the bacteria-containing protrusion, and could also contribute to the endocytosis of this structure by neighboring cells. Ned Tijdschr Geneeskd, 2000 Aug 26, 144(35), 1688 - 92 {Shigellosis in Amsterdam, 1991-1998: incidence and results of contact tracing}; Vermaak MP et al.; OBJECTIVE: To determine the incidence of reported shigellosis in Amsterdam (1991-1998) and the factors related to the secondary attack rate of shigellosis . DESIGN: Retrospective . METHODS: From an automated data bank data were collected on all registered shigellosis patients in Amsterdam in the period 1991-1998, and their contacts found by contact tracing . RESULTS: 567 Patients with shigellosis had been reported and a Shigella infection has been diagnosed in 96 of the 983 contacts . The annual incidence decreased statistically significantly from 13 to 8/100.000 . 329 males and 334 females were infected . The highest number of cases was found in children from 0-4 years and among adults from 20-34 years . Of the 663 patients 106 (16%) required hospitalization; for children under 7 this proportion was 30% (32/108) . Among the contacts (excluding the 14 contacts with the same foreign source and the same first sickness day as the index patient) the secondary attack rate was 8% . Young age, of both the patient and the contact was independently related to a higher secondary attack rate . CONCLUSION: With contact tracing for shigellosis a large number of secondary infections were found . Given the high infection rate and large number of hospitalisations among children, contact tracing should focus at this young age group. Trans R Soc Trop Med Hyg, 2000 May-Jun, 94(3), 323 - 6 Treatment of bacillary dysentery in Vietnamese children: two doses of ofloxacin versus 5-days nalidixic acid; Vinh H et al.; Nalidixic acid (NA: 55 mg/kg daily for 5 days) is the recommended treatment for uncomplicated bacillary dysentery in areas where multidrug-resistant Shigella are prevalent . An open randomized comparison of this NA regimen with 2 doses of ofloxacin (total 15 mg/kg) was conducted in 1995/96 in 135 Vietnamese children with fever and bloody diarrhoea . Sixty-six children with a bacterial pathogen isolated were eligible for analysis . Of the 63 Shigella isolates, 39 (62%) were resistant to multiple antibiotics . Resolution times for fever and diarrhoea were similar in the 2 groups, but excretion time of stool pathogen was significantly longer in the NA recipients {median (range) days 1 (1-9) vs 1 (1-2), P = 0.001} . There were 9 (25%) treatment failures in the NA regimen and 3 (10%) in the ofloxacin group; P = 0.1 . Two patients had NA-resistant Shigella flexneri . One of these isolates was selected during NA treatment . From a clinical and public health standpoint a 2-dose regimen of ofloxacin is preferable to nalidixic acid in the treatment of bacillary dysentery. Eur J Biochem, 2000 Sep, 267(18), 5769 - 76 Characterization of the interaction of IpaB and IpaD, proteins required for entry of Shigella flexneri into epithelial cells, with a lipid membrane; De Geyter C et al.; Entry of Shigella flexneri into epithelial cells and lysis of the phagosome involve the IpaB, IpaC, and IpaD proteins, which are secreted by type III secretion machinery . We report here the purification of IpaB and IpaD and the characterization of their lipid-binding properties as a function of pH . The interaction of IpaB with the membrane was quite independent of the pH whereas that of IpaD took place only at low pH . To support the data obtained with the purified proteins, we designed a system in which protein secretion by live bacteria was induced in the presence of liposomes, thereby allowing interaction of proteins with lipids directly after secretion and bypassing any purification step . In these conditions, both IpaB and IpaC, as well as minor amounts of IpaA and IpgD, were associated with the membrane and the ratio of IpaB to IpaC was modulated by the pH . The relevance of these results with respect to the dual roles of IpaB, IpaC and IpaD in induction of membrane ruffles and lysis of the endosomal membrane is discussed. Cent Eur J Public Health, 2000 Aug, 8(3), 150 - 1 New serovars of Plesiomonas shigelloides--1992-1998; Aldova E; Most of the 26 new O (O77-O102) and 10 new H (H42-H50) and H1a1d antigens were found in various P . shigelloides strains isolated from man and other mammals, birds, fish, and water and water insects, not only in the Czech Republic but also in 12 foreign countries. Microbes Infect, 2000 Jun, 2(7), 813 - 9 Diversion of cytoskeletal processes by Shigella during invasion of epithelial cells; Bourdet-Sicard R et al.; Shigella, the causative agent of bacillar dysentery, invades colonic epithelial cells and moves intracellularly to spread from cell to cell . The processes of Shigella entry, determined by the Ipa proteins, and of actin-based motility, dependent on the IcsA/VirG protein, represent different levels of bacterial manipulation of the cell cytoskeleton. Proc Natl Acad Sci U S A, 2000 Sep 12, 97(19), 10567 - 72 Multiple independent origins of Shigella clones of Escherichia coli and convergent evolution of many of their characteristics; Pupo GM et al.; The evolutionary relationships of 46 Shigella strains representing each of the serotypes belonging to the four traditional Shigella species (subgroups), Dysenteriae, Flexneri, Boydii, and Sonnei, were determined by sequencing of eight housekeeping genes in four regions of the chromosome . Analysis revealed a very similar evolutionary pattern for each region . Three clusters of strains were identified, each including strains from different subgroups . Cluster 1 contains the majority of Boydii and Dysenteriae strains (B1-4, B6, B8, B10, B14, and B18; and D3-7, D9, and D11-13) plus Flexneri 6 and 6A . Cluster 2 contains seven Boydii strains (B5, B7, B9, B11, B15, B16, and B17) and Dysenteriae 2 . Cluster 3 contains one Boydii strain (B12) and the Flexneri serotypes 1-5 strains . Sonnei and three Dysenteriae strains (D1, D8, and D10) are outside of the three main clusters but, nonetheless, are clearly within Escherichia coli . Boydii 13 was found to be distantly related to E . coli . Shigella strains, like the other pathogenic forms of E . coli, do not have a single evolutionary origin, indicating convergent evolution of Shigella phenotypic properties . We estimate the three main Shigella clusters to have evolved within the last 35,000 to 270,000 years, suggesting that shigellosis was one of the early infectious diseases of humans. Infect Immun, 2000 Sep, 68(9), 4884 - 92 Attenuated Shigella flexneri 2a vaccine strain CVD 1204 expressing colonization factor antigen I and mutant heat-labile enterotoxin of enterotoxigenic Escherichia coli; Koprowski H 2nd et al.; A multivalent live oral vaccine against both Shigella spp . and enterotoxigenic Escherichia coli (ETEC) is being developed based on the hypothesis that protection can be achieved if attenuated shigellae express ETEC fimbrial colonization factors and genetically detoxified heat-labile toxin from a human ETEC isolate (LTh) . Two detoxified derivatives of LTh, LThK63 and LThR72, were engineered by substitution-serine to lysine at residue 63, or lysine to arginine at residue 72 . The genes encoding these two derivatives were cloned separately on expression plasmids downstream from the CFA/I operon . Following electroporation into S . flexneri 2a vaccine strain CVD 1204, coexpression of CFA/I and LThK63 or LThR72 was demonstrated by Western blot analysis, GM(1) binding assays, and agglutination with anti-CFA/I antiserum . Hemagglutination and electron microscopy confirmed surface expression of CFA/I . Guinea pigs immunized intranasally on days 0 and 15 with CVD 1204 expressing CFA/I and LThK63 or LThR72 exhibited high titers of both serum immunoglobulin G (IgG) and mucosal secretory IgA anti-CFA/I; 40% of the animals produced antibodies directed against LTh . All immunized guinea pigs also produced mucosal IgA (in tears) and serum IgG anti-S . flexneri 2a O antibodies . Furthermore, all immunized animals were protected from challenge with wild-type S . flexneri 2a . This prototype Shigella-ETEC hybrid vaccine demonstrates the feasibility of expressing multiple ETEC antigens on a single plasmid in an attenuated Shigella vaccine strain and engendering immune responses against both the heterologous antigens and vector strain. Infect Immun, 2000 Sep, 68(9), 4856 - 64 Structural analysis of phage-borne stx genes and their flanking sequences in shiga toxin-producing Escherichia coli and Shigella dysenteriae type 1 strains; Unkmeir A et al.; The stx-flanking regions of 49 Shiga toxin-producing Escherichia coli strains and nine Shigella dysenteriae serotype 1 strains containing either stx, stx(1), stx(2), or stx(2) variant genes, were examined . We analyzed these regions by PCR using a set of primers with one primer specific for the respective stx gene and a second primer complementary to sequences of Stx phages H-19B and 933W . We further characterized the amplification products by restriction endonuclease digestion and nucleotide sequencing . PCR products of stx(1)-containing E . coli strains of serogroups O157, O26, and 0103 showed the same lengths and similar restriction patterns . However, we failed to amplify the 3' stx-flanking region in stx(1)-harboring E . coli O111:H(-) strains . Stx2-producing E . coli strains revealed amplification products of different lengths and restriction patterns, suggesting greater heterogeneity than in stx(1)-positive strains . We also obtained specific PCR products for two Stx2c-producing and seven Stx2f-producing E . coli strains when they were subjected to PCR analysis . In nine S . dysenteriae type 1 strains, H-19B- and 933W-specific primers amplified only the 3' stx-flanking region . The results of our study demonstrate that the stx genes of all strains investigated are continuous with phage sequences . Whereas almost all strains except E . coli O111:H(-) strains were associated with a S-like gene, association with Q could not be demonstrated in nine S . dysenteriae type 1 strains and three E . coli strains . Furthermore, we showed that the organization of the stx-flanking regions is similar in all strains investigated, whereas fine-structure analysis showed subtle differences among the sequences examined . Our results support the hypothesis that stx genes in E . coli and S . dysenteriae are generally phage-borne. J Appl Microbiol, 2000 Jul, 89(1), 70 - 5 Prevalence and virulence properties of Vibrio cholerae non-O1, Aeromonas spp . and Plesiomonas shigelloides isolated from Cambé Stream (State of Paraná, Brazil); Gibotti A et al.; The incidence of Vibrio cholerae, Aeromonas spp . and Plesiomonas shigelloides was determined in water samples from Cambe Stream . The samples were collected from seven different sites . The serogroups, virulence markers and drug resistance profiles were also evaluated . Twelve Aer . hydrophila, 12Aer . caviae, eight Aer . sobria, seven Ple . shigelloides and two V . cholerae non-O1 were isolated . They belonged to different serogroups and all produced haemolysis in different assays . Five of the Aeromonas strains and one of V cholerae non-O1 were positive for enterotoxin activity . Haemagglutination and its inhibition, using erythrocytes of different origins, was variable for Aeromonas spp . and V . cholerae, while none of the Ple . shigelloides haemagglutinated in association with any type of erythrocyte . All isolates exhibited multiple drug resistance . These results indicate that the occurrence of V . cholerae non-O1, Aeromonas spp . and Ple . shigelloides, in water used for vegetable irrigation, human recreation and animal consumption, among others, represents a potential risk for humans. Microbiol Immunol, 2000, 44(6), 529 - 31 Expression of fimbriae and hemagglutination activity in Shigella boydii; Utsunomiya A et al.; This report describes the presence of type 1 fimbriae on Shigella boydii 5 which agglutinate guinea pig erythrocytes and feature mannose-sensitive adherence . Morphologically, the fimbriae were thin, rigid cylinders 2-5 microm in length and 35 nm in diameter, and the organella retained axial holes . This is the first study to have revealed the existence of type 1 fimbriae on S . boydii. Rev Latinoam Microbiol, 1999 Oct-Dec, 41(4), 259 - 61 Four biochemical tests for identification of probable enteroinvasive Escherichia coli strains; Flores Abuxapqui JJ et al.; Enteroinvasive Escherichia coli (EIEC) share important features with Shigella spp., but EIEC strains are difficult to identify because their biochemical reactions are variable, and Sereny tests or other biological and molecular assays are expensive or hard to perform . The aim of this work was to detect probable enteroinvasive E . coli strains by using four biochemical tests, in children under 5 years of age with and without acute diarrhea . 330 strains of E . coli isolated from children with diarrhea, and 660 strains from children without diarrhea were studied . All strains were tested with the following tests: mucus , lysine and ornithine decarboxylase and motility . The strains which were negative to the four tests were tested by Sereny assay . Twelve strains (3.6%) isolated from children with diarrhea were negative to the tests proposed; eleven were lactose positive and only one was lactose negative . Three strains (0.5%) from children without diarrhea were negative to the tests proposed and were lactose positive . All the 15 strains (100%) were positive in Sereny assay . We recommend the use of these four biochemical tests for initial detection of EIEC strains, because their cost is very low and it is feasible carry out them in small diagnostic laboratories. Microbiology, 2000 Aug, 146 ( Pt 8), 2039 - 49 Analysis of the beta-glucoside utilization (bgl) genes of Shigella sonnei: evolutionary implications for their maintenance in a cryptic state; Kharat AS et al.; The pattern of expression of the genes involved in the utilization of aryl beta-glucosides such as arbutin and salicin is different in the genus Shigella compared to Escherichia coli . The results presented here indicate that the homologue of the cryptic bgl operon of E . coli is conserved in Shigella sonnei and is the primary system involved in beta-glucoside utilization in the organism . The organization of the bgl genes in S . sonnei is similar to that of E . coli; however there are three major differences in terms of their pattern of expression . (i) The bglB gene, encoding phospho-beta-glucosidase B, is insertionally inactivated in S . sonnei . As a result, mutational activation of the silent bgl promoter confers an Arbutin-positive (Arb(+)) phenotype to the cells in a single step; however, acquiring a Salicin-positive (Sal(+)) phenotype requires the reversion or suppression of the bglB mutation in addition . (ii) Unlike in E . coli, a majority of the activating mutations (conferring the Arb(+) phenotype) map within the unlinked hns locus, whereas activation of the E . coli bgl operon under the same conditions is predominantly due to insertions within the bglR locus . (iii) Although the bgl promoter is silent in the wild-type strain of S . sonnei (as in the case of E . coli), transcriptional and functional analyses indicated a higher basal level of transcription of the downstream genes . This was correlated with a 1 bp deletion within the putative Rho-independent terminator present in the leader sequence preceding the homologue of the bglG gene . The possible evolutionary implications of these differences for the maintenance of the genes in the cryptic state are discussed. Eur J Biochem, 2000 Aug, 267(16), 5136 - 41 31P-NMR and 13C-NMR studies of mannose metabolism in Plesiomonas shigelloides . Toxic effect of mannose on growth; Rager MN et al.; The metabolism of mannose was examined in resting cells in vivo using 13C-NMR and 31P-NMR spectroscopy, in cell-free extracts in vitro using 31P-NMR spectroscopy, and by enzyme assays . Plesiomonas shigelloides was shown to transport mannose by a phosphoenolpyruvate-dependent phosphotransferase system producing mannose 6-phosphate . However, a toxic effect was observed when P . shigelloides was grown in the presence of mannose . Investigation of mannose metabolism using in vivo 13C NMR showed mannose 6-phosphate accumulation without further metabolism . In contrast, glucose was quickly metabolized under the same conditions to lactate, ethanol, acetate and succinate . Extracts of P . shigelloides exhibited no mannose-6-phosphate isomerase activity whereas the key enzyme of the Embden-Meyerhof pathway (6-phosphofructokinase) was found . This result explains the mannose 6-phosphate accumulation observed in cells grown on mannose . The levels of phosphoenolpyruvate and Pi were estimated by in vivo 31P-NMR spectroscopy . The intracellular concentrations of phosphoenolpyruvate and Pi were relatively constant in both starved cells and mannose-metabolizing cells . In glucose-metabolizing cells, the phosphoenolpyruvate concentration was lower, and about 80% of the Pi was used during the first 10 min . It thus appears that the toxic effect of mannose on growth is not due to energy depletion but probably to a toxic effect of mannose 6-phosphate. EMBO J, 2000 Aug 1, 19(15), 3876 - 87 Supramolecular structure of the Shigella type III secretion machinery: the needle part is changeable in length and essential for delivery of effectors; Tamano K et al.; We investigated the supramolecular structure of the SHIGELLA: type III secretion machinery including its major components . Our results indicated that the machinery was composed of needle and basal parts with respective lengths of 45.4 +/- 3.3 and 31.6 +/- 0.3 nm, and contained MxiD, MxiG, MxiJ and MxiH . spa47, encoding a putative F(1)-type ATPase, was required for the secretion of effector proteins via the type III system and was involved in the formation of the needle . The spa47 mutant produced a defective, needle-less type III structure, which contained MxiD, MxiG and MxiJ but not MxiH . The mxiH mutant produced a defective type III structure lacking the needle and failed to secrete effector proteins . Upon overexpression of MxiH in the mxiH mutant, the bacteria produced type III structures with protruding dramatically long needles, and showed a remarkable increase in invasiveness . Our results suggest that MxiH is the major needle component of the type III machinery and is essential for delivery of the effector proteins, and that the level of MxiH affects the length of the needle. MMWR Morb Mortal Wkly Rep, 2000 Jun 30, 49(25), 565 - 8 Outbreak of gastroenteritis associated with an interactive water fountain at a beachside park--Florida, 1999. {Diarrheal disease among caregivers at children's day-care centers . A retrospective and seroepidemiologic study} Koton S, Cohen D, Green MS. Israel Center for Disease Control, Ministry of Health, Tel HashomerDiarrheal disease tends to spread from infected children to their families . Due to the increased risk of exposure, children's caregivers in day-care centers may have a higher incidence of diarrhea, particularly when caring for very young children . We therefore examined the incidence of diarrhea and antibodies to Shigella among caregivers in day-care centers, according to age groups of children in their care (< 18, 18-34, and > 35 months) and in comparison with the general population . 2 studies with a retrospective cohort and seroepidemiological cross-sectional design were carried out . Questionnaires were completed by 401 caregivers in 36% of all WIZO day-care centers . As a measure of past exposure to Shigella, levels of S . sonnei and S . flexneri antibodies were examined in the blood of 110 caregivers (ELISA method) . There was a higher incidence of diarrhea among young children, increasing the potential exposure to diarrheal agents among their caregivers . Nevertheless, no statistically significant differences in diarrhea incidence were found among caregivers of the various age groups during the previous year (p = 0.768) and during the previous month (p = 0.319), nor in absenteeism due to diarrhea during the last month (p = 0.761) . Levels of Shigella antibodies were similar among caregivers in all 3 groups . Diarrheal incidence was higher among women in the population control group than among caregivers during the previous year (p = 0.005) and month (p = 0.067) . No statistically significant differences in levels of S . sonnei and S . flexneri antibodies were found between caregivers and women in the control group . There was no evidence that diarrhea is an occupational hazard for caregivers of young children . An explanation may be the development of protective immunity against common diarrheal agents due to recurrent exposure. FEBS Lett, 2000 Jul 7, 476(3), 129 - 33 High-conductance channel induced by the interaction of phage lambda with its receptor maltoporin; Berrier C et al.; Bacteriophage lambda that binds to liposomes bears its receptor maltoporin (LamB) and is able to inject its DNA into the internal space . During this process, the liposomes are permeabilized, suggesting that a transmembrane channel has formed (Roessner and Ihler (1986) J . Biol . Chem . 261, 386-390) . This pore possibly constitutes the pathway used by lambda DNA to cross the membrane . We reconstituted purified LamB from Shigella in liposomes that were incubated with lambda phages . Addition of this mixture to a bilayer chamber resulted in the incorporation in planar bilayers of high-conductance channels whose conductance, kinetics and voltage dependence were totally different from those of maltoporin channels. Cent Afr J Med, 1998 Nov, 44(11), 272 - 6 Predictors of mortality in children hospitalized with dysentery in Harare, Zimbabwe; Nathoo KJ et al.; OBJECTIVES: To document the pattern of complications and identify risk factors for subsequent mortality in a hospitalized paediatric population during a Shigella dysenteriae type 1 epidemic . DESIGN: Hospital based prognostic study . SETTING: Paediatric wards in Harare and Parirenyatwa tertiary referral hospitals . SUBJECTS: All children aged one month to 12 years admitted to the wards with a history of bloody diarrhoea between January 1993 to June 30 1994 were included in the study (n = 312) . MAIN OUTCOME MEASUR