Isolation and Culture of Slime Molds Essay Sample
Zoospores are motile. nonsexual spores holding one or more scourges. They may be monoploid or diploid and are formed inside a spore case. discharged from the spore case to go a free swimming phase. and after the motile phase. the zoospore encysts in a suited substrate or host. ( Lange & A ; Olson. 1983 ) . Teasing technique is a method that allows farther growing of fungous mycelia on prepared come-ons. Major groups of Fungis isolated utilizing teasing techniques are those from Kingdom Chromista. peculiarly in the Class Oomycetes. The Oomycetes or “egg fungi” . besides called H2O casts were antecedently grouped with Fungis based on their filiform growing and the presence of coenocytic hyphae. one of the morphological traits and feature of Fungi. However. their life rhythm. unlike that of many Fungis. includes a diploid. asexually or sexually reproducing stage. ( Madigan. et Al. . 2009 ) . Oomycetes differ from Fungis in other cardinal ways every bit good. The cell walls of Oomycetess are typically made of cellulose. non chitin. as for Fungi. and they have flagellated cells. which are missing in all but a few Fungi. ( Madigan. et Al. 2009 ) Zoospores are produced by about all Oomycetes.
They are biflagellate organic structures. holding one whiplash scourge directed rearward and another tinsel type scourge directed frontward. The scourge are attached anteriorly or laterally. ( Sharma. 2011 ) . Interestingly. zoosporic fungi belonging to Phylum Chytridiomycota of Kingdom Eumycota are besides normally isolated utilizing the baiting technique. Chytridiomycetes. or chytrids. are the earliest diverging line of descent of Fungi with about 100 genera consisiting of about 1000 species. Their name refers to the construction of fruiting organic structure ( “little pot” ) which contains zoospores. Cell wall contains chitin like other true Fungis. However they differ from other Fungis. in bring forthing motile. flagellated spores called zoospores. a possible leftover of their version to aquatic environment. largely freshwater and moist dirts. where they are normally found. ( Madigan. et Al. . 2009 ) .
Harmonizing to JW Deacon & A ; G Saxena ( 1997 ) . Chytridiomycetes are really common as saprotrophs. facultative parasites and compel parasites in moist dirt and fresh water home grounds. They depend on their zoospores for dispersion and site-selection. In fact. this is how the chytridiomycota can be detected – by puting come-ons such as works seedlings. pollen grains. insect exoskeletons etc. in H2O or wet dirt. because the zoospores encyst on these stuffs and so give rise to microscopic settlements. This survey was designed with two to three aims. The first was to measure the possible beginnings of fungi turning on the prepared come-ons. second is to discourse how these zoosporic Fungis reach their substrates and thirdly. to qualify the fungal isolates based on their come-ons. The ultimate end of this survey is to insulate and civilization zoosproic fungi from different come-ons like dead insects. droppingss. and pollen grains. Morphological and generative word picture will besides be conducted on the stray Fungi.
A. Isolation and civilization of zoosporic Fungis from pollen and snake tegument utilizing baiting technique
Baits such as pollen grains ( Pinus or any other flower-bearing workss ) and snake or lizard tegument ( 2 pieces. ? inches squares ) were obtained and brought to the research lab. Pond Waterss with attach toing dirt or works dusts were collected and placed inside a disposable petri home base for each come-ons. Baits ( one come-on for one home base ) were placed on the pool water-filled disposable petri dishes. The civilizations were incubated at room temperature under cool conditions. All of the come-ons were examined under the microscope after 2 yearss and at day-to-day intervals thenceforth. Pollen grains were examined 12 hours after civilization and thenceforth. To detect the civilizations. the group obtained a little part of their come-ons. placed them each on a glass slide and was straight observed under the microscope. The hyphae. generative constructions and zoospores were to detect. Observations were recorded by taking images seen under the microscope.
B. Isolation of Zoosporic Fungi from dead insects and animate being droppings utilizing Baiting technique
Dead insects ( 2-3 pieces of cockroaches ) and dried shrimp exoskeleton ( 2-3 decalcified spots. ? inch square ) were obtained and brought inside the research lab. Fresh dirt samples were collected and placed inside a disposable petri home bases. Baits ( one come-on for one home base ) were placed on the soil-filled disposable petri dishes. The civilizations were incubated at room temperature under cool conditions for 24 hours. After 24 hours. the civilizations were observed and the dirt was moistened utilizing a wash bottle. Moist environment will assist for the growing of the Fungi. All of the come-ons were examined under the microscope after 2 yearss and at day-to-day intervals thenceforth. To detect the civilizations. the group obtained a little part of their come-ons. placed them each on a glass slide and was straight observed under the microscope. The hyphae. generative constructions and zoospores were to detect. Observations were recorded by taking images seen under the microscope.
C. Preparation of RDA and PDA for pure isolates of the civilizations
Correct sum of murphy dextroglucose agar was measured utilizing the ternary beam balance. The media was autoclaved before pouring into glass petri home bases. The figure of PDA home bases that were prepared was harmonizing to the figure of fungous zoospores observed on the civilizations of shrimp exoskeleton and cockroach. The said zoospores were inoculated aseptically on the prepapred PDA home bases. RDA media was besides prepared by boiling one coney droppings per 100 milliliter of distilled H2O. Two coney droppingss were boiled in a beaker incorporating 200 milliliter distilled H2O for 10 proceedingss and filtered utilizing a filter paper. Correct sum of Bacto agar was measured. with the given 15g/L. This was added to the filtered droppings H2O to function as solidifying agent of the media. The prepared media was subjected to autoclave before pouring on home bases. The spores from the coney dung damp civilization were inoculated on the RDA home bases. All of these will be stored for 1-2 yearss under cool conditions. The turning pure civilizations were observed macroscopically and under the compound or dissecting microscope.
As the consequence of the experiment described above. the civilizations were successfully incubated at room temperature and were observed under compound or dissecting microscope. Isolates of the chitinolytic Fungi ( civilizations of shrimp exoskeleton and cockroach ) and coprophilous Fungis were produced utilizing Potato Dextrose Agar and Rabbit Dung Agar. severally. The civilizations were inoculated two to three times in PDA home bases until purified. and so inoculated on trial tubings with the same media. However. the consequences observed under the microscope shows merely the resting spores. hyphae. vegetive hyphae and zoospores. The images taken under HPO below shows the pure isolates of the civilizations.
Fig. 1 Snakeskin civilization under HPO Fig. 2 Pollen grain civilization under HPO
Fig. 3 Cockroach civilization: Pure isolate inside a trial tubing and the civilization under HPO
Fig. 4 Shrimp Exoskeleton civilization under HPO
Fig. 5 Rabbit Dung civilization under HPO
Teasing technique was used to happen chytrids on dirt or H2O samples. Chytrids are zoosporic Fungis classified in the category Chytridiomycetes. Chytridiomycetes are true Fungis that require H2O to scatter throughout their environment. Chytrid zoospores are readily dispersed in the presence of free H2O but the demand for H2O does non curtail them entirely to aquatic home grounds. Besides being normally found in lakes. watercourses. pools. roadside ditches and coastal Marine environments. chytrids besides are present in dirt. [ 1 ] Accompanying organic stuff such as dirt can besides be used to wash the set-up to be used as a beginning of fungous growing.
The type of zoosporic Fungi that will turn on the come-on will depend on the substrate used and what sort of chytrid you are seeking ( keratinolytic Fungi. chitinolytic Fungi. cellulitic Fungi ) . As members of tellurian and aquatic microbic communities. chytrids play an of import ecological function in apparently omnipresent biodegraders of fractious stuffs like ceratin. chitin. and cellulose. [ 1 ] Hence. they play a function in alimentary cycling. Chytrids live saprobically or as parasites in. or on. a figure of different beings and substrates such as in our instance. pollen grains. amphibious tegument and exoskeleton of runt and cockroaches.
The fungous isolates from the pollen grain are saprotrophic Fungis that should degrade polymers. The fungous isolates from the snakeskin are keratinolytic Fungis that should degrade ceratin. However. observations of civilizations shown under the microscope were non successful on turning Fungis. may be because of the undermentioned grounds: 1 ) excessively much H2O flooded the whole snakeskin and pollen grain set-up ; 2 ) the temperature is non ideal for the zoosporic Fungi to turn. However. the fungous isolates from shrimp exoskeleton. cockroaches and coney droppings were successful. Fungal isolates from droppingss are coprophilous Fungis that degrade the waste and other foods in the animate being droppings. On the other manus. fungous isolates from exoskeleton are chitinolytic Fungis. Coprophilous and chitinolytic Fungis were successfully isolated. Their cottony growing were observed on the said substrates.
The writers are thankful to the staffs of Botanical Garden and Microbiology research lab of the University of Santo Tomas for aid in obtaining the stuffs. The writers would besides wish to thank their professors. Sir Thomas Edison dela Cruz and Sir Mike Valdez. for their ageless counsel and forbearance.
[ 1 ] Longcore. J. E. Morphology and Zoospore Ultrastructure of Entophlyctis luteolus sp. nov. ( Chytridiales ) : Deductions for chytrid taxonomy. Mycologia. 87: 25-33. 1995.
JW Deacon & A ; G Saxena ( 1997 ) . Orientated zoospore fond regard and cyst sprouting in Catenaria anguillulae. a facultative parasite of roundworms. Mycological Research 101. 513-522.
Madigan. M. . Martinko. J. . Dunlap. P. & A ; Clark. D. ( 2009 ) . Brock Biology of Microorganisms. Twelfth Ed. Jurong. Singapore: Pearson Education South Asia Pte Ltd. 531-539. Sharma. OP ( 2011 ) . Fungi and Allied Microbes. West Patel Nagar ( New Delhi ) : Tata Mcgraw Hill Education Private Limited.
Isolation and Culture of Plasmodial Slime Molds from Barks of Acasia tree utilizing Moist Chamber Method
Rillera DP. Talibsao K. Talucod AC and Tan P
Department of Biological Sciences. College of Science. University of Santo Tomas. Espana Street. Manila 108