Academia Journal of Microbiology Research 4(4): 053-056, April 2016
DOI: 10.15413/ajmr.2016.0305
ISSN: 2315-7771
©2016 Academia Publishing
Research Paper
Bacteriological Quality of Bottled Water Brands in Riyadh Saudi Arabia
Accepted 25th April, 2016
ABSTRACT
A Mohammad. Al-Sulaiman
Community College, Huraimla, Shargra
University, P. O. Box 300, Huraimla 11962,
Saudi Arabia.
*Corresponding author e-mail:
mohazizsu@yahoo.com.
In the present study, bacteriological qualityof 448 different mineral water bottle
brands marketed in Riyadh w a s analyzed from July, 2013 t o January, 2014. All
the collected samples were analyzed for the presence of total coliform bacteria,
Escherichia coli bacteria and Pseudomonas aeruginosa. 24(5%) of these samples
were contaminated with pseudomonas spp and 4 ( 0 . 8 % ) contaminated with E.
coli. The presence of pseudomonas spp and total coliform and E. coli bacteria in
drinking water suggests the possible presence of pathogenic enteric microorganisms, thus, rendering the water unsafe for drinking. The data presented here
clearly raised the concerns regarding the quality of drinking mineral water and
highlights the danger posed to public health.
Key words: Coliform bacteria, E. coli,P. aeruginosa, bottled water.
INTRODUCTION
Safe and hygienic water is nowadays of great concern to
consumers. This is due to the risk of waterborne disease
like cholera, diarrhea, typhoid transmission (Mavridou,
1992). Consequently, the demand for bottled water has
tremendously increased in recent years along with the
necessary measures for its quality (Venieri et al., 2006).
Samadi et al. (2009) reported an annual consumption of
1.3 × 1011 L of bottled water worldwide.
In the last decade in North America the consumption of
bottled water increased by 25% (Bharath et al., 2003).
Bottled water may contain strains of micro-organism that
already exist in their source, that is, spring and bore wells
(Warburton, 1993). Coliform bacteria, E. coli or
Pseudomonas are the most common microbiological agents
in bottled water that greatly affect public health. Infants,
children, elderly people and immuno-compromised people
especially are at high risk of waterborne diseases due to
such micro-organism ingestionevenif at low infective
doses (WHO, 2005).
Moreover, Pseudomonas aeruginosa is a drug resistant
bacterium that can transfer its drug resistant genesto other
bacteria in the human body (Salvatore et al., 1995). Victoria
and Galvan (2001) reported cases of gastroenteritis in
children from Mexico as a result of water contamination by
Escherichia coli and Pseudomonas. Detection of E. coli in
water is linked to recent fecal pollution and thus, its
presence is an indicator for such pollution (Victoria and
Galvan, 2001). Pseudomonas is an opportunistic bacterium
that can induce infection for immune-compromised
patients who suffer open wounds, soft tissue or respiratory
system infections (Zamberlan et al., 2008) and hence, its
presence is unacceptable (Romling et al., 1994).
The Saudi Arabian Standards Organization (SASO) has set
standards for bottled water quality based upon the WHO
measures (WHO, 1971; SASO, 1984). Regarding
microbiological contamination, the WHO has recommended
that potable water should be devoid of Coliform bacteria,
fecal coliform (E. coli, entrococci and P. aeruginosa) and
contain less than 20 CFU/ml heterotrophic bacteria count
(Romling et al., 1994). Al-Zahrani and El-Hamshary (2013)
detected Staphylococcus aureus and P. aeruginosa in bottled
water in Jeddah, while Al et al. (2013) reported the same
for bottled water from Riyadh.
The main objectives of the current study are to analyze
the bacteriological quality and bacterial contamination of
bottled drinking water from Riyadh markets and to check
Academia Journal of Microbiology Research; Al-Sulaiman.
their compliance with the standard. Also, the effect of
storage and other factors like temperature, light and
temperature upon bottled water samples was also
investigated.
MATERIALS AND METHODS
054
this second incubation were used to statistically determine
the MPN from the appropriate reference chart. P.
aeruginosa was cultured in blood agar plate where 1.0 ml
was added and then incubated at 37°C for 24 h. The
presence of P.aeruginosa was confirmed by oxidase test
where the appearance of a dark purple color indicated a
positive reaction.
Sample collection and storage
RESULTS AND DISCUSSION
A total of 448 bottled water samples from seven brands
commonly sold in Saudi Arabia were bought from retail
outlets from North Riyadh, Saudi Arabia. The samples
were of 330 and 600 ml and 20.0 L bottles and 125 and
250 ml cups respectively. Samples were labeled using
alphabet letters instead of brand names for commercial
considerations. The samples were stored from 24 th July,
2013 up to 24th February, 2014 in four rooms under
different conditions:
1) The first room (L1) is an air conditioned chamber in
which the temperature was fixed to 24°C without any
access to sunlight;
2) The second room (L2) was a closed chamber with
neither air conditioning nor sunlight;
3) The third room (L3) was an open top chamber exposed
to direct sunlight without air conditioning;
4) The fourth room (L4) was semi-open chamber in which
the sunlight partially penetrates to the samples.
Bacterial analysis
For enumeration of heterotrophic bacteria, the pour plate
methods were employed (Martin et al., 2004). 1.0 ml of
bottled water sample was mixed with melted plate count
agar. Two plates were prepared for each sample where
one plate was incubated aerobically at 37°C for 24 h and
the other at 22°C for 72 h. Colonies were then counted as
CFU/ml.
The Multiple Tube Fermentation (MTF) technique is
useful in determining the fecal coliform density in most
water, solid or semisolid samples. The technique is based
on the most probable number (MPN) of bacteria present in
a sample which produce gas in a series of fermentation
tubes with various volumes of diluted sample. The MPN is
obtained from charts based on statistical studies of known
concentrations of bacteria. About1.0 ml sample from each
dilution was taken to inoculate three series of three test
tubes containing MacConkey broth (company) containing
litmus indicator in double and normal slant concentration
with Durham's tubes dilutions first incubated in lauryl
(sulfonate) tryptose broth at 37°C for 24 to 48 hto
determine total coilform. Positive samples were then
transferred to EC broth and incubated at 44°C for an
additional 24 h to determine E. coli. Positive samples from
Riyadh city is a highly urbanized area of Saudi Arabia
where approximately 200 brands of bottled water are
marketed. To the best of our knowledge, very few studies
have been conducted to analyze the quality of bottled
mineral water from Riyadh (Al et al., 2013; Abd and
Alwakeel, 2007).
Therefore, this study was undertaken to analyze the
bacteriological quality of bottled water from markets and
to check their compliance with the standards. According to
the
guidelines
of
national
and
international
recommendations, all the specimens were checked for the
following parameters: presenceof total coliform bacteria,
fecal coliforms, E. coli and P. aeruginosa.
Heterotrophic bacteria are usually present in water
along with food, soil and air as they consume organic
nutrients for their growth (Martin et al., 2004). Table 1
shows that the samples have HPC less than 20 under both
22 and 37°C indicating that all water samples comply with
the Saudi and WHO set standards that restrict HPC of less
than 20. Similar studies were carried out by Tsai and Yu
(1997) in Taiwan where 51.1% of domestic and 60.4% of
imported exceeded 200 CFU/ml HPC/ml level. While in
Fiji, Zeenat et al. (2009) found that 28 to 68% of samples
analyzed were not complying with HPC standards.
Zamberlan et al. (2008) reported 87% of the bottled water
analyzed to contain more than 500 CFU/ml HPC limit set
by Brazilian quality standards. Their justification to such
high value was the ineffectiveness of disinfection process
to eliminate such bacteria. Many authors attributed the
high level of HPC either to the presence of bacteria in the
water source or contamination during bottling processes
(Sticker, 1989; Kassenga, 2007). It can be concluded that
all samples analyzed in this work are likely to have noncontaminated sources and /or proper bottling processes.
The results in Figure 1a and Table 1 indicates that no P.
aeruginosa growth was observed in L1 during all of the
storage time. On the other hand, P. aeruginosa started to
grow in a detectable quantity in the third month into water
samples in other chambers. The percentage of bottles
becoming contaminated by P. aeruginosa ranged between
4.69 (3/64) and 9.38% (6/64) respectively. In L1, there
was no Pseudomonas detected as the bottles were stored in
air-conditioned room with low temperature unfavorable
for bacterial growth. The conditions in the other rooms
were conducive for bacterial growth by providing warmth
Academia Journal of Microbiology Research; Al-Sulaiman.
055
Table 1: Pseudomonas and E. Coli growth at the different conditions rooms
Local
Month 1
Month 2
Month 3
Month 4
Month 5
Month 6
Month 7
L1
0
0
0
0
0
0
0
Pseudomonas
L2
L3
0
0
0
0
3
3
3
5
5
5
5
5
6
6
E. coli
L4
0
0
3
3
4
5
5
L1
0
0
0
0
0
0
0
L2
0
0
0
0
0
0
0
L3
0
0
0
2
2
2
2
L4
0
0
0
1
1
1
1
Figure 1. 1a) Bacterial growth in blood agar; b) Bacterial growth in MacConkey.
and light. Many factors such as available oxygen, rise in
temperature and nutrients degrading from bottles led to
bacterial growth in bottled water (Warburton and Konkle,
1992).
Bacteria can grow to the extent of affecting human
health due to storage of bottled water (Warburton, 2000).
Al-Zahrani and M El-Hamshary (2013) reported 3.9% P.
aeruginosa contamination in bottled water from Jeddah,
Saudi Arabia. They attributed the presence of bacteria to
the colonization of bottling plant equipment or disinfecting
detergents that may be organic nutrients.
Coliform organisms are used as water quality indicator
due to the ease of their detection and enumeration (WHO,
2005).Their presence is a clue to pathogen presence in
water as well as, an indicator for inadequacy of water
treatment
process
and/or
distribution
system
contamination. From the data presented in Figure 1b and
Table 1, no E. coliwas detected during the first three
months in all of the storage room. During the fourth
month, 1 and 2 samples out of 64 developed E. coli in
storage chambers of L4 and L3 respectively giving 1.56
and 3.13% contamination in the same sequence.Values of
1.2 and 4.0% bottled water contamination were reported
by Warburton and Konkle (1992) and Richard et al. (1994)
respectively. Zamberlan et al. (2008) reported less
contamination of municipal tap water as compared to
bottled water. This finding supports the hypothesis that
contamination taking place during the bottling process can
lead to bacterial growth especially if water is stored under
inappropriate conditions.
Contamination can be from the source of water subject
to human interaction. Zabed et al. (2014) reported positive
correlation between total coilform, fecal coliform and E.
coliwith the mass bathing, while Hong and Liang (2010)
measured higher population of coliform bacteria in urban
and industrial areas compared to remote areas. Venieri et
al. (2006) reported 11% total coliform and 1% E. coli in
bottled water marketed in Greece whereas, 5.2 and 1.5%
of total coliform and E. colirespectively were detected in
bottled water from Trinidad (Bharath et al., 2003). High
percentage up to 40% contaminated bottled water was
found in Brazil (American Public Health Association,
2005). At the bright side, some studies were carried out in
Taiwan (Tsai and Yu, 1997) and Greece (Kokkinkis et al.,
2009).
Physicochemical properties of water were found to
affect the growth and death of bacteria. For example,
bacteria cannot survive at high pH (Curtis et al., 1992) but
can have considerable growth in the presence of total
suspended solids to which bacteria can absorb and avoid
environmental factors such as UV-light and toxicity of
dissolved metals (Davies et al., 1995; An et al., 2002).
Academia Journal of Microbiology Research; Al-Sulaiman.
In all sample studied, the pH range between 7.0 and 8.0
was optimum for bacterial growth. Moreover, the
conductivity (data not included) was found to increase
with storage time from 220 to 290 uS. This observation
may indicate a release of ions from the PET bottle such as
Sb used as a catalyst for PET polymerization. Such process
will be accompanied with disintegration of the wall of
water bottle and availing organic nutrients on which
bacteria feed and grow. These two factors may contribute
to the bacterial growth observed as storage time increases.
It can be seen that the storage of water bottles for
extended intervals can lead to contamination with
bacteria. In a similar study, the storage of bottled water for
months facilitated the proliferation of micro-organisms
(Leclerc and Moreau, 2002).
Conclusions
Bottled water microbial analysis was performed under
different environmental conditions and time interval. The
bottled water was found to comply with the standards set
for drinking water quality. The results obtained indicated
that under normal low temperature the samples didnot get
contaminated with bacteria,while rise in temperature and
light led to a considerable bacterial growth. As a result, the
study recommends that bottled water should be kept at
low temperature, away from sunlight and consumed
within three months from bottling date.
REFERENCES
Abdand KF, Alwakeel SS (2007). Mineral and microbial content of bottled
and tap water in Riyadh, Saudi Arabia. Middle East J. Sci, Res. 2(3-4):
151-156.
Al-Hazzani AA, Al-Farra WL, Asran A, Shehata AI, Moubayed NMS (2013).
Bacterial quality of domestic and imported brands of bottled water in
Saudi Arabia. J. Toxicol. Environ. Health Sci. 5(10): 178-184.
Al-Zahrani HAA, El-Hamshary OIM (2013). Microbial quality of bottled
water and their molecular characterization in Jeddah, Saudi Arabia.
Life Sci. J. 10(4): 731-736.
American Public Health Association (APHA) (2005). Standard methods
for the Examination of water and wastewater, 21st Ed.
An YI, Kampbell DH, Breidenbach GP (2002). Escherichia Coli and total
choliform in water and sediments at lake Marina. Environ. Pollut. 120:
771-778.
Bharath J, Mosodeen M, Motilal S, Sandy S, Sharma S, Tessaro T, Thomas
K, Umamahesswaran M, Simon D, Adesiyun A (2003). Microbial quality
of domestic and imported brands of bottled water in Trinidad. Int. J.
Food Microbiol. 81: 53-62.
Curtis TP, Mora DD, Silva SA (1992). Influence of pH, oxygen and humic
substances on ability of sunlight to damage fecal coliform in waste
stabilization pond water. Appl. Environ. Microbiol. 58(4): 1335-1343.
Davies CM, Long JAH, Donald M, Ashbolt NJ (1995). Survival of
microorganisms in marine and freshwater sediments. Appl. Environ.
Microbiol. 61(5): 1888-1896.
Hong H, Qiu J, Liang Y (2010). Environmental factors influencing the
distribution of total and fecal coliform bacteria in six water storage
reservoirs in Pearl river delta region in China. J. Environ. Sci. 22(5):
663-668.
Kassenga GR (2007). The health-related microbiological quality of bottled
056
drinking water sold in Dar es Salaam, Tanzania. J. Water Health. 5: 179185.
Kokkinkis EN, Fragkiadakis GA, Kokkinaki N (2009). Monitoring
microbiological quality of bottled water as suggested by HACCP
methodology. Food Control. 19: 957-961.
Leclerc H, Moreau A (2002). Microbiological safety of natural mineral
water. FEMS Microbiol. Rev. 26: 207-222.
Martin JA, Stephen CE, Donald JR (2004). Heterotrophic plate count
bacteria-what is their significance in drinking water. Int. J. Food
Microbiol. 92: 265-274.
Mavridou A (1992). Study of the bacterial flora of non-carbonated natural
mineral water. J. Appl. Bacteriol. 73: 355-361.
Richard P, Le Floch R, Chamoux C, Pannier M, Espaze E, Richet H (1994).
Pseudomonas aeruginosa outbreak in a burn unit: role of
antimicrobials in the emergence of multiply resistant strains. J. Infect.
Dis. 170(2): 377–383.
Romling U, Wingender J, Muller H, Tummler B (1994). A major P.
aeruginosa clone common to patients and aquatic habitat . Appl.
Environ. Microbiol. 60: 1734-1738.
Salvatore M, Maurizio P, Michele F, Laura G (1995). Drug resistant
bacteria in non-carbonated mineral water. Microbiol. Res. 150: 403408.
Samadi MT, Rahmani AR, Sedehi M, Sonboli N (2009). Evaluation of
chemical Quality in 17 Brands of Iranian Bottled Drinking waters. J.
Res. Health Sci. 9: 25-31.
SASO (1984). Bottled and Unbottled drinking water, SSA 409/1984 2nd
ed. ISSN 1319-230.
Sticker DJ (1989). The microbiology of bottled mineral waters. J. Royal
Soc. Promot. Health. 109: 118-124.
Tsai GT, Yu SC (1997). Microbial evaluation of bottled uncarbonated
mineral water in Taiwan. Int. J. Microbiol. 37: 137-143.
Venieri D, Vantarakis A, Komninou G, Papapertropoulou M (2006).
Micobiological evaluation of bottled non-carbonated water from
domestic brands in Greece. Int. J. Food Micobiol. 107: 68-72.
Victoria J, Galvan M (2001). Pseudomonas aeruginosa as an indicator of
health risk in water for human consuption. Water Sci. Tech. 43(12): 4952.
Warburton DW (1993). A review of the microbiological quality in bottled
water sold in Canada Part 2. The need for more dtringent standards
and regulations. Can. J. Microbiol. 39: 158-168.
Warburton DW (2000). Methodology for screening bottled water for the
presence of indicator and pathogenic bacteria. Food Microbiol. 17: 312.
Warburton DW, Bowen B, Konkle A (1992). The survival of Pseudomonas
aeruginosa and its effect upon Samonellae in water: Methodology to
test bottled water in Canada. Can. J. Microbiol. 40: 987-992.
WHO (1971). European Standards for drinking water 3rd edition Geneva.
WHO (2005). Guidelines for drinking water quality. Health criteria and
other supporting information. 2 Geneva.
Zabed H, Suely A, Faruq G, Sahu JN (2014). Water quality assessment of
an unusual ritual well in Bangladesh and impact of mass bathing on
this quality. Sci. Total Environ. 472: 363–369.
Zamberlan da Silva ME, Santana RG, Guilhermitti M, Filho IC, Endo EH,
Ueda-Nakamura T (2008). Comparison of the bacteriological quality of
tap water and bottled mineral water. Int. J. Hyg. Environ. Health, 211:
504-409.
Zeenat A, Hatha AAM, Viola L, Vipra K (2009). Bacteriological quality and
risk assessment of the imported and domestic bottled water sold in
Fiji. J. Water Health. 7: 642-649.
Cite this article as:
Al-Sulaiman AM (2016). Bacteriological Quality of Bottled Water
Brands in Riyadh Saudi Arabia. Acad. J. Microbiol. Res. 4(4): 053-056.
Submit your manuscript at
http://www.academiapublishing.org/ajmr