Open Access Original Research Article

Specific Gram-Positive Antibacterial Activity of 4-hydroxy-3-(3-methyl-2-butenyl) Acetophenone Isolated from Senecio graveolens

Javier Santander, Caitlin Otto, Dave Lowry, Mauricio Cuellar, Marco Mellado, Cristian Salas, Francisco Rothhammer, Carlos Echiburu-Chau

Microbiology Research Journal International, Page 94-106
DOI: 10.9734/BMRJ/2015/11934

Background: Resistance to antimicrobial drugs has become an increasingly global problem, and is the main reason for an extended search for new drugs to treat microbial infections. Senecioneae is one of the largest tribes of Asteraceae, comprised of about 150 genera and 3000 plant species. Senecio graveolens, commonly called Chachacoma, is highly used as a medicinal plant for altitude sickness by the natives of the Andes Mountains around the Atacama Desert. Previous studies have demonstrated that S. graveolens extracts possess antibacterial properties, but its active compound and molecular mechanisms are still unknown.
Methods: Form the ethanolic extract of S. graveolens the main compound 4-hydroxy-3-(3-methyl-2-butenyl)acetophenone (4-H-3-(MB)AP) was identify and purified by nuclear magnetic resonance (NMR). Antibacterial activity of (4-H-3-(MB)AP) was assayed on Gram-positive and Gram-negative bacteria by microbiological techniques. Possible mechanisms of action of (4-H-3-(MB)AP) were explored by microbiological, flow cytometry and electron microscopy techniques.
Results: Here we determined that S. graveolens extract has specific antibacterial activity against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus, and Mycobacterium smegmatis. The most abundant compound from S. graveolens extract, 4-H-3-(MB)AP, showed broad antibacterial activity against Gram-positive but no activity against Gram-negative strains. We determined that 4-H-3-(MB)AP permeabilizes bacterial membranes and precludes cell division by disrupting Gram-positive bacteria divisome, suggesting that the synthesis of teichoic acid is inhibited.
Conclusions: We conclude that 4-H-3-(MB)AP is one of the active compounds of S. graveolens extract responsible for its antibacterial activity. 4-H-3-(MB)AP is a candidate for further chemical modification studies and practical approaches to design antimicrobial drugs.

Open Access Original Research Article

A Cross Sectional Study on Prevalence of Antibiotic Resistance and Role of Efflux Pumps in Fluoroquinolone Resistance by using Efflux Pump Inhibitors in Isolated Cultures from Poultry, Dairy Farms and MTCC Strains from Reservoirs

Leena Seasotiya, Priyanka Siwach, Pooja Bharti, Sheema Bai, Anupma Malik, Ravinder Kumar, Sunita Dalal

Microbiology Research Journal International, Page 107-116
DOI: 10.9734/BMRJ/2015/12342

Aims: Emergence of antibiotic resistance in bacterial strains has always remained a crucial concern. Mutations in antibiotic target sites, over expression of efflux pump are the major modes of development of bacterial antibiotic resistances. The present study was conducted to determine antibiotic resistance and role of efflux pumps in fluoroquinolone resistance by using efflux pump inhibitors.
Place and Duration of Study: The Research was conducted during June 2011 to March 2012 at Department of Biotechnology, Kurukshetra University, Kurukshetra, Haryana – India.
Methodology: Out of 57 bacterial strains 19 were procured from collection centres (reservoirs) and 38 were isolated from dairy (n = 10) and poultry farms (n = 28) and screened against 12 antibiotics of different groups by well assay. Further, fluoroquinolone sensitive/resistant strains were tested to observe the decline in minimum inhibitory concentration (MIC) levels in the presence/absence of efflux pump inhibitors.
Results: Antibiotic resistance in tested strains was higher against nitrofurantoin, lincomycin, cefixime and chloramphenicol. Majority of the bacterial strains (94.74%) showed resistance to two or more antibiotics. Isolated bacterial strains were exhibiting more antibiotic resistance than reservoir strains indicating their exposure to antibiotics. Reduction in MIC (2-4 folds) was observed when Piperine (28.07%) or Plumbagin (19.29%) was used in combination with fluoroquinolones. The findings emphasized that majority of efflux pump inhibitors are active against Gram-positive bacteria. Overexpression of efflux pump and higher antibiotic resistance was also observed in subgroup III exhibiting resistance to combination of cefixime/nitrofurantoin.
Conclusion: Efflux mediated resistance appears to contribute significantly to fluoroquinolone resistance and multidrug resistance in organisms, which may be due to involvement of active efflux pumps of both MFS and RND Family.

Open Access Original Research Article

Prevalence of Multiple Antibiotic Resistant Escherichia coli Serotypes in Cow Raw Milk Samples and Traditional Dairy Products in Osun State, Nigeria

Thonda Olukemi Abike, Oluduro Anthonia Olufunke, Kolawole Deboye Oriade

Microbiology Research Journal International, Page 117-125
DOI: 10.9734/BMRJ/2015/12868

Aims: Isolation and characterization of multiple antibiotic resistant Escherichia coli serotypes in cow raw milk and traditional dairy products in Osun state, Nigeria was reported.
Study Design: Experimental based study of raw milk of lactating cows, local cheese and yoghurt.
Place and Duration of Study: Samples were collected at different markets in Ile-Ife, Modakeke, Edun-abon, and Akinlalu in Osun State, Nigeria, between June and August, 2011.
Methodology: Samples of cow raw milk, cheese and yoghurt were enumerated bacteriologically on MacConkey agar at 37ºC for the total coliform. Isolation of E. coli was done on eosin methylene blue agar at 37ºC using pour plate technique. The isolate identity was further confirmed by biochemical tests and serotyping. Antibiotic susceptibility of the isolates was determined by the disk diffusion technique. Molecular typing of Escherichia coli was performed using polymerase chain reaction (PCR) technique.
Result: The coliform count ranged from 1.0 to 6.00×105cfu/g in raw milk, 1.30 to 9.60×105cfu/g in cheese and 1.3 to 5.0×105cfu/g in yoghurt. The presence of E. coli was significantly low in cow raw milk compared to yoghurt and cheese (p< 0.05). Resistance to antibiotics varied among the E. coli isolates with the highest resistance to tetracycline (88.6%), cotrimoxazole (77.2%) and nitrofurantoin (6.81%). Most E. coli isolates were multiple antibiotic resistant types displaying 17 different multiple antibiotic resistance patterns. All the E. coli strains belonged to 9 serogroups (‘O’) and 17 serotypes (‘H’). The ‘O’ serogroups identified include O26 (2), O55 (4), O86 (3), O111 (11), O114 (1), O119 (1), O127 (5), O128 (1) and O142 (4) with 0111 (35.48%) being the most prevalent serotype.
The flagellin (fliC) gene restriction analysis of E. coli serotypes showed that 72.7% were motile with 17 H-type while 27.3% were non-motile (O55 (1), O86 (1), O111 (5), O119 (2), O127 (2), O142 (1) ). The polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) patterns for the amplified fliC gene in the E. coli strains, produced bands of sizes ranging from 0.9 to 1.8 kb.
Conclusion: The identification of E. coli O55:H27, O111:H8, O127:H42 and O142:H6 serotypes in the dairy products is of great public health concern.

Open Access Original Research Article

Plasmid Borne Resistance in Bacteria to Common Household Spices

Anoor Pawan Kumar, Karthik Rajkumar, M. Swathi Raju, A. Nichita, Sandeepta Burgula

Microbiology Research Journal International, Page 126-138
DOI: 10.9734/BMRJ/2015/11066

Aim: To examine the antimicrobial properties of various spices commonly used in Indian cuisine and to check if the resistance in bacteria is plasmid borne.
Study Design: To demonstrate the antimicrobial properties in various household spices like Cinnamon (Cinnamomum zeylanicum), Clove (Syzygium aromaticum), Black Pepper (Piper nigram) and Turmeric (Curcuma longa) and to examine their effects on the pathogenic bacteria Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus.
Place and Duration of Study: Department of Microbiology, Osmania University, India during June 2013- January 2014.
Methodology: Antimicrobial assay was performed to determine the antimicrobial activity of these spices. Plasmids were isolated from bacteria showing resistance to these spices to confirm if the resistance to these spices was plasmid borne. The isolated plasmids were used to transform E. coli DH5α which was previously observed to be sensitive to these spices.
Results: On transformation with plasmids isolated from K. pneumoniae and P. aeruginosa, E. coli DH5α cells were observed to gain resistance to all the four spices tested in the original strains of K. pneumoniae and P. aeruginosa.
Conclusion: The resistance to spices seems to be a plasmid borne feature which may be transferred to sensitive bacterial strains.

Open Access Original Research Article

Microbial Contamination of Ghanaian Cedi Notes from Traders of the Tamale Central Market, Ghana

P. Luure, W. Asare, S. J. Cobbina, A. B. Duwiejuah, M. Nkoom

Microbiology Research Journal International, Page 139-145
DOI: 10.9734/BMRJ/2015/12551

The study was to determine the microbial contamination of Ghanaian cedi notes from traders in the Tamale Central Market, Ghana. A total of ninety (90) currency notes of three lower denominations mainly in circulation (Ghanaian cedi 1, 2 and 5) were collected into sterile paper envelopes. Membrane filtration technique was used to determine total coliform bacteria, Salmonella spp and Escherichia coli. Microbes isolated from the cedi notes were Escherichia coli (19.1%), Salmonella species(3.8%), Bacillus species (0.4%), Staphylococcus aureus (0.0%) and Total coliform (76.6%). The five Ghana Cedi notes had the highest microbial load (746) followed by the two Ghana cedi notes (593) and one Ghana cedi notes (44) recorded the least microbial load. There was also a strong positive correlation (0.97*) between the GH¢5 and GH¢2 notes at 1% significant level which indicate common source of microbial contamination of the cedi notes. Five non-circulated notes denomination were used as control recorded no microbial count. The study revealed that handling of Ghana cedi notes cannot be risk free. It is therefore recommended that individuals should improve upon their personal health consciousness by washing hands after handling of currency notes.

Open Access Original Research Article

Bacteriological Assessment of Hawked Sorrel Drink (Zobo Drink) in Aba, South-East Nigeria

O. R. Ezeigbo, S. Uhiara, J. A. Nwodu, M. U. Ekaiko

Microbiology Research Journal International, Page 146-151
DOI: 10.9734/BMRJ/2015/13027

Bacteriological investigation of hawked Sorrel drink (zobo drink) was carried out in Aba, South-east Nigeria, between January and March, 2014. Zobo drink is a non-alcoholic local beverage made from different varieties of dried petals, acid succulent aqueous extract of calyx of roselle, Hibiscus sabdariffa. It is consumed in Nigeria and other parts of the world. Samples of zobo drink were randomly purchased from the open markets, for a period of three months and analyzed for bacteriological qualities using standard methods. The result revealed a total aerobic bacterial count ranging from 0.3 × 106 cfu/ml to 4.4× 106 cfu/ml while the total coliform count ranged from 0.1 × 105cfu/ml to 6.5 × 105 cfu/ml. The control sample gave 0.8 ×106 cfu/ml and 0.1 × 105cfu/ml for aerobic bacterial and coliform count respectively. All the screened samples including the control sample had total aerobic bacterial count above the acceptable limit of <104cfu/ml. The contamination of the control samples could likely come from the spices (grounded) and additives which are usually added raw, since the appropriate hygienic standards were maintained during preparation. A total of five bacterial isolates were identified, which include Escherisha coli, Staphylococcus spp, Lactobacillus spp, Bacillus spp and Pseudomonas spp. Personal and environmental hygiene is required during production, packaging and preservation of zobo to avoid food borne illnesses.

Open Access Original Research Article

Morphological, Molecular and Biochemical Comparative Studies of Two Novel Fungal Honey Isolates

Amany Lotfy Kansoh, Zeinab Hassan Kheiralla, Hala Abd Elmonem Ahmed, Tarek Kahil, Eman Karam Abd El-Hameed, Mona Abdeltawab Esawy

Microbiology Research Journal International, Page 152-168
DOI: 10.9734/BMRJ/2015/12695

Aims and Study Design: The stress honey medium expected to have spores with unique feature. Within this context, two honey isolates were identified, characterized, and evaluated as secondary metabolite producers.
Place and Duration of Study: The study was undertaken in the National research center, 33 Al Behous, Dokki, Giza, Egypt. The duration of study was during the period of January 2012 to February 2013.
Methodology: Two strains were isolated from mountain honey, Yemen. They were identified based on morphological characterization and 18S rRNA sequence analysis. Validation of novelty was supported further by their protein profile and randomly amplified polymorphic DNA PCR patterns (RAPD). Growth parameters were studied to determine optimum growth factors. In addition, they were undergone to enzymatic, antibiotic, antioxidant, ochratoxins and aflatoxins testes.
Results: Two novel honey isolates were identified and designed as Aspergillus niger EM77 and Aspergillus awamori EM66. Intracellular protein profiles showed clear difference among the honey isolates and their references. The RAPD referred to 51% coincidence between the two isolates and showed different RAPD patterns between the references and honey isolates. Different parameters which affected isolates growth such as pH, temp, carbon and nitrogen sources were studied. Effect of NaCl concentrations referred to the halo-tolerant feature of both isolates. Both strains showed strong antimicrobial activity against Candida albicans, C. tropicalis and Pseudomonas species. Reducing power assay reported 82% and 83% antioxidant activity of Aspergillus niger EM77 and A. awamori EM66, respectively. Aflatoxins and ocratoxins tests revealed that both isolates had negative results. In addition, two fungal honey isolates showed relatively similar enzymatic activities.
Conclusion: This research continues in highlights the honey as a new reservoir of unique isolates. In addition, it was suggested that the honey osmophilic stress carried spores have novel properties.

Open Access Original Research Article

Prevalence and Antibiotic Susceptibility of Gram-negative Aerobic Bacteria Cultured from the Intestine and Hepatopancreas of Blue Land Crab (Cardisoma guanhumi) in Grenada, West Indies

Victor A. Amadi, Ross Peterson, Vanessa Matthew-Belmar, Ravindra Sharma, Harry Hariharan

Microbiology Research Journal International, Page 169-179
DOI: 10.9734/BMRJ/2015/13620

Aim: To evaluate the prevalence of aerobic bacteria associated with the intestine and hepatopancreas of blue land crab and the susceptibility of the bacteria to a panel of antimicrobials that included some drugs used for the treatment of bacterial infections in the human and veterinary clinics in Grenada.
Study Design: The tested crabs were collected during a three month period from November 2011 to February 2012 from six parishes of Grenada and analyzed in the bacteriology lab in the Pathobiology Department, School of Veterinary medicine, St. George’s University, Grenada, West Indies.
Methodology: A total of 65 blue land crabs were examined for the presence of Gram-negative aerobic bacteria in their intestines and hepatopancreas by culture. The isolated bacterial species were tested against 12 antibiotics using the disc diffusion method.
Results: Eighty-nine percent of crabs were culture positive. Klebsiella pneumoniae was the most common species (60%), followed by Citrobacter freundii (28%), Enterobacter cloacae (17%), Salmonella spp. (17%), and Escherichia coli (12%). Vibrio isolates included V. alginolyticus (8%), V. parahaemolyticus (5%), and V. fluvialis (3%). Antimicrobial susceptibility tests against 12 drugs showed susceptibility of all K. pneumoniae, C. freundii, Enterobacter cloacae, and Escherichia coli isolates to enrofloxacin, gentamicin, and imipenem. Resistance to ciprofloxacin, chloramphenicol, and trimethoprim-sulfamethoxazole was ≤ 7%. All K. pneumoniae isolates were resistant to ampicillin, and all C. freundii isolates were resistant to cephalothin. Resistance to tetracycline was highest (33%) in E. cloacae, and ≤ 13% in the other three major species. Susceptibility of Salmonella serotypes has been published, and no resistance was seen among any of the isolates.
Conclusion: This study showed that the blue land crabs of Grenada, commonly used as food, can serve as reservoirs of potential human pathogens, and may carry bacteria resistant to antimicrobial drugs used for treatment in human medicine.

Open Access Original Research Article

Review and Follow up Findings on HIV Serodiscordance among Couples in Benue State Nigeria

Odimayo Michael Simidele, Ochejele Silas, Nwadioha Samuel Iheanacho

Microbiology Research Journal International, Page 180-185
DOI: 10.9734/BMRJ/2015/11863

Background: Sexual contact with an HIV infected individual is a significant risk in HIV transmission. We observed occurrence of HIV sero-discordance among some couples in our environment and went further to determine the prevalence of such discordance in Benue state, Nigeria.
Aim: To report HIV discordance rate among couples attending some antenatal care clinics in Benue state, Nigeria in order to stimulate search for factors responsible for such discordance.
Methodology: This study was carried out in collaboration with the Site Coordinators of Prevention-of-Mother-to-Child-Transmission (PMTCT) programs in Benue State, Nigeria. Antenatal care clients attending HIV/AIDS Counseling &Testing were screened for HIV from January 2006 to December 2008. At screening HIV testing was done using paired commercial HIV rapid tests run in parallel. Fourth generation enzyme immunoassays (EIA) HIV kits Manufactured by Alere Medical Company Limited and Trinity Biotech were used. To confirm partner’s positivity, paired rapid-assay were also used. Manufacturers’ instructions were fully followed. Each of the HIV positive patients was requested to come along with her spouse at the next follow up visit for partner counseling and screening. Socio-demographic data were obtained from all subjects. A group of discordant couples were followed up till December, 2012.
Results: A total of 3,508, 5,531, and 4,475 women were counseled and screened annually from 2006 to 2008. HIV positive patients recorded in those years were 15.8%, 17.4% and 46.5% respectively. The peak HIV prevalence occurred in the 21-30 years age group. Among the positive patients, the following percentages accepted partner notification; 53.5% (2006), 72.0% (2007), 67.3% (2008) and came back on consequent follow up visit with their partners while 25%, 15.7%, 47.0% partners accepted to be screened in 2006, 2007 and 2008 respectively. Among the partners screened, there was a sero-discordance of 46% (n=34/74) in 2006. 57% (n=62/109) in 2007 and 45.3% (n= 298/658) in 2008. The highest incidence of HIV sero-discordance occurred among couples < 5 years old in marriage(40%), couples with history of sexually transmitted infections in the past one year(55%) and male circumcision(30% in uncircumcised, 70% in circumcised). Among a cohort of 20 discordant couples consecutively followed up, 80% of the couples were still seen and sero discordant four years after the initial discovery.
Conclusion: We recommend risk-reduction behavior, empowerment of vulnerable groups, effective life planning skills as well as behavioral and cultural change among couples. This group of patients can be a major study point for identification of possible factors responsible for prevention of HIV transmission. These findings should guide prevention interventions in order to achieve maximal impact.

Open Access Original Research Article

Multiple-Antibiotic Resistance in Salmonella enterica Serovars Isolated in Iran Harboring Class 1 Integrons

Bahareh Rajaei, Nahid Sepehri Rad, Farzad Badmasti, Mohamad Reza Razavi, Mohammad Reza Aghasadeghi, Raheleh Saboohi, Taraneh Rajaei, Arfa Moshiri, Seyed Davar Siadat

Microbiology Research Journal International, Page 186-193
DOI: 10.9734/BMRJ/2015/9303

Aims: This research was carried out to detect the content and distribution of class 1 integrons in multidrug resistant Salmonella isolates.
Materials and Methods: Eighty four clinical isolates of Salmonella serovars were subjected to molecular detection of class 1 integrons following the antimicrobial susceptibility test using disk diffusion method and MIC determination.
Results: Eleven isolates (13.1%) which were resistant to at least 4 groups of antimicrobial agents considered as MDR (multidrug resistant) Salmonella serovars. The intI1 gene and internal variable regions (IVRs) of class 1 integron were detected in 50 (59.5%) and 35 (70%) of Salmonella clinical isolates respectively. Analysis of the sequence data revealed four gene cassette arrays including the dhfr7 (0.8 kb), aadA1 (1kb), blaP1 (1.2 kb), dhfr1-aadA1 (1.6 kb) with eight IVR distribution patterns.
Conclusion: Detection of class 1 integron carrying gene cassettes which confer resistance to different classes of antibiotics such as aminoglycosides, ß-lactams and trimethoprim confirms that integron-mediated antimicrobial gene cassettes are prevalent in Salmonella serovars isolated in Iran.