A comparison of PCR and LAMP methods for detecting shiga toxin producing escherichia coli / Монголын Анагаах Ухаан

Introduction@#PCR to detect and amplificate the virulence genes of STEC is specific and more sensitive, however, it takes five to six hours for whole test procedure and requires special lab instruments such as thermocycler. Loop-mediated isothermal amplification is a simple, rapid, specific and cost-effective nucleic acid amplification method by using four to six primers when compared to PCR, nucleic acid sequence-based amplification, self-sustained sequence replication and strand displacement amplification.@*Goal@#Detection and comparison of STEC by PCR and LAMP@*Materials and Methods@#In our study, we analyzed comparison of PCR and LAMP results on standard strain used quality control strain solution which diluted 1pg/µL DNA, 10 pg/µL DNA, 100 pg/µL DNA, 1 ng/μL DNA, 10 ng/μL DNA, and 100 ng/μL DNA concentration from LB agar cultures. @*Research ethics@#Permission to submit the survey was granted by the Ethics Review Committee of the MNUMS and the survey was conducted in accordance with the rules and regulations.@*Result@#Sensitivity of Stx1 and stx2 genes in PCR results are positive in 10 pg/µL DNA solution and negative in 1pg/µL DNA. In LAMP test results showed that positive for all concentration. It shows that LAMP method sensitivity is 10 times more than PCR. @*Conclusion@#It shows that LAMP method sensitivity is 10 times more than PCR. All in allLAMP test is cost effective test with sensitive for detection STEC.

A comparison of PCR and LAMP methods for detecting shiga toxin producing Escherichia coli / Монголын Анагаах Ухаан

Introduction@#PCR to detect and amplificate the virulence genes of STEC is specific and more sensitive, however, it takes five to six hours for whole test procedure and requires special lab instruments such as thermocycler. Loop-mediated isothermal amplification is a simple, rapid, specific and cost-effective nucleic acid amplification method by using four to six primers when compared to PCR, nucleic acid sequence-based amplification, self-sustained sequence replication and strand displacement amplification.@*Materials and Methods@#In our study, we analyzed comparison of PCR and LAMP results on standard strain used quality control strain solution which diluted 1pg/µL DNA, 10 pg/µL DNA, 100 pg/µL DNA, 1 ng/μL DNA, 10 ng/μL DNA, and 100 ng/μL DNA concentration from LB agar cultures. @*Research ethics@#Permission to submit the survey was granted by the Ethics Review Committee of the MNUMS and the survey was conducted in accordance with the rules and regulations. @*Goal@#Detection and comparison of STEC by PCR and LAMP@*Result@#Sensitivity of Stx1 and stx2 genes in PCR results are positive in 10 pg/µL DNA solution and negative in 1pg/µL DNA. In LAMP test results showed that positive for all concentration. It shows that LAMP method sensitivity is 10 times more than PCR. @*Conclusion@#It shows that LAMP method sensitivity is 10 times more than PCR. All in allLAMP test is cost effective test with sensitive for detection STEC.

Presence and antibiotic resistance of diarrheagenic Escherichia coli in ready-to-eat salads / Монголын Анагаах Ухаан

Introduction@#Foodborne diseases are a major public health concern worldwide. The report, which estimates the burden of foodborne diseases – states that each year as many as 600 million, or almost 1 in 10 people in the world, fall ill after consuming contaminated food. Of these, 420 000 people die, including 125 000 children under the age of 5 years. The 20.3% of diarrhea and 27.5% of die caused by contaminated foods are diarrheagenic Escherichia coli (DEC).@*Aim@#To identify of DEC and determine their antibiotic resistance from ready-to-eat salads@*Material and Methods@#A total of 40 bagged salad mix samples were collected from food markets in Ulaanbaatar, Mongolia. Escherichia coli (E.coli) strains were determined on the basis of MNS 6308:2012 standard and confirmed by polymerase chain reaction (PCR) in samples. DEC was identified using multiplex PCR. Bacterial susceptibility to antimicrobial agents determined by the Kirby Bauer disk diffusion method.@*Results@#Our results showed the presence of E. coli in 19 samples (47.5%). DEC isolates identified by multiplex PCR were defined as follows: the presence of eae and bfp for EPEC, the presence of lt for ETEC, the presence of ipaH for EIEC, the presence of stx1 and stx2 for EHEC, the presence of aap and aggR for EAEC, and the presence of daaE for DAEC. The multiplex PCR assays detected EHEC 6 (31.6%), EPEC 5 (26.3%), EIEC 1 (5.3%). EAEC and ETEC were not detected in samples. The E.coli isolates were 73.7% resistant to chloramphenicol as the first choice of treatment of diarrhea and high resistance (68.4-94.7%) to the cephalosporins. In our country, cephalosporins are widely used in medical practice for the treatment of infectious diseases.@*Conclusion@#In this study, about half of ready-to-eat salads are contaminated with E. coli. The three types (EHEC, EPEC, EIEC) of DEC pathotypes were identified in the ready-to-eat salads and high prevalent of antimicrobial resistance. Future research is required to track the contamination sources and develop appropriate steps that should be taken by industry and retailers to reduce microbial contamination in ready-to-eat salads.

Detection result of virulence factors in Staphylococcus aureus isolated from retail beef / Монголын Анагаах Ухаан

Introduction@#In the United States, Staphylococcus aureus (S.aureus) is considered one of the top five pathogens causing domestically acquired foodborne diseases and is responsible for an estimate of 241,000 illnesses per year. Foods that have been frequently implicated in Staphylococcal food-borne disease are meat, meat products, egg products, milk, dairy products, salads and bakery products. β-lactam antibiotics are routinely prescribed for treating S. aureus caused infections, but antibiotic resistance is increasing at an alarming rate.@*Aim@#Detection of virulence factors and antibiotic resistance in S.aureus isolated from retail beef@*Materials and Methods @#A total of 100 meat samples were collected from markets including Kharkhorin 28, Bars 4, Bayanzurkh 15, Huchit shonkhor 33, Denjiin myanga 4 and Bumbugur 16. S.aureus strains were determined on the basis of MNS 6308:2012 standard using Baird-Parker selective agar and confirmed by polymerase chain reaction (PCR) in retail beefs. Bacterial susceptibility to antimicrobial agents determined by the Kirby Bauer disk diffusion method.@*Results@#Overall, 81% meat samples were contaminated with staphylococcal of which 54.3% were low, 28% were moderate, 11.1% were high and 6.1% were very high. PCR amplification of the thermostable nuclease-encoding nuc gene using the gene-specific primers and the chromosomal DNA preparation yielded a 270 bp amplicon, as expected and 35 (43.2%) confirmed as S. aureus. According to the findings of the current study, S.aureus strains isolated from the beef were high resistant (88.6% -97.1%) to antibiotics of penicillins group and low resistant (8.6%) to chloramphenicol. In total, 48.6% of isolates were multidrug resistant.@*Conclusion@#The contamination of staphylococcal was high in retail beef in Ulaanbaatar. Most S.aureus isolates exhibited resistance to a antibiotics of penicillin group. The half of the isolates were multidrug resistant and high virulence.

Detection of virulent of Klebsiella isolates from ñlinical samples / Монголын Анагаах Ухаан

IntroductionKlebsiella spp is a well-known opportunistic pathogen associated with nosocomial infections such asurinary tract, septicaemia and pneumonia number of multi-drug resistant strains and infections causedby Klebsiella has progressively increased, causing treatment limitations.GoalIdentify of phenotype of Klebseilla isolates from ñlinical samplesMaterials and MethodsA total of 112 Klebsiella strains were isolated from clinical samples in State Central First Hospital and StateCentral Third Hospital from July 2015 through December 2015. The bacterial isolates were identifi edaccording to cultural characteristics, biochemical test and API20E. The serum resistance, capsule andhypermucoviscosity, cell surface protein (curly), a-hemolysin and ability to form biofi lm were sought byphenotypic assays. Antimicrobial susceptibility was tested by diffusion method.ResultA total of 112 Klebsiella samples were collected. The bacterial isolates were identifi ed according tocultural characteristics, biochemical test and API20E, the results revealed that 16.1 percent isolateswere identifi ed as K.oxytoca all of them 83.9 percent isolates were belong to K.pneumonia. Therewere observed for ampicillin (99 percent), nitrofurantoin (53.6 percent), cepalotin (50.6 percent) and51 percent of isolates were considered as a multiple drug resistant. Serum resistance properties ofK.pneumoniae was resistance 89.4 percent, intermediately susceptible 4.3 percent, sensitive 6.4percent and for K.oxytoca resistance 88.9 percent, intermediately susceptible 5.6 percent, sensitive 5.6percent. The hemolysin àalpha was detected in 32.2 percent, and gamma, beta in 66.96 percent, 0.9percent respectively. The capsule was observed in 46.5 percent and hypermucoviscosity in 27.7 percentof isolates. The cell surface protein (curly) and biofi lm were detected in 100 percent.Conclusion:Both K.pneumoniae and K.oxytoca isolates from clinical samples have similar virulent properties, andthe a-hemolysin and hypermucoviscosity positive isolates were more resistance to antibiotics.

Relation between adherence factors and the phylogenetic group of extraintestinal pathogenic Escherichia coli / Шинэ санаа Шинэ нээлт

Extraintestinal pathogenic Escherichia coli (ExPEC), the specialized strains ofE.coli that cause most extraintestinal infections, represent a major but littleappreciated health threat. Phylogenetic analysis has shown that ExPEC is composedof four main phylogenetic groups (A,B1, B2, and D) and that virulent extraintestinalstrains mainly belong to groups B2 and D.In this study, we aimed to assess therelation between adherence virulence and phylogenetic groups of ExPEC.A total of 161 E.coli samples were collected. Out of these 17 (10.6%) werefrom pus, 66 (41 %) from urine, 78 (48.4%) from cervical swab. The phylogeneticgroups and 6 virulence genes (fimH, papC, papGII, papGIII, fa/draBC,andSfa/focDE) encoding adhesins were identified by triplex PCR. Phylogeneticgroups distribution was as follows: B1 10.5%, A 24.7%, B2 25.3%, and D 38.9%. Virulence genes prevalence was fimH 90.1%, papC 23%, papGII 16.8%, papGIII1.9%, Afa/draBC 11.8%, andSfa/focDE 5.6%. The cell surface protein (curli) wasdetected 50,3% by Congo red agar. In conclusion: The most isolated strainsbelonged to the phylogenetic group B2 and D. The phylogenetic groups weresignificantly associated with some genes encodingadhesins (fimH, papC) and cellsurface protein (curli).

Relation between adherence factors and the phylogenetic group of extraintestinal pathogenic Escherichia coli / Шинэ санаа Шинэ нээлт

Extraintestinal pathogenic Escherichia coli (ExPEC), the specialized strains ofE.coli that cause most extraintestinal infections, represent a major but littleappreciated health threat. Phylogenetic analysis has shown that ExPEC is composedof four main phylogenetic groups (A,B1, B2, and D) and that virulent extraintestinalstrains mainly belong to groups B2 and D.In this study, we aimed to assess therelation between adherence virulence and phylogenetic groups of ExPEC.A total of 161 E.coli samples were collected. Out of these 17 (10.6%) werefrom pus, 66 (41 %) from urine, 78 (48.4%) from cervical swab. The phylogeneticgroups and 6 virulence genes (fimH, papC, papGII, papGIII, fa/draBC,andSfa/focDE) encoding adhesins were identified by triplex PCR. Phylogeneticgroups distribution was as follows: B1 10.5%, A 24.7%, B2 25.3%, and D 38.9%. Virulence genes prevalence was fimH 90.1%, papC 23%, papGII 16.8%, papGIII1.9%, Afa/draBC 11.8%, andSfa/focDE 5.6%. The cell surface protein (curli) wasdetected 50,3% by Congo red agar. In conclusion: The most isolated strainsbelonged to the phylogenetic group B2 and D. The phylogenetic groups weresignificantly associated with some genes encoding adhesins (fimH, papC) and cellsurface protein (curli).

Identification of Enteroaggregative Escherichia coli in Mongolia / Монголын Анагаах Ухаан

Introduction: Enteroaggregative Escherichia coli (EAEC) is an important agent of acute and persistent diarrhea worldwide. Few cases have been reported in healthy children. EAEC strains are characterized by aggregative adherence (AA) to HEp-2 cells, wherein bacteria are seen in “stacked brick” aggregates attaching to HEp-2 cells and usually to the glass surface between cells. Goal: To identify Enteroaggregative Escherihia coli using multiplex polymerase chain reaction (PCR) and HEp-2 adherence assay in Ulaanbaatar, Mongolia Materials and Methods: A total of 329 E. coli strains were isolated from stool with diarrhea in National Center for Communicable Diseases from July 2012 through September 2014. All specimens were processed by routine microbiological and biochemical tests in the bacteriological laboratories to identify Salmonella spp., Shigella spp. All specimens in our study were negative for these bacterial and parasitic pathogens. The biofilm formation was evaluated by the growth rate of E.coli on plastic surface. PCR assays were used to detect genes of five types of diarrheagenic E.coli (DEC). All of the DEC strains showed mannose-resistant adherence to HEp-2 cells, and aggregative adherence was predominant in these isolates. Bacterial susceptibility to antimicrobial agents determined by the Kirby Bauer disk diffusion method on Muller Hinton agar. Results: EAEC (31.9%) was the most prevalent by PCR and HEp-2 assay comparing with others. EAEC by multiplex PCR in samples (11, 3.3%), followed by enteropathogenic E.coli (EPEC) seen in 2.1%. Enterohemorrhagic E.coli (EHEC) and enteroinvasive E.coli (EIEC) were found in 7 (2.1%) and 1 (0.3%) of the samples. Enterotoxigenic E.coli (ETEC) and diffusely adhering E.coli were detected in 2 (0.6%), respectively. The evaluation of bacterial biofilm formation using 96 well plates showed 309 negative (93%), 15 weak biofilm (4.6%) and 8 moderate biofilm (2.4%) formation for E.coli and no strong biofilm forming strain was detected. Above 50% of antibiotic resistance was observed for ampicillin, trimethoprim/sulfamethoxazole, cefuroxime and cephalotin. Also, 95.4% of isolates were resistant to at least three different classes of antimicrobial agents and considered as multidrug resistance. Conclusion: EAEC is most prevalent pathogen among DEC in our samples. It is necessary to implement EAEC identifying method on Hep-2 assay in our laboratory practice.

The detection of adherence factorsby Escherichia coli cause of urinary tract infectionsin Ulaanbaatar, Mongolia / Монголын Анагаах Ухаан

INTRODUCTION: Urinary tract infections among the most common bacterial infectious diseases encountered at all ages. Escherichia coli are being the etiologic agent in 50–80%. Therefore, it is an important public health problem. E.coli causing urinary tract infections express pilli, fimbriae and others adherence virulence factors. GOAL: To detect the some adherence virulence factors of Uropathogenic Escherichia coli (UPEC) in Ulaanbaatar, Mongolia MATERIALS AND METHODS: A total of 76E.colisampleswere collected. These samples were positive bacteriological examination of urine, performed at the bacteriological laboratory of the State Central Third Hospital and State Central First Hospital, Ulaanbaatar, Mongolia. The biofilm formation was evaluated by the growth rate of E.coli on plastic surface.The detection of the virulence factors type 1 fimbriae (fimA gene) and P-fimbriae (papC) was performed by multiplex PCR using gene specific primers.Curli expression was determined by using congo red agar. RESULTS: The evaluation of bacterial biofilm formation using 96 well plates showed 40 negative (52.6%), 32 weak biofilm (42.1%) and 4 moderate biofilm (5.3%) formation for E.coli and no strong biofilm forming strain was detected. The cell surface protein (curli) was detected by Congo red agar. The result was 71% positive for studied E.coli strains. The detection result of pili genes by multiplex PCR showed that fimH gene detected for 73 (96.1%) and papC gene detected for 18 (23.7%) E.coli cultures. CONCLUSION: Almost half of surveyed Uropathogenic E.coli isolated in Ulaanbaatar, Mongolia had ability of biofilm formation and it has been determined by the bacterial surface protein (curli), which is one of bacterial adherence factors, may cause biofilm formation.