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    <h3 class="center">The Environmental Impact of Urbanization within Abu Dhabi on the Microbial Profile of Man-Made Beaches</h3>

    <p class="body-bold center">Fatima Al Marzooqi and Fatme Al Anouti</p>

    <p class="body center">Zayed University, College of Sustainability Science and Humanities,
Department of Natural Science and Public Health
Khalifa City, Abu Dhabi, UAE
<br> U2880028@zu.ac.ae, Fatme.AlAnouti@zu.ac.ae</p>

    <p class="body"><b>Abstract</b> - <i>The United
Arab Emirates (UAE) is witnessing a significant development in several areas.
Many new construction projects are currently underway. One of the main project
aims at establishing new beaches around the coastal areas of the UAE. Particularly,
in Abu Dhabi the capital of UAE, a new man-made beach has been recently
developed in the north coast of the city. The urbanization of this newly
developed beach might have a significant impact on the environment. This study
aims to investigate the effects of urban development of beaches within Abu
Dhabi on the microbial profile of the soil. It is hypothesized that microbial
growth would be higher in man-made beaches as compared to natural beaches. Methodology
involved collection of 16 soil samples from two different beaches with emphasis
on two elements: water content in soil and depth of soil. Subsequently, samples
were processed and used for microbial cultivation using selective and differential
growth media for the identification of some commonly encountered microorganisms
namely "yeast, faecal coliforms,
Escherichia coli and Staphylococcus aureus". Polymerase Chain
Reaction (PCR) and electrophoresis were also used for further analysis. Results
of the study revealed the existence of Staphylococcus aureus,
Escherichia. coli and coliforms in the soil
samples isolated from the man-made beach. Moreover, the hypothesis was
supported by the results which showed higher microbial growth (CFU/g) for both
factors: (depth of the soil and water content) for man-made beaches as compared
to natural beaches.</i></p>

    <p class="body"><b><i>Keywords:</i></b> Microbiology, Beach, Abu Dhabi,
Environment, Urbanization, Bacteria</p>

    <p class="body">© Copyright 2015 Authors - This is an Open Access article published under the <a href="http://creativecommons.org/licenses/by/3.0" class="body-link" target="_blank" class="body-link">Creative Commons Attribution License terms</a>. Unrestricted use, distribution, and reproduction in any medium are permitted, provided the original work is properly cited.</p>

    <p class="body">Date Received: 2013-10-30 <br />

Date Accepted: 2014-04-11  <br />

Date Published: 2014-10-03</p>

  <div class="border"></div>

  <div class="indent">
<h4>1. Introduction</h4> 

<p class="body">Microorganisms have huge
effects on ecosystem processes that sustain life on earth. They produce oxygen,
store carbon and serve as essential factors in the nutrient cycle [14]. Species
usually live together and depend on each other to survive. Rarely can we find
ecosystems with little diversity. Each ecosystem has its own combination or
mixture of microorganisms. For example, bacteria often occur at higher levels
in the soil than in water. Also, they survive in the soil longer than they do
in the water [29]. Nevertheless, few studies show that the bacteria survive
longer in wet soil as compared to dry soil [30]. Many factors such as
temperature, depth, moisture, air, organic matter availability, light availability,
pH and size of soil particles play an important role in promoting the growth of
microorganisms. </p>

<p class="body">To maintain the normal cycle
of life, the ecosystem should function normally. However, if the ecosystem loses
one of it is vital components, detrimental effects could result. Many studies
related to soil microbiology have attempted to explore the potential pathogenicity
of microorganisms in beach soil. The World Health Organization (WHO) in 2003 [35]
indicated that there was no epidemiological evidence to show the correlation
between soils contaminated with microorganisms and human infection. Furthermore,
Prado et al., [27]
and Brand&atilde;o et al., [5] in their studies demonstrated a lack of correlation
between health and potentially pathogenic fungi inhabiting beach soil. However,
Brand&atilde;o et al., [5] mentioned that people
sometimes are responsible for contaminating beach sands by carrying microorganisms
that are left on the soil. Also humans could play a role by discarding organic
waste that serves as food for the development of yeasts, bacteria, and molds. To
some extent, those microorganisms will not cause severe effects to human unless
they reach a certain level. Therefore, the level of contamination in beach soil
should be known to ensure public safety. </p>

<p class="body">To determine the level of
contamination in beach soil, the faecal index of organisms is often used as a
reliable indicator of the level of non-potentially pathogenic microorganism.
These microorganisms are usually present in higher numbers as compared to the potentially
pathogenic microorganisms and are easy to isolate, identify and count. Species
included in the faecal index organisms are <i>coliforms</i> which consist of total <i>coliforms, thermotolerant
coliforms and E. coli</i>;
<i>intestinal enterococci,
clostridia and bacteriophages</i>. One of the recent studies conducted in Portugal regarding
microbial quality of beach soilrevealed the presence of potentially pathogenic
fungi, yeasts, <i>dermatophytes,
total coliforms, intestinal enterococci</i> and <i>E. coli</i> in soil samples taken from thirty three
public beaches around the country. In this study, the investigators proposed a
safe threshold for the potentially pathogenic microorganisms. They proposed 15
colony forming unit per gram (CFU/g) of soil for yeast, 17 CFU/g for potentially
pathogenic fungi, 8 CFU/g for <i>dermatophytes</i>, 84 CFU/g for <i>coliforms</i>, 250 CFU/g for <i>E. coli</i>,
and 100 CFU/g for intestinal <i>enterococci</i>. Potential health risks for human could occur if those
thresholds are exceeded [30].</p>

<p class="body">Bacterial species are
important to determine the potential risk level of beach soil. <i>Staphylococcus spp</i>. is a flora which is widely
spread in beach soil [8]. Examination of soil samples from two popular beaches
in Chile showed that from a total of 85 strains of Gram-positive bacteria detected;
cocci were the highest in number [27]. This was speculated to be the result of
human activity in beaches as some individuals might carry this species on their
skin. Another study in Portugal isolated <i>P. aeruginosa</i> and <i>Clostridium perfringens</i> from the soil of different
beaches around the country. The investigators suggested that Clostridium<i> perfringens</i> could be a good faecal index
microorganism [21]. </p>

<p class="body">With regards to potential
pathogenicity of fungi in beach soil, most fungal diseases are associated with immune-compromised
conditions [6, 21]. In the central coastal area of Portugal, <i>dermatophytes</i>
showed a high percentage of around 42% in the beach soil. While others were <i>T. rubrum, Trichophyton
mentagrophytes and Microsporum nanum</i>, all isolated from soil with low organic
nutrient [33]. These fungi are all potentially pathogenic and could hence have
serious implications on the health of beach users. </p>

<p class="body">Another important factor to
determine when studying microorganisms in beach soil is the water content in
the soil. Brand&atilde;o et al., [5] showed that the microbiological contamination is
higher in dry soil than in wet soil. Several others also showed that
contamination of soil was the highest in dry soil and during the summer than
during spring [20, 31]. However, others also showed that microorganisms survive
longer in wet soil [30]. Therefore, there is no clear conclusion whether growth
of microorganisms is strictly higher in dry or wet soil. Concerning the
correlation between species on wet and dry soil, it was found in Italy by Bonadonna
et al., 2002 that a correlation exist between <i>E. coli</i> and <i>enterococci;</i>
yeasts and molds; clostridial spores and <i>staphylococci</i> and between <i>enterococci</i> and spores of sulfite-reducing
Clostridium [35]. While, Papadakis <i>et al.</i>
[26] found no correlation between <i>Staphylococcus aureus</i> counts and the indices of faecal
pollution counted from the wet soil of the beach [35]. </p>

<p class="body">All those species detected in
the beach soil are associated with several factors such as tidal phenomena, nature
of the beach, seasonal variation, animals and human presence [35]. The nature
of the beach has an important effect on the pathological species which could exist
there. Wet soil which has organic substances can be a favourable environment
for enteric bacteria [26]. Mendes <i>et al.</i>
[21] studied the effects of tides on the number of pathogens and faecal index
microorganisms in the soil without finding any clear differences. However,
Obiri-Danso &amp; Jones [25] showed a very small difference for pathogens and
faecal index microorganisms count after and before tidal phenomena. Moreover,
in the same study campylobacters showed different counts with different
seasons, with the highest rate in winter 100%, followed by spring where it
reached around 33-67%. However, in autumn it increased again to around 67-78%.
The lowest counts were found in summer. </p>

<p class="body">Furthermore, when organisms
move from one environment to another many factors including temperature, water
availability, pH, and nutrient change. For instance, these microorganisms might
be exposed to higher temperatures and salinity or receive little rainfall in
their new environment. Some species can adapt to all the changes in the new
environment while others cannot and thus would not survive. Some adaptations could
cause microorganisms to be more potentially pathogenic or less productive than
others in their environment. </p>

<p class="body"> In Abu Dhabi, the Corniche
beach which is a new man-made beach had been recently developed in the northern
coast of the city. Although construction developments had been designed based
on an environmental framework which ensures sustainability [1] studies which examine
the microbial profile of soil from this newly developed beach are still lacking.
This study
explored the difference between Abu Dhabi man-made and natural beaches in terms
of soil microbial profile. Differences were examined with regards to soil water
content and depth. To
our knowledge, this is the first study which attempts to provide an idea about the
presence of both potentially pathogenic and non-pathogenic microorganisms
inhabiting the soil beach within Abu Dhabi.</p>

<h4>2. Materials and Methods</h4>

<p class="subhead">2.1. Site Description </p>

<p class="body">Sampling sites for collection
of soil samples in this study were chosen to be man-made and natural. The
man-made beach selected was the Abu Dhabi Corniche beach because it is not a natural
beach. It is located along the north-western shore of the capital of the UAE.
It is a newly developed beach that was awarded in 2009 to National Marine
Dredging Company (NMDC) for recreational development. It consists of the
development of a 3.5 km beachfront half of which is new [23]. It is protected
from erosion caused by tidal forces with a 3.5 km submerged rock sill below the
water, with sand in the top layer of beach equivalent to any internationally
acclaimed beach. Beach sand was imported to that beach and about 3.5 million m<sup>3</sup>
were dredged from the sea bottom and used [23]. At different period of time,
the sand had to be replaced and replenished with sand from other sources. Thus,
many differences between natural and exotic microorganisms could have been introduced
due to this process on the other hand, Al Yas beach which is located outside
Abu Dhabi Island was used as natural resource of beach soil. This area does not
undergo developmental changes and maintenance. Therefore, microorganisms would
be assumed to live in their natural habitats. Both environments are characterized
by hot weather, moisture and lack of rainfall during the summer. During winter,
the weather becomes cooler and drier with a small chance of rainfall. </p>

<figure>
<img src="IJEPR%20012_files/image001.jpg" class="article-img">

<figcaption>
Figure 1. Abu Dhabi city Map (Abu Dhabi Geospatial Portal, n.d</figcaption>
</figure>



<p class="subhead">2.2. Sample collection and field measurements</p>

<p class="body">Beaches were visited 2 times
at 1-2 weeks intervals from November to December 2012 (Wheeler Alm., et. al.,
2003). Sampling occurred over a 3 hour period between 12:00 pm and 3:00 pm (UAE
local time) to control the differences in solar insulation [32]. The number of
humans within 100 m during the sampling period was recorded. Also, air and soil
temperature; and humidity were measured to set the ambient temperature [15]. General
observations of the site were recorded, and pictures were taken for
documentation. Sieves, gloves and sample containers were used to collect the soil
samples from the beach. Wet (collected from the swash zone) and dry (collected
away from swash zone) soil samples were collected from each site and the
distance between the transects was recorded [2, 9]. This was done in order to
compare the microbial profile of wet and dry soil samples in the same beach. Soil
was collected from different depths (5 and 15 cm below the surface) by using
manual soil-coring devices in order to preserve the integrity of the inhabiting
microorganisms. This allowed collection of soil without mixing and hence
measurement of each depth separately. Three sub-samples were collected for each
sampling criteria to represent the beach as a whole. Those sub-samples were taken
to ensure proper representation of the microbial profile of the whole beach. All
collected and transported samples were placed in sterile; dry and cold
environment (around 4 &deg;C) to avoid contamination and further growth of
inhabiting microorganisms. In addition, deep freezing was done to stabilize the
microbial DNA used later for PCR. </p>

<p class="subhead">2.3. Sample Analysis Strategies</p>

<p class="subhead"><i>2.3.1. Cultivation Analysis</i></p>

<p class="body">General procedures of
detecting microorganisms were followed using cultivation methods. A total of 16
samples were collected from both beaches depending on different parameters like
water content and depth of soil (cm). Water content depended on distance from
sea water and varied from 3-10m for wet soil and 20-24m for dry soil. The depth
of the soil sample was fixed for 5 cm and 15 cm. Control samples of <i>enterococci</i>, yeast, <i>faecal coliforms, Escherichia
coli, Staphylococcus</i>
<i>aureus, </i>were inoculated in all types
of growth media used. <i>Enterococci,
Escherichia coli, and Staphylococcus aureus</i> were cultivated in MacConkey, EMB, Mannitol
Salt Agar (MSA) and then incubated for 18-72 h at 30-35 &deg;C [11]. Yeast and
fungi control samples for Malt Salt and Sabouraud Dextrose Agar (SDA) were retrieved
from the University of ADELAIDE mycology online database. Negative controls
were made for all growth media and left for few days to test for growth. All
media were prepared as per the manufacturer's instruction and sterilized.</p>

<p class="body">Cultivation of soil samples
followed the previously established method by Sabino, et. al., [30] with some
modification depending on experimental circumstances. Bacteriological analysis
started with having 6 tubes each containing 1 g of the soil sample it was was
diluted in 2 mL of Luria broth and, agitated for 6 min at 500 rpm. Two
different concentrations (0.25 mL and 1 mL) were later taken using sterilized
pipettes, and poured each in separate MacConkey growth media. This step was repeated
for all other growth media such as EMB, MSA, Malt Extract and SDA. Sterilized spreaders
were then used to spread the liquid in the growth plates which were incubated at
37 &deg;C for 24-72 hour. All bacterial, yeast and fungal growth analysis was reported
as colony forming units (CFU) per g of soil. All the physical characteristics
of the colonies and growth media were checked and noted. </p>

<p class="subhead"><i>2.3.2. Electrophoresis-based Polymerase
Chain Reaction </i></p>

<p class="body">Polymerase Chain Reaction (PCR)
was used for more specific identification of some microorganisms based on the
detection of unique gene sequences [22]. Preparation of PCR sample solution
from beach soil samples followed TAKARA manual [34] for the bacterial screening
PCR kit protocol 1.B. by preparing a heat extracted sample directly from
bacteria. A small amount of the bacteria from the colony was extracted and
suspended in 100 &micro;L volume of Chelex solution. Then the mixture was heated at
99 &deg;C for 5 minutes, this was quickly followed by cooling down by placing on
ice for 1 minute or more. The mixture was later centrifuged at 12000 rpm for 1
minute. The centrifuged supernatant was used as the DNA sample solution for
PCR. </p>

<p class="body">PCR analysis of <i>enterococci, Escherichia coli </i>and <i>Salmonella</i> was performed according to
the TAKARA manual [34]. The 2X premix solution, primer mix ENT, DNA sample
solution were all mixed together for analysis. The Negative controls were also
subjected to PCR using the specific primers and dH<sub>2</sub>O instead of DNA
sample solution. Moreover, positive controls were amplified as per the
instructions specified by the kit. ENT primers were used to identify <i>Enterobacteriaceae</i> family including <i>E. coli</i>. PCR primers were used for
targeting specific <i>Enterobacteriaceae</i> 16S rRNA gene markers<i> </i>(Figure1). Thermal cycler (PXE 0.2
Thermal Cycler) was used for amplification, and the PCR parameters were as
such: 95&deg;C for 60 seconds, then 95&deg;C,59&deg;C, 72&deg;C all for 30 seconds, finally 72&deg;C,
4&deg;C respectively for 60 seconds. </p>

<figure>
5'-AGAGTTTGATCCTGGCTCAG-3'

<figcaption>
Figure 2. ENT
16S rRNA primer</figcaption>
</figure>


<p class="body">Following PCR, gel electrophoresis
was used to identify the bands specific for each species family or genus.
Agarose and TBE buffer preparation was according to the TAKARA manual [34] for
bacterial screening PCR kit. In this experiment, 15 &micro;l of PCR amplicons were added
to 3 &micro;L of loading buffer. Promega 100bp DNA Ladder was used as a marker to indicate
the size of DNA bands. A 1.8% agarose gel was run at 65 V for 40 minutes and then
stained in ethidium bromide and finally placed under UV transilluminator. A band
of approximately 420 bp evident with the ENT primers indicated that the samples
contained a member of the <i>Enterobacteriaceae</i> family including <i>E. Coli. </i></p>

<h4>3. Results</h4>

<p class="subhead">3. 1. Cultivation Analysis</p>

<p class="body">Yas beach is a natural beach
located outside Abu Dhabi Island and is considered as the control beach for
this study. The average ambient temperature of the two trials was 31&deg;C and average
humidity was 60%. Results from the 2 trials for Yas beach showed a mean growth)
value of 11.71&plusmn; 13.81CFU/g of soil. Moreover, soil samples from this beach showed
growth in Mannitol Salt Agar (MSA) which was indicative of the presence of <i>Staphylococcus aureus</i> and other <i>Staphylococci</i>. Growth on MacConkey medium indicated
the possible presence of <i>Pseudomonas
aeruginosa, Shigella, Enterobacter and Klebsiella</i> in Yas beach soil. Fungal and yeast
cultivation results revealed the likely growth of <i>Candida guilliermondii, Candida
parapsilosis, Trichophyton equinum, Trichophyton terrestre</i>, <i>Malbranchea sp.</i>,<i> Aspergillus fumigates, </i><i>Trichophyton schoenleinii
Acremonium sp. </i>and<i>
Apophysomyces elegans </i>(Table
1). </p>

<center>
<div class="widetable">
  <p class="body">Table 1. Average
Microbial Growth (CFU/g) for Yas Beach Soil Samples (N=2)</p>
<table>
<thead>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray" colspan="2"><b>Criteria</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Mannitol Salt</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>MacConkey</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>EMB</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Sabouraud Dextrose</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Malt Salt</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray"><b>Depth</b></td>
  <td style="padding:5px; border:2px solid gray">Water content</td>
 </tr>
</thead>
<tbody>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray" rowspan="3"><b>5 cm</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray">Dry</td>
  <td style="padding:5px; border:2px solid gray">4</td>
  <td style="padding:5px; border:2px solid gray">31</td>
  <td style="padding:5px; border:2px solid gray">2</td>
  <td style="padding:5px; border:2px solid gray">1</td>
  <td style="padding:5px; border:2px solid gray">3</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray">Wet</td>
  <td style="padding:5px; border:2px solid gray">7</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">8</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray" rowspan="3"><b>15 cm</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray">Dry</td>
  <td style="padding:5px; border:2px solid gray">2</td>
  <td style="padding:5px; border:2px solid gray">2</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">1</td>
  <td style="padding:5px; border:2px solid gray">-</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray">Wet</td>
  <td style="padding:5px; border:2px solid gray">5</td>
  <td style="padding:5px; border:2px solid gray">33</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">38</td>
  <td style="padding:5px; border:2px solid gray">27</td>
 </tr>
</tbody>
</table>
</div>
</center>


<p class="body"> Dhabi Corniche beach which
is a man-made beach is located on the north-western shore of Abu Dhabi island.
The average ambient temperature of the two trials was 27 &deg;C and average humidity
was 60%. Results from the examination of 8 samples of soil (both wet and dry)
from the Corniche beach showed a mean growth value of 21.57 &plusmn; 27.73 CFU/g of
soil. It revealed growth with a total of 47.5%. MacConkey and MSA plates revealed
significant growth (CFU/g) especially for wet soil. The minimum growth average value
was 7 CFU/g of soil while the maximum value was 38 CFU/g of soil. <i>Staphylococcus aureus,</i> <i>Pseudomonas aeruginosa, Shigella, </i><i>Enterobacter, </i><i>Salmonella </i><i>and E. coli </i>all potentially existed in
Corniche beach soil<i>.
</i>For yeast and
fungi, the possibilities were <i>Malbranchea sp.</i>, <i>Aspergillus
niger</i>, <i>Trichophyton equinum, Paecilomyces
variotii,</i> <i>Aspergillus carbonarius </i>and <i>Candida glabrata </i>(Table 2). </p>


<center>
<div class="widetable">
  <p class="body">Table 2. Average Microbial Growth (CFU/g) for Corniche Beach Soil Samples (N= 2)</p>
<table>
<thead>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray" colspan="2"><b>Criteria</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Mannitol Salt</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>MacConkey</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>EMB</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Sabouraud Dextrose</b></td>
  <td style="padding:5px; border:2px solid gray" rowspan="2"><b>Malt Salt</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray"><b>Depth</b></td>
  <td style="padding:5px; border:2px solid gray">Water content</td>
 </tr>
</thead>
<tbody>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray" rowspan="3"><b>5 cm</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray">Dry</td>
  <td style="padding:5px; border:2px solid gray">13</td>
  <td style="padding:5px; border:2px solid gray">5</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">15</td>
  <td style="padding:5px; border:2px solid gray">4</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray">Wet</td>
  <td style="padding:5px; border:2px solid gray">40</td>
  <td style="padding:5px; border:2px solid gray">88</td>
  <td style="padding:5px; border:2px solid gray">3</td>
  <td style="padding:5px; border:2px solid gray">25</td>
  <td style="padding:5px; border:2px solid gray">2</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray" rowspan="3"><b>15 cm</b></td>
 </tr>
 <tr align="center">
  <td style="padding:5px; border:2px solid gray">Dry</td>
  <td style="padding:5px; border:2px solid gray">3</td>
  <td style="padding:5px; border:2px solid gray">64</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">1</td>
 </tr>
  <tr align="center">
  <td style="padding:5px; border:2px solid gray">Wet</td>
  <td style="padding:5px; border:2px solid gray">70</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">-</td>
  <td style="padding:5px; border:2px solid gray">38</td>
 </tr>
</tbody>
</table>
</div>
</center>


<p class="body">The average growth (CFU/g) for
wet and dry soil isolates from Corniche beach (38 &plusmn; 32.19 CFU/g and 6.00 &plusmn; 5.69
CFU/g, respectively) was 10-60% higher than that for isolates from Yas beach
(13.00 &plusmn; 14.69 CFU/g and 5.75&plusmn; 10.25 CFU/g, respectively, figure 3). Moreover,
the average growth (CFU/g) for isolates for 5 and 15 cm depths from Corniche
beach (21.6 &plusmn; 27.82 CFU/g and 35.2 &plusmn; 32.61 CFU/g, respectively) was 50-70% higher
than that for isolates from Yas beach (8 &plusmn; 10.46 CFU/g and 15.43 &plusmn; 16.48 CFU/g,
respectively, figure 4).</p>


<figure>
<img src="IJEPR%20012_files/image002.gif" class="article-img">

<figcaption>
Figure 3.
Average Microbial Growth (CFU per g of soil g) for Samples from Corniche and
Yas Beaches According to Distance from Swash Zone (N = 16)</figcaption>
</figure>


<figure>
<img src="IJEPR%20012_files/image003.jpg" class="article-img">

<figcaption>
Figure 4.
Average Microbial Growth (CFU per g of soil g) for Samples from Corniche and
Yas Beaches According to Depth from Soil Surface (N = 16)</figcaption>
</figure>



<p class="subhead">3. 2. Electrophoresis-based Polymerase
Chain Reaction (PCR) </p>

<p class="body">The PCR analysis was performed to confirm the
presence of <i>Enterobacteriaceae</i> family strains in soil
samples from either beach. Samples used for PCR and electrophoresis were all
gram negative bacteria from MacConkey and EMB growth media. Figure 2 shows the
matching in size for the positive control in lane 8 and samples from MacConkey
plate in lane 5. The results showed positive growth for bacteria isolated from MacConkey
plate from Corniche beach soil samples indicating the presence of <i>E. coli, salmonella and coliforms</i> in the soil. However, the
other sample from MacConkey from Yas beach showed a negative result for <i>coliforms</i> using the same ENT primers
which means that other gram negative bacteria present. Moreover, in another
electrophoresis gel, wet
sample from the Yas beach (MacConkey, 15 cm depth, 1 ml concentration)
demonstrated also negative result for coliforms using the same ENT primers.
This also suggested that other gram negative bacteria (but not <i>E. coli</i>) could be present.<i> </i></p>

<figure>
<img src="IJEPR%20012_files/image004.jpg" class="article-img">

<figcaption>
Figure 5. Results from Agarose gel electrophoresis Analysis
1. Lane 1: 100bp DNA Ladder (Promega, USA) 2. Lane 2: <i>E. coli</i> positive control 3. Lane 3: MacConkey growth sample from Yas beach (dry soil) 4. Lane 4: MacConkey growth sample from Corniche beach (dry soil) 5. Lane 5: MacConkey growth sample from Corniche beach (wet soil) 6. Lane 6: EMB growth sample from Corniche beach (wet soil) 7. Lane 7: EMB growth sample from Corniche beach (wet soil) 8. Lane 8: ENT positive control</figcaption>
</figure>



<h4>4. Discussion</h4>

<p class="body">Results of the study showed
some similarities between bacteria that inhabited both man-made and natural
beaches. The most frequent gram positive isolates were species of <i>Staphylococcus</i><i>,</i> whereas gram-negative genera from both
beaches were <i>Pseudomonas</i>,<i> Enterobacterriaeae sp. </i>The only difference between
the two beaches was with regards to <i>E. coli</i> which was detected in all samples from Corniche
beach but only in one sample from Yas beach. <i>E. coli</i> could have inhabited the soil from human
skin, infection or animal. In
a study done by Anderson et al., [3], similar findings were reported about <i>E.coli</i> and the reason was suggested
to be the increase of beach users. In Corrniche beach there was at least 1 to 2
humans each 5m<sup>3</sup> whereas in Yas beach there was none. Moreover,
growth of the algae <i>Cladophora</i> could have provided a
favourable environment for the stability and growth of <i>E. coli</i> [10]. This could be a
reason for the greater growth of <i>E. coli</i>
in Corniche beach. An additional cause could be related to the source of the
soil itself. <i>E.
coli</i> could be
considered a health risk to beach users. Moreover, <i>Staphylococcus sp.</i> outnumbered other bacteria in
all soil samples. Dowidart &amp; Abdel-Monem [8] found that <i>Staphylococcus sp.</i> predominate over other flora
in the sand. Classification of the study indicated a total of 20-30% growth
for <i>S. aureus</i>, 70-80% growth <i>for</i> other<i> Staphylococcus sp. </i>Similarly, a study done by
Prado et al., [27] on two popular beaches in Chile revealed 24% growth as <i>S. aureus</i> and 76% as other <i>Staphylococcus sp.</i> The presence of these
bacteria could be related to human activity. Corniche beach showed more CFUs
for <i>Staphylococcus
sp.</i> as it had
more frequent human activity as compared to Yas beach. Papadakis et al., [26] demonstrated
that the presence of<i>
S. aureus</i>
correlated with the presence of yeasts of human origin in soil samples. This
was related to this study findings as yeast was isolated from samples taken
from both Corniche and Yas beaches. </p>

<p class="body">Regarding results of yeast and
fungal growth, yeast like<i>
Candida</i> sp.
were found in 1.25% of samples from both beaches, whereas <i>Candida albicans</i> was not detected at all.
Other researchers also had reported the absence or low incidence of <i>C. albicans</i> [28, 12, 4]. Furthermore, in
this study all yeast from both beaches were <i>Candida</i> species which is comparable to results
found in a research study done by Sabino, et. al. [30] which demonstrated that
67.5% of the detected yeast were Candida species. Other fungi were <i>C. guilliermondii</i> and <i>C. parapsilosis</i>; which were also isolated by
other researchers from seashore soil [4]. Moreover, comparison of the fungal
profile growth in media plates between man-made (4%) and natural beach (6%)
indicated that the man-made beach had less growth than natural beach. This
could be attributed to the low level of human activity and soil maintenance in
Yas beach (natural) in contrast to Corniche beach (man-made) which is subjected
to constant maintenance and has laws to prohibit the presence of animals.<i> </i>Related to the safe CFU/g threshold,
based on Sabino, et. al. [30] study, 15 CFU/g, is the calculated threshold for the
yeasts. In this study, all the isolated yeast showed less than 15 CFU/g,
therefore, would not be considered as a health risk for beach users. </p>

<p class="body"> Potentially pathogenic
fungi such as <i>Aspergillus
sp.</i> were
found in more abundance in wet soil samples as compared to dry soil samples probably
because fungi have
higher resistance to solar exposure compared to bacteria [3]. A study by Izquierdo et al., [16] revealed that
saprophytic fungi such as <i>A.
Fumigates</i>, <i>A. Candidus</i> and <i>A. ochraceus</i> were isolated from wet soil areas
of the beach in high tidal conditions. Whereas in other investigations such as that of Sabino,
et.al., [30] these fungi, were found in dry soil with high average growth of
CFU/g than what was found in our study for Yas and Corniche soil beaches. Furthermore,
<i>Trichophyton
sp.</i> showed higher
growth percentage than yeast and bacteria and was detected in all the samples
collected. Salvo, &amp; Fabiano, 2007 in
Italy, Ulfig, et al., 1997 in Spain, Vidal, et al., 1966 in Sweden and Sabino et. al. in Portugal all showed the same conclusions
for this species. Also, Harris
et al. 1995; Bater 1996; Van der Heijden et al. 1998; Caruso et al. 2008 indicated
that the growth
of fungi could be caused by the
extinction of the native species [14].
Hence, this could be a reason for the disappearance of some significant species
from beach soil such as <i>E.
coli</i> in Yas
beach. It has
been said by Sabino,
et. al. in 2011 that 17 CFU/g is the threshold for potentially pathogenic
fungi, and 8 CFU/g for dermatophytes. In comparison, both beaches (man-made
and natural) had higher number of CFU/g of soil hence suggesting a potential health
risk for human health. More
studies are needed however to confirm or generalize this finding. </p>

<p class="body"> The hypothesis of the study
suggested that the growth of microorganisms in man-made beach soil would be higher
as compared to natural beach soil. Results of the study showed a high average
growth CFU/g for Corniche beach for both parameters (water content and depth
from the soil surface). The
results demonstrated that average growth was higher in wet soil as compared to dry
soil. This suggested the existence of a correlation between water content and
growth of bacteria in soil. Our study is supported by two other studies which showed
the same results. Su, Jianping. et. al., 2004, found that wetlands showed more
abundant microbial population as compared to dry lands. Also, Kaneko and Atlas,
[17] found that the density of bacterial communities positively correlated with
the water content in the soil. Therefore,
the studied man-made beach has both similarities and differences compared to natural
beaches with some differences the natural beach of the study. Many reasons support this finding
as one of the microbial growth features is the availability of water content [24]. Papadakis et al., [26] stated
that the wet soil is enriched with organic substances which provide a harbor
environment for enteric bacteria allowing it to survive longer than in other
areas. The natural beach of the study might have been affected by the
surrounding construction which might have in turn affected the microbial
population living there. However, a study performed by Brand&atilde;o et al., [5]
showed that the microbiological growth was higher in dry soil as compared to
wet soil. This could be due to the preferences of beach users who often enjoy
sitting in the dry soil more than the wet soil. </p>

<p class="body"> The other parameter used in
this investigation between the man-made beach and the natural beach was the depth
of the soil from the surface. The finding of this study showed a higher average
growth of microorganisms in the depth of around 15 cm. That finding is
supported by a study by Papadakis et al., [26] which stated that enteric
bacteria survive for short duration on the surface of dry soil as it gets
destroyed mostly by environmental pressure. Therefore, its favorable conditions
are in deeper layers of soil. Also, it was stated by Othman, 2003 that
variations in temperature of the soil occur in surfaces and deeper depths. Soil
in deep depth does not exhibit very dramatic daily and seasonal variations in
temperature, so growth of microorganisms, gets more abundant and stable [13]. However,
the only exception was that <i>E.
coli</i> was
isolated only from the upper 5 cm of soil in this study, while growth in the
depth of 15 cm was not detected. This could be related to the need of this particular
species to sunlight and oxygen which are found abundantly in the upper layers
of soil. This
finding is supported by the Kleinheinz, [19] and Desmarais, et. al., [7] who demonstrated
that with deeper
depth, microorganisms become less abundant. </p>

<p class="body">PCR results demonstrated the presence
of <i>E. coli,
Salmonella and coliforms</i>
in Corniche beach but not in Yas beach for the same species. The presence of members of the
<i>Enterobacteriaeae</i> species could indicate recent
faecal pollution [14]
Therefore, this could be an indication of some pollution in the Corniche beach.
A study by Hafez, E, &amp; Elbestawy, E.,
[14] demonstrated
that the dominant gram negative bacteria in the soil of a number of beaches
they assessed in their study were identified as <i>Shigella flexneri</i> and <i>Escherichia sp.</i> using 16S rRNA primers. However,
direct extraction from soil was used in the previous study while in this study
manual DNA extraction from culture was utilized. PCR was used in the
identification of human enteric pathogens in two South-western Lake Michigan
recreational beaches. The results indicated the presence of <i>Salmonella </i>(0.7% of isolates) and <i>Campylobacter</i> (22.7%) [18]. Our results
similarly revealed that <i>Salmonella,
which is potentially pathogenic to human health,</i> inhabited Corniche beach soil. </p>


<h4>5. Conclusion </h4>

<p class="body">In conclusion, the man-made beach
(Corniche beach) showed a higher microbial growth in terms of water content and
depth from the soil surface as compared to the natural beach (Yas beach). Furthermore,
<i>E. coli </i>and <i>coliforms </i>had been detected<i> </i>in Corniche beach only. So regardless of
the great effort invested for maintaining a good soil quality in man-made
beach, there could be some factors which allow microbes to grow and thrive in the
soil of such beaches. Even if these microorganisms have been introduced
unnaturally to the beach by different means including the collection of soil from
various locations and contamination from the skin/infection of humans and
animals, they could adapt to the new environment with time and become part of
the ecological soil diversity. Future research studies could investigate
whether there could be differences within the soil microbial diversity over a
longer period of time taking into consideration the seasonal and environmental
changes. </p>


<h4>Acknowledgement</h4>

<p class="body">Especial thanks to the NSPH
Department at Zayed University for the help and support throughout the study.</p>

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