<!DOCTYPE html>
<html xmlns="http://www.w3.org/1999/xhtml">
	<head>
		<meta charset="utf-8" />
		<title>AAVS_MH20150609100653_ Goharrizi et al</title>
		<link href="http://nexusacademicpublishers.com/uploads/cssfiles/AAVS_MH20150609100653_ Goharrizi et al.css" rel="stylesheet" type="text/css" />
	</head>
    <style> 
.pt{
	background-color:#BDF;
	border-top-style:solid;
	
	border-top-color:#d3d1d1;
	border-top-width: 1px;
	border-bottom-style: solid;
	
	border-bottom-color:#d3d1d1;
	border-bottom-width: 1px;
	height:90px;
	}
	  
.img_display{
	float:left;
	padding:5px;
	
	}
	.img_text{ padding:10px;
		
		}
		
		.Txt_Dis{
			padding-right:20px;
			}
</style>

<body id="AAVS_MH20150609100653_-Goharrizi-et-al" xml:lang="en-GB">
		<div class="Basic-Text-Frame">
			<p class="Type-of-Article">&nbsp;</p>
			<p class="Type-of-Article">&nbsp;</p>
			<p class="Type-of-Article">Research Article</p>
		  <p class="Type-of-Article">&nbsp;</p>
    </div>
        <!-- Start CrossMark Snippet v1.5 -->
<meta name="dc.identifier" content="doi:10.14737/journal.aavs/2015/3.7.406.412">
<script type="text/javascript" src="http://smithandfranklin.com/js/jquery.min.js" ></script>
<script type="text/javascript" src="http://smithandfranklin.com/js/jquery-ui.min.js" ></script>
<script type="text/javascript" src="https://crossmark.crossref.org/javascripts/v1.5/crossmark.min.js"></script>
              <a id="open-crossmark" href="javascript:" style="padding: 3px 0 13px 0; float:right;"><img style="border: 0; display: none;" id="crossmark-icon" src="https://crossmark.crossref.org/images/crossmark_button.png" /></a>
              <div id="crossmark-dialog"  title="">
              <!-- the external CrossMark data is loaded inside this iframe -->
              <iframe id="crossmark-dialog-frame" frameborder="0"></iframe>
              </div>
			<!-- End CrossMark Snippet -->

<div class="Basic-Text-Frame">
			<p class="title- ParaOverride-1">&nbsp;</p>
			<p class="title- ParaOverride-1">The Study on Effect of Temperature Stress on Occurrence of Clinical Signs Caused by <i>Aeromonas hydrophila</i> in <i>Capoeta damascina</i> in <i>In Vitro </i>Condition</p>
		</div>
		<div class="Basic-Text-Frame">
			<p class="Authors ParaOverride-1">&nbsp;</p>
			<p class="Authors ParaOverride-1"><span class="CharOverride-2">Laleh Yazdanpanah Goharrizi</span><span class="CharOverride-3">1*</span><span class="CharOverride-2">, Mohammad Ebrahim Jalil Zorriehzahra</span><span class="CharOverride-3">2</span><span class="CharOverride-2">, Milad Adel</span><span class="CharOverride-3">3</span></p>
		</div>
		<div class="Basic-Text-Frame">
			<p class="Affiliations ParaOverride-1"><span class="CharOverride-4">1</span>Animal Sciences Dept., Kerman Agricultural and Natural Resources Research Center, Agricultural Research Education and Extension Organization (AREEO), Kerman; <span class="CharOverride-4">2</span>Aquatic Animal Health &amp; Diseases Dept., Iranian Fisheries Sciences Research Institute (IFSRI); <span class="CharOverride-4">3</span>Aquatic Animal Health and Diseases Dept., Caspian Sea Ecology Research Center, Iranian Fisheries Science Research Institute (IFSRI), Agricultural Research Education and Extension Organization (AREEO),Tehran, I.R. Iran. </p>
		</div>
		<div>
			<p class="Abstract ParaOverride-1">&nbsp;</p>
		  <p class="Abstract ParaOverride-1"><span class="CharOverride-5">Abstract</span> | <span class="CharOverride-6">Aeromonas hydrophila</span> is one of the most important microbial pathogens affecting several fish species worldwide and economical loss of related mortality should be considered in world aquaculture industry. In this study, the effects of temperature stress on occurrence of clinical signs caused <span class="CharOverride-6">A. hydrophila </span>on <span class="CharOverride-6">Capoeta damascina </span>was carried out. For this purpose, 240 <span class="CharOverride-6">C. damascina</span> were captured from Baft’s rivers in Kerman province in spring season of 2014 and transported to central laboratory. After adaptation period in first stage, 0.1 ml of 3× 10<span class="CharOverride-7">8</span> cfu/ml<span class="CharOverride-7"> </span>of <span class="CharOverride-6">A. hydrophila </span>was injected intraperitoneal to each fish. Then, fish were divided into 4 groups (0-10, 10 - 20, 20 - 30 and to over 30<span class="CharOverride-7">o</span>C). After 2 weeks, clinical signs and mortality of each group were recorded. The results revealed that water temperatures can play an important role to pathogenicity of <span class="CharOverride-6">A.hydrophila</span> on <span class="CharOverride-6">C. damascina</span>. Most clinical signs and mortality were observed in 20-30<span class="CharOverride-7">o</span>C and above groups, although, at 10-20<span class="CharOverride-7">o</span>C and below 10<span class="CharOverride-7">o</span>C only 40% and 1% of fish revealed typical clinical signs, respectively.</p>
		  <p class="Abstract ParaOverride-1">&nbsp;</p>
			<p class="Abstract ParaOverride-1"><span class="CharOverride-5">Keywords</span> | Iran, <span class="CharOverride-6" xml:lang="en-GB">Aeromonas hydrophila</span>, <span class="CharOverride-6" xml:lang="en-GB">Capoeta damascina</span>, Temperature stress</p>
		  <p class="Abstract ParaOverride-1">&nbsp;</p>
		</div>
		<div class="Basic-Text-Frame">
			<p class="Editor----Citation"><span class="CharOverride-5">Editor</span> | Kuldeep Dhama, Indian Veterinary Research Institute, Uttar Pradesh, India.</p>
			<p class="Editor----Citation"><span class="CharOverride-5">Received</span> | April 28, 2015; <span class="CharOverride-5">Revised</span> | June 09, 2015; <span class="CharOverride-5">Accepted</span> | June 10, 2015; <span class="CharOverride-5">Published</span> | June 19, 2015&#9;&#9;</p>
			<p class="Editor----Citation"><span class="CharOverride-5">*Correspondence</span> | Laleh Yazdanpanah Goharrizi, Agricultural Research Education and Extension Organization (AREEO), Kerman, I.R. Iran; <span class="CharOverride-5">Email:</span> zorrieh@yahoo.com</p>
			<p class="Editor----Citation"><span class="CharOverride-5">Citation</span> | Goharrizi LY, Zorriehzahra MEJ, Adel M (2015). The study on effect of temperature stress on occurrence of clinical signs caused by <span class="CharOverride-18">Aeromonas hydrophila</span> in <span class="CharOverride-18">Capoeta damascina</span> in <span class="CharOverride-18">in vitro</span> condition. Adv. Anim. Vet. Sci. 3(7): 406-412.  </p>
			<p class="Editor----Citation"><span class="CharOverride-5">DOI</span> | <a href="http://dx.doi.org/10.14737/journal.aavs/2015/3.7.406.412"><span class="Hyperlink">http://dx.doi.org/10.14737/journal.aavs/2015/3.7.406.412</span></a></p>
			<p class="Editor----Citation"><span class="Editor---Citation CharOverride-5">ISSN (Online)</span> | 2307-8316; <span class="Editor---Citation CharOverride-5">ISSN (Print)</span> | 2309-3331</p>
			<p class="Editor----Citation"><span class="CharOverride-5">Copyright</span> © 2015 Goharrizi et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.</p>
		</div>
		<div class="Basic-Text-Frame">
			<p class="Heading-1--Introduction----">&nbsp; </p>
		  <p class="Heading-1--Introduction----">Introduction</p>
			<p class="Caps-on-First-Para ParaOverride-1">&nbsp;</p>
			<p class="Caps-on-First-Para ParaOverride-1"><span class="_idGenDropcap-1">A</span><span class="CharOverride-6">eromonas </span><span class="CharOverride-6">hydrophila</span> is a ubiquitous gram-negative, motile and rod-shaped bacterium with wide distribution in aquatic habitats (<a href="#Krovacek-K--Pasquale-V--Baloda-SB--Soprano-V--Conte-M--Dumontet-S--1994"><span class="Hyperlink">Krovacek et al., 1994</span></a>; <a href="#Kersters-I--Van-Vooren-L--Huys-G--Janssen-P--Kersters-K--Verstraet-W--1995"><span class="Hyperlink">Kerster et al., 1995</span></a>; <a href="#Maalej-S--Denis-M--Dunkan-S--2004-"><span class="Hyperlink">Maalej et al., 2004</span></a>). <span class="CharOverride-6">A. hydrophila</span> is an opportunist human pathogen which is widely distributed in aquatic environments (<a href="#Rahman-MH--Suzuki-S--Kawai-K--2001"><span class="Hyperlink">Rahman et al., 2001</span></a>). This bacterium commonly isolated from fresh water ponds and is also a normal inhabitant of the gastrointestinal tract of fish.&#160;<span class="CharOverride-6">Aeromonas </span>species in aquatic environments is hazardous, especially for fish and shellfish species as it was the causative agent of several infectious diseases in aquaculture (<a href="#Santos-JA--Gonza-lez-CJ--Otero-A--Garc--a-Lo-pez-ML--1999"><span class="Hyperlink">Santos et al., 1999</span></a>). Some strains of <span class="CharOverride-6">Aeromonas </span>are more virulent, which means that they possess special properties which enable them to cause more serious disease outbreaks (<a href="#Bullock-GL--1968-"><span class="Hyperlink">Bullock, 1968</span></a>). If these more damaging strains become endemic in a population of fish (which means that they are there all of the time and the fish develop an immunity to them), it becomes difficult to introduce new fish into the water body without suffering major losses of newly-stocked fish (<a href="#Colwell-RR--2000-"><span class="Hyperlink">Colwell, 2000</span></a>).</p>
		  <p class="Caps-on-First-Para ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">Motile <span class="CharOverride-6">Aeromonas</span> Septicemia (MAS) is a common bacterial disease, caused by <span class="CharOverride-6">Aeromonas</span>, which affects warm water fish, both in commercial production systems and in natural waters (<a href="#Austin-B--Stobie-DA--1993-"><span class="Hyperlink">Austin and Stobie, 1993</span></a>). Frequently, MAS outbreaks are stress-related and elimination of the underlying stress factor may be sufficient to resolve the disease outbreak (<a href="#Shelton-JL--2000-"><span class="Hyperlink">Shelton, 2000</span></a>). </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">The disease primarily affects freshwater fish such as catfish, bass, and many species of tropical or ornamental fish. Fish infected with <span class="CharOverride-6">A. hydrophila</span> may have many dif&#173;ferent clinical signs (<a href="#Chakrabaiti-NM--2006-"><span class="Hyperlink">Chakrabaiti, 2006</span></a>). These range from sudden death to in appetence, swimming abnormalities, pale gills, bloat and skin ulcerations. The skin ulcers may occur at any site on the fish and often they are surrounded by a bright red rim of tissue. Because of the variability of these clinical signs, the diagnosis of this disease based only upon the clinical presentation of the fish is highly unreliable and may be economically disastrous to the fish produc&#173;er with intensive fish farming systems, whether these systems are outdoor ponds or indoor aquaria and tanks, predisposing factors are primarily responsible for the precipitation of this disease (<a href="#Elbour-M--Attia-H--Elhilli--Mraouna--Ayari-W--2001"><span class="Hyperlink">Elbour et al., 2001</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text">Any time an <span class="CharOverride-6">Aeromonas</span> infection persists as a chronic problem, it is important to make efforts to determine whether an underlying stress factor causes the fish to have insufficient immune protection from the bacteria (<a href="#Chakrabaiti-NM--2006-"><span class="Hyperlink">Chakrabaiti, 2006</span></a>). </p>
			<p class="Body-Text ParaOverride-1">Several authors noted that the survival of <span class="CharOverride-6">A. hydrophila</span> in aquatic environment was inhibited by entrance into a viable but non-cultivatable (VBNC) state, due to stress induced by aquatic conditions for the bacterial cell, which remained metabolically activity, but unable to undergo the sustained cellular division required for growth in  different conditions (<a href="#Maalej-S--Denis-M--Dunkan-S--2004-"><span class="Hyperlink">Maalej et al., 2004</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text">Stress is the one of the most im&#173;portant predisposing factor associated with this disease. Common sources of stress are water temperature, water poor quality, overcrowding, or rough handling. Temperature is one of the most common environmental stressors in aquaculture ecosystems. Temperature is a stressor when it ranges to near the fish’s high, long-term tolerance level and when it fluctuates rapidly by more than a few degrees (e.g. 3 to 5°C in less than one hour), especially if temperature was increased. A relationship between changes in water temperature and the incidence of <span class="CharOverride-6">Aeromonas </span>spp. has been reported (<a href="#Kersters-I--Van-Vooren-L--Huys-G--Janssen-P--Kersters-K--Verstraet-W--1995"><span class="Hyperlink">Kersters et al., 1996</span></a>).</p>
		  <p class="Body-Text">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">The main objective of this study was to investigat the effects of temperature stress on occurrence of clinical signs of <span class="CharOverride-6">Aeromonas</span> infections in <span class="CharOverride-6">C. damascina</span> as native fish from rivers of Baft region in Kerman province, Iran. </p>
			<p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">Materials and Methods </p>
			<p class="Heading-2--History-in-MM-">&nbsp;</p>
		  <p class="Heading-2--History-in-MM-">Fish</p>
			<p class="Body-Text">About 240<span class="CharOverride-6"> C. damascina</span> were captured from a native river of Baft region in spring season of 2014 in Kerman province and shifted to central laboratory immediately.  In first stage, fish were acclimated to the condition of the laboratory for 7 days. In this time, fish were fed twice daily with commercial pellet (Mazan, Mazandaran). During the study, water temperature, pH and dissolved oxygen were daily measured as 18±2<span class="CharOverride-7">o</span>C, 7-8 and 5.2- 5.8ppm, respectively.</p>
		  <p class="Body-Text">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Isolation of <span class="CharOverride-1">A. hydrophila</span>  </p>
			<p class="Body-Text ParaOverride-1">From Baft River, about 25 infected <span class="CharOverride-6">C. damascina</span> were used for isolation of bacteria. Kidney sample were collected from infected fish and transferred to (tryptic soy agar) TSA medium and were then incubated for 48 h at 25<span class="CharOverride-7">o</span>C. After 48 h, <span class="CharOverride-6">A. hydrophila </span>were identified based on biochemical test (<a href="#Mohajeri-J--Afsharnasab-M--Jalali-B--Kakoolaki-S--Sharifrohani-M--Haghighi-A--2011"><span class="Hyperlink">Mohajeri et al., 2011</span></a>). Cell densities were photometrical obtained at 540 nm wavelength (<a href="#Saulnier-D--Haffner-P--Goarant-C--Levy-P--Ansquer-D--2000-"><span class="Hyperlink">Saulnier et al., 2000</span></a>), based on the Mcfarland standard and 3×10<span class="CharOverride-7">8</span> CFUmL<span class="CharOverride-7">-1</span> of bacteria was prepared. </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Challenge Assay</p>
			<p class="Body-Text ParaOverride-1">About 20 fish from each subgroup were randomly selected for the challenge assay. The challenge was by intraperitoneal injection with 0.1 ml<span class="CharOverride-7">−1 </span>suspensions of <span class="CharOverride-6">A. hydrophila</span> in 0.9% (w/v) saline containing 3×10<span class="CharOverride-7">8</span> CFUmL<span class="CharOverride-7">-1</span>.  </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Experimental Group </p>
			<p class="Body-Text ParaOverride-1">The study was designed as three treatments, 0-10, 10 - 20, 20 - 30 and to over 30<span class="CharOverride-7">o</span>C and control (30<span class="CharOverride-7">o</span>C) in triplicates. Twelve glass aquariums were divided to the groups. Also, 20 <span class="CharOverride-6">C. damascina</span> were kept to each aquarium for 2 weeks and clinical signs and mortality were recorded to each group. Aquarium heater was used to regular temperatures. The erosions of caudal and dorsal fins and kidney of moribund fish were selected as target tissues for isolation and confirmation of bacteria. Nutrient and Blood agar were used as primary media cultures. After separating and preparing the pure culture, some biochemical tests including: SIM, Nitrate, Indol, Motility, MR, VP, O/F, Oxidase, Urease, Maltase, Gelatinase, Hemolysin, Catalase, Esculine, Lysine and Arginine have been employed to characterization of  <span class="CharOverride-6">A. hydrophila</span>. </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Statistical Analysis</p>
			<p class="Body-Text ParaOverride-1">All the obtained data were subjected to statistical analysis using the SPSS software, version 18. The statistical analysis was done by using one-way analysis of variance (ANOVA) followed by Duncan’s multiple range tests. P-value of &lt;0.05 was considered significant.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Results</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">In this study, different temperature ranges were examined to survey its impacts on pathogenicity of <span class="CharOverride-6">A. hydrophila</span> in <span class="CharOverride-6">C. damascina</span>.<span class="CharOverride-6"> </span>The results of biochemical tests of<span class="CharOverride-6"> A. hydrophila</span> isolated from erosions of fins and internal organs of moribund fish were revealed in <a href="#Table-1-"><span class="Hyperlink">Table 1</span></a>. </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">In this study, clinical signs including: exophthalmia, blackening of the skin, abdominal distension, hemorrhages in the eyes, base of the fins or on the skin and internal organs such as liver (<a href="#Figure-1-"><span class="Hyperlink">Figure 1</span></a>), kidney and spleen were observed. Internal signs were included fluid in the abdomen, swollen liver and spleen, fluid-filled and distended intestines.  The results revealed that water temperatures play a major role to pathogenicity of <span class="CharOverride-6">A. hydrophila</span> in <span class="CharOverride-6">C. damascina</span>. Most clinical signs and mortality were observed in 20-30<span class="CharOverride-7">o</span>C and above groups, although, at 10-20<span class="CharOverride-7">o</span>C and below 10<span class="CharOverride-7">o</span>C only 40% and 1% of fish showed clinical signs, respectively (<a href="#Figure-2-"><span class="Hyperlink">Figure 2</span></a>, <a href="#Table-2-"><span class="Hyperlink">Table 2</span></a> and <a href="#Table-3-"><span class="Hyperlink">3</span></a>).  </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Table-1-"></a>Table 1: </span>The result of biochemical test for bacterial isolation (<span class="CharOverride-6">Aeromonas hydrophila)</span></p>
			<table width="659" height="232" class="Table-Style-1" id="table-1">
				<colgroup>
					<col />
					<col />
					<col />
					<col />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Gram</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">-</span></p>
						</td>
						<td />
						<td />
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Nitrate</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">NO</span><span class="CharOverride-12">2</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Urease</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">- </span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">SIM</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">- or +</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Maltase</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">H2S</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Gelatinase</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Indol</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">TSI</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">A/A</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Motility</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Hemolysin</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">MR</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">-</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Catalase</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">VP</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Esculine</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">O/F</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Lysine</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+SH2</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Oxidase</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">+</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Arginine</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">- </span></p>
						</td>
					</tr>
				</tbody>
			</table>
		  <p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
		  <p class="Figure--and-Table-Heading ParaOverride-1"><a id="Table-2-"></a><span class="CharOverride-5">Table 2:</span> Variance analysis of infected fish with clinical signs of <span class="CharOverride-6">Aeromonas hydrophila</span> in <span class="CharOverride-6">Capoeta damascina</span></p>
			<table width="658" height="94" class="Table-Style-1" id="table-2">
				<colgroup>
					<col />
					<col />
					<col />
					<col />
					<col />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Sources                        alteration</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Degree of freedom    (df)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Sum squares</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Mean Squares</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">F</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Treatment</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">3</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">329/67</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">109/89</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">0</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Error</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">8</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">5/33</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">0/667</span></p>
						</td>
						<td />
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">Total</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">11</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">335</span></p>
						</td>
						<td />
						<td />
					</tr>
				</tbody>
			</table>
			<p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
		  <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Table-3-"></a>Table 3:</span> Mean Comparisons with Duncan’s multiple range test  </p>
			<table width="659" height="50" class="Table-Style-1" id="table-3">
				<colgroup>
					<col />
					<col />
					<col />
					<col />
					<col />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-1">
						<td width="107" height="21">
							<p class="Basic-Paragraph"><span class="CharOverride-13">Temperature</span></p>
						</td>
						<td width="139">
							<p class="Basic-Paragraph"><span class="CharOverride-13">T1  </span><span class="CharOverride-13">(0-10</span><span class="CharOverride-13">°C)</span></p>
						</td>
						<td width="120">
							<p class="Basic-Paragraph"><span class="CharOverride-13">T2 </span><span class="CharOverride-13">(10-20°C)</span></p>
						</td>
						<td width="120">
							<p class="Basic-Paragraph"><span class="CharOverride-13">T3 </span><span class="CharOverride-13">(20-30°C)</span></p>
						</td>
						<td width="149">
							<p class="Basic-Paragraph"><span class="CharOverride-13">T4 </span><span class="CharOverride-13">(above30°C)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td height="21">
							<p class="Basic-Paragraph"><span class="CharOverride-11">Average</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">0.67</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">7</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">12</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">14.33</span></p>
						</td>
					</tr>
				</tbody>
			</table>
			<p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
			<p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-14">*Means in rows with different superscripts are significantly different at the level of (P&lt;0.05).</span></p>
			<p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
		  <p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
			
            <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150621033442.png" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150621033442.png" width="80" height="80"></a>
            
          <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Figure-1-"></a>Figure 1: </span><a href="http://nexusacademicpublishers.com/uploads/figures/20150621033442.png">Hemorrhagic liver due to <span class="CharOverride-6">Aeromonas hydrophila</span> infection</a></p>
       </div>

			
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">Discussion</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text"><span class="CharOverride-6">A. hydrophila</span> and other motile <span class="CharOverride-6">aeromonads</span> are among the most common bacteria in freshwater habitats throughout the world, and these bacteria frequently cause infectious diseases among cultured and feral fishes (<a href="#Maalej-S--Denis-M--Dunkan-S--2004-"><span class="Hyperlink">Maalej et al., 2004</span></a>). Speculated that septicemic infections in fish caused by motile <span class="CharOverride-6">Aeromonads</span> that were common throughout Europe during the middle ages. Although the bacterial etiology of these early reports was inconclusive, the pathology was similar to that observed with red leg disease in frogs, in which <span class="CharOverride-6">A. hydrophila</span> was identified as the causal organism. Because many bacteria isolated from fish with hemorrhagic septicemias in fish were often misidentified, it is now recognized that certain isolations of bacteria ascribed to the genera <span class="CharOverride-6">Pseudomonas, Proteus, Bacillus, Aerobacter, </span>and<span class="CharOverride-6"> Achromobacter</span> actually belonged to the genus <span class="CharOverride-6">Aeromonas</span>. The exact etiology of disease involving <span class="CharOverride-6">Aeromonads</span> is complicated by the diverse genetic, biochemical, and antigenic heterogeneity that exists among members of this group (<a href="#Mary-P--Chihib-NE--Charafeddine-O--Defives-C--Hornez-JP--2002"><span class="Hyperlink">Mary et al., 2002</span></a>). Consequently, motile Aeromonads are often referred to as a complex of disease organisms that are associated with bacterial hemorrhagic septicemias and other ulcerative conditions in fishes (<a href="#Post-E--1989-"><span class="Hyperlink">Post, 1989</span></a>). Although motile aeromonads appropriately receive much notoriety as pathogens of fish, it is important to note that these bacteria also compose part of the normal intestinal microflora of healthy fish (<a href="#Trust-TJ--Bull-LM--Currie-BR--Buckley-JT--1974-."><span class="Hyperlink">Trust et al., 1974</span></a>). Therefore, the presence of these bacteria, by itself, is not indicative of disease and, consequently, stress is often considered to be a contributing factor in outbreaks of disease caused by these bacteria. Such stressors are most commonly associated with environmental and physiological parameters that adversely fish under intensive culture. Furthermore, <a href="#Cipriano-RC--Bullock-GL--Pyle-SW--1984"><span class="Hyperlink">Cipriano et al. (1984)</span></a> have shown that the prevalence of motile Aeromonad septicemia in cultured and wild Nile tilapia (<span class="CharOverride-6">Oreochromis niloticus)</span> was 10.0% and 2.5% respectively; it was 18.75% and 6.25% in cultured and wild Karmout catfish, respectively. <a href="#Ventura-MT--Grizzle-JM--1988"><span class="Hyperlink">Ventura and Grizzle (1987)</span></a> produced systemic infections more readily among channel catfish (<span class="CharOverride-6">Ictalurus punctatus</span>) by abrading their skin prior to exposing the fish to the bacterium. Under controlled laboratory conditions, <a href="#Defigvereido-J--Plumb-JA--2005-"><span class="Hyperlink">Defigueredo and Plumb (2005)</span></a> found that strains of motile <span class="CharOverride-6">aeromonads</span> isolated from diseased fish were more virulent to channel catfish than were those isolated from pond water. <a href="#Lallier-R--Leblanc-D--Mittal-KR--Olivier-G--1981"><span class="Hyperlink">Lallier et al. (1981)</span></a> performed studies on rainbow trout <span class="CharOverride-6">(Oncorhyncus mykiss</span>, formerly <span class="CharOverride-6">Salmo gairdneri</span> to compare the relative virulence of <span class="CharOverride-6">A. hydrophila</span> and <span class="CharOverride-6">A. sobria</span>, as taxonomically described. Their results indicated that strains of <span class="CharOverride-6">A. hydrophila</span> isolated from either healthy or diseased fish were more virulent than strains of <span class="CharOverride-6">A</span>. <span class="CharOverride-6">sobria</span>. Additionally, <span class="CharOverride-6">A. sobria</span> was not isolated from fish with clinical signs of motile <span class="CharOverride-6">aeromonad</span> septicemia (<a href="#Bogosian-G--Aardema-ND--Bourneuf-EV--Morris-PJL--O-Neil-JP--2000-"><span class="Hyperlink">Bogosian et al., 2000</span></a>; <a href="#Paniagua-C--Rivero-O--Anguita-J--Naharro-G--1990-."><span class="Hyperlink">Paniagua et al., 1990</span></a>), for example, collected <span class="CharOverride-6">aeromonad</span> isolates along the River Porma, Leon Province (Spain) and found that their isolates grouped within three species; <span class="CharOverride-6">A</span>  <span class="CharOverride-6">hydrophila </span>(n=74 strains), <span class="CharOverride-6">A. sobria</span> (n =11 strains), and <span class="CharOverride-6">A. caviae</span> (n = 12 strains). The authors additionally observed that 72.02% of <span class="CharOverride-6">A. hydrophila</span> isolates and 63% of <span class="CharOverride-6">A. sobria</span> isolates were virulent for fish by intramuscular challenge, but all of the strains of <span class="CharOverride-6">A. caviae</span> were avirulent. Pathologic conditions attributed to members of the motile <span class="CharOverride-6">Aeromonad</span> complex may include dermal ulceration, tail or fin rot, ocular ulcerations, erythrodermatitis, hemorrhagic septicemia, red sore disease, red rot disease, and scale protrusion disease (<a href="#Desnues-B--Cuny-C--Gre-gori-G--Dukan-S--Aguilaniu-H--Nystrom-T--2003"><span class="Hyperlink">Desnues et al., 2003</span></a>).</p>
		  <p class="Body-Text">&nbsp;</p>
			<p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Table-4-"></a>Table 4:</span> Principal technology components and their relationship to stress management of fish in cage culture technology</p>
			<table id="table-4" class="Table-Style-1">
				<colgroup>
					<col />
					<col />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Technology component </span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-13">Stress management component </span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">1. Select a high quality, relatively stable environment for culture </span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">1. Water quality variables fluctuate well within tolerance ranges of fish </span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">2. Construct rectangular or square cages of 1- to 4-m3 volume; enclose with net with &gt; 13mm mesh</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">2. Water exchanges maintain water quality inside cages within tolerance ranges of fish </span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">3. Place cages in open water, </span></p>
							<p class="Basic-Paragraph"><span class="CharOverride-11">with individual cages spaced apart </span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">3.  Water exchanges maintain water quality inside cages within tolerance ranges of fish</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-1">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">4. Stock select, healthy fish </span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">4. Fish have natural resistance to stressors and pathogens </span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-2">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">5. Feed proper allowance of nutritionally complete feed of good physical quality in special feed enclosure </span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-11">5. Nutritional requirements are for growth and good health. Minimum pollution potential wastes are lost to the environment </span></p>
						</td>
					</tr>
				</tbody>
			</table><br>
			
          <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150621031144.png" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150621031144.png" width="80" height="80"></a>
            
          <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Figure-2-"></a>Figure 2: </span><a href="http://nexusacademicpublishers.com/uploads/figures/20150621031144.png">Number of infected fish to <span class="CharOverride-6">Aeromonas hydrophila</span> in 3 different temperature ranges (<span class="CharOverride-5">1</span>: 0-10˚C; <span class="CharOverride-5">2</span>: 10-20˚C; <span class="CharOverride-5">3</span>: 20- 30˚C; <span class="CharOverride-5">4</span>: Over 30˚C)</a></p>
       </div>
       
			
		  
			<p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">These researchers further revealed that <span class="CharOverride-6">A. hydrophila</span> infected internal organs through the digestive tract or uninjured skin under conditions of crowding (13.1 g of fish/L) and high temperature (24<span class="CharOverride-7">o</span>C). Such infections did not occurring when catfish were held at a lower density (5.2 g of fish/L) and temperature (18<span class="CharOverride-7">o</span>C). In another study (<a href="#Peters-G--Faisal-M--Lang-T--Ahmed-I--1988"><span class="Hyperlink">Peters et al., </span><span class="Hyperlink">2000</span></a>) subjected subordinate rainbow trout (<span class="CharOverride-6">Oncorhynchus mykiss)</span> to social stresses of cohabitation with dominant cohorts and then exposed these fish to infection by <span class="CharOverride-6">A. hydrophila</span>. By comparison to their dominant counterparts, the subordinate trout revealed physical evidence of stress based on elevated plasma glucose concentrations and increased leukocyte volumes. Following exposure to the pathogen, the bacterium was also recovered from more organs and with greater prevalence among the subordinate fish than from their dominant cohorts. In feral fish, other factors may be important stressors that trigger motile <span class="CharOverride-6">Aeromonad</span> septicemias. For example, described the occurrence of such an epizootic associated with spawning stress among gizzard shad (<span class="CharOverride-6">Dorosoma cepedianum</span>) in the Potomac River (Maryland, U.S.A.). It is important to submit fish suspected of being infected with <span class="CharOverride-6">Aeromonas </span>to a diagnostic laboratory to confirm the disease, and to determine the antibiotic (<a href="#Cipriano-RC--2001"><span class="Hyperlink">Cipriano, 2001</span></a>). Sensitivity of the strain of <span class="CharOverride-6">Aeromonas</span> can cause some problems. In addition, because <span class="CharOverride-6">Aeromonas</span> infections is stress-mediated disease, it is not unusual to find that infected fish are heavily parasitized or on currently infected with another systemic disease agents. When MAS, or any bacterial infection, is suspected in your fish, you should immediately submit a live, sick fish to the nearest diagnostic facility (<a href="#Vila-J--Marco-F--Soler-L--ChaconM--Figueras-MJ--2002-."><span class="Hyperlink">Vila et al., 2002</span></a>). If <span class="CharOverride-6">Aeromonas</span> is diagnosed, you need to know what legal drug the isolate is sensitive to and whether or not other infectious agents are present. In addition, if the affected system is an indoor or closed system, good sanitation is essential to decrease the number of bacteria in the system. Techniques for preventing and minimizing stress are the principal techniques of any aquaculture technology system (<a href="#Maalej-S--Denis-M--Dunkan-S--2004-"><span class="Hyperlink">Maalej et al., 2004</span></a>). Principal technology components of high density cage fish culture and their purpose relative to stress management are presented in <a href="#Table-4-"><span class="Hyperlink">Table 4</span></a> By managing those specific chemical, biological and physical factors discussed earlier that may cause  stress, fish health is almost assured but not guaranteed. Diseases do occur in fish that have not been stress mediated (<a href="#Bogosian-G--Bourneuf-E--2001-"><span class="Hyperlink">Bogosian and Bourneu, 2001</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">The behavior of <span class="CharOverride-6">A. hydrophila</span>, like that of other bacteria, depends on the cells’ history and their ability to respond to environmental changes (<a href="#Yesmin-SMH--Rahman-M--Hussain-A--Khan-AR--Pervin-F--Hossain-MA--2004"><span class="Hyperlink">Yesmin et al., 2004</span></a>). The organism is ubiquitous in nature and is even found in the intestinal tract of the fish. In natural situations, infections of fish with <span class="CharOverride-6">A. hydrophila</span> are probably a minor problem. However, with intensive fish-fanning systems, whether these systems are outdoor ponds or indoor aquaria or tanks, other factors must be considered. The common occurrence of this disease relates to stress conditions or factors of the fish (<a href="#Holmes-B--Farmer-JJ--2009"><span class="Hyperlink">Holmes et al., 2005</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
             <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150621035344.png" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150621035344.png" width="80" height="80"></a>
            
          <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-5"><a id="Figure-3-"></a>Figure 3: </span><a href="http://nexusacademicpublishers.com/uploads/figures/20150621035344.png">Generalized illustration of how warm water freshwater fish might respond to specific environmental factors under certain conditions</a></p>
       </div>
       
		  
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
		  <p class="Body-Text ParaOverride-1">Fish experts agree that fish are easily stressed when mishandled, overcrowded, transported under poor conditions, are on a poor level of nutrition, have poor water quality (<a href="#Shelton-JL--2000-"><span class="Hyperlink">Shelton, 2000</span></a>). Experimental demonstration shows that fish which are in poor environments, low water quality such as high nitrite levels, low levels of dissolved oxygen (DO) or high levels of carbon dioxide (CO<span class="CharOverride-16">2</span>) are more susceptible to infection by <span class="CharOverride-6">Aeromonas hydrophila</span>. Additionally, a seasonal incidence of a higher number of rep-ted fish deaths in the spring is associated with decreased water temperatures (<a href="#El-Mejri-S--El-Bour-M--Boukef-I--Mraouna-R--Boudabbous-A--2008-"><span class="Hyperlink">El Mejri et al., 2008</span></a>). Based on these observations, we suggest that low densities of <span class="CharOverride-6">A. hydrophila</span> in internal organs, coupled with high water temperatures and stress induced by extreme trapping procedures and handling, are sufficient to create physiologic conditions conducive to rapid proliferation of the bacteria. Similar to our results, the results of the <a href="#Rahman-MH--Suzuki-S--Kawai-K--2001"><span class="Hyperlink">Rahman et al. (2001)</span></a> indicated that temperature affects both the cell surface structure of <span class="CharOverride-6">A. hydrophila</span> and the phagocytic activity of goldfish macrophages, resulting in different mortality rates of the fish at different temperatures.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">In this study, it was clear that at 20-30 and above 30°C, the bacteria were more pathogenic than at 10-20 and lower than 20°C in <span class="CharOverride-6">C. damascina</span>. This finding was similar to results that observed in goldfish at 17 and 25°C, the bacteria were more pathogenic than at 10°C (<a href="#Rahman-MH--Suzuki-S--Kawai-K--2001"><span class="Hyperlink">Rahman et al., 2001</span></a>). Survey of <a href="#O-Reilly-T--Day-DF--1983-"><span class="Hyperlink">O’Reilly and Day (1983)</span></a> revealed that stimulation of the extracellular proteolytic activity of <span class="CharOverride-6">A. hydrophila</span> was influenced by temperature. It is possible that <span class="CharOverride-6">A. hydrophila</span> may have lost its virulence at 10°C and below. At 10°C, the metabolism and potential virulence mechanisms of <span class="CharOverride-6">A. hydrophila</span> are decreased. On the other hands,  <a href="#Ishiguro-EE--Kay--WW--Anisworth-T--Chamberlain-JB--Austen-RA--Buckley-TJ--Trust-TJ--1981"><span class="Hyperlink">Ishiguro et al. (1981</span></a>) reported that <span class="CharOverride-6">A. salmonicida</span> lost its virulence when cultured at high temperatures. Results of <a href="#Sautour-M--Mary-P--Chihib-NE--Hornez-JP--2003"><span class="Hyperlink">Sautour et al. (2003)</span></a> and <a href="#Koeypudsa-W--Jongjareanjai-M--2010-"><span class="Hyperlink">Koeypudsa and Jongjareanjai (2010)</span></a> showed among the three factors tested including temperature, water activity and pH, temperature has the main influence on growth and survival of <span class="CharOverride-6">A. hydrophila</span> in microcosm water. These results confirmed current findings.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">It was suggest that the increase level of <span class="CharOverride-6">A. hydrophila</span> in internal organs during increasing the water temperature was related to the production of sufficient quantities of lytic toxins that caused histopathology changing and subsequent mortality. So, <span class="CharOverride-6">A. hydrophila</span>, in combination with stress factors, could increase mortality. It was recommended that during trapping, handling and manipulation of fish increasing of water temperatures should be eliminated.  </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Acknowledgement</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">We would like to thanks Mr. Mohamadreza Sabetpay for his kind collaboration in this study.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Author’s cONTRIBUTION</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text">Authors declare that there is no conflict of interest.</p>
		  <p class="Body-Text">&nbsp;</p>
			<p class="Heading-1--Introduction----">Conflict of Interest</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">Laleh Yazdanpanah Goharrizi participated in the design and coordination of the study, Milad Adel and Mohammad Ebrahim Jalil Zorriehzahra participated in field sampling and conducted the laboratory work. Milad Adel analyzed data of this study and Mohammad Ebrahim Jalil Zorriehzahra reviewed the manuscript. All authors read and approved the final manuscript.  </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Reference</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		
			  <li class="References ParaOverride-2" xml:lang="en-US"><a id="Austin-B--Stobie-DA--1993-"></a>Austin B, Stobie DA (1993). Bacterial fish pathogens: Disease of farmed and wild fish. Ellise Horwood Chichester. Pp. 35-39.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Bogosian-G--Aardema-ND--Bourneuf-EV--Morris-PJL--O-Neil-JP--2000-"></a>Bogosian G, Aardema ND, Bourneuf EV, Morris PJL, O’Neil JP (2000). Recovery of hydrogen peroxide sensitive culturable cells of <span class="CharOverride-6">Vibrio vulnificus</span> gives the appearance of resuscitation from a viable but nonculturable state. J. Bacteriol. 182: 5070–5075. <a href="http://dx.doi.org/10.1128/JB.182.18.5070-5075.2000"><span class="Hyperlink">http://dx.doi.org/10.1128/JB.182.18.5070-5075.2000</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Bogosian-G--Bourneuf-E--2001-"></a>Bogosian G, Bourneuf E (2001). A matter of bacterial life and death. EMBO Rep. 2: 770–774. <a href="http://dx.doi.org/10.1093/embo-reports/kve182"><span class="Hyperlink">http://dx.doi.org/10.1093/embo-reports/kve182</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Bullock-GL--1968-"></a>Bullock GL (1968). The bacteriology of brook trout with tail rots, Progressive Fish-Culturist. J. Dis. Aqua. Organ. 30: 19-22.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Chakrabaiti-NM--2006-"></a>Chakrabaiti NM (2006). Diseases of cultivable fresh water fishes and their control<span class="CharOverride-6">. </span>International Books and Periodicals Supply Service Dilni-110034(India). Pp. 149.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Cipriano-RC--Bullock-GL--Pyle-SW--1984"></a>Cipriano RC, Bullock GL, Pyle SW (1984). <span class="CharOverride-6">Aeromonas hydrophila</span> and motile aeromonad septicemias of fish. Fish Disease Leaflet 68. </li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Cipriano-RC--2001"></a>Cipriano RC (2001). Revision of Fish Disease Leaflet 68 (1984), “Aeromonas Hydrophila And Motile Aeromonad Septicemias Of Fish,” by R. C. Cipriano, G. L. Bullock, and S. W. Pyle. </li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Colwell-RR--2000-"></a>Colwell RR (2000)<span class="CharOverride-6">.</span> Nonculturable microorganisms in the environment. pp. 325–342. Edited by R. R. Colwell &amp; D. J. Grimes. Washington: American Society for Microbiology. <a href="http://dx.doi.org/10.1007/978-1-4757-0271-2"><span class="Hyperlink">http://dx.doi.org/10.1007/978-1-4757-0271-2</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Defigvereido-J--Plumb-JA--2005-"></a>Defigvereido J, Plumb JA (2005). Virulence of different isolates of <span class="CharOverride-6">Aeromonas hydrophila</span> in channel catfish. Aquaculture. 11: 349-54. </li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Desnues-B--Cuny-C--Gre-gori-G--Dukan-S--Aguilaniu-H--Nystrom-T--2003"></a>Desnues B, Cuny C, Gre´gori G, Dukan S, Aguilaniu H, Nystrom T (2003). Differential oxidative damage and expression of stress defence regulations in culturable and nonculturable cells of <span class="CharOverride-6">Escherichia coli</span>. EMBO Rep. 4: 400–404. <a href="http://dx.doi.org/10.1038/sj.embor.embor799"><span class="Hyperlink">http://dx.doi.org/10.1038/sj.embor.embor799</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Elbour-M--Attia-H--Elhilli--Mraouna--Ayari-W--2001"></a>Elbour M, Attia H, Elhilli, Mraouna, Ayari W (2001). Bacterial study of mesophillic aeromonads distribution in shellfish.<span class="CharOverride-6"> </span>In: Proceedings of the fifth international conference on the Mediterranean coastal environment, MEDCoASt, 1, 557-565.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="El-Mejri-S--El-Bour-M--Boukef-I--Mraouna-R--Boudabbous-A--2008-"></a>El Mejri S, El Bour M, Boukef I, Mraouna R, Boudabbous A (2008). Effect of previous incubation of <span class="CharOverride-6">Aeromonas hydrophila</span> in wastewater prior to its transfer into marine water microcosms.  J. Water Sci. 21(3): 259-266. <a href="http://dx.doi.org/10.7202/018774ar"><span class="Hyperlink">http://dx.doi.org/10.7202/018774ar</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Holmes-B--Farmer-JJ--2009"></a>Holmes B, Farmer JJ (2009). International committee on systematics of prokaryotes subcommittee on the taxonomy of Aeromonadaceae, Vibrionaceae and related organisms<span class="CharOverride-6">. </span>Minutes of the Meetings, 25 July (2005), San Francisco, CA, USA. Int. J. Syst. Evol. Microbiol. 59: 2633-2637. <a href="http://dx.doi.org/10.1099/ijs.0.017145-0"><span class="Hyperlink">http://dx.doi.org/10.1099/ijs.0.017145-0</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Ishiguro-EE--Kay--WW--Anisworth-T--Chamberlain-JB--Austen-RA--Buckley-TJ--Trust-TJ--1981"></a>Ishiguro EE, Kay, WW, Anisworth T, Chamberlain JB, Austen RA, Buckley TJ, Trust TJ (1981). Loss of virulence during culture of <span class="CharOverride-6">Aeromonas salmonicida</span> at high temperature. J. Bacteriol. 148: 333-340.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Kersters-I--Van-Vooren-L--Huys-G--Janssen-P--Kersters-K--Verstraet-W--1995"></a>Kersters I, Van Vooren L, Huys G, Janssen P, Kersters K, Verstraet W (1995). Influence of temperature and process technology on the occurrence of <span class="CharOverride-6">Aeromonas</span> species and hygienic indicator organisms in drinking water production plants. Microbial. Ecol. 30(2): 203-218. <a href="http://dx.doi.org/10.1007/BF00172575"><span class="Hyperlink">http://dx.doi.org/10.1007/BF00172575</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Kersters-I--Van-2010"></a>Kersters I, Van Vooren L, Huys G, Janssen P, Kersters K, Koeypudsa W, Jongjareanjai W (2010). Effect of water temperature on hematology and virulence of Aeromonas hydrophila in hybrid catfish (<span class="CharOverride-6">Clarias gariepinus</span> Burchell x <span class="CharOverride-6">C. macrocephalus</span><span class="CharOverride-1"> </span>Gunther). Thai J. Vet. Med. 40(2): 179-186.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Koeypudsa-W--Jongjareanjai-M--2010-"></a>Koeypudsa W, Jongjareanjai M (2010). Effect of water temperature on hematology and virulence of <span class="CharOverride-6">Aeromonas hydrophila</span> in hybrid catfish (<span class="CharOverride-6">Clarias gariepinus</span> Burchell x <span class="CharOverride-6">C. macrocephalus</span> Gunther). Thai J. Vet. Med. 40(2): 179-186.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Krovacek-K--Pasquale-V--Baloda-SB--Soprano-V--Conte-M--Dumontet-S--1994"></a>Krovacek K, Pasquale V, Baloda SB, Soprano V, Conte M, Dumontet S (1994). Comparison of putative virulence in <span class="CharOverride-6">Aeromonas hydrophila </span>strains isolated from the marine environment and human diarrhoeal cases in southern italy. Appl. Environ. Microbiol. 60: 1379-1382.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Lallier-R--Leblanc-D--Mittal-KR--Olivier-G--1981"></a>Lallier R, Leblanc D, Mittal KR, Olivier G (1981). Serogrouping of motile <span class="CharOverride-6">Aeromonas</span> species isolated from healthy and moribund fish. J. Appl. Environ. Microbiol. 42: 56-60.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Mary-P--Chihib-NE--Charafeddine-O--Defives-C--Hornez-JP--2002"></a>Mary P, Chihib NE, Charafeddine O, Defives C, Hornez JP (2002). Starvation survival and viable but non culturable states in <span class="CharOverride-6">Aeromonas hydrophila.</span> Microbiol. Ecol. 43: 250-258. <a href="http://dx.doi.org/10.1007/s00248-001-0046-4"><span class="Hyperlink">http://dx.doi.org/10.1007/s00248-001-0046-4</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Maalej-S--Denis-M--Dunkan-S--2004-"></a>Maalej S, Denis M, Dunkan S (2004). Temperature and growth-phase effects on <span class="CharOverride-6">Aeromonas hydrophila</span> survival in natural seawater microcosms: role of protein synthesis and nucleic acid content on viable but temporally nonculturable response. Microbiology. 150: 181-187. <a href="http://dx.doi.org/10.1099/mic.0.26639-0"><span class="Hyperlink">http://dx.doi.org/10.1099/mic.0.26639-0</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><span class="Hyperlink"><a id="Mohajeri-J--Afsharnasab-M--Jalali-B--Kakoolaki-S--Sharifrohani-M--Haghighi-A--2011"></a></span>Mohajeri J, Afsharnasab M, Jalali B, Kakoolaki S, Sharifrohani M, Haghighi A (2011). Immunological and histopathological changes in <span class="CharOverride-6">Penaeus semisulcatus</span> challenged with <span class="CharOverride-6">Vibrio harveyi</span>. Iranian J. Fish. Sci. 10(2): 254-265.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="O-Reilly-T--Day-DF--1983-"></a>O’Reilly T, Day DF (1983). Effect of cultural conditions on protease production by <span class="CharOverride-6">Aeromonas hydrophila</span>. Appl. Environ. Microbiol. 45: 1132-1135.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Paniagua-C--Rivero-O--Anguita-J--Naharro-G--1990-."></a>Paniagua C, Rivero O, Anguita J, Naharro G (1990). Pathogenicity factors and virulence for rainbow trout (<span class="CharOverride-6">Salmo gairdneri</span>) or motile <span class="CharOverride-6">Aeromonas </span>spp. isolated from a river. J. Clin. Microbiol. 28: 350–355.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Peters-G--Faisal-M--Lang-T--Ahmed-I--1988"></a>Peters G, Faisal M, Lang T, Ahmed I (1988). Stress caused by social interaction and its effect on susceptibility to Aeromonas hydrophila infection in rainbow trout <span class="CharOverride-6">Salmo gairdneri</span>.  Dis. Aqua. Organ. 4: 83 - 89. <a href="http://dx.doi.org/10.3354/dao004083"><span class="Hyperlink">http://dx.doi.org/10.3354/dao004083</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Post-E--1989-"></a>Post E (1989). Text book of fish health. T.F.H. Publication Inc. 10: 293P. </li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Rahman-MH--Suzuki-S--Kawai-K--2001"></a>Rahman MH, Suzuki S, Kawai K (2001). The effect of temperature on <span class="CharOverride-6">Aeromonas hydrophila</span> infection in goldfish, <span class="CharOverride-6">Carassius auratus</span>. J. Appl. Ichthyol. 17: 282–285.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Sautour-M--Mary-P--Chihib-NE--Hornez-JP--2003"></a>Sautour M, Mary P, Chihib NE, Hornez JP (2003). The effects of temperature, water activity and pH on the growth of <span class="CharOverride-6">Aeromonas hydrophila</span> and on its subsequent survival in microcosm water. J. Appl. Microbiol. 95: 807–813. <a href="http://dx.doi.org/10.1046/j.1365-2672.2003.02048.x"><span class="Hyperlink">http://dx.doi.org/10.1046/j.1365-2672.2003.02048.x</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><span class="Hyperlink"><a id="Santos-JA--Gonza-lez-CJ--Otero-A--Garc--a-Lo-pez-ML--1999"></a></span>Santos JA, Gonza´lez CJ, Otero A, Garcı´a-Lo´pez ML (1999). Haemolytic activity and siderophore production in different Aeromonas species isolated from fish. Appl. Environ. Microbiol. 65: 5612–5614.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Saulnier-D--Haffner-P--Goarant-C--Levy-P--Ansquer-D--2000-"></a>Saulnier D, Haffner P, Goarant C, Levy P, Ansquer D (2000). Experimental infection models for shrimp vibriosis studies: are view. Aquaculture. 191: 133-144.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Shelton-JL--2000-"></a>Shelton JL (2000). Trout aquaculture. Cooperative State Research Service and Extension Service, U.S. Department of Agriculture, under Special Project No. 87-Exca-3-0836.</li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Trust-TJ--Bull-LM--Currie-BR--Buckley-JT--1974-."></a>Trust TJ, Bull LM, Currie BR, Buckley JT (1974). Obligate anaerobic bacteria in the gastrointestinal microflora of the grass carp (<span class="CharOverride-6">Ctenopharyngodon idella</span>), goldfish (<span class="CharOverride-6">Carassius auratus</span>), and rainbow trout (<span class="CharOverride-6">Salmo gairdneri</span>). J. Fish. Res. Board Canada. 36: 1174 - 1179. <a href="http://dx.doi.org/10.1139/f79-169"><span class="Hyperlink">http://dx.doi.org/10.1139/f79-169</span></a><span class="Hyperlink"> </span></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><span class="Hyperlink"><a id="Ventura-MT--Grizzle-JM--1988"></a></span>Ventura MT, Grizzle JM (1988). Lesions associated with natural and experimental infections of <span class="CharOverride-6">Aeromonas hydrophila </span>in channel catfish<span class="CharOverride-6">, Ictalurus punctatus </span>(Rafinesque). J. Fish Dis. 11: 397-407.</li>
		  <li class="References ParaOverride-2" xml:lang="en-US"><a id="Vila-J--Marco-F--Soler-L--ChaconM--Figueras-MJ--2002-."></a>Vila J, Marco F, Soler L, ChaconM, Figueras MJ (2002). <span class="CharOverride-6">In vitro</span> antimicrobial susceptibility of clinical isolates of <span class="CharOverride-6">Aeromonas caviae</span>, <span class="CharOverride-6">Aeromonas hydrophila</span> and <span class="CharOverride-6">Aeromonas veronii</span> biotype <span class="CharOverride-6">sobria</span>. J. Antimicrob. Chemother. 49: 701-102. <a href="http://dx.doi.org/10.1093/jac/49.4.701"><span class="Hyperlink">http://dx.doi.org/10.1093/jac/49.4.701</span></a></li>
				<li class="References ParaOverride-2" xml:lang="en-US"><a id="Yesmin-SMH--Rahman-M--Hussain-A--Khan-AR--Pervin-F--Hossain-MA--2004"></a>Yesmin SMH, Rahman M, Hussain A, Khan AR, Pervin F, Hossain MA (2004). <span class="CharOverride-6">Aeromonas hydrophila</span> infection in fish of swamps in Bangladesh. Pakistan J. Biol. Sci. 7(3): 409-411. <a href="http://dx.doi.org/10.3923/pjbs.2004.409.411"><span class="Hyperlink">http://dx.doi.org/10.3923/pjbs.2004.409.411</span></a></li>
                <p>&nbsp;</p>
                <p>&nbsp;</p>
                <p>&nbsp;</p>
                <p>&nbsp;</p>
			
		</div>
	</body>
</html>
