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			<p class="Type-of-Article">&nbsp;</p>
			<p class="Type-of-Article"><span class="CharOverride-1">Research Article</span></p>
	</div>
		<div class="Basic-Text-Frame">
			<p class="title- ParaOverride-1">&nbsp;</p>
			<p class="title- ParaOverride-1">Comparison of Phenotypic Characteristics and Virulence Traits of Klebsiella pneumoniae Obtained from Pneumonic and Healthy Camels (Camelus dromedarius)</p>
		</div>
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			<p class="Authors ParaOverride-1">&nbsp;</p>
			<p class="Authors ParaOverride-1"><span class="CharOverride-3">Sandeep Kumar Sharma</span><span class="CharOverride-4">1*</span><span class="CharOverride-3">, Naresh Kumar Mudgal</span><span class="CharOverride-4">2</span><span class="CharOverride-3">, Pradeep Sharma</span><span class="CharOverride-4">2</span><span class="CharOverride-3">, Brij Nandan Shrngi</span><span class="CharOverride-4">3</span></p>
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			<p class="Affiliations ParaOverride-1"><span class="CharOverride-5">1</span>Department of Veterinary Microbiology &amp; Biotechnology, Post Graduate Institute of Veterinary Education and Research- Jaipur, Rajasthan University of Veterinary and Animal Sciences, Bikaner, India; <span class="CharOverride-5">2</span>Department of Animal Husbandry; <span class="CharOverride-5">3</span>Department of Veterinary Microbiology &amp; Biotechnology, College of Veterinary &amp; Animal Science- Bikaner, Rajasthan University of Veterinary and Animal Sciences, Bikaner, India.</p>
		</div>
		<div>
			<p class="Abstract ParaOverride-1">&nbsp;</p>
			<p class="Abstract ParaOverride-1"><span class="CharOverride-6">Abstract</span> | The present study was aimed to compare <span class="CharOverride-7">Klebsiella pneumoniae </span>isolates obtained from nasal discharge of pneumonic and healthy camels in context of phenotypic characteristics and virulence traits. Total 65, 16S-23S rDNA internal transcribed spacer (ITS) region based confirmed isolates were included with species-specific amplicon of 130 bp in this investigation. Out of which 47 were from pneumonic and 18 from healthy camels. All <span class="CharOverride-7">K. pneumoniae </span>isolates showed typical inherent phenotypic pattern except six isolates from pneumonic camels were found ability to produce indole. For virulence traits, all isolates were non hemolytic with prominent capsule. All the isolates from pneumonic camels were urease positive and most of (93.6%) isolates positive on first three days while all isolates from healthy camels were urease negative on first three days. None of the isolates from healthy camels showed hypermucoviscosity while 25 (53.19%) isolates from pneumonic camels showed existence of virulent hypermucoviscosity trait. It may indicate differences in virulence properties of studied isolates and further suggest some genotypic studies regarding virulence characterization of<span class="CharOverride-7"> K. pneumoniae</span>.</p>
		  <p class="Abstract ParaOverride-1">&nbsp;</p>
			<p class="Abstract ParaOverride-1"><span class="CharOverride-6">Keywords </span>| <span class="CharOverride-7">Klebsiella pneumoniae, </span>Camels, Phenotypic characteristics and virulence traits</p>
		  <p class="Abstract ParaOverride-1">&nbsp;</p>
		</div>
		<div class="Basic-Text-Frame">
			<p class="Editor----Citation"><span class="CharOverride-9">Editor</span> | Kuldeep Dhama, Indian Veterinary Research Institute, Uttar Pradesh, India.</p>
			<p class="Editor----Citation"><span class="CharOverride-6">Received</span> | December  22, 2014; <span class="CharOverride-6">Revised</span> | January 10, 2015; <span class="CharOverride-6">Accepted</span> | January 12, 2015; <span class="CharOverride-6">Published</span> | January 21, 2015&#9;&#9;</p>
			<p class="Editor----Citation"><span class="CharOverride-6">*Correspondence</span> | Sandeep Kumar Sharma, Rajasthan University of Veterinary and Animal Sciences, Bikaner, India; <span class="CharOverride-6">Email: </span>drsharmask01@hotmail.com</p>
			<p class="Editor----Citation"><span class="CharOverride-6">Citation </span>| Sharma SK, Mudgal NK, Sharma P, Shrngi BN (2015). Comparison of phenotypic characteristics and virulence traits of Klebsiella pneumoniae obtained from pneumonic and healthy camels (Camelus dromedarius). Adv. Anim. Vet. Sci. 3(2): 116-122.  </p>
			<p class="Editor----Citation"><span class="CharOverride-9">DOI</span> | <a href="http://dx.doi.org/10.14737/journal.aavs/2015/3.2.116.122"><span class="Hyperlink">http://dx.doi.org/10.14737/journal.aavs/2015/3.2.116.122</span></a></p>
			<p class="Editor----Citation"><span class="Editor---Citation CharOverride-6" lang="en-US">ISSN </span><span class="Editor---Citation CharOverride-6" lang="en-US">(</span><span class="Editor---Citation CharOverride-6" lang="en-US">Online</span><span class="Editor---Citation CharOverride-6" lang="en-US">) | 2307-8316; <span class="Editor---Citation CharOverride-6" lang="en-US">ISSN </span><span class="Editor---Citation CharOverride-6" lang="en-US">(Print) </span> | 2309-3331</p>
			<p class="Editor----Citation"><span class="CharOverride-9">Copyright </span>© 2015 Sharma 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----">&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">K</span><span class="CharOverride-7">lebsiella </span><span class="CharOverride-7">pneumoniae</span> was recognized in 1887, when <a href="#Trevisan-V--1887-"><span class="Hyperlink">Trevisan (1987)</span></a> has described a bacterium from the lungs of a patient who had died of pneumonia. <span class="CharOverride-7">K. pneumoniae</span> is a lactose-fermenting, facultative anaerobic, non-motile with a prominent capsule bacillus in the family <span class="CharOverride-7">Enterobacteriaceae</span>. This opportunistic pathogen found in a variety of environmental sources such as soil, vegetation, water and industrial wastes including mucosal surfaces of mammals with common association of a broad range of infections <span class="CharOverride-7">viz.</span> peritonitis, septicemia, pneumonia, urinary tract infections, and meningitis (<a href="#Podschun-R--Ullmann-U--1998-"><span class="Hyperlink">Podschun and Ullmann, 1998</span></a>).<span class="CharOverride-7"> </span>It is an important pulmonary pathogen for camels, horses, dogs, foals, sow and calves (<a href="#Arora-RG--Kalra-DS--1973-."><span class="Hyperlink">Arora and Kalra, 1973</span></a>; <a href="#Boguta-L--Gradzki-Z--Borges-E--Maruin-F--Kodjo-A--WIniarczyk-S--2002-"><span class="Hyperlink">Boguta et al., 2002</span></a>; <a href="#Henton-MM--Coetzer-JA--Thomson-GR--Tustin-RC--1994-."><span class="Hyperlink">Henton et al., 1994</span></a>; <a href="#Selbitz-HJ--Rolle-M--Mayr-A--2002-."><span class="Hyperlink">Selbitz et al.,</span><span class="Hyperlink"> 2002</span></a>). In camels, <span class="CharOverride-7">K. pneumoniae</span> reported from acute destructive bronchopneumonia and community-acquired bacterial pneumonia with increased tendency to develop abscess, cavitation and empyema (<a href="#Arora-RG--Kalra-DS--1973-."><span class="Hyperlink">Arora and Kalra, 1973</span></a>; <a href="#Zubair-R--Khan-AMZ--Sabri-MA--2004-"><span class="Hyperlink">Zubair et al., 2004</span></a>; <a href="#Kane-Y--Kadja-MC--Bada-Alambedji-R--Bezeid-OE--Akakpo-JA--Kaboret-Y--2005-."><span class="Hyperlink">Kane et al., 2005</span></a>; <a href="#Abubakar-MS--Fatihu-MY--Ibrahim-NDG--Oladele-SB--Abubakar-MB--2010-"><span class="Hyperlink">Abubakar et al., 2010</span></a>).</p>
		  <p class="Caps-on-First-Para ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1"><span class="CharOverride-7">Klebsiella pneumoniae</span> strains exhibit many virulence factors such as capsular polysaccharides (CPS), lipopolysaccharide (LPS), iron-scavenging systems (siderophores), adhesins and hypermucoviscosity (<a href="#Podschun-R--Ullmann-U--1998-"><span class="Hyperlink">Podschun and Ullmann, 1998</span></a>) along with other virulence factors such as urease production, hemolysins, protein-tyrosine kinase, phosphotyrosine-protein phosphatase and heat-labile &amp; heat-stable endotoxins (<a href="#Dworkin-M--Falkow-S--Rosenberg-E--Schleifer-K--Stackebrandt-E--2006"><span class="Hyperlink">Dworkin et al., 2006</span></a>). The capsule is an important virulence factor to govern pathogenesis<span class="CharOverride-7">, </span>which protects the bacteria from phagocytosis and bactericidal effects of serum factors (<a href="#Highsmith-AK--Jarvis-WR--1985-."><span class="Hyperlink">Highsmith and Jarvis, 1985</span></a>, <a href="#Albesa-I--Eraso-AJ--Frigerio-CI--Lubetkin-AM--1980-."><span class="Hyperlink">Struve and Krogfelt, 2003</span></a>). Sometime <span class="CharOverride-7">K. pneumoniae</span> also exhibits large amounts of mucopolysaccharide web of capsular and extracapsular polysaccharides to produce more virulent hypermucoviscosity strain (<a href="#Wiskur-BJ--Hunt-JJ--Callegan-MC--2008-."><span class="Hyperlink">Wiskur et al., 2008</span></a>). <span class="CharOverride-7">Klebsiella</span> is a non-hemolytic for human red blood cells but found to be hemolytic for rabbit blood agar and urease production of this organism has significant role in colonization and survivability in various systemic infections (<a href="#Dworkin-M--Falkow-S--Rosenberg-E--Schleifer-K--Stackebrandt-E--2006"><span class="Hyperlink">Dworkin et al., 2006</span></a>; <a href="#Maroncle-N--Rich-C--Forestier-C--2006-.-Th"><span class="Hyperlink">Maroncle et al., 2006</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">Camel (<span class="CharOverride-7">Camelus dromedarius</span>) is a uniquely morphological &amp; physiological adapted animal in desert ecosystem with good potential to thrive well on meager resources under extreme climatic conditions. It has important utilities in human society such as drought, farming, milking and many other important farming purposes but sometime certain sudden environmental variations make this animal susceptible to various infections (<a href="#Kebede-F--Gelaye-E--2010-"><span class="Hyperlink">K</span><span class="Hyperlink">eb</span><span class="Hyperlink">e</span><span class="Hyperlink">d</span><span class="Hyperlink">e </span><span class="Hyperlink">a</span><span class="Hyperlink">n</span><span class="Hyperlink">d</span><span class="Hyperlink"> </span><span class="Hyperlink">G</span><span class="Hyperlink">e</span><span class="Hyperlink">la</span><span class="Hyperlink">y</span><span class="Hyperlink">e, </span><span class="Hyperlink">20</span><span class="Hyperlink">1</span><span class="Hyperlink">0</span></a>). In India<span class="CharOverride-7">,</span> <span class="CharOverride-7">Klebsiella pneumoniae</span> has been also reported from cases of <span class="CharOverride-7">“</span><span class="CharOverride-7">khurak”</span> (local name of pneumonia) in camels. <span class="CharOverride-7">“Khurak”</span> is an infectious respiratory disease characterized by alveolar serofibrinous exudation, capillary hyperaemia and microbial bronchopneumonia with respiratory distress, pyrexia, prolonged cough and septicemia in camels. It is a highly morbid, causing considerable loss of production and deaths especially in adult camels (<a href="#Arora-RG--Kalra-DS--1973-."><span class="Hyperlink">Arora and Kalra, 1973</span></a>; <a href="#Sharma-SK--2012-."><span class="Hyperlink">Sharma, 2012</span></a>). <span class="CharOverride-7">K.</span><span class="CharOverride-7"> </span><span class="CharOverride-7">p</span><span class="CharOverride-7">n</span><span class="CharOverride-7">eu</span><span class="CharOverride-7">m</span><span class="CharOverride-7">on</span><span class="CharOverride-7">iae</span><span class="CharOverride-7"> </span>found as commensal in healthy individual as well as pathogenic agent in diseased animals. So it is required to detect variations among <span class="CharOverride-7">Klebsiella pneumoniae</span> strains obtained from healthy and diseased individuals. Thus this study has designed to compare <span class="CharOverride-7">Klebsiella pneumoniae </span>isolates obtained from pneumonic and healthy camels on the basis of primary and secondary biochemical characters and virulence properties.</p>
			<p class="Body-Text ParaOverride-1">&nbsp;</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Materials and Methods</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Sample Collection and Identification</p>
			<p class="Body-Text ParaOverride-1">A total of 163 severe nasal discharge samples from pneumonic (96) and healthy (67) camels were collected. The pneumonic and healthy camels were determined on the basis of clinical symptoms <span class="CharOverride-7">viz.</span> the camel with severe nasal discharge, respiratory distress, depression and pyrexia up to 41°C temperature in early stages. After a week or so, the temperature tends to drop down to normal, but cough persists for a long time was considered as pneumonic camel. While the camels, which were not show any clinical and medication history since last six month considered as healthy. Samples of pneumonic camels has been collected from clinical complex of college of veterinary and animal science, Bikaner (Rajasthan) and samples of healthy camel has been from surrounding villages of Bikaner area. All samples were collected without any discrimination of age, sex and breed. The samples were collected directly from nasal cavity aseptically with the help of sterile cotton swabs. And proceeded on selective medium Simmon’s citrate agar with 1% inositol (SCAI) (<a href="#Van-Kregten-E--Westerdaal-NAC--Willers-JMN--1984-.-N"><span class="Hyperlink">Van Kregten et al., 1984</span></a>) and single yellow dome shaped colonies were further checked for genotypic confirmation with 16S-23S rDNA internal transcribed spacer (ITS) region based specific primers (<a href="#Liu-Y--Liu-C--Zheng-W--Zhang-X--Yu-J--Gao-Q--Hou-Y--Huang-X--2008-."><span class="Hyperlink">Liu et al., 2008</span></a>). For genotypic confirmation genomic DNA was used as template and isolation was carried out by method of<a href="#Chen-WP--Kuo-TT--1993-"><span class="Hyperlink"> Chen and Kuo (1993)</span></a>. All amplified PCR products were analyzed by 8% polyacrylamide gel electrophoresis (PAGE).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-2--History-in-MM-">Phenotypic Characterization and Detection of Virulence Traits</p>
			<p class="Body-Text ParaOverride-1">Genotypically confirmed 65 isolates were included, out of which 47 from pneumonic and 18 isolates were from healthy camels. All were subjected to various primary and secondary biochemical reactions such as catalase, oxidase and motility then checked for lactose fermenter mucoid colonies on MacConkey agar (MCA) and nonmetallic dark colored mucoid growth on EMB agar further secondary biochemical characters such as IMViC pattern, growth on TSI, gelatin liquefaction, aesculin hydrolysis, nitrate reduction, malonate utilization, lysine decarboxylation and ONPG (o-Nitrophenyl-<span class="CharOverride-13">β</span>-d Galactopyranoside) tests were conducted (<a href="#Quinn-PJ--Carter-ME--Markey-BK--Carter-GR--1994-."><span class="Hyperlink">Quinn et al., 1994</span></a> and <a href="#Garcia-LS--Isenberg-HD--2007"><span class="Hyperlink">Garcia and Isenberg, 2007</span></a>). For phenotypic detection of virulence traits, Maneval’s capsular staining method was used for capsular detection, haemolysin production was observed on sheep blood agar, urease production was tested by change in color of media from yellow to red observed for seven days in urease broth as described in literature and virulent hypermucoviscosity trait checked by string test (<a href="#Wiskur-BJ--Hunt-JJ--Callegan-MC--2008-."><span class="Hyperlink">Wiskur et al., 2008</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			
            <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150123033220.jpg" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150123033220.jpg" width="80" height="80"></a>
            
            <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-6"><a id="Figure-1"></a>Figure 1:</span><a href="http://nexusacademicpublishers.com/uploads/figures/20150123033220.jpg"> <span class="CharOverride-7">Klebsiella pneumoniae</span> showing Yellow, dome shaped mucoid colonies on SCAI</a></p>
       </div>

			
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">Results </p>
			<p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">Only 65 samples (47 were from pneumonic and 18 from healthy camels) showed typical yellow dome shaped colonies (<a href="#Figure-1"><span class="Hyperlink">Figure 1</span></a>) on Simmon’s citrate agar with 1% inositol (SCAI) from 163 nasal discharge samples of pneumonic and healthy camels and further these 65 isolates were confirmed with 16S-23S rDNA internal transcribed spacer (ITS) region based specific primers showing species-specific amplicon of 130 bp (<a href="#Figure-2"><span class="Hyperlink">Figure 2</span></a>). In the present study, without any variation all isolates showed pink lactose fermenter mucoid colonies on MacConkey agar (MCA). Nonmetallic dark colored mucoid growth on EMB agar, non-hemolytic grey-white, mucoid colonies on sheep blood agar (<a href="#Figure-3"><span class="Hyperlink">Figure 3</span></a>). And mucoviscosity observed in all other growth medium with large capsule. All isolates showed typical IMViC pattern (--++)  and other biochemical characters (<a href="#Table-1-"><span class="Hyperlink">Table 1</span></a>) as described in literature, except for six isolates from pneumonic camels that were found ability to produce indole. 100% isolates from pneumonic camels were urease producing and most of isolates of pneumonic camels were urease producers within first three days of observation while out of 18 isolates from healthy camels, three (16.66%) were urease negative and rest of the 15 isolates showed positive results after three days of incubation (<a href="#Table-2"><span class="Hyperlink">Table 2</span></a>). Only 25 (53.19%) isolates from pneumonic camels showed existence of virulent hypermucoviscosity trait (<a href="#Figure-4"><span class="Hyperlink">Figure 4</span></a>) while none of the <span class="CharOverride-7">K. pneumoniae</span> isolates from healthy camel exhibited hypermucoviscosity. </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			
            <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150123033121.jpg" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150123033121.jpg" width="80" height="80"></a>
            
           <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-6"><a id="Figure-2"></a>Figure 2:</span><a href="http://nexusacademicpublishers.com/uploads/figures/20150123033121.jpg">  <span class="CharOverride-7">Klebsiella pneumonaie</span> isolates showing typical 130bp size product with specific primers on 8% polyacrylamide gel</a></p>
       </div><br>
			
		  <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150123030524.jpg" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150123030524.jpg" width="80" height="80"></a>
            
          <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-6"><a id="Figure-3"></a>Figure 3:</span><a href="http://nexusacademicpublishers.com/uploads/figures/20150123030524.jpg">  <span class="CharOverride-7">Klebsiella pneumoniae</span> showing non-hemolytic grey-white, mucoid colonies on sheep blood agar</a></p>
       </div>
			
		  <p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
		  <p class="Figure--and-Table-Heading ParaOverride-1">&nbsp;</p>
		  <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-6"><a id="Table-1-"></a>Table 1: </span>Secondary biochemical tests of <span class="CharOverride-7">Klebsiella pneumoniae</span> isolates from pneumonic and healthy camels</p>
			<table id="table-1" class="Table-Style-1">
				<colgroup>
					<col class="_idGenTableRowColumn-1" />
					<col class="_idGenTableRowColumn-2" />
					<col class="_idGenTableRowColumn-3" />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-4">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-16">S. No.</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-16">Test</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-16">Reaction</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">1</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on 10°C</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-6">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">2</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on 44.5°C</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-7">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">3</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on SCAI</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Yellow mucoid dome shaped colonies</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-4">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">4</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on EMB agar</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Dark mucoid non-metallic colonies</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-7">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">5</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on BCP agar</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Yellow mucoid colonies</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-4">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">6</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Growth on TSI agar</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Acid/Acid/No H</span><span class="CharOverride-18">2</span><span class="CharOverride-17">S (Y/Y/-)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">7</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Gelatin Liquefaction</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-6">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">8</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Aesculin Hydrolysis</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">9</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Nitrate Reduction</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-6">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">10</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Malonate Utilization</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">11</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Arginine Hydrolysis</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-4">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">12</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Lysine Decaboxylation</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-7">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">13</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Phenylalanine Deamination</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-8">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">14</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">ONPG (o-Nitrophenyl-</span><span class="CharOverride-13">β</span><span class="CharOverride-17">-d Galactopyranoside)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive</span></p>
						</td>
					</tr>
				</tbody>
			</table><br>
		  <div class="pt" > <a href="http://nexusacademicpublishers.com/uploads/figures/20150123030827.png" target="new"><img class="img_display" src="http://nexusacademicpublishers.com/uploads/figures/20150123030827.png" width="80" height="80"></a>
            
         <p class="Figure--and-Table-Heading ParaOverride-1"><span class="CharOverride-6"><a id="Figure-4"></a>Figure 4:</span><a href="http://nexusacademicpublishers.com/uploads/figures/20150123030827.png">  <span class="CharOverride-7">Klebsiella pneumoniae</span> showing virulent hypermucoviscosity trait (string test)</a></p>
       </div>
			 
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
		  <p class="Heading-1--Introduction----">Discussion</p>
			<p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">In the present study, we have examined the primary and secondary biochemical characters and virulence traits of <span class="CharOverride-7">Klebsiella pneumoniae </span>isolates<span class="CharOverride-7"> </span>obtained from severe deep nasal discharge of pneumonic and healthy camels. Samples which were showing yellow dome shaped colonies on Simmon’s citrate agar with 1% inositol (SCAI) also found confirmed with <span class="CharOverride-7">K. pneumoniae</span> species specific primers. Similar to present study, <a href="#Sharma-SK--Kataria-AK--Shringi--BN--Nathawat-P--Bhati-T--Mohammed-N--2013-."><span class="Hyperlink">Sharma et al. (2013)</span></a> has also reported specificity of these species specific primers for detection of <span class="CharOverride-7">K. pneumoniae</span> from acute respiratory tract infections of camels. In addition, <a href="#Liu-Y--Liu-C--Zheng-W--Zhang-X--Yu-J--Gao-Q--Hou-Y--Huang-X--2008-."><span class="Hyperlink">Liu et al. (2008)</span></a> was also detected this organism with same primers and similar conditions from infant formula food sources. It proves genotypic specificity of this method for <span class="CharOverride-7">K. pneumoniae</span> detection not only from food sources but also from animal origin. It also indicates the good discriminative capacity of SCAI agar therefore SCAI agar can be used as a selective medium for isolation of <span class="CharOverride-7">K. pneumoniae.</span> <a href="#Van-Kregten-E--Westerdaal-NAC--Willers-JMN--1984-.-N"><span class="Hyperlink">Van Kregten et al. (1984)</span></a> had also reported selectivity of SCAI agar for <span class="CharOverride-7">K. pneumoniae</span> with similar results, who developed a medium based on the presence of two carbon sources, citrate and myo-inositol, without any inhibitor because <span class="CharOverride-7">K. pneumoniae</span> has ability to utilize both citrate and myo-inositol (<a href="#Dworkin-M--Falkow-S--Rosenberg-E--Schleifer-K--Stackebrandt-E--2006"><span class="Hyperlink">Dworkin et al., 2006</span></a>). </p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text">In agreement with present study, <a href="#Smith-JA--Ngui-Yen-JH--1980-"><span class="Hyperlink">Smith and Ngui-Yen (1980)</span></a> were also found non hemolytic <span class="CharOverride-7">K. pneumoniae</span> while <a href="#Albesa-I--Eraso-AJ--Frigerio-CI--Lubetkin-AM--1980-."><span class="Hyperlink">Albesa et al. (1980)</span></a> reported hemolytic <span class="CharOverride-7">K. pneumoniae</span> in rabbit blood agar. In support of present study,<span class="CharOverride-19"> </span>indole production in <span class="CharOverride-7">K. pneumoniae</span> has been also reported by <a href="#Brown-C--Seidler-RJ--1973-."><span class="Hyperlink">Brown and Seidler (1973)</span></a>, <a href="#Rennie-RP--Duncan-IBR--1974-."><span class="Hyperlink">Rennie and Duncan (1974)</span></a> and <a href="#Edwards-PR--Ewing-WH--1972-"><span class="Hyperlink">Edwards and Ewing (1972)</span></a> with 28%, 16% and 6% frequency respectively. None of the significant difference was observed in hemolytic pattern of <span class="CharOverride-7">K. pneumoniae</span> isolates obtained from pneumonic and healthy camels in present study but variations were observed in indole production. This ability may contribute in virulence and antibiotic resistance since indole producing ability can act as a signalling molecule to regulate the expression of adhesion and biofilm-promoting factors (<a href="#Martino-PD--Fursy-R--Bret-L--Sundararaju-B--Phillips-RS--2003-"><span class="Hyperlink">Martino et al., 2003</span></a>; <a href="#Nishida-M--Asano-H--Kamimura-T--Yokota-Y--1978-."><span class="Hyperlink">Nishida et al., 1978</span></a>). Other secondary biochemical characters in present study were found similar as described earlier literature (<a href="#Eguchi-M--Yokomizo-Y--Kuniyasu--C--1987"><span class="Hyperlink">Eguchi et al., 1987</span></a>; <span class="Hyperlink">Rennie and Duncan, 1974</span>). These similar biochemical characters may explain through previous studies, who reported similar biochemical properties of <span class="CharOverride-7">K. </span><span class="CharOverride-7">pneumoniae</span> from various sources without significant variations (<a href="#Bouvet-OMM--Lenormand-P--Grimont-PAD--1989-"><span class="Hyperlink">Bouvet et al., 1989</span></a>; <a href="#Munoz-MA--Welcome-FL--Schukken-YH--Zadoks-RN--2007-."><span class="Hyperlink">Munoz et al., 2007</span></a>). Thus it may also conclude that source of bacterial isolation not too much affects primary and secondary biochemical reaction of <span class="CharOverride-7">K. pneumoniae </span>isolates<span class="CharOverride-7">.</span></p>
			<p class="Body-Text">&nbsp;</p>
		  <p class="Body-Text">&nbsp;</p>
			<p class="Figure--and-Table-Heading"><span class="CharOverride-6"><a id="Table-2"></a>Table 2: </span>Urease production in <span class="CharOverride-7">Klebsiella pneumoniae </span>isolates from pneumonic and healthy camels</p>
			<table id="table-2" class="Table-Style-1">
				<colgroup>
					<col class="_idGenTableRowColumn-9" />
					<col class="_idGenTableRowColumn-10" />
					<col class="_idGenTableRowColumn-11" />
					<col class="_idGenTableRowColumn-12" />
					<col class="_idGenTableRowColumn-13" />
				</colgroup>
				<tbody>
					<tr class="_idGenTableRowColumn-14">
						<td rowspan="2">
							<p class="Basic-Paragraph"><span class="CharOverride-16">Observation day</span></p>
						</td>
						<td colspan="2">
							<p class="Basic-Paragraph ParaOverride-3"><span class="CharOverride-16">Pneumonic camels</span></p>
						</td>
						<td colspan="2">
							<p class="Basic-Paragraph ParaOverride-3"><span class="CharOverride-16">Healthy camels</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-15">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive isolates</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative isolates</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Positive isolates</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Negative isolates</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-6">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">First</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">37 (78.72%)</span></p>
						</td>
						<td rowspan="7">
							<p class="Basic-Paragraph ParaOverride-3"><span class="CharOverride-17">-</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">-</span></p>
						</td>
						<td rowspan="7">
							<p class="Basic-Paragraph ParaOverride-3"><span class="CharOverride-17">3 (16.66%)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Second</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">4 (8.51%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">-</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-16">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Third</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">3 (6.38%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">-</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-5">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Fourth</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">1 (2.12%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">4 (22.22%)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-17">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Fifth</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">2 (4.25%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">7 (38.55%)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-15">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Sixth</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph ParaOverride-1"><span class="CharOverride-17">-</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">3 (16.66%)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-6">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Seventh</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">-</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">1 (5.55%)</span></p>
						</td>
					</tr>
					<tr class="_idGenTableRowColumn-18">
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">Total</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">47 (100.0%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph ParaOverride-3"><span class="CharOverride-17">-</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">15 (83.33%)</span></p>
						</td>
						<td>
							<p class="Basic-Paragraph"><span class="CharOverride-17">3 (16.66%)</span></p>
						</td>
					</tr>
				</tbody>
			</table>
			
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
		  <p class="Body-Text ParaOverride-1">In accordance to present study, <a href="#Mobley-HLT--Warren-JW--1996-"><span class="Hyperlink">Mobley and Warren (1996)</span></a> and <a href="#Wilson-GS--Miles-A--1975-."><span class="Hyperlink">Wilson and Miles (1975)</span></a> also reported that clinical isolates of <span class="CharOverride-7">K. pneumoniae</span> were 100% urease producing. And present investigation also found that <span class="CharOverride-7">K. pneumoniae</span> isolates from pneumonic camels were faster urease producing as compared to isolates from healthy camels. Urease production enables the bacteria to grow in acidic environment (<a href="#Mobley-HLT--Warren-JW--1996-"><span class="Hyperlink">Mobley and Warren, 1996)</span></a>). Thus it has significant role in colonization, survival and pathogenesis of several bacterial species including <span class="CharOverride-7">K. pneumoniae </span>(<a href="#Burne-RA--Chen-YY--2000-."><span class="Hyperlink">Burne and Chen, 2000</span></a>; <a href="#Maroncle-N--Rich-C--Forestier-C--2006-.-Th"><span class="Hyperlink">Maroncle et al., 2006</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">Results of hypermucoviscosity trait in this study are almost comparable with finding of previous studies <span class="CharOverride-7">viz.,</span> <a href="#Yu-WL--Ko-WC--Cheng-KC--Lee-HC--Ke-DS--Lee-CC--Fung-CP--Chuang-YC--2006"><span class="Hyperlink">Yu et al. (2006)</span></a> reported 38% prevalence of virulent hypermucoviscosity trait in clinical isolates of <span class="CharOverride-7">K. pneumoniae.</span> And <a href="#Whitehouse-CA--Keirstead-N--Taylor-J--Reinhardt-JL--Beierschmitt-A--2010"><span class="Hyperlink">Whitehouse et al. (2010)</span></a> reported absence of virulent hypermucoviscosity trait in captive (apparently healthy) vervets. Thus the absence of virulent hypermucoviscosity trait among isolates from healthy may underset. In support of present study, <a href="#Edwards-PR--1928-"><span class="Hyperlink">Edwards (1928)</span></a> also reported that extensive capsule production is necessary to produce metritis in mares and <a href="#Simoons-Smit-AM--Verweiji-van-Vught--AMJJ--MacLaren-DM--1986-."><span class="Hyperlink">Simoons-Smit et al. (1986)</span></a> established the relationship between the size of capsule of <span class="CharOverride-7">K. pneumoniae </span>and pathogenicity in experimental lobar pneumonia. Since virulent hypermucoviscosity trait has results from overproduction of extra capsular polysaccharide mucoviscous web, it has significant role in virulence because these are resistant to phagocytosis by neutrophils and from serum killing by complement system and are commonly associated with community acquired pneumonia, bacteremia and distinct invasive syndromes such as primary liver abscesses, meningitis, and endophthalmitis (<a href="#Vila-A--Cassata-A--Pagella-H--Amadio-C--Yeh-KM--Chang-FY--Siu-LK--2011"><span class="Hyperlink">Vila et al., 2011</span></a>; <a href="#Lin-YT--Jeng-YY--Chen-TL--Fung-CP--2010-."><span class="Hyperlink">Lin et al., 2010</span></a>).</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">In the present study, we found some differences regarding indole production, urease production and virulent hypermucoviscosity trait of isolates from pneumonic and healthy camels. It may indicates the differences in virulence and suggest further genotypic characterization of isolates to find genotypic mechanisms of virulence of <span class="CharOverride-7">Klebsiella pneumoniae</span> to make proper diseases diagnosis and prevention.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">Acknowledgments</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			<p class="Body-Text ParaOverride-1">We acknowledge the support and facilities provide by Head of department of veterinary microbiology and biotechnology, Dean of college of veterinary and animal science, Bikaner and Dean of Post Graduate Institute of Veterinary Education and Research- Jaipur for this study.</p>
		  <p class="Body-Text ParaOverride-1">&nbsp;</p>
			<p class="Heading-1--Introduction----">References</p>
		  <p class="Heading-1--Introduction----">&nbsp;</p>
			
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