Journal of Applied Sciences Research, 2(11): 1037-1041, 2006

© 2006, INSInet Publication

A Studdy on Mesophilic Aeromonas in Milk and Some Milk Products in Port Said City

Nahla, T. korashy

Department of Microbiology, Port-Said Lab., Animal Health Research Institute, Dokki, Giza, Egypt.

Abstract: One hundred twenty samples (30 each of raw milk, pasteurized milk, yoghurt and kareish cheese)
were collected from super markets, street peddlers and dairy shops in Port-Said City. The samples were
examined for enumeration and isolation of mesophilic Aeromonas species. The mesophilic Aeromonas species
could be isolated from 80% and 86.7% of the examined raw milk samples using Starch ampicillin agar (SAA)
and MacConky Mannitol ampicillin agar (MMA) with mean value of count 8.6x106 and 9.2x106 CFU/ml
respectively. The contamination rate of pasteurized milk samples were 16.7% and 16.7% using (SAA) and
(MMA) with mean value of count 4.7x104 and 5.2x104 CFU/ml respectively. In case of yoghurt and kareish
cheese samples, the percentage of positive samples were 73.3% and 66.7% using (SAA) with mean value of
count 3.9x106 and 6.3x105 CFU/ml respectively, while using (MMA) the percentage of positive samples were
76.7% and 70% with mean value of count 5.1x106 and 8.3x105 CFU/ml respectively. The total number of
mesophilic Aeromonas isolates from 120 examined samples was 103 isolates of which 71 (69%) isolates were
A. hydrophila, 21 (20.4%) isolates were A. caviae and 11 (10.6%) isolates were A. sorbia. The highest number
of isolates 39(37.9%) were recovered from raw milk samples while the lowest number of isolates 10(9.7%)
were recovered from pasteurized milk samples.

Key words: Aeromonas, Milk and Some milk products.

INTRODUCTION urinary tract infections, hepatobiliary, endocarditis and

Genus Aeromonus contains two well separated
groups the first group consists of a single psychrotrophic,
non motile species, Aeromonus salmoncida, which not a
pathogenic of man Popoff[1] while the second group
consists of mesophilic motile strains including A.
hydrophila, A. Sorbia and A. caviae and described by
popoff et al.,[2]. Motile aeromonads are wide spread in the
environment and cause food related illness and become of
concern to food microbiologist as it has been isolated
from variety of food including raw milk Palumbo et al.,[3]
and Greenway[4]. The pathogencity of the organism has
been correlated to its ability to secrete several extra
cellular virulent products including enterotoxin,
cytotoxins, haemolysin, lipase and proteases Trust and
Chipman[5] and Ljungh and Wadstrom[6]. In humans,
motile Aeromonas organisms are recognized as the cause
of gastroenteritis Wadstrom and Ljungh[7] and could
disseminating infections in the immunocompromised
patients Ellison and Mostow[8]. Gastrointestinal
infections due to Aeromonas are of two types, a
cholera-like and dysentery-like illness. Moreover it
can produce a wide rang of extra intestinal infections as
septicemia and meningitis Davis et al.,[9] and Ellison
and Mostow[8] cellulites, wound infections and other
unusual manifestations include eye infections, pneumonia,
ear infection Koneman et al.,[10]. Aeromonas were
acquired through the consumption of contaminated
food and water Chopra and Houston[11]. The organism has
been associated with spoilage of refrigerated foods of
animal origin other than raw milk Saad et al.,[12]. So the
aim of this study was to determine the prevalence of
mesophilic Aeromonas species in raw milk, pasteurized
milk, yoghurt and kareish cheese and its role as a public
health hazard.
MATERIALS AND METHODS
1-Samples: A total of 120 samples of raw milk and milk
products (30 each of raw milk, pasteurized milk, yoghurt
and kareish cheese) were collected form super markets,
street peddlers and dairy shops in Port-Said city. All
samples were collected under aseptic precautions and
transferred to the laboratory as soon as possible to
examined bacteriologically for presence of mesophilic
Aeromonas species.
2-Bacteriological examination:
a-Preparation of samples: milk samples were tested for
heat treatment and shaken thoroughly while kareish
cheese and yoghurt were grind well in sterile mortar
Richardson[13].

Corresponding Author: Nahla, T. korashy, Department of Microbiology, Port-Said Lab., Animal Health Research Institute,
Dokki, Giza, Egypt.

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 J. Appl. Sci. Res., 2(11): 1037-1041, 2006
b-Enrichment of the samples: Ten ml of raw and
pasteurized milk and 10 gram of yoghurt and kareish
cheese were homogenized in 90 ml Trypticase soya broth
containing 10 µg ampicillin/ml and blended for 2 minutes,
then incubated at 28EC for 24 hours Villari et al.,[14].
c-Enumeration of Aeromonas species: The enrichment
cultures were serially diluted up to 10-6, then 0.1 ml from
each dilutions was spread over starch ampicillin agar
(SAA) and MacConkey Mannitol ampicillin agar (MMA)
with sterile bent glass rod and incubated at 28EC for
24 hours then the numbers of isolated Aeromonas were
calculated and typical colonies were picked into triple
sugar iron (TSI) and nutrient agar slants for biochemical
identification EL-prince[15].
d-Identification of Aeromonas species: Presumptive
aeromonad were identified by biochemical tests
according to Koneman et al.,[10].
Statistical analysis: Minimum, maximum, mean,
standard deviation and standard error of mean were used
to describe data. T-test was used to evaluate the efficacy
of both media used for Aeromonas isolation and counting.
P value was considered significant if less than 0.05 and
0.01 at 95% and 99% respectively. These tests were
analyzed using the Statistical Package for Social
Scientists (SPSS) for windows 10.0 (SPSS Inc., Chicago,
IL and USA).
RESULTS AND DISCUCSIONS
In the recent years, Aeromonas has received
increasing attention as an agent of food borne diarrheal
diseases in healthy people Gracey et al.,[16] and
Palumbo et al.,[3]. The ability of aeromonads to grow at
low temperature is of great concern, particularly in foods,
which are stored under refrigeration. This explains that
Aeromonas could a potential food born pathogen and
dairy products may represent an important vehicle of its
transmission Freitas et al.,[17].
Table 1: Efficacy of solid specific media for isolation of mesophilic Aeromonas from different examined samples.
Types of samples Number of samples
Raw milk 30
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Pasteurized milk 30
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Yoghurt 30
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Kareish cheese 30
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Total 120
Table (1) revealed that the contamination rate of raw
milk by mesophilic Aeromonas species using enrichment
technique on (SAA) and (MMA) were (80%) and (86.7%)
respectively. While the count rang were ranged from
0.00 up to 2.5 x107 with the mean count value 8.6x106
CFU/ml and 0.00 up to 2.9x107 with the mean count
value 9.2 x106 on (SAA) and (MMA) respectively, as
shown in Table 2. Similar finding was reported by Hafez
and Halawa[18] and Khalil[19]. Low incidence of
Aeromonas in raw milk had reported by Enany et al.,[20]
and Amany Shalaby[21]. Lower finding 2.44 %, 9.7 % and
14.9 % were recorded by Banerjee and Block[22],
Ergullu[23] and Shweizer et al.,[24] respectively. The higher
incidence of Aeromonas in raw milk could be attributed
to its wide distribution in the nature; Aeromonas
microorganism is commonly present in farms, in feeding
stuff, water, faeces and soil El-Shenawy and Marth[25].
The organisms can invade the udder tissues; multiply in
mammary tissues and subsequently discharge in milk.
Also the contaminated water used for washing milking
equipments is considered as a significant source of
contamination. So presence of mesophilic Aeromonas in
a high level in raw milk samples is indicative to bad
hygienic measures of milk production and distribution.
Concerning the contamination rate of pasteurized
milk by mesophilic Aeromonas species using (SAA) and
(MMA) were 16.7 % and 16.7 % respectively as in
Table 1. While the count rang from 0.00 up to 8.5 x105
with the mean value 4.7x104 CFU/ml and 0.00 up to 9x105
with the mean value 5.2x104 on (SAA) and (MMA)
respectively as in Table 3, Zienab and Amany[26] found
that 22% of the examined pasteurized milk samples
positive for A. hydrophila. Lower incidence recorded
by Kirov et al.[27]. The process of pasteurization was
done at 71-75EC for 15 second. Aeromonas organisms are
sensitive to temperature above 48EC Palumbo et al.,[28]
and the strain of Aeromonas should have been killed
during this process. So presence of mesophilic Aeromonas
species in the pasteurized milk may be due to inefficient
pasteurization or post pasteurization contamination during
packaging process of pasteurized milk. The contamination
of the pasteurized milk may be introduced by subsequent
handling of the milk.
Positive samples
--------------------------------------------------------------------------------------------------------------------
Starch ampicillin agar MacConkey Mannitol ampicillin agar
----------------------------------- ------------------------------------------------
No % No %
24 80 26 86.7
5 16.7 5 16.7
22 73.3 23 76.7
20 66.7 21 70
71 59.1 75 62.5
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 J. Appl. Sci. Res., 2(11): 1037-1041, 2006

Table 2: Statistical analytical results of total count of mesophilic Aeromonas species CFU/ml of raw milk samples on solid media.
Static’s
----------------------------------------------------------------------------
Types of samples Media used No. of samples Min Max Mean SE SD
Raw milk Starch ampicillin agar 30 0 2.5x107 8.6x106 1.7x106 9.1x106
----------------------------------------------------------------------------------------------------------------------------------------------------------
MacConkey Mannitol ampicillin agar 30 0 2.9x107 9.2x106 1.7x106 9.4x106
Min=Minimum, Max=Maximum, SE=Standard Error, SD=Standard Deviation

Table 3: Statistical analytical results of total count of mesophilic Aeromonas species CFU/ml of Pasteurized milk samples on solid media.
Static’s
-----------------------------------------------------------------------
Types of samples Media used No. of samples Min Max Mean SE SD
Pasteurized milk Starch ampicillin agar 30 0.00 8.5x105 4.7x104 3.2x104 1.8x105
----------------------------------------------------------------------------------------------------------------------------------------------------------
MacConkey Mannitol ampicillin agar 30 0.00 9x105 5.2x104 3.3x104 1.8x105

Table 4: Statistical analytical results of total count of mesophilic Aeromonas species CFU/g of Yoghurt samples on solid media.
Static’s
-----------------------------------------------------------------------
Types of samples Media used No. of samples Min Max Mean SE SD
Yoghurt Starch ampicillin agar 30 0.00 1.2x107 3.9x106 7.8x105 4.3x106
----------------------------------------------------------------------------------------------------------------------------------------------------------
MacConkey Mannitol ampicillin agar 30 0.00 4x107 5.1x106 1.4x106 7.7x106

Table 5: Statistical analytical results of total count of mesophilic Aeromonas species CFU/g of Kareish cheese samples on solid media.
Static’s
-----------------------------------------------------------------------
Types of samples Media used No. of samples Min Max Mean SE SD
Kareish cheese Starch ampicillin agar 30 0.00 3x106 6.3x105 2.1x105 1.2x106
----------------------------------------------------------------------------------------------------------------------------------------------------------
MacConkey Mannitol ampicillin agar 30 0.00 5x106 8.3x105 2.5x105 1.4x106

Table 6: Correlation between the total count of mesophilic Aeromonas species in both solid media in the different examined samples at p. 0.05 (95%)
and 0.01 (99%).
Starch ampicillin agar
----------------------------------------------------------------------------------------------------------------
Raw milk Pasteurized milk Yoghurt Kareish cheese
MacConkey Mannitol Raw milk 0.021 (*) ---- ---- ----
ampicillin agar --------------------------------------------------------------------------------------------------------------------------------------------------
Pasteurized milk ---- 0.032 (*) ---- ----
--------------------------------------------------------------------------------------------------------------------------------------------------
Yoghurt ---- ---- 0.084 ----
--------------------------------------------------------------------------------------------------------------------------------------------------
Kareish cheese ---- ---- ---- 0.087
(**) = Highly significant result at 0.01 (99%).
(*) = Significant result at 0.05 (95%).

Table 1 and 4 revealed that 73.3 % and 76.7% of the
examined yoghurt samples contained mesophilic
Aeromonas species with mean counts of 3.9x106 and
5.1 x106 CFU/g using (SAA) and (MMA) respectively.
Lower incidences were reported by Sami[29] who found
that the rates of recovery of mesophilic Aeromonas
species recovered from yoghurt were 12% on (SAA) and
16 % on ampicillin dextrin agar medium. While
Enany et al.,[20] found that contamination rate of yoghurt
by Aeromonas species using enrichment technique on
(SAA) was 17.8%.
Yoghurt is one of the most popular milk products
specially for young and old people. Result of this study
indicate that mesophilic Aeromonas species may be
present in raw milk or contaminate it during preparation
of yoghurt, also storage of the product at refrigeration
temperature may result in high number of Aeromonas
which could cause food borne illness. Result of the
examination of kareish cheese samples revealed that
mesophilic Aeromonas species were detected in 20 out of
30 (66.7 %) and 21 out of 30 (70 %) kareish cheese
samples examined using (SAA) and (MMA) presented in
Table (1 and 5) with mean counts of 6.3x105 and 8.3x105
CFU/g respectively. Similar finding was reported by
EL-Prince[15], Khalil[19] and Enany et al.,[20]. Higher
incidence (92%) was recorded by AL-Ashmawy[30]. On
the other hand, Abdel-Hady et al.,[31] reported that
isolation rate of Aeromonas species from fresh skim milk
salt cheese using (SAA) was 20%.
Kareish cheese is considered the main protein
supplement for average class population due to low price
and low fat content so presence of mesophilic Aeromonas
species in this level could cause food born illness. The
high incidence of mesophilic Aeromonas isolates may be

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 J. Appl. Sci. Res., 2(11): 1037-1041, 2006

Table 7: Incidence of mesophilic Aeromonas isolates recovered from different samples.

Aeromonas species
-----------------------------------------------------------------------------------------
Number of isolates A. hydrophila A. cavia A. sorbia
----------------------- ------------------------ ------------------ --------------------
Types of samples Number of samples No % No % No % No %
Raw milk 30 39 37.9 26 66.7 9 23.1 4 10.2
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Pasteurized milk 30 10 9.7 6 60 3 30 1 10
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Yoghurt 30 30 29.1 22 73.3 5 16.7 3 10
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Kareish cheese 30 24 23.3 17 70.8 4 16.7 3 12.5
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Total 120 103 100 71 69 21 20.4 11 10.6

attributed to contamination of used raw milk, unhygienic prevent growth of Aeromonas organisms in milk during
practices during manufacture improper handling and storage.
storage and low salt content.
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