Microbial Analysis of Soil Essay Example for Free

Microbial Analysis of Soil Essay Abstract: soil samples were collected fortnightly from area near Dahisar River, A river in suburb of Mumbai. laboratory analysis started from July 2010 to September 2010. Total bacterial and fungal count were estimated by standard spread plate isolation. Isolated bacteria were subject to colony characterization and were estimated by their morphological and biochemical characters. As being a monsoon the occurrence of variation of different species were high. The microorganisms isolated from the soil were of staphylococcus strain and were gram positive, aerobic, coccus shaped bacteria. The fungal species were also identified, of which Aspergillus and Penicillium were dominant, followed by mucur, as sub dominant .This project aims to find out the water and soil quality of River and as it is flowing through an industrial area, to find out if it is getting affected by the Industrial pollutants. Introduction: Soil is the region on the earth’s crust where geology and biology meet, the land surface that provides a home to plant animal and microbial life (Pelczar et al., 1993). Soil teems with microscopic life (bacteria, fungi, algae, protozoa and viruses) as well as macroscopic life such as earthworms, nematodes, mites, and insects, and also the root systems of plants. The numbers and kinds of micro- organisms present in soil depend on many environmental factors: amount and type of nutrients available, available moisture, degree of aeration, pH, temperature etc (Prescott et al., 1999). Soil bacteria and fungi play pivotal roles in various biochemical cycles and are responsible for the recycling of organic compounds (Wall and Virginia, 1999). Soil microorganisms also influence above- ground ecosystems by contributing to plant nutrition, plant health, soil structure and soil fertility (O’Donnell et al., 2001). Soil is generally a favorable habitat for the proliferation of microorganisms, with micro colonies, developing around soil particles. Numbers of micro organism . In soil habitats normally are much higher than those in fresh water or marine habitats (Atals and Bartha, 1998). Bacteria make up the most abundant group of micro- organisms in the soil (3.0 x 106 – 5.0 x 108) per gram of soil, followed by the actinomycetes (1.0 x 106 – 2.0 x 107), fungi (5.0 x 103 – 9.0 x 106), yeast (I.0 x 103 – 1.0 x 106), algae and protozoa (1.0 x 103- 5.0 x 105) and nematodes (50 – 200) counts per gram of soil are wide differences in the relative proportions of individual bacteria genera found in particular soils (Atals and Bartha, 1998). Soil fungi may occur as free-living organisms or in mycorrhizal association with plant roots. Fungi are found primarily in the top 10 cm of the soil and are rarely found below 30 cm. They are most abundant in well-aerated and acidic soils (Domsch et al., 1980). Most fungi in soil are opportunistic (zymogenous). They grow and carry out active metabolism when conditions are favorable which implies adequate moisture, adequate aeration and relatively high concentrations of utilizable substrates (Postage, 1994; Miyanoto et al., 2002). In this research we isolate culturable heterotrophic bacteria and fungi from different top soil samples MATERIALS AND METHODS Laboratory analysis Preparation of materials The materials needed for this experiment include; glass wares (conical flasks, bijou bottles, pipettes, petri-dishes) and they were washed with detergents. These glass wares were rinsed thoroughly with clean distilled portable water and left to air dry before sterilizing them in the autoclave at 15ââ€" ¦C for 1 hour. Also, the laboratory cabinets on which the work would be carried out was swabbed with cotton wool soaked in methylated spirit to sterilize it before any microbiological analysis was carried out to avoid the growth and isolation of other organisms not present in the samples. After sterilization, the plates were allowed to cool to about 45 degrees before they were used. Microbiological evaluation Ten (10) grams of the soil sample for microbiological evaluation was weighed into 9ml of sterile water. Preparation of serial dilution goes thus: 1ml of the original stocks solution was poured into 9ml sterile distilled water and mixed thoroughly to give 10-2 of the original sample and this was done for each sample and the bottles labeled according to date of collection Isolation and Enumeration of Micro-organisms. 1gram of the samples was homogenized in 9mls of distilled water to obtain a ratio of 1:9 and the second diluted of each sample was plated using the pour plate technique. Sterile molten nutrient agar (NA), potato dextrose agar (PDA), macconky’s agar,(MA) manitol salt agar (MSA) and deoxycholate astrate agar (DCA) were used{the potato dextrose agar (PDA) was acidified). These agars were then added and left to solidify undisturbed. These plates were incubated 37oC for 24hours (incubation was aerobic) and the procedure was repeated using 10-2 finally the number of colonies per plates were counted and recorded. The acidified PDA was incubated at 25C for 3-7 days for microbial growth. Total Bacterial counts (Cfu/g) The total bacteria count for each sample was determined with the pour plate techniques using nutrient agar. The plates were incubated between 24hours at 370C and all colonies appearing on the end of the incubation period were counted using digital unlimited colony counter and the counts were expressed in colony forming unit per gram {CFU/g} of the sample. Colonies of bacteria developing on the plates were observed, isolated and reisolated on a fresh media until pure culture was obtained. Preparation of Pure Culture It is necessary to isolate organisms in pure culture before studying and identifying them because a pure culture originates from one cell. Characteristics colonies from the original culture on the plates were picked with a sterile wire loop (using surface streaking method) and this loop was used to make streak of the colony on the surface of newly prepared sterile agar plates of NA,MA MSA. These streak will space out the inoculants and discrete colony of a particular specie of organism and then incubated at 35-37oC for 24hours to enhance microbial growth. Distinct colonies were re-inoculated on another fresh agar plates in order to obtain a pure culture. The isolates were picked with sterile loop and streaked into prepared agar slants, labeled and incubated for growth after which they were kept in the refrigerator for future use and identification. Identification of Isolates These isolated bacteria were identified using both morphological culture characteristics (i.e. the color, shape, elevation, capacity, consistency, edge) and biochemical test (i.e. citrate, oxidase, indole, sugar fermentation, test etc.)and the bacteria were identified based on the results obtained from the above mentioned biochemical characterization results and the procedures include. Grams Staining Techniques A drop of distilled water was placed on a clean glass slide. The inoculating wire loop was sterilized by flaming until it was red hot (this is to prevent the invasion of unwanted micro- organisms that might be inhabiting the wire loop) in the blue flame of a Bunsen burner. The loop was allowed to cool and the small portion of each colony of microorganisms to be gram stained was picked and smeared in the drop of water (distilled) on the glass slide and then spread into a thin smear along the slide. The smear was air dried and passed through the blue flame. The smear was stained with 1%crystal violet and left for 1minutes (60secs) and then washed with running distilled water it was then stained again with Lugols iodine for another 60secs and also washed with running distilled water. The slide was decolorized rapidly with 75% alcohol in order to present the organism from having the color of the primary reagent and it was washed immediately with distilled water. The slide finally was flooded with a counter stain safranine (a secondary stain) for 60secons and also washed off with distilled water and allow to air dry. The slide was covered with a cover slide and observed under the microscope using oil immersion x 100 objective lens with immersion oil. The gram reaction of the isolated arrangement and the shape of the cell were observed and recorded. Gram positive (+ve) bacterial were characterized by a purple color (i.e. the primary stain) while the gram negative (-ve) bacteria were characterized by red color (i.e. the secondary stain) .This procedure is actually used to ascertain the component of each organisms cell wall. Motility Motility was determined by hanging drop techniques. Using loop, a little part of the colony of the organisms were grown in peptone water for 18hours and then placed in the grease free slide and covered with a Vaseline bound cover slip and then observed under x100 objective lens. A motile organism is then seen moving in the drop of liquid. Identification Of Mold Isolates Mold isolated was identified using cultural and morphological characteristics and according to (Fawole and Oso, 2001), microscopic observation was carried out using lacto phenol blue stain. Procedure for Mold Staining A drop of lacto phenol blue stain was dropped on a clean grease free sterilized glass slide and after this a sterile inoculating wire loop was used to pick the mycelium unto the glass slide from the mold culture .The mycelium was spread evenly on the slide. Teasing was carried out to separate the mycelium in order to get a homogenous mixture and the mixture was then covered with cover slips gently and then allowed to stay for some seconds before observing under x40 under the microscope. The microscope examination of actively growing mold was on the basis of structures bearing spores, presence or absence of septate. BIOCHEMICAL TESTS Catalase Test Catalase test demonstrates the presence of catalase enzyme by aerobic microorganisms. Catalase is an enzyme that catalysis the release of oxygen from hydrogen peroxide (H2O2). To test for catalase, a drop of 3% hydrogen peroxide solution was added to a slide and the organism to be tested for catalase production is brought in contact with the hydrogen peroxide. The production of gas bubbles however indicates a positive reaction and this shows that catalase enzyme is produced.(FawoleOso, 2001) Oxidase test This was carried out by placing a clean filter paper on the working bench or petri dishes and 2-3 drops of freshly prepared oxidase reagent was added to the isolate using a sterile inoculating wire loop. After this, a few quantity of oxidase reagent was added and a purple coloration was observed within 10-15minutes which indicated that the organisms is oxidase positive and according to Olutiola et al, 1991, a positive reaction is dependent on the presence of cytochrome. This test is also useful for the separation of Neisseria in mixed culture and in differentiating Pseudomonas from enteric bacteria. Indole test Olutiola et al, 1991, describes the test as one which is important in the differentiation of colonies and it depends on the production of indole from tryptophan by the organism. An inoculating loop was used to inoculate the organism into a test tube containing decarboxylase medium becomes violet. An uninoculated test tube serves as a control (i.e. remained yellow) Sugar fermentation test The ability of the isolates to utilize certain sugar as energy source was tested. If the organism does ferment a particular sugar, acid will be produced and gas may be produced or not. Acid production is indicated by color change of the medium from red to yellow and acid presence could also be detectable with a ph. indicator in the medium while the production of gas is indicated by a void produced in a Durham tube. The fermentation medium was prepared by 0.1g of sodium chloride and 0.1g of fermentable sugar (glucose) in 10ml of distilled water. An amount of 9ml of the medium was pipette into a test tube containing Durham’s tubes in replicates. 5ml of phenol red indicator was immediately discharged into the test tubes. The test tubes containing medium were sterilized in an autoclave at 121 o for 15minutes.After sterilization, each isolate were incubated in glucose Medium. An uninoculated test tube was also incubated for glucose to serve as a control. The test was also carried out using maltose, lactose, galactose, manitol, sucrose, fructose and mannose.(Olutiolaet al., 1991) Discussion: The abundance of bacteria and fungi in this study were typical of environment with high species richness and functional diversity. Despite the fact that it is possible that a number of bacteria and fungi may be missed in this study, the isolates could be readily assigned dominant (e.g. Bacillus sp, Aspergillus sp) or transient/succession roles in the isolation of organisms form different seasons, which form the basis of this study. In additions to the implications of the determination of the number of microorganisms during soil sampling, one should consider the qualitative aspect of the preservation of important species and groups of microorganisms and of the changes in these biochemical characteristics resulting from the variations in these counts. Although the results of this study would not be considered to be exhaustive, as it was done within the limits of facilities available in the laboratory, an insight into the population dynamics and distribution of culturable aerobic bacteria and fungi diversity has been elucidated. This is without prejudice to the possible influence which a substantial proportion of bacteria and fungi that are not culturable in vitro could have on the overall picture of event. It would require more modern technology (nuclei acid probes) to obtain such detailed overview of microbial diversity. This should be a subject of extension of this investigation in future. Conclusion Through this project, if emphasis is made on public health, the observation and findings show striking predominance of Salmonella typhi. And E.coli. E.coli being an enterobacter cause dysentery and S.typhi poses a great risk of typhoid. Health inspector and municipal authorities should look into this matter for further investigation and if possible improvement Acknowledgement Investigators are grateful to the Principal Management of S.V.K.M’s Mithibai College for constant encouragement support. And head of department of zoology Prof. V.V. Dalvie for providing me opportunities and Prof. Radhika D’souza, under whose guidance the project was successfully completed References 1 .Atals RM, Bartha R (1998). Microbial Ecology: Fundamentals and Applications. 4th Edition. Benjamin Cummings Publishing Company Inc. Addison Wesley Longman Inc. pp. 300 – 350. 2. Miyanoto T, Igaraslic T, Takahashi K (2002). Lignin–degradation ability of litter decomposing basidomycetes from picea forest of Hokkaida Myco.sci. (41): 105 – 110. 3. Domsch KH, Gaws W, Anderson TH (1980). Compendium of soil fungi 4. O’ Donnell AG, Seasman M, Macrae A, Waite I, Davies JT (2001). Plants and Fertilizers as drivers of change in microbial community structure and function in soil. Plant Soil (232): 135 – 145. 5. Pelczar MJ, Chan ECS, krieg NR (1993). Microbiology: Concept and Application International edition McGraw-Hill, USA. Pp 281-324. 6. Wall DH, Virginia RA (1999). Controls on soil biodiversity insights from extreme environments. Appl. Soil Ecol. (13): 137–150. 7. Fawole and Oso, 2001

Essay --

Thus â€Å"in the thinking about post-conflict reconstruction it is that policy oriented work, which primarily reflects an institutional capacity approach to state development†¦this state will have a monopoly over the legitimate use of violence, maintain public order, generate employment, stabilize the economy, and provide essential services†( Krasner 2010:6). This gives insight that institutions in their premises posses legitimate and authoritative power to govern. Imposed in proper way, trained and informed, this state settlement is licensed to implement new political agenda. Solely they consist tools and methods to implemented new codes of conduct and make civil society more effective. Economic growth mainly depends on their accuracy and effectiveness. Recapitulate, indeed institutions should be taken as primary concern in state building process before any upcoming change. Arguing about democracy, it is seen as exogenous phenomena which certainly deems most appropriate political settlement for the country but according to above mentioned, it cannot operate without beforehand installation of good endogenous components-institutions. Recommendation that strikes from above mentioned is that at most basic level democracies and capitalism presuppose a functioning of state apparatus but in state building agenda which is oriented solely toward promotion of democratization and marketization in intuitionally weak post-conflict environments is counterproductive.(Paris 2004:205). Paris in this regard proposes strategy IBL that addresses to phenomenon of institutionalization, hence advantages of installing proper institutions before liberalizing the field. The dilemma of IBL (Institutionalization before Liberalization) IBL solution and its p... ...nce, on one side the actions taken by UN peace mission come up to $19.9 billion and on the other side during same period costs up to $6.9 trillion were used for military enforcement in different parts of the world, which when it comes to costs and values of human lives neglects the fact of lavish international assistance in terms of state building process.(Paris 2004: 211). From this derives that all this dangers could be easily maneuvered with patient and prioritized actions. Ultimately by promoting† gradual controlled liberalization combined with the immediate construction of domestic institutions that are capable of managing destabilizing effects of democratization and marketization†, IBL strategy seems more compatible and harmless for state building process.(Paris 2004:211). As such prioritization of actions encountered under its umbrella should be considered.

Juveniles Tried as Adults Essay

In the United States, anyone who is charged for committing a crime before the day of their 18th birthday is considered a juvenile and depending on the severity of the case shall be tried as a juvenile. There are some cases; however, where the juvenile justice system should be harder on the juvenile, but in most cases they should not go to an adult prison. There are most certainly some cases in which the juvenile should face the adult justice system, but for petty instances, a juvenile court will suffice. I find it hard to agree that a juvenile convicted for crimes dealing with drugs, alcohol, traffic violations, etc. should be tried in an adult court to receive punishment; however, I do believe that someone who commits rape, murder, kidnapping, or any other major crimes of the sort should be taken to an adult court. â€Å"Old enough to do the crime, old enough to do the time,† is a quote I remember hearing as I was growing up, but I was not taught that it applied to small or m inor crimes, but often serious ones involving the harming of another individual. Placing juveniles in adult prisons can cause them to be put in danger, when in reality many of them can be â€Å"fixed† through the juvenile justice system. Juvenile offenders sometimes commit crimes that are equal to or of higher quality than those of adults; however, punishing them as adults in adult prisons will do no justice; they are less competent to stand trial, adult prisons can harm them mentally, physically, and emotionally, and they more often than not choose the actions they do because of someone who is of influence to them. Juvenile offenders are often less competent than adults to stand trial making it ineffective to sentence them as if they were adults. Juvenile offenders are classified as â€Å"childish, infantile, and young,† according to dictionary.com. â€Å"Some studies have examined the understanding that youths’ have on trial procedures and the overall basic knowledge of trials† (Grisso et al). During these studies they found that there was no compassion to the basic knowledge of trials and trial procedures to that of adults. One study conducted showed that 55% of the juveniles they interviewed could not accurately describe what the Miranda laws meant when read to them except the section that says, â€Å"you have the right to remain silent,† according to the National Center of Juvenile Justice. The National Center of Juvenile Justice also stated that, â€Å"juveniles from the ages of 11-15 are very incompetent and that 16-24 year  olds have similar levels of competence.† The juveniles who are younger, or in the 11-15 range, are less likely to understand the risks and consequences of the adult justice system, and therefore may not benefit from it. They also possess weaker decision-making skills. Since they lack decision-making skills, they are more likely to make poor decisions when committing crimes, but also when agreeing to sentencing or plea agreements, leading them to an unfair trial because of the unfair advantages that justice system would have over them. Adult prisons are very harmful to one’s mental, physical, and emotional capabilities especially when they are juvenile. Being placed in an adult prison can make them susceptible to sexual harassment, physical harassment, and psychological harassment from other inmates. They could also face longer, rou gher sentences than they would have if they would have stayed in the juvenile system. A study that was done on 946 juveniles found that 87% of them faced longer sentences than they would have if they had stayed in a juvenile justice system, according to Mulvey and Schubert. In 2005, 21% of all inmates that were sexually victimized by another inmate were under the age of 18, states Mulvey and Schubert. The risk of a juvenile being physically abused in an adult prison is much higher than that of an adult in the same system because juveniles are â€Å"easier† targets and less likely to create a struggle. â€Å"Doing the time for doing the crime might be seen as fair, but doing much worse time because the crime was done while an adoles ­cent seems to tip the balance beyond even-handed justice† (Mulvey et al 846). Adult prisons also have a different effect on juveniles than they do on adults when it comes to their development; since juveniles are receiving the punishment they are at a younger age it can cause problems for them in the future. According to Mulvey and Schubert, â€Å"Adolescents in the adult system may be at risk for disruptions in their personal development, identity formation, relationships, learning, growth in skills and competencies, and positive movement into adult status.† Identity formation is just one of the aspects in which their developmen t can be affected. Identity formation is when you find out who you are as a person, this is often discovered through learning from your parents, friends, peers, etc., but when you are placed in a facility like an adult prison you are surrounded by people who have all committed a crime and are bad influences to you and cause you to create your true self around that type of behavior. Juveniles in the adult systems also lose great opportunities such as their ability to learn about all aspects of life and the responsibilities and goals they should have. They are instead learning about the inside of a prison, jail, etc. Being in these facilities causes juveniles to miss out on learning the responsibility of a job, school, family, values, goals, finding qualities in someone that could be a potential spouse, making new friends who could be positive influences, and a. All of these statistics prove that adult prisons are very harmful to juveniles, especially mentally, physically, and emotionally. It is also often found that juveniles will make the choices they do based upon the choices they watched their close peers make or just the types of people they are surrounded by in general which shows that they are immature and very easily influenced. It is hard to prove that most, or all, of juveniles have a full understanding of the justice system and the courts, making it di fficult to believe that they should be tried as adults in this system. Many juveniles have the ability to change their behavior through the programs that we have specifically for them not through adult prisons. A child who comes from a broken home, or a home without both parents, a family that is part of the lower class, or from a family that is rather large in size have been found to be the majority of juveniles facing time in juvenile or adult systems. Families who are large in size and of a lower class often find that the children are more likely to grow up without any values or goals because they are often left home alone or there is at least less supervision over each individual child, therefore causing them to be hurt and sometimes wanting to inflict pain upon someone or something in hopes to make themselves feel better. â€Å"Family relationships, duties, responsibilities and privileges, and the amount of control exercised over children all play roles in forming character and influencing behavior. The attitudes and actions of parents can create an important influence in the lives of children. Families in crisis will most likely affect the behavior of juveniles. If one member of a family becomes sick, schizophrenic, or alcoholic, a child may react based on the family’s structural problems,† according to Joseph Wickliffe. Families who contain an unstable parent(s) can create a child that is more likely to be disobe dient, especially if the way the rules are portrayed is too aggressive, too passive, or just unclear. According to a study that Joseph Wickliffe talks  about, â€Å"It was discovered that 4.1 percent of fathers were found to use sound discipline practices; 26.7 percent, fair; and 69.3 percent, unsound. Sound – consistent and firm control but not so strict as to arouse fear and antagonism, fair – control which is indefinite: sometimes strict, sometimes lax, and unsound – extremely lax or extremely rigid control by the parents, which, on the one hand, gives unrestrained freedom of action and, on the other hand, restricts to the point of rebellion.† Juveniles are also prone to make decisions based off of what the people with authority want, for instance, they are more likely to confess or accept a plea agreement if their lawyer is telling them that they should do so. After learning of all of the negative consequences that come with placing a juvenile in th e adult courts and/or prisons, I have concluded that for most cases a juvenile should be processed through the juvenile system and take the punishment given there.

About Seppuku and Samurai Ritual Suicide

Seppuku, also known less formally as harakiri, is a form of ritual suicide that was practiced by the samurai and daimyo of Japan.  It usually involved cutting the abdomen open with a short sword, which was believed to immediately  release the samurais spirit to the afterlife. In many cases, a friend or servant would serve as a second, and would ritually decapitate the samurai to provide release from the terrible pain of the abdominal cuts. The second needed to be very skillful with his sword to achieve the perfect decapitation, known as  kaishaku, or embraced head. The trick was to leave a small flap of skin attached at the front of the neck so that the head would fall forward and look like it was being cradled by the dead samurais arms. Seppukus Purpose Samurai committed seppuku for a number of reasons, in accordance with bushido, the samurai code of conduct. Motivations could include personal shame due to cowardice in battle, shame over a dishonest act, or loss of sponsorship from a daimyo. Often times samurai who were defeated but not killed in battle would be allowed to commit suicide in order to regain their honor. Seppuku was an important act not only for the reputation of the samurai himself but also for his entire familys honor and standing in society. Sometimes, particularly during the Tokugawa shogunate, seppuku was used as a judicial punishment. Daimyo could order their samurai to commit suicide for real or perceived infractions. Likewise, the shogun could demand that a daimyo commits seppuku. It was considered far less shameful to commit seppuku than to be executed, the typical fate of convicts from further down the social hierarchy. The most common form of seppuku was simply a single horizontal cut. Once the cut was made, the second would decapitate the suicide. A more painful version, called  jumonji giri, involved both a horizontal and vertical cut. The performer of jumonji giri then waited stoically to bleed to death, rather than being dispatched by a second. It is one of the most excruciatingly painful ways to die. Location for the Ritual Battlefield seppukus were usually quick affairs; the dishonored or defeated samurai would simply use his short sword or dagger to disembowel himself, and then a second (kaishakunin) would decapitate him. Famous samurai who committed battlefield seppuku included Minamoto no Yoshitsune during the Genpei War  (died 1189); Oda Nobunaga  (1582) at the end of the Sengoku Period; and possibly Saigo Takamori, also known as the Last Samurai  (1877). Planned seppukus, on the other hand, were elaborate rituals. This might be either a judicial punishment or the samurais own choice.  The samurai ate a last meal, bathed, dressed carefully, and seated himself on his death cloth. There, he wrote a death poem. Finally, he would open the top of his kimono, pick up the dagger, and stab himself in the abdomen.  Sometimes, but not always, a second would finish the job with a sword. Interestingly, ritual seppukus were usually performed in front of spectators, who witnessed the samurais last moments. Among the samurai who performed ceremonial seppuku were General Akashi Gidayu during the Sengoku (1582) and forty-six of the 47 Ronin in 1703. A particularly horrifying example from the twentieth century was the suicide of Admiral Takijiro Onishi at the end of World War II. He was the mastermind behind the  kamikaze  attacks on Allied ships. To express his guilt over sending some 4,000 young Japanese men to their deaths, Onishi committed seppuku without a second. It took him more than 15 hours to bleed to death. Not for Men Only Seppuku was by no means a solely male phenomenon. Women of the samurai class often committed seppuku if their husbands died in battle or were forced to kill themselves. They also might kill themselves if their castle was besieged and ready to fall, so as to avoid being raped. To prevent an unseemly posture after death, women would first bind their legs together with a silk cloth. Some cut their abdomens as male samurai did, while others would use a blade to slit the jugular veins in their necks instead. At the end of the Boshin War, the Saigo family alone saw twenty-two women commit seppuku rather than surrendering. The word seppuku comes from the words setsu, meaning to cut, and fuku meaning abdomen.