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Bacteriological Quality Assessment of Milk in College of Veterinary Medicine (Cvm) Dairy Farm and Kalamino Dairy Farm in Mekelle, Tigray, Ethiopia
Faisal Said Mohamed* and Ahmed Ali Farah
Consultant Research of Ministry of Livestock and Animal Husbandry, Mekelle University, Ethiopia
Submission: October 28, 2018;Published: November 20, 2018
*Corresponding author: Faisal Said Mohamed, Mekelle University, College of Veterinary Medicine, Mekelle, Ethiopia
How to cite this article: Faisal S d, Ahmed Ali F. Bacteriological Quality Assessment of Milk in College of Veterinary Medicine (Cvm) Dairy Farm and Kalamino Dairy Farm in Mekelle, Tigray, Ethiopia. Dairy and Vet Sci J. 2018; 8(2): 555734. DOI:10.19080/JDVS.2018.08.555734
Milk is an important source of nutrients to human and animals, but due to its high-water activity and nutritional value, it serves as an excellent medium for growth of many kinds of microorganisms under suitable conditions. The present cross-sectional study was conducted to assess hygienic practices, determination of bacterial quality of milk and isolation and identification of bacterial pathogens in milk at each critical control point throughout the value chain in MU-CVM and Kalamino dairy farms in Mekelle from August to November 2017. A total of 40 respondents were interviewed and subsequently, 84 milk samples were collected for laboratory analysis including bacterial load assessment and isolation and identification of bacteria. Total bacterial plate counts from milk were conducted. Isolation and identification of the bacteria in the milk was also conducted following standard methods. Results showed that, attendants of study dairy farms, managing their cattle in moderate hygienic environments and practicing intensive farming system.
The mean total bacterial plate counts of raw milk samples analyzed were 2.15x108 cfu/ml (udder), 3.4x108 cfu/ml (storage area in the farm) and 5.96x108cfu/ml (distribution center milk container). The increment of both counts at each critical control points was observed statistically significant (P=0.000). However, there was no significant variation between the two farms (P= 0.084) in the mean total bacterial count. About 26.7%, 50%and 62.5% from teat, storage area in the farm and distribution center milk containers, respectively at MU-CVM and 40.9% from the teat, 62.5% from milk storage area and 75% from distribution center milk containers, respectively at Kalamino dairy farm were graded of poor quality. In the course of this study, the frequent bacterial pathogens isolated from raw milk samples taken from different critical points include: E. coli, S. aureus and Streptococcus spp. From the result there were hygienic practices in the study farms, but the quality of milk used for human consumption area was found inferior quality according the standard level. Thus, it is important practice proper and restricts hygienic practices, proper transportation and storage and reducing the milk storage time, raising the awareness of dairy workers to enhance the quality of milk used for human consumption.
Cow milk has long been considered a highly nutritious and valuable human food and it is consumed by millions daily in a variety of different products . Raw milk of good hygienic quality meets the nutritional needs of body better than any single food as it contains essential food constituents such as fat, proteins, carbohydrates, minerals and vitamins . As a result of the presence of these nutritional components, milk is an excellent culture medium for many microorganisms, especially bacterial pathogens . Milk is often prone to early contamination and spoilage if not handled properly .
Microorganisms present in milk can be classified into two main groups: pathogenic and spoilage organisms, although some may play a dual role for example Bacillus cereus. Pathogenic organisms are those capable of inducing food poisoning, thus posing a threat to public health . These pathogenic microbial contaminants in milk have been a major factor for public health concern since the early days of dairy industry . There is a constant challenge to those involved in milk production to prevent or minimize the entry and subsequent growth of microorganisms in milk . These is mainly due to the importance of producing milk of good hygienic quality, which is necessary to milk product of superior quality and prolonged shelf-life thereby to provide a safe and wholesome food for the consumers . Bacterial contamination can generally occur from three main sources; within the udder, outside the udder and from the surface of equipment used for milk handling and storage .
Once milk is secreted out of the udder of the cow, the
retention of milk requires cleanliness, sanitation and cooling .
Fresh milk drawn from a healthy cow normally contains a low
microbial load of less than 103cfu/ml . However, the bacterial
load may increase up to 100-fold or more if stored for sometimes
at ambient (30 to 35°C) temperature . Milk produced under
hygienic conditions from healthy animals should not contain
more than 1 x 105cfu/ml.
Ethiopia possesses the largest livestock population in
Africa. Estimates for farmer holding in rural areas indicate
that the country has about 53.99 million heads of cattle, 24.6
million goats, 25.5 million sheep and 0.92 million camels .
In Ethiopia dairy production depends mainly on indigenous
livestock genetic resources; more specifically on cattle, goats,
camels and sheep. Cattle has the largest contribution (81.2%) of
the total national annual milk output, followed by goats (7.9%),
camels (6.3%) and sheep (4.6%).
While the industry is growing at a rapid rate, no milk quality
standards currently exist, therefore, it is important to establish
milk quality standards that focus on food safety measures in order
to improve public health. This is one reason why milk testing and
quality control include hygiene as well as microbial qualities in
addition to testing for fat content and heat stability . Prior
to the discovery and widespread adoption of pasteurization
for instance, raw milk and its products were responsible for
serious bacterial infections such as diphtheria, scarlet fever and
tuberculosis . Consumers all over the world are increasingly
concerned about the safety of their food in general and milk and
milk products. Therefore, quality should not be ignored at all
stages of the dairy value chain.
There is limited data on hygienic practices throughout the
dairy production system in Ethiopia and standard milking
procedures do not exist. A recent study in Ethiopia showed
many farmers do not properly clean teats prior to milking.
The study also showed a trend of farmers either not using a
towel at all for disinfection or using a collective towel for two
or more cows . This practice can clearly lead to the spread
of contagious pathogens. Raw milk is an important vehicle for
the transmission of milk-borne pathogens to humans, as can
be easily contaminated during milking and handling . Poor
or improper handling of milk can exert both a public health
and economic constraints thus requiring hygienic vigilance
throughout the milk value chain .
In some parts of the world including developing countries
like Ethiopia, milk is still a significant source of these infections
and other FBDs . Consumption of raw milk and its derivatives
is common in Ethiopia , which is not safe from consumer
health point of view as it may lead to the transmission of various
diseases. Raw or processed milk is a well know food medium that
supports the growth of several microbes with resultant spoilage
of the product or infections (intoxications) in consumers .
Even though milk represents an important place in the
nutrition of consumers as well as nutrition and income of
producers, there is limited work so far undertaken regarding
assessment of bacteriological quality of raw cow milk in northern
Ethiopia in general and dairy farms in the Mekelle particular.
Determining the current status of bacteriological quality of milk
at MU-CVM dairy farm and Kalamino dairy farm would create
awareness on the bacterial safety in the milk and support for
strengthening the hygienic standards practiced at different levels
of the production chain. Thus, the present study is designed with
the following objectives
a. To evaluate the hygienic practices and sanitary
standards at MU-CVM and Kalamino dairy farms.
b. To determine the bacteriological quality of milk at the
study dairy farms.
c. To isolate major bacterial species from milk samples
having high bacterial load.
MU-CVM and Kalamino dairy farms are in southern part of
Mekelle city Tigray region, 13032’N and 39033’E, and the city is
2200m above sea level. The climate is semi-arid with an average
annual rain fall of 600mm/Hg and the temperature ranges
between 12 to 27.1 °C.
Both farms are intensive dairy cows, MU-CVM dairy farm
is owned by the College of Veterinary Medicine of Mekelle
University and Kalamino dairy farm is owned by Tigray
Development Association (TDA). MU-CVM farm was established
in August 2009 with 11 cows of HF cross breed, in 2012 it was
modernized. During this study period 110 cattle were in the farm
of which 84 were exotic breeds and 2 cross-breeds, and 30 of
them were lactating cows. Kalamino dairy farm was established
in 1997(GC). On its establishment the farm has begun the project
by 61 cross breed cows of HF and currently the farm has 87 HF
cows, with 22 lactating cows.
A cross-sectional study was conducted from August to
November 2017, to assess the general sanitary conditions
of milking areas in dairy farms and examine bacteriological
quality of milk. The study involved MU-CVM and Kalamino dairy
farms and study unit was lactating cows in these farms where
questionnaires were administered, and raw cow milk was
collected for bacteriological analysis.
First, a pilot survey was made in order to gather information
on lactating cows of study dairy farms, fifty-two lactating cows
were identified in the study farms (30 in MU-CVM and 22 in
Kalamino dairy farms). In the farms there was a different critical
control points throughout the value chain, such as milk collecting
areas during milking. Both farms have milk distribution centers.
The milk samples were collected from udder of all lactating cows,
milk containers in the collection area and utensils of distribution
center of farms. Thus, a total 84 samples (52 milk direct from
teats in all lactating cow were taken, 16 (8 in each farm) from
milk containers in collecting area immediately after milking and
16 (8 in each farm) from distribution sites immediately before
distribution of milk samples were collected by random selection
using sterile bottles. At all levels of sampling, the sampling bottles
were capped, labeled with a permanent marker and transported
to MU-CVM microbiology laboratory in an ice box, and stored at 4
°C. Culturing was conducted within 24 hours after sampling .
Structured questionnaire (Annex1) was used to collect
information from workers of study dairy farms, such as animal
health workers, individuals involved in milking as well as
handlers of the milk. Twenty respondents in each farm were
interviewed with pre-coded response choices (closed-ended
questions). The questionnaires were designed to get information
on possible risk factors for bacterial contaminations in milk. Risk
factors considered in the current study were sanitary conditions
of the barn/milking environment, hygiene of milking cows’
udder and milk handlers, hygiene of milking equipment with
special emphasis to hygiene of milking procedures and milk
handling practices, milk containers used for milking, storage and
transportation. The questionnaire was administered through
face to face interview. While administering questionnaires, direct
observation on general cleanliness and hygienic conditions and
practices about milk were also done and noted.
The total bacterial count was made by adding 1ml of sample
into sterile test tube having 9ml normal saline solution (NSS).
After thoroughly mixing, the sample was serially diluted up to
1:10-6 and 1:10-7 then duplicate samples (1ml) were pour
plated using 15-20 ml standard plate count agar solution and
mixed thoroughly. The plated sample could solidify and then
incubated at 37 °C for 48 hours. Finally, counts were made using
a colony counter after incubation of the cultures at 37 °C for 48
hours. All colonies including those of pin point size were counted
on selected plates using colony counter. Results from plates,
which contained 30 to 300 colonies per plate were recorded.
Plates with more than 300 colonies could not be counted and
were designating as TMTC (too many to count) while plates
with fewer than 30 colonies were designate as TFTC (too few to
count). The plate counts were expressed as colony forming unit
of the suspension (CFU/ml)  and the average for each sample
were recorded as CFU/ml. Samples were graded as very good if
the total bacterial count did not exceed 2x105 CFU/ml, good if it
was between 2x105 and 1x106 CFU/ml and fair if the count was
between 1x106 and 5x106 CFU/ml. Samples having bacterial
count above 5x106 CFU/ml were graded as poor quality .
Milk samples that were graded as poor quality were
considered for bacterial isolation and identification. Isolation
and identification of bacterial species was carried out based on
conventional culture technique and biochemical assays. After
thorough mixing of each milk samples, a loop full of the milk
sample was streaked on the blood agar base enriched with 7%
sheep blood agar and MacConkey agar. Both agar plates were
incubated at 37 °C and examined for bacterial growth after 48
hours and then bacterial colonies were identified. Pure culture
colonies were selected and sub-cultured on nutrient agar (Oxoid
UK) and incubated aerobically at 37 °C for 24 - 48 hours for
biochemical testing. The identification of the bacterial species
was performed using biochemical tests and culturing on selective
media. All media were prepared according to the manufacturer’s
The data was entered excel spread sheet and analyzed
using SPSS version 16.0 a statistical software. Percentages were
also used to express the proportion of bacterial isolation and
milk quality grade based on Indian standards. The differences
in bacterial load between the samples from dairy farm, milk
distribution center, hands of milkers and surface milk containers
The floor of both farms was constructed with concrete
materials (cement) where it facilitates easy cleaning. About
80%of the respondents at MU-CVM farm indicated they clean
the barn two times daily, while in Kalamino farm 85% reported
that they clean one’s day. Both farms use tap water for cleaning
the farm (Table 1). In both farms milking is done manually twice
a day. The study farms use aluminum container (buckets) during
milking while MU-CVM and Kalamino farms use 75% and 95%
metal pots, respectively for storage and transportation. Most of
the respondents practiced washing of their milk utensils daily;
while MU-CVM However, the cleaning is moderate in both but
not efficient and utensils are not well dried. It was observed that
milkers dip their fingers in the milking vessel to moisten teats of
the cows with the intention of facilitating milking (Table 2).
In the present study, majority of the respondents across the
2 farms wash their hands before milking, while around 80% of
the millers at MU-CVM wash their hands before and after every
cow milking, the rate of hand cleaning at Kalamino farm was
60%. About 90% and 75% of the respondents at MU-CVM and
Kalamino farms respectively indicated washing udder before
milking. Generally, it was observed that the person involved in
milking was not clean; also, the hygiene milking environments
was moderate, although Kalamino farm was low hygiene than the
MU-CVM farm and the utensils for milking and transportation
were looked well in both, but there were the possibilities
indicated for microbial contaminations of milk.
In both farms most of the milk was transported to their own
milk distribution centers where their customers collect the milk.
The milk distribution center of MU-CVM dairy farm is within the
campus, but distribution centers of Kalamino dairy farm is in the
down town of Mekelle city, therefore after milking takes time
to reach the milk distribution center and there is opportunity
increase of microbial contamination of milk.
Total bacterial load: The mean for total bacterial plate
counts (TBPC) of raw milk samples at three critical points
within the two farms are shown in Table 3. The overall mean
TBC was 2.15x108, 3.4x108 and 5.96x108 CFU/ml for milk
samples collected directly from the udder, milking container
in temporary storage area in the farm and milk containers in
distribution centers, respectively. There was an increasing
trend of total bacterial count as the milk passed through udder,
milking storage area and distribution centers. Accordingly, the
count increased by 1.25x108 CFU/ ml from point of production
(milk sampled directly from the teat) to milk samples taken from
milking bucket in storage area at the farm.
SA = Storage area in farm; DC = Distribution center in a farm; critical control points of sampling.
Likewise, TBPC increased by 2.56 x108 CFU/ ml from
milking bucket in storage area at the farm level to distribution
center milk container. The increase from point of production
to distribution center milk containers was 3.81x108 CFU/ ml.
There was a difference in mean of total bacterial count between
the farms where 2.98x108 and 4.4x108 CFU/ml for milk samples
collected from MU-CVM and Kalamino dairy farms, respectively.
However, the difference was not statistically significant (p=
0.084), while analysis of variance indicated that there were
statistically significant differences in total bacterial count (p<
0.001) between the critical points.
Quality of Milk Samples: Milk samples having total bacterial
counts higher than acceptable level (2x106CFU/mL) according
to Sherikar  were considered as poor quality. Out of the
total milk samples collected from MU-CVM dairy farm, 26.7%,
50%and 62.5% of the samples from direct cow’s teat (udder),
burkes in milk storage area and milk containers in distribution
center, respectively were considered as poor quality while
40.9%, 62.5% and 75% milk samples collected from Kalamino
dairy farm from direct cow’s teat, buckets in milk storage area
and milk containers at distribution center, respectively were
graded as poor quality (Figure 1).
The major bacteria isolated out of from milk samples having
high bacterial load include S. aureus, Staphylococcus. species other
than S. aureus and Streptococcus species, E. coli, other coliform
bacteria, Salmonella, Klebsiella and Pseudomonas (Table 4).
The overall purpose of this study was to assess the hygienic
condition and bacterial quality of raw cow milk in MU-CVM and
Kalamino dairy farms in Mekelle. The result of questionnaire
survey and observation in the study farms showed that milk was
generally produced by dairy producers under moderate hygienic
environmental condition. This study further revealed that both
dairy farms managed their cattle in shaded cattle houses that are
cleaned daily. Attendants try to clean environments but there are
sources of milk contaminations. The present study also showed
that most of the persons involved in milking activities were also
not clean with their body and clothes during milking and milking
utensils although better than traditional way, these possibilities
predisposed milk to microbial contaminations at farm level.
It was further found that factors that were likely sources of
microbial contamination in milk include hand milking in a same
times dirty animal house, not washing udder and/or teats before
and after milking with clean water and some milkers not drying
the udder before milking. Most of the respondents (80%) clean
the barn on daily basis by removing the feces except weekends.
Similar finding was also reported in Sidama highlands of
Southern Ethiopia .
Milking was also performed in the same place after cleaning.
Although, the high proportion of dairy cows their barn floors,
clean, dry and comfortable bedding condition is important to
minimize the growth of pathogenic microorganisms. Practices
that expose the teat end to these organic bedding sources, wet
and muddy pens increase the risk of occurrence of mastitis and
milk contamination . Unless properly handled, milk can be
contaminated by microorganisms at any point from production to
consumption. Producers should therefore make udder washing a
regular practice in order to minimize contamination and produce
good quality milk. Most of the respondents who practiced udder
and teat drying use towel (80% in CVM 75% in Kalamino). It
was reported by , that pre-milking udder preparation and
teat sanitation play important part in the microbial load of milk,
infection with mastitis, and environmental contamination of raw
milk during milking. Cleaning the udder of cows before milking
is important since it could have direct contact with the ground,
urine, dung and feed refusals while resting. Lack of washing
udder before milking can impart possible contaminants into the
Production of milk of good hygienic quality for consumers
requires good hygienic practices (clean milking utensils,
washing milker’s hands, washing the udder and use of individual
towels) during milking and handling, before delivery to
consumers or processors . In the study area, most of the
respondents practiced washing of their milk utensils. However,
some of worker of study farms not uses detergents for cleaning
and utensils are not properly dried. Surfaces such as milking
equipment’s’ and hands meeting milk if not clean enough may
cause milk contaminations. During the current study, aluminum
containers were the major utensils for collection and storage
of milk. In such a situation, microorganisms can rapidly build
up in potentially nutritious milk residues of storage containers
consequently contaminating the milk on subsequent uses.
Contrary observations were also reported by Shija , who
observed high microbial load in milk which was correlated with
narrow necked plastic containers used in handling of milk.
The increment of bacterial load could be attributed to
contamination of the milk throughout the value chain from
production by different environmental factors through different
exposures of contamination like pooling of milk from different
animals together with unhygienic handling and leaving the milk
without cooling. In the present study highest bacterial load was
in distribution center and followed by bulks in storage area and
least bacterial load was from milk collected directly from cow’s
teats (udder). This result agreed with the report by Zewdu  in
Sidama highlands of Southern Ethiopia and Farah  in Somalia
who also reported an increase in bacterial counts through milk
value chain showing the highest count from bulk milk of cow
stored for 24 hrs. without cooling. After milk passed one critical
control point to another (from udder to temporary storage area)
due to interactions of milk handles, transfer of milk in different
containers and multiple sources the milk contamination rate will
rise. This was also evidenced in the present study.
The mean total plate bacterial counts from teat milk samples
was lower than mean value reported by Alehegne . Similarly,
the mean value of total plate bacterial counts for milk samples
taken from the milking bucket in temporary storage area at
farm level (3.4x108 CFU/ml) were lower than that of previous
reports for the same sample source . The probable reason
for the difference could be the use of aluminum containers which
are easy to clean and better hygienic standards in the farms
considered in the present study.
In general, present study is lower TPBC than majority of
the previews study reported in Ethiopia due to improvement of
hygiene on study farm and also these farms are generally proper
milk hygiene and milking activities compared with reported dairy farming systems in Ethiopia in previous studies, although
mean of TPBC observed in the current study was higher than the
maximum recommended level of 2.0 x 106 CFU/ml. That mean
as observed during sampling, high TPBC obtained in the current
study might be related to the overall sanitary conditions followed
which causes bacterial contamination in a milk.
Out of the total milk samples collected from MU-CVM dairy
farm, 26.7%, 50% and 62.5% of the samples from direct cow’s
teat (udder), burkes in milk storage area and milk containers in
distribution center, respectively were considered as poor quality
while 40.9%, 62.5% and 75% milk samples collected from
Kalamino dairy farm from direct cow’s teat (udder), buckets
in milk storage area and milk containers at distribution center,
respectively were graded as poor quality, similarly by Redda 
poor quality milk in dairy farms, cafeterias and wholesalers in
Adigrat town of Tigray region.
The most predominant bacteria isolated in the present study
were Staphylococcus spp. (37.25%), Streptococcus spp. (13.7%),
Escherchia coli (17.6%), other coliform bacteria spp. (11.8%),
Salmonella spp. (5.9%), Shigellas pp. (3.9%) and Pseudomonas
spp. (3.9%). These bacterial groups are the most common
contaminants of raw milk and responsible for causing mastitis
in dairy farms [33-36].
Milk intended for human consumption must be free from
pathogens and must, if conditions permit, contain no or few
bacteria. Clean milk could only be obtained if effective sanitary
measures are taken starting from the point of milk withdrawn
from the cow until it reaches the consumers . From the
findings of this study, it is concluded that; the milk produced
by at MU-CVM and Kalamino dairy farms generally had higher
TBPCs according to the international acceptable limits. Although,
the mean TBPCs in the study farms were lower than previous
reports, it calls for immediate action to improve the hygiene and
sanitary measures at different critical points where milk could
get contaminated. Majority of raw milk samples from distribution
center containers for consumers had higher TBPCs, hence, its
keeping quality would be lower and some of the pathogens
present in the milk have public health significance.
Based on the findings of the present study, the following
recommendations are made:
a. Raising the awareness of farm workers on good
hygiene and sanitary measures could significantly reduce the
b. Ensuring proper transportation and storage of the
milk, and reducing the milk storage time, at milk distribution
centers should be considered.
c. Routine assessment of milk quality by the farms should
be performed on a regular basis to ensure the supply of good
quality milk to consumers.