The Influence of Agitation Rates, pH and Calcium Carbonate on L-lysine Production by Bacillus subtilis using Agricultural Products as Carbon and Nitrogen Sources

L-Lysine is an essential amino acid that is required in the diet of humans and animals. It is utilized in human medicine, cosmetics and pharmaceutical industry. ’The influence of agitation rates, pH and calcium carbonate on L-lysine production by Bacillus subtilis using agricultural products as carbon and nitrogen sources was studied. The L-lysine-producing bacteria had already been isolated from Nigerian soil. They were purified and Identified as B. subtilis PR13 and B. subtilis PR9, using cultural, biochemical and molecular characteristics. Optimization of some parameters which included agitation rates, pH values and CaCO3 concentrations, on L-lysine production by the Bacillus species was carried out. The L-lysine was produced in 250 ml flasks containing fermentation media (FM1 and FM2). The findings revealed that, enhanced L-lysine yield of 2.10 and 1.33 mg/ml was observed at agitation rate of 180 rpm for B. subtilis PR13 and PR9 respectively. There was a positive correlation between agitation rates and L- lysine production by B. subtilis PR13 and PR9 (r = 0.96 and 0.83 respectively). The pH of 7.5, stimulated optimum L- lysine yield of 2.27 mg/ml for PR13 and 1.38 mg/ml for PR9. There was a positive correlation between pH values and L-lysine production by B. subtilis PR13 and PR 9 (r = 0.63 and 0.50 respectively). The supplementation of 40g/l of CaCO3, enhanced optimum L-lysine yield of 2.18 mg/ml for B. subtilis PR 13 and 1.30 mg/ml for B. subtilis PR9. There was a positive correlation between varying concentrations of calcium carbonate and L-lysine production by the B. subtilis PR13 (r =0.35), while negative correlation was observed for B. subtilis PR 9 (r = -0.10). The results obtained in the study illustrated that the optimization of process parameters could increase the L-lysine yield from agricultural products by B. subtilis PR13 and B. subtilis PR9.

Protease Production by Submerged Fermentation in Shake Flasks Using Bacillus sp. Isolated from the Soil

Proteases are one of the most industrially important enzymes, which account for about 60% of total enzyme market. Protease production by submerged fermentation in shake flasks using Bacillus sp. isolated from the soil was studied. Soil samples were collected from different locations within Chukwuemeka Odumegwu Ojukwu University, Uli, Anambra state. The soil samples were serially diluted and inoculated on sterilized skim milk agar plates. The plates were incubated at 30oC for 72 h. A clear zone around the colonies gave an indication of protease-producing bacteria isolates. The selected protease producers were subsequently used for shake flask fermentation in 50 ml sterile medium. Optimization study was conducted to determine the effect of carbon sources, nitrogen sources, trace elements, agitation rates and pH. Twenty one bacteria isolates were found to be active protease producers and isolates RS-5 and OS-9 had the highest zone of clearance of 13.5 and 12.1 mm respectively. The result of submerged production of protease by the bacteria isolates revealed that the isolates RS-5 and OS-9 accumulated maximum protease yield of 3.23 and 2.71 U/ml respectively. The isolates were Gram positive and endospore formers, and were identified as Bacillus sp. RS-5 and OS-9.The addition of Starch and maltose stimulated optimum protease production of 3.47 and 2.77 U/ml by Bacillus sp. RS-5 and OS-9 respectively. Beef extract enhanced maximum enzyme yield of 3.35 and 2.90 U/ml for Bacillus sp. RS-5 and OS-9 respectively. Maximum protease yield of 3.28 U/ml for Bacillus sp. RS-5 and 2.85 U/ml for Bacillus sp. OS-9 was obtained by the supplementation of 0.4 g/l of FeS04 respectively. The maximum protease yield was observed at agitation rate of 200 rpm for Bacillus sp. RS-5 and 170 rpm for Bacillus sp. OS-9. At pH8, protease accumulation was highest for Bacillus sp. RS-5 and OS-9. The study revealed that the soil harbours some protease-producing bacteria strains and protease production can be greatly enhanced through optimization of process parameters.

Amylase Production by Solid State Fermentation of Agro-industrial Wastes Using Bacillus species

This study evaluated amylase production by Bacillus species employing the solid state fermentation (SSF) method using five agro-industrial wastes namely corn cobs, potato peel and maize straw, groundnut husk and corn chaff. Five Bacillus species were tested for amylase production abilities and Bacillus subtilis showed the highest amylase production ability after incubation. Corn chaff gave maximum enzyme production (3.25 U/ml) while the least enzyme was recorded on groundnut husk (2.35 U/ml) at 25. Potato peel had maximum enzyme production by Bacillus subtilis (3.05 U/ml) at pH 7.0 while the least enzyme production was from groundnut husk (2.84 U/ml) at pH 4.0.Thus there was an increase in enzyme production with corresponding increase in substrate concentration. The results obtained in this study support the suitability of using agro-industrial wastes as solid state fermentation substrates for high production of amylase. It’s also a means of solving pollution problems thus making solid state fermentation an attractive method.

Screening and Characterisation of Keratin-Degrading Bacillus sp. from Feather Waste

Aim: This study focuses on the screening and characterisation of keratin-degrading Bacillus species from feather waste. Methods: Nine bacteria were isolated from feather waste obtained from a poultry layout at Egbeda local government secretariat, Ibadan, Nigeria. These bacteria were grown in basal medium with feather as primary source of carbon, nitrogen, sulfur and energy. Feather degrading bacteria were screened for both proteolytic activity and keratin degradation on skimmed milk agar and keratin azure medium respectively. They were also screened for their ability to degrade other keratin substrates such as hair and nail. Results: Three of the isolates with higher feather degradation levels also showed high proteolytic activity and release of azure dye. They were selected and identified phenotypically and genotypically using 16S rRNA sequencing as Bacillus licheniformis-K51, Bacillus subtilis-K50 and Bacillus sp.-K53. The bacteria were capable of degrading other keratin-containing substrates such as nail and hair. Bacillus subtilis-K50 and Bacillus licheniformis-K51 showed significant difference (P) in degradation among the three different keratin sources used yielding higher degradation with feather as keratin source with respective optical densities of 0.07 and 0.11 followed by hair and least in nails with optical densities of 0.05 and 0.07 respectively. Highest degradation of all the three keratin substrates was observed in Bacillus licheniformis-K51. Conclusion: The three isolated bacteria possess the ability to degrade keratin and utilize feather as keratin substrate. As a result, these can be considered as potential candidates for degradation and utilization of feather keratin.

Preliminary Detection of Bacillus species in Commercial Honey.

Aim: This research is aimed at detecting the presence of Bacillus species in honey using morphological characteristics, biochemical tests and preliminary molecular studies. Place and Duration of Study: The study was carried out in the department of Molecular Biology, Institute of Medical Research Yaba Lagos, Nigeria. The study was carried out from June to July 2012. Methodology: A total of 33 honey samples were used for this study, twenty- eight of the honey samples were of local origin while 5 were of international origin. Twenty-eight commercial honey samples were obtained from the six geographical regions in Nigeria from commercial retailers. The five foreign honey samples were obtained from the supermarkets namely: Friz fruit, Blossom, Forever, Aloe Vera and Rose honey, all of international origin. The honey samples were inoculated into sterile agar, blood and tryptone soy plates using the spread plate technique. Isolates obtained were purified and subjected to morphological tests, biochemical tests and further identification using polymerase chain reaction. Results: All the honey samples had microbial growth in them, higher counts were observed in the commercial honeys from retailers than the foreign honey samples. Forty isolates suspected to be Bacillus from biochemical tests were subjected to PCR, 14 from the 40 were confirmed to be Bacillus spp. Conclusion: Microorganisms in honey cannot be identified fully with morphological and biochemical examinations alone, but combine use of morphological, biochemical tests and PCR technique is more accurate and reliable method of identification.

Production of peptide antibiotics by Bacillus sp: GU 057 indigenously isolated from saline soil

A total of 112 soil samples were taken from differents areas of district D.I.Khan and Kohat (KPK) Pakistan and screened for production of antibiotics against the Micrococcus luteus and Staphylococcus aureus. Widest zone of inhibition (18mm) was produced by microorganism isolated from saline soil. The strain was later identified as Bacillus GU057 by standard biochemical assays. Maximum activity (18mm inhibition zone) was observed against Staphylococcus aureus after 48 hours of incubation at pH 8 and 4% concentration of glucose. The antibiotic was identified by autobiography as bacitracin. The Bacillus strain GU057 was confirmed as good peptide antibiotic producer and can effectively be indulged as biocontrol agent.

A Case of Bacillus licheniformis Spondylitis and Bacteremia in a Patient with Lung Cancer

Bacillus licheniformis is an aerobic, gram-positive, spore-forming rod bacteria usually found in the environment. Infections with B. licheniformis are rare and usually associated with an immunocompromised state, trauma, and an indwelling catheter. We report a case of bacteremic B. licheniformis spondylitis following vertebroplasty in a patient with lung cancer.

Xylanase production by a thermo-tolerant Bacillus species under solid-state and submerged fermentation

Effects of xylose on xylanase production by a thermophilic Bacillus sp showed diverse patterns on corn cob (CC) and wheat bran (WB) as sole carbon sources in solid- state fermentation (SSF) and submerged fermentation (SmF). Supplementation of these media with either mineral salt solution (MSS) or yeast extract peptone (YEP) also exerted variable effects. While under SSF, xylose stimulated xylanase synthesis by 44.01 percent, on wheat bran supplemented with MSS, it decreased the enzyme activity by 12.89 percent with YEP supplementation. In SmF, however the enzyme synthesis was stimulated by xylose on supplementation with both MSS and YEP by 41.38 percent and 27.47 percent, respectively. On corn cob under SSF, xylose repression was significant both with MSS (26.92 percent) and YEP (23.90 percent) supplementation. Repression by xylose also took place on corn cob and YEP (19.69 percent) under SmF, while significant stimulation (28.55 percent) was observed by MSS supplementation. The possible role of media composition and fermentation conditions in the regulation of xylanase synthesis by xylose is discussed.

In Vitro Antagonistic Characteristics of Bacilli Isolates against Trichoderma spp. and Three Species of Mushrooms

Twenty isolates of Bacillus species obtained from livestock manure composts and cotton-waste composts were tested for their antagonistic effects in vitro against three green mold pathogens of mushrooms (Trichoderma harzianum, T. koningii, and T. viridescens). However, there exists a possibility Bacillus species may have antagonistic effects against mushrooms themselves, and thus the same 20 isolates were tested in vitro against three species of mushrooms (Flammulina velutipes, Lentinus edodes, and Pleurotus ostreatus). Of the 20 Bacillus species isolates tested, two inhibited mycelial growth of T. harzianum, seven that of T. koningii, and eight that of T. viridescens. Importantly, the bacterial isolates M27 and RM29 strongly inhibited mycelial growth of all the Trichoderma spp. isolates tested. The isolate M27 was subsequently identified as the most effective in inhibiting mycelial growth of all the Trichoderma species. Interesting results of the effect Bacillus isolates had upon the mushroom species followed. It was found that most Bacillus isolates except 5T33 at least somewhat inhibited mycelial growth of the three mushroom species or some of the mushrooms. Furhermore, the antagonistic effects of the bacterial isolates against the three species of mushrooms varied depending on the mushroom species, suggesting a role for mushroom type in the mechanism of inhibition. The bacterial isolates M27 and RM29 were identified as having the most antagonistic activity, inhibiting mycelial growth of all the Trichoderma spp. as well as mycelial growth of the three species of mushrooms. These results suggest that the bacterial isolates and their antagonistic effects on green mold pathogens should be further studied for their practical use for biological control of green mold in the growing room of the mushrooms.

A cross sectional study of microbial contamination of medical students’ white coat

The objective of this study is to determine the incidence of microbial contamination on medical students’ white coats, the way they handle and clean their white coats and their perception towards contamination. For this purpose, cross sectional survey of the bacterial contamination of white coats in a medical college has been carried out in 3 different locations; Royal College of Medicine, Perak, University of Kuala Lumpur and a private college attached to Ipoh General Hospital. It was found that the incidence of Staphylococus aureus, was 32% on short-sleeved and 54% on long-sleeved white coats. Bacillus species was the second most common type of bacteria found. Male collars and female pockets had higher microbial contaminations (p=0.01, 0.03 respectively). Clinical students’ white coats were significantly less contaminated than non-clinical students (p=0.001) although they tend to wear it for a longer period (5.75 ± 2.19 h vs. 2.32 ± 0.81 h) (p=0.001). Clinical students owned more short-sleeved coats (p=0.001) and washed their coats more often (p=0.01) than non-clinical ones. More than eighty one percent of clinical students wear their white coats in the college the majority of whom were females (p=0.005). Perception of clinical and non-clinical students towards white coat contamination was similar. Medical students’ white coats are contaminated with bacteria and they are potentially source of cross infection. Student’s way of handling and washing white coats should be corrected by issuing and following standard guidelines. Students should be bared from wearing white coats in non-clinical areas. Washing hands and using plastic aprons is highly recommended before examining wounds

Regulation and structure of aspartokinase in the genus Bacillus.

The aspartate pathway of amino acid biosynthesis in bacteria serves as paradigm for the evolution of patterns of enzyme regulation in response to specific physiological requirements. In Bacillus species, the first step in the pathway is catalyzed by multiple forms of aspartokinase, which differ in their structure and feedback regulation. One form of aspartokinase (V-type) functions primarily during cell growth, another form (S-type) during sporulation. The V-type aspartokinase from Bacillus subtilis and Bacillus polymyxa is discussed in some detail on account of its complex pattern of regulation by the pathway endproducts lysine and threonine and its unusual subunit structure. The enzyme is composed of two dissimilar subunits, the smaller of which corresponds to the carboxyl-terminal domain of the larger subunit. The coding sequence for the subunits of Bacillus subtilis aspartokinase has recently been cloned in Escherichia coli. The study of its structure and mode of expression has revealed that the two aspartokinase subunits are encoded by in-phase overlapping genes. These unusual features of aspartokinase suggest that important aspects of the regulation of the aspartate pathway are yet to be discovered.