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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


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

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

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b-Enrichment of the samples: Ten ml of raw and Table (1) revealed that the contamination rate of raw
pasteurized milk and 10 gram of yoghurt and kareish milk by mesophilic Aeromonas species using enrichment
cheese were homogenized in 90 ml Trypticase soya broth technique on (SAA) and (MMA) were (80%) and (86.7%)
containing 10 µg ampicillin/ml and blended for 2 minutes, respectively. While the count rang were ranged from
then incubated at 28EC for 24 hours Villari et al.,[14]. 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
c-Enumeration of Aeromonas species: The enrichment value 9.2 x106 on (SAA) and (MMA) respectively, as
cultures were serially diluted up to 10-6, then 0.1 ml from shown in Table 2. Similar finding was reported by Hafez
each dilutions was spread over starch ampicillin agar and Halawa[18] and Khalil[19]. Low incidence of
(SAA) and MacConkey Mannitol ampicillin agar (MMA) Aeromonas in raw milk had reported by Enany et al.,[20]
and Amany Shalaby[21]. Lower finding 2.44 %, 9.7 % and
with sterile bent glass rod and incubated at 28EC for
14.9 % were recorded by Banerjee and Block[22],
24 hours then the numbers of isolated Aeromonas were
Ergullu[23] and Shweizer et al.,[24] respectively. The higher
calculated and typical colonies were picked into triple incidence of Aeromonas in raw milk could be attributed
sugar iron (TSI) and nutrient agar slants for biochemical to its wide distribution in the nature; Aeromonas
identification EL-prince[15]. microorganism is commonly present in farms, in feeding
stuff, water, faeces and soil El-Shenawy and Marth[25].
d-Identification of Aeromonas species: Presumptive The organisms can invade the udder tissues; multiply in
aeromonad were identified by biochemical tests mammary tissues and subsequently discharge in milk.
according to Koneman et al.,[10]. Also the contaminated water used for washing milking
equipments is considered as a significant source of
Statistical analysis: Minimum, maximum, mean, contamination. So presence of mesophilic Aeromonas in
standard deviation and standard error of mean were used a high level in raw milk samples is indicative to bad
to describe data. T-test was used to evaluate the efficacy hygienic measures of milk production and distribution.
of both media used for Aeromonas isolation and counting. Concerning the contamination rate of pasteurized
P value was considered significant if less than 0.05 and milk by mesophilic Aeromonas species using (SAA) and
0.01 at 95% and 99% respectively. These tests were (MMA) were 16.7 % and 16.7 % respectively as in
analyzed using the Statistical Package for Social Table 1. While the count rang from 0.00 up to 8.5 x105
Scientists (SPSS) for windows 10.0 (SPSS Inc., Chicago, with the mean value 4.7x104 CFU/ml and 0.00 up to 9x105
IL and USA). 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
RESULTS AND DISCUCSIONS
positive for A. hydrophila. Lower incidence recorded
by Kirov et al.[27]. The process of pasteurization was
In the recent years, Aeromonas has received done at 71-75EC for 15 second. Aeromonas organisms are
increasing attention as an agent of food borne diarrheal sensitive to temperature above 48EC Palumbo et al.,[28]
diseases in healthy people Gracey et al.,[16] and and the strain of Aeromonas should have been killed
Palumbo et al.,[3]. The ability of aeromonads to grow at during this process. So presence of mesophilic Aeromonas
low temperature is of great concern, particularly in foods, species in the pasteurized milk may be due to inefficient
which are stored under refrigeration. This explains that pasteurization or post pasteurization contamination during
Aeromonas could a potential food born pathogen and packaging process of pasteurized milk. The contamination
dairy products may represent an important vehicle of its of the pasteurized milk may be introduced by subsequent
transmission Freitas et al.,[17]. handling of the milk.

Table 1: Efficacy of solid specific media for isolation of mesophilic Aeromonas from different examined samples.
Positive samples
--------------------------------------------------------------------------------------------------------------------
Starch ampicillin agar MacConkey Mannitol ampicillin agar
----------------------------------- ------------------------------------------------
Types of samples Number of samples No % No %
Raw milk 30 24 80 26 86.7
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Pasteurized milk 30 5 16.7 5 16.7
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Yoghurt 30 22 73.3 23 76.7
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Kareish cheese 30 20 66.7 21 70
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Total 120 71 59.1 75 62.5

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

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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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