International Journal of Advance Study and Research Work (2581-5997)/ Volume 2/Issue 7/July 2019

Bacterial, Fungal, and Protozoal Microflora of Hay

Ibadete Ismajli1, Osman Fetoshi2*, Albona Shala Abazi3, Pajtim Bytyçi4, Alban Hyseni5, Qendrim Ramshaj6
1
University Clinical Center of Kosovo, Prishtina 10000, Kosovo
2
Faculty of Tourism and Environment, University of Applied Sciences, Ferizaj 70000, Kosovo
3
Faculty of Management in Tourism, Hotels and the Environment, University “Haxhi Zeka”, Pejë 30000, Kosovo
4,5,6
Faculty of Mathematics and Natural Sciences, University of Prishtina ”Hasan Prishtina”, Prishtina 10000,
Kosovo
*E-mail ID: osmanfetoshi@hotmail.com

DOI: 10.5281/zenodo.3355410

Abstract

Herbivores feed on plant structural carbohydrates such as lignin, cellulose, and hemicellulose, which they are not able to
utilize themselves because of their inability to produce respective hydrolytic enzymes. Therefore, symbiotic microorganisms
are established in their alimentary tracts that can hydrolyze these compounds to generate energy for themselves, as well as
for the host animal. This study aimed to the qualitative determination of bacterial, fungal and protozoal microflora in hay
(dry grass) harvested from the lowland meadow in Bradash village, municipality of Podujeva. Based on the results, the
number of microflora represented differs in the first, second, and third test. The number of microflora, bacteria, yeasts, and
molds has been low in the first test, respectively high in the third test.

Keywords: Bacteria, Fungi, Hay, Microbial interactions, Microflora.

Introduction
Bacteria comprise a diverse group of microorganisms that display a wide range of physiologies and live in a variety of habitats.
Their diversity allows them to participate in almost every biological activity on Earth [1]. Higher organisms could not exist for
long without their native bacteria, that is, the bacteria that normally reside in or on the body. The planet also relies almost
entirely on bacteria to cycle nutrients from sediments through animal and plant life, then through the atmosphere and back to
the earth. Bacterial numbers reach enormous levels in many habitats on Earth. More bacterial cells live on or in the human,
plant and animals cells. Bacteria play roles in nature, inhuman and animal health, in association with plants and insects.
Bacteria and fungi in soil and water both degrade organic wastes and recycle the elements carbon, nitrogen, phosphorus, and
sulfur, among others in the earth’s biogeochemical cycles.
In the process of degrading the earth’s organic matter, some bacteria produce the gases methane, carbon dioxide, hydrogen,
oxygen, and nitrous oxide, which convert to nitrogen gas. Bacteria, therefore, are major contributors to the composition of the
atmosphere, and they carry out these reactions even when they are inside plants or animals. Many live in commensal
relationships with higher organisms and affect the health of their hosts, and mutual organisms [3]. Some of these microbes
interact with each other in a synergistic relationship to extract energy while producing highly active lignocellulolytic enzymes
supporting digestion of the host [2] [4] [14]. In such a relationship, the bacteria help their hosts by digesting nutrients and secreting
compounds made during their normal metabolism, which the host then absorbs and uses in its own metabolism. The most
important of these secretions are acids, alcohols, antibiotics, proteins, and amino acids, enzymes, and vitamins. This research
presents the determination of bacterial, fungal and protozoal microflora in the hay, and how this microflora helps herbivores
indigestion.

Material and Methods

As a research material was served the samples of hay that were taken in the lowland meadow in the village of Bradash,
Municipality of Podujeva, during the winter season, in January, February and March (Test-1, Test-2, Test-3). The samples taken
were placed in a jar of water that stayed 7 (seven) days at room temperature 18-22C°, then their examination was done.
At the Laboratory of Microbiology at Faculty of Geosciences and Technology in Mitrovica where microbiological analyses
have been conducted. Microbiological analysis and examination of hay were carried out in order to determine the total number
of bacteria: heterotrophic bacteria, yeast, and molds as relevant microbiological parameters.

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Plate Count Agar used for cultivation and enumeration of microorganisms from potable water sources, swimming pools, and
other water specimens by the membrane filter method and heterotrophic plate count technique. Wort Agar used for cultivation
and enumeration yeasts and molds [5][6].
At the point of the sample observed in the light microscope, various types of bacteria were found, as well as the presence of
some, paramecium, rotifers, and parasitic worms. The native preparation is prepared from a sample that it was taken from the
bottom glass sample and then it is observed to detect density and mobility of microorganisms. To perform the microbiological
examination, are used indirect methods such as the agar method according to Choh and membrane filter methods were applied.
Initially, the samples were diluted with the dilution series of 10 1-106. Determining the number of cells is done by finding the
number of colonies. It starts from one cell give one colony. Determination of the cells is done according to the formula [7][15].

Where:
No of cells/100 - total number in 100 mL.
CFU - colony-forming unit.
SD - serial of dilution.
100 - Volume (mass) in which the number of microorganisms is required.
VS - The volume of the sample.

Results and Discussions

The results obtained from the research are presented in (Figure: 1, 2, 3, 4). Serving on contemporary research methods (such as
membrane filters), the results have shown the large density of micro flora.

45000000
40000000
35000000
30000000
25000000
20000000
15000000
10000000
5000000
0
Serial Colony No of cells
dulition forming unit /100 ml
Heterotrophic bacteria 10000 43 430,00,000
Yeasts and Molds 100000 9 9,00,000

Fig 1: Results of microbiological parameters, January 2014 (first test)

Thus, the number of Heterotrophic bacteria in 100 mL of the sample during the first test was for a total of 43 million cells. It is
worth pointing out that the bacteria found to originate from the dry meadow grass (hay) that survived during the dry process.
Here we can talk about bacteria that have passed into the dormant and sporogenic phase. Here, we must not ignore the presence
of heterotrophic mesophile bacteria from the water supply with which is softened hay.
Hay dedicated for animal feed also had a very rich fungal micro flora. The results show for 900,000 (nine hundred thousand)
cells for 100 mL of the sample. From these results, it can be concluded heterotrophic bacteria in the digestive tract of herb
dominating than yeast and molds that are in a low number.

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 Fig 2: Colonies of yeast and molds (left) and Heterotrophic bacteria (right)

Yeast and molds, after cultivation and incubation period, in-room temperature, 18-22 ºC, have given different size, color, and
consistency. Heterotrophic bacteria, after cultivation and incubation period, in-room temperature 18-22 ºC, have given different
size, color, and consistency.

Fig.3: Paramecium (left) and parasitic worms (right)

As we can see from figures bacterial and fungal microflora, there are also present of protozoa and some individuals of metazo -
parasitic worms (Figure 3).

50000000
45000000
40000000
35000000
30000000
25000000
20000000
15000000
10000000
5000000
0
Serial Colony No of cells
dulition forming unit /100 ml
Heterotrophic bacteria 100000 47 470,00,000
Yeasts and Molds 10000 11 11,00,000

Fig 4: Results of microbiological parameters, February 2014 (second test)

While in the first test, 43 million (forty-three million) cells were recorded from heterotrophic bacteria while in the second test
the number of heterotrophic bacteria showed an increase of 4 (four) million cells. On the other hand, yeasts and molds were
increased by 200 (two hundred) thousand cells. These results indicate the creation of favorable conditions at the time of
sampling for the second test. Bacterial and fungal microflora of from meadow has increased a result of humidity and feed in the
field.

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 50000000
45000000
40000000
35000000
30000000
25000000
20000000
15000000
10000000
5000000
0
Serial Colony No of cells
dulition forming unit /100 ml
Heterotrophic bacteria 100000 50 500,00,000
Yeasts and Molds 10000 14 14,00,000

Fig 5: Results of microbiological parameters, Marc 2014 (third test)

If we compare the results obtained from the first and second test with the third test, it is noted that heterotrophic bacteria and
yeasts –molds have continued to increase in the maximum number of heterotrophic bacteria was 50 (fifty) million cells per 100
mL of the sample, respectively the largest of the tested fungi tests, was 1.400.000 (one million four hundred thousand) per unit
of measurement. It should be noted here that the colonization of heterotrophic bacteria, yeasts, and molds in hay has been from
the soil, [12] the surface water, plants, and pollution. So it is evident that animals get the largest microflora from hay but they also
get a large amount of microflora from the water that they also take in large quantities. There should also be no exclusion of
livestock feed in the form of grains of grain which will also have a more densely considered microflora. More than 60 % of
starch is also digested by the rumen microbes, depending on the amount fed and its rate of passage through the rumen. Almost
all the sugars are digested in the rumen [8] [9] [10] [11].

Conclusions
Based on the results of presented in this research we can conclude as follows:
The heterotrophic bacterial microflora and the fungal micro flora (yeasts and molds) form hay are very common in nature. The
site of spreading and colonization microflora is water, soil, plants, animals (on the surface and inside). Microorganisms of
grams that represent most of the livestock grass respectively are heterotrophic inhabitants. There are also protozoa, worms.
Ruminants enter into symbiosis with bacterial, fungal, protozoal microflora mutually beneficial. Microflora possesses cellulose
enzymes that successfully digest cellulose matter, while from ruminant gain source energy, vitamins, minerals, temperature.

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