The Presence of Micrococcus luteus in Canis lupus familiaris


Microorganisms consist of a single cell or cell cluster, and can also include viruses, which are considered to not be cellular (Madigan et al. 2015). The number of microorganisms in the world is too enormous to count or even comprehend, but if we look at a smaller sample, studying these microbes becomes somewhat easier. The sample in this study was taken from the mouth of a dog. It is known that a wide variety of microbes inhabit the mouth of both humans and dogs, but in a study done by Elliot et al. (2005), they found a significant difference in the cultivable oral microbes found in human and dog mouths. They found that the genera most frequently isolated from dog’s saliva were Actinomyces, Streptococcus, and Granulicatella, and the genera most frequently isolated from plaque were Porphyromonas, Actinomyces, and Neisseria (Elliot et al. 2005).

Micrococcus luteus has one of the smallest genomes of free-living actinobacteria sequenced to date (Young et al. 2010). It is a Gram-positive coccus shaped bacterium that belongs to the family Micrococcaceae, it is commonly found on human and animal skin as well as in water and soil. Because M. luteus is a part of the bacteria flora of humans it is not thought of as a pathogenic bacterium, but it is an opportunistic pathogen. In some rare cases, it has shown to give infections in immunocompromised patients (Smith et al. 1999). The genome of micrococcus lutes has shown many similarities with Kocuria rhizophila a closely related organism, and of late the ATCC 9341 strain known as M. luteus was reclassified as K. rhizophila (Tang et al. 2003).

In this experiment, I set out to isolate, characterize and identify a bacterium from an environmental sample. The sample was taken from a dog’s mouth and through a series of physiological test, genome sequencing, and antibiotic testing, characterization of the bacterial sample to the species level was achieved.


In order to identify our environmental sample, I started out with collecting my isolate from a dog’s mouth and placing it on TSA agar (Trypticase soy agar). The plate was inoculated at room temp, without light for five days until the first quadstreak was performed. After every quadstreak the samples were incubated at 37oC for 2-3 days, until the next quadstreak was performed, and then placed in a refrigerator to inhibit growth. In order to try and isolate the bacterial strain a total of five quadstreaks were performed. After the bacterium was isolated it was Gram stained (Lab 4, Gram staining protocol) in order to classify the bacterium and to determine if the culture was pure. Following the Gram staining, genomic DNA was extracted from the latest quadstreak, and sent to sequencing. In order to extract the DNA from the sample we used the PowerSoil DNA Isolation Kit (Lab 5, DNA extraction protocol), and the samples were sequenced in the UAF Core Lab on the MiSeq Illumina Sequencer. In order to analyze our genome sequence datasets, I used BaseSpace Ilumina for the genomic assembly I used the SPAdes genome assembler, to determine taxonomic assignment we used Kraken metagenomics, and to look at the functional gene annotation I used Prokka genome annotation (Lab 7, Bioinformatics protocol).

In order to further characterize my sample several physiological tests (Lab 6, physiological test protocol) were performed. A fluid thioglycolate test in order to determine the oxygen class of the bacterium, an oxidase test to determine if the strain had cytochrome C oxidase, a catalase test to see if the strain had the enzyme catalase, and a API 20 E test strip was used to look at 21 different physiological processes. The API 20 E test strip was incubated for 3 days in 37oC, while the test tubes from the fluid thioglycolate test were stored I room temp for 3 days.

The last procedure that was performed was an antibiotic disc diffusion test (Lab 9, Disk diffusion test protocol), to assess the susceptibility or resistance of our isolate to a variety of different antibiotics. Eight different antibiotics were tested: oxacillin, gentamicin, piperacillin, amikacin, clindamycin, trimethoprim, cefoperazone, and vancomycin.

Most of the data were collected and analyzed over a span of several weeks. DNA analysis of the isolate gave 98.86 % confidence to the species level for Micrococcus luteus, and only 19% of the sample was unclassified (Figure 1). All the analysis performed by BaseSpace showed results that were either equal to or above the guideline.

Figure 1. Results from the genomic sequencing using Kraken Metagenomics and Krona, in BaseSpace Illumina.


The identification test performed on the isolate are summarized in table 1.   The isolate tested positive for both the catalase test and the oxidase test, indicating that the isolate contained the enzyme catalase and cytochrome c oxidase. The gram stain came out red, but because both rods and cocci were observed, the culture was considered to not yet be pure, and more isolation attempts were needed. From the 8 different antibiotics tested, the isolate was susceptible to 6 of them, it was resistant to oxacillin and intermediate resistant to trimethoprim. The zone measurement for oxacillin was 5 mm while the zone measurement for trimethoprim was 13 mm. The other antibiotics tested were gentamicin, piperacillin, amikacin, clindamycin, cefoperazone, and vancomycin.

Table 1. Results for various identification test for the isolate.

Identification test Result
Gram Staining Mixed culture (red)
Catalase test Positive
Oxidase test Positive
Fluid thioglycollate test Facultative
Antibiotic resistance: Disk diffusion test Resistant to oxacillin and intermediate resistance to trimethoprim.
API 20 E test strip See Figure 2

From the last physiological test done, the API 20 E (Figure 2), 7 of the 23 tests performed were positive. The positive test was the LDC, URE, TDA, GEL, SOR, OX, and NO2 tests (Figure 2).

Figure 2. Results from the API 20 E test strip


M. luteus is described as the type species of the genus Micrococcus (Stackebrandt et. al. 1995), and is commonly found on human skin, and on animals, it can also be found in the mouth, and in the upper respiratory tract of animals and humans. The bacterium can likewise inhabit many other areas in the environment, like water, dust, and soil (Kocur et al. 2006).

In relation to the literature, the physiological test results were mostly consistent with information found. Nevertheless, there was one inconsistency found with the fluid thioglycollate test. I observed the bacterium to be facultative, although literature describes the bacteria as an obligate aerobe (Kocur et al. 2006). This may be due to human error or to the sample being impure.

The genus Micrococcus was first described more than a hundred years ago, but since then the description has been revised several times (Stackebrandt et. al. 1995). Today it is clear that this genus of bacteria is gram-positive, cocci shaped and catalase positive (Stackebrandt et. al. 1995). During our research, I found our isolate to be a mixed culture, and not gram positive as literature states. When observing the isolate under the microscope after gram staining it appeared red, with both rods and cocci present. Several isolation attempts were done, but the two bacteria observed seemed to live in symbiosis which complicated the isolation process. However, isolation was achieved after several quadrant streaks.

Numerous interesting studies have been done on the genus Micrococcus and their traits. Dib et al. (2013) discussed how genes present on the plasmids of Micrococcus bacteria can give advantageous features to their respective hosts like antibiotic and heavy metal resistances, the ability to degrade cholesterol, and osmotolerance. A study conducted by Greenblatt et al. (2004) looked at survival of Micrococcus in extreme environments. They found that M. luteus and other closely related cocci that are non-spore-forming seem well suited to extreme environments, all due to special individual factors (Greenblatt et al. 2013). Research has been done on M. luteus superior ability to absorb radiation through pigments that absorb long-wave UV radiation, between 350-475 nanometers, and the researchers hope to implement this into sunscreen and other cosmetic products (SINTEF 2013).

For further research, it would be interesting to look more closely at some of the traits that make M. luteus so good at surviving in extreme conditions, and absorb such long wavelengths of radiation. I only looked at some physiological traits of the bacterium, it would be interesting to see how these aid the bacterium in extreme settings, like the ones previously mentioned.

Our research looked at bacteria within our environment, and our sample was taken from a dog’s mouth where we found the bacterium M. luteus. We conducted several physiological and antibiotic tests, along with genome sequencing in order to understand and characterize the bacterium. Most of the test that we conducted were consistent with literature, but there were some variances within gram staining and the fluid thioglycollate test. However, the tests gave us important information about the bacterium and a better understanding of the environment it lives in.


Literature Cited

Dib, J., Liebl, W., Wagenknecht, M., Farías, M., & Meinhardt, F. (2013). Extrachromosomal genetic elements in Micrococcus. Applied Microbiology & Biotechnology, 97(1), 63-75.

Elliott, D. R., Wilson, M., Buckley, C. M., & Spratt, D. A. (2005). Cultivable Oral Microbiota of Domestic Dogs. Journal of Clinical Microbiology, 43(11), 5470-5476.

Greenblatt, C. L., Baum, I., Klein, B. Y., Nachshon, S., Koltunov, V., & Carlo, R. J. (2004). Micrococcus luteus – Survival in Amber. Microbial Ecology, 48(1), 120-127.

Kocur, M., Klosss, W. E., & Schliefer, K. (2006). The Genus Micrococcus. Prokaryotes, 3, 961-971.

Madigan, M. T., Martinko, J. M., Bender, K. S., Buckley, D. H., & Stahl, D. A. (2015). Brock biology of microorganisms (Fourteenth edition.). Boston: Pearson.

SINTEF. (2013). Super sunscreen from fjord bacteria. ScienceDaily. Retrieved April 17, 2017 from

Smith, K. J., Neafie, R., Yeager, J., & Skelton, H. G. (1999). Micrococcus folliculitis in HIV-1 disease. British Journal of Dermatology, 141(3), 558-561.

Stackebrandt, E., Koch, C., Gvozdiak, O., & Schumann, P. (1995). Taxonomic Dissection of the Genus Micrococcus: Kocuria gen. nov., Nesterenkonia gen. nov., Kytococcus gen. nov., Dermacoccus gen. nov., and Micrococcus Cohn 1872 gen. emend. International Journal of Systematic Bacteriology, 45(4), 682-692.

Tang, J. S., & Gillevet, P. M. (2003). Reclassification of ATCC 9341 from Micrococcus luteus to Kocuria rhizophila. International Journal Of Systematic And Evolutionary Microbiology, 53(4), 995-997.

Young, M., Artsatbanov, V., Beller, H. R., Chandra, G., Chater, K. F., Dover, L. G., … Greenblatt, C. L. (2010). Genome Sequence of the Fleming Strain of Micrococcus luteus, a Simple Free-Living Actinobacterium . Journal of Bacteriology, 192(3), 841—860.


Art Project

Planet of Microbes by Aasne Hoveid

I took base in one of the first pictures that were on the syllabus and I wanted to create something that related  to the diversity and abundance of microbes in the world. Therefore I chose to make a mosaic, or use many small photos to make this one large photo. I chose to use photos that I have taken of various microbes along with several other photos of microbes in order to make this big picture. My idea was to try and show the diversity of microbes and how they shape and have shaped the world we live in.


Microbes in The News: Transplanted gut microbes may protect babies from infection

Transplanted gut microbes may protect babies from infection

Jana Howden: Cosmosmagazine, 21. April  2017

A  new study in mice showed that particular gut bacteria can provide newborns with a vital protection against infection. A bacterium known as Clostridia helped the mouse pups to digest food and to be protected from infection. The researchers then exposed these mice to C. rodentium and found that only the mice transplanted with added Clostridia were able to resist infection. The team is now conducting further research to uncover the role of Clostridia in defending newborns against infection, human clinical trials are a possibility if the bacterium’s protective potential bears fruit in further animal studies.

I think this relates to a lot of things that we have discussed in this class and many  especially to the lectures on immunology and on pathogens. I found this article  to be very well written and with a lot of relevant and interesting information. I thought  the article did a good job at relaying the information.

Why and how does  Clostridia  protect newborns  from infections?

Microbial Worlds


Impermafrost by Gail Priday

I really liked Gail’s art pieces I thought they were simple but that they also really captured the essence of microbial worlds and the idea that microbes are everywhere. I liked that she took concepts that we see a lot in Alaska, like permafrost, and applied that into her work. I think what mostly captured me by her work, was her use of colors, which I thought was done very well and in a simplistic way, I think it really captures the diversity of microbes.

I definitely think she captured the idea of permafrost, and the vast majority of microbes that are frozen in the permafrost, but because of global warming they are now starting to become exposed, and how that affects the global climate.


Beach Plastic by Margo Klass

I thought Margo Klass’s peace on plastic waste was very interesting and a good way of giving people a visual of how the increased need for plastic, is also responsible for the accumulation of it, and how it has become a global concern. Margo collected plastic waste form beaches in Oregon for three years and made this art piece to try and draw attention to the problem (plastic pollution). In her statement, she also talked about how scientist form China and the US recently discovered that polystyrene and Styrofoam can be biodegraded by mealworms.

From my perspective, I think her art piece is successful in what she is trying to do, raise awareness for plastic accumulation in the environment. I think that the information provided makes the whole piece come together.


EMERGENCE: The Warming Climate is Waking Up Sleeping Microbes by Nancy Hausle-Johnson

I think Nancy’s piece connects well with the lecture we had on ecology and the carbon cycle, but also to lectures on microbial diversity. When permafrost melts many of the microbes that have been lying dormant suddenly emerge and become a part of the bigger ecosystem. These microbes also become available to bigger organisms meaning that they get brought back into the carbon cycle.

Nancy’s piece features microbes that are emerging due to the melting of permafrost, and how at different stages of thawing influenced the diversity of microbes they saw.


What sort of piece might you have created?

I’m not sure what I would work with, but I do enjoy taking pictures and the idea of how microbes can help us work against global warming and climate change. Therefore I think it would have to have something to do with that.

Microbes in the News.

These bugs could help Seattle’s poop spill. But they’re hibernating.

KUOW Seattle news and information: Tuesday, April 4th

The West Point treatment plant

The West Point Treatment Plant in Seattle  was crippled by a flood last month, and now it continues to spit solid waste into Puget Sound. In order to clean up after the spill the treatment plant relies on microorganisms to  break down the solid organic material that’s in the waste water. The microorganisms need heat and food to survive, but the flood damaged the boilers that heat the plant. Therefore, the microorganisms have been dormant or in hibernation ever since the treatment plant got damaged, and until the boilers are fixed and the heat restored, the microbes will remain in hibernation.

I think this best ties to lectures that we had earlier this semester on growth and nutrition, but also to lectures about microbial metabolism and physiology. I did find this story pretty amusing, although it wasn’t a very technical article about microbes and their function I do think that it gives relevant and decent information.  I thought it was interesting how the microbes could go into a dormant state, and also how they used the microbes to clean up after the spill, but how exactly do the microbes clean up after a spill like that?



Extra Credit: Dan Stinchcomb Seminar

Dan Stinchcomb lecture was titled “Development of Vaccines for Mosquito Borne Tropical Diseases”. He talked about different viruses and how they can spread rapidly and how human density favors the emergence of new viral pathogens. He mentioned Dengue fever, West Nile Virus, Chikungunya disease, Zika Virus and how they worked to develop effective vaccines for these diseases. He went through the distribution of the disseases and the cause and effect and how we could work to prevent them, mostly talking about vaccines. He gave a lot of information on the development of vaccines, trial methods, safety, etc. he also talked about IDRI and the promise of RNA vaccines.

I thought it was a really good seminar, although some parts of his lecture was very technical, i did understand most of what he was talking about, and i think it is a very important topic. This connects closely to the virology proportion of our class, although his lecture went more in depth than what we did in class. Is RNA vaccines the future? Can they (in the future) replace the other vaccines that we have today?

Microbes in the News: Researchers Uncover Clue about How Tiny Microbes Self-mutate.

Laboratory Equipment: Monday, April 3rd 2017

Researchers just discovered a genetic element that enables a group of unidentified microorganisms to self-mutate. Researchers found genetic elements, called diversity-generating retroelements (DGRs), that enable the microbes to target their own genes for accelerated mutation. They found that a majority

An ultra-small cell of a bacterium that may be a relative of the self-mutating microbes.

of ac certain class of Archaea, as well as some yet-to-be-characterized categories of organisms closely related to bacteria, appear to have DGRs. The DGRs target the nucleotide adenine to initiate a new mutation. Much is still unknown about the newly discovered microorganisms.

This ties in with a lot of subjects that we have talked about in class, especially microbial evolution, because this shows how fast evolution and adaptation occurs in microbes.

I did find this story very interesting, and although much is still unknown about this subject, i think the article gave sufficient evidence and facts in order for the reader to understand the concept. I thought this was very interesting and i look forward to seeing further research on this subject.

The biggest question I had after reading this was, how? How can a microorganism self mutate? This is probably the biggest question that the researchers had as well.


Painting with Microbes

I did not really have any intent, but I was looking at the different colors of bacteria provided and I noticed that there was a lot of red and
white/yellow. I asked my lab partner what things I could draw that were white and red, and she suggested a candy cane, so that’s what I attempted to paint.

I did my painting on the TSA agar which is a complex medium, which we know is used as a general growth medium for many different typed of bacteria. I used the S. marcescens to create the red color and I think it was the s. enteritidis that I used for the white color.

Extra Credit: Eric Collins

In Eric Collins seminar, he presented some very interesting results from studies done in Arctic marine environments. He presented a lot of interesting and unknown (to me) facts about the Arctic ice and how the global environment is affecting it. From the topic of ice, Collins moved into the topic of microbial diversity in arctic environments. He talked about how different techniques were used in order to investigate the diversity and function of Arctic bacteria, archaea, viruses, fungi, etc. and how this was important for the several ongoing projects that are being conducted in the Arctic, which in turn will be used to build a conceptual model of diversity flux.

I think this was a very interesting seminar, and I really liked how he presented the information with the help of graphs and figures to better the understanding of what he was talking about. In class, we have talked about different factors that affect microbial growth (temperature, nutrient availability, etc.), which Collins also mentions about in his seminar.

Because the ice in the Artic is melting so quickly lots of microbial diversity is lost, is there any way of measuring this loss?

A1: Intro post

Hi! My name is Aasne Hoveid, im am a junior studying biology. I am born in a small town called Alta, in the very north of Norway. Im not shure what i want to do after im done with my undergrad, if il stay in Alaska or go back to Norway. Im really looking forward to this class!This is a picture of me and my dad, rafting down Copper River.