Open Access Short Research Article

Proteolytic Activities Expressed by Gastrointestinal Pathogens Bacillus cereus, Listeria monocytogenes and Enterococcus faecium in Different Growth Phases

Carmen M. Abfalter, Thomas P. Schmidt, Silja Wessler

Microbiology Research Journal International, Page 62-70
DOI: 10.9734/BMRJ/2015/16402

Aims: Bacterial proteases are implicated in protein quality control, biofilm formation or might have a direct function in pathogenesis by processing virulence factors or cleaving host factors. In recent years, knowledge of proteases expressed by Gram-negative pathogens remarkably increased. However, investigation of proteases from Gram-positive bacteria is rather rare, but required for the analysis of pathogenesis-relevant proteases. In this study, we extracted and detected proteases from the gastrointestinal pathogens Bacillus cereusListeria monocytogenes, and Enterococcus faecium in different growth phases.
Methodology: Bacteria were grown to logarithmic or stationary phases, harvested and extracted by sonication and French press. For the detection of active proteases, zymography analyses were performed using casein and gelatin as substrates to monitor caseinolytic and gelatinolytic activities.
Results: We observed different active proteases with different intensities in bacteria grown to logarithmic or stationary phases. Strong activities as gelatinases were detected in B. cereus and distinct caseinolytic proteases exhibiting molecular weights of > 170 kDa, 70 kDa and 45 kDa were shown in L. monocytogenes and E. faecium, respectively. Interestingly, detected proteases were differentially regulated in bacteria grown to logarithmic or stationary phases.
Conclusion: In summary, the data clearly indicated proteases that are differentially regulated in the Gram-positive pathogens B. cereus, Lmonocytogenes, and E. faecium, which might contribute to bacterial pathogenesis.

Open Access Short Research Article

Effect of Inoculum Size and Culture Age on the Cellular Properties and Host-Pathogen Interactions of Cryptococcus neoformans

Khi Khi Choo, Pei Pei Chong, Anthony Siong Hock Ho, Phelim Voon Chen Yong

Microbiology Research Journal International, Page 100-108
DOI: 10.9734/BMRJ/2015/16464

Aims: Cryptococcus neoformans is a fungal pathogen which infection caused devastating morbidity and mortality in immunocompromised patients. The present study investigated the effect of culture starting inoculum size and culture age towards cellular properties of C. neoformans and its interactions with mammalian host alveolar epithelial cells.
Methodology: C. neoformans H99 was cultured at different starting inoculum sizes and collected at varied culture ages to examine the morphology of the yeast cells and agar invasion property. The interaction with host alveolar epithelial cells was assessed in vitro using A549 cells as the host cell model.
Results: Visual observation demonstrated that cryptococci cultured with higher starting inoculum sizes and longer incubation periods displayed flocculation properties, aberrant morphologies with lysed cell structure attached to intact yeast cells, release of capsule material to the culture medium, as well as changes in FITC staining of cell surface proteins. The changes in cryptococcal cellular morphology did not affect agar invasion, as the encapsulated cryptococci did not invade agar under all conditions tested. Lysed cell material on cryptococcal cells adhered to host alveolar epithelial cells, which induced localised actin reorganisation at the host-pathogen interface.
Conclusion: Our results indicate that the differences in starting inoculum size and culture age of C. neoformans H99 resulted in yeast cells with distinctive morphologies, which affected he pathogen’s association with host alveolar epithelial cells.

Open Access Original Research Article

In vitro Assessment of the Antibacterial Activity of Matricaria chamomile Alcoholic Extract against Pathogenic Bacterial Strains

Hayder M. Alkuraishy, Ali I. Al-Gareeb, Ali K. Albuhadilly, Salah Alwindy

Microbiology Research Journal International, Page 55-61
DOI: 10.9734/BMRJ/2015/16263

An alarming increasing in the occurrences of antimicrobial resistance inside the existing clinical use and so the recent appearance of multidrug resistant bacteria that attempts the treatment of infections necessities to find out novel antimicrobial agents. In the attendance of Chamomile there is quite a lot of information on various studies concerning the antibacterial effects of this herb and fractioned bioassay conducted with the consent of energetic standards. Chamomile powder (Matricaria chamomilla L.) was purchased from private pharmacies. Fifty grams of chamomile powder was extracted with 250 ml 10% ethanol by soxhelt device at 45ÄŠ. In vitro determination of antibacterial activity of alcoholic extract of Matricaria chamomilla was done. The chamomile alcoholic extract showed higher action against Klebsiella pneumoniae (35±1.57 mm) inhibition zone and lower effect alongside Enterococcus faecalis (10±1.43 mm), but all the results were significant because inhibition values were near the result of active control. MIC values of chamomile alcoholic extract near values of tetracycline mainly against Klebsiella pneumoniaeEscherichia coli and Staphylococcus aureus. While MBC values of the chamomile alcoholic extract were in corresponding with tetracycline values similar to MIC, except that chamomile produced less corresponding values with tetracycline against Enterococcus faecalis and Pseudomonas aeruginosa.
Conclusion Chamomile possesses significant and potent antibacterial activity against various bacterial strains like Pseudomonas aeruginosaEnterococcus faecalisStaphylococcus aureusEscherichia coli and Klebsiella pneumoniae.

Open Access Original Research Article

Detection of Indicator Organisms in Drinking Water by Membrane Filtration Method in Open Defaecation Free VDCs of Kaski District, Nepal

Dhaka Raj Pant, Niraj Chaudhary, Ishor Sharma, Chiranjivi Adhikari, Dipesh Kumar Karki, Niraj Manandhar, Sworup Shrestha, Shraddha Upadhyaya

Microbiology Research Journal International, Page 81-92
DOI: 10.9734/BMRJ/2015/16162

Aims: The aim of the project is to detect and enumerate the presence of indicator organisms in drinking water sources of open defaecation free (ODF) village development committees (VDCs) of Kaski district; to compare distribution of Total coliform (TC) and Faecal count (FC) among various water sources.
Study Design: Cross-sectional Study.
Place and Duration of Study: Department of Medical Laboratory Science, School of Health and Allied Sciences, Pokhara University, December 2013 to March, 2014.
Methodology: The study was conducted to detect and enumerate indicator organisms in ODF VDCs of Kaski district, Nepal. 44 water samples were collected from reservoir / distribution tank, tapstand and spring. The bacteriological water quality was analyzed by using membrane filter method to detect the presence and its risk level of fecal contamination to human health in study area. The study mainly focuses on two part of the study; laboratory analysis for water samples and questionnaire survey. Total water samples collected were 44, out of which 32 were from tap, 3 from reservoir tank and 9 from spring. Chlorine disinfection treatment was not done in any VDCs.
Results: All the drinking water samples from Bharatpokhari and Kalika VDCs (100%) were found to be fecally contaminated and the number of total coliforms and fecal coliform per 100ml water were above WHO guidelines for drinking water. None of the water sources from both the VDCs were potable for drinking. Mann-Whitney test was used to compare mean range between samples of two VDCs and mean rank of total coliforms of Bharatpokhari and Kalika VDC were found to be 29.18 and 13.17 and that of Bharatpokhari and Kalika VDCs were found to be 29.80 and 12.89 respectively. Tap water, reservoir/distribution tank and spring water were analyzed to detect indicator organisms and Kruskal Wallis test suggest no significant difference in fecal contamination level among various sources of water was found (p>0.05). The median concentration of fecal coliform of gravity water (spring) in Bharatpokhari was found to be 95 CFU/100ml and that of Kalika is 40 CFU/100 ml. E. coli was detected in every water samples of both the VDCs.
Conclusion: The water bodies used for drinking purpose in Bharatpokhari and Kalika VDC are heavily contaminated with total and fecal coliform bacteria even though those VDCs have been declared ODF. The water bodies used for drinking purpose in Bharatpokhari is found to be more contaminated than Kalika VDC. The distribution of total coliforms and fecal coliforms are same across categories of source of sample. E. coli was isolated from every water sample under study.

Open Access Original Research Article

Caenorhabditis elegans-Aspergillus fumigatus (Nematode-mould) Model for Study of Fungal Pathogenesis

Ikechukwu Okoli, Elaine M. Bignell

Microbiology Research Journal International, Page 93-99
DOI: 10.9734/BMRJ/2015/15838

Aims: Caenorhabditis elegans nematode pathosystem has been used to study both bacterial and fungal pathogenesis. Apart from Candida and Cryptococcus, studies using this model for other fungal infections especially filamentous fungi however, are still lacking. This work aimed at developing a C. elegans-Aspergillus fumigatus (nematode-mould) killing assay model.
Study Design: Infection model of Caenorhabditis elegans with Aspergillus fumigatus.
Place and Duration of Study: Centre for Molecular Microbiology and Infection, Imperial College London, London SW7 2AZ, United kingdom, between October 2011 and April 2012.
Methodology: Double mutant glp-4;sek-1 strain of C. elegans worms were propagated and maintained on nematode growth medium (NGM) with Escherichia coli non-pathogenic strain HB101 used as food prior to a fungal killing assay. L4 stage of the worms were infected with spores of A. fumigatus wild-type strain AF293, and incubated in 30% brain heart infusion (BHI) in M9 buffer at room temperature for 72 h. The survival of the worms was studied within this period.
Results: The scenario presented after killing of the worms by A. fumigatus appears to be the same as previously reported for Candida albicans, except for the position of the protruded filaments on the worms.
Conclusion: This model provides a platform for future studies of fungal pathogenesis, “curing” experiments, as well as for discovery of new antifungal agents.

Open Access Review Article

Resurgence of Pandemic A/H1N1 2009 Viruses during Influenza Season 2013-2014

Omar Mestoui, Mohammed EL Mzibri, Sanaâ Lemriss, Amal Barakat, Saâd EL Kabbaj, Abdelkarim Filali Maltouf

Microbiology Research Journal International, Page 71-80
DOI: 10.9734/BMRJ/2015/15781

In April 2009, the world has known an emergence of a triple-reasserting influenza virus, pandemic A/H1N1, causing a pandemic around the world. The virus has continued to circulate during influenza seasons from 2010 to 2013, but with low frequencies. However, during the influenza season 2013-2014, we have assisted to the resurgence of this same virus worldwide. A comparison of the 2009-2010 and the 2013-2014 influenza seasons revealed a notable change regarding the epidemiology of this virus, in term of hospitalizations and deaths associated with pandemic A/H1N1: A gradual transition of hospitalizations and deaths for older ages related to influenza has been reported.
The resurgence of pandemic A/H1N1 is critical and its explanation is still difficult, but could be related to the viral genetic and/or the host susceptibility. Therefore, the best management of the disease and the effective control of the virus can be attempted by vaccination of all persons without contraindications and strengthening the surveillance systems of influenza.