Saturday, April 2, 2016

Garden Creation Story

In the beginning God created heaven and earth. I do not believe the biblical creation story is a historical account of the beginning of life, but the myth of Adam and Eve still has deep meaning. Whenever I have to weed the garden I always think, “Cursed is the ground for thy sake… Thorns also and thistles shall bring forth to thee; and thou shalt eat the herb of the field; In the sweat of thy face shalt thou eat bread, till thou return unto the ground; for out of it wast thou taken: for dust thou art, and unto dust shalt thou return.“  The thorns and thistles that we are punished with are the little challenges of life. I’ve never heard a sermon on that but I like to think that is what it means. I kind of enjoy weeding the garden. Sometimes I like to think that Eve and Adam were quite bored in a garden that required no work. Imagine all the plants coming up by themselves like winning some sort of garden lottery without real appreciation for the work put in to the planting and caretaking. Gardening would be dull and unchallenging without weeds, thorns, bugs and sometimes bad weather. The weeds of life and the weeds of the garden work in the same way because they keep us challenged and distracted enough to keep us out of trouble.

Gardens are mentioned many times in the Bible. In the Song of Solomon, the woman’s body is compared to various plants. The Garden of Gethsemane literally means “oil press.” I’ve read that there are very old olive trees in that area that can be 900 years old. In the book of Esther the king’s gardens are mentioned quite a bit. In Ecclesiastes, Solomon talks about the beautiful gardens that he had planted. The city of Babylon is written about in the Bible, but I’m not sure that there is mention of the Hanging Gardens of Babylon. Historians are not sure if the Hanging Gardens were real or poetry. I like to think that the garden was real. There are people who have gardens with only plants that are mentioned in the Bible. I find that an interesting project and it would be a challenge.

Friday, April 1, 2016

The Smell of Soil

Pea plant wrapping itself around the previous year vine
Soil has a particular aroma but it is not the soil itself but an actinobacteria which produces geosmin. I like the earthy taste of freshly dug potatoes or carrots. There is a saying that a good farmer can know when to start planting by picking up a handful of soil and smelling it. This is a true statement considering the actinobacteria only thrives in warm soil. Since most seeds need a soil temperature between 50-77F which is optimum for the life cycle of the species of Streptomyces that I studied for a semester in microbiology. Below is an excerpt from my essay:   

Description of an actinobacterium isolate from soil sample taken in the vicinity of the Oconaluftee River, Great Smoky Mountains National Park
12 December 2012


There are over 500 recognized species of Streptomyces, which is a testament to how important the genus is to humans. Many Streptomyces species play important roles in the decomposition of organic matter and are ubiquitous in soil accounting for 106-107 per gram of soil. [10] The genus is an important source of antibiotics for both animals and plants. The genome of Streptomyces are typically large and that is attributed to the high number of genes required for antibiotic synthesis.[7] Chromosomes are linear, much like eukaryotes, and the chromosomal ends functions similar to the function of telomeres in eukaryotic cells. [11]

The isolated colony used for the study on the R2A agar was 1-2mm in size with an opaque consistency, circular appearance and curled edges. There was a zone of inhibition around the first colony which was undetectable with subsequent inoculated plates. The isolated colony appeared to change color from white to dark pink-purple with time, possibly due to nutrient depletion of the agar. The cause for the coloration of the colony is due to pigmented conidia and sporophores. [7] Depending on the composition of the medium, the colony can appear blue, gray, green, red, violet or yellow. [10] The life cycle of Streptomyces involves sporophores which form cross-walls in the multinucleate sporophores produce conidia also known as conidiospores or spores. [7] Conidiospores germinate to make substrate mycelium. [10] Dormant spores can readily be dispersed by air currents. [9] The colony had a particular earthy odor which was caused by a group of volatile organic compounds known as geosmins. [10]

Gram staining techniques showed the Streptomyces spp. to be gram-positive. Because gram-positive microorganism have cell walls a thicker layer of peptidoglycan, the crystal violet-iodine complex gets trapped in the cell wall making the cells appear dark purple. A negative staining technique was also performed to enable viewing the microorganism morphology and cellular arrangement. With the aid of a compound light microscope, the microorganism appeared filamentous with diameter sizes of approximately 1µm and a variable length size of up to 80µm. According to data collected about Streptomyces, the filament sizes are typically 0.5-1µm in diameter and indefinite in length. [7]    

Environmental parameter tests were performed to find the optimal environment which the microorganism could grow. The tests showed Streptomyces spp. to be a facultative anaerobe, but the growth was slower than aerobic environments. Temperature parameter testing was used to determine if the Streptomyces spp. could thrive with a range of temperatures. The results showed the species does not tolerate extreme temperatures of 4°C or 50°C and only two of the three streaks grew in 37°C. The microorganism thrived in the control condition of 25°C. A salt tolerance test was performed using concentrations of 0%, 5%, 10% and 15% NaCl. The Streptomyces spp. is halointolerant and only grew under the control conditions of 0% NaCl. A pH test also conducted and it was found the microorganism can tolerate a range of pH and growth occurring from pH 5-9. The pH test results show the growth of the microorganism in alkaline to neutral soils is more favorable than acidic.

Results from the environmental parameters test demonstrate the variable soil pH the Streptomyces can tolerate. Since the microorganism can tolerate lower oxygen levels as a facultative anaerobe, the depth at which the organism can grow varies. In one study Streptomyces was found in a three different locations of prairie soil at four depths. The study used variation in the 16S rDNA sequence to determine the genetic variation among the Streptomyces strains. There was little variation found in the genetic diversity from the isolates taken at differing soil depths. [5]  An extreme Streptomyces has been isolated from the hyper-arid Atacama Desert. Streptomyces desertai can grow in temperatures from 10°C to 35°C between pH 4 and 11 and in the presence of 4% NaCl. [12]

A modified catalase test was used to determine if the organism had the enzyme. Because the genus does not typically react with the standard amount used for a slide test, a modification of using more of the sample directly on the R2A plate was used. The Streptomyces spp. tested positive for catalase, which means the microorganism has the ability to convert hydrogen peroxide into water and gaseous oxygen. The microorganism does not have cytochrome c oxidase as a respiratory enzyme because it the results were negative for oxidase test. [1]

An Enterotube II was used to determine the metabolic characteristics of Streptomyces spp. The test included possible reactions with glucose, adonitol, lactose, arabinose, sorbitol and dulcitol fermentation, lysine and ornithine decarboxylation, sulfur reduction, indole production, acetin production from glucose fermentation, phenylalanine deamination, urea hydrolysis, or citrate utilization. The organism tested positive for the characteristic to ferment glucose and had a slow reaction with hydrolyzing urea.[1] The Streptomyces spp. uses the carbohydrate, glucose, as a sources of carbon and energy. A positive result for the enzyme urease means that the organism hydrolyzes urea to ammonia, CO2 and water. Despite the Streptomyces spp. inability to react with the majority of medium found in the Enterotube II, the microorganism has the ability to decompose biopolymers like lignocellulose, starch, chitin, pectin and latex. [8][10]

The rhizosphere zone in the soil profile is defined as the zone of soil that adheres to plant roots. Streptomyces thrive in the rhizosphere zone with the aerobic conditions and loose loamy soil. [9] With 106-107 Streptomyces per gram of soil, the microorganism is ubiquitous in the rhizosphere.[10]  Because of the filamentous mode of growth, the Streptomyces has a competitive advantage in colonizing around plant roots. Streptomyces decompose organic matter such as plant residues, but also benefit plants by growing in close associating with the root system where they protect the plant roots from potential invasion by fungal pathogens. Studies have shown with legumes, the Streptomyces assimilate iron from the soil and transfer it into root nodules where it is assimilated by bacteroids. [8] Streptomyces virginiae has been found to be beneficial in the prevention of tomato wilt caused by a soil-borne plant pathogen called, Ralstonia solanacearum. [3] The use of microorganisms to combat plant pathogens is only in the beginning stages, but using biocontrol microbes is considered to be a less invasive alternative to pesticides and fertilizers. The natural antibiotics produced in the rhizosphere are thought to cause less stress on the indigenous microbes when compared with chemical fungicides. Streptomyces can easily colonize the plant root surfaces because of the filamentous growth pattern, they can help protect roots against pathogens and can decompose organic matter. The spores can also be dried, packaged into powdered products making it a potential candidate for future commercial use. [8]

Within Actinobacteria there are many organisms that have antibacterial and antifungal characteristics. In a study involving fungus-growing termites, antibiotic screening has shown that most Actinobacteria throughout the termite nests produced molecules with antifungal activity. [6]

The zone of inhibition around the colony in the original first colony collected for this study demonstrated the antibiotic characteristics of the Streptomyces spp. It is interesting to note an organism is sensitive to the antibiotics made by other streptomycetes. The production of antibiotics is poorly understood, but it is linked with sporulation which may be a mechanism to inhibit the growth of nearby organisms competing for resources. There are over 500 distinct antibiotics produced by stretomycetes. [7] Over half of the commercially produced antibiotics originated from the genus. [10] Streptomycin and is often used in patients who are allergic to penicillin. The antibiotic Daptomycin is produced by Streptomyces and is used against pathogenic staphylococci and streptococci. Streptomyces platensis produces the antibiotic, platensimycin, which is effective against methicillin-resistant Staphylococcus aureaus and vancomycin resistant enterococci. The first treatment used against tuberculosis originated from Streptomyces griseus. [7] It is unclear if Streptomyces produce antibiotics in their natural habitat.[10]

A possible change to the study could involve using more cells for the metabolic tests. For example, by using more cells than usually needed for the catalase test, a positive reaction with hydrogen peroxide occurred. Using mediums other than those found in the Enterotube II may result in finding more metabolic characteristics of Streptomyces spp. Studying why the color of the colony changed with age would or possible advantages for the organism to have a variety of colors may be an interesting focus for future studies. Further studies may also include why humans are able to smell the compound and how it could have been useful to humans from an evolutionary perspective.