Isolation of a given bacteria is an important procedure for the scientific ad medical fields.  Isolating pure cultures helps find, study, identify, organize, and produce valuable resources that occur naturally in the environment.  The primary bacteria in this lab is Streptomyces which often produces a variety of antibiotics that have huge beneficial qualities. Antibiotics play a huge role in medicine.  They help prevent and kill many microbial diseases that humans are susceptible.  Studying Streptomyces helps scientists find new and important medicines to kill off dangerous microbes.

    Streptomyces that produce antibiotics are generally found in soil. There are no specific qualities that the soil needs to have in order to support the bacteria.  We believe that by gathering a variety of soil samples, we will be able to find, isolate and correctly identify that we have gathered a pure sample of Streptomyces.  Some but not all of the strains of Streptomyces that have been collected will produce antibiotics. However, we are confident that with our media and protocol we will be able to isolate the an antibiotic producing strain of Streptomyces.

    We gathered samples of soil from all over the University of Maryland. Yet we did not isolate Streptomyces from all of the sampled collected. Samples from Denton Community, by the stadium, South Campus and from underneath uprooted trees were taken.  It was only found though from soil taken from potted plants kept in an apartment.  This suggests that a potting soil of manufactured and altered soil sample is better for finding and isolating Streptomyces.  This does not mean that Streptomyces was not present in the other samples, but perhaps our protocol was not the best method for isolating Streptomyces in natural soil samples.  The samples containing Streptomyces were dark brown black soil with a strong pungent odor.

    First we heated the soil samples for 1.5 hours at 60 degrees Celsius  to allow for sporulation and to perhaps kill off other bacteria that is heat sensitive.  We prepared serial dilutions  and plates with soil sprinkled over for each of eight collected samples, incubating at two temperatures. References suggested 25-30 degrees, so we incubated at room temperature (25 degrees) and then in the dark at 30 degrees.  Both temperatures showed growth, yet the 30 degree incubation plates showed more growth over a given time.  An interesting inconvenience is that even though cyclohexane was added to the plate medium to prevent the growth of mold, large amounts of mold did grow on a majority of the plates.

    After our first incubation, there was a wide range of bacteria growing on the plates.  Each varied in size texture and color.  This occurred because the medium was not a selective media.  Instead, the media was designed to support Streptomyces, yet as a side effect, many other bacteria were able to grow in the same media.  After some research, we selected from opaque, rough, dry colonies that were difficult to sample from.  There were both white and red colonies that fit these characteristics.  These colonies were re-streaked to obtain pure cultures for further testing.  After incubation of the pure cultures, only four produced the selected phenotype we were hoping for.  Of these four, all were gram positive, yet one sample was not pure.  The major problem faced with the contaminated sample was mold contamination.  Two samples were then rejected with two remaining, both from soil samples that came from a processed potting soil.  MacConkey¹s Agar plated confirmed the gram stains of the remaining samples because neither of
the.  The bacteria was filamentous branched with aerial mycelium which is a very distinct characteristic of Streptomyces.

    Streptomyces contains catalase in order to break down the toxins that oxygen produces.  We performed a catalase test on the samples and all were found to be positive for catalase.  An oxidase test was also performed and all of the tests were negative for that.

    Many species of Streptomyces produce antibiotics.  This is a characteristic that allows Streptomyces to survive and compete with other organisms over resources and  survive the presence of some bacteria.  We tested for the presence of antibiotics in out samples of identified Streptomyces.  We punched out circular sections of media from the samples. It was assumed that after a given incubation period that the Streptomyces would have seeped into the media, so that if antibiotics were produced they too would be in the media.  The circular cut outs of media were put on a lawn of e.coli and incubated for 4-7 days.  After the incubation, there was a zone of inhibition around one of the circled of media, proving that sample as producing antibiotics.  The inability to conduct additional tests kept us from proving that the other samples did in fact produce antibiotics.  This could have been done by testing other bacteria with the Streptomyces. There may also be more than one strain of antibiotic for each of the samples, but given time constraints, this could not be tested.

    We concluded that we did in fact isolate one strain of Streptomyces that produces antibiotic.  The other samples of Streptomyces may in fact be antibiotic producing, but more testing is required to discover the variety of strains present and the strength of each antibiotic produced by Streptomyces.