Theory and Applications of Microbiology and Biotechnology Vol. 1

An aerosol is a suspension of solid or liquid particles in air or other gaseous environment. Sources of bacterial aerosols exist within and outside the dental clinic. The generation of bacterial aerosols and splatters appears to be highest during dental procedures. The use of rotary dental and surgical instruments and air-water syringes generates visible infectious spray, that enclose large-particle spatter of water, saliva, microorganisms, blood, and other debris. Several infectious diseases could be transmitted to staff and patients by airborne bacterial and other contaminants in the dental clinic. The vigilant use of barriers along with appropriate immunizations procedures could safe guard the dental fraternity from the ill-effects of the aerosols.

Two wastes from animal origin, viz., human hairs and egg shells were evaluated for different manifestations of white oyster mushroom, Pleurotus florida. The mushroom utilized both the substrates for their growth and sporophore formation. The mycelial growth was significantly (P=0.05) faster on egg shell (18 days) as compared to human hair (23 days). The crop of mushroom was harvested in four flushes where human hairs showed higher yield and biological efficiency of mushroom (165 gm, 33%) than egg shells (155 gm, 31%), respectively. In respect of yield parameters such as yield, biological efficiency, number of mushroom fruit bodies and average weight of sporophores, both the substrates were statistically at par to each other. Utilization of human hairs and egg shells by P. florida reveals a new strategy for mycoremediation of these wastes.

This research work aimed to present the activity guided fractionation with antimicrobial evaluation from both crude extract and various fractions obtained from Pergularia tomentosa L. whole plant. The P. tomentosa L. whole plant was extracted with 95% aqueous ethanol; fractionated into acidic, basic, polar and nonpolar fractions. All fractions with the crude extract were screened for both antimicrobial and minimum inhibitory concentration (MIC) potentialities. For crude extract, all concentrations (1.5, 0.75, 0.35, & 0.168 mg/ml) indicated marginal antibacterial activity with range of 17, 20, 14 mm zone of inhibition for S. aureus, E. coli and C. albicans. While, both basic fraction showed highest activity against E. coli and C. albicans at 15 mm & 15 mm; along with acidic and methanolic fraction haven large spectrum against S. aureus, E. coli and C. albicans at 13, 12, 12 mm. Moreover hexane did not showed antimicrobial activity for both S. aureus and C. albicans except for E. coli which showed activity at 12 mm. The study clearly indicated that basic fraction showed highest antimicrobial activity for selected micro-organisms with lower minimum inhibitory concentration which ranges from 18.75 µg/ml to 150 µg/ml. Followed by wider spectrum of antimicrobial activity for acidic and methanolic fraction against all tested organisms with minimum inhibitory concentration from 75 µg/ml to 150 µg/ml; while 300 µg/ml (MIC) stand for hexane fraction. Thus, P. tomentosa L. particularly, the basic fraction (alkaloid) and, both acidic and neutral fractions will be centered areas for further scientific research findings in isolating an active antimicrobial component therein.

Fungi are potential profounding cause of aeroallergens which are creating a appalling scenario culminating into various fatal diseases to man, livestock and plant biodiversity such asperglosis, mycosis and as smut and rust, respectively. In the present study, air-borne mycoflora of Baghsar Fort Samahni (BFS) and its allied forest (Azad Kashmir, Pakistan) was spatially and temporally analyzed. Spores were collected from five different sampling sites (S.1 S.2, S.3, S.4, S.5) by air-trapper and grown on two different culture media viz. potato dextrose agar (PDA) and water agar (WA), and isolated by using culture plate technique (CPT). A total of seven different fungal species of six genera namely Aspergillus niger, Alternaria solani, Cladosporium sp., Fusarium sp., Penicillium sp. and Curvularia sp. were isolated and identified. On PDA medium, Cladosporium sp. was found to be the most abundant (21.3%) fungal species followed by A. niger (17.8%), A. solani (15.4%), Alternaria alternata (14.9%), Fusarium sp. (13.3%), Penicillium sp. (9.0%) and Curvularia sp. (8.3%). In other WA medium, Cladosporium sp. was again dominating and most prevalent (23.2%), and other species occurrence was as: A. niger (19.2%), A. solani (18.1%), A. alternata (14.1%), Fusarium sp. (11.3%), Curvularia sp. (8.5%) and Penicillium sp. (5.6%). PDA proved a better medium for capturing and perpetuation of fungi. High incidence of Cladosporium spp. is a major cause of fruit rots and immature seed falling and ultimately leads to economic loss to the area.

Aims: This work aimed to determine the toxicity of Bauhinia monandra leaf lectin (BmoLL) to mice and Artemia salina.

Study design: BmoLL was purified from the leaves of B. monandra through affinity chromatography and tested against mice and A. salina in order to evaluate its toxicity.

Methodology: The lectin purification was performed through saline precipitation and chromatography onto a guar gel column. The acute toxicity of BmoLL was evaluated using female mice during 14 days. Toxicity of BmoLL to A. salina larvae was also evaluated.

Results: BmoLL did not induce death or weight loss of mice at 2000 mg/kg b.w.; additionally, the lectin did not affect the survival of A. salina at the assayed concentrations (250–1,000 µg/mL).

Conclusion: BmoLL is an interesting molecule to investigate pharmacological applications due to its low toxicity according to the two models used in this work.

The purpose of this study was to detect biofilm formation and to examine the correlation between biofilm and Metallo-β-lactamases (MBL) production in Pseudomonas aeruginosa. A total of 64 P. aeruginosa isolates were identified using standard microbiological methods and antimicrobial susceptibility testing (AST) was performed on them according to Clinical and Laboratory Standards Institute (CLSI) guidelines. The isolates were screened for biofilm production using both qualitative and quantitative methods. The presence of MBL genes were checked by multiplex PCR assay.  Out of all 30 meropenem resistant P. aeruginosa, 2 isolates were found producing all the three genes (i.e. blaIMP, blaVIM, blaSIM) for MBL production and they were found to produce biofilm. Resistant to four antibiotics such as aztreonam (85.7% vs 11.1%, P< 0.000), Cefepime (82.1% vs 2.8%, P<0.000) gentamycin (82.1% vs 27.8%, P< 0.000) and Pipercillin/Tazobactum was also high (28.6% vs 2.8% P< 0.003) was comparatively higher among biofilm producers than non biofilm producers. In biofilm production, both qualitative method and quantitative plate method showed 16 isolates (53.3%) as biofilm producers for MBL genes. Out of these 16, only 9 isolates showed MBL production along with biofilm production having significant association (P<0.004).

The prevalence of MBLs has been increasing worldwide, particularly among P. aeruginosa, leading to severe limitations in the therapeutic options for the management. Presence of MBL genes has a role in inducing biofilm production and significant association in P. aeruginosa isolates. Overall, drug resistance was found to be more in biofilm producing isolates than non biofilm isolates.

Aims: This work investigated the population density of endophytic microorganisms from Moringa oleifera leaves collected in three localities at the State of Pernambuco (northeastern Brazil): Urban (campus from the Federal University of Pernambuco, UFPE) and forest (botanical garden) areas at Recife city and an urban area (industrial district) at Caruaru city.

Place and Duration of Study: Department of Antibiotics and Department of Biochemistry from Federal University of Pernambuco, during four years.

Methodology: Sodium hypochlorite was used to disinfect the leaves, which were macerated in PBS buffer and separately sowed on seven culture media supplemented with antibacterial or antifungal agents.

Results: The majority of endophytes isolated were bacteria and the highest density was found in leaves from the forest area. Regarding to fungi isolation, there was no statistical difference between the density in leaves from UFPE campus and the botanical garden while no fungal isolates was obtained from leaves collected in Caruaru. The highest diversity of endophytes was found in the leaves from the botanical garden, with 111 different isolates. A total of 71, 94 and 50 bacterial isolates were obtained from leaves of UFPE campus, botanical garden and Caruaru, respectively. The number of fungal isolates were 17 (UFPE campus) and 12 (botanical garden).

Conclusion: In conclusion, the methodology employed in this work was effective for the isolation of endophytes; it is still worth to mention that climatic and geographical conditions may interfere in density and diversity of endophytes from M. oleifera leaves.

Staphylococcus aureus is considered an infectious agent of great clinical importance, responsible for many different types of infection. Strains of methicillin-resistant Staphylococcus aureus (MRSA), Panton-valentine leukocidin producers, are considered more invasive, presenting clinical sequelae related to abscesses and infection in skin and soft tissues. The use of invasive techniques in hospital environment, such as the introduction of intravascular catheter in immunocompromised patients, has contributed to this microorganism spreading through the bloodstream, causing bacteremia, necrotizing pneumonia and increasing the number of septic patients in intensive care units with high mortality. In this report, atypical infections in Swiss mice using experimental model of sepsis was presented.

Introduction: Many drugs have been isolated from fungal species. This study aims at identifying fungal species isolated in Anguilla to determine the antimicrobial effect of their secondary metabolites from pure culture against Staphylococcus aureus by demonstrating the presence of a zone of inhibition in the culture plate.

Methods: Samples were cultured and sub-cultured to isolate pure culture on potato dextrose yeast agar (PDYA) and chosen for further studies by the presence of surface exudates. Those cultures that produced copious amounts of surface exudates were examined for antimicrobial effects by further testing.

Results: Antimicrobial testing of MB0725C (P. digitatum) samples did not result in any evidence of antimicrobial properties. However, MB0725A (P. chrysogenum) punch biopsy-agar overlay produced an 11 mm ZOI, whereas crude exudates testing resulted in a 27 mm ZOI. Crude culture filtrate of potato dextrose broth (PDB) did not result in any ZOI for either MB0725A or C. Sensitivity testing on samples collected from Yeast Extract Lactose Broth (YELB) on Day 3, Day 6, Day 9 and Day 12 resulted in ZOI of 11 mm, 13, 15 and 17 mm respectively. The change in pH for MB0725A liquid culture in PDB versus YELB was significantly different (N=12, P<0.0001).

Conclusion: MB0725A was an excellent producer of surface exudates and further experiments showed that its secondary metabolites had antimicrobial effects against Staphylococcus aureus using Kirby-Bauer sensitivity testing.

Aim: The present study was designed to evaluate the antibacterial activity of Mentha piperita (L) leaf extract against clinical isolates of urinary tract infections.

Introduction: M. piperita L. (Peppermint) is a strongly scented herb belonging to family Lamiaceae. The plant is stimulant, aromatic and used for headache, vomiting and allaying nausea. In India the leaves are used to relieve sore throat. The most common form of bacterial infections is urinary tract infections (UTIs). They affect people of all age groups throughout their lifespan.

Methodology: The M. piperita ethanolic extract (MPEE) was prepared by cold maceration. The presence of phytoconstituents was determined using standard protocols. Clinical isolates of UTI pathogens such as Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa were isolated from urine samples and identified by biochemical tests. The antibacterial property was determined by agar well diffusion method.

Results and Discussion: The preliminary phytochemical screening revealed the presence of amino acids, carbohydrates, alkaloids, proteins, glycosides, steroids, tannins and flavonoids. MPEE exhibited pronounced antibacterial activity against tested microorganisms. The extract exhibited antibacterial activity at 1000 µg concentration against S. aureus (21.50±1.22 mm), E. coli (19.33±0.81 mm), P. aeruginosa (15.33±1.69 mm) from high to low respectively. The minimum inhibitory concentration was ranged from 62.5 to 125 mg/ml.

Conclusion: The results of antibacterial studies confirm that MPEE was found to contain bioactive constituents that exhibited notable antibacterial activity. However, further isolation and characterization of phytoconstituents will be needed to evaluate the antimicrobial activities against a wider range of microbial pathogens.

Sulfate reducing bacteria (SRB) was successfully isolated from Estuary Dam in Suwung Denpasar, Indonesia. This estuary catches highly polluted water from Badung River which runs across and hence carries pollution due to waste disposal from Denpasar City. SRB was studied in detail for their ability to reduce sulfate to sulfide with organic material as an oxidizing agent. SRB exploration of the estuary ecosystem of the contaminated dam was accomplished through isolation, selection and characterization of the isolates obtained. The result of this study found superior SRB named DPS 1711, DPS 1705 and DPS 1703. The bacteria have the ability to grow at pH 3, room temperature and uses compost as organic substrate. This ability is an important factor for the application of isolates in the treatment of acid mine waste. Isolates have optimum optical density under the pH range of 4 to 7 and the best at pH 5 have a growth rate profile at a temperature range of 25 to 40°C. The isolates observed were Gram-negative stem, motile bacteria which only grow in anaerobic condition. Physiological-biochemical characterization showed the three isolates, namely DPS 1703, DPS 1705 and DPS 1711 were SRB groups identified as Desulfotomaculum orientis.

Biodegradation of hydrocarbons by microorganisms represents one of the primary mechanisms by which petroleum and other hydrogen pollutants are eliminated from the environment. This work was carried out on the effect of microorganisms on the biotreatment of oil in crude oil contaminated soil.

Microorganisms were isolated from two experimental soil samples contaminated with Bonny Crude and normal uncontaminated soil as a control over a period of seven months. The microbial as well as the physico-chemical parameters of the soil samples were all analyzed using standard methods. Changes in total petroleum hydrocarbon level were measured appropriately. Treatments used were the microbial isolates.

Forty-four microorganisms were isolated from the contaminated soils and identified as species of Pseudomonas (7), Flavobacterium (6), Bacillus (8), Proteus (4), Klebsiella (1), Pencillium (5), Aspergillus (7), Fusarium (3), Trichypton (2) and Neurospora (1). Ten of the forty-four isolates had ability to degrade crude oil in the laboratory. On contamination a value of 1.0X10⁵ cfu/g in microbial counts were obtained followed by a subsequent increase in population levels after a period of 2months with a value of 1.0X10⁶ cfu/g. Oil application to the soil resulted in an increase in total petroleum hydrocarbon from 0.31 ppm to 5.53 ppm; organic matter from 0.41% to 7.34%; available phosphorus from 1.75 ppm to 2.84 ppm. The treatment measures all showed progressive decrease in oil concentration in the soil. Mixture of bacterial and fungal isolates as a treatment measure proved to be more favourable above all others, it brought the concentration from 5.53 ppm to 0.31 ppm after a period of 5 weeks of treatment, which is same value with the normal soil (uncontaminated).

Species of Pseudomonas, Bacillus, Flavobacterium, Proteus, Klebsiella, Penicillium, Aspergillus, Fusarium, Trichyphyton and Neurospora had potential for the degradation of bonny crude oil. They could therefore be employed in environmental cleanup of petroleum spill site.

Urinary Tract Infection (UTI) is one of the commonest infectious disease in pregnancy, and in pregnancy we have very limited number of antibiotics to treat the UTI. This study was conducted on 151 patients who attended the gynecology clinic during the study period. Nineteen UTI proven cases of UTI were studied for prevalence of microorganism and sensitivity pattern against different antibiotics. Among the bacteria isolated, Escherichia coli (73.68%) and Staphylococcus aureus (10.52%) were the most prevalent Gram negative and Gram positive bacteria respectively. To know the resistance pattern of microorganism we used commercially available discs of different antibiotics. Gram negative bacteria showed more resistance as compared to Gram positive one. It is observed that the most effective antibiotic for Gram negative isolates is Ceftriaxone (87.5%), followed by Amoxicillin + Clavulanic acid (81.25%), Amikacin (75%), Cefuroxime (75%), Cefixime (68.75%) and Mezlocillin (62.5%). For the Gram positive bacteria, Ceftriaxone, Amikacin and Amoxicillin + Clavulanic acid were the most effective antimicrobials (100%). Multidrug resistance Gram negative bacteria were also tested for Extended-spectrum Beta Lactamase (ESBL), 35.71% of E. coli isolates were ESBL producer.

Aim: Potential risk factors and prevalence associated with Helicobacter pylori (H. pylori) infection in apparently healthy children in Nigeria were studied.

Study Design: To investigate the current potential risk factors associated with recent prevalence of H. pylori in apparently healthy children in Nigeria.

Place and Duration of Study: The study was conducted in two Local Government Areas, Alimosho and Ajeromi, of Lagos State, Nigeria between March and September 2014.

Methodology: Seroprevalence status of 185 asymptomatic children made up of 93 males and 92 females, aged between 2-16 years were selected by randomized stratified sampling with descriptive questionnaire. Serum immunoglobulin G H. pylori antibody of the individual subjects was determined using DiaSpot H. pylori kit while fecal samples of same group were analysed for HpSAg using immunoassay test kit of Helicobacter pylori Stool Antigen (HpSAg).

Results: Of 185 children tested for H. pylori antigen, 134 (68.7%) and 51(26.2%) were classified as seropositive and fecal HpSA positive respectively. Highest rate of 40.0% and 34.6% of the children weighing between 21 and 40 kg were positive while 29.2% and 32.5% children of parents that were traders were positive to serum H. pylori antigen and fecal HpSA respectively. Only 12.4% and 14.1% children from artisan parents were positive but different age group have no association with the infectivity or prevalence of fecal H. pylori antigen (OR=0.67, CI=0.142-0.152). Significant higher percentage of seropositivity of 59.0% and fecal positivity of 55.7% was recorded among children from 5-8 people in a room (p>0.05), while Households with regular potable water supply have lower H. pylori seropositivity and fecal positivity of 11.9% and 7.6% compared with households that sometimes have water supply. The Households that never had water supply had highest number of seropositivity of 40.0% and 18.4%, respectively. Sewage nearness to kitchen indicates 30.8% and 28.7% H. pylori seropositive and fecal positivity rate among children.  

Conclusion: Paediatric H. pylori prevalence is highly associated with water borne infection and poor sanitary practices. There is need for achievable interventions and improvement in environmental sanitation.

Growth response profile of three species of Pseudomonas isolated from pesticide contaminated soils within Uyo, Nigeria was studied using standard microbiological and analytical techniques. The ability of the isolates to tolerate varying concentrations of commercially available pesticides namely; Decis, DD force and Cyperforce was assessed over a 96 hour period. Selective enrichment cultures with graded concentrations of the pesticides were used to assay their growth response profile and the absorbance determined using CO75 digital colorimeter. The results showed that the Pseudomonas species differed biochemically. Their growth response at pesticide concentrations 0.0001, 0.001, 0.01, 0.1, 1 and 10% v/v differed significantly (P<0.05) at 24 hours interval for four days. At 10% concentration of Decis pesticide over 96 hour, the absorbance were 0.20, 0.23 and 0.30 for Pseudomonas from Agriculture research farm, hospital dumpsite and municipal waste dump site respectively. This ability therefore offers a veritable tool for use in the bioremediation and ultimate restoration of pesticide contaminated soils but however requires further evaluation.

Vegetables are usually consumed raw. This implied best hygienic conditions from the harvest to the processing because of the gastro-enteritis that they could provoke. This study was conducted with the aim to appreciate microbiological quality of raw tomatoes, endives and ready-to-eat products sold in markets. Samples were taken randomly in two markets of Abidjan. A microbiological analysis was done in order to identify and enumerate fecal coliforms, Escherichia coli, Enterococcus and Pseudomonas. A decontamination treatment based on washing samples with running water and sodium hypochlorite solution I° chlorymetric was also applied to tomatoes and endives. The results indicated that, for tomatoes and endives, the average load was 1.5.10⁴ CFU/g of Enterococcus, 1.3.10³ CFU/g of Pseudomonas and 1.7.10² CFU/g of fecal coliforms. In ready-to-eat products, the load was 9.3.10¹ CFU/g for Enterococcus, l.03.10¹ CFU/g for Pseudomonas and 9.9.10¹ CFU/g for fecal coliforms. The disinfection with a sodium hypochlorite solution l° chlorymetric reduced Enterococcus and fecal coliforms load to 98% and Pseudomonas load to 97% as compared to the washing with running water in which Enterococcus was only reduced to 80%, fecal coliforms to78% and Pseudomonas to 73%. Escherichia coli were isolated in 28 samples as follow: 15 stumps from endives (54%), 10 stumps from tomatoes (36%) and 3 stumps from ready-to-eat products (10%). Results showed that before consumption, vegetables need to be washed, cleaned and disinfected. This will avoid sanitary hazard.