Category: Microbiology

The uniqueness of microbial groups

Among a large number of microorganisms, there are seven macro groups with hundreds of thousands of representatives. These include bacteria, archaea, algae, protozoa, fungi, viruses, and parasitic helminths. Despite their microscopic size, each of the groups described represents a unique biological diversity and is therefore identified as a separate cohort. From all groups of microscopic organisms, representatives of the bacterial and archaean world do not have a nucleus, for this reason, bacteria and archaea are considered prokaryotic cells. However, other membrane organelles, such as mitochondria, are not included in such cells. In addition, these two groups are the most common classes of living organisms, and their habitats are represented by all possible options, from reservoirs to boiling volcanic vents. Some of the bacterial representatives demonstrate parasitic properties such as C. tetani, B. burgdorferi, and S. typhi (1a The Main Themes). At the same time, archaeas have unique biochemical properties, which allows them to use chemical reserves as energy sources. The representatives of this group can be Methanogens and Haloarchaeas.

Moreover, parasitic properties are presented at a group of helminths  these are microscopic worms, which have a variety of forms that cause damage to animals. Compared to other groups of microscopic forms, helminths can be considered the most progressive because they have sexual dimorphism and a complex body. Trichinella is a parasite for mammals by settling in the small intestine and Fluke parasites in animals and snails.

Other groups of microorganisms, except for viruses, are considered eukaryotes because their cells have a formalized nucleus. Viruses are not living in nature and represent integration between inanimate objects and organisms, which is their main feature. The effect is simple particle structure  they do not have most of the known organelles and the capsid contains only genetic material (Moelling and Broecker 2). Well-known representatives of viruses are Tobacco mosaic virus, Ebola virus, and Coronavirus.

Microscopic fungi are beneficial cultures of living organisms for humans: Penicillium and Saccharomyces are used in the production of medicines and culinary products. Fungi do not have chlorophyll, but the cellular structure and features of biochemical processes allow them to correlate with animals and plants simultaneously. Unlike other representatives of microorganisms, fungi are unable to move actively but lead an attached lifestyle.

Among all groups, microscopic algae  photosynthesizing plants of the simplest form  stand out especially. Euglena, a transitional form between animal and plant, has a mixotrophic feeding. The majority of algae move with the help of flagellates or cilium. Congestions of microscopic algae are found in glaciers, in snowy areas, in damp soils, but most of them inhabit water bodies.

The remaining class of microorganisms is protozoa: as a rule, this group collects creatures on the principle of residue, so representatives of the simplest are most diverse. Vorticella has the shape of a bell and is attached to crayfish covers, while Paramecia have cilium that actively moves around the water column. The simplest are usually predators, and their characteristic lifestyles are individual or colonial.

The Golden Age of Microbiology

Since Leeuwenhoek first discovered living microscopic organisms, research into the smallest cells has begun to gain momentum. Simple observation of microorganisms shifted to predicting their properties and characteristics and then to conscious editing to create economically important forms (1b Main Themes). Thus, the science of biotechnology was born, aimed at using biological systems in human activity. The father of biotechnology development can be rightfully considered Edward Jenner, who created the vaccine against smallpox. There is a legend that the young boy Edward, interested in the medical sciences, was approached by a dairymaid and asked about the ulcers that appeared on her body. Jenner assumed it was smallpox, but the milkmaid answered that she could not get sick because she had recently suffered from cowpox. This reaction shocked the boy, and he decided to look into the matter. This legend may be fiction, but it does not negate the fact that Jenner had the intention of developing a cure for this disease. The revolutionary idea he proposed was that a person who had contracted cowpox was not exposed to a more serious type of viral infection. Jenner decided to carry out a risky experiment in which he initially infected an eight-year-old boy with cowpox, and a few days later with smallpox (Morabia 255). As a result, the child was resistant because his immunity had already developed antibodies to the disease, and Jenners work became a source of vaccines against hundreds of infections.

Elizabeth Bugie was another researcher who made a useful contribution to microbiology. In particular, the woman studied soil bacteria to find antimicrobial agents to combat plant infections. In 1944, during a series of laboratory experiments, Bugie accidentally discovered the fascinating effect of Gram-positive S. griseus  they produce a metabolic product with antimicrobial action against M. tuberculosis (Elizabeth Bugie). This is how the first milestones in antimicrobial research were reached and how a branch of medicine aimed at combating tuberculosis has developed.

Rebecca Lancefields classification helped to understand the existing diversity of microscopic organisms. The researcher has devoted her scientific career to an in-depth study of the structure of cell walls of streptococcal bacteria: she found that there are differences in the carbon composition of antigens (Rebecca Lancefield). Thus, were not only developed a fundamentally new concept of serological classification, but also found that there are several pathogenic forms of bacteria causing scarlet, erysipelas, and sore throat. Lancefield has been awarded the title of President of the American Immunologists Association for its achievements in microbiology.

Works Cited

1a The Main Themes of Microbiology. Section Biol 004 or 103. PowerPoint Lecture.

1b Main Themes of Microbiology. Section Biol 004 or 103. PowerPoint Lecture.

Elizabeth Bugie. Britannica, n.d., 2020. Web.

Moelling, Karin, and Felix Broecker. Viruses and Evolution  Viruses First? A Personal Perspective. Frontiers in Microbiology, vol. 10, 2019, pp. 1-13.

Morabia, Alfredo. Edward Jenners 1798 Report of Challenge Experiments Demonstrating the Protective Effects of Cowpox Against Smallpox. Journal of the Royal Society of Medicine, vol. 111, no. 7, 2018, pp. 255-257.

Rebecca Lancefield. Britannica, n.d., 2020. Web.

Methods

Isolation of pure culture took place on BIOLOG media; gram stain was then performed to illustrate the testing protocol. Inoculum is prepared at a specified cell density with the use of a turbid meter. Inoculation is done on a micro-plate, which is then read, and identity is determined.

The pure culture used for identification was tested by gram stain. We did this to determine if the organism was bacteria, gram-positive or negative. One must be very alert for variables in gram, which indicated the different microbes present. We realized those good techniques were of great significance, and mistakes in determining the shape and gram reaction could put an individual on the wrong track.

Aerobic bacterias Microplates were read after a period of four to six-hour incubation. The resulting pattern could be read manually with the naked eye or using a microplate reader at 590nm. White coverer was removed and placed on a white sheet. Dark purple wells meant positive while clear wells indicated negative.

The next step was manually entering the reactions into the onscreen microplate. A purple circle signified a positive result. Whereas white circles meant negative results, we individually changed the wells positive to negative by clicking on all positive.

Results

The system identified the microorganisms by the use of extensive database software. The microplate indicated a good match with the organism an ID appeared in the bar area, which is green in color, at the middle of the screen which is at the pinnacle of the results area.

The software demonstrated microbes on a list that is ranked with the entry number one topping up, as the best matched chosen from the database. A (+) sign found in a negative well implied that the BIOLOG database was positive for that particular test. Gram-positive Rod was also used to determine if it was spore-forming bacillus, as special handling was required in spore formers as they often grow at a faster rate.

Directions and protocols must be followed meticulously, we discovered this can be a weakness, if not properly guarded. On the other hand, grams positive bacteria were a bit challenging to identify. It is mandatory to incubate anaerobic bacteria with a container together with a headspace of nitrogen. Some organisms are capable of producing nutrient-rich slime coating, and cells can actually feed on them, this, however, breeds false negatives.

Conclusion

It is imperative to ensure that the culture is first filtered into non-selective media. With this in place, all wells can stand positive. Use of proper aseptic techniques is always necessary. Disposable sterile plastic ware, as well as glassware, should be employed when handling cell solutions and suspensions. Glass cell that is already washed may have some traces of detergent which can affect the findings with sensitive strains.

Failure to use the right media, atmosphere, and temperature for incubation influences the accurateness of the database generated, which results in BUG media. Not following the correct protocol or cutting corners and using old sporulating or unhealthy cells to make a suspension. By using worn-out cells, the system tests the metabolic properties of the cells that are living because some of them are liable to losing their metabolic strength by subjecting the cells to stress, even for a short period.

Works Cited

BIOLOG 2011. Web.

Introduction

Bacteria are microscopic single celled organisms which are said to be the smallest yet most ancient forms of cellular life. Bacterial cells are prokaryotic and wide spread. Complex behaviors often associated with the organisms have prompted scientists around the world to keenly monitor the characteristics of various types of bacteria in an attempt to control them (Havey and Cling 42). This field of research has in the past gained popularity especially in the health and medical sectors.

Materials and Methods

Microscopic and scientific technology has been vital in the study of bacteria and other microscopic organisms. This has been achieved through the invention of various test methods that are known to produce results which are specific to a given class of bacteria (Hwang and Ederer 51). Results obtained from the various tests can then be applied to a specific type of bacteria. Most of these tests involve staining to identify the various features of the microscopic bacteria being observed under a microscope.

Organisms/Colonies Morphology Test

A clean and uncontaminated petri dish is removed from a plastic bag and placed on a level surface (Greenwood and Pickett 45). This was done carefully to avoid touching the inside of the petri dish or expose it before the experiment. A 1ml pipette was then filled with water and emptied into the bacteria medium bottle. The medium bottle was inverted twice to mix water with the medium. The mixture of water and medium was then poured into the petri dish and swirled to completely cover the surface. The petri dish was then covered. The dish was then incubated for 48 hours under room temperature. The contents were then observed.

Gram Stain Test

A sample of the unknown bacteria was picked from a colony based on the colour. A drop of purple crystal violet dye was then placed on an uncontaminated slide. A colony from the petri dish was then transferred on to the cover slip and immediately inverted onto the crystal violet dye. Alcohol was then used to wash the slide thus washing the dye out of the cell surface. A counter stain containing pink dye was then used to stain the cells and the cells observed.

Catalase Test

A drop of Hydrogen Peroxide was placed on a clean slide. A colony of the unknown bacteria was identified from the contents in the petri dish. A colony was then placed on the cover slip and immediately inverted onto the hydrogen peroxide.

Slide Coagulase Test

Two separate portions of concentrated homogenous suspensions were prepared on a slide (Greenwood and Pickett 45). Three loopfuls of citrated plasma at room temperature were placed next to one of the solutions. The suspension was then carefully mixed with the plasma and left for about 10 seconds. The two solutions were then compared after the 10 seconds.

Oxidase Test

A strip of a filter paper was impregnated with 1% Tetramethyl-p-phenylenediamine dihydrochloride solution. Forceps were then used to place the filter paper onto a glass slide. A glass rod was then used to pick a colony from a freshly grown culture. The colony was then placed onto the impregnated strip of the filter paper. Forceps were then used to touch the impregnated strip of the filter paper onto the colony. The colour of the colony was then observed.

Tube Coagulation Test

A volume of 24 hour culture was prepared. The volume was then placed in a sterile test tube containing an equal volume of sterile oxalated human plasma and the solution swirled to mix (Havey and Cling 42). The solution was then incubated for 4 hours and was regularly checked at intervals of 30 minutes. The solution was then left for 24 hours.

Acid and Mannitol

0.5 millilitres of 50% sulphuric acid was added into two millilitres of a medium containing the unknown bacteria and then observations made.

Acetone Production

A sample of the bacteria colony was inoculated and incubated for 48 hours at a temperature of 37 degrees centigrade. 8 to 10 drops of naphthol, 5 percent ethanol and 8 to 10 drops of 40% potassium hydroxide containing 0.3% creatine were added into a 1 millilitre quantity of the culture and shaken well.

Urease Test

A culture medium was heavily inoculated in a medium bottle. The solution was then incubated for 24 hours at 37 degrees centigrade and observed (Havey and Cling 42). The solution was then incubated for seven days upon negative results.

Deoxyribonuclease (Danse) Test

The unknown bacteria were spot inoculated on the surface of the agar with heavy growth. The bacteria were then incubated for around 18 to 24 hours at around 37 degrees centigrade.

Resistance and susceptibility

Novobiocin and Polymyxin antibiotic solutions were placed in two separate test tubes. Colonies of the unknown bacteria were then introduced into the solutions and observations made.

Results

The cells observed in a microscope were circular in shape. This signified that the bacteria were morphologically cocci. Upon completion of the gram stain test, the bacteria were observed to still possess the purple colour, the colour of the primary stain. This signified that the unidentified bacteria were Gram positive cocci. Bubbles were also observed at the end of the Catalase test signifying the production of oxygen.

The bacteria were thus classified as Catalase positive. At the end of the slide Coagulase test, a uniform suspension was observed signifying that the unidentified bacteria were Slide Coagulase negative. Similar results were obtained in the tube Coagulase test with no coagulation noted in the test tube. This indicated that the bacteria were tube Coagulase negative. The Oxidase test did not signify any colour changes implying that the unknown bacteria were Oxidase negative.

There was no formation of crystal precipitation in the solution in the acid/ Mannitol test making the bacteria acid/Mannitol negative. The absence of colour change during the acetone production test signified that the bacteria did not have any neutral products. There was no colour change noted at the end of the Urease test. This indicated that the bacteria do not produce urea which would have otherwise been indicated by a colour change to bright pink.

A clear zone was observed to surround the colony at the end of the Deoxyribonuclease test. This indicated that the bacteria were Deoxyribonuclease negative. Introduction of Novobiocin and Polymyxin antibiotic solutions produced uncertain observations and the effect of the antibiotics could not be identified as a result of malfunctioning antibiotic disk. Based on these observations, it is clear that the bacteria in question were Streptococcus haemolyticus.

Discussion

Though the tests were carried out effectively, the microscopic size of the bacteria has continued to be a challenge in the classification of bacteria. Care must also be taken to prevent the individuals carrying out the test from being infected (Greenwood and Pickett 45). The abundance of bacteria in the atmosphere and in the environment may end up contaminating cultures, a situation that would give wrong results. Observation of bacteria under extremely high magnification powers can help in the classification of the organisms.

Works Cited

Greenwood, Girolamo, and Johnson Pickett. Clinical Microbiology, New York: CBS Interactive Network, 2004. Print.

Havey, Cohn, and Johnson Cling. Microbiology, New York: Henry and Holt, 2010. Print.

Hwang, Padmanabh, and Nahavira Ederer. Clinical Microbiology, Harvard: Harvard University Press, 2005. Print.

Overview

The research was done to assess the impacts of edge effects, forest age, and invertebrate exclusion on decomposition rates in a newly planted forest. The study found out that there were no correlations and interactions amongst the parameters that the research undertook to establish. The parameters used in the research were mean mass remaining, edge of the forest, age of the forest, and the abundance of the invertebrate organisms. The research method involved the use of three transects which each had six notches marked at 10 meters intervals and acted as data collection points. The data collected on various parameters were analyzed and results generated.

Introduction

To better understand the rate of decomposition, it is important to understand what decomposition generally means. When organisms such as animals, vegetations, insects, and other living organisms die, their bodies get broken down into several tiny pieces. The process by which the dead organisms are broken down into pieces either through mechanical or chemical means is referred to as decomposition (Swift, 1979, p.1). The process involves the activities of bacteria, certain kinds of worms and this may also involve some chemicals. The organisms involved in the process of decomposition are known as decomposers (Bharatdwaj, 2006, p. 43). The decomposers do not perform the task of decomposing the dead organisms as their primary goal, but the process is a way of their ingestion of food. The process of them ingesting dead bodies is what is referred to as decomposition (The University of Michigan, 1971, 72). The decomposers depend on the dead organisms to develop, grow and have life sustenance (JSTOR 1956, p. 25). It is important to note that even the decomposers

die and also eaten by other living organisms like bacteria (Limnological Society of Southern Africa, 1986, p.96). This means that the decomposers also get decomposed after they die. The implication is that decomposition is a cycle that will never end; when the decomposers breed they bring to existence new decomposers. After a complete decomposition, the remaining particles of the dead organism integrate into the ground and become part of the soil.

The rate of decomposition of forest litter coupled with the formation of various kinds of humus is greatly dependent on numerous factors. One of the most important of these factors is the climate. In the rate of decomposition is slowed in a cool climatic condition and in many cases, there can be found accumulation of remains of dead organisms in on the forest floor. Cool condition can exists due thick canopy which prevents the solar rays from reaching the forest floors hence is also a factor determining the rate of decomposition. The process of decomposition also depends on such factors as edaphic conditions and the quality of the soil fauna and the forest litter. The significance of each of these factors is divergent in terms of spatial and sequential scale (Agricultural Institute of Canada, 2008, pp. 443-867).

Literature Review

The process of decomposition is one of the most important processes of the ecosystem. Through decomposition nutrients go back into the soil to be used by plants in the manufacture of new food (Ecological Society of Australia, 1978, pp. 15-27). The plants are in turn food to many living organisms. Ecologists have argued that for new organisms to be born and survive the old one ones must die and decay. It therefore means that death and decomposition are integral part of the sustenance of the ecosystem. Through scientific study, it has been proven that decomposition of dead organisms is facilitated by decomposers (Gowariker, 2009, p. 181). The decomposers include, but not limited to, bacteria, some types of earthworms and insects. It therefore means that the rate of decomposition is a factor of the decomposers; the rate increases when the number of decomposers increases. Decomposers like bacteria increase in number when conditions such as hot humid conditions exist. Research literature indicates that forest experiencing hot humid conditions experience availability of large numbers of decomposers hence rate of decomposition in such forests are high.

Ecological scientific researchers have distinguished two types of decomposition: aerobic decomposition require oxygen while anaerobic decomposition takes place in the absence of oxygen. In view of this, aerobic decomposition can only be facilitated by the decomposers that require oxygen for their physiological functions. While anaerobic decomposition requires decomposers that can survive even in the absence of oxygen (Society for Applied Bacteriology, 1992, p.9). Available literature also cites the importance of moisture in the process of decomposition. For faster rate of decomposition, the amount of moisture in the decomposing organism of heap must be as high as possible and should allow free infiltration of air so as to benefit aerobic bacteria. Research findings also indicate that the minimum amount of moisture that should support bacterial activities in the decomposition process should be around 12% to 15% (Martin, 1992, p.33).

Scientific studies have proven that temperature is such a vital part of decomposition. The findings argue that low temperatures during winter season slow down the rate of decomposition while high temperatures facilitates the rate of decomposition and this happens mostly during summer. The microbes known to be responsible for decomposition of raw organic substances are basically categorized into two: mesospheric microbes live and grow in temperatures ranging from 10⁰ Celsius to 46⁰ Celsius and thermophillic microbes live and grow in temperatures ranging from 46⁰ Celsius to 70⁰ Celsius. Ecologists have identified that decomposition taking place at high temperatures is of great benefit to gardeners since the temperatures kill germs and weed seeds that may be dangerous to the vegetables. Earthworm lives in the soil and its activities are very vital to decomposition of dead organisms or heap. Its barrowing tendency helps to create spaces that make aeration within the decomposing heap possible. Earthworms also derive their food from the decomposing organic matter and hence facilitate decomposition in the process (Consultants Bureau, 1972, p.317).

Research studies have also proven that the rate at which decomposition take places in within the forest soil depends on exposure to open air, moisture and water. The findings reveal that organic matters that are placed underground take more time to decompose than those exposed on top of the soil. The findings further state that the rate of decomposition reduces with the depth of burial of organic matter. This, the researchers say, can be explained by reduced exposure to air or oxygen; the deeper the burial of decomposing organic matter, the less the availability of air to facilitate decomposition and the more time taken for complete decomposition (National Research Council of Canada, 1987, p.2030). Insects have also been found to play the role of decomposers. They are specifically important for decomposition of organic matters that are found on the surface of the earth within the forest. Just like other decomposers, insects also derive some of their food from the dead or cheap or organic matter. Amongst the catalysts of decomposition is light. Light from the sun is a great source of heat energy that is necessary for decomposition; it also provides the plants with energy for photosynthesis within the forest. It is important that note that the availability of light to forest soil is determined by the canopy layers formed by the trees above. The more the canopy layers the less the amount of light reaching the floor of the forest (Mullen, 2009, P.40).

Critical studies that have been conducted have revealed that de composition in the forests is an important stage in the nutrient cycle. Through decomposition nutrients are taken back into the soil to be used by trees again in the manufacture of new food through photosynthesis. The tree leaves are eaten by both micro-organisms and the invertebrates found within the forest (Pacific Northwest Research Station, Nd, p. 74). So it is natural that the invertebrates are naturally attracted to the decomposition sites within the forest by the availability of food materials, which are the fallen leaves from the forest trees. Research literatures further indicates that decomposition process is facilitated differently by the decomposers. The process is commenced by the bacterial activities then the rest follow and then the process continues to the point where the remnants are integrated into the soil as parts of the soil (Tasmania, 1939, p. 14). The role of the invertebrates in the decomposition process is to sectionalize the litter into smaller pieces and also mixing the litter with the mineral soil hence divulging a wider surface area for microbial colonization (Bloem, 2006, p.25).

Problem Statement

There is a great importance to the study of decomposition of forest soil with regard to its role in maintaining the ecosystem. The research studies that have always been done have not revealed the nature of decomposition that take place within the Motutapu and scrubland soils. It can be argued that decomposition of organic matter is the same in ecosystems experiencing similar climatic conditions coupled with the availability of necessary decomposers. However, it is important to note that different ecosystems experience different climatic conditions. The distribution of the decomposers also varies from one ecosystem to the other depending on the availability of necessary conditions for survival. There is a difference of decomposition in old forests and the new ones. It is believed that there are impacts of edge effects, forest age and invertebrate exclusion on decomposition rates in a newly planted forest. It is therefore important to study the Motutapu and scrubland soils to determine whether this is the case. The research question that was during the research study therefore was, Is it true that there are impacts of edge effects, forest age and invertebrate exclusion on decomposition rates in a newly planted forest?

Research Objectives

The main objective of the research was to find out if there are impacts of edge effects, forest age and invertebrate exclusion on decomposition rates in a newly planted forest.

Hypotheses

1. Decomposition rate will increase as forest ages;
2. The rate of decomposition increases with distance from the edge of the replanted forests; and
3. When the size of invertebrates that is able to access litter increases, the rate of decomposition also increases.

Methodology

Three transects of 60 meters each in length 50 meters from the upper age of the replanted forest and continued toward the centre of the forest. Each of the three transects had six notches each marked at intervals of 10 meters. The notches served as data collection points. The first transect was located within the trees that were 8 years old, the second one was placed through trees of ten years old and the last transect was located within trees that were fifteen years of age. Litter bags of two mesh sizes measuring 1mm and 5mm was each placed at ten-metre interval of all transects except the first one; there was high possibility that the tall and thick grass coverage present with that study area would most likely cause unprecedented damages to the litterbags and adversely obstruct significant data collection. Each of the 36 litterbags contained ten mature freshly pulled out leaves.

Moreover, five more standardized litter bags were filled with other leaves collected from the same region the same day; the mass of the leaves were determined after which they were laced in oven to dry at 70^o Celsius for a period of 24 hours. Thereafter the dry weight measurement of the leaves was obtained. The 36 litterbags were retrieved after a two-month period; they were dried and then their contents weight again and then the dry weight that was lost each litterbag was calculated and the percentage remaining mass of the contents was established.

Ambiotic measurement was also obtained from transect points, a light meter was used to measure the light intensity reaching the ground level, soil temperature and moisture was measured by use of a probe and the pH of the soil that lies immediately beneath the litter layer was examined. The litter within 25mm² quadrat around the litterbag was weighed; invertebrates were juddered from it and the numerically determined to in order to approximate the mega and macro-faunal abundance in each of the regions. A point centred quarter stick was placed at each transect point and the species of the four nearest trees of greater than 15cm in height was recorded.

Data Analysis

The data analysis process used the R commanders for two-way ANOVA for fine and coarse against age; fine/coarse against Location, and Location against Age. For other mystifying factors, a multiple regression was done.

Results

The table 2 below indicates p-values from ANOVA outputs from R for mass remaining for: A  mesh type, age, mesh type and age interaction, B  mesh type and age, C- mesh type.

The result obtained from the study, as shown in the table2 above indicated that there was no interaction for the mass remaining between mesh type and age. It also indicated that there is no difference in the mean mass remaining between the different ages of the sites. However, it indicates that there is a significant difference in the mass remaining between the different mesh types. In other words, table two shows that with 95 percent confidence there was no interaction between mesh type and the location on thee transect (p-value=0.13107). There was also no evidence against the null hypothesis that the underlying means for the location on thee transect (edge, middle and centre) are the same (p-value=0.20098). And lastly there was evidence against the null hypothesis that the underlying means for mesh type are the same (p-value=0.00158).

The table 3 below shows the p-values from ANOVA outputs from R for Mass remaining for: A  age, location, age and location interaction, B  age and location.

According to the table it is evident that there is no interaction for the mass remaining between mesh type and location, the table also shows that there is no difference in the mass remaining means between the different locations on the transect and there is no interaction for the mass remaining between age and location. In other words, the table indicates that with 95 percent confidence there was no interaction between the age of the site and the location on thee transect (p-value=0.99384). There was also no evidence against the null hypothesis that the underlying means for the age of the site (old, middle and new) are the same (p-value=0.14404). Lastly there was no evidence against the null hypothesis that the underlying means for the location on thee transect (edge, middle and centre) are the same (p-value=0.27687).

As for the confounding factors, the only difference in the mean mass remaining, occurred between the fine and coarse mesh decomposition bags, therefore, a multiple regression analysis was to be completed to observe if there was an impact on the mass remaining by other factors such as pH, canopy coverage, litter depth, litter weight, invertebrate abundance and the number of orders. The results of the multiple regression is summarised as in the table below:

P-values from multiple regression outputs from R for mass remaining for coarse and fine mesh.

The following are the trends in times and space according to the research outcome:

1. Invertebrate abundance typically greatest at edges and in young forest
2. Invertebrate diversity typically greatest at edge
3. Increase in litter weight with time
4. Increase in canopy cover with time
5. Decrease in pH with time

Discussion

After the whole process of research we found out that, there is no significant difference that exists between the rate of decomposition and the age of the forest. It also came out clear that there is no considerable difference in the rate of decomposition in relation to the edge, middle and the centres of the used transects taking into account both within a transect and between one transect and the other. This is in consistence with the other research findings indicating that there is no significant change in the rate at which leaf-litter decomposition took place as compared from the interior to outward edges of the forest. It is almost impossible to explain the insignificant edge difference. The research study also found out that the rate at which decomposition took place was not correlated with the difference in air temperature, the depth of litter, moisture contents, and densities of invertebrates. These findings were in relation to the edge of transects and across all the sites used during the study. This finding is in contrast with other research findings indicating that edge difference affects the rate at which decomposition takes place. However, the removal of some lea-litter by the feeding termites is the cause for the effects of edge difference.

There was also a significant variation between the mass remaining in the coarse and fine litterbags. The greater mass that was remaining in the fine bags shows that the rates of decomposition increase when larger invertebrates are able to access the litter, which is in consistence with other research findings. We also found out the presence of mesofauna, microfauna and macrofauna has an impact on decomposition within a restored NZ forest system.

Reference List

Agricultural Institute of Canada. (2008). Canadian journal of soil science, Volume 88, Pages 443-867. Canadian Society of Soil Science, pp. 443-867.

Bharatdwaj, K. (2006). Physical Geography. Biogeography, Discovery Publishing Houses, P.43.

Bloem, J. et al. (2006). Microbiological methods for assessing soil quality. CABI Publishing Series. United Kingdom: CABI, P.25.

Consultants Bureau. (1972). The Soviet journal of ecology, Volume 2. Consultants Bureau, p.317.

Ecological Society of Australia. (1978). Australian journal of ecology, Volume 3, Issues 2-4. The University of California, pp. 15-17.

Gowariker, V. (2009). The Fertilizer Encyclopedia. New York: John Wiley and Sons, p.181.

JSTOR (Organization). (1953). Quarterly review of biology electronic edition, Volume 28. University of Chicago Press. Journals Division, University of Chicago Press for the State University of New York at Stony Brook, p.25.

Limnological Society of Southern Africa. (1986). Journal of the Limnological Society of Southern Africa, Volumes 12-14. Limnological Society of Southern Africa, p. 96.

Martin, D. (1992). The Rodale book of composting. Rodale, p.33.

Mullen, G. (2009). Medical and Veterinary Entomology. Academic Press, P.40.

National Research Council of Canada. (1987). Journal canadien de botanique, Volume 65, Issues 9-12, p. 2030.

Pacific Northwest Research Station. (Nd). Fall River long-term site productivity study in coastal Washington site characteristics, methods, and biomass and carbon and nitrogen stores before and after harvest, p.74.

Society for Applied Bacteriology. (1992). The Journal of applied bacteriology, Volume 72. Symposium series. Blackwell Scientific, p.9.

Swift, J. et al. (1979). Decomposition in terrestrial ecosystems, Volume 1979, Part 2. Volume 5 of Oakland Project Series. United States: University of California Press. pp. 1-12.

Tasmania. (1939). The Tasmanian journal of agriculture, Volumes 8-9. Tasmanian Dept. of Agriculture, p.14.

The University of Michigan. (1971). Journal of theoretical biology, Volume 30. Academic Press, p.72.

Zygomycota is mostly terrestrial in habitat. They thrive from decaying animal or plant decays. Some Zygomycota forms a symbiotic association with plants where they depend on each other for their living, while others are parasitic and depend on insects, animals, and plants for their living (Benny, 2000). A black bread mold is a form of Zygomycota that spreads on the surface of bread as well as other decaying food sources. Its hyphae penetrate inside the food to absorb nutrients.

Zygomycota produces through the asexual as well as the sexual process. For asexual production to happen, there must be the development of bulbous black sporangia first at the tip of standing hyphae. Normally, the hyphae will contain spores. On the other hand, for sexual production to take place, there must be an association between haploids of various mating types at proximity.

This leads to the growth of gametangia that results in the fusion of the cytoplasm. Subsequently, the fusion of the nuclei then occurs. The zygosporangium that is formed from this fusion is diploid and has thick walls which are highly resilient to harsh environmental conditions as well as metabolically inert. The zygosporangium germinates under favorable conditions to form vegetable hyphae. For asexual reproduction of Zygomycota to occur, sporangia and sporangiospores must develop first. Thereafter, any further development of the sporangium occurs via sporangial cytoplasm courtesy of the internal cleavage.

At maturity, the walls of sporangial disintegrate, thereby making the spores free which are then dispersed by wind or water (Alexopoulos and Blackwell, 1996).

Ascomycota is usually produced sexually as well as asexually. Sexual production occurs through ascospores or meiospores while asexual reproduction occurs through conidia. Some Ascomycota can outbreed others self-breed while others reproduce from both processes. The sexual reproduction method occurs through vegetative reproductive spores that are referred to as conidia.

Usually, conidiophores contain a single nucleus. Conidiophores come about as a result of mitotic cell divisions. Their genetic makeup is also similar to that of mycelium,

Ascomycota usually undergoes asexual reproduction. These Ascomycota are to be found in conidia, at the end of the hyphae. Usually, the hyphae are called conidiophores. This is different from the zygomycetes that undergo asexual reproduction.

They take any liquid as long as there is some water present in it (Berbee, 2001).

Basidiomycota reproduces sexually as well as asexually. When underground environmental conditions are favorable for Basidiomycota, Basidiomycota forms reproductive structures that facilitate the formation of spores that are formed at the tip of the Basidiomycota in a structure referred to as Basidiomycota (McKerracher, 1985). The formation of these spores results from meiosis that splits the genetic code of the Basidiomycota in its half.

When two basidiospore combines, a new organism is formed which sprout more Basidiomycota to continue with the reproduction process. Basidiospores on the other hand can also reproduce asexually from their underground structures by dividing themselves to form duplicates of themselves which makes them spread their hyphae out as they look for compatible basidiospore. Once they come into contact with their compatible partner, they join to continue with their reproduction cycle (Lichtwardt, 1986).

Zygomycota, Ascomycota as well as Basidiomycota reproduce both sexually as well as asexually. Ascomycota and Basidiomycota have similar reproductive cycles, although they have different structures for reproduction. Basidiomycota produces their spores in basidia cells, while Ascomycota produces their spores in asci which are tube-like cells. The asci have to burst to disperse the top of the spore continues with the asexual reproduction which later merges to form sexually producing structures.

Reference List

Alexopoulos, C. J. and Blackwell, M. (1996). Introductory to Mycology. New York: John Wiley and Sons.

Benny, G. L. (2000). Amoebidium parasitismis a protozoan, not a Trichomycete. Cambridge: Cambridge University Press.

Berbee, M. L. (2001). Fungal molecular evolution: gene trees and geologic time. New York: Springer-Verlag.

Lichtwardt, R. W. (1986). The Trichomycetes, fungal associates of arthropods. New York: Springer-Verlag.

McKerracher, L. J. (1985). The structure and cycle of the nucleus-associated organelle in two species of Basidiobolus. New York: Prentice Hall.

Abstract

Staphylococcus aureus is a round-shaped bacterium belonging to the Firmicutes phylum of bacteria. The escalation in the prevalence and gravity of staphylococcal contagions necessitates assessing the burden of asymptomatic carriage of Staphylococcus aureus in the community setting. The aim of this study was to establish the nasal carriage rates of S. aureus in microbiology students at RMIT. Nasal swabs were collected from 766 microbiology students at RMIT between 2013 and 2018. The swabs were inoculated on mannitol salt agar (MSA) and identified by their morphological characteristics. Tube coagulase tests were done to ascertain the identity of the pathogens. About 26.11% (200) samples tested positive for S. aureus. This value was comparable to those found in previous studies conducted in the community, hospital and food industry settings. The experimental nasal carriage rates varied from year to year, which could be attributed to inconsistencies in the streaking methods and biochemical tests. It was concluded that S. aureus was a prevalent normal flora in humans. As a result, it is important to observe high standards of hygiene to lower the risk of contracting infections attributed to the pathogen.

Aim and Significance

The purpose of this study was to conduct a survey of nasal carriage of Staphylococcus aureus in microbiology students at RMIT from 2013 to 2018. Nasal swabs from the subjects were collected and subjected to bacteriological analysis. Carriage of S. aureus plays a vital role in the epidemiology and development of infection. Eradication of carriage is a viable pre-emptive measure in at-risk individuals. Therefore, it is important to determine the carriage of S. aureus as part of developing appropriate measures to reduce the risk of S. aureus infection.

Introduction

S. aureus is an important pathogen as it has been connected to a wide range of infections. This bacterium is commonly found on the skin as well as in the respiratory tract and nose (Kobayashi, Malachowa & DeLeo, 2015). Numerous S. aureus infections happen because of its far-reaching virulence factors. The importance of this bacterium also rests on its high rates of resistance to antimicrobial agents as well as prevalence as a nosocomial disease-causing agent.

S. aureus possesses surface proteins such as laminin and fibronectin, constituents of the extracellular matrix that enhance its attachment to the host. Blood clots as well as epithelial and endothelial surfaces also contain fibronectin, which promotes the attachment of S. aureus to injured tissues via a fibrin-binding protein. Some virulence factors employed by S. aureus include antigens, enzymes and toxins. Capsules and adhesins are examples of antigens, while coagulase, hyaluronidase, nuclease and staphylokinase are examples of enzymes that promote the virulence of S. aureus. Virulent toxins include enterotoxins, P-V leukocidin and alpha, beta and delta toxins (Tong, Davis, Eichenberger, Holland & Fowler, 2015). S. aureus exhibits multifactorial virulence because of the collective action of numerous virulence factors and extracellular and cell wall constituents.

S. aureus causes many human diseases, particularly skin and soft tissue illnesses. Examples of minor skin infections include impetigo, pimples, boils, folliculitis, cellulitis, scalded skin syndrome and carbuncles. In addition, life-threatening disorders caused by S. aureus encompass meningitis, pneumonia, endocarditis, osteomyelitis, bacteraemia, toxic shock syndrome and sepsis (Kobayashi et al., 2015).

S. aureus is responsible for several nosocomial infections that plague health-care settings. This problem is aggravated by antibiotic resistance. Health-care workers are the major carriers of S. aureus. Studies indicate that nasal carriage of S. aureus in health-care staff ranges from 16.8 to 56.1% (Rongpharpi, Hazarika & Kalita, 2013). Moreover, S. aureus carriage has a significant impact on the food industry as indicated by the prevalence of food-borne illnesses. The World Health Organization (WHO) estimates that approximately 30% of the population in developed countries are afflicted by food-borne diseases annually (Rongpharpi et al., 2013). In contrast, 2 million deaths associated with food poisoning occur in developing countries every year. S. aureus may be present on the nose or skin of food handlers and is transferred to humid cooked foods that then become poisonous if unrefrigerated. High carriage rates of S. aureus in the community are linked to the appearance and universal dissemination of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA; Tong et al., 2015).

In Australia, approximately 1962 respiratory infections, 3946 surgical site infections and 1100 hospital-onset S. aureus bacteraemia cases were reported between 2010 and 2016 (Mitchell, Shaban, MacBeth, Wood & Russo, 2017). However, these numbers were considered underestimations because other disorders such as pneumonia and bloodstream infections had not been included in the statistics.

Other researchers have examined S. aureus carriage rates through cross-sectional studies (Bojang et al., 2017; Rongpharpi et al., 2013) and population-based surveys (Pires et al., 2014). A common technique employed in all investigations is obtaining nasal swabs and subjecting them to conventional bacteriological analysis.

Many cases of drug-resistant S. aureus have been reported over the last few years. One example of a reported strain is Methicillin-resistant S. aureus (MRSA), barely a year following the introduction of methicillin. Multiple drug-resistant S. aureus has also been isolated from food items and human nasal mucosa. These strains are a significant public health concern because they easily circulate in the environment.

Materials and Methods

About 766 microbiology students at RMIT were selected to participate in the study from 2013 to 2018. Nasal swabs were obtained from the participants using aseptic methods by rolling an applicator stick dampened with normal saline. A nasal swab from each student was inoculated onto mannitol salt agar (MSA, CM0085; Oxoid, UK) and incubated for 24 hours at 35 to 37 °C (Bojang et al., 2017). Isolates were identified as S. aureus by their growth features on MSA. Gram staining was conducted using conventional procedures (Microbiology Unit RMIT University, 2015). The tube coagulase test was performed as a biochemical test to verify the identity of the isolates (Grando & Paras, 2009).

Results

Yellow colonies with yellow pigmentation were produced in MSA medium, indicating the growth of S. aureus. The bacterial cells stained purple in the Gram staining test. All S. aureus isolates coagulated the plasma in the coagulase test. Among the 766 healthy microbiology students, the overall prevalence of nasal carriage of S. aureus was 200 (26.11%). Table 1 indicates the annual prevalence of S. aureus carriage from 2013 to 2018.

Table 1: S. aureus nasal carriage rates of RMIT Students from 2013 to 2018. Source: T. Istivan, personal communication, 2018.

Discussion

A nasal carriage prevalence of 26.11% was reported over a six-year period in 766 subjects. These findings were within the range of published data on nasal S. aureus incidence in health-care settings, the food industry and the community. Eke, Eloka, Mgbachi, Nwobodo and Ekpen-Itamah (2015) investigated the nasal carriage of S. aureus among food handlers in Nigeria. About 100 nasal swabs were collected from participants aged 21 to 40 years and evaluated using conventional methods. The prevalence rate was 60%, with males having higher incidence (58%) than females (42%). Rongpharpi et al. (2013) assessed health-care staff in Assam. Samples were collected from 315 subjects between August 2009 and July 2010. The swabs were inoculated on MSA, 10% sheep blood agar medium and MacConkey’s agar. Five biochemical tests were used: catalase test, tube and slide coagulase tests, mannitol fermentation and the modified Hugh and Leifson test (Rongpharpi et al., 2013). About 70 samples (22.22%) were positive for S. aureus.

Three studies checked the nasal presence of S. aureus in the general community with varying outcomes. Pires et al. (2014) studied an urban Brazilian population using 686 subjects aged one year and older. Nasal swabs were grown on Baird Parker agar. Species identification entailed Gram staining and biochemical tests (catalase, coagulase and fermentation of mannitol, trehalose and maltose). The isolates further underwent genotypic classification by Polymerase Chain Reaction (PCR). Younger subjects (average age 28 years) who had reported recent skin infections were more likely to carry the pathogen than their older counterparts. Bojang et al. (2017) conducted a similar investigation in rural Gambia using 1264 nasal swabs from 232 children aged between 5 and 10 years. The samples were analysed using conventional methods (MSA and coagulase test). Prevalence of S. aureus carriage was 25.9%. Chen et al. (2017) tested nasal swabs from 295 volunteers living on a medical campus. The samples were subjected to molecular means of S. aureus identification. Approximately 24.7% of the isolate samples were positive for the bacterium.

Methodological differences that accounted for the differences in the findings of previous studies included the use of assorted biochemical tests and different isolation media (Pires et al., 2014; Rongpharpi et al., 2013). Pires et al. (2014) and Chen et al. (2017) also used molecular methods, known to have high accuracy levels, to confirm their identification. The sampling also differed from one study to another.

The prevalence rates ranged from 22% to 26.11% in this experiment and four other studies. Only the Nigerian study by Eke et al. (2015) reported a higher prevalence of S. aureus than this experiment (60%). However, the method used by Eke et al. (2015) did not differ substantially from those used in this investigation. The high prevalence in this study could be attributed to differences in the distribution of S. aureus in developed and developing countries.

The prevalence of S. aureus at RMIT fluctuated over the years (Figure 1). This observation could be attributed to the lack of standardization of the methods used in swabbing nostrils, streaking techniques and performing confirmatory tests. Antibiotic-resistance strains of S. aureus were not identified in this study because antibiotic sensitivity was not done.

Conclusion

This study determined the nasal carriage of S. aureus in microbiology students at RMIT from 2013 to 2018, which was 26.11%. While this value was within the reported ranges in the community, food industry and hospital settings, these rates indicated a significant risk of contracting S. aureus infections in these three areas. Therefore, the study results reveal the need to observe high standards of hygiene to prevent the transmission of the bacterium and disease development.

References

Bojang, A., Kendall, L., Usuf, E., Egere, U., Mulwa, S., Antonio, M.,… Roca, A. (2017). Prevalence and risk factors for Staphylococcus aureus nasopharyngeal carriage during a PCV trial. BMC Infectious Diseases, 17(1), 588.

Chen, B. J., Xie, X. Y., Ni, L. J., Dai, X. L., Lu, Y., Wu, X. Q.,… Huang, S. Y. (2017). Factors associated with Staphylococcus aureus nasal carriage and molecular characteristics among the general population at a Medical College Campus in Guangzhou, South China. Annals of Clinical Microbiology and Antimicrobials, 16(1), 28.

Eke, S. O., Eloka, C. C. V., Mgbachi, N., Nwobodo, H. A., & Ekpen-Itamah, U. J. (2015). Nasal carriage of Staphylococcus aureus among food handlers and restaurant workers in Ekpoma Edo State, Nigeria. International Journal of Community Research, 4(1), 7-14.

Grando, D., & Paras, C. (2009). G.E.R.M.M. Melbourne, Australia: RMIT University.

Kobayashi, S. D., Malachowa, N., & DeLeo, F. R. (2015). Pathogenesis of Staphylococcus aureus abscesses. The American Journal of Pathology, 185(6), 1518-1527.

Microbiology Unit RMIT University. (2015). Medical microbiology techniques manual. Melbourne, Australia: RMIT University.

Mitchell, B. G., Shaban, R. Z., MacBeth, D., Wood, C. J., & Russo, P. L. (2017). The burden of healthcare-associated infection in Australian hospitals: A systematic review of the literature. Infection, Disease & Health, 22(3), 117-128.

Pires, F. V., de Souza, M. D. L. R., Abraão, L. M., Martins, P. Y., Camargo, C. H., & Fortaleza, C. M. C. B. (2014). Nasal carriage of Staphylococcus aureus in Botucatu, Brazil: A population-based survey. PloS One, 9(3), e92537.

Rongpharpi, S. R., Hazarika, N. K., & Kalita, H. (2013). The prevalence of nasal carriage of Staphylococcus aureus among healthcare workers at a tertiary care hospital in Assam with special reference to MRSA. Journal of Clinical and Diagnostic Research: JCDR, 7(2), 257-260.

Tong, S. Y., Davis, J. S., Eichenberger, E., Holland, T. L., & Fowler, V. G. (2015). Staphylococcus aureus infections: Epidemiology, pathophysiology, clinical manifestations, and management. Clinical Microbiology Reviews, 28(3), 603-661.

Introduction

To begin with, Escherichia coli (E Coli) are members of the Enterobacteriaceae family, which are gram-negative and rod-shaped. Further, E Coli is among the non-spore-forming bacteria with the flagella cell structures. According to Luna Guevara et al. (2019), E Coli bacteria can live in different environments with them having the capacity to survive with or without air. The optimal pH range of the E Coli O157:H7 bacteria is depicted to be 6.5 to 7.5 which depends on the temperature of their environments (Valilis et al., 2018). The common carriers of E Coli are contaminated water and foods, especially undercooked vegetables and foods, while the gastrointestinal tract is its reservoir.

Point of Entry

Majorly, individuals and animals contract the E Coli bacteria through ingestion; hence it is the main portal of entry. E Coli bacteria are found in contaminated water and foods from which individuals can pass to other people. Also, coming into contact with either animal or human feces or stool can lead to individuals getting E Coli bacteria. E Coli is a common type of bacteria that is found in the intestines of human beings and the gut of several animals (Valilis et al., 2018). The normal microbiota is found in the digestive system and plays an essential role in the defense of the body against microbial pathogens.

Mechanism of Pathogenicity

E coli have many virulence factors that include toxins, adhesins, and invasins that are normally encoded on plasmids together with other genetic elements. E Coli evades body defenses by producing proteins that alter the immune responses of the body (Nawell & La Ragione, 2018). Further, the produced proteins prolong the survival of the bacteria by being persistent in the immune system. Conversely, the symptoms of infection of the E Coli bacteria in an individual start appearing after three days of exposure. However, they vary across patients, with some becoming ill immediately after exposure to the bacteria.

Epidemiology of the Infectious Disease

Escherichia coli is characterized as a bacteria in the class of gammaproteobacteria and the family of Enterobacteriaceae. Research conducted by the Centers for Disease and Control (CDC) in the United States revealed that around 265,000 new cases of E Coli are reported every year hence portraying the significance of this disease across the country. However, the actual number of individuals diagnosed with E Coli each year is expected to be higher since the majority of patients do not go to the hospital for this illness but rather to the laboratory. E Coli is spread across individuals in scenarios where an infected person fails to clean their hands and comes into contact with other people hence facilitating person to person spreading through a continuous common source.

Characteristics of the Infectious Disease

Generally, the signs and symptoms of E coli patients might vary they might appear to some patients immediately after contraction while others take almost 5 days. The common symptoms among E Coli patients include abdominal pains, diarrhea, stomach cramping, nausea, and vomiting, which can be experienced for about two weeks (Nawell & La Ragione, 2018). E Coli infections are contagious for the period that the patient is going through diarrhea, while it may take longer among some patients. Further, E Coli patients have a higher chance of survival with side effects of developing heart and kidney problems.

Diagnosis, Treatment, and Preventative Measures

E Coli infections are diagnosed by conducting laboratory tests of the patient’s stool to look for the presence of E Coli bacteria. Fortunately, the majority of E Coli cases among patients heal on their own without the significant intervention of medical professionals. Treatment of this bacterium involves ample rest for the patient and increased consumption of fluids to prevent fatigue and dehydration for patients (Luna Guevara et al., 2019). In this scenario, antimicrobial drugs are not used since they may worsen an individual’s illness. Conclusively, there are no vaccines and medications that can be used to protect individuals from contracting E Coli. Individuals should avoid risky foods, and dirty water and maintain cleanliness to prevent the contraction of these bacteria.

References

Luna-Guevara, J. J., Arenas-Hernandez, M. M., Martínez de la Peña, C., Silva, J. L., & Luna-Guevara, M. L. (2019). The role of pathogenic E. coli in fresh vegetables: Behavior, contamination factors, and preventive measures. International journal of microbiology, 2019.

Newell, D. G., & La Ragione, R. M. (2018). Enterohaemorrhagic and other Shiga toxin‐producing Escherichia coli (STEC): Where are we now regarding diagnostics and control strategies?. Transboundary and emerging diseases, 65, 49-71.

Valilis, E., Ramsey, A., Sidiq, S., & DuPont, H. L. (2018). Non-O157 Shiga toxin-producing Escherichia coli—a poorly appreciated enteric pathogen: systematic review. International Journal of Infectious Diseases, 76, 82-87.

Purpose

The purpose of this academic report is to summarize the results of a study of the cultivation of two cell cultures, Gram-positive S. epidermidis, and Gram-negative E. coli, on three nutrient media obtained in a previous experiment. This involved the study of cell morphology and colonial growth efficiency. In addition, microbiological analysis of the cultivation of the Unknown bacterial specimen on two new media, namely Columbia CNA Agar and Eosin Methylene Blue Agar, was performed.

Columbia CNA Agar

Columbia CNA Agar (or C-CNA for short) is a classic example of a selective media enriched with two antibiotics, colistin and nalidixic acid, and a defibrinated sheep’s blood (5%) as a cell differentiation factor for hemolysis studies. This type of medium is standardly used to isolate Gram-negative bacteria, with a source of nutrients in the form of carbon and nitrogen from the casein hydrolysate that is part of the C-CNA. The enriched nature of the nutrient medium creates unique opportunities for the growth of fastidious cultures: due to inhibition of growth properties of Gram-negative bacteria, C-CNA is selective against Gram-positive bacteria in the presence of whole blood. The selectivity is due to two sources of antimicrobial effect at once, namely colistin and nalidixic acid. The principle of action of both is based on the destruction of the cell wall of Gram-negative organisms and blocking the replication processes of bacterial DNA (Zhang et al., 2019). As a result, bacteria with a thin cell wall cannot be grown on C-CNA-type medium. If, after incubation time (usually 18-24 hours), the plate shows apparent growth in the form of a stroke, this indicates the ability of the microorganism to survive in the selective medium. Consequently, the bacterium is gram-positive. On the contrary, if the streak does not appear after a period of time, this signals the sensitivity of the strain to the action of the antibiotic. Consequently, the bacterium is gram-negative.

Eosin Methylene Blue Agar

Eosin Methylene Blue agar is the second nutrient medium studied in this study: it is both a selective and a differential medium for the isolation of fecal intestinal bacterial forms. EBM agar is a complex medium because it does not contain growth or suppression factors as such. At the same time, the Peptic digest of animal tissue is a source of both carbon and nitrogen necessary for bacterial growth. The selective nature of EBM is justified by the possibility of separating bacteria according to their metabolic ability to ferment lactose. The separation is realized through the use of the pigments Eosin and Methylene Blue, which act as indicators. In other words, Eosin Methylene Blue agar is effective against Gram-negative bacteria because the toxic effect of Methylene Blue tends to inhibit the growth of Gram-positive prokaryotes: this is due to the porous structure of the cell wall of such bacteria without a protective membrane. As a result of the differential nature, some bacteria have a dark blue or black coloration when grown. Hence, these are the ones that can ferment lactose. On the other hand, if bacterial colonies are grown but are not colored, this indicates that they cannot ferment lactose.

Reference

Zhang, Y., Wang, X., Li, X., Dong, L., Hu, X., Nie, T.,… & You, X. (2019). Synergistic effect of colistin combined with PFK-158 against colistin-resistant Enterobacteriaceae. Antimicrobial agents and chemotherapy, 63(7), e00271-19.

Purpose

The purpose of this laboratory work was to learn laboratory skills through culturing procedures and studying the results of incubation. Two microorganisms (E. coli and S. epidermidis) were used for this work and were inoculated on three different media: broth, slanted agar, and agar plate by the streak-plate method. Based on the results of these inoculations, the differences in culturing the organisms were studied, and the advantages and disadvantages of each inoculation method were discussed.

Inoculation on Broth

Inoculation on broth is the cultivation of microorganisms on non-solid surfaces. The main advantage of this microbiological method is the different concentrations of nutrients and oxygen at different levels of the depth of the beaker: this allows bacteria with different aerotolerant needs to grow. In addition, it allows for a large stock of microorganisms for the studies. In contrast, through the broth, it becomes impossible to determine the purity of the culture grown and separate the bacteria from each other qualitatively. Tryptic Soy Broth (TSB) medium was chosen for inoculation in this study: both E. coli and S. epidermidis were cultured in this broth at 37℃ under constant agitation for 18-24 hours. To prepare TSB, 3 g of TSB powder is dissolved in 100 mL of distilled water while stirring on a magnetic stirrer as standard. Then, 10 mL of medium is placed in a glass test tube and autoclaved at 121℃ for 15 minutes: after that, the test tubes are cooled without direct exposure to sunlight. For cultivation, the broth temperature should be equal to room temperature.

Inoculation on Agar Slant

Inoculation on slant agar is used to establish a stock of bacteria for future studies, but its applicability is also realized for mixture differentiation. The advantages of this method are the creation of sterile conditions — by the aseptic technique — for long-term storage of microorganisms without the possibility of drying out. The key disadvantages of growth on slant agar are less visibility of the culture and, consequently, worse observability of their dynamics, and the inability to obtain large stocks of bacterial cultures. For culturing E. coli and S. epidermidis, Tryptic Soy Agar Slants medium was chosen, whose preparation does not differ from TSB. The same ingredients in the same amount are used to make the TSA medium, but the broth is poured into tubes at an inclination for solidification after autoclaving is completed. It is acceptable to add defibrinated pureblood without bubbles if hemolysis is investigated in a clinical setting. The two pathogens are cultured at 37℃ for 18-24 hours.

Inoculation on Agar Plate by Streak-Plate Method

Finally, cultivation on an agar plate by streak-plate method to isolate individual bacteria on a nutrient medium. The advantage of this method is that separate and independent colonies can grow on a plate; hence it saves laboratory resources and allows more efficient use of the dishes. Consequently, it is possible to distinguish pure colonies. However, disadvantages of this method include the possibility of cross-infection, difficulty in counting concentrations, and the inability to grow obligate organisms using this method (Tankeshwar, 2013). In this work, Tryptic Soy Agar medium was again used to cultivate E. coli and S. epidermidis with the difference that the liquid broth was poured into Petri dishes, and cultivation was performed via the streak-plate method. Consequently, the ingredients for preparation were not different, although the procedure for creating the medium was slightly modified. Incubation conditions are also the same and are 37℃ for 18-24 hours.

Reference

Tankeshwar, A. (2013). Chocolate agar: Composition, preparation, uses. Microbe Online. Web.

The flow of electrons takes place in redox reactions. They move electrons from a donor to an acceptor. This movement consists of the protons through a cell membrane. It works by creation of a concentration gradient. The difference in concentration helps in this transfer. Electrons move from a higher concentration of carbon irons to a lower concentration. This takes place through an electron chain. The carbon irons are transferred from the carbon donor to a recipient. Oxygen is used as an acceptor. Electrons can go into the chain through two levels. Either cytochrome or quinine levels are used. Ferrous is another donor. We see that they get into the chain through the former level. This is because iron is not an organ donor. Iron ions contain protons which are used in the transfer.

The transfer normally involves a series of reactions. These reactions are a combination of oxidation and reduction reactions. Sulfur also gives out electrons. These are in turn used in the transport chain. It should always be more negative than that which is accepting the electrons. It also works by creating a proton gradient. When there is difference in concentration between to membranes there is a possibility of movement of elements. These elements are electrons. Since the transfer takes place in a reaction there is always emission of energy.

The transfer occurs against a concentration gradient. Therefore there is the use of force. This brings about production of energy. The form of energy produced is in most cases in form of heat. The energy is helps in driving ATP. Iron will conserve the least amount of energy as compared to other donors. This is because more force used during the transfer (Astruc 389). Biomining is usually performed by micro-organisms. It involves degrading of the elements of minerals which are poisonous. This takes place in a mining process under some conditions. The pH should be neutral that is a Ph of 7. The acceptor should be present in order to take the electrons. Another condition is the concentration of one molar.

Pathogen evolves by obtaining new genes. This can happen through the contact between different bacteria. They develop resistance by combining their genes with the acquired ones. Metallosphaera sedula obtain genes from their environment. This takes place when they take up the DNA molecules. An example of these molecules is plasmid. The transformation of bacteria enables the cloning in genetics. The most likely mechanism is transduction. Another one is conjugation and lastly transformation. Bacterial transduction is a process by which a DNA is introduced though a virus. A virus gets access into the cell. Then it multiplies inside while transferring genes. Pathogens are very mobile. This made them spread faster and also the speed at which they reproduce was faster (Jackson 157).

Single nucleotide polymorphism, SNP analysis is a distinction at one part in DNA. It is the main distinction in genome. It is normally performed to find us disease vulnerability genes. It does this for every individual to determine genes. A clade is a group of components. It comprises an organism and all its components. It also has taxons. It is like a tree and its branches. That particular group is given a scientific name. The clade is normally nested in levels. These levels define the levels of classification. Node is where two branches meet, showing the likelihood with a pathogenic tree.

Works cited

Astruc, Didier. Organometallic chemistry and catalysis. New Mexico: Springer, 2007.

Jackson, Robert. Plant pathogenic bacteria: genomics and molecular biology. Norwich: Horizon Scientific Press, 2009.