Chemistry Research Journal, 2016, 1(4):132-140

Available online www.chemrj.org

ISSN: 2455-8990
Research Article CODEN(USA): CRJHA5

Microbiological and Chemical Examinations of Water and Fish Obtained From River Nile
of Damietta Governorate, Egypt

Husain A El-Fadaly, Sherif M El-Kadi, Salah E El-Kholy

Agric. Microbiol. Dept., Fac. Agric., Damietta Univ., Damietta, Egypt

Abstract This research was carried out at Microbiol. Lab., Faculty of Agric., Damietta Univ. to evaluate the water
and fish of River Nile. Water and fish samples were collected from the same site of River Nile in Damietta
governorate namely Farskour city during spring, summer, autumn and winter at 2014. All fishes obtained from Rive
Nile were belonging to one genus of fish namely, Oreochromis niloticus (Nile tilapia). Long, weight and color of
examined fish were recorded. Temperature, electrolyte conductivity (EC) and dissolved oxygen (DO) of River Nile
water were determined. Obtained results proved that the highest value of Biochemical Oxygen Demand (BOD 5) was
found during winter being 7.0 mgO2/L and the lowest value was during summer being 4.7 mgO 2/L. Cadmium was
detected in summer, autumn and winter 0.001, 0.003 and 0.007, respectively. In case of spring these minerals were
not detected. The highest value of total bacteria, total fungi, staphylococci, Aeromonas and coliform count in the
water were 6.121, 4.231, 4.342, 3.301 and 5.997 log cfu/ml, respectively. The highest values of bacterial groups of
muscles, intestine and surface of fish were in spring and the lowest values were in autumn. Twenty bacterial isolates
were identified; 2 colonies were considered as Staphylococcus sp., other 2 were considered as Micrococcus sp. Six
colonies were considered as Bacillus sp. Another five colonies were considered as Aeromonas sp. Three fungal
isolates were identified as Aspergillus oryzae.

Keywords Heavy metals, BOD and Microbial populations.

Introduction
River Nile represents more than 90% of the Nile basin’s water resources. Industrial waste waters are considered
among the major sources of environmental pollution, endangering public health through direct use as well as feeding
fish that live in the polluted streams. It is estimated that more than 400 factories continue to discharge more than 2.5
million m3 per day of untreated effluent into Egypt’s waters [1].
Nowadays, the demand of fish food is increasing throughout the world due to the recognition of its nutritional value.
Oreochromis niloticus (Nile tilapia) dominates in fresh water due to its superior performance in this environment.
Tilapias are considered as the best species for culture because of their high tolerance to both adverse environmental
conditions and relatively poor water quality[2].
BOD is the total amount of dissolved oxygen required (milligrams per liter or parts per million, ppm) by
microorganisms for biodegradation of organic matters. It is a common indicator used to measure organic water
pollutants [3]. Fish and other aquatic animals depend on DO (the oxygen present in the water) to live. The amount of
DO in streams is dependent on the water temperature, the quantity of sediment in the stream, the amount of oxygen
taken out of the system by respiring and decaying organisms, and the amount of oxygen put back into the system by
photosynthesis and aeration [4]. The optimum value of good water quality is 4 to 6 mg/l of DO, which ensures
healthy aquatic life in a water body. The seasonal BOD values were slightly higher in summer low during winter
and rainy season. Higher values of BOD in summer season due to higher microbial activity and elevated temperature
[5].

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Bacterial populations of fish skin ranged from 102 to104 (cfu/cm2). Gill tissue has been found to harbour high
bacterial populations, e.g., up to 106 cfu/g. Muscle was considered to be sterile [6]. Bacterial populations in the
digestive tract can be up to 108 cfu/g. For aerobic heterotrophs and 105 cfu/g. For anaerobic bacteria Fish eggs may
be populated by high numbers of bacteria 103–106 cfu/g. Incidentally, the digestive tract of newly hatched larvae
contains scant bacterial populations, but are quickly colonized [7]. Fish of a good quality should have counts of total
bacteria less than 10/g. Faecal coliforms and total coliforms should not exceed 10/g and 100/g, respectively. Total
coliform count in water is not higher than of World Health Organization standard (WHO) (1.0x10 3 cfu/100 ml) [8].
Metals are a major category of globally-distributed pollutants and are natural elements that have been extracted from
earth and harnessed for human industry and products for millennia. Heavy metal is a term used to define metallic
elements with atomic weight higher than 40.0. Metals are notable for their wide environmental dispersion from such
activity; their tendency to accumulate in selected tissues of human body; and their overall potential to be toxic even
at relatively minor levels of exposure. Some metals, such as copper and iron, are essential to life and play an
irreplaceable role in the function of critical enzyme systems. Other metals have no useful role in human physiology
(and most other living organisms) and, even worse, as in the case of lead and mercury may be toxic even at trace
level of exposure. Metals that are essential have the potential to be toxic at very high level of exposure. One
reflection of the importance of metals relative to other potential hazard is their ranking by the United State Agency
for Toxic Substances and Disease Registry (ATDSR), which lists all hazards present in toxic waste site according to
their prevalence and the severity of their toxicity The first, second, third, and sixth hazards on the list are heavy
metals: lead, mercury, arsenic and cadmium, respectively [9].
Therefore, the present study was aimed to determine the physical and chemical state of water and fish obtained from
River Nile, Egypt. Also, the microbiological examination including the identifying of bacterial and fungal isolates
obtained.

Materials and Methods

Physical and Chemical Examination
Electrolyte Conductivity and Temperature
Electrolyte conductivity (EC) and Temperature were determined using a conductivity meter (CM) (Model: CD-
4301, Power: 9 V battery, Lutron Electronic Enterprise Co., LTD Made in Taiwan) [10].
Biological Oxygen Demand (BOD5)
Dissolving oxygen was determined using a dissolved oxygen meter (Model: YK-22DO, Power: 9 V battery, Lutron
Electronic Enterprise Co., LTD Made in Taiwan). The initial dissolving oxygen (initial DO) was determined using a
dissolved oxygen meter directly in the site. Water samples (125 ml) were collected from 20 cm below the water
surface to avoid floating material using brown glass bottles. These samples were firmly covered and placed in an
incubator in the dark for 5 days at 20°C. At the end of this time, the dissolved oxygen level was determined and
considered as final DO. BOD5 was calculated by the method described by Stirling [11]. (BOD205) mg/L = (Initial
DO - Final DO) x dilution factor
Heavy Metals
These analyses were carried out at Central Laboratory of Damietta. To determine Lead (Pb), Cadmium (Cd),
Stannum (St), Arsines (As) and Copper(Co) concentrations, collected water samples were conducted according to
the methods of Gloterman et al. [12] using Perkin – Elmer atomic absorption spectrophotometer (A.A.S 2) with
hydride generation system Perkin – Elmer model PinAAcle 900T, serial No. PTCS12032601.
Microbiological Examinations
Samples Collection and Preparation
Water and fish samples were collected in three replicates from the same site of River Nile in Damietta governorate
namely Farskour city during spring, summer, autumn and winter of 2014. Water samples were collected in 100 ml
sterile glass bottles and then transferred to the microbiological laboratory into the icebox. One ml of water samples
(each is mixed one of the three bottles) or one gram of each fish intestine or fish muscles sample were aseptically
transferred to 9 ml of sterile buffer phosphate PH7. For the microbiological examination of fish surface, 10 ml of
sterile water were aseptically transferred to a plastic bag containing the tested fish and samples were shaken
manually for 2 min, the suspension was collected aseptically in sterilized test tube. The suspension of all samples
were shaken for 10 min using a vortex (VM-300 power: 220 VAC, 50Hz, 0.16A/Made in Taiwan-Associated with
Cannic, in the U.S.A.) to homogenate the obtained solution. Serial dilutions were performed and one ml of each last
three dilutions was used for microbiological examinations [13].

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Total Bacterial Count

For total bacterial count of all samples (water and fish), poured plate method was used After preparing suitable
serial dilutions of water samples, 1 ml was transferred into sterile glass Petri dish in triplicates. Approximate 15 ml
of melted nutrient agar medium at 45-50 °C was poured in each plate, then thoroughly mixed and left for
solidification. The plates were incubated at 37 °C for 72 hours in a digital incubator (Switc, MPM Instruments
S.R.L., Bernareggio/Made in Italy). After the incubation period, developed separated colonies were counted per
each plate of the same dilution and the mean value was calculated [14].
Counting, Isolation and Maintenance of Some Pathogenic Bacteria
One ml of the last three dilutions of all samples (water and fish) were transferred into Petri dishes in three replicates
and approximately 15.0 ml of a specific cultivation medium (Staph. 110 medium, Aeromonas selective agar medium
or S. S. agar medium) was added and left to hardness. Petri dishes were placed upturned in incubator at 37°C for 72
h. The obtained colonies which were produced yellow-orange pigment on Staph. 110 medium was monitored as
Staphylococcus sp. Also, The colonies which were a yellow color on Aeromonas selective agar medium were
considered as Aeromonas sp. The colonies which were black-center colonies or pink to red colonies were monitored
as Salmonella sp or Shigella sp. All typical colonies were isolated on the same specific cultivation for maintenance
and identification [14].
The following microbiological methods were carried out to identify the obtained bacterial isolates according to Holt
et al. [15]. Shape, arrangement of the cells, the Gram reaction, spore stain and acid fast stain were microscopically
examined in stained preparations of 24-48 hrs old bacterial cultures. Presence of spores were recognized in stained
smears using Schaeffer and Fulton’s method after 2 days old cultures [16]. The colonies count per ml or gram of
samples was calculated as follows: The bacterial or fungal count (cfu/ml or cfu/g) = average number of triplicates of
the same dilution x reciprocal of the dilution used [17].
Coliform counts were detected using the most probable number (MPN) technique [14]. Three decimal dilutions for
each sample in three replicated tubes were employed. One ml of each suitable dilution was added to test tubes
containing MacConkey broth medium and Durham tubes, then incubated at 37ºC for 48 hours. The number of
positive tubes showing acid and gas were recorded. The MPN of coliform bacteria per gram of sample was
calculated from standard Table according to Sutton [18].
Total Fungal Count, Isolation, Maintenance and Identification
One ml of suitable serial dilutions of all water or fish samples were inoculated onto three plates using poured plate
method [14]. Approximately fifteen ml of potato dextrose agar (PDA) medium at about 50°C was poured in each
plate, then thoroughly mixed and left for solidification. The plates were incubated at 25°C for 5 days. After the
incubation period, developed separated colonies were counted per each plate and the mean count of 3 plates was
recorded to represent fungal count. Single different developed colonies were isolated on a PDA medium slant for
identification tests. The fungal isolates were subcultured then maintained on PDA slants at 5°C till use [19]. Fungal
isolates were identified by morphological characteristics of colonies in PDA medium. In addition, the vegetative and
reproductive features observed using a light microscope (Olympus CX31 Binocular Halogen Microscope, Made in
Japan) with a magnification power 400x, was also used. The taxonomic keys of Chung and Bennett [20] were used.
Statistical analysis
The differences between means were calculated by Duncan`s Multiple Range Test. Regression coefficient was
analyzed with original data [21].

Results and Discussion

Morphological examination of fish obtained from River Nile during seasons of 2014
All fishes obtained from Rive Nile were belonging to one genus of fish namely, Oreochromis niloticus (Nile tilapia).
Table 1: Morphological examination of Nile tilapia (Oreochromis niloticus) obtained from River Nile during four
seasons of 2014
Seasons Long (cm) Weight (g) Fish color
Spring 7 37 Green with red color in abdomen region
Summer 10 33 Black with yellow and red parts in abdomen region
Autumn 7 22 Gray with white parts in abdomen region
Winter 4 15 Dark gray with yellow and red parts in abdomen region
BOD and EC values of water obtained from River Nile during seasons of 2014
The mean value of long of three individuals were 7, 10, 7 and 4 cm in spring, summer, autumn and winter,
respectively (Table 1) Each value represents the mean of 3 individuals. The highest weight of fishes were obtained
during spring and summer being 37 and 33 gm, respectively, and the lowest weight were in case of autumn and

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winter being 22 and 15 gm, respectively, It was observed that, the color of fishes were differed according to seasons,
where it was green with red color in abdomen region (Fig. 1a) during spring. On the other hand, fish color was black
with yellow and red parts in abdomen region (Fig. 1b) during summer. It was grey with white parts in abdomen
region (Fig. 1c) in the autumn, and dark grey with yellow and red parts in abdomen region (Fig. 1d) in the winter.
Data in Table 2. showing that, temperature varied between 18 and 25°C. Initial DO were 6.6, 6.0, 7.0 and 7.5
mgO2/L in spring, summer, autumn and winter, respectively. Final DO were 5.6, 4.7, 5.9 and 7.0 mgO2/L in spring,
summer, autumn and winter, respectively. The highest value of BOD520 was during winter being 7.0 mgO2/L and the
lowest value was during summer being 4.7 mgO2/L. The highest value of EC was round in winter being 1.95
mhos/cm and the lowest value was in spring being 1.48 mhos/cm. The DO of this study was higher than that of
Surendraraj et al. [22] who detected the DO of feeder canal water and they found that, the values ranged between
0.89-3.53 ppm. BOD and salinity were lower which were ranged between 4.83-13.6 ppm and 0.07-0.23 mhos/cm.
a b

U U
s s
i d
i
c
n n
gU g
U
s s
B iB Nile during
i Figure 1: Oreochromis niloticus obtained from River
a) spring, b) summer, c) autumn and d) winter
rn and EC values of water obtained from Damietta governorate
Table 2: BOD nr during four seasons of 2014
20
Seasons o Temperature DO, mgO2/L o BOD EC
(2014) g (°C) Initial Final g 5
(mgO2/L) (mhos/cm)
Spring w 20 6.6 5.6 w 2.6 1.48
Summer 25 6.0 4.7 3.1 1.58
Autumn n 22 7.0 5.9 Bn 2.0 1.60
Winter
B 18 7.5 7.0 1.5 1.95
r
Heavy metals values of water and fish muscles during seasons of 2014. r
Lead and cupper were not detected in all seasons samples except in spring being 0.006 and 0.106 ppm, respectively.
bo ob
On the other hand, Cadmium was detected in all seasons to be 0.001, 0.003 and 0.007 during summer, autumn and
winter, respectively, while in spring it was not detected. Stannum and arsines did not present in all seasons. Also,
ow wo
heavy metals were determined in all fish muscles samples obtained from River Nile. Cadmium, Cupper, Arsines and
tn nt
Stannum did not found in all seasons. The values of lead were 0.088 and 0.040 ppm in spring and summer
,respectively, while they not detected in autumn and winter. These results are lower than the permissible levels (1
t t
mg/L) permitted by the Egyptian Organization for Standardization [23]. Also, Pb concentration did not exceed the
Egyptian Standards of the Environmental Laws No. 48/1982 [24]. which the maximum Pb concentration in water
was 0.05 mg/L. eb be
The Results of this study are lower than those obtained by Ali et al. [25] who tested heavy metals in Nile Tilapia
lo ol
(Oreochromis niloticus) from two sites of River Nile at Aswan (water and fish) and they found that, the
concentrations of heavy metals (Cu, Ni, Pb and Cd) of water in the first site were 0.26, 1.81, 2.63 and 1.04 ppm,
.t t.
respectively. In the site II the values were 0.41, 1.9, 2.59 and 0.13 ppm, respectively. On the other hand, the
concentrations of heavy metals of fish muscles in site I and II were 1.85, 5.2, 7.7, 1.0 and 3.15, 6.4, 8.0, 0.35 ppm,
.
respectively. t . t
Ali et al. [25] also explain that, pb can find its way to the water of the River Nile through the leaching of gasoline
e e
from the fishery boats and the tour ships travels. Moreover the increasing of heavy metals concentrations at River
Nile can be attributed to the huge quantities of sewage and industrial wastes via drains.
l l
Microbiological examination of water taken from River Nile (Damietta governorate) during seasons of 2014
. .
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Results in Table 3 showing the highest value of total bacteria count was during spring being 6.121 log cfu/ml, while
the lowest value was in winter being 2.477 log cfu/ml. The high bacterial load may be enplaned by the observation
of Ali et al. [1] who reported that, the richness of the effluent in organic carbon exerted a specific enrichment effect
on the microbial population. Table 3 also showed that, the highest value of total fungal count was in summer and
spring being 4.231 and 4.041 log cfu/ml, respectively. the total fungal count was in the lowest value in the autumn
and winter being 1.0845 and 1.845 log cfu/ml, respectively. It was observed that, there were no bacterial growth on
SS agar medium. The highest value of Staphylococci count was in spring being 4.342 log cfu/ml, while it was in the
lowest value in autumn being 1 log cfu/ml. The highest value of Aeromonas count in summer being 3.301 log
cfu/ml, but the lowest values were during spring, autumn and winter to be 2.797, 2.602 and 2.544 log cfu/ml,
respectively. The highest values of coliform count were in spring and the summer being 5.997 and 4.964 log cfu/ml,
respectively, while coliform count was in the lowest value in the autumn and winter being 0.477 and 0.301 log
cfu/ml, respectively.
These results are in good agreement with those of Osman [26] who showed that, the microbiological quality of the
River Nile was carried out from three different sites i.e. Helwan, El-Giza and Shoubra. The highest average log
number of total coliform, faecal coliform and faecal streptococci were 4.38, 3.29 and 2.58, respectively. Those
values are lower than that obtained by Ali et al. [25]. who reported that, the total bacterial count of the samples
ranged from 2.3 x104-2.19 x 105 cfu/g. Also, these results did not similar to that obtained by Ali et al. [25] who
reported that, the Salmonella and Shigella counts ranged from 2 and 57 cfu/g. This study took the same trend of Ali
et al. [1] who found that, the total microbial load of various bacterial groups being highest in autumn and lowest in
winter. They also found that, water load of bacterial indicators of pollution, in particular faecal coliforms, was
higher in spring compared to other seasons. Moreover El-Kadi and El-Morsy [27] studied the microbiological
populations of water of Nile tilapia and they found that, the maximum value of total bacterial count and total yeast
and fungal in the water of fish were 2.88×10 4 and 7.3 ×102 CFU/ml, respectively.
Table 3: Microbiological values (Log cfu/ml) of water taken from River Nile (Damietta governorate) during seasons
of 2014
Seasons °C Count of
(2014) Total bacteria Total fungi Staphylococci Aeromonas Coliform
Spring 23 6.121 4.041 4.342 2.797 5.997
Summer 28 4.114 4.231 3.000 3.301 4.964
Autumn 20 3.301 1.845 1.000 2.602 0.477
Winter 14 2.477 1.840 1.041 2.544 0.301
Microbiological examination of fish obtained from River Nile (Damietta governorate) during seasons of 2014.
The total bacterial count of fish muscles were 3.403, 5.602 , 0.954 and 2.000 log cfu/ml in spring, summer, autumn
and winter (Table 4) ,respectively. On the other hand, the highest value of total bacterial count of fish intestine was
8.598 log cfu/ml in spring, but the lowest value was in the autumn being 1.398 log cfu/ml. Total bacterial count of
fish surface were 6.297, 6.447, 3.114 and 3.114 log cfu/ml in spring, summer, autumn and winter, respectively. The
highest values of fungal count in muscles, intestine and fish surface were during autumn, summer and spring being
1.477, 7.903 and 5.121 log cfu/g, respectively. On the other hand, all lowest fungal counts of muscles, intestine and
surface were during winter being 0.699, 5.301 and 1.845, respectively.
Table 4: Microbiological examination (Log cfu/gm) of fish obtained from River Nile (Damietta governorate) during
seasons of 2014
Seasons Count of
(2014) Fish part Total bacteria Total fungi Staphylococci Aeromonas Coliform
Muscles 3.403 1.041 2.083 2.422 4.218
Spring Intestine 8.598 7.899 6.246 7.172 9.083
Surface 6.297 5.121 4.917 3.513 5.083
Muscles 5.602 0.954 0.778 0.778 1.230
Summer Intestine 7.447 7.903 1.301 5.301 4.477
Surface 6.447 5.086 1.114 0.602 1.041
Muscles 0.954 1.477 0.945 0.477 0.301
Autumn Intestine 1.398 6.114 1.477 2.301 1.322
Surface 3.114 2.477 1.176 2.301 0.699
Muscles 2.000 0.699 0.845 2.114 0.903
Winter Intestine 3.000 5.301 2.301 3.398 1.361
Surface 3.114 1.845 2.301 4.778 4.633

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It was observed that, there were no bacterial growth on SS agar medium. The highest values of Staphylococcal
counts of fish muscles, intestine and fish surface were in spring being 2.083, 6.246 and 4.917 log cfu/g, but the
lowest values were in summer being 0.778, 1.301 and 1.114 log cfu/g, respectively.
The highest count of Aeromonas was in fish muscles during spring being 2.422 log cfu/g and the lowest value was
during autumn being 0.477 log cfu/g. The highest count was in fish intestine during spring being 7.172 log cfu/gm
and the lowest value (2.301 log cfu/gm) was during autumn. Aeromonas count on fish surface ranged between 4.778
and 0.602 during winter and summer, respectively. Coliform was found in the highest values in all fish parts in
spring being 4.218, 9.083 and 5.083 log cfu/g in muscles, intestine and fish surface, on the other hand, coliform
count was in the lowest values in the autumn.

These results are higher than those obtained by Surendraraj et al. [22]. who reported that, the total bacterial
counts of muscles of carp farm fish ranged between log 4.19 – 4.85 cfu/g. Also, they found that, the total coliform of
farmed fish ranged between log 2.0 – 3.4 cfu/ml. Mohammed and Hamid [28] studied the bacterial load of fresh fish
(Oreochromis niloticus and Clarias lazera), at the period from November 2009 to March 2010 in the Sudan
University of Science and Technology, Department of Fisheries and Wildlife Science. The results were 8.4×10 5 and
1.7×105 cfu/g for fresh Tilapia and fresh catfish, respectively. These results are higher than obtained results. Jimoh
et al. [29] studied the microbial flora of the gastro-intestinal tract of Clarias gariepinus caught from river Dandaru
Ibadan, Nigeria. They found that, the total bacterial count, total fungal count and total coliform count were 6.5 x 10 5,
3.0 x 103 and 1.9 x104 cfu/g., respectively. In the same trend, El-Kadi and El-Morsy [27] reported that, the
maximum value of total bacterial count and yeast and fungal count in fish intestine were 1.96×106 and 12.2 ×102
(CFU) /g, respectively.

The relationship between the log of total bacterial count and the biological oxygen demand
The correlation coefficient between the log of total bacterial count (LTBC) in the water of River Nile and the BOD
was calculated (Fig. 2). Statistical analysis indicated a low correlation (r = 0.552) between the log of total bacterial
count and BOD. Kagalou et al. [30] reported that, a positive correlation between BOD and total bacterial count in a
river ecosystem highly polluted by industrial pollutants. In contrast, other author [1] found a negative correlation
between both total and faecal coliforms with BOD.

8 4
Log Total Bacterial Count

BOD5 mg O2/L water

6 3

4 2

2 1
Spring Summer Autumn Winter

LTBC BOD5

Figure 2: The relationship between the log of total bacterial count and the biological oxygen demand

Characterization and Identification of Bacterial Isolates

Table 5 showing bacterial isolates numbers, characterization and its sources. Seven different bacterial isolates were
isolated from nutrient agar medium, 9 isolates were isolated on Aeromonas agar medium and 4 isolates were found
on Staph 110. medium. Eleven isolates were isolated from water, 6 from muscles, 2 from intestine and 4 from
surface of fish. Among 20 bacterial isolates, only 4 were coccoid shape, 10 isolates were short rods and 6 isolates
were long rods. Ten isolates were Gram negative and 10 isolates were Gram positive. Six isolates were spore
formers and 14 isolates were non spore formers. All isolates gave negative results in acid fast stain.

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Table 5: Bacterial isolates numbers, characterization and sources obtained from fish and water of River Nile
(Damietta governorate) during seasons of 2014
Sources Cultivation Isolates Characterization of isolates
(2014) media Nos. Shape Arrangement Gram stain Spore stain Acid fast stain
9 Short rods Single - - -
11 Short rods Single - - -
Nutrient Agar 22 Long rods Pair + + -
23 Long rods Chain + + -
39 Short rods Single - - -
Water 6 Long rods Single + + -
42 Long rods Single + + -
Aeromonas
77 Short rods Single - - -
78 Short rods Single - - -
33 Coccoid Staph + - -
Staph 110.
37 Coccoid Clusters + - -
40 Short rods Single - - -
Nutrient Agar
59 Short rods Single - - -
Fish
16 Long rods Single + + -
Muscles Aeromonas
76 Short rods Single - - -
Staph 110. 15 Coccoid Clusters + - -
Fish Single
Aeromonas 75 Short rods - - -
Intestine
41 Long rods Single + + -
Fish Aeromonas
74 Short rods Single - - -
Surface
Staph 110. 21 Coccoid Pair + - -
Two typical colonies representing Staphylococcal growth, orange color on Staph. 110 medium, colonies were picked
up and streaked onto slant of the same medium. After growth, the morphological characteristics under light
microscope were done. The cells were spherical, Gram positive, arranged in irregular clusters. Isolate No. 15 and 37
were considered as Staphylococcus sp. according to Bergey’s Manual of Determinative Bacteriology [15]. Isolates
Nos. 21 and 33 were considered as Micrococcus sp. This results are in harmony with those published by Ali [1] who
isolated and identified of Staphylococcus sp from fish of fresh water in Mosul city. Five species of Staphylococcus
sp, S. saprophyticus, S. epidermidis, S. hyicus, S. aureus, and S. intermedius were identified. The percentages of
Staphylococcus isolates from skins was 35.5%, 17.7% in muscles, 25.8% in livers and 21% in intestines.
Six colonies which were white, yellow or orange color, were picked up and streaked onto nutrient agar slant. After
growth, the morphological characteristics under light microscope were done. The cells were long rods, Gram
positive, spore formers and non acid fast. Its arrangement were single, pair or chains. Isolates Nos. 6, 16, 22, 23, 41
and 42 were considered as Bacillus sp. according to Bergey’s Manual of Determinative Bacteriology [15]. Jimoh et
al. [29] isolated Bacillus alvei and Bacillus megaterium from the microbial flora of the gastro-intestinal tract of
Clarias gariepinus caught from river Dandaru Ibadan, Nigeria.
Five colonies isolated on Aeromonas agar medium that gave yellow color were picked up and streaked onto the
same medium slant. After growth, the morphological characteristics under light microscope were done. The cells
were short rods, Gram negative, non spore formers and non acid fast. Its arrangement was single. Isolates Nos. 74,
75, 76, 77 and 78 were considered as Aeromonas sp. according to Bergey’s Manual of Determinative Bacteriology
[15]. Similar results were obtained by Rokibul et al [31] and Jimoh et al. [29] who isolated Aeromonas sp.
Colonies isolated on nutrient agar medium which were white or yellow color were picked up and streaked onto
nutrient agar slant. After growth, the morphological characteristics under light microscope showed that. The cells of
isolates Nos. were 9, 11, 39, 40 and 59. were short rods, Gram negative, non spore formers and non acid fast. Its
arrangement was single.
Characterization of Fungal Isolates
Three fungal isolates were isolated from all samples. Characterization of the isolates showed that, colonies yellow
green on PDA medium (Fig. 3), conidiophores colorless, long, coarsely roughened. conidial heads typically radiate,

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conidia globose to subglobose. From these characteristics, isolates Nos. 38, 60 and 61 were identified as Aspergillus
oryzae [20]. Jimoh et al. [29] isolated Aspergillus niger, Aspergillus flavus, from the gastro-intestinal tract of
Clarias gariepinus. In addition, El-Kadi and El-Morsy [27] isolated five fungal isolates and identified it as
Aspergillus ochraceus, A. oryzae, A. niger, Geotrichum candidum and Penicillium sp.

Figure 3: Yellow green fungal colony on PDA medium

Conclusion
Obtained results proved that the highest value of BOD was during winter and the lowest value was during summer.
Heavy metals are lower than the permissible levels permitted by the Egyptian Organization for Standardization.
Also, Pb concentration did not exceed about the Egyptian Standards of the Environmental Laws No. 48/1982. The
highest value of total bacteria, total fungi, staphylococci, Aeromonas and coliform count in the water. The highest
values of bacterial groups of muscles, intestine and surface of fish were in spring and the lowest values were in
autumn. Staphylococcus sp., Micrococcus sp., Bacillus sp., Aeromonas sp. and Aspergillus oryzae were isolated and
identified from water and fish.

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