Atypical Yersinia virulence markers isolated from children with diarrhea

Background and Objective: Yersinia is a gram-negative bacillus that cause diarrhea through consumption of contaminated food and water. This study was performed to identify the atypical Yersinia virulence markers isolated from children with diarrhea

Methods: This descriptive cross -sectional study was done on 384 fecal samples of 0- 14 years old children admitted at children medical center from August 2011 to August of 2012. Fecal samples, for the enrichment, after 21 days of incubation in alkaline buffer with pH=7.2 at 4degree C, on days 7,14 and 21 samples were cultured on CIN agar and Mac agar and then confirm the differentiation atypical Yersinia from other typical Yersinia species from fermentation of different sugars. Isolates were tested for marker of virulence including calcium dependence, auto agglutination, Congo red uptake and binding of crystal violet

Results: Out of 384 stool samples, 4 [1.04%] were infected with Yersinia [Yersinia frederikseni, Yersinia kristensenii and Yersinia enterocolitica]

Out of these three, only two samples in association was positive with virulence markers

Conclusion: Phenotypic markers can be used to study the properties of phenotypic strains of Yersinia

Evaluation of anti-microbial activity of Lactobacillus acidophillus and Lactobacillus ruteri against entero-pathoges by in vitro and in vivo methods

Background and Objective: Probiotics are beneficial organisms therapeutic within microbial flora. Shigella, Escherichia coli and Salmonella are the most common cause of intestinal infectious diseases that lead to morbidity and mortality in infant and children worldwide. The aim of this study was to evaluate anti-microbial activity of Lactobacillus acidophillus and Lactobacillus ruteri against entero-pathoges by in vitro and in vivo methods

Methods: In this experimental study, the therapeutic effect of the lactobacillus acidophilus ATCC 4356 and ruteri ATCC 23272 against Shigella sonnei ATCC 9290, Escherichia coli ATCC 25922 and Salmonella enterica BAA-708 were evaluated by in vitro [spot agar] and in vivo [BALB/c mice] methods. Weight improvment and survival rate in mice were recorded

Results: Lactobacillus acidophillus and ruteri had protective and therapeutic effect against diarrhea caused by pathogenic bacteria. Probiotics reduced the weight, colonization of pathogens and increased the survival rate of animals [P<0.05]

Conclusion: Lactobacillus acidophillus and ruteri has anti-microbial activity and their consumption can be effective in the prevention and also the treatment of intestinal disease

[Antimicrobial effect of zataria multiflora and rosemarinus officinalis on antibiotic-resistant staphylococcus aureus strains isolated from food]

Staphylococcus aureuse is one of the important pathogens which transmitted by food and has majority of habitant in human and animal community as a pathogen and normal flora. Antibiotic resistant among Staphylococcus aureuse strains is a global health challenge. Regarding to the different therapeutic and antimicrobial effect of Shirazi Zataria multiflora and Rosemarinus officonalis in present work the antibacterial effect of this extract and its synergistic effect with routine antibiotics was investigated. In this in vitro study the antimicrobial effect of Shirazi Zataria multiflora and Rosemarinus officonalis extract on methicillin resistant Staphylococcus aureus [MRSA] and other antibiotic resistant strains to tetracycline, erithromycine, trimethoprim, sulfamethoxazol, together with its MIC and MBC were determined. Also synergistic effect of these extracts with these antibiotics was investigated by paper disc method. Shirazi zataria multiflora, had a significant antibacterial effect against MRSA, and other Staphylococcus aureus resistant strain to tetracycline, erithromycine, trimethoprim, and sulfametoxazol, isolated from food. Production of a suitable herbal medicine with few side effects will give rise to a promising outlook in the treatment of infections caused by antibiotic resistant strains of Staphylococcus aureus

[Pattern of serotyping and antibiotic resistance of Salmonella in children with diarrhea]

Gastroenteritis due to Salmonella is common in human and considered as a global dilemma of public health. This study was done to determine the Pattern of serotyping and antibiotic resistance of Salmonella in children with diarrhea in Iran. In this laboratory study, 306 stool samples were collected from children with diarrhea in public health centers in Robat-karim, Tehran province, Iran. The specimens were enriched in Selenite F medium and then cultured on Hekton agar. The identification of Salmonella was carried out by conventional method and antimicrobial susceptibility testing was performed according to CLSI procedures. Out of 306 stool samples, 7.2 % were identified as Salmonella species, as follow: 7 Salmonella typhi, 6 Salmonella paratyphi B, 3 Salmonella paratyphi C, 2 Salmonella paratyphi A and 4 samples were not identifiable. There was a significant relation between presence of WBC in fecal and salmonellosis [P<0.05]. In drug sensitivity trends, 92.3% of Salmonella species were sensitive to chloramphenicol, ceftizoxime, Nalidixic acid and Amikacin. This study showed that Salmonella was the cause of children diarrhea in 7.2% in this region

[Efficacy of Ciprofloxacin, ceftizoxims and carbenicillin on klebsiella species isolated from hospital specimens]

Klebsiella species are gram-negative bacteria with positive voges proskauer [VP] reaction. Klebsiella species are found as commensal in human digestive and respiratory system. This group of organisms can create a serious health hazards in hospitalized patients, and their ability to drug resistance is a major health problems. This study was done to evaluate the efficacy of Ciprofloxacin, Ceftizoxims and Carbenicillin on Klebsiella species isolated from hospital specimens. In this laboratory study, 1200 clinical samples were isolated from patients in Imam Khomeini hospital, Tehran, Iran. The identification Klebsiella species were carried out according to conventional biochemical tests. The minimum inhibitory concentration [MIC] of carbenicillin, ceftizoxime, and ciprofloxacin antibiotics were determined using Macrodilution broth test. Out of 1200 isolated samples, 25% were identified as Klebsiella species. 73% of identified Klebsiella were obtained from urine samples. Klebsiella.peumoniae with rate of 94% was the most abundant among other species. The results of MIC and minimum bactericidal concentration by using standard microdilution method showed drug resistance range of 16-1024 micro g/ml, 4-256 microg/ml and 0.25-16 micro g/ml for carbenicillin, ceftizoxime, and ciprofloxacin, respectivley. In general, 94%, 6% and 1% of species were resistance to carbenicillin, ceftizoxime and ciprofloxacin, respectively. Ciprofloxacin and Ceftizoxime are suitable for the treatment of infections due to Klebsiella species

[ trend of drug resistance in toxin and non-toxin producing Shigella Spp. isolated from stool of children with diarrhea]

Diarrhea is recognized as one of the main causes of morbidity and mortality in developing countries. Gastrointestinal diseases can lead to death of many children of less than 5 years of age. The aim of this study was to evaluate the drug resistance pattern in Shigella toxin and non-toxin producing strains in children. In this descriptive analytic study a total of 80 Shigella strains, 60 strains isolated from stool samples of children with diarrhea from Loghman, Emam and Tebi Koodakan Centre Hospitals, and 20 national collection strains isolated and reserved during the last years. The isolates were evaluated for cytotoxin production by using cell culture technique [Hela cell]. Our study included 54 strains of S. flexneri, 14 strains of S.sonnei, 10 strains of S. boydii and 2 strains of S. dysenteriae. Data were analyazed by means of chi-square and Fisher's exact test. Of 80 strains 9 [11.25%] showed cytotoxic effect. Chi-square test showed no significant difference between the isolated and national collection strains [P >/= 0.05].There was no correlation between the cytotoxic activity and clinical symptoms such as abdominal pain, nausea, and frequency of passing stools / day, but other symptoms like fever and presence of blood in the stool had correlation with cytotoxin production. The results of this study showed that there was no significant difference in the antimicrobial resistance pattern between toxin and non-toxin producing Shigella strains isolated from the clinical samples and the standard national collection

[Evaluation of antibacterial properties of human amniotic membrane in vitro]

Amniotic membrane is the inner-most layer of the three fetal membranes. The membrane is consisted of three layers; epithelial layer, basal membrane, and connective tissue. Owing to expression of mRNA of elafin, HBD 1-3, and secretory leukocyte protease inhibitors, amniotic membrane has antimicrobial properties. The aim of the current study was to evaluate the in vitro antibacterial effect of human amniotic membrane on standard bacterial species of Salmonella enterica BAA-708, E.coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Klebsiella pneumoniae ATCC 7881, and Enterococcus faecalis ATCC 29212. Fresh amniotic membranes were obtained from Organ Transplant Bank of Imam Khomeini Hospital in Tehran. The membranes were obtained from pregnant women who had negative tests for HIV, HBV, HCV, and syphilis after elective Cesarian section. The membranes were cut into 1.5× 1.5 cm pieces under sterile conditions. The membrane pieces were placed on Müller-Hinton agar medium containing the bacterial suspensions and then incubated at 37 [degree sign]C for 24 hours. The antibacterial properties of amniotic membrane against Salmonella enterica and E. coli were demonstrated by development of the no growth halo, but for Pseudomonas aeruginosa only a very narrow halo was observed. The halo was not developed for Klebsiella pneumoniae and Enterococcus faecalis. Amniotic membrane showed antibacterial effects against a wide spectrum of bacteria. With regard to the increasing antibiotic resistance, use of amniotic membrane against pathogenic bacteria can be considered valuable

Evaluation of culture and pcr methods for diagnosis of group b streptococcus carriage in Iranian pregnant women

Group B streptococcus [GBS] is one of the most important cause of morbidity and mortality among newborns especially in developing countries. It has been shown that the screening approach rather than the identification of maternal clinical risk factors for early-onset neonatal GBS disease is more effective in preventing early-onset GBS neonatal disease. The objective of this study was to detect GBS among clinical samples of women using PCR and standard microbiological culture. Samples were taken from 375 women at 28-38 weeks of gestation during six month from January 15 till June 15, 2011 from a hospital in Tehran, Iran. Samples were tested by standard culture using Todd- Hewitt broth, blood agar and by PCR targeting the cfb gene. Among the 375 women, 35 [9.3%] were identified as carriers of group B streptococci on the basis of the results of the cultures of specimens, compared to 42 [11.2%] on the basis of PCR assay. We found that GBS can be detected rapidly and reliably by a PCR assay in vaginal secretions from women at the time of delivery. This study also showed that the rate of incidence of GBS is high in Iranian women

frequency of extended spectrum beta lactamase and CTX M-I of Escherichia coli isolated from the urine tract infection of patients by phenotypic and PCR methods in the city of Khoy in Iran

The production of Extended Spectrum Beta Lactamases [ESBLs] by Escherichia coli is the main cause of resistance to Cephalosporins. In the past decade, CTX-M enzymes have become the most prevalent ESBLs in Europe, Canada, and Asia. In this study, the frequency of ESBL- producing E.coli and molecular detection of the CTX-M-I group was investigated. A total of 400 urine samples were collected from both hospitalized and out-patients in Khoy's hospitals between November 2009 and April 2010. Out of these samples, 188 were identified as E.coli by standard biochemical tests. The antibiotic Susceptibility tests to 10 antibiotics were performed by the-disk-agar diffusion [DAD] method. ESBL production was screened by phenotypic test that including disk diffusion agar and combined disk as recommended by the Clinical and Laboratory Standards Institute [CLSI] Screened isolates were investigated by PCR assay for detection of CTX-M-I group genes. The results show that out of 188 E.coli isolates identified, 56 [29.8%] were producing ESBls by phenotypic test. All isolates were sensitive to imipenem. Overall, 49 [87.5%] isolates were confirmed as CTX-M-I producer by PCR. The results of this study showed that about 30% of the identified E.coli were producing ESBL Therefore, we recommend to use molecular methods in such researches

[Synergistic effect polymyxin B sulphate and trimethoprim on yersinia enterocolotica and closely related species]

Previous studies have shown that polymyxin B sulfate and trimthoprim antibiotics are not individually effective on Yersinia enterocolitica and their closely related species. The aim of this study was to evaluate the synergistic effect of above antibiotics on Y. enterocolitica and their closely related species, from the clinical and the natural environment specimen collected in Iran, and compare them with the isolates that that were obtained from the Pasteur institute collection in France. In total, 73 species from Iran and 25 from the Pasteur institute in France were tested. The microdilution method was used for the MIC according to the standard protocol. The synergistic effect was seen in all tested samples. However, the human species from the Pasteur institute were more sensitive than the Iranian human and the environmental species were less sensitive than clinical specimens [1.6+16 micro gr, 0.4+4 micro gr in French Samples]. The Y. enterocolitica isolates were less sensitive than the related species such as Y. intermedia, Y. fredriksenii, and Y. kristensenii. The synergistic effect polymyxin B sulfate and trimthoprim were more evident on other closely related Yersinia species Y. enterocolitica

Antimicrobial effect of Zataria multiflora on antibiotic-resistant Staphylococcus aureus strains isolated from food

Methicillin -resistant strains of Staphylococcus aureus [MRSA] have created a major problem in the treatment of diseases. MRSA colonization in nose can lead to increased rate of nosocomial infections and mortality. Zataria multiflora is a plant which is widely used in the world for medical purposes. The aim of this study was to evaluate the in vitro antimicrobial effect of zataria multiflora extract on MRSA strains isolated from food. In this in vitro study the antimicrobial effect of Shirazi Zataria multiflora extract on MRSA and other staphylococci aureus strains resistant to tetracycline,erithromycine,trimethoprim,sulfamethoxazol,and methiclline, together with its MIC and MBC were determined. Shirazi zataria multiflora, had a significant affects against MRSA, and other Staphylococcus aureus resistant to methicillin, tetracycline,erithromycine,trimethoprim,and sulfametoxazol ,isolated from food. Production of a suitable herbal medicine with few side effects will give rise to a promising out-look in the treatment of infections caused by Staphylococcus aureus

Identification of P fimbriae virulence factor in uropathogenic Escherichia coli by PCR in Shaherkord Hospitals

Urinary tract infection is one of the most common bacterial diseases in human being.The most important etiologic agent is Eschrichia coli and the most important virulence factor in uropathogenic E.coli is P fimberiae. Construction of P fimbriae is coded by pap operon. The importance of P fimbriae is related to the incidence of urinary tract infection, especially infection of the upper tracts. The aim of this study was to identify and determine P fimbriae prevalence in isolated uroparhogenic E.coli. A total of 182 isolates of E.coli were evaluated .Bacterial DNA was extracted by boiling method and subsequently DNA samples were examined for pap operon [pap C gene] by use of PCR method. Then PCR products were stained and identified by polyacrylamid gel electrophoresis. Based on the results of PCR, among 182 isolated strains, 66 samples [36.3%] had pap operon[pap C gene]. The results indicated that prevalence of pap operon in isolated strains was within the global range. Pyelonephritis was more common than cystitis and asympthomatic bacteriuria

[Antibacterial effects of Rosmarinus officinalis on methicillin - resistant Staphylococcus aureus isolated from patients and foods]

Methicillin-resistant Staphylcoccus aureus [MRSA] is considered a major problem in the world. This strain colonizes nose and causes increased incidence of nosocomial infections, mortality and morbidity. Rosemary [Rosmarinus officinalis] is a herbal medicine widely used all over the world. The aim of this study was to evaluate the in vitro antimicrobial activity of rosemary essence on MRSA isolated from patients and food. 200 strains of MRSA, 100 from patients and 100 from food samples, were collected and analyzed in Tehran, during the last year. 28 MRSA strains and multi drug resistant [MDR] strains were isolated. The antimicrobial activity of the rosemary essence against different isolates of the microorganism was evaluated by disk diffusion and macro broth dilution methods. MRSA isolates belonged to 25% and 60% of food and clinical samples, respectively. The results showed effective and similar antimicrobial activity of Rosmarinus officinalis on broth clinical and food samples with an inhibition zone of 20mm in diameter. The minimum inhibitory concentration [MIC] and minimum bactericidal concentration [MBC] in our study were 1.40 mg/ml and 2.81 mg/ml, respectively. Overuse of antibiotics has led to extensive bacterial resistance to antibiotics, which demonstrates the need for use of new antimicrobial agents. Considering increasing prevalence of MRSA strains and the beneficial effect of rosemary essence on these strains, this essence can be recommended for the treatment of MRSA infections

[ evaluation of probiotic effect of L.acidophilus on the immune responses in BALB/C mice against transplanted tumor derived from breast tissue]

Antitumor effect of lactic acid bacteria have been shown in many studies, this effect maybe due to the immunomodulatory properties of these bacteria. In present work we have studied the effect of Lactobacillus [L] acidophilus on the immune responses of BALB/c mice against transplanted tumor derived from breast tissue. 6-8 week-old in-bred BALB/c mice, each weighing 25-30 g, were used. The mice were divided into two groups each consisted of 9 mice as test and control groups. The L.acidophilus ATCC4356 strain was used in this study. It was inoculated in MRS agar and cultivated overnight under anaerobic conditions then collected and resuspended in PBS. After preparation of proper amount of this suspension it was orally [2.7 x 10[8] CFU/ml] administered to the mice with a gastric feeding 2 weeks before tumor transplantation and 3 weeks after that, with 3 days break and 7 days administration. The control mice received an equal volume of PBS during the study. Results of the present work showed that L. acidophilus can increase the production of immunomodulatory cytokine IL-12 and decrease the TGF-alpha which can suppress immune response. Moreover, the growth rate of tumor in group which received L. acidophilus were decreased and the results of delayed type hypersensitivity [DTH] of this group in 48h were better than control group. The results of our study suggest that daily use of L. acidophilus can regulate immune response with Th1 dominance and may be helpful for cancer immunotherapy, but further studies are needed to investigate the other mechanisms of this effect

[ probiotic effects of Lactobacillus casei in BALB/c mice on the Tumor growth rate bearing breast cancer]

Antitumor effect of lactic acid bacteria [LAB] have been shown in many studies, this effect maybe as a result of immunomodulatory properties of these bacteria. In present work, we have studied the effect of Lactobacillus casei on the tumour growth rate in BALB/c mices bearing breast cancer. 6-8 week-old In-bred BALB/c mice, each weighing 25-30 g, were used. There are two experimental group consisted of 9 mices that they were used as controls in each assay. The L.casei ATCCT 39392 strain used in this study was inoculated in MRS broth and cultivated for a day at 37 °C under anaerobic conditions, collected by centrifugation and resuspend in PBS. After preparation of proper amount of these suspension it was orally administered to the mice with a gastric feeding, Control mice received an equal volume of PBS in duration of study. Results of this study showed that oral administration of L.casei can inhibit the tumour growth and increased the local inflammation in DTH assay as a result of increase in immune responses efficiency. In conclusion oral administration of Lactobacillus casei may regulate immune responses skewed Th1 balance and maybe helpful for cancer immunotherapy, but further studies is needed to investigate the other mechanisms of this effect

[Detection of E.coli strains containing shiga toxin [stx1/2] gene in diarrheal specimens from children less than 5 years old by PCR technique and study of the patterns of antibiotic resistance]

Shiga toxin- producing Escherichia coli [STEC] is an emerging bacterial pathogen in developing countries that causes several diseases such as diarrhea, hemorrhagic colitis [HC] and hemolytic uremic syndrome [HUS], particularly in children. Aim of the research was detection of STEC in diarrheal specimens from under 5 year olds and study of the patterns of antibiotic resistance of these strains. In the study, 300 fecal samples were collected from children with diarrhea referring to Ali Asghar Hospital. E.coli species were isolated by standard bacteriological and biochemical tests. Presence of shiga toxin genes [stx 1/2] was investigated by PCR technique [Qiagen]. Antibiogram test for strains containing the toxin gene was performed using 16 different antibiotic discs [MAST] by disc diffusion agar [Kirby- Bauer] method. From 39 E.coli isolates, 9[23.1%] strains were detected by PCR to contain stx 1/2 gene. One strain was resistant to all 16 antibiotics. All the STEC strains were sensitive to meropenem [MRP], imipenem [IMI], gentamycin [GEN] and nitrofurantoin [NI]. 4[44.44%] strains showed multi-drug resistant pattern. All these 4 strains were resistant to cotrimoxazole [SxT]. Also, 6[66.66%] strains were resistant to at least one antibiotic. In Iran, shiga toxin-producing Escherichia coli [STEC] may be a commonly bacterial pathogen causing diarrhea, particularly in children. Therefore, we should use new techniques for investigation of these strains. Increase in number of emerging and new strains that could be resistant to classic antibiotics such as cortimoxazole may be foreseen. It is suggested that antibiotics prescription programs in treatment of diarrhea causing E.coli strains be updated

[Prevalence and antibiotic resistance pattern of Staphylococcus aureus strains isolated from food stuff]

Staphylococcus aureus is considered to be one of the leading causes of food-borne illnesses. Foodstuff contamination may occur directly from contaminated food-producing animals or may result from poor hygiene during food production processes, or the retail and storage of foods, since humans may carry the microorganism. The number of S. aureus strains that exhibits antimicrobial-resistance properties has increased, together with the potential risk of transmitting the same properties to the human micro flora via food or inducing infections hard to be treated. The aim of this study was to evaluate the occurrence of S. aureus in various food samples and determination of antibiotic resistance pattern in this isolates. A total of 1047 food samples were analysed from July 2006 to December 2007. To determine the presence of S.aureus, the samples were analysed according to the guidelines of Iran standard instructions [No. 1194]. S.aureus isolates were tested for susceptibility to a panel of 11 antimicrobics using the agar disc diffusion method on Muller-Hinton agar. Of 1047 samples analysed 100 [9.5%] were contaminated with S.aureus. Among these contaminated samples, 31% showed antimicrobial resistance properties to at least one of the antibiotic tested and 15 antibiotypes were determined. According to the observed prevalence of S.aureus strains in food samples and their antibiotic resistance pattern, more attention should be paid in foodstuff industry to prevent contamination and transmission of resistant strains

[Prevalence of Yersinia spp. in meat and chicken marketed in southern Tehran]

Yersinia is an important water- and food-borne bacterium causing gastroenteritis in humans. From December 2002 to July 2003, a total of 250 samples -including 158 meat samples and 92 chicken samples- were taken from butcheries and poultry shops operating under the supervision of Tehran University of Medical Sciences. We used a two-step enrichment procedure: phosphate buffer saline was used as primary enrichment within 3 weeks in refrigerator [cold enrichment]. Then we applied KOH treatment as secondary enrichment and performed cultures on CIN agar. In this study, 44.4% of all samples showed Yersinia contamination. The prevalence of Yersinia was 29.1% in meat and 70.7% in poultry. Of the 155 Yersinia isolates, 53 [34.2%] were identified as Y. enterocolitica, 47[30.3%] as Y. intermedia, 42 [27%] as Y. fredriksenii and one [0,6%] as Y. kristensenii. Biotyping of Y. enterocolitica showed that 51 strains [39.7%] corresponded to biotype 1A, 13 strains [24.6%] to biotype 1B, one [1.8%] to biotype 2, three [5,7%] to biotype 3 and one [1.8%] to biotype 4. Fourteen strains [26.4%] could not be classified. The high prevalence rates in meat and poultry implies that these products could be widely contaminated with Yersinia, thus serving as important vehicles for transmission to humans

Evaluation of PCR method for diagnosis of Group B Streptococcus carriage in pregnant women

Group B Streptococcus [GBS] [Streptococcus agalactiae] is the leading cause of morbidity and mortality of newborn infants and accounted as a leading factor causing septicemia after birth in mothers. Infections in infants are usually acquired by contact with the genital tract of the mothers during labor and delivery. In two last decades, significant progress toward detection, prevention and treatment of pregnant women carrying GBS has been achieved. A rapid screening test for GBS that could accurately identify pregnant women carrying the bacteria at the time of delivery would obviate the need for prenatal screening. The standard method for the diagnosis of GBS colonization consists of culturing vaginal and anal secretions in a selective broth medium which inhibits the growth of other microorganisms. Today, it is accepted that PCR has a high sensitivity and specifically in diagnosis. The goal of this study was to screen pregnant woman carrying GBS by PCR. Samples were taken from anal and vaginal mucus of 125 pregnant women who were at 28-38 weeks of ingestion by swab. Samples were tested by standard culture using Todd Hewitt Broth and Blood Agar and also by PCR using primers specific for cfb gene. Culture identified 10 [8%] women as carriage of GBS out of 125 women tested. On the other hand, the PCR assay could identify 12 [9/6%] women positive for GBS. In comparison to culture results, sensitivity, NPV, specificity, and PPV of PCR were 100%, 100%, 98%, and 83%, respectively. The time required for PCR assay and culture were 2h and 36h, respectively. We found that GBS can be detected rapidly and reliably by a PCR assay using combined vaginal and anal secretions from pregnant women at the time of delivery. Also this study shows that the rate of incidence of GBS is high in Iranian pregnant women. We, therefore, recommend screening of pregnant women for detecting of GBS emphatically