Shigella

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 MEASURES: Patterns of complications and predictors of mortality . RESULTS: The peak age group of the patients was 12 to 23 months . Ninety five children died giving an overall case fatality rate (CFR) of 30.4% (95% CI, 25.3 to 35.6) . One third (n = 107) had haemolytic uraemic syndrome (HUS), among whom the CFR was 43% . Other complications included altered consciousness (35%), seizures (12%), coma (4.5%), severe dehydration (14%), abdominal distention (17%), rectal prolapse (5%) and ileus (6%) . In a multivariate analysis, temperature < 36 degrees C (OR 2.12; 95% CI, 1.33 to 3.39), severe dehydration (OR, 1.70; 95% CI, 1.15 to 2.53), hyperkalemia (> 5.5 mmol/L) (OR, 1.74; 95% CI, 1.01 to 1.97), hyponatremia (< 120 mmol/L) (OR, 1.57; 95% CI, 1.17 to 2.11), urea (> 8 mmol/L) (OR, 1.74; 95% CI, 1.29 to 2.36), and abdominal distention (OR, 1.67; 95% CI, 1.16 to 2.41) were found to be predictors of mortality . CONCLUSION: Some of the major clinical and laboratory features allowing the early identification of children at increased risk of dying during dysentery outbreaks have been delineated . The high fatality associated with complicated dysentery highlights the need for preventive strategies which are sustainable in the region.

Commun Dis Public Health, 2000 Jun, 3(2), 135 - 6
Family outbreak of dysentery caused by a rhamnose non-fermenting, ONPG-negative strain of Shigella sonnei phage type 6; Old DC et al.; Three members of a Scottish family, with no history of foreign travel but who had recently visited Bristol, were infected by a strain of Shigella sonnei of phage type 6 (PT 6) that did not ferment rhamnose and was negative for o-nitrophenyl-beta-D-galactopyranoside (ONPG) . The incident exposed limitations associated with commercial systems for the identification of strains of S . sonnei with atypical biochemical properties.

Infect Immun, 2000 Aug, 68(8), 4706 - 13
Ultrastructure of Rickettsia rickettsii actin tails and localization of cytoskeletal proteins; Van Kirk LS et al.; Actin-based motility (ABM) is a mechanism for intercellular spread that is utilized by vaccinia virus and the invasive bacteria within the genera Rickettsia, Listeria, and Shigella . Within the Rickettsia, ABM is confined to members of the spotted fever group (SFG), such as Rickettsia rickettsii, the agent of Rocky Mountain spotted fever . Infection by each agent induces the polymerization of host cell actin to form the typical F (filamentous)-actin comet tail . Assembly of the actin tail propels the pathogen through the host cytosol and into cell membrane protrusions that can be engulfed by neighboring cells, initiating a new infectious cycle . Little is known about the structure and morphogenesis of the Rickettsia rickettsii actin tail relative to Shigella and Listeria actin tails . In this study we examined the ultrastructure of the rickettsial actin tail by confocal, scanning electron, and transmission electron microscopy . Confocal microscopy of rhodamine phalloidin-stained infected Vero cells revealed the typhus group rickettsiae, Rickettsia prowazekii and Rickettsia typhi, to have no actin tails and short (approximately 1- to 3-micrometer) straight or hooked actin tails, respectively . The SFG rickettsia, R . rickettsii, displayed long actin tails (>10 micrometer) that were frequently comprised of multiple, distinct actin bundles, wrapping around each other in a helical fashion . Transmission electron microscopy, in conjunction with myosin S1 subfragment decoration, revealed that the individual actin filaments of R . rickettsii tails are >1 micrometer long, arranged roughly parallel to one another, and oriented with the fast-growing barbed end towards the rickettsial pole . Scanning electron microscopy of intracellular rickettsiae demonstrated R . rickettsii to have polar associations of cytoskeletal material and R . prowazekii to be devoid of cytoskeletal interactions . By indirect immunofluorescence, both R . rickettsii and Listeria monocytogenes actin tails were shown to contain the cytoskeletal proteins vasodilator-stimulated phosphoprotein profilin, vinculin, and filamin . However, rickettsial tails lacked ezrin, paxillin, and tropomyosin, proteins that were associated with actin tails of cytosolic or protrusion-bound Listeria . The unique ultrastructural and compositional characteristics of the R . rickettsii actin tail suggest that rickettsial ABM is mechanistically different from previously described microbial ABM systems.

Antimicrob Agents Chemother, 2000 Aug, 44(8), 2034 - 8
beta-lactamases in Shigella flexneri isolates from Hong Kong and Shanghai and a novel OXA-1-like beta-lactamase, OXA-30; Siu LK et al.; Ninety-one ampicillin-resistant Shigella flexneri strains from Hong Kong and Shanghai were studied for production of beta-lactamases . TEM-1-like and OXA-1-like enzymes were identified in 21 and 79% of the strains, respectively, by isoelectric focusing (IEF) . No difference in the pattern of beta-lactamase production was found between strains from Hong Kong and Shanghai . Four ribotypes were detected . Over 88% of OXA-producing strains had the same ribotype . All TEM-1-like strains harbored a plasmid which hybridized positively with the bla(TEM) probe . Total DNA from OXA-1-like strains failed to hybridize or only hybridized weakly with an OXA probe . The OXA resistance was not transferable . OXA-1-like enzymes exhibited substrate and inhibition profiles similar to that of OXA-1 and were shown to have a pI of 7.3 by further IEF using a narrow-range ampholine gel . The gene encoding the OXA-1-like enzyme from one isolate (CH-07) was cloned, sequenced, and found to differ from bla(OXA-1) at codon 131 (AGA-->GGA; Arg to Gly), resulting in the novel designation OXA-30 . The predominance of OXA-type enzymes in ampicillin-resistant S . flexneri suggests host preference for specific beta-lactamases.

Indian J Pediatr, 2000 May, 67(5), 329 - 33
Clinical risk factors for fatal diarrhea in hospitalized children; Uysal G et al.; It is important to determine the specific factors for diarrheal deaths in infants & young children to enable the intervention and reduce the mortality rates . This study aimed to identify these factors in children under five years of age, hospitalized with diarrheal complaints . Four hundred diarrheal children were included in the study . Twenty-seven (6.75%) of them died and 373 (93.25%) survived . The nutritional status of the patients was determined using weight for height for age as percentage of Harward Standard . It was found that severe malnutrition (p = 0.000 for weight for height ration < 70% and p = 0.036 for height for age < 85%), co-existent sepsis (p = 0.000), shigella infection (p = 0.0014), hypoalbuminemia (p = 0.0000), hypoglycemia (p = 0.0002), hyponatremia (p = 0.016), hypokalemia (p = 0.0041) and metabolic acidosis (p = 0.0069 for pH < 7.35 and p = 0.000 for HCO3 < 20 moL/l) were significant risk factors for diarrheal deaths in the univariate analysis . In the multivariate analysis, young age (under 6 months of age) inverted question markOdds ratio (OR) 10.49, 95% confidence interval (CI) 1.75, 62.75) inverted question mark, moderate or severe dehydration (OR 8.17, 95% CI 1.53, 43.67), severe malnutrition (OR 0.04, 95% CI 0.00, 0.22 of weight for height < 70% and OR 0.03, 95% CI 0.00, 0.36 for height for age < 85%), co-existent sepsis (OR 37.26, 95% CI 6.94, 200.06), shigella infection (OR 23.01, 95% CI 3.08, 171.98), hypoalbuminemia (OR 0.11, 95% CI 0.02, 0.54), metabolic acidosis (OR 0.03, 95% CI 0.00, 0.33 of HCO3 < 20 mMol/L) were significant risk factors . It is concluded that, in addition to electrolyte and fluid treatments, prevention of malnutrition, continuation of feeding which lessens weight loss and may prevent fatal hypoglycemia, and early detection and treatment of probable sepsis are important in reducing diarrheal deaths.

Nat Cell Biol, 2000 Jul, 2(7), 441 - 8
A complex of N-WASP and WIP integrates signalling cascades that lead to actin polymerization; Moreau V et al.; Wiskott-Aldrich syndrome protein (WASP) and N-WASP have emerged as key proteins connecting signalling cascades to actin polymerization . Here we show that the amino-terminal WH1 domain, and not the polyproline-rich region, of N-WASP is responsible for its recruitment to sites of actin polymerization during Cdc42-independent, actin-based motility of vaccinia virus . Recruitment of N-WASP to vaccinia is mediated by WASP-interacting protein (WIP), whereas in Shigella WIP is recruited by N-WASP . Our observations show that vaccinia and Shigella activate the Arp2/3 complex to achieve actin-based motility, by mimicking either the SH2/SH3-containing adaptor or Cdc42 signalling pathways to recruit the N-WASP-WIP complex . We propose that the N-WASP-WIP complex has a pivotal function in integrating signalling cascades that lead to actin polymerization.

J Immunol, 2000 Jul 15, 165(2), 903 - 14
Invasive Shigella flexneri activates NF-kappa B through a lipopolysaccharide-dependent innate intracellular response and leads to IL-8 expression in epithelial cells; Philpott DJ et al.; The pathogenesis of Shigella flexneri infection centers on the ability of this organism to invade epithelial cells and initiate an intense inflammatory reaction . Because NF-kappa B is an important transcriptional regulator of genes involved in inflammation, we investigated the role of this transcription factor during S . flexneri infection of epithelial cells . Infection of HeLa cells with invasive S . flexneri induced NF-kappa B DNA-binding activity; noninvasive S . flexneri strains did not lead to this activation . The pathway leading to NF-kappa B activation by invasive S . flexneri involved the kinases, NF-kappa B-inducing kinase, I kappa B kinase-1, and I kappa B kinase-2 . NF-kappa B activation was linked to inflammation, because invasive S . flexneri activated an IL-8 promoter-driven reporter gene, and the kappa B site within this promoter was indispensable for its induction . Microinjection of bacterial culture supernatants into HeLa cells suggested that LPS is responsible for NF-kappa B activation by S . flexneri infection . In conclusion, the eukaryotic transcription factor NF-kappa B was activated during S . flexneri infection of epithelial cells, which suggests a role for this transcriptional regulator in modulating the immune response during infection in vivo.

Pediatr Infect Dis J, 2000 Jun, 19(6), 522 - 6
Treatment of shigellosis with cefixime: two days vs . five days; Martin JM et al.; BACKGROUND: Although the recommended standard course of therapy for shigellosis is 5 days of oral ampicillin or trimethoprim-sulfamethoxazole therapy, successful outcome has been reported in adults treated with abbreviated courses of antibiotics . The purpose of this study was to compare short course (2-day) vs . 5-day therapy with cefixime for treatment of diarrheal disease caused by Shigella sonnei in children . METHODS: This was a prospective, randomized, double blind, placebo-controlled study . Patients were eligible if they were at least 6 months of age and presented to the Children's Hospital of Pittsburgh during an outbreak of diarrhea caused by S . sonnei, with (1) a history of fever and diarrhea (at least three loose or watery stools per day), (2) bloody diarrhea or (3) diarrhea and known exposure to an individual with documented shigellosis . Patients were randomized to receive either 2 days of cefixime (8 mg/kg(day) given once daily followed by 3 days of placebo or 5 days of cefixime . Telephone follow-up was performed on Days 3, 7 and 14 after enrollment . Follow-up stool cultures were obtained on Day 7 to assess bacteriologic cure . There were standardized definitions for cure, improvement, failure and relapse . RESULTS: Forty-seven patients were enrolled . Eleven were eliminated from analysis because their stool cultures were not positive for S . sonnei . There were 36 evaluable patients, 21 in the 2-day group and 15 in the 5-day group . Patients ranged in age from 6 months to 17 years . Forty-four percent of the subjects were male . Symptoms were improved or had resolved by Day 3 of therapy in all patients . There were 8 patients who experienced a clinical relapse: 5 of 21 (24%) patients in the 2-day treatment group and 3 of 15 (20%) in the 5-day group . There were 13 patients who experienced a bacteriologic failure (defined as the occurrence of a positive culture at the Day 7 follow-up visit), 11 of 20 (55%) in the 2-day group and 2 of 14 (14%) in the 5-day group (P < 0.02) . CONCLUSION: Two- and 5-day treatment courses with cefixime for treatment of diarrheal disease caused by S . sonnei result in similar rates of clinical cure and clinical relapses; however, there was a higher rate of bacteriologic failure with shorter course therapy.

Zh Mikrobiol Epidemiol Immunobiol, 2000 Jan-Feb, (1), 57 - 60
{The cyclicity and prognosis of shigellosis morbidity in Russia}; Kartsev AD; The epidemic process (EP) of shigellosis morbidity proceeds in cycles . As shown in this study, cyclic curves are the sum of polyharmonic fluctuations, being the axis of EP . The duration of one cycle is 18 years, and its configuration is stably retained for a long time . In Nizhny Novgorod shigellosis morbidity is strongly correlated with air temperature is summer at the peak of cyclic rises and moderately correlated in the years of low morbidity . In some cities of the Russian Federation fluctuations of air temperature have been found of occur synchronously with disturbances of the magnetic field of the earth . The character and stability of EP cycles provide grounds for the prognostication of shigellosis morbidity . The current cycle will end in the year 2000, and then natural preconditions for a new cyclic rise will appear.

Zh Mikrobiol Epidemiol Immunobiol, 1999 Nov-Dec, (6), 42 - 5
{Shigellosis and other acute intestinal infections in the Republic of Tajikistan: the epidemiological patterns}; Rafiev KhK; Acute enteric infections (AEI), including shigellosis, are characterized by their extremely wide spread in Tajikistan . Their hyperendemicity is due to a high activity of the water route of transmission under the conditions of hot climate . The morbidity level of AEI both in the republic as a whole and in its individual territories has been found to directly depend on the degree of the potential epidemiological danger of the water supply system, as well as on the quality of water used by the population in the presence of its pronounced deficiency . The important role of breast feeding for the prophylaxis of AEI among young children is shown.

Philos Trans R Soc Lond B Biol Sci, 2000 May 29, 355(1397), 575 - 86
The pathogenesis of Shigella flexneri infection: lessons from in vitro and in vivo studies; Philpott DJ et al.; Shigella flexneri is a Gram-negative facultatively intracellular pathogen responsible for bacillary dysentery in humans . More than one million deaths occur yearly due to infections with Shigella spp . and the victims are mostly children of the developing world . The pathogenesis of Shigella centres on the ability of this organism to invade the colonic epithelium where it induces severe mucosal inflammation . Much information that we have gained concerning the pathogenesis of Shigella has been derived from the study of in vitro models of infection . Using these techniques, a number of the molecular mechanisms by which Shigella invades epithelial cells and macrophages have been identified . In vivo models of shigellosis have been hampered since humans are the only natural hosts of Shigella . However, experimental infection of macaques as well as the murine lung and rabbit ligated ileal loop models have been important in defining some of the immune and inflammatory components of the disease . In particular, the murine lung model has shed light on the development of systemic and local immune protection against Shigella infection . It would be naive to believe that any one model of Shigella infection could adequately represent the complexity of the disease in humans, and more sophisticated in vivo models are now necessary . These models require the use of human cells and tissue, but at present such models remain in the developmental stage . Ultimately, however, it is with such studies that novel treatments and vaccine candidates for the treatment and prevention of shigellosis will be designed.

J Biol Chem, 2000 Sep 15, 275(37), 28893 - 901
Profilin is required for sustaining efficient intra- and intercellular spreading of Shigella flexneri; Mimuro H et al.; The ability of Shigella to mediate actin-based motility within the host cell is a prominent pathogenic feature of bacillary dysentery . The ability is dependent on the interaction of VirG with neural Wiskott-Aldrich syndrome protein (N-WASP), which in turn mediates recruitment of Arp2/3 complex and several actin-related proteins . In the present study, we show that profilin I is essential to the rapid movement of Shigella in epithelial cells, for which the capacity of profilin to interact with G-actin and N-WASP is critical . In COS-7 cells overexpressing either mutated profilin H119E, which failed to bind G-actin, or H133S, which is unable to interact with poly-l-proline, Shigella motility was significantly inhibited . Similarly, depletion of profilin from Xenopus egg extracts resulted in a decrease in bacterial motility that was completely rescued by adding back profilin I but not H119E or H133S . In COS-7 cells overexpressing a N-WASP mutant lacking the proline-rich domain (Deltap) unable to interact with profilin, the actin tail formation of intracellular Shigella was inhibited . In N-WASP-depleted extracts, addition of Deltap but not full-length N-WASP was unable to restore the bacterial motility . Furthermore, in a plaque formation assay with Madin-Darby canine kidney cell monolayers infected by Shigella, Madin-Darby canine kidney cells stably expressing H119E, H133S, or Deltap reduced the bacterial cell-to-cell spreading . These results indicate that profilin I associated with N-WASP is an essential host factor for sustaining efficient intra- and intercellular spreading of Shigella.

Mol Cell Probes, 2000 Jun, 14(3), 181 - 9
PCR detection of Giardia lamblia in stool: targeting intergenic spacer region of multicopy rRNA gene; Ghosh S et al.; A PCR based detection that amplifies the 552-bp intergenic spacer (IGS) region of multicopy rRNA gene of Giardia lamblia and 320-bp internal sequences to first PCR product has been used in diagnosis of giardiasis in stool sample . The primers were found highly specific to Giardia spp . only, because no amplification was observed with DNAs from other enteric pathogens like Escherichia coli, Shigella dysenteriae and Entamoeba histolytica . The test could detect even less than 2 pg of genomic DNA from Giardia trophozoites . In direct diagnosis of Giardia lamblia in stool samples, it was observed that PCR amplification of IGS followed by nested PCR could enhance the sensitivity and specificity of the tests manifold and the system was able to detect as low as 10 parasites in 100 microl of stool . The comparative evaluation of the present system with conventional microscopy, CIEP and ELISA in the diagnosis of giardiasis from diarrhoeic stool samples and control subjects demonstrated a 100% correlation among nested PCR, microscopic examination and ELISA in patients suggestive of giardiasis (Group I) and control subjects (Group II) . In Group I cases (patients suffering from other than giardiasis), CIEP, ELISA and nested PCR showed better results than microscopic examination . However, among them, PCR was found most sensitive and specific because 20% positivity was noticed by PCR whereas CIEP and ELISA showed only 7.14% and 12.85%, respectively . Break-up results showed that all the samples which were positive by CIEP or ELISA, also found positive by PCR . The present observation clearly suggests the use of PCR that amplifies the intergenic spacer region of multicopy rRNA gene of Giardia lamblia followed by nested PCR for routine, quick and reliable detection of Giardia lamblia in stool samples .

Zh Mikrobiol Epidemiol Immunobiol, 1999 May-Jun, (3), 74 - 7
{The rapid identification of bacteria by agglutination on a slide of antibody-sensitized erythrocytes}; Karal'nik BV et al.; The method for the identification of Brucella (genus) and shigellae (species) by using slide agglutination of antibody erythrocytic immune reagents is substantiated . Such reagents, obtained from polyclonal nonadsorbed immune sera, ensure high specificity of taxon identification . The use of the proposed method may greatly accelerate bacteriological analysis.

APMIS, 2000 Apr, 108(4), 251 - 60
Disease-dependent changes in T-cell populations in patients with shigellosis; Islam D et al.; In shigellosis, bacterial infection is associated with an extensive inflammation of the rectal mucosa, resulting in bloody dysentery . The role of T-cell-mediated pro-inflammatory mechanisms has been implicated in this process, but the specific role of T-cell subsets is still not well understood . In this study we attempted to identify the changes in T-cell populations in patients with shigellosis during the disease course . The T-cell subset distribution was analyzed by immunohistochemistry in the rectal mucosa and by immuno-flow cytometry in the peripheral blood . Blood and rectal biopsies were studied from patients with Shigella dysenteriae 1 (n= 11) and S . flexneri (n= 11) infection and 20 healthy age-matched controls . We found an expansion of gammadelta+T cells in the rectal mucosa, but a decrease in the percentage of gammadelta+T cells in the blood in acute shigellosis . There was also a preferential increase in CD8+ T cells in the surface epithelium of rectal tissue in patients infected with S . dysenteriae 1, but not in patients infected with S . flexneri . Our findings suggest that the rectal mucosal inflammation in shigellosis is associated with an expansion of T cells, in particular CD8+ and gammadelta+T-cell subsets in the gut mucosa, which may be of importance for the pathogenesis of shigellosis.

Immunity, 2000 May, 12(5), 581 - 90
Caspase-1 activation of IL-1beta and IL-18 are essential for Shigella flexneri-induced inflammation; Sansonetti PJ et al.; Caspases are intracellular proteases that mediate mammalian cell apoptosis . Caspase-1 (Casp-1) is a unique caspase because it activates the proinflammatory cytokines interleukin (IL)-1beta and IL-18 . Shigella flexneri, the etiological agent of bacillary dysentery, induces macrophage apoptosis, which requires Casp-1 and results in the release of mature IL-1beta and IL-18 . Here we show that casp-1(-/-) mice infected with S . flexneri do not develop the acute inflammation characteristic of shigellosis and are unable to resolve the bacterial infection . Using casp-1(-/-) mice supplemented with recombinant cytokines and experiments with IL-1beta(-/-) and IL-18(-/-) mice, we show that IL-1beta and IL-18 are both required to mediate inflammation in S . flexneri infections . Together, these data demonstrate the importance of Casp-1 in acute inflammation and show the different roles of its substrates, IL-1beta and IL-18, in this response.

J Exp Med, 2000 Jun 5, 191(11), 1905 - 20
Rho family GTPase Cdc42 is essential for the actin-based motility of Shigella in mammalian cells; Suzuki T et al.; Shigella, the causative agent of bacillary dysentery, is capable of directing its movement within host cells by exploiting actin dynamics . The VirG protein expressed at one pole of the bacterium can recruit neural Wiskott-Aldrich syndrome protein (N-WASP), a downstream effector of Cdc42 . Here, we show that Cdc42 is required for the actin-based motility of Shigella . Microinjection of a dominant active mutant Cdc42, but not Rac1 or RhoA, into Swiss 3T3 cells accelerated Shigella motility . In add-back experiments in Xenopus egg extracts, addition of a guanine nucleotide dissociation inhibitor for the Rho family, RhoGDI, greatly diminished the bacterial motility or actin assembly, which was restored by adding activated Cdc42 . In N-WASP-depleted extracts, the bacterial movement almost arrested was restored by adding exogenous N-WASP but not H208D, an N-WASP mutant defective in binding to Cdc42 . In pyrene actin assay, Cdc42 enhanced VirG-stimulating actin polymerization by N-WASP-actin-related protein (Arp)2/3 complex . Actually, Cdc42 stimulated actin cloud formation on the surface of bacteria expressing VirG in a solution containing N-WASP, Arp2/3 complex, and G-actin . Immunohistological study of Shigella-infected cells expressing green fluorescent protein-tagged Cdc42 revealed that Cdc42 accumulated by being colocalized with actin cloud at one pole of intracellular bacterium . Furthermore, overexpression of H208D mutant in cells interfered with the actin assembly of infected Shigella and diminished the intra- and intercellular spreading . These results suggest that Cdc42 activity is involved in initiating actin nucleation mediated by VirG-N-WASP-Arp2/3 complex formed on intracellular Shigella.

Carbohydr Res, 2000 May 5, 325(4), 245 - 52
Synthesis of the tetrasaccharide related to the repeating unit of the antigen from Shigella dysenteriae type 5; Mukherjee I et al.; Starting from L-rhamnose, D-mannose and 2-amino-2-deoxy-D-glucose hydrochloride, two disaccharide blocks, namely, ethyl 2,4-di-O-benzyl-3-O-{(R)-1-(methoxycarbonyl)ethyl}-alpha-L-rhamnopyranos yl-(1-->3)-2-O-acetyl-4,6-di-O-benzyl-1-thio-alpha-D-mannopyranoside and 2-(trimethylsilyl)ethyl 2-O-acetyl-3,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-->3)-4,6-di-O-benzy l-2-deoxy-2-phthalimido-beta-D-glucopyranoside, were synthesised and then allowed to react in the presence of N-iodosuccinimide and trifluoromethane sulfonic acid to give a tetrasaccharide derivative . This compound was converted into 2-(trimethylsilyl)ethyl 2,4-di-O-benzyl-3-O-{(R)-1-(methoxycarbonyl)ethyl}-alpha-L-rhamno- pyranosyl-(1-->3)-2-O-acetyl-4,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-- >4)-2-O-acetyl-3,6-di-O-benzyl-alpha-D-mannopyranosyl-(1-->3)-2-acetamid o-4,6-di-O-benzyl-2-deoxy-beta-D-glucopyranoside, which on hydrogenolysis, afforded the methyl ester 2-(trimethylsilyl)ethyl glycoside of the tetrasaccharide related to the repeating unit of the O-antigen from Shigella dysenteriae type 5.

J Antimicrob Chemother, 2000 Jun, 45(6), 877 - 80
beta-lactamase expression in Plesiomonas shigelloides; Avison MB et al.; We have examined the production of beta-lactamases by 11 clinical and nine environmental isolates of Plesiomonas shigelloides from Czechoslovakia, the Czech Republic and Cuba . Ten isolates (50%) expressed detectable amounts of a chromosomally encoded, non-inducible beta-lactamase, though all isolates showed a broadly similar resistance profile: low-level resistance to ampicillin and higher-level resistance to carbenicillin . All strains were susceptible to cephalosporins and meropenem . Three clinical isolates expressed a beta-lactamase similar to a class 2c carbenicillinase, with a pI of 5.2 and three expressed an enzyme similar to a class 2d oxacillinase, with a pI of 5.3 . The environmental isolates produced a variety of penicillinases, indicating that there is a reservoir of heterogeneous beta-lactamase genes in this species.

Scand J Infect Dis, 2000, 32(2), 207 - 11
Shigellosis of childhood in northern Greece: epidemiological, clinical and laboratory data of hospitalized patients during the period 1971-96; Kavaliotis J et al.; The aim of this study was to evaluate epidemiological, clinical and laboratory data of shigellosis in children from northern Greece, hospitalized in our department during the period 1971-96 . In total, 422 cases of shigellosis, aged 1 month to 14 y (238M, 184F) were hospitalized during the study period . The annual distribution was approximately stable until 1990, the mean number of cases per year being about 20 . During the last 4 y the incidence significantly decreased . Shigella was serotyped in 138/422 cases . Seventy six of the strains were S . flexneri (55%) and 56 S . sonnei (40%) . In the majority of cases the clinical picture was mild . Severe dehydration was seen in only 6 patients . Ninety four patients (22%) had extra-intestinal manifestations . Most common of these were convulsions (16%) and, less frequently, disturbances of consciousness (n = 26), rash (n = 9), shock and disseminated intravascular coagulopathy (n = 2), nerve paralysis (n = 2), severe anaemia (n = 2) and haemolytic-uraemic syndrome (n = 1) . Nine patients had acute encephalopathy of 12 h to 12 d duration . It is important to note that all these cases recovered completely with no residual neurological deficit, except for 1 girl who developed temporal epilepsy 8 y later . Spinal fluid was normal in all 42 examined patients . Antibiotics were given to 212 of 422 patients, mainly during the first half of the study period . Shigella resistance to antibiotic was significant for cotrimoxazole (24%) and ampicillin (16%) . All patients were cured . Shigellosis is a mild disease in our area, with a decreasing prevalence.

J Food Prot, 2000 May, 63(5), 568 - 72
Fate of Shigella sonnei on parsley and methods of disinfection; Wu FM et al.; Outbreaks of shigellosis associated with chopped parsley used as a garnish for foods occurred in four states in the United States and in two Canadian provinces in 1998 . This prompted a study to determine survival and growth characteristics of Shigella sonnei inoculated onto raw parsley . Two inoculum levels (approximately 10(3) and 10(6) CFU/g) were applied to parsley leaves, portions of which were then chopped . Inoculated whole and chopped parsley leaves were held at 4 degrees C or 21 degrees C for up to 14 days . Initial populations of the organism on chopped parsley receiving high or low levels of inoculum increased by approximately 3 log10 CFU/g, within 1 day at 21 degrees C . Populations of S . sonnei on inoculated chopped or whole parsley leaves held at 4 degrees C decreased by 2.5 to 3.0 log10 CFU/g during a 14-day storage period . The pathogen multiplied, without a lag phase, on inoculated (2.72 log10 CFU/g) chopped parsley held at 21 degrees C, exceeding 6 log10 CFU/g within 24 h . Treatment of inoculated whole parsley leaves with vinegar containing 5.2% (vol/vol) acetic acid or 200 ppm free chlorine for 5 min at 21 degrees C reduced the population of S . sonnei by more than 6 log10 CFU/g, whereas treatment with vinegar containing 7.6% acetic acid or 250 ppm free chlorine reduced initial populations of 7.07 and 7.26 log10 CFU/g, respectively, to undetectable levels (<0.6 log10 CFU/g) . These studies revealed that S . sonnei can grow rapidly on chopped parsley held at ambient temperature and remain viable for at least 14 days at 4 degrees C . Treatment of contaminated parsley with vinegar or chlorinated water offers a simple method to reduce markedly or eliminate the pathogen in food-service or home settings.

Rapid Commun Mass Spectrom, 2000, 14(10), 911 - 7
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometric detection of bacterial biomarker proteins isolated from contaminated water, lettuce and cotton cloth; Holland RD et al.; Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectra of bacterial proteins were obtained from water, lettuce and cloth samples contaminated with Shigella flexneri, Escherichia coli, and Aeromonas hydrophila . Spectra were obtained using proteins directly isolated from water (or water used for rinsing samples) without culturing the bacteria . For S . flexneri and E . coli, two marker ions for specific proteins associated with a virulence-related property (acid resistance) were easily detected . For A . hydrophila, ions from two specifically selected marker proteins, as well as ions from the larger group of proteins isolated from pure cultures, all matched spectra from a contaminated water sample, providing strong evidence that A . hydrophila was the bacterial contaminant . Rinse water from contaminated lettuce and cloth samples showed the same marker ions as the contaminated water samples .

Acta Biochim Pol, 1999, 46(4), 853 - 61
Plasmid-mediated suppression of the mutational activation of the bgl operon in Shigella sonnei; Kharat AS et al.; SSOR, a clinical isolate of Shigella sonnei which exhibits a Salicin-negative phenotype, is unable to mutate to give rise to Sal+ derivatives although a homolog of the Escherichia coli bgl operon is retained by the strain . This was correlated to the presence of an endogenous plasmid in the strain . A plasmid-cured derivative, AK711, could give rise to Sal+ mutants in two steps . Introduction of the plasmid DNA, extracted from SSOR, into various strains of E . coli and S . sonnei, resulted in ampicillin resistant transformants . Interestingly, the presence of the plasmid suppressed the mutational activation of the bgl operon in the transformants . This was further substantiated by the observation that, transformants that have lost the plasmid regained the ability for mutational activation of the bgl operon . Preliminary characterisation of the plasmid indicated a size of 3.8 kb with an origin of replication resembling that of ColE1 replicons and the bla gene homolog of Tn3 . Observations of the mutation frequency at the srl and lac loci in the presence of the plasmid indicate that there is a reduction in the mutation frequency, suggesting an antimutator activity associated with the plasmid.

Infect Immun, 2000 Jun, 68(6), 3710 - 5
Interaction of Shigella flexneri IpaC with model membranes correlates with effects on cultured cells; Tran N et al.; Invasion of enterocytes by Shigella flexneri requires the properly timed release of IpaB and IpaC at the host-pathogen interface; however, only IpaC has been found to possess quantifiable activities in vitro . We demonstrate here that when added to cultured cells, purified IpaC elicits cytoskeletal changes similar to those that occur during Shigella invasion . This IpaC effect may correlate with its ability to interact with model membranes at physiological pH and to promote entry by an ipaC mutant of S . flexneri.

Infect Immun, 2000 Jun, 68(6), 3608 - 19
Shigella flexneri IpaH(7.8) facilitates escape of virulent bacteria from the endocytic vacuoles of mouse and human macrophages; Fernandez-Prada CM et al.; The behavior of Shigella flexneri ipaH mutants was studied in human monocyte-derived macrophages (HMDM), in 1-day-old human monocytes, and in J774 mouse macrophage cell line . In HMDM, strain pWR700, an ipaH(7.8) deletion mutant of S . flexneri 2a strain 2457T, behaved like the wild-type strain 2457T . This strain caused rapid host cell death by oncosis, and few bacterial CFU were recovered after incubation in the presence of gentamicin as previously described for 2457T-infected HMDM . However, analysis of bacterial compartmentalization within endocytic vacuoles with gentamicin and chloroquine indicated that more pWR700 than 2457T was present within the endocytic vacuoles of HMDM, suggesting that ipaH(7.8) deletion mutant transited more slowly from the vacuoles to the cytoplasm . In contrast to findings with HMDM, CFU recovered from pWR700-infected mouse J774 cells were 2 to 3 logs higher than CFU from 2457T-infected J774 cells . These values exceeded CFU recovered after infection of J774 cells with plasmid-cured avirulent strain M4243A1 . Incubation with gentamicin and chloroquine clearly showed that pWR700 within J774 cells was mostly present within the endocytic vacuoles . This distribution pattern was similar to that seen with M4243A1 and contrasted with the pattern seen with 2457T . Complementation of pWR700 with a recombinant clone expressing ipaH(7 . 8) restored the intracellular distribution of bacteria to that seen with the wild-type strain . Strains with deletions in ipaH(4.5) or ipaH(9.8), however, behaved like 2457T in both HMDM and J774 cells . The distribution profile of pWR700 in 1-day-old monocytes was similar to that seen in J774 cells . Like infected J774 cells, 1-day-old human monocytes demonstrated apoptosis upon infection with virulent Shigella . These results suggest that a role of the ipaH(7 . 8) gene product is to facilitate the escape of the virulent bacteria from the phagocytic vacuole of monocytes and macrophages.

Isr Med Assoc J, 2000 Feb, 2(2), 86 - 90
Induction of mRNA for tumor necrosis factor-alpha and interleukin-1 beta in mice brain, spleen and liver in an animal model of Shigella-related seizures; Nofech-Mozes Y et al.; BACKGROUND: The pathogenesis of neurological symptoms, the most common extraintestinal complication of childhood shigellosis, is unclear . To elucidate the mechanisms involved, we developed an animal model and demonstrated that TNF alpha and IL-1 beta play a role . OBJECTIVES: To determine whether TNF alpha and IL-1 beta genes are expressed in the brain following peripheral administration of Shigella dysenteriae 60R . METHODS: Expression of mRNA for TNF alpha and IL-1 beta was examined in the brain structures (hypothalamus and hippocampus) and peripheral organs by reverse transcriptase polymerase chain reaction, at different time points after intraperitoneal injection of S . dysenteriae sonicate . RESULTS: In our animal model of Shigella-related seizures, TNF alpha and IL-1 beta mRNA were induced in the brain, spleen and liver already 1 hour after injection of S . dysenteriae sonicate . The expression of TNF alpha and IL-1 beta mRNA in spleen, hippocampus and hypothalamus decreased after 6 h and increased again at 18 h post-injection . CONCLUSIONS: Local production of TNF alpha and IL-1 beta in the brain may be involved in the enhanced seizure response of mice after administration of S . dysenteriae . It is possible that intracerebral production of TNF alpha and IL-1 beta plays a role in neurological disturbances of human shigellosis.

Immunogenetics, 2000 Apr, 51(4-5), 314 - 25
A high incidence of Shigella-induced arthritis in a primate species: major histocompatibility complex class I molecules associated with resistance and susceptibility, and their relationship to HLA-B27; Urvater JA et al.; The human major histocompatibility complex (MHC) class I gene, HLA-B27, is a strong risk factor for susceptibility to a group of disorders termed spondyloarthropathies . Rodents that express HLA-B27 develop spondyloarthropathies, implicating HLA-B27 in the etiology of these disorders . To determine whether an HLA-B27-like molecule was associated with spondyloarthropathies in nonhuman primates, we analyzed the MHC class I cDNAs expressed in a cohort of rhesus macaques that developed reactive arthritis after an outbreak of shigellosis . We identified several cDNAs with only limited sequence similarity to HLA-B27 . Interestingly, one of these MHC molecules had a B pocket identical to that of HLA-B39 . Pool sequencing of radiolabeled peptides bound by this molecule demonstrated that, like HLA-B27 and HLA-B39, it could bind peptides with arginine at the second position . However, extensive analysis of the MHC class I molecules in this cohort revealed no statistically significant association between any particular MHC class I allele and susceptibility to reactive arthritis . Furthermore, none of the rhesus MHC class I molecules bore a strong resemblance to HLA-B27, indicating that reactive arthritis can develop in this animal model in the absence of an HLA-B27-like molecule . Surprisingly, there was a statistically significant association between the rhesus macaque MHC A locus allele, Mamu-A*12, and the absence of reactive arthritis following Shigella infection.

Diagn Microbiol Infect Dis, 2000 May, 37(1), 11 - 6
Detection of PCR products of the ipaH gene from Shigella and enteroinvasive Escherichia coli by enzyme linked immunosorbent assay; Sethabutr O et al.; PCR techniques applied to diarrheal stools reliably diagnose Shigella and enteroinvasive Escherichia coli (EIEC) infections . Identification of PCR products using agarose gel electrophoresis (AGE) and hybridization with DNA probes has several shortcomings . Automated methods of identifying PCR products that process larger numbers of specimens can facilitate epidemiologic studies and standardize results . In this study, we used ELISA following PCR to detect ipaH gene sequences of Shigella and EIEC from 89 diarrheal stools . Results of ELISA were compared with AGE with and without DNA probe, and with culture . Two specimen preparation methods were compared as well: boiling/centrifugation, and purification with silicon dioxide (SiO(2)) . Both PCR product-detection methods identified significantly more infections than did culture . PCR-ELISA detected significantly more infections than PCR-AGE when processed using SiO2 (P = 0.014) . PCR-ELISA allows screening of larger numbers of specimens, automates test results, and avoids use of mutagenic reagents . PCR-ELISA is faster than PCR-AGE when testing large numbers of specimens, although not when testing small numbers of specimens.

J Appl Microbiol, 2000 May, 88(5), 845 - 52
Evaluation of the environmental impact of microbial aerosols generated by wastewater treatment plants utilizing different aeration systems; Brandi G et al.; Using three sampler devices (SAS, Andersen Six-Stages and All Glass Impinger), the environmental impact of bacterial and fungal aerosols generated by municipal wastewater treatment plants operating with different methods of sludge oxygenation were evaluated . The highest microbial concentrations were recovered above the tanks (2247 cfu m-3) and in downwind positions (1425 cfu m-3), where a linear correlation (P < 0.05) was found between the quantity of sewage treated and the entities of microbial aerosol dispersion . Moreover, an exponential increase (P < 0.05) in the bacteria recovered from the air occurred at increasing times of treatment . However, after long-term plant operation, high bacterial and fungal concentrations were found in almost all of the sites around the plant . Coliforms, enterococci, Escherichia coli and staphylococci were almost always recovered in downwind positions . Considerable fractions (20-40%) of sampled bacteria were able to penetrate the final stages of the Andersen apparatus and thus, are likely to be able to penetrate the lungs . The plant operating with a fine bubble diffused air system instead was found to generate rather low concentrations of bacteria and fungi; moreover, staphylococci and indicator micro-organisms were almost absent . Finally, salmonellae, Shigella, Pseudomonas aeruginosa and Aeromonas spp . were not detected in either of the plants . The results indicate a remarkable dispersion of airborne bacteria and fungi from tanks in which oxygen is supplied via a mechanical agitation of sludge, and suggest the need to convert them to diffused aeration systems which pose a lesser hazard for human health.

Microbiol Immunol, 2000, 44(3), 161 - 72
The complete DNA sequence of the O antigen gene region of Plesiomonas shigelloides serotype O17 which is identical to Shigella sonnei form I antigen; Chida T et al.; We cloned and determined the sequence of a DNA region of approximately 15-kb containing the cluster of genes required for O17 antigen expression in the Escherichia coli K-12 strain from the chromosome of Plesiomonas shigelloides serotype O17:H2 strain . The sequencing analysis revealed that the minimum essential region of the P . shigelloides O17 antigen gene cluster had a size of approximately 11.5-kb and contained 9 contiguous open reading frames (ORFs), which were almost identical to the corresponding ORFs of Shigella sonnei form I antigen gene region, except for IS630 sequence, at the DNA as well as amino acid levels . The putative function of most of the ORFs could be determined on the basis of amino acid sequence similarities and characteristics . In addition, the G+C content of the P . shigelloides O17 antigen genes was lower than that of the chromosomal DNA of P . shigelloides and S . sonnei, suggesting that both P . shigelloides O17 and S . sonnei form I antigen genes had been derived from the same origin with a low G+C content.

J Biol Chem, 2000 Jul 21, 275(29), 21946 - 52
GRB2 links signaling to actin assembly by enhancing interaction of neural Wiskott-Aldrich syndrome protein (N-WASp) with actin-related protein (ARP2/3) complex; Carlier MF et al.; Proteins of the Wiskott-Aldrich Syndrome protein (WASp) family connect signaling pathways to the actin polymerization-driven cell motility . The ubiquitous homolog of WASp, N-WASp, is a multidomain protein that interacts with the Arp2/3 complex and G-actin via its C-terminal WA domain to stimulate actin polymerization . The activity of N-WASp is enhanced by the binding of effectors like Cdc42-guanosine 5'-3-O-(thio)triphosphate, phosphatidylinositol bisphosphate, or the Shigella IcsA protein . Here we show that the SH3-SH2-SH3 adaptor Grb2 is another activator of N-WASp that stimulates actin polymerization by increasing the amount of N-WASp . Arp2/3 complex . The concentration dependence of N-WASp activity, sedimentation velocity and cross-linking experiments together suggest that N-WASp is subject to self-association, and Grb2 enhances N-WASp activity by binding preferentially to its active monomeric form . Use of peptide inhibitors, mutated Grb2, and isolated SH3 domains demonstrate that the effect of Grb2 is mediated by the interaction of its C-terminal SH3 domain with the proline-rich region of N-WASp . Cdc42 and Grb2 bind simultaneously to N-WASp and enhance actin polymerization synergistically . Grb2 shortens the delay preceding the onset of Escherichia coli (IcsA) actin-based reconstituted movement . These results suggest that Grb2 may activate Arp2/3 complex-mediated actin polymerization downstream from the receptor tyrosine kinase signaling pathway.

J Bacteriol, 2000 May, 182(9), 2416 - 21
The stability region of the large virulence plasmid of Shigella flexneri encodes an efficient postsegregational killing system; Sayeed S et al.; The large virulence plasmid pMYSH6000 of Shigella flexneri contains a determinant that is highly effective in stabilizing otherwise unstable plasmids in Escherichia coli . Expression of two small contiguous genes, mvpA and mvpT (formerly termed STBORF1 and STBORF2), was shown to be sufficient for stability . Mutations in mvpT abolished plasmid stability, and plasmids expressing only mvpT killed the cells unless mvpA was supplied from a separate plasmid or from the host chromosome . When replication of a plasmid carrying the minimal mvp region was blocked, growth of the culture stopped after a short lag and virtually all of the surviving cells retained the plasmid . Thus, the mvp system stabilizes by a highly efficient postsegregational killing (PSK) mechanism, with mvpT encoding a cell toxin and mvpA encoding an antidote . The regions that surround the mvp genes in their original context have an inhibitory effect that attenuates plasmid stabilization and PSK . The region encompassing the mvp genes also appears to contain an additional element that can aid propagation of a pSC101-based plasmid under conditions where replication initiation is marginal . However, this appears to be a relatively nonspecific effect of DNA insertion into the plasmid vector.

East Afr Med J, 1999 May, 76(5), 255 - 9
Intrafamilial person-to-person spread of bacillary dysentery due to Shigella dysenteriae in southwestern Saudi Arabia; el Bushra HE et al.; OBJECTIVE: To identify the factors that influence transmission of bacillary dysentry (BD) within families during a propagated outbreak of bacillary dysentery . DESIGN: A retrospective cohort study . SETTING: Eighteen neighbouring villages in rural Gizan, southwestern Saudi Arabia . SUBJECTS: Two hundred and thirty three cases of BD were identified among seventy nine families . RESULTS: Secondary cases of BD occurred in 57 of 79 families with a primary case of BD . The secondary attack rate per cent (AR%) within families ranged between 7.7% and 80% . Age of primary cases did not correlate with degree of secondary AR% in exposed families (p > 0.04; p > 0.05); however, within households, the age of the first secondary cases (median = two years) was usually less than the age of the primary case (median = six years) . Children under five years of age constituted 43% of secondary cases . The median interval between successive cases within a house ranged from three and seven days . Two hundred and twenty cases (94.4%) gave history of close contact within another case of BD . Cases of BD were exposed to close relatives with BD (79.1%), neighbours (11.4%), and friends (9.5%) . Risk factors influencing the spread of BD within families included two rooms or fewer per house (OR = 4.3, 9.5% CI 1.3-14.3), family size of five or more (p = 0.012, two-tailed Fisher's exact test), and presence of more than two persons per room (OR = 11.2, 95% CI 3.1-42.4) . CONCLUSION: Person-to-person secondary transmission can amplify the spread of bacillary dysentery within households and neighbouring villages . Crowding was a risk factor that amplified transmission of BD within families.

Rev Panam Salud Publica, 2000 Feb, 7(2), 113 - 7
{Antimicrobial resistance of Shigella spp . in Córdoba, Argentina, during the period 1990-1997}; Suarez ME et al.; This study analyzed the evolution of antimicrobial resistance in 771 isolates of Shigella spp . obtained from a total of 9,195 feces cultures done between 1990 and 1997 in a children's hospital in Cordoba, Argentina . S . flexneri, which was responsible for 73% of the Shigella infections, was the species with the greatest resistance . The frequency of S . flexneri resistance to the three antibiotics most used (ampicillin, trimethoprim-sulfamethoxazole, and chloramphenicol) increased from 10% in 1990 to 58% in 1997 (P < 0.001) . Considering each of the drugs individually, the resistance to ampicillin increased from 60% to 100% (P < 0.001), the resistance to chloramphenicol from 13% to 71% (P < 0.001), and the resistance to trimethoprim-sulfamethoxazole from 79% to 84% (P = 0.22) . For S . sonnei, the increase in resistance to ampicillin (from 36% in 1990 to 54% in 1997) was not statistically significant (P = 0.20), nor was the reduction in resistance to trimethoprim-sulfamethoxazole, which went from 82% in 1990 to 55% in 1997 (P = 0.08) . Only two S . sonnei isolates were found that were resistant to chloramphenicol, one in 1995 and another in 1997; two S . sonnei isolates were found with resistance to all three antibiotics . We consider it essential to carry out susceptibility tests of each Shigella clinical isolate, to detect changes in the resistance profile and thus modify empiric treatment.

Eur J Clin Microbiol Infect Dis, 2000 Feb, 19(2), 108 - 11
Antimicrobial resistance trends in Shigella serogroups isolated in Israel, 1990-1995; Mates A et al.; From a total of 31319 Shigella strains isolated in Israel between 1990 and 1996, 17574 were sent to the National Shigella Reference Center for typing . Of these, 15287 were identified as Shigella sonnei, 1833 as Shigella flexneri, 327 as Shigella boydii and 127 as Shigella dysenteriae . In all, 4395 strains were tested for sensitivity to ampicillin, trimethoprim-sulfamethoxazole, tetracycline, chloramphenicol, nalidixic acid and ofloxacin . All strains tested were sensitive to ofloxacin, and only three strains were resistant to nalidixic acid . Only 113 of 3240 (3.5%) Shigella sonnei strains, 172 of 970 (17.7%) Shigella flexneri strains and 45 of 185 (24.3%) Shigella boydii strains tested were sensitive to four other antibiotic agents . The rates of resistance of Shigella sonnei, Shigella flexneri and Shigella boydii to trimethoprimsulfamethoxazole were 94.4%, 51.3% and 61.6%, respectively . Rates of resistance to ampicillin among these species were 73.4%, 63.5% and 21.4%, respectively . The proportion of strains exhibiting multiple drug resistance was higher for Shigella sonnei than for the other serotypes studied . These results emphasize the need to reassess the use of antibiotic agents in the treatment of shigellosis.

Cell Motil Cytoskeleton, 2000 Apr, 45(4), 272 - 8
Shigella actin-based motility in the presence of truncated vinculin; Southwick FS et al.; Mounting evidence supports the role of truncated vinculin in the intracellular actin-based motility of Shigella flexneri . Vinculin's role was recently questioned by Goldberg {1997: Cell Motil Cytoskeleton 37:44-53} who observed Shigella motility in mouse embryonal carcinoma 5.51 cells, a genetically modified cell line that reputedly lacked vinculin . That challenge implicitly relied on the assumption that 5.51 cells had no detectable vinculin polypeptide and lacked full-length vinculin mRNA . Despite the appearance of being an unambiguous test of vinculin's role in Shigella motility, 5.51 cells were shown to contain adequate amounts of truncated vinculin (as well as the corresponding mRNA transcript) to support bacterial locomotion . We also examined Shigella locomotion in gamma229 cells, a related embryonal carcinoma cell line containing approximately one-half the vinculin content found in 5.51 cells . We observed that there was a commensurate twofold decrease in the Shigella motility rate, as compared to 5.51 cells; this finding raises the possibility that vinculin can become a rate-limiting factor under some circumstances . Immunofluorescence microscopy using vin 11-5 monoclonal antibody directed against the vinculin head domain showed intense staining of Shigella rocket tails in both gamma229 and 5.51 cells . Our findings clearly demonstrate that motility in 5.51 cells cannot be regarded as a valid criterion for evaluating the role of truncated vinculin in Shigella motility .

Infect Dis Clin North Am, 2000 Mar, 14(1), 41 - 65, viii
Shigella and Shiga toxin-producing Escherichia coli causing bloody diarrhea in Latin America; Lopez EL et al.; In Latin America, Shigella and shiga toxin-producing Escherichia coli are the two leading agents in the cause of bloody diarrhea . The already high and increasing antimicrobial resistance of Shigella also is a significant problem . Shiga toxin-producing E . coli is an emerging disease with life-threatening complications: hemolytic uremic syndrome . Although E . coli O157:H7 remains the most commonly recognized serotype, recently emerging, non-O157 bacteria may be the cause of a similar spectrum of disease in humans.

Carbohydr Res, 2000 Jan 29, 324(1), 53 - 65
NMR spectroscopy, molecular dynamics, and conformation of a synthetic octasaccharide fragment of the O-specific polysaccharide of Shigella dysenteriae type 1; Coxon B et al.; A synthetic octasaccharide fragment (2) of the O-specific polysaccharide (1) of Shigella dysenteriae type 1 has been studied as its methyl glycoside by one- and two-dimensional homo- and heteronuclear NMR spectroscopy . Complete 1H and 13C NMR assignments have been generated, and the 13C spin-lattice relaxation times have been measured for the octasaccharide 2 . A congener (6) of this octasaccharide containing one D-galactose residue with a specific 13C label at C-1 has been synthesized and used to measure interglycosidic 13C-1H coupling by the 2D J-resolved 1H NMR method . From the NMR data, three types of conformational restraints were developed: (a) 29 inter-residue, distance restraints; (b) 48 intra-residue, ring atom dihedral angle restraints, and (c) one heteronuclear, inter-residue dihedral angle restraint . The use of these restraints in a restrained molecular dynamics computation with simulated annealing yielded a conformation resembling a short, irregular spiral, with methyl substituents on the exterior.

Clin Infect Dis, 2000 Mar, 30(3), 515 - 9
Emergence of antimicrobial-resistant shigellosis in Oregon; Replogle ML et al.; Ampicillin and trimethoprim-sulfamethoxazole (TMP-SMZ) are currently considered acceptable empirical therapy for shigellosis in developed countries . However, there are few recently reported studies on antimicrobial resistance among shigellae isolated in the United States . We examined the epidemiology of shigellosis and the antimicrobial susceptibility of Shigella species isolated in Oregon from July 1995 through June 1998 . Of 430 isolates, 410 were identified to the species level: Shigella sonnei accounted for 55% of isolates, and Shigella flexneri, for 40% . The overall annual incidence of shigellosis was 4.4 cases per 100,000 population . Children aged <5 years (annual incidence, 19.6 cases per 100,000 population) and Hispanics (annual incidence, 28.4 cases per 100,000 population) were at highest risk . Of 369 isolates tested, 59% were resistant to TMP-SMZ, 63% were resistant to ampicillin, 1% were resistant to cefixime, and 0.3% were resistant to nalidixic acid; none of the isolates were resistant to ciprofloxacin . Thirteen percent of the isolates had multidrug resistance to ampicillin, chloramphenicol, streptomycin, sulfisoxazole, and tetracycline . Infections due to multidrug-resistant shigellae are endemic in Oregon . Neither ampicillin nor TMP-SMZ should be considered appropriate empirical therapy for shigellosis any longer; when antibiotics are indicated, a quinolone or cefixime should be used.

Microbios, 2000, 101(398), 37 - 46
Effects of caffeic acid, chlorogenic acid and ferulic acid on growth and arylamine N-acetyltransferase activity in Shigella sonnei (group D); Tsou MF et al.; Arylamine N-acetyltransferase (NAT) activities with 2-aminofluorene (2-AF) as substrates were determined in Shigella sonnei (group D) collected from patients with diarrhoeal disease . The NAT activity was determined using an acetyl CoA recycling assay and high pressure liquid chromatography . Inhibition of growth studies from S . sonnei (group D) demonstrated that caffeic acid (CA), chlorogenic acid (CGA) and ferulic acid (FA) elicited a dose-dependent bactericidal effect in S . sonnei (group D) cultures, i.e . the greater the concentration of CA, CGA and FA, the greater the inhibition of growth of S . sonnei (group D) . Cytosols or suspensions of S . sonnei (group D) with and without selected concentrations of CA, CGA and FA co-treatment showed different percentages of 2-AF acetylation . The data indicated that there was reduced NAT activity associated with increased CA, CGA and FA in Shigella dysenteriae (group D) cytosols and intact cells . For the cytosol and intact bacteria examinations, the apparent values of K(m) and Vmax decreased after being co-treated with 400 microM CA, CGA and FA . This report is the first demonstration of plant phenolic inhibition (CA, CGA and FA) of arylamine NAT activity and growth in the bacterium S . sonnei (group D).

Vaccine, 2000 Apr 28, 18(21), 2193 - 202
DeltaguaBA attenuated Shigella flexneri 2a strain CVD 1204 as a Shigella vaccine and as a live mucosal delivery system for fragment C of tetanus toxin; Anderson RJ et al.; The DeltaguaBA Shigella flexneri 2a vaccine candidate, CVD 1204, was evaluated as a delivery system for the non-toxic C-terminal of tetanus toxin (fragment C), either as a polypeptide expressed in the bacteria or as a DNA vaccine . CVD 1204 was transformed with plasmid pTETnir15 which encodes the fragment C gene (tetC) under the control of the inducible prokaryotic nir15 promoter or a DNA vaccine plasmid pcDNA3tetC which encodes tetC under the eukaryotic hCMV promoter . Guinea pigs immunised intranasally (i.n.) with either recombinant strain mounted a secretory immune response against S . flexneri 2a Lipopolysaccharide (LPS) and were protected against ocular challenge with wild-type S . flexneri 2a . Both strains were effective in eliciting a serum IgG response against fragment C in guinea pigs following i.n . immunisation . Furthermore, serum from guinea pigs immunised with CVD 1204(pTETnir15) contained tetanus toxin neutralising antibodies . These results demonstrate that this S . flexneri 2a vaccine candidate can serve as a vehicle for the delivery of foreign antigens to the systemic immune system while retaining its capacity to serve as a mucosal Shigella vaccine.

Mol Microbiol, 2000 Mar, 35(5), 974 - 90
The development of a FACS-based strategy for the isolation of Shigella flexneri mutants that are deficient in intercellular spread; Rathman M et al.; In the disease course of bacillary dysentery, pathogenic Shigella flexneri invade colonic epithelial cells and spread both within and between host cells . The ability to spread intercellularly allows the organism to infect an entire epithelial layer without significant contact with the extracellular milieu . Using fluorescence activated cell sorter (FACS)-based technology, we developed a rapid and powerful selection strategy for the isolation of S . flexneri mutants that are unable to spread from cell to cell . The majority of mutants identified using this strategy harbour mutations that affect the structure of their lipopolysaccharide or the ability of the bacteria to move intracellularly via actin-based motility; both factors have previously been shown to be essential for cell-to-cell spread . However, using a modified strategy that eliminated both of these types of mutants, we identified several mutants that provide us with evidence that bacterial proteins of the type III secretion system, which are essential for bacterial entry into host cells, also play a role in cell-to-cell spread.

Eur J Biochem, 2000 Mar, 267(6), 1672 - 9
Structural studies of the O-specific polysaccharide from Plesiomonas shigelloides strain CNCTC 113/92; Czaja J et al.; The structure of the O-specific side chain of the lipopolysaccharide (LPS) of Plesiomonas shigelloides, strain CNCTC 113/92 has been investigated by NMR spectroscopy, matrix-assisted laser desorption/ionization time of flight mass spectrometry and sugar and methylation analysis . It was concluded that the polysaccharide is composed of a hexasaccharide repeating unit with the following structure: in which D-beta-D-Hepp is Dglycero-beta-Dmanno-heptopyranose and 6d-beta-D-Hep is 6-deoxy-beta-Dmanno-heptopyranose . This structure represents a novel hexasaccharide repeating unit of bacterial O-antigen that is characteristic and unique to the Plesiomonas shigelloides strain . Using the high-resolution magic angle spinning technique, 1H-NMR spectra were also obtained for the O-polysaccharide components of isolated LPS and in their original form directly on the surface of bacterial cells.

Mikrobiol Z, 1999 Nov-Dec, 61(6), 36 - 41
{The role of the structural components of bacterial lipopolysaccharide in its inductive immunosuppressive activity}; Borisova EV; The inductive immunosuppressive activity of lipopolysaccharides (LPS) and structural parts (O-chains,cor,lipid A) obtained from bacteria of the genus Shigella and Escherichia coli was studied . LPS preparations were extracted by phenol-water method . Its structural components were obtained by the acetic acid hydrolysis and gel filtration . It has been shown that all these preparations did not change the level of delayed type hypersensitivity (DTH) to test-antigen in mice in the case of intraperitoneal injection . They did not also induce immunosuppressive activity in avirulent bacterial strains . After the redox treatment all LPS preparations acquired the ability to induce immunosuppressive property in avirulent bacteria . As a result of redox treatment the immunosuppressive activity appeared in lipid A preparations . O-chains and cor preparations were not active . Immunosuppressive activity was lost after treatment with phenol . It is supposed that active chemical groups are located in lipid A.

J Immunol, 2000 Mar 15, 164(6), 3214 - 21
Blockade of CD14 increases Shigella-mediated invasion and tissue destruction; Wenneras C et al.; Shigella is a diarrheal pathogen that causes disease through invasion of the large intestinal mucosa . The endotoxin of the invading bacterium may play a key role in the disease process by causing inflammation and tissue injury during infection . Earlier studies have shown that various animal species lacking functional CD14 were protected against endotoxin-mediated shock . Rabbits experimentally infected with Shigella were used to test the hypothesis that blockade of endotoxin-induced cell activation with anti-CD14 mAb would diminish inflammation and thus disease severity . Unexpectedly, we observed that the intestinal mucosa of anti-CD14-treated animals exhibited a 50-fold increase in bacterial invasion and more severe tissue injury compared with controls . Despite higher bacterial loads in treated animals, the numbers of polymorphonuclear leukocytes that were recruited to the infection site were similar to those in controls . Furthermore, the phagocytic cells of CD14-blocked animals produced IL-1 and TNF-alpha . Moreover, in vitro blockade of CD14 did not impede bactericidal activity . Thus, anti-CD14 treatment interfered with host defense mechanisms involved with removal/eradication of Shigella.

Med Trop (Mars), 1999, 59(3), 259 - 65
{Acquired antibiotic resistance in Madagascar: first evaluation}; Decousser JW et al.; The purpose of this study was to evaluate the incidence of acquired resistance to antibiotics in Madagascar . Testing was carried out on total of 1267 strains of medically significant bacteria isolated from specimens sent to the Pasteur Institute of Madagascar in Antananarivo between October 1997 and October 1998 . Antibiograms were performed using the diffusion technique on gel media with antibiotic disks . Results were read according to the criteria of the Antibiogram Committee of the French Society of Microbiology . Preliminary findings documented a high incidence of resistance to widely available, low-price antibiotics including penicillin G and tetracycline for which 84 p . 100 and 65 p . 100 of Staphylococcus aureus respectively demonstrated resistance; tetracyclin to which 80 p . 100 of streptococcus were resistant; and ampicillin, cotrimoxazole, and phenicoles to which 60 p . 100, 60 p . 100 and 28 p . 100 of Escherichia coli respectively and 77 p . 100, 83 p . 100, and 71 p . 100 of Shigella sp . respectively were resistant . Second-line antibiotics including penicillin M, macrolides, nalidixic acid, and nitrofuranes were still relatively active, thus providing an effective alternative . Newly developed antibiotics such as fluoroquinolones and third-generation cephalosporines were highly effective but a few resistant strains were observed . Although not representative of Madagascar as a whole, the findings of this preliminary study indicate that acquired resistance must be taken into account in designing simplified decision charts for front-line laboratories, that appropriate information must be made available to health care workers, and that further testing is needed to monitor the evolution of antibiotic resistance.

Nat Med, 2000 Mar, 6(3), 265 - 70
A new biological agent for treatment of Shiga toxigenic Escherichia coli infections and dysentery in humans; Paton AW et al.; Gastrointestinal disease caused by Shiga toxin-producing bacteria (such as Escherichia coli O157:H7 and Shigella dysenteriae) is often complicated by life-threatening toxin-induced systemic sequelae, including hemolytic-uremic syndrome . Such infections can now be diagnosed very early in the course of the disease, but at present no effective therapeutic intervention is possible . Here, we constructed a recombinant bacterium that displayed a Shiga toxin receptor mimic on its surface, and it adsorbed and neutralized Shiga toxins with very high efficiency . Moreover, oral administration of the recombinant bacterium completely protected mice from challenge with an otherwise 100%-fatal dose of Shiga toxigenic E . coli . Thus, the bacterium shows great promise as a 'probiotic' treatment for Shiga toxigenic E . coli infections and dysentery.

Mol Microbiol, 2000 Feb, 35(4), 924 - 35
Transfer RNA modification, temperature and DNA superhelicity have a common target in the regulatory network of the virulence of Shigella flexneri: the expression of the virF gene; Durand JM et al.; Full expression of the virulence genes of Shigella flexneri requires the presence of two modified nucleosides in the tRNA {queuosine, Q34, present in the wobble position (position 34) and 2-methylthio-N6-isopentenyladenosine (ms2i6A37, adjacent to and 3' of the anticodon)} . The synthesis of these two nucleosides depends on the products of the tgt and miaA genes respectively . We have shown that the intracellular concentration of the virulence-related transcriptional regulator VirF is reduced in the absence of either of these modified nucleosides . The intracellular concentration of VirF is correlated with the expression of the virulence genes . Overproduction of VirF in the tgt and the miaA mutants suppressed the less virulent (tgt) or the avirulent (miaA) phenotypes respectively, caused by the tRNA modification deficiency . This suggests that the primary result of undermodification of the tRNA is a poor translation of virF mRNA and not of any other mRNA whose product acts downstream of the action of VirF . Shigella showed no virulence phenotypes at 30 degrees C, but forced synthesis of VirF at 30 degrees C induced the virulence phenotype at this low temperature . In addition, removal of the known gene silencer H-NS by a mutation in its structural gene hns increased the synthesis of VirF at low temperature and thus induced a virulent phenotype at 30 degrees C . Conversely, decreased expression of VirF at 37 degrees C induced by the addition of novobiocin, a known inhibitor of gyrase, led to an avirulent phenotype . We conclude that tRNA modification, temperature and superhelicity have the same target - the expression of VirF - to influence the expression of the central regulatory gene virB and thereby the virulence of Shigella . These results further strengthen the suggestion that the concentration of VirF is the critical factor in the regulation of virulence in Shigella . In addition, they emphasize the role of the bacterial translational machinery in the regulation of the expression of virulence genes which appears here quantitatively as important as the well-established regulation on the transcriptional level.

Infect Immun, 2000 Mar, 68(3), 1289 - 96
Induction of necrosis in human neutrophils by Shigella flexneri requires type III secretion, IpaB and IpaC invasins, and actin polymerization; Francois M et al.; Infection by Shigella flexneri is characterized by infiltration of neutrophils in the intestinal mucosa and by a strong inflammatory reaction . Although neutrophils are constitutively programmed to die by apoptosis, we show that isolated human neutrophils undergo necrosis 2 h after infection with virulent S . flexneri strain M90T but not with the virulence plasmid-cured strain BS176 . This was demonstrated by the release of azurophil granule proteins concomitant with the release of lactate dehydrogenase (LDH), disruption of the plasma membrane, and absence of DNA fragmentation . Mutants with the mxiD1 gene, coding for an essential component of the secretion type III machinery, or the genes coding for IpaB or IpaC invasins deleted were not cytotoxic . Neutrophil necrosis occurred independently of the bacterial ability to leave phagosomes, and it involved actin polymerization, as the addition of cytochalasin D after phagocytosis of Shigella inhibited the release of LDH . In conclusion, Shigella kills neutrophils by necrosis, a process characterized by the release of tissue-injurious granular proteins . This probably contributes to disruption of the epithelial barrier, leading to the dysentery observed in shigellosis and allowing Shigella to enter its host cells.

Infect Immun, 2000 Mar, 68(3), 1034 - 9
Shigella flexneri 2a strain CVD 1207, with specific deletions in virG, sen, set, and guaBA, is highly attenuated in humans; Kotloff KL et al.; A phase 1 clinical trial was conducted among 35 healthy adult volunteers to evaluate the safety, immunogenicity, and shedding of different doses of CVD 1207, a live attenuated Shigella flexneri 2a vaccine candidate with specific deletion mutations in virG, sen, set, and guaBA . CVD 1207 retains the ability to invade epithelial cells but cannot effectively spread intercellularly after invasion (DeltavirG), does not produce enterotoxin (Deltasen and Deltaset), and has limited proliferation in vivo (DeltaguaBA) . In a consecutive fashion, groups of three to seven subjects ingested a single oral dose of CVD 1207 at an inoculum of either 10(6), 10(7), 10(8), 10(9), or 10(10) CFU . CVD 1207 was remarkably well-tolerated at inocula as high as 10(8) CFU . In comparison, one of 12 subjects who received 10(9) CFU experienced mild diarrhea and another experienced a single episode of emesis . One of five subjects who received 10(10) CFU experienced watery diarrhea and emesis . All subjects who ingested doses of 10(8) to 10(10) CFU excreted the vaccine; in 23 of 25, the duration of excretion was </=3 days . A dose-related, immunoglobulin A antibody-secreting cell (ASC) response to S . flexneri 2a O-specific lipopolysaccharide was seen, with geometric mean peak values of 6.1 to 35.2 ASCs/10(6) peripheral blood mononuclear cells (PBMC) among recipients of 10(7) to 10(10) CFU . The cytokine response to Shigella-specific antigens observed in volunteers' PBMC following vaccination suggested a Th1 pattern with stimulation of gamma interferon and absence of interleukin 4 (IL-4) or IL-5 . CVD 1207 represents a Shigella live oral vaccine strain prepared from wild-type S . flexneri 2a by rational use of recombinant DNA technology that achieves a remarkable degree of attenuation compared with earlier recombinant strains, even when administered at high dosage.

Comp Immunol Microbiol Infect Dis, 2000 Jan, 23(1), 45 - 51
Isolation, biochemical and serological characterisation of Plesiomonas shigelloides from freshwater in Northern Europe; Krovacek K et al.; Isolation and characterisation of Plesiomonas shigelloides from fresh water in Northern Europe is reported in this study . The organisms were isolated from two lakes and a river in Sweden . All isolates of P . shigelloides showed an identical biochemical profile and belonged to different serotypes, namely, O18, O23, O26, O58 and O60 . The study indicates that P . shigelloides may occur in the aquatic environment of cold climates and as a result, it is likely to be associated with human infections caused by waterborne pathogens in geographical areas with similar climatic conditions.

Cent Eur J Public Health, 1999 Nov, 7(4), 172 - 5
Plesiomonas shigelloides in water and fish; Aldova E et al.; Plesiomonas shigelloides strains isolated from aquariums, surface water and fish were serotyped using the international antigenic scheme and a provisional scheme designed for environmental strains, called the "Schubert" scheme . A remarkable variety of serovars represented in both schemes was found, namely in surface water samples and in fish.

J Immunol, 2000 Feb 15, 164(4), 2221 - 32
Production of IFN-gamma and IL-10 to Shigella invasins by mononuclear cells from volunteers orally inoculated with a Shiga toxin-deleted Shigella dysenteriae type 1 strain; Samandari T et al.; Volunteers were orally administered invasive, non-Shiga toxin-producing Shigella dysenteriae 1 to establish a challenge model to assess vaccine efficacy . In stepwise fashion, four separate groups were given 3 x 10(2), 7 x 10(3), 5 x 10(4), or 7 x 10(5) CFU . Using PBMC, proliferative responses and cytokine production were measured to S . dysenteriae whole-cell preparations and to purified recombinant invasion plasmid Ags (Ipa) C and IpaD . Anti-LPS and anti-Ipa Abs and Ab-secreting cells were also evaluated . Preinoculation PBMC produced considerable quantities of IL-10 and IFN-gamma, probably secreted by monocytes and NK cells, respectively, of the innate immune system . Following inoculation, PBMC from 95 and 87% of volunteers exhibited an increased production of IFN-gamma and IL-10, respectively, in response to Shigella Ags . These increases included responses to IpaC and IpaD among those volunteers receiving the lowest inoculum . No IL-4 or IL-5 responses were detected . Whereas there were no Ab or Ab-secreting cell responses in volunteers receiving the lowest inoculum, other dose groups had moderate to strong anti-LPS and anti-Ipa responses . These results suggest that in humans, type 1 responses play an important role in mucosal and systemic immunity to S . dysentariae 1.

Appl Environ Microbiol, 2000 Feb, 66(2), 844 - 9
Selective and sensitive method for PCR amplification of Escherichia coli 16S rRNA genes in soil; Sabat G et al.; A set of PCR primers targeting 16S rRNA gene sequences was designed, and PCR parameters were optimized to develop a robust and reliable protocol for selective amplification of Escherichia coli 16S rRNA genes . The method was capable of discriminating E . coli from other enteric bacteria, including its closest relative, Shigella . Selective amplification of E . coli occurred only when the annealing temperature in the PCR was elevated to 72 degrees C, which is 10 degrees C higher than the optimum for the primers . Sensitivity was retained by modifying the length of steps in the PCR, by increasing the number of cycles, and most importantly by optimizing the MgCl(2) concentration . The PCR protocol developed can be completed in less then 2 h and, by using Southern hybridization, has a detection limit of ca . 10 genomic equivalents per reaction . The method was demonstrated to be effective for detecting E . coli DNA in heterogeneous DNA samples, such as those extracted from soil.

Mol Med Today, 2000 Feb, 6(2), 66 - 71
Recent advances with recombinant bacterial vaccine vectors; Shata MT et al.; Bacille Calmette-Guerin (BCG), Listeria monocytogenes, Salmonellae and Shigellae have shown promise as vaccine vectors in experimental animal models . Although disappointing results in humans and non-human primates stalled the development of this vaccination strategy, interest in this approach was reinvigorated recently by the development of bacterial DNA-vaccine-vectors . The purpose of this review is to highlight the strengths and weaknesses of bacterial vaccine vectors, and to discuss the future prospects of these vaccine delivery systems.

J Microbiol Immunol Infect, 1999 Dec, 32(4), 278 - 82
{Relatedness of Shigella sonnei isolates from six outbreaks in Tao-Yuan area, Taiwan}; Lin CS et al.; Between November 1995 and January 1996, one larger-scale outbreak of Shigella sonnei infection occurred in the area of Tao-Yuan, Taiwan . Subsequently, five outbreaks of S . sonnei infection occurred in the same area during the period from October 1998 to February 1999 . Fifty-five isolates obtained from these six outbreaks were differentiated by antimicrobial susceptibility testing, plasmid profile analysis (PPA) and pulsed-field gel electrophoresis (PFGE) . They were classified into two and three types by antimicrobial susceptibility testing and PPA, respectively . However, only one pattern was observed by PFGE . The molecular epidemiology by PFGE in this study indicated that they were closely-related . These data suggest that the clinical isolates obtained during the period of January 1998 to January 1999 were probably derived from the outbreak strain of 1995.

FEMS Immunol Med Microbiol, 2000 Feb, 27(2), 139 - 45
Effect of erythromycin on Shigella infection of Caco-2 cells; Honma Y et al.; Erythromycin (EM), one of the macrolides, shows a dose-dependent effect on Shigella flexneri invasion of Caco-2 cells even at concentrations less than the minimum inhibitory concentration (subMIC) . LS13, a strain of S . flexneri 1b, invaded Caco-2 cells in vitro . When the strain was treated with subMIC of EM, the invasion efficiency decreased . The carrier rate of the invasion plasmid containing virulence genes was reduced by EM treatment, as determined by the colony pigmentation test on Congo red agar plates . Presence of the invasion plasmid was found to increase susceptibility of the organisms to EM . The growth of virulent organisms carrying the invasion plasmid was inhibited at 25 microg ml(-1) of EM, whereas the growth of organisms without the plasmid was inhibited at 100 microg ml(-1) of EM . This was supported by the finding that the MIC of EM for a virulent isolate of S . flexneri 2a YSH6000 (6.25 microg ml(-1)) and for the mutant strain del-17 (50 microg ml(-1)), carrying the type III apparatus, impaired plasmid . These findings suggested that EM passed through the type III apparatus and suppressed the growth of invasive organisms selectively . This mechanism may account for the clinical effect of EM on shigellosis.

Trends Microbiol, 2000 Jan, 8(1), 17 - 23
Serotype-converting bacteriophages and O-antigen modification in Shigella flexneri; Allison GE et al.; O-antigen modification (serotype conversion) in Shigella flexneri, which is an important virulence determinant, is conferred by temperate bacteriophages . Several serotype-converting phages have been isolated and preliminary characterization has identified the genes involved in O-antigen modification, and has also provided insight into the molecular biology of these phages.

Bioorg Med Chem Lett, 2000 Jan 3, 10(1), 19 - 21
Synthesis of the repeating unit of the O-specific polysaccharide of Shigella sonnei and quantitation of its serologic activity; Toth A et al.; The chemical synthesis of the zwitterionic disaccharide 2 is described that corresponds to the repeating unit of the O-specific polysaccharide (1) of the gram-negative human pathogen Shigella sonnei . Passive hemolysis inhibition tests using a hyperimmune rabbit serum raised against S . sonnei showed that the serologic activity of the disaccharide 2 is nearly 2- to 3-fold higher than those of its component monosaccharides . NMR data of 2 are in support of the proposed structure of the O-specific polysaccharide.

Kansenshogaku Zasshi, 1999 Nov, 73(11), 1130 - 9
{Outbreak case caused by different colicin type of Shigella sonnei in a day nursery in Tokyo (1998)}; Yamada S et al.; From October through December, 1998, person to person infection caused by Shigella sonnei had occurred in the day nursery in Hachioji-city Tokyo, and a total of 41 patients including 3 suspected cases and 3 carriers had been confirmed . Although one case was imported case which is one-year-old kindergarten child, the remaining was domestic case . Of patients, 33 cases (80.5%) were a three-year-old kindergarten children and their families, accounting for 20 and 13 cases, respectively . From the data of symptom onset of patients, epidemic could distinguish to the former part and the later part . The recur or reinfection, and re-detection case was observed in the later part . Clinical symptom of the patients was diarrhea (100%), fever (80%) and abdominal cramps (70%), and LVFX for adult or FOM for child was used for the therapy . In the colicin typing test and the antibiotic susceptibility test for 9 reagents, almost isolates in the former part were type 0 and susceptible, and MBC for FOM ranged from 6.25 to 100 micrograms/ml, whereas those in the later part was type 2 and resistant for the TC, and showed highly MBC for FOM with 50 or 100 micrograms/ml . However, both isolates showed same patterns in plasmid profiles and DNA fingerprints by RAPD analysis . On the other hand, the strain from imported case was also colicin 0, but it was different as regards resistant for ABPC and ST and two genetic analysis.

J Mol Biol, 2000 Jan 21, 295(3), 605 - 12
HDEA, a periplasmic protein that supports acid resistance in pathogenic enteric bacteria; Gajiwala KS et al.; The X-ray crystal structure of the Escherichia coli stress response protein HDEA has been determined at 2.0 A resolution . The single domain alpha-helical protein is found in the periplasmic space, where it supports an acid resistance phenotype essential for infectivity of enteric bacterial pathogens, such as Shigella and E . coli . Functional studies demonstrate that HDEA is activated by a dimer-to-monomer transition at acidic pH, leading to suppression of aggregation by acid-denatured proteins . We suggest that HDEA may support chaperone-like functions during the extremely acidic conditions .

FEMS Microbiol Lett, 2000 Jan 1, 182(1), 93 - 8
I-CeuI fragment analysis of the Shigella species: evidence for large-scale chromosome rearrangement in S . dysenteriae and S . flexneri; Shu S et al.; I-CeuI fragments of four Shigella species were analyzed to investigate their taxonomic distance from Escherichia coli and to collect substantiated evidence of their genetic relatedness because their ribosomal RNA sequences and similarity values of their chromosomal DNA/DNA hybridization had proved their taxonomic identity . I-CeuI digestion of genomic DNAs yielded seven fragments in every species, indicating that all the Shigella species contained seven sets of ribosome RNA operons . To determine the fragment identities, seven genes were selected from each I-CeuI fragment of E . coli strain K-12 and used as hybridization probes . Among the four Shigella species, S . boydii and S . sonnei showed hybridization patterns similar to those observed for E . coli strains; each gene probe hybridized to the I-CeuI fragments with sizes similar to that of the corresponding E . coli fragment . In contrast, S . dysenteriae and S . flexneri showed distinct patterns; rcsF and rbsR genes that located on different I-CeuI fragments in E . coli, fragments D and E, were found to co-locate on a fragment . Further analysis using an additional three genes that located on fragment D in K-12 revealed that some chromosome rearrangements involving the fragments corresponding to fragments D and E of K-12 took place in S . dysenteriae and S . flexneri.

J Food Prot, 1999 Dec, 62(12), 1475 - 7
Hemolytic and elastolytic activities influenced by iron in Plesiomonas shigelloides; Santos JA et al.; The aim of this study was to determine the presence of hemolytic and elastolytic enzymes in several strains of Plesiomonas shigelloides in relation to the availability of iron in culture media . Hemolytic activity and elastolytic activity were detected in strains of P . shigelloides and were enhanced when the strains were grown in an iron-depleted medium and lost after thermal treatment at 100 degrees C for 10 min . Also, elastolytic activity was inactivated by phenylmethyl sulfonyl fluoride, an inhibitor of serine proteases . Hemolytic activity was detected extracellularly in cell-free supernatants, whereas elastin degradation activity was cell associated . Both activities may be related to the virulence of P . shigelloides.

Infect Immun, 2000 Jan, 68(1), 382 - 6
Interaction of enteropathogenic or enterohemorrhagic Escherichia coli with HeLa cells results in translocation of cortactin to the bacterial adherence site; Cantarelli VV et al.; Infection of cultured HeLa epithelial cells with enteropathogenic Escherichia coli (EPEC) or enterohemorrhagic E . coli (EHEC) O157:H7 results in accumulation of cortactin under the adherent bacteria . Tyrosine phosphorylation of cortactin is not induced following HeLa cell infection with EHEC or EPEC, contrary to what has been reported to occur with Shigella flexneri.

Emerg Infect Dis, 1999 Nov-Dec, 5(6), 820 - 3
HIV infection as a risk factor for shigellosis; Baer JT et al.; We investigated cases of shigellosis in San Francisco and Alameda Counties identified during 1996 by active laboratory surveillance to assess the role of HIV infection as a risk factor for shigellosis . Dramatically elevated rates of shigellosis in HIV-infected persons implicate HIV infection as an important risk factor for shigellosis in San Francisco.

J Bacteriol, 1999 Dec, 181(24), 7614 - 7
Promiscuous origin of a chimeric sequence in the Escherichia coli O157:H7 genome; LeClerc JE et al.; A novel sequence of 2.9 kb in the intergenic region between the mutS and rpoS genes of Escherichia coli O157:H7 and closely related strains replaces a sequence of 6.1 kb in E . coli K-12 strains . At the same locus in Shigella dysenteriae type 1, a sequence identical to that in O157:H7 is bounded by the IS1 insertion sequence element . Extensive polymorphism in the mutS-rpoS chromosomal region is indicative of horizontal transfer events.

Drugs, 1999, 58 Suppl 2, 55 - 9
Quinolone use in the developing world: state of the art; Tupasi TE; The development of bacterial resistance to previously recommended standard therapy for common childhood infections in developing countries, notably dysentery and invasive salmonellosis, has left treatment options limited to the quinolones . A number of randomised controlled trials of the quinolones in shigellosis have demonstrated their efficacy and safety in children . Strategies to make these drugs available in developing countries for specific indications should be devised . Quinolones, however, should be limited to specific indications in order to prevent the emergence of bacterial resistance and thus preserve their clinical efficacy for important childhood infectious diseases in developing countries.

Mol Microbiol, 1999 Dec, 34(5), 1058 - 69
Spontaneous tandem amplification and deletion of the shiga toxin operon in Shigella dysenteriae 1; McDonough MA et al.; Only one species of Shigella, Shigella dysenteriae 1, has been demonstrated to produce Shiga toxin (Stx) . Stx is closely related to the toxins produced by Shiga toxin-producing Escherichia coli (STEC) . In STEC, these toxins are often encoded on lambdoid bacteriophages and are major virulence factors for these organisms . Although the bacteriophage-encoded stx genes of STEC are highly mobile, the stx genes in S . dysenteriae 1 have been believed to be chromosomally encoded and not transmissible . We have located the toxin genes of S . dysenteriae 1 to a region homologous to minute 30 of the E . coli chromosome, within a 22.4 kbp putative composite transposon bracketed by IS600 insertion sequences . This region is present in all the S . dysenteriae 1 strains examined . Tandem amplification occurs via the flanking insertion sequences, leading to increased toxin production . The global regulatory gene, fnr, is located within the stx region, allowing deletions of the toxin genes to be created by anaerobic growth on chlorate-containing medium . Deletions occur by recombination between the flanking IS600 elements . Lambdoid bacteriophage genes are found both upstream and within the region, and we demonstrate the lysogeny of Shigella species with STEC bacteriophages . These observations suggest that S . dysenteriae 1 originally carried a Stx-encoding lambdoid prophage, which became defective due to loss of bacteriophage sequences after IS element insertions and rearrangements . These insertion sequences have subsequently allowed the amplification and deletion of the stx region.

J Cell Sci, 2000 Jan, 113 ( Pt 1), 71 - 80
Src tyrosine kinase activity down-regulates Rho-dependent responses during Shigella entry into epithelial cells and stress fibre formation; Dumenil G et al.; Invasion of epithelial cells by Shigella, the causative agent of bacillary dysentery, is dependent upon the formation of characteristic membrane ruffles that engulf the bacteria in a macropinocytic-like process . We show here that Cdc42 and Rac GTPases, but not Rho;, are critical for actin polymerisation, whereas Rho; is necessary for the recruitment of ezrin and Src at the site of entry . Remarkably, cells expressing constitutively active Src did not show ezrin recruitment at Shigella entry foci . In these cells, formation of stress fibres induced by LPA stimulation, or microinjection of activated Rho; (V14Rho), was inhibited . Src-mediated tyrosyl-phosphorylation of p190RhoGAP correlated with changes in the ability of p190RhoGAP to interact with Rho;, suggesting that Src regulates Rho; function via p190RhoGAP . We propose that Rho; activation is required for proper organisation of Shigella entry foci and for Src recruitment, and that Src tyrosine kinase activity, in turn, down-regulates the function of Rho; at the site of Shigella entry . The significance of this negative regulatory loop on Rho;-dependent responses is discussed.

FEMS Microbiol Lett, 1999 Dec 15, 181(2), 289 - 95
Cell density dependent acid sensitivity in stationary phase cultures of enterohemorrhagic Escherichia coli O157:H7; Datta AR et al.; Escherichia coli O157:H7, the causative agent of hemorrhagic colitis and hemolytic uremic syndrome, can survive in a highly acidic environment . The acid resistance of this organism, as measured by its ability to survive in low pH, depended on the density of the cells present during the assay . At low cell densities (</=2 x 10(7) ml(-1)), about 100% of the stationary phase cells survived in Luria broth pH 2.5 at 37 degrees C for at least 7 h . The same cultures at high cell densities (2-5 x 10(9) ml(-1)) were about 1000-fold more sensitive under identical conditions . Exponential phase cultures did not exhibit the cell density effect . The increased acid sensitivity at high cell densities was absent in the stationary phase cultures of a rpoS mutant (rpoS::pRR10) of an E . coli O157:H7 strain . Cell density dependent acid sensitivity of the stationary phase cultures was also observed in other enterohemorrhagic E . coli and Shigella strains . The increased acid sensitivity at high cell densities was absent in Gram-positive organisms.

FEMS Microbiol Lett, 1999 Dec 15, 181(2), 267 - 75
Adhesion and invasion of a mutant Shigella flexneri to an eukaryotic cell line in absence of the 220-kb virulence plasmid; Guhathakurta B et al.; A Shigella flexneri strain, cured of the large 220-kb virulence plasmid, expresses adhering and invading ability in confluent monolayers of HeLa cells similar to its parent strain . Invasion by both the parent and the cured strains resulted in alteration of the monomeric actin (G) in the total actin pool of HeLa cells . Other indicators of invasive characteristics of virulent Shigella strains such as production of keratoconjunctivitis in guinea pig eye in vivo, Congo red binding and expression of contact hemolysin however, indicated loss of invasive properties in the plasmid cured strain . Further, pretreatment of bacterial cells with para-bromophenacyl bromide (p-BPB), a specific chemical inhibitor of phospholipase A, adversely affected adhesion to and invasion of HeLa cells in vitro, irrespective of the presence of the 220-kb plasmid indicating the possible involvement of the enzyme phospholipase A in the invasion process . Adherence of both the strains to guinea pig colonic epithelial cells (CECs) in vitro was reduced significantly on pretreatment of bacteria or CECs with p-BPB . Expression of exocellular enzymes viz . protease, elastase, phospholipase A and phospholipase C were not related to the large plasmid.

Microbes Infect, 1999 Dec, 1(14), 1221 - 8
DsbA: a protein-folding catalyst contributing to bacterial virulence; Yu J et al.; DsbA, a periplasmic thiol:disulphide oxidoreductase, catalyses the folding of various factors, among which are virulence determinants or the components of type III secretory machinery . It is also necessary for intracellular survival and cell-to-cell spread of the intracellular pathogen Shigella flexneri.

Nature, 1999 Oct 28, 401(6756), 926 - 9
Actin-based motility of vaccinia virus mimics receptor tyrosine kinase signalling; Frischknecht F et al.; Studies of the actin-based motility of the intracellular pathogens Listeria monocytogenes and Shigella flexneri have provided important insight into the events occurring at the leading edges of motile cells . Like the bacteria Listeria and Shigella, vaccinia virus, a relative of the causative agent of smallpox, uses actin-based motility to spread between cells . In contrast to Listeria or Shigella, the actin-based motility of vaccinia is dependent on an unknown phosphotyrosine protein, but the underlying mechanism remains obscure . Here we show that phosphorylation of tyrosine 112 in the viral protein A36R by Src-family kinases is essential for the actin-based motility of vaccinia . Tyrosine phosphorylation of A36R results in a direct interaction with the adaptor protein Nck and the recruitment of the Ena/VASP family member N-WASP to the site of actin assembly . We also show that Nck and N-WASP are essential for the actin-based motility of vaccinia virus . We suggest that vaccinia virus spreads by mimicking the signalling pathways that are normally involved in actin polymerization at the plasma membrane.

J Leukoc Biol, 1999 Nov, 66(5), 753 - 6
Intracellular niches for extracellular bacteria: lessons from Helicobacter pylori; Allen LA; Helicobacter pylori colonizes the gastric epithelium of humans and plays a causative role in peptic ulcer disease and perhaps gastric cancer . H . pylori proliferates in the mucus layer over the epithelium and is not cleared by the host immune response . Although the mucus layer is the major reservoir of H . pylori in vivo, a growing body of evidence suggests that H . pylori can persist in multiple intracellular locales . Clinical isolates of H . pylori invade epithelial monolayers at least as well as Shigella . The intracellular organisms are cytotoxic, and bacterial microcolonies form on the exposed basement membrane . Both mononuclear phagocytes and neutrophils phagocytose unopsonized H . pylori . However, the internalized organisms are not killed efficiently and our recent data suggest that H . pylori disrupts phagosome maturation . Collectively, the data support the hypothesis that intracellular H . pylori represent a reservoir of organisms that contributes to bacterial persistence, host tissue damage, and treatment failure.

J Biol Chem, 1999 Dec 3, 274(49), 35278 - 82
Activation of Src family kinase yes induced by Shiga toxin binding to globotriaosyl ceramide (Gb3/CD77) in low density, detergent-insoluble microdomains; Katagiri YU et al.; Shiga toxin (Stx) is an enterotoxin produced by Shigella dysenteriae serotype 1 and enterohemorrhagic Escherichia coli, which binds specifically to globotriaosylceramide, Gb3, on the cell surface and causes cell death . We previously demonstrated that Stx induced apoptosis in human renal tubular cell line ACHN cells (Taguchi, T., Uchida, H., Kiyokawa, N., Mori, T., Sato, N., Horie, H., Takeda, T and Fujimoto, J . (1998) Kidney Int . 53, 1681-1688) . To study the early signal transduction after Stx addition, Gb3-enriched microdomains were prepared from ACHN cells by sucrose density gradient centrifugation of Triton X-100 lysate as buoyant, detergent-insoluble microdomains (DIM) . Gb3 was only recovered in DIM and was associated with Src family kinase Yes . Phosphorylation of tyrosine residues of proteins in the DIM fraction increased by 10 min and returned to the resting level by 30 min after the addition of Stx . Since the kinase activity of Yes changed with the same kinetics, Yes was thought to be responsible for the hyperphosphorylation observed in DIM proteins . Unexpectedly, however, all of the Yes kinase activity was obtained in the high density, detergent-soluble fraction . Yes was assumed to be activated and show increased Triton X-100 solubility in the early phase of retrograde endocytosis of Stx-Gb3 complex . Since Yes activation by the Stx addition was suppressed by filipin pretreatment, Gb3-enriched microdomains containing cholesterol were deeply involved in Stx signal transduction.

Eur J Biochem, 1999 Dec, 266(2), 460 - 6
Structural studies of the O-antigen polysaccharide from the enteroinvasive Escherichia coli O164 cross-reacting with Shigella dysenteriae type 3; Linnerborg M et al.; The structure of the O-antigen polysaccharide from Escherichia coli O164 has been determined . Nuclear magnetic resonance spectroscopy together with component and methylation analyses of lipid free polysaccharide were the principal methods used . The sequence of the sugar residues could be determined by NOESY and heteronuclear multiple bond connectivity NMR experiments . It is concluded that the polysaccharide is composed of a pentasaccharide repeating unit with the following structure: {structure: see text} . Matrix assisted laser desorption ionization mass spectrometry (MALDI-MS) was performed on intact lipopolysaccharide and from the resulting molecular mass, the O-antigen part was estimated to contain approximately 24 repeating units . The nature of the previously reported cross-reactivity of this O-antigen to those of Escherichia coli O124 and Shigella dysenteriae type 3 is discussed.

Infect Immun, 1999 Dec, 67(12), 6364 - 8
Role of nitric oxide in the enhancement of pentylenetetrazole-induced seizures caused by Shigella dysenteriae; Balter-Seri J et al.; Convulsions and encephalopathy are frequent complications of childhood shigellosis . We studied the role of nitric oxide (NO) in Shigella-related seizures in an animal model . Pretreatment of mice with Shigella dysenteriae 60R sonicate elevated serum NO levels and enhanced the convulsive response to pentylenetetrazole (PTZ), as indicated by a higher mean convulsion score and a higher number of mice responding with seizures . Treatment of the mice with S-methylisothiourea sulfate (SMT), a potent inhibitor of inducible NO synthase (NOS), prevented the elevation of serum NO levels and concomitantly reduced the enhanced response to PTZ . The mean convulsion scores were 0.7, 0.7, 1.3, and 0.8 for mice treated with saline, saline and SMT, S . dysenteriae 60R sonicate, and S . dysenteriae 60R sonicate with SMT, respectively (P = 0.001 for 60R sonicate versus saline and P = 0.013 for 60R sonicate versus 60R sonicate with SMT) . The corresponding seizure rates were 40, 44, 75, and 47% for saline, saline with SMT, S . dysenteriae 60R sonicate, and S . dysenteriae 60R sonicate with SMT, respectively (P = 0.0004 for 60R sonicate versus saline and P = 0.005 for 60R sonicate versus 60R sonicate with SMT) . In contrast, injection of N-nitro-L-arginine, a selective inhibitor of constitutive NOS, neither abolished the elevation of serum NO nor attenuated the enhancement of seizures . These findings indicate that NO, induced by S . dysenteriae 60R sonicate, is involved in enhancing the susceptibility to seizures caused by S . dysenteriae.

Mikrobiol Z, 1999 Jul-Aug, 61(4), 64 - 71
{The immunobiological properties of the lipopolysaccharides of bacteria in the genus Shigella with contrasting virulence and their chemically modified analogs}; Borisova EV; The antibody formation and expression of delayed type hypersensitivity (DTH) of several antigens under the conditions of their injection together with bacterial lipopolysaccharide (LPS) in mice have been studied . Water-phenol LPS extract from S . sonnei was used . The LPS was treated with the redox system (LPS-R) or not (LPS-PW) . Besides, LPS was obtained by immunoaffinity method from avirulent (LPS-A) or virulent (LPS-V) S . sonnei . It had been found that the mice reacted by more intensive increase antibody formation to LPS or sheep erythrocytes if LPS-R or LPS-V were injected alone or together with erythrocytes . More intensive increase of lymph nodes was shown as well . When specified LPS were injected in mice together with E . coli they induced the immunosuppression of DTH . The LPS-PW and LPS-A caused the most feeble reaction of immune system in mice . It has been suggested that the differences of immune reaction are the reflection of conformation processes in LPS molecules.

Mol Microbiol, 1999 Nov, 34(4), 675 - 89
A system for identifying post-invasion functions of invasion genes: requirements for the Mxi-Spa type III secretion pathway of Shigella flexneri in intercellular dissemination; Schuch R et al.; Invasion and intercellular spread are hallmarks of Shigella pathogenicity . Invasion of the eukaryotic cell cytosol requires a type III secretion system (Mxi-Spa) and its cognate set of secreted Ipa invasins . Once intracellular, the IcsA protein directs a form of actin-based motility that helps to drive intracellular bacterial movement, formation of cellular protrusions and cell-to-cell spread . Work in our laboratory has focused on identifying additional factors required for this intercellular form of dissemination . In this study, we sought to identify novel contributions of the type III secretion pathway to post-invasion-specific processes, distinct from its previously characterized roles in invasion . Studies of post-invasion Ipa and Mxi-Spa functions are complicated by an absolute requirement for these virulence proteins in invasion . To circumvent this problem, we developed a system called TIER (for test of intracellular expression requirements), whereby specific ipa, mxi or spa loci are transiently expressed before infection of tissue culture cell monolayers (thus supporting invasion), but then repressed after invasion in the intracellular environment . Such invasive type III secretion mutants (called TIER mutants) were severely restricted in their ability to spread intercellularly and form plaques in confluent tissue culture cell monolayers . Intercellular spread defects were associated with the repression of most type III pathway components examined, including structural (MxiM and Spa33), secreted effector (IpaB, IpaC and IpaD) and regulatory elements (VirF and VirB) . A kinetic analysis of bacterial growth in L2 cell monolayers showed that each of the TIER mutants was defective with respect to long-term intracellular proliferation and viability . Examination of TIER mutant-infected monolayers by electron microscopy revealed that the type III pathway was required for a late step in intercellular spread - bacterial escape from protrusion-derived, double-membrane-bound vacuoles . The TIER mutants were eventually degraded in a process involving vacuolar acidification . Based on these findings, we propose that Ipa secretion via Mxi-Spa is required in the protrusion vacuole for double-membrane lysis.

J Clin Pathol, 1999 Jun, 52(6), 415 - 8
A survey of infections in United Kingdom laboratories, 1994-1995; Walker D et al.; AIMS: To identify the number and type of infections occurring in United Kingdom clinical laboratories during 1994 and 1995, following similar surveys covering 1970 to 1989 . METHODS: A retrospective questionnaire survey was undertaken of 397 responding UK clinical laboratories covering 1994 and 1995 . A follow up telephone survey was undertaken with each of the laboratories from which a questionnaire had been received indicating a possible or probable laboratory acquired infection during 1994 or 1995 . RESULTS: Questionnaires were sent to 659 laboratories or organisations which were thought to have laboratories, of which 557 responded (response rate of 84.5%) . Of these, only 397 were from organisations with laboratories . Over 55,000 person-years of occupational exposure were covered, and only nine cases identified, giving an infection incidence rate overall of 16.2/100,000 person-years, compared with 82.7 infections/100,000 person-years found in a similar survey covering 1988 and 1989, reported previously . Infections were commonest in females, in relatively young staff, in microbiology laboratory workers, and in scientific/technical employees . Gastrointestinal infections predominated, particularly shigellosis, but few specific aetiological factors relating to working practices were identified . No hepatitis B cases were reported . CONCLUSIONS: The small number of cases identified indicates high standards of infection control, though there is still room for improvement . Periodic studies of this kind are not adequate for comprehensive monitoring of the incidence of laboratory acquired infections . That will require the introduction of a routine, active surveillance programme or prospective survey which has the support and commitment of the laboratories themselves.

Zentralbl Hyg Umweltmed, 1999 Sep, 202(5), 383 - 8
Cell adhesion of Plesiomonas shigelloides; Schubert RH et al.; Plesiomonas shigelloides evidences an inconsistent behaviour in the INT-407 cell adhesion test . Isolates from clinical material evidence regular and pronounced cell adhesion which, however, is low compared with that of clinical Aeromonas isolates . In some of the P . shigelloides strains isolated from clinical material, the cell adhesion is markedly more pronounced than in others . The P . shigelloides strains isolated from the natural habitat from both pond water, mud samples and water insects manifest either no or only slight cell adhesion.

J Cell Biol, 1999 Nov 1, 147(3), 683 - 93
The tripartite type III secreton of Shigella flexneri inserts IpaB and IpaC into host membranes; Blocker A et al.; Bacterial type III secretion systems serve to translocate proteins into eukaryotic cells, requiring a secreton and a translocator for proteins to pass the bacterial and host membranes . We used the contact hemolytic activity of Shigella flexneri to investigate its putative translocator . Hemolysis was caused by formation of a 25-A pore within the red blood cell (RBC) membrane . Of the five proteins secreted by Shigella upon activation of its type III secretion system, only the hydrophobic IpaB and IpaC were tightly associated with RBC membranes isolated after hemolysis . Ipa protein secretion and hemolysis were kinetically coupled processes . However, Ipa protein secretion in the immediate vicinity of RBCs was not sufficient to cause hemolysis in the absence of centrifugation . Centrifugation reduced the distance between bacterial and RBC membranes beyond a critical threshold . Electron microscopy analysis indicated that secretons were constitutively assembled at 37 degrees C before any host contact . They were composed of three parts: (a) an external needle, (b) a neck domain, and (c) a large proximal bulb . Secreton morphology did not change upon activation of secretion . In mutants of some genes encoding the secretion machinery the organelle was absent, whereas ipaB and ipaC mutants displayed normal secretons.

EMBO J, 1999 Nov 1, 18(21), 5853 - 62
Binding of the Shigella protein IpaA to vinculin induces F-actin depolymerization; Bourdet-Sicard R et al.; Shigella flexneri, the causative agent of bacillary dysentery, enters into epithelial cells by a macropinocytic process . IpaA, a Shigella protein secreted upon cell contact, binds to the focal adhesion protein vinculin and is required for efficient bacterial uptake . IpaA was shown here to bind with high affinity to the N-terminal residues 1-265 of vinculin . Using co-sedimentation and solid-phase assays, we demonstrated that binding of IpaA to vinculin strongly increases the association of vinculin with F-actin . We also characterized a depolymerizing activity on actin filaments associated with the vinculin-IpaA complex both in vitro and in microinjected cells . We propose that the conformational change of vinculin induced by IpaA binding allows interaction of the vinculin-IpaA complex with F-actin and subsequent depolymerization of actin filaments.

Eur J Biochem, 1999 Nov, 266(1), 246 - 51
Structural studies utilizing 13C-enrichment of the O-antigen polysaccharide from the enterotoxigenic Escherichia coli O159 cross-reacting with Shigella dysenteriae type 4; Linnerborg M et al.; The structure of the O-antigen polysaccharide from Escherichia coli O159 has been determined using primarily NMR spectroscopy of the 13C-enriched polysaccharide . The sequence of the sugar residues could be determined by heteronuclear multiple bond connectivity NMR experiments . The polysaccharide is composed of a pentasaccharide repeating unit with the following structure: {sequence: see text} Matrix assisted laser desorption ionization mass spectrometry was performed on intact lipopolysaccharide and from the resulting molecular mass the O-antigen part was estimated to contain approximately 23 repeating units . Cross-reactivity of this O-antigen to that of Shigella dysenteriae type 4 was confirmed using enzyme-linked immunoabsorbant assay.

Kansenshogaku Zasshi, 1999 Sep, 73(9), 923 - 9
{The molecular epidemiological analysis of Shigella sonnei isolates from outbreak cases in Yokohama}; Yamada M et al.; We conducted a molecular epidemiological analysis to evaluate the epidemiologic patterns of Shigella sonnei isolates from outbreak cases in Yokohama to clarify the epidemiologic linkages by contact tracing and sources of infection . In the first case (case A), all of the 6 isolates were the colicin 0 type and resistant to both streptomycin (SM) and trimethoprim/sulfamethoxazole (ST) . The 5 isolates have plasmid of 230 kb . By RAPD analysis with 2 kinds of primers specific for Shigella, every 6 isolates showed the same pattern . But the DNA fingerprint analysis by PFGE that was performed according to 2 standardized restriction endonucleases revealed a discriminative pattern . However, the resemblance of all isolates, which was calculated by the UPGMA methods, was 0.90 or higher . In the second case (case B), all of the 14 isolates were the colicin 6 type and sensitive to 16 drugs . The serotype of 13 isolates was phase I . The 11 isolates have plasmids of 230 kb and 3 kb . The resemblance of all isolates, which was calculated by the UPGMA methods, was 0.89 or higher . The analysis with a combination of the plasmid, RAPD analysis and PFGE profiles may be effective in investigating detailed epidemiological features of isolates.

Infect Immun, 1999 Nov, 67(11), 6056 - 66
Increased interleukin-1 (IL-1) and imbalance between IL-1 and IL-1 receptor antagonist during acute inflammation in experimental Shigellosis; Arondel J et al.; Infection by the enteric bacterial pathogen Shigella results in intense mucosal inflammation and destruction of the colonic and rectal epithelium in infected humans . Initial bacterial translocation occurs through the follicle-associated epithelium . Previous experiments suggest that interleukin-1 (IL-1) is crucial to trigger inflammation, particularly in the follicular zones . During the first 4 hours of infection in a rabbit ligated-loop model of intestinal invasion, there are two salient characteristics: (i) a high concentration of IL-1alpha and IL-1beta, both in infected Peyer's patch tissue and in the corresponding efferent mesenteric blood, and (ii) a very low level of expression of IL-1 receptor antagonist (IL-1ra) . These may reflect a combination of regulation of expression and secretion of IL-1alpha, IL-1beta, and IL-1ra by both resident and recruited phagocytes and the induction of mononuclear phagocyte apoptosis by Shigella . This low IL-1ra/IL-1 ratio likely accounts for the rapid, uncontrolled inflammation characteristic of shigellosis.

Infect Immun, 1999 Nov, 67(11), 5841 - 7
Native and mutant forms of cholera toxin and heat-labile enterotoxin effectively enhance protective efficacy of live attenuated and heat-killed Shigella vaccines; Hartman AB et al.; Both native and mutant forms of cholera toxin (CT) and heat-labile enterotoxin (LT) are effective adjuvants for antigens and killed whole-cell preparations . To determine whether these toxin molecules could also boost the immunogenicity and efficacy of live attenuated vaccines directed against shigellosis, the guinea pig keratoconjunctivitis model was used to evaluate the adjuvant effect of these toxin molecules on EcSf2a-3, a DeltavirG DeltaaroD Escherichia coli-Shigella flexneri 2a hybrid vaccine strain that was previously found to be less protective than its parent strain in the guinea pig model . Experiments using native and mutant toxin molecules showed that both CT and LT and mutant derivatives were effective as an adjuvant for EcSf2a-3 and that the mutant toxin molecules, which were developed to retain adjuvanticity without the toxicity associated with the native molecules, were as effective as the native toxin molecules as adjuvants . Protective efficacy was enhanced for both the oral and intranasal routes of immunization . Serum antibody response to the S . flexneri 2a O antigen, the primary antigen for protective immunity, was not dependent on the addition of an adjuvant . However, enumeration of the O-antigen-specific immunoglobulin G (IgG) and IgA antibody-secreting cells in the spleen and draining lymph nodes following intranasal immunization suggested that enhancement of the local immune response by the toxin molecules may contribute to the observed increase in protective efficacy . The efficacy of heat-killed S . flexneri 2a was enhanced only by mutant LT molecules . These results suggest that the best candidates for enhancing the efficacy of both live attenuated and heat-killed Shigella vaccines with minimal reactogenicity are the mutant toxin molecules.

Clin Infect Dis, 1999 Nov, 29(5), 1303 - 6
Dr . Kiyoshi Shiga: discoverer of the dysentery bacillus; Trofa AF et al.; The clinical manifestations of dysentery have been described for centuries, and the prototypic bacterial agent, Shigella dysenteriae, was identified 100 years ago . In the English language there has been remarkably little written about Dr . Kiyoshi Shiga, discoverer of the dysentery bacillus . We submit a brief biography of Dr . Shiga and the circumstances leading to his discovery, which proved the bacterial etiology of nonamebic dysentery.

Nature, 1999 Oct 7, 401(6753), 613 - 6
Reconstitution of actin-based motility of Listeria and Shigella using pure proteins; Loisel TP et al.; Actin polymerization is essential for cell locomotion and is thought to generate the force responsible for cellular protrusions . The Arp2/3 complex is required to stimulate actin assembly at the leading edge in response to signalling . The bacteria Listeria and Shigella bypass the signalling pathway and harness the Arp2/3 complex to induce actin assembly and to propel themselves in living cells . However, the Arp2/3 complex alone is insufficient to promote movement . Here we have used pure components of the actin cytoskeleton to reconstitute sustained movement in Listeria and Shigella in vitro . Actin-based propulsion is driven by the free energy released by ATP hydrolysis linked to actin polymerization, and does not require myosin . In addition to actin and activated Arp2/3 complex, actin depolymerizing factor (ADF, or cofilin) and capping protein are also required for motility as they maintain a high steady-state level of G-actin, which controls the rate of unidirectional growth of actin filaments at the surface of the bacterium . The movement is more effective when profilin, alpha-actinin and VASP (for Listeria) are also included . These results have implications for our understanding of the mechanism of actin-based motility in cells.

J Clin Microbiol, 1999 Nov, 37(11), 3608 - 11
Prevalence of Shigella enterotoxins 1 and 2 among Shigella strains isolated from patients with traveler's diarrhea; Vargas M et al.; Shigella spp . are known primarily as a cause of bacillary dysentery . However, in an initial phase, numerous patients exhibit watery diarrhea that may or may not be followed by dysentery . New virulence factors associated with the species of Shigella have recently been described . These are enterotoxins 1 and 2 of Shigella (ShET-1 and ShET-2, respectively) . The aim of the present study was to determine the prevalence of ShET-1 and ShET-2 in species of Shigella isolated from patients with traveler's diarrhea . During the period from 1993 to 1998, stool samples from 500 travelers with diarrhea were cultured for the isolation of Shigella spp . and other enteropathogens . The detection of ShET-1 and ShET-2 was performed by a PCR technique with specific primers . Among a total of 51 strains of Shigella isolated during this period (22 S . flexneri, 26 S . sonnei, and 3 S . dysenteriae strains), at least one enterotoxin was detected in 31 (60.78%) strains; 2 (9.09%; both of which were S . flexneri strains) produced only ShET-1, while 21 (41.17%; 3 S . flexneri, 15 S . sonnei, and 3 S . dysenteriae strains) produced ShET-2 . Furthermore, 8 (15.69%) of 22 S . flexneri strains presented both enterotoxins . Our results show that the prevalence of ShET-2 was high in all the Shigella species studied and confirm that ShET-1 is detected only in S . flexneri.

Indian J Exp Biol, 1999 Jul, 37(7), 681 - 90
Purification and characterization of Klebsiella pneumoniae cytotoxins; Singh BR et al.; Isolation, purification and characterization of 3 new cytotoxins of a K . pneumoniae strain isolated from ready to eat pork sausage are reported . Purification process involved extraction of cytotoxins with polymyxin B sulphate, salt precipitation, gel filtration and anion exchange chromatography . Klebsiella cytotoxin (KCT) I, a glycoprotein of about 65 kDa was verocytotoxic, enterotoxic and dermonerotic . KCT II was erythemogenic, verocytotoxic and enterotoxic protein of co 55 kDa, while KCT III was about double in MW (110 kDa) hadverocytotoxicity but neither enterotoxicity nor dermatotoxicity . KCT I and II caused granulation, conglomeration, shrinkage, detachment and lysis of MDBK and Vero cells, while KCT III induced enlargement, vacuolation, granulation, multinucleolation and syncytia formation in exposed cells . All the three cytotoxins induced specific neutralizing antibodies and cytotoxins were detectable in nanogram quantities with enzyme-linked immunosorbant assay using homologous antibodies . None of the anticytotoxin cross-reacted with either heterologous Klebsiella cytotoxins or with verocytotoxic preparations of Shigella dysenteriae.

Indian J Exp Biol, 1999 Jul, 37(7), 671 - 5
Potentiality of tricyclic compound thioridazine as an effective antibacterial and antiplasmid agent; Radhakrishnan V et al.; Thioridazine (Th), which is therapeutically used in psychiatric patients, was found to possess conspicuous antimicrobial activity when tested against 316 strains belonging to a number of Gram positive and Gram negative bacteria . Although Staphylococcus aureus, Vibrio chloerae and V . parahaemolyticus were found to be most sensitive, Th was highly bactericidal against S . aureus and bacteriostatic for vibrios and other Gram negative organisms . In the study of antiplasmid/curing effect of Th on twelve multiply antibiotic and Th resistant bacteria, it was observed that elimination of R plasmids was facilitated by choice of optimal concentration of Th . Significant elimination of single and combined antibiotic resistance occurred in E . coli and Shigella flexneri and not in S . aureus.

Bull World Health Organ, 1999, 77(8), 651 - 66
Global burden of Shigella infections: implications for vaccine development and implementation of control strategies; Kotloff KL et al.; Few studies provide data on the global morbidity and mortality caused by infection with Shigella spp.; such estimates are needed, however, to plan strategies of prevention and treatment . Here we report the results of a review of the literature published between 1966 and 1997 on Shigella infection . The data obtained permit calculation of the number of cases of Shigella infection and the associated mortality occurring worldwide each year, by age, and (as a proxy for disease severity) by clinical category, i.e . mild cases remaining at home, moderate cases requiring outpatient care, and severe cases demanding hospitalization . A sensitivity analysis was performed to estimate the high and low range of morbid and fatal cases in each category . Finally, the frequency distribution of Shigella infection, by serogroup and serotype and by region of the world, was determined . The annual number of Shigella episodes throughout the world was estimated to be 164.7 million, of which 163.2 million were in developing countries (with 1.1 million deaths) and 1.5 million in industrialized countries . A total of 69% of all episodes and 61% of all deaths attributable to shigellosis involved children under 5 years of age . The median percentages of isolates of S . flexneri, S . sonnei, S . boydii, and S . dysenteriae were, respectively, 60%, 15%, 6%, and 6% (30% of S . dysenteriae cases were type 1) in developing countries; and 16%, 77%, 2%, and 1% in industrialized countries . In developing countries, the predominant serotype of S . flexneri is 2a, followed by 1b, 3a, 4a, and 6 . In industrialized countries, most isolates are S . flexneri 2a or other unspecified type 2 strains . Shigellosis, which continues to have an important global impact, cannot be adequately controlled with the existing prevention and treatment measures . Innovative strategies, including development of vaccines against the most common serotypes, could provide substantial benefitsPIP: This article presents a review of the literature published between 1966 and 1997 on Shigella infection . The purpose of the review is to provide data on the global morbidity and mortality caused by the infection and to plan strategies of prevention and treatment . The data obtained from this literature were used to calculate the number of Shigella infection cases and the associated mortality occurring worldwide each year, by age and by clinical category . The burden of Shigella infection was also estimated by serogroup and serotype . A sensitivity analysis was performed to estimate the high and the low range of morbid and fatal cases in each category (mild cases remaining at home, moderate cases requiring outpatient care and severe cases demanding hospitalization) . The result of the calculations and analysis revealed that the annual number of Shigella infections throughout the world was estimated to be 164.7 million . 163.2 million occurred in developing countries, with 1.1 million deaths, and 1.5 million occurred in industrialized countries . More than half of the episodes and death affects children under 5 years of age . In comparing developing countries against industrialized countries, the median of isolates are S . flexneri (60% vs . 16%), S . sonnei (15% vs . 77%), S . dysenteriae (6% vs . 1%), and S . boydii (6% vs . 2%) . The predominant serotype of S . flexneri in developing countries is 2a, followed by 1b, 3a, 4a, and 6, while in industrialized countries most isolates are S . flexneri 2a and unspecified type 2 strains .

Microb Pathog, 1999 Oct, 27(4), 223 - 30
Preincubation of recombinant Ipa proteins of Shigella sonnei promotes entry of non-invasive Escherichia coli into HeLa cells; Terajima J et al.; Invasion plasmid antigens of Shigella sonnei, IpaB, C, D, were expressed as fusion proteins either with maltose-binding protein (MBP) or Strept-tag sequence . Affinity-purified IpaB and IpaD were separated from MBP by digestion with Factor Xa . Recombinant IpaC having Strept-tag sequence at its C-terminal was also purified by avidin affinity column chromatography . These recombinant proteins showed the ability to cause non-invasive Escherichia coli K-12 to internalize HeLa cell only when all of the proteins were preincubated with the bacterial prior to the inoculation of the mixture into HeLa cell culture . Electron microscopy also showed internalized bacteria within HeLa cells suggesting that functional complex of invasins (IpaB, C and D) were formed in vitro .

Infect Immun, 1999 Oct, 67(10), 5526 - 9
Treatment with succinic anhydride improves the immunogenicity of Shigella flexneri type 2a O-specific polysaccharide-protein conjugates in mice; Pavliakova D et al.; Seroepidemiological data and a clinical trial with a Shigella sonnei O-specific polysaccharide (O-SP)-Pseudomonas aeruginosa recombinant exoprotein A (rEPA) conjugate provide evidence that a critical level of immunoglobulin G (IgG) lipopolysaccharide (LPS) antibodies in serum confers protection against shigellosis . We evaluated the immunogenicity of conjugates whose carrier proteins and O-SPs were treated with succinic anhydride (SA), which reacts with amino groups at neutral pH to form amide-linked carboxyls (succinylation) . Conjugates were synthesized with either of two genetically inactivated medically useful toxins, the diphtheria protein CRM9 or rEPA, bound to the O-SP of Shigella flexneri type 2a . Conjugates composed of the succinylated protein, succinylated O-SP, or both succinylated components were administered to mice by a clinically relevant scheme, and their levels of serum IgG anti-LPS and anti-proteins were assayed 7 days after the second and third injections . CRM9 served as a more immunogenic carrier than rEPA . Conjugates composed of succinylated components were more immunogenic than the conjugates composed of the native components . SA treatment of both the carrier protein and the O-SP did not confer an advantage over the succinylated protein alone . Conjugates prepared with native proteins, in general, elicited slightly higher levels of IgG protein antibodies than conjugates composed of the SA-treated proteins.

J Cell Biol, 1999 Sep 20, 146(6), 1319 - 32
Activation of the CDC42 effector N-WASP by the Shigella flexneri IcsA protein promotes actin nucleation by Arp2/3 complex and bacterial actin-based motility; Egile C et al.; To propel itself in infected cells, the pathogen Shigella flexneri subverts the Cdc42-controlled machinery responsible for actin assembly during filopodia formation . Using a combination of bacterial motility assays in platelet extracts with Escherichia coli expressing the Shigella IcsA protein and in vitro analysis of reconstituted systems from purified proteins, we show here that the bacterial protein IcsA binds N-WASP and activates it in a Cdc42-like fashion . Dramatic stimulation of actin assembly is linked to the formation of a ternary IcsA-N-WASP-Arp2/3 complex, which nucleates actin polymerization . The Arp2/3 complex is essential in initiation of actin assembly and Shigella movement, as previously observed for Listeria monocytogenes . Activation of N-WASP by IcsA unmasks two domains acting together in insertional actin polymerization . The isolated COOH-terminal domain of N-WASP containing a verprolin-homology region, a cofilin-homology sequence, and an acidic terminal segment (VCA) interacts with G-actin in a unique profilin-like functional fashion . Hence, when N-WASP is activated, its COOH-terminal domain feeds barbed end growth of filaments and lowers the critical concentration at the bacterial surface . On the other hand, the NH(2)-terminal domain of N-WASP interacts with F-actin, mediating the attachment of the actin tail to the bacterium surface . VASP is not involved in Shigella movement, and the function of profilin does not require its binding to proline-rich regions.

J Clin Microbiol, 1999 Oct, 37(10), 3113 - 7
Typing and characterization of mechanisms of resistance of Shigella spp . isolated from feces of children under 5 years of age from Ifakara, Tanzania; Navia MM et al.; Eighty-six strains of Shigella spp . were isolated during the dry season from stool samples of children under 5 years of age in Ifakara, Tanzania . The epidemiological relationship as well as the antimicrobial susceptibility and mechanisms of resistance to ampicillin, chloramphenicol, and co-trimoxazole were investigated . Four different epidemiological tools, pulsed-field gel electrophoresis (PFGE), repetitive extragenic palindromic (REP)-PCR, plasmid analysis, and antibiogram, were compared for typing Shigella strains . Seventy-eight (90%) strains were Shigella flexneri and were distributed into four groups, by either PFGE or REP-PCR, with 51, 17, 7, and 3 strains . The four strains of Shigella dysenteriae belonged to the same group, and the four strains of Shigella sonnei were distributed in two groups with three and one strain each . Plasmid analysis showed a high level of heterogeneity among strains belonging to the same PFGE group, while the antibiogram was less discriminative . REP-PCR provided an alternative, rapid, powerful genotyping method for Shigella spp . Overall, antimicrobial susceptibility testing showed a high level of resistance to ampicillin (81.8%), chloramphenicol (72.7%), tetracycline (96.9%), and co-trimoxazole (87.9%) . Ampicillin resistance was related to an integron-borne OXA-1-type beta-lactamase in 85.1% of the cases and to a TEM-1-type beta-lactamase in the remaining 14.8% . Resistance to co-trimoxazole was due to the presence of a dhfr Ia gene in all groups except one of S . flexneri, where a dhfr VII gene was found within an integron . Chloramphenicol resistance was associated in every case with positive chloramphenicol acetyltransferase activity . All strains were susceptible to nalidixic acid, ciprofloxacin, ceftazidime, cefotaxime, and cefoxitin . Therefore, these antimicrobial agents may be good alternatives for the treatment of diarrhea caused by Shigella in Tanzania.

Rev Esp Salud Publica, 1999 May-Jun, 73(3), 393 - 401
{Outbreak of shigellosis in a lower-class district}; Chover Lara JL et al.; BACKGROUND: The outbreaks of Shigella sonnei in our environment frequently involve day care centers and elementary schools . An outbreak of shigellosis in a lower-class district is reported . The purpose of this study is that of pinpointing the center of infection, the means of contagion, the characteristics of those infected and of assessing the suitability of the measures taken . METHODS: For monitoring the outbreak over time, a combined observational timeline study was conducted within a territory the bounds of which were marked by means of the conventional epidemiological monitoring variables (time, place and individual) . The infectivity of the center of infection (contagion rate) is analyzed by age, gender and school; rate ratio and percentage attributable thereto . RESULTS: On a time-related basis, the outbreak in question started on week 46/97 and ended on week 8/98 . This outbreak involved 218 individuals (110 males and 108 females) totaling 5.46% of the district . The highest frequency was found among the 0-4 age group (43.6%), 29.4% in day care (70.32% contagion rate) with a relative risk of 3.9 (95% CI: 2.57-5.93) and 74.36% attributable percentage . The rate ratio between day care and the schools in the district in question is 5.62 (95% CI: 4.33-7.31) . Stool cultures were taken and analyzed in 84 cases (38.5%), Shigella sonnei being detected in 38 cases (17.4%) . Antibacterial treatment (amoxicillin-clavulan) was set out and individual and group health and safety measures were employed . CONCLUSIONS: The long communicability period and the small number of viable microorganisms necessary for causing this disease fostered its being passed on from one person to another at school and in the home . The measures employed effectively confined the contagion of the infectious agent at the schools.

Comp Immunol Microbiol Infect Dis, 1999 Oct, 22(4), 221 - 30
Verocytotoxins (Shiga-like toxins) produced by Escherichia coli: a minireview of their classification, clinical presentations and management of a heterogeneous family of cytotoxins; Agbodaze D; Bacterial virulence usually requires the interaction of multiple factors in order to cause disease . The enterotoxins produced by certain strains of bacteria are proteins which vary in their mode of action, but do fall into two general groups; the cytotoxic and the cytotonic enterotoxins . While cytotoxic enterotoxins typically kill eucaryotic cells (eg . by inhibiting protein synthesis), cytotonic enterotoxins derange cell metabolism in specific ways (eg . by elevating cyclic nucleotide levels) . Some strains of Escherichia coli produce protein toxins that are biologically, structurally and antigenically related to a cytotoxin (Shiga toxin) (ShT) produced by Shigella dysenteriae type 1 . Although this group of related, but not necessarily identical toxins have been referred to as Vero cell toxins or Verocytotoxins (VTs), the term Shiga-like toxins (SLTs) has been widely accepted . ShT and SLTs have been implicated as a cause of diarrhoea as well as haemorrhagic colitis (HC) and haemolytic uremic syndrome (HUS) in humans, whilst SLTs have been implicated as causal agents of oedema disease and HC in weaner pigs and calves, respectively . While S . dysenteriae is an invasive organism, the SLT-producing strains of E . coli have not been reported to be invasive, but cause diarrhoea that may contain blood and mucus . Thus, SLTs can be considered an important "new" type of enterotoxins whose role in the pathogenesis of diarrhoea, HC and HUS is beginning to emerge, not only in certain geographical settings, but worldwide . This mini review focuses on this family of SLTs, because of recent advances which have been made towards their detection, nomenclature, pathogenesis and possible management of their clinical presentations.

Vaccine, 1999 Aug 6, 17(23-24), 3109 - 15
Quantitative analysis of IgG class and subclass and IgA serum response to Shigella sonnei and Shigella flexneri 2a polysaccharides following vaccination with Shigella conjugate vaccines; Robin G et al.; It has been recently reported that a conjugate vaccine composed of the O-specific polysaccharide of S . sonnei bound to Pseudomonas aeruginosa recombinant exoprotein A (rEPA) conferred 74% protection against S . sonnei shigellosis . In the present study affinity purified Shigella antibodies were used as standards to quantify and characterize the serum antibody response to vaccination with Shigella sonnei or Shigella flexneri 2a polysaccharide conjugated to rEPA . The geometric mean concentrations of antibodies at the pre-vaccination stage were 3.8 microg/ml for IgG anti-S . sonnei LPS and 11.26 microg/ml for IgG anti-S . flexneri 2a LPS . Vaccination with S . sonnei-rEPA and S . flexneri 2a-rEPA induced the production of specific IgG antibodies to levels of 115.8 microg/ml and 126.5 microg/ml, respectively . The levels of specific antibodies above the pre-vaccination values persisted for at least 2 years . The IgG response to S . flexneri 2a-rEPA conjugate was almost entirely represented by the IgG2 subclass . The concentration of IgG1 anti-S . sonnei LPS was significantly higher than that of IgG2 14 days after vaccination with the homologous conjugate, but decreased to similar levels to those of IgG2 6, 12 and 24 months after immunization . Since the only difference between the S . sonnei and S . flexneri 2a conjugates lies in the different polysaccharides of the two Shigella serogroups (the protein rEPA, is identical in both cases), it follows that the different pattern of IgG subclass response is a result of the different structures of the two O-polysaccharides of S . sonnei and S . flexneri 2a.

FEMS Immunol Med Microbiol, 1999 Aug 15, 25(3), 283 - 8
Immune response to rotavirus VP4 expressed in an attenuated strain of Shigella flexneri; Loy AL et al.; An attenuated strain of Shigella flexneri was utilised to express viral protein (VP) 4 of rotavirus and the immunogenicity of the recombinant constructs was studied in BALB/c mice . VP4 was expressed as a fusion with maltose binding protein (MBP) in both the cytoplasm and periplasm, with a much higher level of expression occurring in the former . While all constructs induced a Shigella-specific response in mice, only the construct expressing MBP-VP4 in the cytoplasm of Shigella stimulated an immune response specific to rotavirus . This study demonstrates that Shigella can be used to deliver rotavirus antigens and induces an immune response directed towards both rotavirus and Shigella.

Anal Chem, 1999 Aug 1, 71(15), 3226 - 30
Identification of bacterial proteins observed in MALDI TOF mass spectra from whole cells; Holland RD et al.; Characteristic ions in the MALDI TOF mass spectra from bacterial cells have been associated with four known proteins . The proteins, observed both from cells and in filtered cellular suspensions, were isolated by HPLC and identified on the basis of their mass spectra and their partial amino acid sequence, determined using the Edman method (10-15 residues) . The acid resistance proteins HdeA and HdeB give rise to ions near m/z 9735 and 9060 in MALDI TOF mass spectra from cells and from extracts of both Escherichia coli 1090 and Shigella flexneri PHS-1059 . However, the proteins associated with proteolytic cleavage by the peptidase Lep, rather than the precursor proteins, were observed, both using cells and from cellular extracts . A cold-shock protein, CspA, was associated with the ion near m/z 7643 from Pseudomonas aeruginosa . Similarly, a cold-acclimation protein, CapB, was identified as the source of the ion near m/z 7684 in P . putida . This last protein was homologous with a known CapB from P . fragi . While these experiments involved the detection of known or homologous proteins from typical bacteria, this same approach could also be applied to the detection of unique proteins or biomarker proteins associated with other bacteria of public health significance.

J Med Microbiol, 1999 Aug, 48(8), 781 - 4
Evaluation of pulsed-field gel electrophoresis for typing of Shigella dysenteriae type 1; Talukder KA et al.; Pulsed-field gel electrophoresis (PFGE) has been used successfully to discriminate between strains of many different bacterial species . In this study, digestion of bacterial DNA with the restriction endonuclease NotI and PFGE were evaluated for the typing of isolates of Shigella dysenteriae type 1, an important cause of epidemic dysentery . There were 27 isolates from four outbreaks of dysentery, and 44 isolates from endemic dysentery cases and a laboratory culture collection . The epidemic isolates yielded two types each with two subtypes, whereas the endemic isolates and culture collection yielded eight types with numerous subtypes . These findings suggest that S . dysenteriae 1 can be typed by PFGE.

Trop Med Int Health, 1999 Jun, 4(6), 428 - 32
Risk factors for death in hospitalized dysentery patients in Rwanda; Legros D et al.; To evaluate the management of severe dysentery cases in in-patient facilities during an epidemic of Shigella dysenteriae type 1 (Sd1), and to identify the factors associated with the risk of death, we conducted a prospective cohort study in 10 Rwandese hospitals between September and December 1994 . Data were obtained from 849 cases admitted to hospitals with diarrhoea and visible blood in stools . The proportion of patients with persistent bloody diarrhoea was 51.0% at treatment day 3 and 27.9% at treatment day 5 . At discharge, 79.9% had improved or were cured . The case fatality ratio was 13.2%, higher for patients treated with nalidixic acid than for those treated with ciprofloxacin (12.2% vs . 2.2%, RR = 5.80, 95% CI = 0.83-40.72) . In a logistic regression model three risk factors were significantly associated with an increased risk of death during hospitalization: severe dehydration on admission (adjusted OR = 2.79, 95% CI = 1.46-5.33), age over 50 (adjusted OR vs . 5-49 age group = 3.22, 95% CI = 1.70-6.11) and prescription of nalidixic acid (adjusted OR vs . ciprofloxacin = 8.66, 95% CI = 1.08-69.67) . Those results were consistent with reported high levels of resistance of Sd1 to the commonest antibiotics, including nalidixic acid . Patients belonging to groups with a higher risk of dying should be given special medical attention and supportive care . In areas of high resistance to nalidixic acid, severe cases of dysentery should be treated with fluoroquinolones in order to reduce the mortality associated with these epidemics.

J Lab Clin Med, 1999 Aug, 134(2), 141 - 6
Novel self-sampling culture method to monitor excretion of live, oral Shigella flexneri 2a vaccine SC602 during a community-based phase 1 trial; Teska JD et al.; A culture technique for assessing the excretion of live enteric vaccines was developed and verified during an outpatient safety trial of the Shigella flexneri 2a SC602 vaccine . Preliminary studies showed that SC602 could be recovered on Hektoen enteric (HE) agar plates that had been inoculated with seeded stools in one quadrant, held for up to 48 hours, streaked for isolation, and incubated for 24 +/- 6 hours . Recovery results on HE plates held at 4 degrees C and 25 degrees C were comparable; however, 4 degrees C better inhibited overgrowth before streaking . To prepare for a community-based vaccine trial, volunteers were trained to self-sample fresh stool and to swab-inoculate a single quadrant of HE agar . The trial began with 36 volunteers ingesting 2.5 x 10(4) CFU of SC602 in bicarbonate buffer . During the study, volunteers inoculated HE plates with fresh stool, stored the plates at 4 degrees C, and delivered them to the laboratory within 48 hours . A microbiologist then streaked the HE for isolation, incubated the plates at 35 degrees C +/- 2 degrees C for 24 +/- 6 hours, and identified presumptive S . flexneri colonies by slide agglutination with poly-group B antiserum . The attenuating genetic signature of SC602 was confirmed on selected isolates with the polymerase chain reaction with two specific DNA primer sets . Vaccine was detected from 20% of volunteers on day 1, increasing to 86% by day 4, and all but one vaccinee had excreted SC602 at least once by day 7 . The latest initial SC602 detection occurred on day 7, the longest excretion occurred in one vaccinee on day 33, and excretion throughout the trial was intermittent . The trial was terminated by ciprofloxacin treatment on day 35 . Volunteer compliance with self-sampling and HE plating was excellent because of the convenience of the method, and the advantage of immediate "bedside" plating was evident in the high recovery rate of excreted vaccine . This method can be applied in other trials of live enteric vaccines that require accurate sampling of excreted organisms.

Microbiology, 1999 Jul, 145 ( Pt 7), 1649 - 59
The Shigella flexneri bacteriophage Sf6 tailspike protein (TSP)/endorhamnosidase is related to the bacteriophage P22 TSP and has a motif common to exo- and endoglycanases, and C-5 epimerases; Chua JE et al.; The temperate bacteriophage Sf6 infects Shigella flexneri strains of serotype X or Y, converting them into serotypes 3a or 3b, respectively . The tailspike protein (TSP) of Sf6 possesses endo-1,3-alpha-L-rhamnosidase (endorhamnosidase) activity which results in cleavage of the lipopolysaccharide O-antigen receptor during the adsorption of the phage to the cell surface . When used in Southern hybridization, a P22 gene 9 (encoding P22 TSP) DNA probe hybridized with restriction fragment Pstl-7 of Sf6 . DNA sequencing and analysis of Pstl-7 and the adjacent Pstl-8 fragment revealed an open reading frame (ORF1) of 1872 bp (624 amino acids) bearing amino acid sequence homology to the bacteriophage P22 TSP N-terminal head-binding domain . High conservation of key residues was suggestive of similar secondary and tertiary N-terminal protein structure and a similar function of the Sf6 TSP in this region . In addition, an amino acid sequence motif (DFGX3DGX6AX3A) was identified between residues 164 and 184 which was also found to exist in various prokaryotic and eukaryotic exo-/endoglycanases, C-5 epimerases and bacteriophage proteins . Expression of ORF1 from a T7 promoter produced a 67 kDa protein (detected by L-{35S}methionine labelling and SDS-PAGE) . Assay of heat-treated cytoplasmic extracts containing the ORF1-encoded protein by incubation with whole Sh . flexneri Y cells demonstrated that O-antigen hydrolysis activity was present; ORF1 therefore encodes Sf6 TSP . Sf6 TSP exhibited specific and preferential activity for long-chain Sh . flexneri serotype X or Y O-antigen, cleavage of which resulted in the release of oligosaccharide fragments, consistent with octasaccharides in size, as detected by fluorophore-assisted carbohydrate electrophoresis (FACE).

J Appl Microbiol, 1999 Jun, 86(6), 971 - 8
Detection of Shigella spp . in food with a nested PCR method-sensitivity and performance compared with a conventional culture method; Lindqvist R; A nested PCR method was developed and its performance evaluated by detection of Shigella flexneri in food . The nested PCR amplifies sequences within an invasion-associated locus (ial) on the invasion plasmid specific for Shigella and enteroinvasive Eschrichia coli (EIEC) . The nested PCR detected Sh . flexneri in lettuce inoculated with 2, 20 and 200 cfu g-1 after 1, 7 and 18 d of storage, respectively . In comparison, a culture method (NMKL no . 151) detected 10(5) cfu g-1 after 1 but not after 7 d of storage . The presence of inhibitors in blue cheese and shrimps reduced the sensitivity of the PCR assay . To overcome this inhibition, a sample preparation step based on buoyant density centrifugation was developed . This treatment resulted in a successful recovery of Sh . flexneri in lettuce, milk, shrimp and blue cheese inoculated with 10 cfu g-1 . The proposed method, which includes a combination of enrichment, buoyant density centrifugation and a nested PCR, can be completed in less than two working days.

Southeast Asian J Trop Med Public Health, 1998 Sep, 29(3), 555 - 9
Severe shigellosis in childhood; Sirivichayakul C et al.; A prospective study was performed on 20 bacteriologically proven pediatric cases of severe shigellosis admitted to the Department of Pediatrics, Chulalongkorn Hospital during March 1989 to March 1990 . Fourteen patients were male and six were female . Shigella B was found in 85% and Shigella D in 15% of cases . The major indications for admission were convulsions and dehydration . Fifteen per cent of cases had underlying malignancies and 42.1% had malnutrition . Most patients had a peak of fever between 39.5 and 40.5 degrees C, serum sodium between 128-144 mEq/l . Mild acidosis was detected in 45% and moderate acidosis in 30% of cases . There were no statistical differences in peak of fever and serum sodium between patients who had convulsion and who did not . Shigellemia was found in one case who also had underlying neuroblastoma . One patient died due to necrotizing enterocolitis, septic shock and renal failure . Most of the organisms found resisted to ampicillin and trimethoprim-sulfamethoxazole (TMP-SMX) . However, TMP-SMX was prescribed in most immunocompetent patients and they recovered well . All of three patients with underlying malignancy responded well to ceftriaxone.

Antimicrob Agents Chemother, 1999 Aug, 43(8), 2081 - 3
Molecular characterization of the SHV-11 beta-lactamase of Shigella dysenteriae; Ahamed J et al.; A beta-lactamase with an M(r) of 29,000 and a pI of 7.6 was partially purified from a clinical isolate of Shigella dysenteriae . The bla gene encoded the SHV-11 enzyme carrying the substitution Leu-->Gln at position 35 and was linked to a strong promoter . This variant, unlike the prototype SHV-1 enzyme, hydrolyzed oxacillin, cloxacillin, and oxyiminocephalosporins such as cefotaxime.

J Bacteriol, 1999 Aug, 181(15), 4711 - 8
Serotype 1a O-antigen modification: molecular characterization of the genes involved and their novel organization in the Shigella flexneri chromosome; Adhikari P et al.; The factors responsible for serotype 1a O-antigen modification in Shigella flexneri were localized to a 5.8-kb chromosomal HindIII fragment of serotype 1a strain Y53 . The entire 5.8-kb fragment and regions up- and downstream of it (10.6-kb total) were sequenced . A putative three-gene operon, which showed homology with other serotype conversion genes, was identified and shown to confer serotype 1a O-antigen modification . The serotype conversion genes were flanked on either side by phage DNA . Multiple insertion sequence (IS) elements were located within and upstream of the phage DNA in a composite transposon-like structure . Host DNA homologous to the dsdC and the thrW proA genes was located upstream of the IS elements and downstream of the phage DNA, respectively . The sequence analysis indicates that the organization of the 10.6-kb region of the Y53 chromosome is unique and suggests that the serotype conversion genes were originally brought into the host by a bacteriophage . Several features of this region are also characteristic of pathogenicity islands.

Infect Immun, 1999 Aug, 67(8), 4201 - 7
Dynamics of actin-based movement by Rickettsia rickettsii in vero cells; Heinzen RA et al.; Actin-based motility (ABM) is a virulence mechanism exploited by invasive bacterial pathogens in the genera Listeria, Shigella, and Rickettsia . Due to experimental constraints imposed by the lack of genetic tools and their obligate intracellular nature, little is known about rickettsial ABM relative to Listeria and Shigella ABM systems . In this study, we directly compared the dynamics and behavior of ABM of Rickettsia rickettsii and Listeria monocytogenes . A time-lapse video of moving intracellular bacteria was obtained by laser-scanning confocal microscopy of infected Vero cells synthesizing beta-actin coupled to green fluorescent protein (GFP) . Analysis of time-lapse images demonstrated that R . rickettsii organisms move through the cell cytoplasm at an average rate of 4.8 +/- 0.6 micrometer/min (mean +/- standard deviation) . This speed was 2.5 times slower than that of L . monocytogenes, which moved at an average rate of 12.0 +/- 3.1 micrometers/min . Although rickettsiae moved more slowly, the actin filaments comprising the actin comet tail were significantly more stable, with an average half-life approximately three times that of L . monocytogenes (100.6 +/- 19.2 s versus 33.0 +/- 7.6 s, respectively) . The actin tail associated with intracytoplasmic rickettsiae remained stationary in the cytoplasm as the organism moved forward . In contrast, actin tails of rickettsiae trapped within the nucleus displayed dramatic movements . The observed phenotypic differences between the ABM of Listeria and Rickettsia may indicate fundamental differences in the mechanisms of actin recruitment and polymerization.

Infect Immun, 1999 Aug, 67(8), 4048 - 54
Interaction of Brucella abortus lipopolysaccharide with major histocompatibility complex class II molecules in B lymphocytes; Forestier C et al.; Lipopolysaccharide (LPS), a major amphiphilic molecule located at the outer membrane of gram-negative bacteria, is a potent antigen known to induce specific humoral immune responses in infected mammals . LPS has been described as a polyclonal activator of B lymphocytes, triggering the secretion of antibodies directed against distinct sugar epitopes of the LPS chain . But, how LPS is handled by B cells remains to be fully understood . This task appears to be essential for a better knowledge of the anti-LPS humoral immune response . In this study, we examine the internalization of LPS and its interaction with antigen-presenting major histocompatibility complex (MHC) class II molecules in murine and human B-cell lines . By use of immunofluorescence, we observe that structurally different LPSs from Brucella and Shigella strains accumulate in an intracellular compartment enriched in MHC class II molecules . By use of immunoprecipitation, we illustrate that only Brucella abortus LPS associates with MHC class II molecules in a haplotype-independent manner . Taken together, these results raise the possibility that B . abortus LPS may play a role in T-cell activation.

Infect Immun, 1999 Aug, 67(8), 3970 - 9
Thymic independence of adaptive immunity to the intracellular pathogen Shigella flexneri serotype 2a; Way SS et al.; Shigella flexneri is a facultative intracellular pathogen . While immunity to several intracellular pathogens is mediated by T lymphocytes, it is unknown whether cellular immune responses are important to adaptive immunity to S . flexneri . We show that vaccination with S . flexneri serotype 2a confers protection to mice that lack T lymphocytes or gamma interferon (IFN-gamma), specific depletion of T lymphocytes does not alter the protection, and adoptive transfer of splenocytes from vaccinated mice does not confer protection to naive mice . In contrast, vaccination conferred no protection to mice that lack B lymphocytes and adoptive transfer of immune sera conferred partial protection to naive mice . These data demonstrate that in the mouse bronchopulmonary model, adaptive immunity to S . flexneri 2a is an antibody-mediated, B-lymphocyte-dependent process and can be generated in the absence of T lymphocytes or IFN-gamma.

Rev Assoc Med Bras, 1999 Apr-Jun, 45(2), 121 - 7
{Lethality in hospitalized infants with acute diarrhea: risk factors associated with death}; de Andrade JA et al.; OBJECTIVES: Acute diarrhea is a very frequent disease in developing countries and is the first cause of death in infants under 2 years of age . This study was designed to evaluate the clinical and epidemiological factors associated to the death of 17 out of 511 infants hospitalized owing to severe acute diarrhea, between January 1989 and December 1995 . PATIENTS AND METHODS: The patients were divided into two groups according to their clinical evolution: Group I--Death and Group II--Survival . The following parameters were evaluated: birth weight, sex, age, duration of diarrhea (days) prior to admission, nutritional status, hydration, presence of an enteropathogenic agent in the stools, food intolerance and duration of hospitalization . RESULTS: The analyzed factors have shown a significant association with death for the following variables: age, relative risk (RR) = 4.0 for infants less than 6 months of age, identification of an enteropathogenic Escherichia coli (EPEC) strain in the stools (RR = 3.3), severe malnutrition at admission to the hospital (RR = 4.5), Occurrence of food intolerance during hospitalization (RR = 2.7) . Some enteropathogenic agent was identified in the stools of 253 (54.9%) infants, among the 461 (90.2%) studied . Group I revealed the presence of an enteropathogenic agent in 75% of the cases . The most frequent agents identified in Group I was: EPEC (56.3%) and Shigella (12.5%), while in Group II EPEC was identified in 26.5% of the patients . CONCLUSIONS: The association of some factors such as age less than 6 months, severe malnutrition, food intolerance and the identification of EPEC strains in the stool culture are indicators of high risk of death in infants hospitalized due to severe acute diarrhea.

Eur J Pediatr, 1999 Jul, 158(7), 550 - 2
Lethal encephalopathy complicating childhood shigellosis; Plotz FB et al.; A 6-year-old girl is described who died following rapid neurological deterioration, ending in lethal cerebral oedema . Despite the absence of severe intestinal and metabolic derangement, Shigella was cultured from the stool . Toxic encephalopathy is responsible for death following this rare complication of childhood shigellosis in developed countries . The pathophysiology is unknown . CONCLUSION: Lethal toxic encephalopathy can be caused by Shigella despite the absence of severe intestinal and metabolic derangement . If shigelllosis is suspected, headache may be a first significant sign for the development of toxic encephalopathy . Early recognition and rapid measures to prevent brain oedema may improve outcome.

Arch Latinoam Nutr, 1999 Mar, 49(1), 67 - 71
{Incidence of Plesiomonas shigelloides in tilapia tetrahibrids (Oreochromis sp.)}; Mendoza C et al.; Plesiomonas shigelloides, a member of the family Vibrionaceae, is a Gram negative rod associated with several gastroenteritis outbreaks, especially in tropical and subtropical countries . In same way, it has been related to some septicemia, meningitis and cholecystitis cases . The microorganism is normally found in water, fish and birds . The aim of this work was to study the incidence of Plesiomonas shigelloides in tetrahybrids of Oreochromis sp . (Pink Tilapia) located at the central region of Venezuela . Once the samples were homogenized, the techniques of enrichment and direct streaking were used simultaneously for the isolation of the microorganism . A high incidence of P . shigelloides was determined (73%), being higher in the intestinal tract (60%), followed by the skin (36.7%) and the gills (26.67%), without any correlation among them . In the fish pond, the microorganism isolation frequency was 41.67% . The direct streaking technique presented the highest isolation values in the different Tilapia tissues (60%) and in the water as well (41.60%) . No significant differences were observed on the effectivity of the selective agars used for the isolation of P . shigelloides (Plesiomonas Agar and Inositol-Brilliant Green-Bile Salts Agar) . A positive correlation was observed between the microorganism incidence and the pluviosity levels . A high incidence of E . coli was observed in the samples of Tilapia tissues and the water pond . No correlation was observed between incidence of P . shigelloides and E . coli . Due to the high prevalence of P . shigelloides found in the present study, it is important to assure a proper evisceration, washing and storage at temperatures lower than 8 degrees C, and a proper product cooking to diminish the customeris risk.

Przegl Epidemiol, 1999, 53(1-2), 93 - 101
{Dysentery in 1997}; Stypulkowska-Misiurewicz H et al.; Notifications of dysentery fell to their lowest yearly total since 1918 . The 1997 total figure of 439 was 18% lower than in 1996 (534 cases) and of 1353 cases (75%) lower than the median yearly figure in 1991-95 (1894 cases) . Incidence rate in 1997 was 1.1/100,000 inhabitance, the lowest ever notified . In 1996 incidence rate was 1.4 for 100,000 inhabitants and median incidence rate in 1991-1995 was 4.9 . Dysentery was not diagnosed in 24% administrative regions of Poland . No case of dysentery has been notified in 5/49 regions, only one single patients in 7/49 regions . 80% of patients were children 0-19 years old . The highest incidence rate (15.0 per 100,000) was found in children 3 years old and in children 4 years old (8.9/100,000)-residents of towns . Only bacteriological confirmed Shigella infection was notified as dysentery . In 1997 bacteriological examination for such purpose stopped to be done free of charge . Except one outbreak dispensed in Wroclaw all other outbreaks were limited to the children institution: schools, kindergartens or social care institutions . There were 12 outbreaks, 11 due to S . sonnei infection, and one to S . flexneri 1b . The quality control test performed by bacteriological laboratories of Sanitary Epidemiological Service revealed that not all of them used bacteriological media efficient for all types of Shigella.

Pediatr Infect Dis J, 1999 Jun, 18(6), 500 - 5
Population-based study of the incidence of Shigella diarrhea and causative serotypes in Santiago, Chile; Prado V et al.; BACKGROUND: Shigella is an important cause of diarrheal disease in children in developing countries . The increasing prevalence of antibiotic-resistant strains has stimulated interest in the use of multivalent Shigella vaccines . Because Shigella vaccines under development are based on eliciting immunity to O antigens, monitoring the distribution of serotypes in defined target populations is critical . We initiated health center-based surveillance in a poor semirural community in Colina, Santiago (7489 children <60 months of age) to determine the age-specific incidence of Shigella disease and the responsible serotypes . FINDINGS: Surveillance was maintained at the 2 health centers during warm seasons (November 1 through April 30) for 4 successive years (1994 to 1998) . Shigella was recovered from 54 of 243 cases of dysentery (22%) and from 215 of 3966 cases of nondysenteric diarrhea (5.4%) (P < 0.001) . The peak mean annual incidence of shigellosis occurred among children 12 to 47 months of age (9.0 to 12.6 cases/10(3) children), although the incidence in infants (5.2/10(3)) and children 48 to 59 months of age (6.2/10(3)) was also substantial . During the 1995 through 1996 season, an age-matched healthy control was cultured for every child <60 months of age with diarrhea . Shigella isolation from cases (34 of 576, 5.9%) was >8-fold higher than controls (4 of 576, 0.7%) (P < 0.01) . Four serotypes, Shigella sonnei (45%), Shigella flexneri 2b (19%), S . flexneri 2a (14%) and S . flexneri 6 (11%), accounted for 89% of all cases . INTERPRETATION: Shigella remains an important pediatric pathogen in Santiago . The serotype distribution from Colina, which closely resembles data from a population-based surveillance study in Santiago in the mid-1980s, demonstrates a remarkable degree of serotype stability in Santiago during a 15-year period.

Kansenshogaku Zasshi, 1999 May, 73(5), 414 - 20
{Strains of Shigella sonnei recently isolated in Tokyo}; Matsushita S et al.; A total of 341 Shigella sonnei strains consisting of 94 domestic strains and 247 imported strains isolated during 1990-1997 in Tokyo, were examined regarding their colicine-type, drug-resistance and ornithine-utilization . The colicine typing results showed that the domestic strains were classified into 7 types, and the imported strains were classified into 13 types . Among the colicine-types identified, 8-type, 0-type, 6-type and 12-type were predominant in the domestic strains, whereas 6-type, 0-type, 8-type, 9A-type and 12-type were predominant in the imported strains . The drug-resistance test using 9 drugs (CP, TC, SM, KM, ABPC, ST, NA, FOM and NFLX) showed that 89.4% of the domestic strains and 85.4% of the imported strains were resistant to some of the drugs except FOM and NFLX . Drugs with a high resistant rate were SM, TC and ST for both groups . Drug-resistance patterns of the resistant strains varied in 22 types . Among those, a triple drug-resistance type with TC.SM.ST was found in the most frequent pattern in both groups . The results of the ornithine-utilization test revealed that 28.7% of the domestic strains and 8.1% of the imported strains were negative . The ornithine-negative strains in the same source had a similar plasmid-profile, but generally there was no correlation between the different sources.

Infect Immun, 1999 Jul, 67(7), 3437 - 43
Vaccination against shigellosis with attenuated Shigella flexneri 2a strain SC602; Coster TS et al.; The Shigella flexneri 2a SC602 vaccine candidate carries deletions of the plasmid-borne virulence gene icsA (mediating intra- and intercellular spread) and the chromosomal locus iuc (encoding aerobactin) (S . Barzu, A . Fontaine, P . J . Sansonetti, and A . Phalipon, Infect . Immun . 64:1190-1196, 1996) . Dose selection studies showed that SC602 causes shigellosis in a majority of volunteers when 3 x 10(8) or 2 x 10(6) CFU are ingested . In contrast, a dose of 10(4) CFU was associated with transient fever or mild diarrhea in 2 of 15 volunteers . All volunteers receiving single doses of >/=10(4) CFU excreted S . flexneri 2a, and this colonization induced significant antibody-secreting cell and enzyme-linked immunosorbent assay responses against S . flexneri 2a lipopolysaccharide in two-thirds of the vaccinees . Seven volunteers who had been vaccinated 8 weeks earlier with a single dose of 10(4) CFU and 7 control subjects were challenged with 2 x 10(3) CFU of virulent S . flexneri 2a organisms . Six of the control volunteers developed shigellosis with fever and severe diarrhea or dysentery, while none of the vaccinees had fever, dysentery, or severe symptoms (P = 0 . 005) . Three vaccinees experienced mild diarrhea, and these subjects had lower antibody titers than did the fully protected volunteers . Although the apparent window of safety is narrow, SC602 is the first example of an attenuated S . flexneri 2a candidate vaccine that provides protection against shigellosis in a stringent, human challenge model.

Microbiology, 1999 May, 145 ( Pt 5), 1263 - 73
Functional analysis of the O antigen glucosylation gene cluster of Shigella flexneri bacteriophage SfX; Guan S et al.; Previous studies have shown that Shigella flexneri bacteriophage X (SfX) encodes a glucosyltransferase (GtrX, formerly Gtr), which is involved in O antigen modification (serotype Y to serotype X) . However, GtrX alone can only mediate a partial conversion . More recently, a three-gene cluster has been identified next to the attachment site in the genome of two other S . flexneri bacteriophages (i.e . SfV and SfII) . This gene cluster was postulated to be responsible for a full O antigen conversion . Here it is reported that besides the gtrX gene, the other two genes in the gtr locus of SfX were also involved in the O antigen modification process . The first gene in the cluster (gtrA) encodes a small highly hydrophobic protein which appears to be involved in the translocation of lipid-linked glucose across the cytoplasmic membrane . The second gene in the cluster (gtrB) encodes an enzyme catalysing the transfer of the glucose residue from UDP-glucose to a lipid carrier . The third gene (gtrX) encodes a bacteriophage-specific glucosyltransferase which is largely responsible for the final step, i.e . attaching the glucosyl molecules onto the correct sugar residue of the O antigen repeating unit . A three-step model for the glucosylation of bacterial O antigen has been proposed.

EMBO J, 1999 Jun 15, 18(12), 3249 - 62
IpaC induces actin polymerization and filopodia formation during Shigella entry into epithelial cells; Tran Van Nhieu G et al.; Shigella proteins that are targeted to host cells by a type III secretion apparatus are essential for reorganization of the cytoskeleton during cell invasion . We have developed a semi-permeabilized cell assay that tests the effects of bacterial proteins on the actin cytoskeleton . The Shigella IpaC protein was found to induce the formation of filopodial and lamellipodial extensions in these semi-permeabilized cells . Microinjection of IpaC into cells, or cellular expression of IpaC also led to the formation of filopodial structures . Monoclonal antibodies (mAbs) directed against the C-terminus of IpaC inhibited the IpaC-induced extensions, whereas an anti-N-terminal IpaC mAb stimulated extensive lamellae formation . Shigella induced foci of actin polymerization in the permeabilized cells and these were inhibited by anti-C-terminal IpaC mAbs . Consistent with a role for IpaC in Shigella-induced cytoskeletal rearrangements during entry, stable transfectants expressing IpaC challenged with Shigella showed increased bacterial internalization . IpaC-induced extensions were inhibited by a dominant-interfering form of Cdc42 or the Cdc42-binding domain of WASP, whereas a dominant-interfering form of Rac resulted in inhibition of lamellae formation . We conclude that IpaC leads to activation of Cdc42 which in turn, causes activation of Rac, both GTPases being required for Shigella entry.

Vet Res, 1999 Mar-Jun, 30(2-3), 235 - 57
Shiga/verocytotoxins and Shiga/verotoxigenic Escherichia coli in animals; Mainil J; Vero/Shiga toxins (VT/Stx) have an A-B structure: the A subunit carries the enzymatic activity and the B subunit binds the toxin to the membrane receptor (Gb3 or Gb4) . The VT/Stx inhibit protein synthesis in the target eucaryotic cells, mainly the endothelial cells of blood vessels . The VT/Stx are subdivided into two families . VT1/Stx1 is a homogeneous family of toxins identical to the Stx of Shigella dysenteriae . VT2/Stx2 is a more heterogeneous family of toxins more distantly related to this Stx toxin . The VT2/Stx2 variants can be distinguished by the polymerase chain reaction (PCR) and/or the reaction with monoclonal antibodies . The VT/Stx-producing Escherichia coli are also subdivided into two main groups on the basis of the presence or absence of additional properties: the enterohaemorrhagic E . coli (EHEC) induce the formation of attaching/effacing lesions and carry a 60 MD plasmid encoding a specific haemolysin (the enterohaemolysin); the vero/shiga-toxigenic E . coli (VTEC/STEC) do not show these properties . The EHEC are isolated from humans and ruminants, especially young calves . They are associated with haemorrhagic enterocolitis and its sequelae in humans, the haemolytic-uraemic syndrome (HUS) . The VT/Stx play a role in the occurrence of blood in the faeces and in the HUS by their action on the endothelial cells of blood vessels in the intestinal submucosa and in the renal glomeruli, after resorption through the intestinal walls . The VTEC/STEC are isolated from piglets, calves and humans . In recently weaned piglets, they cause the oedema disease, an enterotoxaemia characterized by subcutaneous, mesenteric and cerebral oedemas, with nervous disorders as main clinical signs . The oedema disease is the consequence of the action of the VT/Stx on the endothelial cells of blood vessels in various organs . In calves and humans, the role in disease of VTEC/STEC is controversial, but they could be associated with some cases of diarrhoea and HUS . The case of the O157:H7 EHEC which are present in healthy cattle of various ages, but are highly virulent for humans is of special interest . The potential zoonotic aspect of VT/Stx-producing E . coli infections in animals is detailed chapter by chapter . Prophylaxis of these infections by vaccination is the subject of the discussion on the future of the research studies on these pathogenic bacteria.

J Clin Microbiol, 1999 Jul, 37(7), 2352 - 3
An untypeable Shigella flexneri strain associated with an outbreak in California; Trevejo RT et al.; Eleven Shigella flexneri (group B) isolates were recovered from epidemiologically linked patrons and food handlers from a restaurant-associated outbreak of shigellosis . Six isolates available for pulsed-field gel electrophoresis were identical . All strains agglutinated in group B and subgroup factor 6 sera but not in group 1 through group 6 sera.

J Cell Sci, 1999 Jul, 112 ( Pt 13), 2069 - 80
Rho family GTPases control entry of Shigella flexneri into epithelial cells but not intracellular motility; Mounier J et al.; Shigella flexneri, an invasive bacterial pathogen, promotes formation of two cytoskeletal structures: the entry focus that mediates bacterial uptake into epithelial cells and the actin-comet tail that enables the bacteria to spread intracellularly . During the entry step, secretion of bacterial invasins causes a massive burst of subcortical actin polymerization leading the formation of localised membrane projections . Fusion of these membrane ruffles leads to bacterial internalization . Inside the cytoplasm, polar expression of the IcsA protein on the bacterial surface allows polymerization of actin filaments and their organization into an actin-comet tail leading to bacterial spread . The Rho family of small GTPases plays an essential role in the organization and regulation of cellular cytoskeletal structures (i.e . filopodia, lamellipodia, adherence plaques and intercellular junctions) . We show here that induction of Shigella entry foci is controlled by the Cdc42, Rac and Rho GTPases, but not by RhoG . In contrast, actin-driven intracellular motility of Shigella does not require Rho GTPases . Therefore, Shigella appears to manipulate the epithelial cell cytoskeleton both by Rho GTPase-dependent and -independent processes.

J Cell Sci, 1999 Jul, 112 ( Pt 13), 2059 - 68
A functional role for ezrin during Shigella flexneri entry into epithelial cells; Skoudy A et al.; Shigella flexneri is an enteroinvasive bacterium responsible for bacillary dysentery in humans . Bacterial entry into epithelial cells is a crucial step for the establishment of the infection . It 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 by a macropinocytic process . We show in this study that the membrane-cytoskeleton linker, ezrin, a member of the ERM (ezrin, radixin, moesin) family, plays an active role in the process of Shigella uptake . Ezrin is highly enriched in cellular protrusions induced by the bacterium and is found in close association with the plasma membrane . In addition, Shigella entry is significantly reduced in cells transfected with a dominant negative allele of ezrin with entry foci showing much shorter cellular protrusions . These results indicate that ezrin not only acts as a membrane-cytoskeleton linker, but may also mediate extension of cellular projections in the presence of signals such as those elicited by invading microorganisms.

Zh Mikrobiol Epidemiol Immunobiol, 1999 Mar-Apr, (2), 42 - 5
{The etiological structure of acute intestinal infections in children in the Republic of Tajikistan}; Rafiev KhK; The Republic of Tajikistan belongs to the territories, traditionally hyperendemic with respect to acute enteric diseases (AED) . The problem of AED still remains topical in recent years, especially among child population . The study of the etiological structure of AED in children has made it possible to establish that more than 70% of these diseases are of infectious nature . The leading role in the etiological structure of these diseases belongs to Shigella infections (32.4%), enterovirus diarrhea (12.1%) and Escherichia infections (8.9%) . No significant differences in the character of the etiological structure of AED in children of urban and rural areas have been detected with the exception of enterovirus diarrhea, found to occur 2.7 times more frequently on children of urban areas (15.9%) than in those of rural areas (5.8%) . The ascertaining of the etiological structure of AED in Tajikistan will make it possible to essentially increase epidemiological surveillance on AED and render it more concrete.

FEBS Lett, 1999 Apr 30, 450(1-2), 72 - 6
A low rate of nucleotide changes in Escherichia coli K-12 estimated from a comparison of the genome sequences between two different substrains; Itoh T et al.; Two genome sequences of Escherichia coli K-12 substrains, one partial W3110 and one complete MG1655, have been determined by Japanese and American genome projects, respectively . In order to estimate the rate of nucleotide changes, we directly compared 2 Mb of the nucleotide sequences from these closely-related E . coli substrains . Given that the two substrains separated about 40 years ago, the rate of nucleotide changes was estimated to be less than 10(-7) per site per year . This rate was supported by a further comparison between partial genome sequences of E . coli and Shigella flexneri.

Rev Cubana Med Trop, 1998, 50(3), 203 - 6
{The antimicrobial sensitivity and plasmid isolation in Plesiomonas shigelloides}; Bravo Farinas L et al.; 30 strains of Plesiomonas shigelloides isolated from patients with acute diarrheal disease at different health centers of the country were studied . The were phenotypically characterized by conventional biochemical tests and the antimicrobial susceptibility to 11 drugs was determined by the Kirby Bauer's method . It was found that the strains of Plesiomonas shigelloides were sensitive to 7 and resistant to 6 of the investigated drugs . The presence of plasmids in 12 of the 29 analyzed strains was determined and the diversity in their plasmidic patterns was proved.

Rev Med Chil, 1998 Dec, 126(12), 1464 - 71
{Antimicrobial multiresistance of Shigella sp strains in a semi rural community of northern Santiago}; Prado V et al.; Appropriate antimicrobial therapy shortens the duration of Shigellosis and significantly reduces the risk of transmission . Shigella strains resistant to common antimicrobials have increased during the past years, determining the need for a periodic surveillance, to guide effective therapy . AIM: To report the results of a surveillance program in a rural community near Santiago (Colina), for Shigella infections . MATERIAL AND METHODS: Between 1995 and 1997, stool samples from 3,534 episodes of diarrhoea, that occurred in Colina, were obtained . Two hundred twenty six Shigella strains were isolated and studied for susceptibility to ampicilin (AM), amoxicillin/clavulanic acid (AMC), cotrimoxazole (STX), chloramphenicol (CAF), tetracycline (TET), furazolidine (FU), ciprofloxacine (CIPR), nalidixic acid (AC NAL), gentamycin (GENT) and cefotaxime (CFTX) . RESULTS: Shigella flexnerii represented 134 of 226 Shigella strains isolated . All strains were susceptible to CIPR, AC NAL, GENT and CFTX . Yearly variation of resistance patterns to other antimicrobials were observed for these strains . Resistance to AM varied from 56 to 76%, to AMC from 25 to 56%, to STX from 21 to 47%, to CAF from 36 to 69%, to TET from 44 to 78% and to FU from 9 to 18% . Overall resistance was higher during 1997 . All 85 strains of S sonnei were susceptible to CIPR, AC NAL and CFTX . Resistance throughout the years varied from 56 to 88% for AM, from 0 to 28% for AMC, from 44 to 53% for STX, from 11 to 40% for CAF, from 11 to 42% for TET and from 5 to 11% for FU . Overall resistance was also higher during 1997, except for AM and STX . Seven S hoydii strains were isolated, only during 1995 . All seven were resistant to AM and TET and none were resistant to FU, CIPR, AC NAL and CFTX . Two strain was resistant to AMC, STX and CAF . CONCLUSIONS: Antimicrobial resistance patterns of Shigella sp isolated in Colina have increased from 1995 to 1997, specially for commonly used antimicrobials . Resistance remains low for furazolidine and all strains remain susceptible to quinolones.

J Cell Biol, 1999 May 17, 145(4), 689 - 98
Trafficking of Shigella lipopolysaccharide in polarized intestinal epithelial cells; Beatty WL et al.; Bacterial lipopolysaccharide (LPS) at the apical surface of polarized intestinal epithelial cells was previously shown to be transported from the apical to the basolateral pole of the epithelium (Beatty, W.L., and P.J . Sansonetti . 1997 . Infect . Immun . 65:4395-4404) . The present study was designed to elucidate the transcytotic pathway of LPS and to characterize the endocytic compartments involved in this process . Confocal and electron microscopic analyses revealed that LPS internalized at the apical surface became rapidly distributed within endosomal compartments accessible to basolaterally internalized transferrin . This compartment largely excluded fluid-phase markers added at either pole . Access to the basolateral side of the epithelium subsequent to trafficking to basolateral endosomes occurred via exocytosis into the paracellular space beneath the intercellular tight junctions . LPS appeared to exploit other endocytic routes with much of the internalized LPS recycled to the original apical membrane . In addition, analysis of LPS in association with markers of the endocytic network revealed that some LPS was sent to late endosomal and lysosomal compartments.

J Cell Sci, 1999 Jun, 112 ( Pt 11), 1697 - 708
A comparative study of the actin-based motilities of the pathogenic bacteria Listeria monocytogenes, Shigella flexneri and Rickettsia conorii; Gouin E et al.; Listeria monocytogenes, Shigella flexneri, and Rickettsia conorii are three bacterial pathogens that are able to polymerize actin into 'comet tail' structures and move within the cytosol of infected cells . The actin-based motilities of L . monocytogenes and S . flexneri are known to require the bacterial proteins ActA and IcsA, respectively, and several mammalian cytoskeleton proteins including the Arp2/3 complex and VASP (vasodilator-stimulated phosphoprotein) for L . monocytogenes and vinculin and N-WASP (the neural Wiskott-Aldrich syndrome protein) for S . flexneri . In contrast, little is known about the motility of R . conorii . In the present study, we have analysed the actin-based motility of this bacterium in comparison to that of L . monocytogenes and S . flexneri . Rickettsia moved at least three times more slowly than Listeria and Shigella in both infected cells and Xenopus laevis egg extracts . Decoration of actin with the S1 subfragment of myosin in infected cells showed that the comet tails of Rickettsia have a structure strikingly different from those of L . monocytogenes or S . flexneri . In Listeria and Shigella tails, actin filaments form a branching network while Rickettsia tails display longer and not cross-linked actin filaments . Immunofluorescence studies revealed that the two host proteins, VASP and (&agr;)-actinin colocalized with actin in the tails of Rickettsia but neither the Arp2/3 complex which we detected in the Shigella actin tails, nor N-WASP, were detected in Rickettsia actin tails . Taken together, these results suggest that R . conorii may use a different mechanism of actin polymerization.

FEMS Microbiol Lett, 1999 May 1, 174(1), 89 - 95
Shigella flexneri YSH6000 induces two types of cell death, apoptosis and oncosis, in the differentiated human monoblastic cell line U937; Nonaka T et al.; Shigella flexneri, but not a non-invasive mutant derivative rapidly induced cell death in human monoblastic U937 cells as well as in differentiated cells pretreated with interferon-gamma (IFN gamma) or retinoic acid (RA) . We investigated the morphological and biochemical characteristics of bacterial invasion-induced cell death in these differentiated U937 cells . IFN gamma-differentiated cells showed morphological changes typical of apoptosis and their DNA was cleaved giving a ladder-like electrophoretic pattern after infection by Shigellae . In contrast, swelling of the cytoplasm and blebbing of the plasma membrane were observed in RA-differentiated and undifferentiated cells invaded by the bacteria . No condensation of nuclei was observed in these cells by light microscopy, and no internucleosomal fragmentation of DNA was detected on agarose gels, which resembled the features of oncosis . Furthermore, cleavage of poly(ADP-ribose) polymerase, a substrate for apoptotic caspases, was seen only in IFN gamma-pretreated cells but not in RA-pretreated or undifferentiated cells . These findings suggested that virulent Shigella flexneri induces distinct types of cell death in U937 cells depending on their differentiation state.

Environ Health Perspect, 1999 Feb, 107 Suppl 1, 191 - 206
Microbiological safety of drinking water: United States and global perspectives; Ford TE; Waterborne disease statistics only begin to estimate the global burden of infectious diseases from contaminated drinking water . Diarrheal disease is dramatically underreported and etiologies seldom diagnosed . This review examines available data on waterborne disease incidence both in the United States and globally together with its limitations . The waterborne route of transmission is examined for bacterial, protozoal, and viral pathogens that either are frequently associated with drinking water (e.g., Shigella spp.), or for which there is strong evidence implicating the waterborne route of transmission (e.g., Leptospira spp.) . In addition, crucial areas of research are discussed, including risks from selection of treatment-resistant pathogens, importance of environmental reservoirs, and new methodologies for pathogen-specific monitoring . To accurately assess risks from waterborne disease, it is necessary to understand pathogen distribution and survival strategies within water distribution systems and to apply methodologies that can detect not only the presence, but also the viability and infectivity of the pathogen.

Mol Microbiol, 1999 Apr, 32(2), 367 - 77
The unipolar Shigella surface protein IcsA is targeted directly to the bacterial old pole: IcsP cleavage of IcsA occurs over the entire bacterial surface; Steinhauer J et al.; Shigella flexneri is an intracellular pathogen that is able to move within the cytoplasm of infected cells by the continual assembly of actin onto one pole of the bacterium . IcsA, an outer membrane protein, is localized to the old pole of the bacterium and is both necessary and sufficient for actin assembly . IcsA is slowly cleaved from the bacterial surface by the protease IcsP (SopA) . Absence of IcsP leads to an alteration in the distribution of surface IcsA, such that the polar cap is maintained and some IcsA is distributed along the lateral walls of the bacillus . The mechanism of unipolar localization of IcsA and the role of IcsP in its unipolar localization are incompletely understood . Here, we demonstrate that cleavage of IcsA occurs exclusively in the outer membrane and that IcsP is localized to the outer membrane . In addition, we show that IcsA at the old pole is susceptible to cleavage by IcsP and that native IcsP is active at the pole . Taken together, these data indicate that IcsP cleaves IcsA over the entire bacterial surface . Finally, we show that, immediately after induction from a tightly regulated promoter, IcsA is expressed exclusively at the old pole in both the icsP- icsA- and the icsA- background . These data demonstrate that unipolar localization of IcsA results from its direct targeting to the pole, followed by its diffusion laterally in the outer membrane.

J Infect Dis, 1999 Jun, 179(6), 1565 - 8
Safety and immunogenicity of Shigella sonnei and Shigella flexneri 2a O-specific polysaccharide conjugates in children; Ashkenazi S et al.; O-specific polysaccharide conjugates of shigellae were safe and immunogenic in young adults, and a Shigella sonnei conjugate conferred protection {1-3} . Shigellosis is primarily a disease of children; therefore, the safety and immunogenicity of S . sonnei and Shigella flexneri 2a conjugates were studied in 4- to 7-year-old children . Local and systemic reactions were minimal . The first injection of both conjugates elicited significant rises in geometric mean levels of serum IgG only to the homologous lipopolysaccharide (LPS) (S . sonnei, 0.32-8.25 ELISA units {EU}; S . flexneri 2a, 1.15-20.5 EU; P<.0001) . Revaccination at 6 weeks induced a booster response to S . flexneri 2a LPS (20.5-30.5 EU, P=.003) . Six months later, the geometric mean levels of IgG anti-LPS for both groups were higher than the prevaccination levels (P<.0001) . Similar, but lesser, rises were observed for IgM and IgA anti-LPS . The investigational Shigella conjugates were safe and immunogenic in children and merit evaluation of their efficacy.

MMWR Morb Mortal Wkly Rep, 1999 Apr 16, 48(14), 285 - 9
Outbreaks of Shigella sonnei infection associated with eating fresh parsley--United States and Canada, July-August 1998.
An epidemic of bloody diarrhea: Escherichia coli O157 emerging in Cameroon?
Centre Pasteur du Cameroun, Yaounde, CameroonBetween November 1997 and April 20, 1998, bloody diarrhea sickened 298 persons in Cameroon . Laboratory investigation of the epidemic (case-fatality rate, 16.4%) documented amoebiasis in one of three patients and three types of pathogens: multidrug-resistant Shigella dysenteriae type 1, S . boydii, and enterohemorrhagic Escherichia coli . We report the first isolation of E . coli O157:H7 in Cameroon and the second series of cases in the Central African region.

Proc Natl Acad Sci U S A, 1999 Apr 27, 96(9), 5194 - 7
Protein conjugates of synthetic saccharides elicit higher levels of serum IgG lipopolysaccharide antibodies in mice than do those of the O-specific polysaccharide from Shigella dysenteriae type 1; Pozsgay V et al.; Our development of vaccines to prevent shigellosis is based on the hypothesis that a critical (protective) level of serum IgG to the O-specific polysaccharide (O-SP) domain of Shigella lipopolysaccharide (LPS) confers immunity . The O-SP is a hapten and must be conjugated to a protein to induce serum antibodies . The O-SP of Shigella dysenteriae type 1 (approximately 27 tetrasaccharide repeat units), prepared by acid hydrolysis of the LPS, was bound to human serum albumin (HSA) by multiple point attachment (O-SP-HSA): The molar ratio of HSA to O-SP was 1.0 . Synthetic saccharides, composed of one or multiples of the O-SP tetrasaccharide, equipped with a spacer at their reducing end, were bound to HSA by a single point attachment: The average molar ratios of the saccharides to HSA ranged from 4 to 24 . Serum IgG anti-LPS, elicited in mice by O-SP-HSA or synthetic tetra-, octa-, dodeca-, and hexadecasaccharide fragments, was measured by ELISA . Outbred 6-week-old female mice were injected s.c . three times at biweekly intervals with 2.5 micrograms of saccharide as a conjugate and were bled 7 days after the second and third injections . Excepting the tetramer, conjugates of the octamer, dodecamer and hexadecamer elicited IgG LPS antibodies after the second injection, a statistically significant rise (booster) after the third injection, and higher levels than those vaccinated with O-SP-HSA (P = 0.0001) . The highest geometric mean levels of IgG anti-LPS were elicited by the hexadecamer with 9 chains or 9 moles of saccharide/HSA (15.5 ELISA units) followed by the octamer with 20 chains (11.1 ELISA units) and the dodecamer with 10 chains (9.52 ELISA units) . Clinical evaluation of these synthetic saccharides bound to a medically useful carrier is planned.

Jpn J Med Sci Biol, 1998 Nov, 51(1), 1 - 12
Prevalence, in-vitro secretory activity, and cytotoxicity of Aeromonas species associated with childhood gastroenteritis in Chennai (Madras), India; Alavandi S et al.; An investigation on the prevalence of Aeromonas in gastrointestinal illnesses of pediatric inpatients 1 month to 3 years of age was conducted from February 1997 through January 1998 in Madras . Sixteen Aeromonas spp . were isolated from 11 male and five female children among the 341 pediatric inpatients suffering from acute diarrhoea . A . caviae, which was isolated from nine cases, was found to be the most predominant isolate, followed by A . veronii biovar sobria, isolated from six cases, and A . hydrophila, isolated from one case . Shigella flexneri was recovered along with Aeromonas veronii biotype sobria serotype 035 from one 5-month-old female child . We did not notice any seasonal pattern in the association between Aeromonas and childhood gastroenteritis . None of the 147 stool samples obtained from age-matched non-diarrhoeic control children yielded Aeromonas spp . Isolation of Aeromonas spp . from patients suffering from gastroenteritis was found to be significant (chi 2 = 7.1312; P = 0.008, < 0.01) . Among the 16 Aeromonas isolates, seven isolates of A . caviae and two isolates of A . veronii biovar sobria induced a secretory response in rabbit intestinal mucosa mounted in Ussing chambers as demonstrated by a significant increase in the short circuit current . Nine of the 16 Aeromonas isolates, including three isolates of A . caviae, five isolates of A . veronii biovar sobria, and the solitary isolate of A . hydrophila were also cytotoxic to CHO cells . Five of the six isolates of A . veronii biovar sobria and the A . hydrophila isolate produced hemolysin . The results of this study indicate that Aeromonas species are important causative agents of diarrhoea in childhood gastroenteritis and are prevalent throughout the year in Madras.

Lik Sprava, 1998 Dec, (8), 144 - 6
{The efficacy of Intetrix in the combined treatment of patients with shigelloses}; Sukhov IuA; Efficacy was studied of Intetrix in the treatment of acute Sonnei's shigellosis and Flexner's shigellosis in 32 patients presenting with mild and moderately severe course of their illness . The Intretrix group patients fared better compared with controls who received furazolidon as an etiotropic drug, with the duration of intoxication symptoms getting shorter, abdominal pains and diarrhea subsiding . Intetrix tended to be associated with a lower incidence of unfavourable outcomes, both immediate and remote ones . The data obtained show expediency of treatment with Intetrix of patients with shigelloses running a mild or moderately severe course.

Clin Infect Dis, 1999 Mar, 28(3), 466 - 75
Rupture of the intestinal epithelial barrier and mucosal invasion by Shigella flexneri; Sansonetti PJ et al.; Invasion of the intestinal barrier by Shigella flexneri involves complex interactions with epithelial and phagocytic cells . Major perturbation of the signals that maintain epithelial integrity permits mucosal invasion, leading to tissue destruction . Expression of this invasive phenotype depends on the secretion of Ipa proteins (invasins), which can trigger entry of the pathogen into epithelial cells by causing massive rearrangement of the host cell cytoskeleton and cause macrophage apoptotic death by direct interaction of IpaB with interleukin-1beta (IL-1beta)-converting enzyme . This results in the killing of defense cells and in the release of IL-1beta . In vivo, bacteria translocate through the epithelial barrier, essentially via M cells of the follicle-associated epithelium in the colonic and rectal mucosa . Apoptotic death of macrophages in subepithelial tissues allows bacterial survival and triggers inflammation, which destabilizes epithelial structures and facilitates further bacterial entry . Once they are intracellular, bacteria multiply within the cytoplasm and move from cell to cell by an actin-dependent process.

FEMS Microbiol Lett, 1999 Mar 15, 172(2), 137 - 43
Analysis, expression and prevalence of the Mycobacterium tuberculosis homolog of bacterial virulence regulating proteins; Gupta S et al.; We have previously reported the identification of a gene from Mycobacterium tuberculosis, H37Rv, which on the basis of its nucleotide sequence encoded a protein product of 38 kDa . This 38-Kda mycobacterial protein designated as VirS exhibits homology with the VirF protein of Shigella, the VirFy protein of Yersinia and the Cfad, Rns and FapR proteins from various enterotoxigenic Escherichia coli strains . In this communication, we show the close sequence and structural similarities of the VirS protein with VirF, VirFy, Cfad, Rns and FapR and describe the results of our studies on the characterization of the virS gene promoter and its expression in E . coli and mycobacteria . virS was present exclusively in the species belonging to the M . tuberculosis complex as revealed by Southern blot and PCR analysis . Our findings suggest the involvement of virS in the regulation of pathogenesis of M . tuberculosis.

Acta Paediatr, 1999 Feb, 88(2), 161 - 5
Killed oral Shigella vaccine made from Shigella flexneri 2a protects against challenge in the rabbit model of shigellosis; Chakrabarti MK et al.; The protective efficacy of an orally administered heat-killed virulent strain of Shigella flexneri 2a (5 weekly oral doses) was evaluated in 25 rabbits (14 immunized and 11 non-immunized controls) against challenge with the same strain of Shigella using the rabbit model of shigellosis . All 11 non-immunized rabbits developed bloody diarrhoea following challenge and 6 (54%) died . None of the 14 immunized rabbits developed diarrhoea (all had pellet stools) but 3 (21%) died from causes not associated with diarrhoea . Protection from diarrhoea and dysentery following oral immunization with a killed Shigella species was 100% and highly significant . Death following challenge was 2.5-fold higher in the non-immunized group (p = 0.115) but was not significant . These promising results suggest that further studies should be undertaken to develop a killed oral vaccine against shigellosis.

J Bacteriol, 1999 Apr, 181(7), 2110 - 7
Binding site recognition by Rns, a virulence regulator in the AraC family; Munson GP et al.; The expression of CS1 pili by enterotoxigenic strains of Escherichia coli is regulated at the transcriptional level and requires the virulence regulator Rns, a member of the AraC family of regulatory proteins . Rns binds at two separate sites upstream of Pcoo (the promoter of CS1 pilin genes), which were identified in vitro with an MBP::Rns fusion protein in gel mobility and DNase I footprinting assays . At each site, Rns recognizes asymmetric nucleotide sequences in two regions of the major groove and binds along one face of the DNA helix . Both binding sites are required for activation of Pcoo in vivo, because mutagenesis of either site significantly reduced the level of expression from this promoter . Thus, Rns regulates the expression of CS1 pilin genes directly, not via a regulatory cascade . Analysis of Rns-nucleotide interactions at each site suggests that binding sites for Rns and related virulence regulators are not easily identified because they do not bind palindromic or repeated sequences . A strategy to identify asymmetric binding sites is presented and applied to locate potential binding sites upstream of other genes that Rns can activate, including those encoding the CS2 and CFA/I pili of enterotoxigenic E . coli and the global regulator virB of Shigella flexneri.

Diagn Microbiol Infect Dis, 1999 Mar, 33(3), 187 - 99
Natural antibiotic susceptibility of Escherichia coli, Shigella, E . vulneris, and E . hermannii strains; Stock I et al.; The natural antibiotic susceptibility of 139 Escherichia coli strains (including 18 enterohemorrhagic E . coli), 73 Shigella strains (S . sonnei (n = 37), S . flexneri (n = 29), S . boydii (n = 6), S . dysenteriae (n = 1)), 23 E . vulneris, and 20 E . hermannii strains toward 71 antibiotics was examined . MICs were determined using a microdilution procedure . All examined taxa were naturally sensitive/intermediate toward tetracyclines, aminoglycosides, some penicillins (amoxycillin/clavulanate, ampicillin/sulbactam, piperacillin {with and without tazobactam}, mezlocillin, azlocillin), cephalosporins, carbapenems, monobactams, quinolones, trimethoprim, cotrimoxazole, and chloramphenicol and were naturally resistant/intermediate toward benzylpenicillin, oxacillin, macrolides, lincosamides, glycopeptides, rifampicin, and fusidic acid . No differences in natural antibiotic susceptibility were seen between enterohemorrhagic and other E . coli strains . Likewise, with one exception, no significant differences in natural antibiotic susceptibility were seen either among the Shigella subgroups or between Shigella and E . coli . The natural population of S . flexneri was slightly more susceptible to chloramphenicol than the natural populations of other taxa within the Shigella-E . coli complex . E . vulneris and E . hermannii showed susceptibility patterns to many antibiotics similar to Shigella and E . coli, but there were other antibiotics toward which there were significant differences in natural susceptibility . E . vulneris and E . hermannii were less susceptible to nitrofurantoin and slightly more susceptible to several aminoglycosides than E . coli and Shigella . E . hermannii was the only species that was naturally resistant/intermediate to ticarcillin and amoxycillin (DIN standard) . The addition of clavulanic acid to the latter resulted in a decrease of seven twofold dilution steps (E . vulneris: four twofold dilution steps, E . coli/Shigella: two twofold dilution steps) of the MICs of the natural population . With the exception of cefoperazone and cefepime, E . hermannii was more susceptible to cephalosporins than strains of the other species . E . vulneris was the species most susceptible to ticarcillin and the only species that was highly resistant to fosfomycin (MIC > 256 micrograms/mL) . The antibiotic susceptibility to fosfomycin was also unique for E . hermannii (MIC 32-128 micrograms/mL), whereas the natural populations of E . coli and Shigella showed lower MIC values . The data of this study represent an assessment of the natural susceptibility of strains of Escherichia spp . and Shigella subgroups to a wide range of antibiotics . These databases can be used for the validation of antibiotic susceptibility test results of Escherichia spp . and shigellae . The conformity of the natural antibiotic susceptibility test results not only among the Shigella subgroups but also between Shigella and E . coli support the status of Shigella as a subgroup of the species E . coli.

Lab Anim Sci, 1998 Aug, 48(4), 330 - 3
Shigella infection in macaque colonies: case report of an eradication and control program; Wolfensohn S; A primate colony comprising three distinct but interrelated units had long-term history of undiagnosed diarrhea and associated deaths for many years . In 1989, the clinical problem was recognized as a confounding factor for the experimental work, and steps were taken to eradicate the disease . This was done by a combined approach involving improved sample collection techniques and microbiological methods, treatment of all animals in the colony, and improvement in management . These management changes included alterations in basic facility and cage design, disinfection procedures, and continuous routine microbiological sampling of all groups of animals on a random basis, as well as sampling of those suspected to be at risk for stress-associated Shigella shedding . Using this approach, we have eliminated clinical cases of shigellosis and have not have any further isolations of Shigella from this colony.

J Commun Dis, 1997 Dec, 29(4), 351 - 4
A study on transferable R-plasmids among Shigella species at Lucknow; Agarwal SK et al.; A total of 500 Shigella strains were included in the present study out of which 233 were isolated at National Shigella Centre of Postgraduate Department of Microbiology, K.G . Medical College, Lucknow during last 5 years and 267 strains were received from different parts of the country . Predominent serotypes were Sheigella flexneri 292 (48.4%) followed by Shigella dysenteriae 153 (30.6%), Shigella boydii 32 (6.4%) and Shigella sonnei 23 (4.6%) . All the strains were sensitive to Nalidix acid, Gentamycin, Kanamycin, Neomycin and Furazolidine . Varying degree of resistance was shown to Streptomycin, Chloramphenicol, Tetracycline and Ampicillin . R . plasmid extraction was done and transfer was studied on E . Coli K-12.

Infect Immun, 1999 Apr, 67(4), 2001 - 4
Adaptive immune response to Shigella flexneri 2a cydC in immunocompetent mice and mice lacking immunoglobulin A; Way SS et al.; Shigella flexneri cydC, which is deficient in cytochrome bd, was rapidly cleared from the lungs of intranasally inoculated mice and was Sereny negative, yet it induced 93% protection against challenge with wild-type S . flexneri . Mice that lack immunoglobulin A (IgA) were fully protected, suggesting that IgA may not be required for adaptive immunity in this model system.

Infect Immun, 1999 Apr, 67(4), 1982 - 91
The mxi-Spa type III secretory pathway of Shigella flexneri requires an outer membrane lipoprotein, MxiM, for invasin translocation; Schuch R et al.; Invasion of epithelial cells by Shigella flexneri is mediated by a set of translocated bacterial invasins, the Ipa proteins, and its dedicated type III secretion system, called Mxi-Spa . We show here that mxiM, part of the mxi-spa locus in the S . flexneri virulence plasmid, encodes an indispensable type III secretion apparatus component, required for both Ipa translocation and tissue culture cell invasion . We demonstrated that mature MxiM, first identified as a putative lipoprotein, is lipidated in vivo . Consistent with features of known lipoproteins, MxiM (i) can be labeled with {3H}palmitate and {2-3H}glycerol, (ii) is associated with the cell envelope, (iii) is secreted independently of the type III pathway, and (iv) requires an intact lipoprotein modification and processing site for full activity . The lipidated form of MxiM was detected primarily in the outer membrane, where it establishes a peripheral association with the inner leaflet . Through analysis of subcellular Ipa distribution in a mxiM null mutant background, MxiM was found to be required for the assembly and/or function of outer, but not inner, membrane regions of Mxi-Spa . This function probably requires interactions with other Mxi-Spa subunits within the periplasmic space . We discuss implications of these findings with respect to the function of MxiM and the structure of Mxi-Spa as a whole.

Cent Eur J Public Health, 1999 Feb, 7(1), 47 - 9
Plesiomonas shigelloides and its serovars in animals in the Czech Republic--region Moravia; Bardon J; In the period of 1994-1997, 4,552 samples of sectional, clinical material and environment were examined for the presence of Plesiomonas shigelloides . In 55 cases (1,21%) Plesiomonas shigelloides was isolated . Within 55 isolated strains, 26 serovars have been successfully identified . Among other secured cases was Plesiomonas shigelloides isolated in pig (Sus scrofa domestica), mouse Mus musculus), binturong (Arctictis binturong) and raccoon (Pocyon lotor) . In mandarin duck (Aix galericulata) the strain of Plesiomonas shigelloides with a new 0 antigen-097 was isolated . In regard of the particular biological classes, the isolation of Plesiomonas shigelloides in fish (2.63%) predominate, followed by reptiles (1.66%), mammals (0.94%) and birds (0.79%) . Plesiomonas shigelloides in amphibians was not demonstrated . This work surveys the incidence of Plesiomonas shigelloides and its serovars in animals in the region Moravia in the Czech Republic.

Antonie Van Leeuwenhoek, 1998 Nov, 74(4), 191 - 7
Molecular bases of epithelial cell invasion by Shigella flexneri; Sansonetti PJ et al.; The pathogenesis of shigellosis is characterized by the capacity of the causative microorganism, Shigella, to invade the epithelial cells that compose the mucosal surface of the colon in humans . The invasive process encompasses several steps which can be summarized as follows: entry of bacteria into epithelial cells involves signalling pathways that elicit a macropinocitic event . Upon contact with the cell surface, S . flexneri activates a Mxi/Spa secretory apparatus encoded by two operons comprising about 25 genes located on a large virulence plasmid of 220 kb . Through this specialized secretory apparatus, Ipa invasins are secreted, two of which (IpaB, 62 kDa and IpaC, 42 kDa) form a complex which is itself able to activate entry via its interaction with the host cell membrane . Interaction of this molecular complex with the cell surface elicits major rearrangements of the host cell cytoskeleton, essentially the polymerization of actin filaments that form bundles supporting the membrane projections which achieve bacterial entry . Active recruitment of the protooncogene pp 60c-src has been demonstrated at the entry site with consequent phosphorylation of cortactin . Also, the small GTPase Rho is controlling the cascade of signals that allows elongation of actin filaments from initial nucleation foci underneath the cell membrane . The regulatory signals involved as well as the proteins recruited indicate that Shigella induces the formation of an adherence plaque at the cell surface in order to achieve entry . Once intracellular, the bacterium lyses its phagocytic vacuole, escapes into the cytoplasm and starts moving the inducing polar, directed polymerization of actin on its surface, due to the expression of IcsA, a 120 kDa outer membrane protein, which is localized at one pole of the microorganism, following cleavage by SopA, a plasmid-encoded surface protease . In the context of polarized epithelial cells, bacteria then reach the intermediate junction and engage their components, particularly the cadherins, to form a protrusion which is actively internalized by the adjacent cell . Bacteria then lyse the two membranes, reach the cytoplasmic compartment again, and resume actin-driven movement.

J Virol, 1999 Apr, 73(4), 2863 - 75
Interactions between vaccinia virus IEV membrane proteins and their roles in IEV assembly and actin tail formation; Rottger S et al.; The intracellular enveloped form of vaccinia virus (IEV) induces the formation of actin tails that are strikingly similar to those seen in Listeria and Shigella infections . In contrast to the case for Listeria and Shigella, the vaccinia virus protein(s) responsible for directly initiating actin tail formation remains obscure . However, previous studies with recombinant vaccinia virus strains have suggested that the IEV-specific proteins A33R, A34R, A36R, B5R, and F13L play an undefined role in actin tail formation . In this study we have sought to understand how these proteins, all of which are predicted to have small cytoplasmic domains, are involved in IEV assembly and actin tail formation . Our data reveal that while deletion of A34R, B5R, or F13L resulted in a severe reduction in IEV particle assembly, IEVs formed by the DeltaB5R and DeltaF13L deletion strains, but not DeltaA34R, were still able to induce actin tails . The DeltaA36R deletion strain produced normal amounts of IEV particles, although these were unable to induce actin tails . Using several different approaches, we demonstrated that A36R is a type Ib membrane protein with a large, 195-amino-acid cytoplasmic domain exposed on the surface of IEV particles . Finally, coimmunoprecipitation experiments demonstrated that A36R interacts with A33R and A34R but not with B5R and that B5R forms a complex with A34R but not with A33R or A36R . Using extracts from DeltaA34R- and DeltaA36R-infected cells, we found that the interaction of A36R with A33R and that of A34R with B5R are independent of A34R and A36R, respectively . We conclude from our observations that multiple interactions between IEV membrane proteins exist which have important implications for IEV assembly and actin tail formation . Furthermore, these data suggest that while A34R is involved in IEV assembly and organization, A36R is critical for actin tail formation.

S Afr Med J, 1999 Jan, 89(1), 59 - 63
Laboratory surveillance of Shigella dysenteriae type 1 in KwaZulu-Natal; Karas JA et al.; OBJECTIVE: To collect data on the antimicrobial susceptibility of Shigella dysenteriae type 1 in KwaZulu-Natal, including the testing of newer therapeutic agents, and to evaluate the ability of laboratories to participate in a provincial surveillance programme . DESIGN: Prospective descriptive study . SETTING: Hospital laboratories in KwaZulu-Natal, including peripheral laboratories and the medical microbiology laboratory of the University of Natal . MAIN OUTCOME MEASURES: Antimicrobial susceptibility pattern of surveillance strains and evaluation of the ability of provincial laboratories to isolate Shigella . RESULTS: All 354 strains tested were resistant to ampicillin, chloramphenicol and tetracycline . Co-trimoxazole resistance was found in 92.2% of strains, and 0.8% of strains were resistant to nalidixic acid . All strains were susceptible to ceftriaxone, ciprofloxacin, ofloxacin, pivmecillinam, azithromycin, loracarbef and fosfomycin . Of the 29 laboratories surveyed, 18 (62.1%) were able to isolate and identify S . dysenteriae correctly, and 9 (32%) were able to serotype it further to S . dysenteriae type 1 . Twenty-seven (93.1%) had appropriate culture media and 26 (89.7%) had antisera for Shigella identification . CONCLUSIONS: There is little variation among strains of S . dysenteriae type 1 in KwaZulu-Natal with regard to their antimicrobial susceptibility pattern . Nalidixic acid should remain the antimicrobial of choice for treatment of dysentery in our region as resistance to it is low . The majority of KwaZulu-Natal laboratories have the expertise and equipment to perform the isolation and identification of Shigella species.

Folia Microbiol (Praha), 1998, 43(6), 707 - 12
Characterization of Shigella dysenteriae type 1 capsular polysaccharide by immunochemical methods; Nasher K et al.; We have isolated the capsular polysaccharide from the strain of Shigella dysenteriae type 1 8337 . The product was purified by ultracentrifugation, treated with enzymes (proteinase K, DNA-RNAase) and analyzed by immunochemical methods . Polyclonal antibodies were obtained from rabbits immunized by whole cell antigens prepared from Shigella by ultrasonic treatment and by purified capsular polysaccharide . Crossed immunoelectrophoresis, PAGE and Western blot analysis showed that this product containing mainly the polysaccharide component also contained glycoprotein and lipopolysaccharide . Double diffusion in agarose gel confirmed that the capsular preparation contained at least three antigens reacting with rabbit polyclonal antiserum.

Curr Opin Microbiol, 1998 Feb, 1(1), 116 - 24
Vaccination against enteric pathogens: from science to vaccine trials; Lindberg AA; Recent advances in scientific research and clinical trials have shown promise for vaccine development against enteric pathogens . Identification of new virulence factors, such as the two distinct shigella enterotoxins, has allowed the development of new immunogen or new attenuated strains . Improved knowledge facilitated the development of safer attenuated live microorganism and construction of multivalent vaccines . Finally, an important advantage is the use of nonreplicating plasmid DNA vectors to express protective antigens in the host.

Curr Opin Microbiol, 1999 Feb, 2(1), 51 - 5
Mechanism of Shigella entry into epithelial cells; Nhieu GT et al.; Shigella, the causative agent of bacillary dysentery, invades epithelial cells by reorganizing the cell cytoskeleton during bacterial entry . This entry process requires the Shigella Ipa proteins that are secreted by a type III secretion apparatus and that act in concert to fine tune cell responses . Actin polymerization at the site of entry is dependent on the IpaB and IpaC proteins, whereas IpaA further modulates cytoskeletal rearrangements by binding to vinculin.

Infect Immun, 1999 Mar, 67(3), 1471 - 80
Interleukin-8 controls bacterial transepithelial translocation at the cost of epithelial destruction in experimental shigellosis; Sansonetti PJ et al.; In shigellosis, the network of cellular interactions mediated by a balance of pro- and anti-inflammatory cytokines or chemokines is clearly tipped toward acute destructive inflammation of intestinal tissues by the bacterial invader . This work has addressed the role played by interleukin-8 (IL-8) in a rabbit model of intestinal invasion by Shigella flexneri . IL-8, which is largely produced by the epithelial cells themselves, appears to be a major mediator of the recruitment of polymorphonuclear leukocytes (PMNs) to the subepithelial area and transmigration of these cells through the epithelial lining . Neutralization of IL-8 function by monoclonal antibody WS-4 caused a decrease in the amount of PMNs streaming through the lamina propria and the epithelium, thus significantly attenuating the severity of epithelial lesions in areas of bacterial invasion . These findings are in agreement with our previous work (31) . In contrast to the PMNs, the bacteria displayed increased transepithelial translocation, as well as overgrowth in the lamina propria and increased passage into the mesenteric blood . By mediating eradication of bacteria at their epithelial entry site, although at the cost of severe epithelial destruction, IL-8 therefore appears to be a key chemokine in the control of bacterial translocation.

Infect Immun, 1999 Mar, 67(3), 1455 - 60
Involvement of tumor necrosis factor alpha and interleukin-1beta in enhancement of pentylenetetrazole-induced seizures caused by Shigella dysenteriae; Yuhas Y et al.; Neurologic manifestations, mainly convulsions, are the most frequent extraintestinal complications of shigellosis . We used an animal model to study the roles of tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1beta) in Shigella-related seizures . Administration of Shigella dysenteriae 60R sonicate enhanced the sensitivity of mice to the proconvulsant pentylenetetrazole (PTZ) within 7 h . This was indicated by a significantly higher mean convulsion score and an increased number of mice responding with clonic-tonic seizures in the Shigella-pretreated group . Preinjection of mice with anti-murine TNF-alpha (anti-mTNF-alpha) or anti-murine IL-1beta (anti-mIL-1beta) 30 min prior to administration of Shigella sonicate abolished their enhanced response to PTZ at 7 h . Mean convulsion scores were reduced by anti-mTNF-alpha from 1.2 to 0.8 (P = 0.017) and by anti-mIL-1beta from 1.3 to 0.7 (P = 0.008) . Preinjection of anti-mTNF-alpha also reduced the percentage of mice responding with clonic-tonic seizures, from 48 to 29% (P = 0.002), and preinjection of anti-mIL-1beta reduced it from 53 to 21% (P = 0 . 012) . Neutralization of TNF-alpha or IL-1beta did not protect the mice from death due to S . dysenteriae 60R . These findings indicate that TNF-alpha and IL-1beta play a role in the very early sensitization of the central nervous system to convulsive activity after S . dysenteriae administration . Similar mechanisms may trigger neurologic disturbances in other infectious diseases.

Curr Biol, 1999 Jan 28, 9(2), 89 - 92
Tyrosine phosphorylation is required for actin-based motility of vaccinia but not Listeria or Shigella; Frischknecht F et al.; Studies of the actin-based motility of pathogens have provided important insights into the events occurring at the leading edge of motile cells {1} {2} {3} . To date, several actin-cytoskeleton-associated proteins have been implicated in the motility of Listeria or Shigella: vasodilator-stimulated phosphoprotein (VASP), vinculin and the actin-related protein complex of Arp2 and Arp3 {4} {5} {6} {7} . To further investigate the underlying mechanism of actin-tail assembly, we examined the localization of components of the actin cytoskeleton including Arp3, VASP, vinculin and zyxin during vaccinia, Listeria and Shigella infections . The most striking difference between the systems was that a phosphotyrosine signal was observed only at the site of vaccinia actin-tail assembly . Micro-injection experiments demonstrated that a phosphotyrosine protein plays an important role in vaccinia actin-tail formation . In addition, we observed a phosphotyrosine signal on clathrin-coated vesicles that have associated actin-tail-like structures and on endogenous vesicles in Xenopus egg extracts which are able to nucleate actin tails {8} {9} . Our observations indicate that a host phosphotyrosine protein is required for the nucleation of actin filaments by vaccinia and suggest that this phosphoprotein might be associated with cellular membranes that can nucleate actin.

J Clin Microbiol, 1999 Mar, 37(3), 873 - 4
Identification of Shigella flexneri subserotype 1c in rural Egypt; El-Gendy A et al.; In a population-based study of diarrhea in rural, northern Egypt, 60 Shigella flexneri strains were identified, of which 10 could not be definitively serotyped . Serological analysis with commercial reagents suggested that they were serotype 1, but the strains failed to react with subserotype 1a- or 1b-specific antibodies . All 10 strains reacted with MASF 1c, a monoclonal antibody specific for a provisional S . flexneri subserotype, 1c, first identified in Bangladesh and not previously detected outside of that region . Our results show that S . flexneri subserotype 1c is not unique to Bangladesh and that the inability to detect it may reflect both the limited use of suitable screening methods and the rarity of this subserotype.

J Infect Dis, 1999 Feb, 179 Suppl 1, S259 - 62
Ebola outbreak in Kikwit, Democratic Republic of the Congo: discovery and control measures; Muyembe-Tamfum JJ et al.; The Ebola epidemic in Kikwit, Democratic Republic of the Congo, was recognized because of a nosocomial outbreak in Kikwit General Hospital . Initially, a diagnosis of shigella infection was suspected because many patients presented with bloody diarrhea . On 4 May 1995, blood samples from 14 acutely ill patients were sent to the Centers for Disease Control and Prevention (Atlanta), and on 9 May, a diagnosis of Ebola hemorrhagic fever was confirmed . The major disease control measures that were undertaken were the isolation of patients in a quarantine ward at Kikwit General Hospital, the distribution of protective equipment to health care workers and family members caring for Ebola patients, the use of barrier nursing techniques, the distribution of health education material, active and passive case finding, and the burying of the deceased in plastic bags by a trained team of Red Cross volunteers who wore gloves and protective clothing.

J Trop Pediatr, 1998 Dec, 44(6), 335 - 7
Contamination of weaning foods: organisms, channels, and sequelae; Afifi ZE et al.; A study was carried out to identify the microbial contaminants of weaning foods in an Egyptian village and the sources of contamination . All 300 households containing infants (< 24 months) were visited . Information regarding food preparation and household sanitation was gathered . Samples of weaning foods (270) were collected and analysed for the presence of Escherichia coli, Bacillus cereus, Shigella and parasites . The first two pathogens were detected in 43.7 per cent and 21.4 per cent of samples respectively . The others were not detected in any sample . The risk of contamination by E . coli and B . cereus was significantly associated with the presence of dung and/or refuse in the house, lack of indoor latrine, non-use of latrine by children, weaning foods not freshly prepared, uncovered storage of foods, and the presence of a case of diarrhoea in the house.

J Bacteriol, 1999 Feb, 181(4), 1229 - 37
Impact of either elevated or decreased levels of cytochrome bd expression on Shigella flexneri virulence; Way SS et al.; Shigella spp . are the major cause of bacillary dysentery worldwide . The pathogenic process involves bacterial invasion and lysis of the phagocytic vacuole, followed by replication and movement within the cell cytoplasm and, ultimately, spread directly into adjacent cells . This study demonstrates that S . flexneri cytochrome bd expression is necessary for normal intracellular survival and virulence . Cytochrome bd is one of two terminal oxidases in the bacterial respiratory chain that reduce molecular oxygen to water, utilizing intermediates shuttled through the electron transport chain . S . flexneri mutants that contain a disruption in the cydC locus, which leads to defective cytochrome bd expression, or in the riboflavin (ribE) or ubiquinol-8 (ubiH) biosynthetic pathway, which leads to elevated cytochrome bd expression, were evaluated in intracellular survival and virulence assays . The cydC mutant formed significantly smaller plaques, had significantly decreased intracellular survival, and had a 100-fold increase in lethal dose for mice compared with the wild type . The ribE and ubiH mutants each formed significantly larger plaques and had a 10-fold decrease in lethal dose for mice compared with the wild type . The data indicate that expression of cytochrome bd is required for S . flexneri intracellular survival and virulence.

JAMA, 1999 Jan 27, 281(4), 361 - 7
Access to essential drugs in poor countries: a lost battle?
Pecoul B, Chirac P, Trouiller P, Pinel J.
Drugs offer a simple, cost-effective solution to many health problems, provided they are available, affordable, and properly used . However, effective treatment is lacking in poor countries for many diseases, including African trypanosomiasis, Shigella dysentery, leishmaniasis, tuberculosis, and bacterial meningitis . Treatment may be precluded because no effective drug exists, it is too expensive, or it has been withdrawn from the market . Moreover, research and development in tropical diseases have come to a near standstill . This article focuses on the problems of access to quality drugs for the treatment of diseases that predominantly affect the developing world: (1) poor-quality and counterfeit drugs; (2) lack of availability of essential drugs due to fluctuating production or prohibitive cost; (3) need to develop field-based drug research to determine optimum utilization and remotivate research and development for new drugs for the developing world; and (4) potential consequences of recent World Trade Organization agreements on the availability of old and new drugs . These problems are not independent and unrelated but are a result of the fundamental nature of the pharmaceutical market and the way it is regulated.

Pediatrics . 1999 Feb;103(2):E18.
Central nervous system manifestations of childhood shigellosis: prevalence, risk factors, and outcome; Khan WA et al.; BACKGROUND AND OBJECTIVE: Alterations in consciousness, including seizures, delirium, and coma, are known to occur during Shigella infection . Previous reports have suggested that febrile convulsions and altered consciousness are more common during shigellosis than with other childhood infections . Those reports, however, have been from locations where S dysenteriae type 1 was not common, thus making it difficult to assess the specific contribution that S dysenteriae type 1 infection, and Shiga toxin, might make to the pathogenesis of altered consciousness in children with shigellosis . In this study we seek to determine the prevalence, risk factors, and outcome of altered consciousness in children with shigellosis in Bangladesh, a country where infection with all four species of Shigella is common . We particularly focus on the importance of metabolic abnormalities, which we have previously shown to be a common feature of shigellosis in this population . METHODS: This study was conducted at the Diarrhea Treatment Centre of the International Centre for Diarrhoeal Disease Research, Bangladesh in Dhaka, Bangladesh, which provides care free of charge to persons with diarrhea . During 1 year, a study physician identified all inpatients infected with Shigella by checking the logs of the Clinical Microbiology Laboratory daily . Study physicians obtained demographic and historical information by reviewing the patient charts and by interviewing patients, or their parents or guardians, to confirm or complete the history of illness obtained on admission . Patients were categorized as being conscious or unconscious based on a clinical scale; having a seizure documented in the hospital; or having a seizure by history during the current illness that was not witnessed by medical personnel . Patient outcome was classified as discharged improved, discharged against medical advice, transferred to another health facility, or died in the Treatment Centre . Laboratory examinations were ordered at the discretion of the attending physician; all such information was recorded on the study form . Clinical management was by the attending physician . Factors independently predictive of a documented seizure, or of unconsciousness, were determined using a multiple logistic regression analysis . For this analysis variables associated with unconsciousness or a documented seizure in the analysis of variance or chi2 analyses were entered into the regression equation and eliminated in a backward stepwise fashion if the probability associated with the likelihood ratio statistic exceeded .10 . RESULTS: During this 1-year study, 83 402 persons with diarrhea came to the Treatment Centre for care, and 6290 patients were admitted to the inpatient unit . Shigella was isolated from a stool or rectal swab sample of 863 (13.7%) of the inpatients . Seventy-one (8%) of the inpatients with shigellosis were >/=15 years old; 61 (86%) were conscious; 10 (14%) were unconscious; none had either a documented seizure or a seizure by history during this illness . Seven hundred ninety-two patients were <15 years old (92%); 654 (83%) were conscious; 73 (9%) were unconscious; 41 (5%) had a documented seizure (compared with >/=15-year age group); 24 (3%) had a seizure by history during this illness . Of the 41 patients with documented seizures, 19 (46.3%) had a seizure at the time of admission, and 22 (53.7%) had a seizure after admission . Twenty-five (61.0%) of the 41 patients with documented seizures were reported to have a seizure during this illness before coming to the Treatment Centre . Clinical features that are known to cause altered consciousness-fever, severe dehydration, hypoglycemia, hyponatremia, or meningitis-were present in 38 (92.7%) of the 41 patients in whom a seizure was witnessed and in 67 (91.8%) of the 73 patients who were unconscious . Nineteen (46 . 3%) of the patients who had a seizure documented had two of these five features, 4 (9.8%) had three, and 1 (2 . (ABSTRACT TRUNCATED)

Ann Trop Paediatr, 1998 Sep, 18(3), 197 - 201
Shigellosis in children: a clinico-epidemiological comparison between Shigella dysenteriae type I and Shigella flexneri; Faruque AS et al.; We reviewed the clinical and epidemiological features of 390 children under 5 years of age infected with either Shigella dysenteriae type I or Shigella flexneri attending a diarrhoea treatment centre from 1993 to 1995 in Dhaka, Bangladesh . Older age (24 months or more), underweight and wasting but not stunting were the host factors significantly more associated with Shigella dysenteriae type I infection than in Shigella flexneri-infected children . Moreover, use of antibiotics at home, use of water from tubewells or pipe-water for drinking and lack of sanitary facilities for defaecation were the behavioural and environmental factors strongly associated with S . dysenteriae type I infection . Children with diarrhoea due to S . flexneri presented with more watery/liquid stools and had a shorter duration of illness . Duration of diarrhoea for 4 or more days was typical of S . dysenteriae type I infection . Interventions to address malnutrition and to promote environmental hygiene would be predicted to offer greater protection against shigellosis due to S . dysenteriae than S . flexneri.

J Bacteriol, 1999 Feb, 181(3), 869 - 78
Functional analysis of a rickettsial OmpA homology domain of Shigella flexneri icsA; Charles M et al.; Shigella flexneri is a gram-negative bacterium that causes diarrhea and dysentery by invasion and spread through the colonic epithelium . Bacteria spread by assembling actin and other cytoskeletal proteins of the host into "actin tails" at the bacterial pole; actin tail assembly provides the force required to move bacteria through the cell cytoplasm and into adjacent cells . The 120-kDa S . flexneri outer membrane protein IcsA is essential for actin assembly . IcsA is anchored in the outer membrane by a carboxy-terminal domain (the beta domain), such that the amino-terminal 706 amino acid residues (the alpha domain) are exposed on the exterior of the bacillus . The alpha domain is therefore likely to contain the domains that are important to interactions with host factors . We identify and characterize a domain of IcsA within the alpha domain that bears significant sequence similarity to two repeated domains of rickettsial OmpA, which has been implicated in rickettsial actin tail formation . Strains of S . flexneri and Escherichia coli that carry derivatives of IcsA containing deletions within this domain display loss of actin recruitment and increased accessibility to IcsA-specific antibody on the surface of intracytoplasmic bacteria . However, site-directed mutagenesis of charged residues within this domain results in actin assembly that is indistinguishable from that of the wild type, and in vitro competition of a polypeptide of this domain fused to glutathione S-transferase did not alter the motility of the wild-type construct . Taken together, our data suggest that the rickettsial homology domain of IcsA is required for the proper conformation of IcsA and that its disruption leads to loss of interactions of other IcsA domains within the amino terminus with host cytoskeletal proteins.

Biochim Biophys Acta, 1998 Dec 8, 1429(1), 45 - 56
Protein-protein interactions in the assembly of Shigella flexneri invasion plasmid antigens IpaB and IpaC into protein complexes; Davis R et al.; Shigella flexneri is a facultative intracellular bacterial pathogen that invades human colonic epithelial cells by a process called pathogen-induced phagocytosis . Pathogen entry requires three virulence plasmid-encoded proteins called invasion plasmid antigens (Ipa) B, C and D which are secreted upon bacterial contact with a host cell . Following their secretion, IpaB and IpaC are found within a complex of proteins that may also contain IpaA and IpaD . Previous work has shown that exogenously added recombinant IpaC is sufficient for promoting the uptake of S . flexneri in gentamicin-protection assays . It is shown here that purified recombinant Ipa proteins can also be used to investigate the formation of Ipa protein complexes in vitro . The protein-protein contacts involved in the formation of Ipa complexes appear to include previously undescribed IpaC-IpaC interactions in addition to a strong association between IpaB and IpaC . IpaD does not appear to interact with either IpaB or IpaC in vitro although it is possible that recombinant IpaD forms homodimers that are stabilized by disulfide bridges involving this protein's single cysteine residue . This investigation represents the first characterization of the biochemistry of Ipa complex assembly.

J Med Microbiol, 1999 Jan, 48(1), 11 - 6
Antibodies to Shiga toxin in the serum of children with Shigella-associated haemolytic uraemic syndrome; Azim T et al.; Antibodies to Shiga toxin (Stx) were measured in the sera of 49 children with Shigella dysenteriae serotype 1 infection, of whom 17 had haemolytic uraemic syndrome (HUS) and 32 had no complications (uncomplicated shigellosis, UCS) . Children with HUS had lower levels of total IgG and IgM and lower IgM titres to Stx than those with UCS . The number of children with neutralising antibodies was similar in the two groups . Of the children with HUS, 11 had HUS on enrolment and six developed HUS subsequent to enrolment . Antibody titres in children who subsequently developed HUS were compared with those in children with UCS to assess whether differences in antibody titres occurred before the development of HUS . IgA titres to Stx were found to be higher in children who subsequently developed HUS than in those with UCS . However, logistic regression analysis revealed that titres of Stx antibodies in the serum were not significant risk factors for the development of HUS . Thus, although the levels of Stx antibodies were different in children with HUS, and higher IgA titres to Stx were identifiable in children who subsequently developed HUS compared with those with UCS, the relevance of these findings in the development of HUS remains to be elucidated.

Infect Immun, 1999 Feb, 67(2), 782 - 8
Strategy for cross-protection among Shigella flexneri serotypes; Noriega FR et al.; Based upon the lipopolysaccharide (LPS) structure and antigenicity of Shigella group B, a strategy for broad cross-protection against 14 Shigella flexneri serotypes was designed . This strategy involves the use of two S . flexneri serotypes (2a and 3a), which together bear the all of the major antigenic group factors of this group . The novel attenuated strains used in these studies were S . flexneri 2a strain CVD 1207 (DeltaguaB-A DeltavirG Deltaset1 Deltasen) and S . flexneri 3a strain CVD 1211 (DeltaguaB-A DeltavirG Deltasen) . Guinea pigs were immunized with an equal mixture of these strains and later challenged (Sereny test) with a wild-type S . flexneri serotype 1a, 1b, 2b, 4b, 5b, Y, or 6 strain of demonstrated virulence in the same model . Guinea pigs that were immunized with these two vaccine strains produced serum and mucosal antibodies that cross-reacted with all the S . flexneri serotypes tested (except of S . flexneri serotype 6) as assessed by enzyme-linked immunosorbent assay, immunoblotting, and slide agglutination . Furthermore, the combination vaccine conferred significant protection against challenge with S . flexneri serotypes 1b, 2b, 5b, and Y but not with serotypes 1a, 4b, or (as predicted) 6.

Indian J Pathol Microbiol, 1998 Oct, 41(4), 453 - 6
Shigella and the fluorinated quinolones; Ballal M et al.; Between July 1996 to June 1997 a total of 531 Stool samples from cases of acute diarrhoea, dysentry, colitis admitted to Kasturba Medical College Hospital, Manipal were investigated . Shigella were isolated from 29 samples (5.46%) . Out of them, Shigella flexneri were 16 (55.17%), Shigella boydii 8 (27.58%), Shigella dysentriae 3 (10.34%) and Shigella sonnei 2 (6.89%) . Five strains of Shigella flexneri and one strain of Shigella dysentriae were found to show resistance to Nalidixic acid and the newer fluoroquinolones viz., Ciprofloxacin, Norfloxacin and Ofloxacin . The in vitro 'R' factor transfer study showed that the resistance was not transferable.

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