Open Access Original Research Article

Degradation Kinetics of Microbial Consortium Isolated from Diesel Oil Impacted Soil in Delta Park, University of Port Harcourt, Nigeria

Ogochukwu Ann Udume, Enobong Effiong, Gideon Orkwagh Abu

Microbiology Research Journal International, Page 1-15
DOI: 10.9734/mrji/2019/v29i530173

Heavy-duty generators constitute a form of hydrocarbon pollution but enrich microorganisms to having degradative ability of hydrocarbons hence can be used for remediation. In this study a laboratory treatability (biodegradation screening) study was employed to investigate the hydrocarbon degradation competence of bacteria and fungi. Culture-dependent microbiological and physicochemical analyses was conducted on the soil samples obtained from the polluted site. The total aerobic heterotrophic bacterial and fungi counts increased from 5.0-7.5 (Log10 cfu/mL) between day 0-14 then reduced to 6.5 Log10 cfu/mL. The hydrocarbon-utilizing bacteria increased from 4.5-5.2 (Log10 cfu/mL). Percentage degradation hydrocarbons, attributed to fungi bacteria and bacteria/fungi consortia, were 42.3%, 54% and 70% respectively while the control had 6.0%. The total petroleum hydrocarbon (TPH) removal rate (K) was modelled using the first order kinetics:  y=-0.0398x+9.79; K=0.0398d-1. These results, correspond to a degradation efficiency of 70% and t1/2 of 17 days for the bacterial/fungal consortium. The K values for the other setups were 0.019 d-1, 0.0261 d-1 and 0.0022 d-1 with the corresponding degradation efficiencies of 42%, 54% and 6.01% and half-life of 37 days, 27 days and 315 days for fungi, bacteria and control respectively. This result indicates that the use of microbial consortia has high potentials in remediation of hydrocarbons and other pollutants of concern.

Open Access Original Research Article

Isolation, Screening and Characterization of Exopolysaccharide Producing Bacteria

I. G. Nwosu, G. O. Abu, K. O. Agwa

Microbiology Research Journal International, Page 1-9
DOI: 10.9734/mrji/2019/v29i530174

Exopolysaccharides(EPSs) are essential metabolites synthesized and excreted by certain microorganisms in response to extreme condition of pH, temperature, salinity, osmotic stress and other contaminants for survival in such adverse environment. The present study focuses on isolation of exopolysaccharide producing bacteria from extreme environment of oil polluted soil of Ogoni land and marine water of Bonny Island. Screening of EPS producing abilities of the selected isolates were estimated using two approaches viz gravimetric analysis of EPS dry weight and quantification assay for total carbohydrate content by phenol sulphuric acid method. Result revealed that a total of forty (40) different colonies were suspected to produce exopolysaccharide after preliminary screening by selecting thick ropy-like colony formers on agar medium. Secondary screening indicated that twelve (12) isolates produced precipitates above 1000mg/l of total dry weight and eight (8) isolates produced highest EPS yield above 1000mg/l of culture media with carbohydrate content determination. Five isolates with the code WAS1, WAS11, SC6, SOS7 and SOS10 produced significantly higher EPS compared to other bacterial colonies isolated and were termed as most potent EPS producers. These isolates were identified based on 16S rDNA sequence as Providencia stuarti, Escherichia coli, Shewanella chilikensis and Bacillus nealsonii. The result of the present study indicated that these strains have the potentials of producing high exopolysaccharides and can be explored in biotechnological industries.

Open Access Original Research Article

Bioremediation Efficiency of Bacillus amyloliquefaciens and Pseudomonas aeruginosa with the Nutrient Amendment on Crude Oil Polluted the Soil

David N. Ogbonna, Renner R. Nrior, Festus E. Ezinwo

Microbiology Research Journal International, Page 1-13
DOI: 10.9734/mrji/2019/v29i530175

Aim: To assess the Bioremediation efficiency of Bacillus amyloliquefaciens and Pseudomonas aeruginosa strain CL 9 with nutrient amendment using bio-stimulating agents such as Fish waste and Goat manure on crude oil polluted soils in Rivers State, Nigeria.

Study Design: The study employs experimental design, statistical analysis of the data and interpretation.

Place and Duration of Study: A portion of Rivers State University demonstration farmland in Nkpolu-Oroworukwo, Mile 3 Diobu area of Port Harcourt, Rivers State was used for this study. The piece of land is situated at Longitude 4°48’18.50’’N and Latitude 6o58’39.12’’E measuring 5.4864 m x 5.1816 m with a total area of 28.4283 m2. Bioremediation monitoring lasted for 56 days, analysis carried out weekly (per 7 days interval).

Methodology: Seven (7) experimental plots were employed using a Randomized Block Design each having dimensions of 100 x 50 x 20 cm (Length x Breadth x Height) were formed and mapped out on agricultural soil and left fallow for 6 days before contamination on the seventh day; after which it was allowed for 21 days for proper contamination and exposure to natural environmental factors to mimic crude oil spill site. Thereafter bio stimulating agents usually referred to as nutrient amendment organics in this study (fish waste and goat manure) and bio-augmenting microorganisms were applied. Soil profile before and after contamination was assayed while parameters like Nitrate, Sulphate, Phosphate, Total Organic Carbon (TOC) and Total Petroleum Hydrocarbon (TPH), were monitored throughout the experimental period. Microbial analyses such as Total Heterotrophic Bacteria (THB), Total Heterotrophic Fungi (THF), Hydrocarbon Utilizing Bacteria (HUB) and Hydrocarbon Utilizing Fungi (HUF) were recorded. Bioremediation efficiency was estimated from percentage (%) reduction of Total Petroleum Hydrocarbon (TPH) from day 1 to the residual hydrocarbon at day 56 of bio augmented/ biostimulation plots with the control.

Results: Results revealed amount of remediated hydrocarbon and % Bioremediation efficiency at 56 days in the different treatment plots (initial TPH contamination value of  9296.83  mg/kg) in a decreasing order as follows: PS+Bac+Pse+GF+FW (8032.825 mg/kg; 86.40%) >PS+GF+FW (6867.825 mg/kg; 73.87%) >PS+Bac+Pse (6587.825mg/kg; 70.86%) >PS+FW (6441.825mg/kg; 69.29%) >PS+GF (5909.825 mg/kg; 63.57%) >CTRL 2 (Polluted soil without amendment) (3604.825mg/kg; 38.78%). Microbiological results showed increased colonial values with increase time exposure. The results observed on day 56 indicate that Polluted soil + Bacillus + Pseudomonas (10.11 Log10 CFU/g) > Polluted soil but un-amended soil (8.76 Log10 CFU/g) > unpolluted soil (8.68 Log10 CFU/g). Comparatively, Polluted soil +Bacillus + Pseudomonas expressed higher heterotrophic bacteria of 9.77 and 9.67 Log10 CFU/g while fungal counts recorded 6.04 and 6.82 Log10 CFU/g.

Conclusion: Study showed that bioremediation of crude oil-polluted soils with bacteria singly is less effective but a combination with other organic nutrients is a better palliative measure. Therefore, amendment with organic nutrients like Goat manure and Fish wastes is recommended for crude oil polluted soils due to its high nutrient content as substrates for biostimulation of indigenous and augmenting biodegrading microbes. This process could be a source of enhanced natural attenuation of oil-contaminated environments in Nigeria.

Open Access Original Research Article

Effects of Nitrogen and Carbon Sources on Biosurfactant Production by Hydrocarbon-utilizing Stenotrophomonas sp.

Victor Ezebuiro, Ipeghan Jonathan Otaraku, Boma Oruwari, Gideon Chijioke Okpokwasili

Microbiology Research Journal International, Page 1-10
DOI: 10.9734/mrji/2019/v29i530177

Aim: This study investigated effects of nitrogen and carbon sources on the production of biosurfactant by a hydrocarbon-utilizing bacterium, Stenotrophomonas sp.

Methodology: The hydrocarbon-utilizing bacterium was isolated with Bushnell Haas (BH) broth using enrichment method. Biosurfactant production was screened by evaluating the following characteristics: Emulsification index (E-24), oil spreading (displacement), tilted glass slide, haemolysis on blood agar, and lipase production. Effects of combination of nitrogen sources (yeast extract and NH4NO3, yeast extract and urea, yeast extract and asparagine, yeast extract and peptone, NaNO3 and peptone, NaNO3 and asparagine, and yeast extract and NaNO3) and carbon sources (glucose, fructose, galactose, cassava peel, soya bran, olive oil, sucrose, crude oil, diesel and glycerol) on biosurfactant production were determined with emulsion stability and surface tension as responses. The bacterium was identified based on phenotypic, microscopic, and biochemical characteristics.

Results: The isolate produced colonies on BH agar containing either naphthalene or hexadecane as sole source of carbon after 48-h incubation. Screening characteristics for the production of biosurfactant by the isolate were as follows: 46% emulsification index, 3.1 cm2 oil displacement, 1.8 cm zone of clearance on tributyrin agar, γ-haemolysis, and positive tilted glass slide. The best carbon source with the highest emulsion stability (51.6%) was fructose whereas the best surface tension reduction (30.85 mN/m) was observed with olive oil as carbon sources after 7 days of incubation. For nitrogen, the combination of yeast extract and NH4NO3 gave the highest emulsion stability (60.7%) and the best surface tension reduction (39.58 mN/m). The data obtained were significant at P<0.05 and the bacterial isolate identified as Stenotrophomonas sp.

Conclusion: This study has demonstrated the ability of the hydrocarbon-utilizing bacterium, Stenotrophomonas sp. to produce biosurfactant, indicated by reduction of surface tension and formation of stable emulsion. This method of biosurfactant production can be further scaled up for industrial purpose. 

Open Access Original Research Article

Isolation, Identification and Antifungal Susceptibility of Dermatophytes Isolated from Clinically Suspected Cases of Tinea Infections in Pakistan

Shumaila Shakir, Sidrah Saleem, Wajhiah Rizvi, Abdul Waheed, Javid Iqbal

Microbiology Research Journal International, Page 1-11
DOI: 10.9734/mrji/2019/v29i530178

Dermatophytosis or tinea is a type of cutaneous infection caused by keratinophilic fungi, infecting the skin, nails and hair. A correct diagnosis is important for epidemiological purposes and initiating appropriate treatment. An increase in the prevalence of fungal infection worldwide is due to abuse of antibiotics, immunosuppressive treatments and numerous medical conditions.

Aim: To isolate, identify, and examine the in-vitro antifungal susceptibility of dermatophytes in clinically suspected cases of tinea infections.

Methodology: After taking informed consent, we took 65 patients suspected of tinea infection and obtained samples from skin, hair and nail, under aseptic precautions, at Department of Microbiology, University of Health Sciences (UHS), Lahore, Pakistan. The identification of dermatophytes was performed using potassium hydroxide (KOH) mounts and culture on Sabouraud Dextrose Agar (SDA) and Dermatophyte Test Medium (DTM). The cultures were incubated at 30ºC for up to 4 weeks in case of SDA and 2 weeks in case of DTM. Lactophenol cotton blue (LCB) stain was used to identify the species morphologically. Susceptibility test was done by agar diffusion method using antifungal disks and zones of inhibition were measured.

Results: More females (55.38%) than males (44.61%) were observed in the study. Most of the cases belonged to the age categories of 1-10 years and 21-30 years. Tinea corporis was the most common clinical type found (27.69%) followed by Tinea capitis (21.53%) and Tinea cruris (12.30%).Trichophyton mentagrophytes was the commonest species isolated (32%)  followed by Trichophyton violaceum (28%) and Trichophyton rubrum (12%). Terbinafine was seen to be the most effective drug against the isolates, followed by clotrimazole. Fluconazole showed least activity.

Conclusion: Fungal culture remains the gold standard in identifying the causative species. Terbinafine promises to be a potent antifungal, whereas fluconazole has low efficacy against such organisms. Disk diffusion method adopted for antifungal susceptibility is cost effective and easily performable in small laboratories not having an established mycology bench.