Anti-Shigellosis Activity of Cola anomala Water/Ethanol Pods Extract on Shigella flexneri-Induced Diarrhea in Rats.

This study was undertaken to evaluate the activities of water/ethanol Cola anomala pods extract. In vitro antimicrobial susceptibility was determined by the disk diffusion method; the minimum inhibitory concentration and minimum bactericidal concentration were determined by agar dilution technique. In vivo, shigellosis was induced in healthy Wistar albino rats by oral administration of Shigella flexneri inoculum, 12 × 108 CFU/mL. At the onset of diarrhea, infected and normal control animals were subdivided into various groups treated with distilled water, with water/ethanol Cola anomala pods extract at 25, 50, or 100 mg/kg, or with ciprofloxacin, 2.5 mg/kg. After one-week treatment, rats were sacrificed, and blood and colon were collected. Blood was used for blood cell count. A portion of the colon served for histological studies while homogenate from the remaining part was centrifuged and the supernatant was collected for the determination of NO, PGE2, IL-1ß, and TNF-α levels. In vitro, water/ethanol Cola anomala pods extract showed to be bactericidal, with a minimum inhibitory concentration of 2.0 mg/mL and a minimum bactericidal concentration of 3.0 mg/mL. In diarrheic rats, the extract significantly (P < 0.01) increased the white blood cells and significantly (P < 0.01) decreased stool Shigella density from the first to the seventh day of treatment. It partially restored the structure of eroded intestine epithelium and prevented weight loss; the dose dependently and significantly (P < 0.001) decreased NO, IL-1ß, and TNF-α production in the colon and was found to have no significant effect on PGE2 production. These results support the use of this plant in traditional medicine in the treatment of gastrointestinal ailments.

A murine model of diarrhea, growth impairment and metabolic disturbances with Shigella flexneri infection and the role of zinc deficiency.

Shigella is one of the major enteric pathogens worldwide. We present a murine model of S. flexneri infection and investigate the role of zinc deficiency (ZD). C57BL/6 mice fed either standard chow (HC) or ZD diets were pretreated with an antibiotic cocktail and received S. flexneri strain 2457T orally. Antibiotic pre-treated ZD mice showed higher S. flexneri colonization than non-treated mice. ZD mice showed persistent colonization for at least 50 days post-infection (pi). S. flexneri-infected mice showed significant weight loss, diarrhea and increased levels of fecal MPO and LCN in both HC and ZD fed mice. S. flexneri preferentially colonized the colon, caused epithelial disruption and inflammatory cell infiltrate, and promoted cytokine production which correlated with weight loss and histopathological changes. Infection with S. flexneri ΔmxiG (critical for type 3 secretion system) did not cause weight loss or diarrhea, and had decreased stool shedding duration and tissue burden. Several biochemical changes related to energy, inflammation and gut-microbial metabolism were observed. Zinc supplementation increased weight gains and reduced intestinal inflammation and stool shedding in ZD infected mice. In conclusion, young antibiotic-treated mice provide a new model of oral S. flexneri infection, with ZD promoting prolonged infection outcomes.

Azithromycin Resistance in Shigella spp. in Southeast Asia.

Infection by Shigella spp. is a common cause of dysentery in Southeast Asia. Antimicrobials are thought to be beneficial for treatment; however, antimicrobial resistance in Shigella spp. is becoming widespread. We aimed to assess the frequency and mechanisms associated with decreased susceptibility to azithromycin in Southeast Asian Shigella isolates and use these data to assess appropriate susceptibility breakpoints. Shigella isolates recovered in Vietnam and Laos were screened for susceptibility to azithromycin (15 µg) by disc diffusion and MIC. Phenotypic resistance was confirmed by PCR amplification of macrolide resistance loci. We compared the genetic relationships and plasmid contents of azithromycin-resistant Shigella sonnei isolates using whole-genome sequences. From 475 available Shigella spp. isolated in Vietnam and Laos between 1994 and 2012, 6/181 S. flexneri isolates (3.3%, MIC ≥ 16 g/liter) and 16/294 S. sonnei isolates (5.4%, MIC ≥ 32 g/liter) were phenotypically resistant to azithromycin. PCR amplification confirmed a resistance mechanism in 22/475 (4.6%) isolates (mphA in 19 isolates and ermB in 3 isolates). The susceptibility data demonstrated the acceptability of the S. flexneri (MIC ≥ 16 g/liter, zone diameter ≤ 15 mm) and S. sonnei (MIC ≥ 32 g/liter, zone diameter ≤ 11 mm) breakpoints with a <3% discrepancy. Phylogenetic analysis demonstrated that decreased susceptibility has arisen sporadically in Vietnamese S. sonnei isolates on at least seven occasions between 2000 and 2009 but failed to become established. While the proposed susceptibility breakpoints may allow better recognition of resistant isolates, additional studies are required to assess the impact on the clinical outcome. The potential emergence of azithromycin resistance highlights the need for alternative options for management of Shigella infections in countries where Shigella is endemic.

Newer insights into the mechanism of action of Psidium guajava L. leaves in infectious diarrhoea.

BACKGROUND: Psidium guajava L., Myrtaceae, is used widely in traditional medicine for the treatment of diarrhoea, dysentery, gastroenteritis, stomachaches, and indigestion. However, the effect of the leaf extract of P. guajava on the pathogenesis of infectious diarrhoea has not been studied. The present study evaluates the effect of a hot aqueous extract (decoction) of dried leaves of P. guajava on parameters associated with pathogenicity of infectious diarrhoea. The aim was to understand its possible mechanism(s) of action in controlling infectious diarrhoea and compare it with quercetin, one of the most reported active constituents of P. guajava with antidiarrhoeal activity. METHODS: The crude decoction and quercetin were studied for their antibacterial activity and effect on virulence features of common diarrhoeal pathogens viz. colonization of epithelial cells and production and action of enterotoxins. Colonization as measured by adherence of enteropathogenic Escherichia coli (EPEC) and invasion of enteroinvasive E. coli (EIEC) and Shigella flexneri was assessed using HEp-2 cell line. The production of E. coli heat labile toxin (LT) and cholera toxin (CT) and their binding to ganglioside monosialic acid (GM1) were studied by GM1-ELISA whereas the production and action of E. coli heat stable toxin (ST) was assessed by suckling mouse assay. RESULTS: The decoction of P. guajava showed antibacterial activity towards S. flexneri and Vibrio cholerae. It decreased production of both LT and CT and their binding to GM1. However, it had no effect on production and action of ST. The decoction also inhibited the adherence of EPEC and invasion by both EIEC and S. flexneri to HEp-2 cells. Quercetin, on the other hand, had no antibacterial activity at the concentrations used nor did it affect any of the enterotoxins. Although it did not affect adherence of EPEC, it inhibited the invasion of both EIEC and S. flexneri to HEp-2 cells. CONCLUSION: Collectively, the results indicate that the decoction of P. guajava leaves is an effective antidiarrhoeal agent and that the entire spectrum of its antidiarrhoeal activity is not due to quercetin alone.

Surveillance of antibiotic susceptibility pattern among shigella flexneri strain isolated in Nagpur district during three years period, January 2009-January 2012.

OBJECTIVE: In the few cases of childhood dirrhea that require the antimicrobial therapy, the correct choice of the drug depends on detailed previous knowledge of local strains and pattern of antimicrobial resistance. Shigellosis is one of the most improtant examples of this group of intestinal infections. In order to establish such parameters in Nagpur city, this study was carried out to determine the antimcrobial resistance profile of Shigella flexneri isolated from patients suffering from diahhrea admitted to Various hoapitals in Nagpur district, India. MATERIALS AND METHODS: The study included 110 stool samples collected from patients during the 3 year period. All the isolates were characterized and confirmed by VITEK® 2 GN ID cards and antimicrobial susceptibility was tested by VITEK® 2 AST test cards. RESULTS: We received 73 positive cultures of S. flexneri out of 110 stool samples during three year periods of January 2009 to January 2012. S. flexneri strains presented a high resistance rate to Ampicillin (100%), Chloramphenicol (76.71%), Trimethoprime-sulfamethaxazole (TMP-SMZ) (68.49%) and low resistance to third- and fourth-generation Cephalosporin. None of the isolates was found to be resistant to Ciprofloxacin (MIC ≥ 4), Norfloxacin (MIC ≥12), and Nalidixic acid (MIC ≥30). CONCLUSION: Our results provide data on antimicrobial resistance to choose a proper antibiotic for the treatment of Shigellosis in our country. According to current findings, Quinolones and Cephalosporins are the drug of choice for the diarrheic patients. In conclusion, systematic monitoring is needed to identify changes in the antimicrobial resistance.

Chronic vulvovaginitis caused by antibiotic-resistant Shigella flexneri in a prepubertal child.

A 7-year 8-month-old girl was diagnosed with a prolonged course of vulvovaginitis caused by Shigella flexneri. The child was symptomatic with intermittent vaginal bleeding, dysuria and foul smelling vaginal discharge for a 3-year period. Initial attempts to resolve the infection with successive courses of antibiotic therapy using ampicillin, trimethoprim-sulfamethoxazole, cefixime and amoxicillin/clavulanic acid failed. The child's infection was finally resolved by a 14-day course of ciprofloxacin.

The mechanism of cell death in Listeria monocytogenes-infected murine macrophages is distinct from apoptosis.

Various pathogenic bacteria with the capacity to live within eukaryotic cells activate an apoptotic program in infected host cells. Induction of apoptosis by Listeria monocytogenes in murine dendritic cells and hepatocytes has been described. Here we address the questions of whether and how the pathogen kills macrophages, its most important habitat. Employing several complementary techniques aimed at discriminating between apoptosis and necrosis, we show that murine bone marrow-derived macrophages (BMM) undergo delayed necrosis but not apoptosis when infected with listeriolysin (Hly)-producing L. monocytogenes. This pathogen failed to elicit apoptotic morphology, DNA fragmentation, and surface annexin V binding of macrophages, in contrast to Shigella flexneri infection or gliotoxin treatment, which were used as positive controls. Furthermore, macrophages infected with L. monocytogenes released lower quantities of interleukin-1beta (IL-1beta) than did Shigella flexneri-infected ones, indicating diminished or even absent activation of IL-1-converting enzyme in macrophages harboring L. monocytogenes. We conclude that murine BMM die by necrosis after several hours of cytoplasmic replication of L. monocytogenes. The pathogen may benefit from this feature by the possibility of taking advantage of cells of "pseudo-healthy" appearance, thus avoiding rapid elimination by other phagocytes.

[A morphological evaluation of the protective properties of noninvasive recombinant strains of Shigella]. / Morfologicheskaia otsenka protektivnosti neinvazivnykh rekombinantnykh shtammov shigell.

In 2-3 weeks after the oral immunization of rabbits, made in one or two administrations, with attenuated two-marker S. dysenteriae 1 strain VS-12 and recombinant S. dysenteriae VS-12/S. sonnei NR-18 and S. flexneri y433/S. sonnei NR-18 pronounced immunological reaction developed in the mucous membrane of the small intestine: blast transformation follicles of Peyer's patches, an increase in the number of lymphoblasts and plasmocytes in the cupolae of follicles and in intestinal villi, and an increase in the number of lymphocytes and macrophages in the intestinal epithelium with their release into the lumen of the intestine after challenge with virulent shigellae. The protective potency of these recombinants after challenge with massive doses of virulent shigellae was found to be high, which was shown by quantitative evaluation of the decrease of adhesion, invasiveness and cytotoxicity, suppression of epithelial lesions and development of inflammation in the intestinal mucosa.

[Enteric dysentery vaccines and their effectiveness in a rabbit intestinal loop model. I. The residual virulence of attenuated vaccinal strains of Shigella administered into the intestinal loops]. / Izuchenie énteral'nykh dizenteriinykh vaktsin i ikh éffektivnosti na modeli kishechnykh petel' krolika. Soobshchenie I. Ostatochnaia virulentnost' attenuirovannykh vaktsinnykh shtammov shigell pri vvedenii v kishechnye petli.

Virulent Sh. flexneri strain 2a, Sh. sonnei strain, attenuated Sh. flexneri vaccine strain 2a 516M, and Sh. sonnei vaccine strain 6S (isolated by Yu. A. Belaya), as well as streptomycin-dependent Sh. flexneri strain 2a 1605/3 (isolated by V. V. Sergeev) were introduced into the ligated loops of the rabbit ileum. The use of light and immunofluorescent microscopy, the measurement of the volume of the fluid in the intestinal loops and the quantitative inoculation of their contents resulted in revealing the differences in the properties of the virulent and vaccine strains. The vaccine strains, in contrast to the virulent strains, did not proliferate in the lumen and did not cause the accumulation of fluid in the intestinal loops. They retained sharply limited, especially in the streptomycin-dependent bacteria, ability to penetrate into enterocytes and, via their cytoplasm, into the basement membrane, but lost their ability to proliferate in the cytoplasm of enterocytes (and probably even deteriorated there) and to cause plurulent ulcerous inflammation. This indicates that vaccine strains have insignificant residual virulence and suggests that the intestinal loop models, together with other models, may be used for testing the safety of vaccines prepared from Shigella strains.