influenza a is one of the most important infectious diseases of humans and is responsible for recurrent seasonal disease epidemics . wild birds are the natural reservoir for all recognized subtypes of influenza a and as such present a potential route for the emergence of new viral strains which can cause human disease . if such newly emerged viruses acquire the ability to transfer effectively from human to human they may lead to pandemic outbreaks with widespread illness and mortality . the h1n1 influenza pandemic of 1918 caused 20 to 50 million deaths on a global scale , making it the single most devastating disease outbreak in human history ( johnson and muller , 2002 ) . this pandemic is believed to have been caused by an avian virus that crossed the species barrier to infect humans and underwent subsequent adaptation to a new host ( taubenberger et al , 2005 ) . the resultant virus acquired exceptional virulence with the ability to replicate in the absence of trypsin , induce death in mice and grow rapidly in human epithelial cells ( tumpey et al , 2005 ) . novel influenza viruses may also arise due to concurrent infection with different virus strains through reassortment of viral rna segments ( horimoto and kawawoka , 2005 ) . reassortment of human and avian influenza a viruses is thought to have generated the pandemic viruses of 1957 and 1968 ( webster et al , 1997 ) . avian influenza viruses are therefore likely to continue to play a significant role in the emergence of new human influenza strains ( perdue and swayne , 2005 ) . more recently , human infections have been caused by the emergence of avian h5n1 viruses , initially in hong kong in 1997 ( claas et al , 1998 ; subbarao et al , 1998 ) . subsequently , h5n1 has re - emerged as a significant threat to human health , with over 385 confirmed cases and 243 deaths ( who data , june 2008 ) . these cases have demonstrated that avian viruses can directly infect humans without the need of intermediate hosts such as pigs . however , the precise molecular basis for the efficient transmissibility of avian influenza viruses to mammals is not fully understood . a major determinant of the ability of influenza viruses to infect cells is the expression of the appropriate host cell receptor to which viral haemagglutinin can bind . consequently , a crucial hurdle that influenza a viruses need to overcome when crossing a species barrier is the acquisition of the ability to utilize alternate host cell receptors ( ito , 2000 ) . influenza virus receptors on host cells are glycosylated oligosaccharides that terminate in sialic acid ( sa ) residues which are bound to glycans through an 2,3 or 2,6 linkage , mediated by sialyltransferases that are expressed in a cell- and species - specific manner ( gagneux et al , 2003 ) . avian influenza viruses have been shown to preferentially bind to sa receptors that are linked to galactose by an 2,3 linkage ( sa2,3-gal ) , while human and classical swine viruses show preference for receptors with an 2,6 linkage ( sa2,6-gal ) ( gagneux et al , 2003 ; matrosovich et al , 2004 ) . among avian influenza viruses , chicken and duck viruses have been shown to further differ in their ability to recognize the structure of the third sugar moiety in sa2,3-gal terminated receptors . a collection of avian influenza viruses from chickens and ducks has been studied for their binding affinities to synthetic receptor analogues in vitro . a marked contrast in preferential binding was noted , where influenza viruses from chickens preferentially bound to synthetic sialylglycopolymer containing neu5ac-(2 - 3)gal-(1 - 4)glcnac , whereas viruses from ducks displayed a higher affinity for neu5ac-(2 - 3)gal-(1 - 3)galnac containing polymer ( gambaryan et al , 2003 ) . ducks and chickens are the major aquatic and terrestrial hosts for a wide variety of influenza viruses and are valuable natural models to study influenza , with ducks displaying resistance to disease , but chickens , in contrast , showing relatively high susceptibility . receptors are important determinants of virus entry and differences in receptor distribution between hosts could therefore account for variation in susceptibility to infection , including the contrasting outcomes following infection with many strains of highly pathogenic avian influenza ( hpai ) . influenza virus receptors in chickens and ducks have previously been studied by virus binding assays using extracted total gangliosides from plasma membranes of respiratory and intestinal epithelial cells ( gambaryan et al , 2002 ) . based on the binding of human influenza virus with sa2,6-gal receptor specificity , it was found that chicken epithelial cells , but not duck epithelial cells , express sa2,6-gal receptors . however , the receptor distribution across intact mucosal surfaces and organ systems was not studied . more recently , chicken trachea and duodenum have been studied using lectin binding assays followed by conventional immunohistochemistry and light microscopy ( wan and perez , 2006 ) . despite this , detailed information on the distribution of influenza virus receptors in these important avian hosts is lacking . to further evaluate the potential role of receptor distribution in ducks and chickens in determining susceptibility to infection , and the potential of these species to support infection of viruses with tropism for sa2,6-gal and sa2 - 3-gal ( and therefore act as  mixing vessels  ) we have examined in detail the anatomical distribution of influenza a receptors in key organs of both species . animals for this study comprised four 35 - 40 wk old , commercial layer chickens ( glenrath farms ltd , east lothian , uk ) , four 4 wk old broilers ( pd hook hatcheries , bampton , oxfordshire , uk ) , two 3 wk old and four 6 wk old pekin ducks ( cherry valley farms , rothwell , lincolnshire , uk ) . the animals were euthanased and samples from trachea , lungs , heart , kidney , brain , skeletal muscle , small and large intestine collected into buffered neutral formalin . tissue samples were dehydrated and cleared using a histokinette ( leica tp 1020 ) before being embedded in paraffin wax . after embedding , the tissues were sectioned using a rotary microtome ( leica rm 2255 ) with a specimen feed of 5m . lectin histochemistry using linkage specific lectins was carried out with minor modifications of a method described previously ( shinya et al , 2006 ) . lectins used in the study were : sambucus nigra agglutinin ( sna ) specific for sa2,6-gal ( shibuya et al , 1987 ) , maackia amurensis i ( maa i ) and maackia amurensis agglutinins ( maa ii ) which are specific for sa2,3-gal(1 - 4)glcnac and sa2,3-gal(1 - 3)galnac respectively ( konami et al , 1994 ) ( all provided by vector laboratories , burlingame , ca ) . sections were pre - soaked in tbs and blocked using a biotin - streptavidin blocking kit ( vector laboratories ) according to manufacturer 's instructions , followed by 4c overnight incubation with fluorescein isothiocyanate ( fitc ) labelled sna or fitc labelled maa i , and biotinylated maa ii lectin each at a concentration of 10g / ml . after three washes with tbs , the sections were incubated with streptavidin - alexa - fluor594 conjugate ( molecular probes inc , eugene , or ) for 2 hrs at room temperature ( rt ) . the sections were washed and then mounted with prolong gold antifade reagent with 4 , 6-diamino-2- phenylindole , dihydro - chloride ( dapi ; molecular probes inc , eugene , or ) . negative controls were performed omitting the primary reagents . to rule out nonspecific binding of the lectins , tissue sections were treated , prior to lectin staining , with sialidase a ( n - acetylneuraminate glycohydrolase ; prozyme , san leandro , ca ) , which cleaves all non - reducing terminal sialic acid residues in the order (2,6 ) > (2,3 ) > differences in receptor distribution on the mucosal lining of tracheae or intestines were quantified using lcs lite software . mean energy values were measured for each fluorochrome in a representative area of the mucosa and corrected by subtracting background energy values . receptor binding assays with h1n1 classical swine strain ( a / sw / iowa/15/30 ) , a subtype closely related to the human 1918 pandemic influenza virus ( tumpey et al , 2004 ) , and a h2n3 low pathogenic avian strain ( a / mallard duck / england/7277/06 ) were performed by a previously reported method , with minor modifications ( couceiro et al , 1993 ) . briefly , paraffin embedded 5m sections of chicken and duck tracheae , small and large intestines were deparaffinised in xylene and rehydrated by alcohol . the sections were washed , blocked with goat serum , and incubated with mouse monoclonal antibody to influenza nucleoprotein ( abcam , cambridge , uk ) at 1:1000 dilution for one hour at rt followed by fitc - labelled goat anti - mouse igg ( abcam , cambridge , uk ) at 1:500 dilution for two hours at rt . after three further washes with tbs , the sections were mounted with prolong gold antifade reagent with dapi and scanned by confocal microscopy ( leica tcs sp2 aobs ) . animals for this study comprised four 35 - 40 wk old , commercial layer chickens ( glenrath farms ltd , east lothian , uk ) , four 4 wk old broilers ( pd hook hatcheries , bampton , oxfordshire , uk ) , two 3 wk old and four 6 wk old pekin ducks ( cherry valley farms , rothwell , lincolnshire , uk ) . the animals were euthanased and samples from trachea , lungs , heart , kidney , brain , skeletal muscle , small and large intestine collected into buffered neutral formalin . tissue samples were dehydrated and cleared using a histokinette ( leica tp 1020 ) before being embedded in paraffin wax . after embedding , the tissues were sectioned using a rotary microtome ( leica rm 2255 ) with a specimen feed of 5m . lectin histochemistry using linkage specific lectins was carried out with minor modifications of a method described previously ( shinya et al , 2006 ) . lectins used in the study were : sambucus nigra agglutinin ( sna ) specific for sa2,6-gal ( shibuya et al , 1987 ) , maackia amurensis i ( maa i ) and maackia amurensis agglutinins ( maa ii ) which are specific for sa2,3-gal(1 - 4)glcnac and sa2,3-gal(1 - 3)galnac respectively ( konami et al , 1994 ) ( all provided by vector laboratories , burlingame , ca ) . sections were pre - soaked in tbs and blocked using a biotin - streptavidin blocking kit ( vector laboratories ) according to manufacturer 's instructions , followed by 4c overnight incubation with fluorescein isothiocyanate ( fitc ) labelled sna or fitc labelled maa i , and biotinylated maa ii lectin each at a concentration of 10g / ml . after three washes with tbs , the sections were incubated with streptavidin - alexa - fluor594 conjugate ( molecular probes inc , eugene , or ) for 2 hrs at room temperature ( rt ) . the sections were washed and then mounted with prolong gold antifade reagent with 4 , 6-diamino-2- phenylindole , dihydro - chloride ( dapi ; molecular probes inc , eugene , or ) . negative controls were performed omitting the primary reagents . to rule out nonspecific binding of the lectins , tissue sections were treated , prior to lectin staining , with sialidase a ( n - acetylneuraminate glycohydrolase ; prozyme , san leandro , ca ) , which cleaves all non - reducing terminal sialic acid residues in the order (2,6 ) > (2,3 ) > differences in receptor distribution on the mucosal lining of tracheae or intestines were quantified using lcs lite software . mean energy values were measured for each fluorochrome in a representative area of the mucosa and corrected by subtracting background energy values . receptor binding assays with h1n1 classical swine strain ( a / sw / iowa/15/30 ) , a subtype closely related to the human 1918 pandemic influenza virus ( tumpey et al , 2004 ) , and a h2n3 low pathogenic avian strain ( a / mallard duck / england/7277/06 ) were performed by a previously reported method , with minor modifications ( couceiro et al , 1993 ) . briefly , paraffin embedded 5m sections of chicken and duck tracheae , small and large intestines were deparaffinised in xylene and rehydrated by alcohol . the sections were washed , blocked with goat serum , and incubated with mouse monoclonal antibody to influenza nucleoprotein ( abcam , cambridge , uk ) at 1:1000 dilution for one hour at rt followed by fitc - labelled goat anti - mouse igg ( abcam , cambridge , uk ) at 1:500 dilution for two hours at rt . after three further washes with tbs , the sections were mounted with prolong gold antifade reagent with dapi and scanned by confocal microscopy ( leica tcs sp2 aobs ) . we conducted an extensive examination of influenza virus receptor distribution in a range of tissues from chickens and ducks . no difference in the reported results was observed due to the age or source of animals , and the receptor distribution was consistent between individual animals within each species . using lectin staining , we found widespread presence of both sa2,6-gal ( sna ) and sa2,3-gal ( maa ii ) receptors in a range of tissues from each species , suggesting that these organs may be potential targets for both avian and human influenza viruses ( figure 1 ) . the expression of sa2,6-gal receptors in duck tissues is in contrast to a previous study using virus binding assays which reported that plasma membranes isolated from duck respiratory and intestinal epithelial cells did not express sa2,6-gal - terminated receptors ( gambaryan et al , 2002 ) . the exact reason for this difference is not clear , however the previous study used mallard ducks rather than pekin ducks used in this study , and the results were based on virus , rather than lectin binding , to isolated cell membranes rather than intact cells . thus it is possible that a difference in the methodology or choice of duck breed may be responsible . although there is widespread distribution of both receptor types in both chickens and ducks , there are clear differences in their spatial distribution within organs between the two species . in the vascular endothelium of the kidney , both sa2,6-gal and sa2,3-gal receptors were found in duck cells , but only the sa2,6-gal receptor type was found in the corresponding region in chicken kidney . the significance of this difference is not clear , however presence of sa2,6-gal receptors in the vascular endothelium in both species indicates that these cells can be potentially infected by mammalian - like influenza viruses and possibly play a role in the haematogenous spread of the virus . tubular cells of duck kidney expressed both sa2,6-gal and sa2,3-gal receptors , whereas chicken kidney tubular cells expressed either sa2,6-gal or sa2,3-gal receptors , but no co - expression was observed . the endocardium , meninges and muscle fibres of both species co - expressed both subtypes of receptor ( figure 1 ) . the expression of both host receptors in such tissues in chickens and ducks suggests that these avian hosts could possibly serve as  mixing vessels  for virus reassortment following co - infection by human and avian viruses . chicken and duck intestine predominantly expressed the sa2,3-gal receptor type across the epithelial lining of villi ( figure 2a ) , as previously reported ( ito and kawaoka , 2000 ; wan and perez , 2006 ) . the measurement of fluorescent mean energy values along the epithelial lining of the mucosa showed no significant presence of sa2,6-gal in intestines of either chicken or duck . this finding reaffirms the avian digestive tract as a major predilection site for avian influenza virus replication . the receptor distribution in the large intestine of both avian species was very similar to the small intestine ( data not shown ) . the major species difference that we observed between chicken and duck in the relative distribution of sa2,3-gal and sa2,6-gal receptors was along the tracheal epithelium . in chicken tracheal epithelium , sa2,6-gal ( with sna binding ) was the dominant receptor type whereas in ducks the sa2,3-gal receptor ( with maa ii binding ) was more abundant in the ciliated cells of the tracheal epithelium ( figure 2b ) . based on mean fluorescent energy values , it was found that the ratio of sa2,6-gal to sa2,3-gal in chicken trachea was approximately 10:1 whereas in duck the ratio was 1:20 . the observed difference in dominant receptor type between chickens and ducks was confined to the upper airway ( trachea ) . in the bronchi and chicken alveolar cells expressed both receptor types ; the precise staining pattern was more difficult to determine in duck alveoli due to the extensive presence of vascular tissue which tended to obscure the alveolar staining , however , both receptors could be seen clearly in some areas of the alveoli ( figure 1 ) . the dominant sa2,6-gal receptor expression pattern in chicken trachea was in contrast to a previous study ( wan and perez , 2006 ) which , using lectin binding , found that 85% of the epithelial cells in chicken trachea were positive for sa2,3-gal receptors , while only 10% were positive for sa2,6-gal receptors . however , it is in agreement with the findings of gambaryan et al ( 2002 ) , who reported that human influenza viruses with sa2,6-gal specificity bound to cell membranes isolated from chicken ( but not duck ) tracheal cell membranes . a possible explanation for the discrepancy in the reported receptor distribution in chicken trachea could be the source of the lectin used . lectins from different suppliers may show different binding specificities ; in particular the source of maa has been shown to significantly affect specificity ( nicholls et al , 2007 ) . glycan microarray screening of lectins from the supplier used for our study confirms that these lectins bind with a high degree of specificity to the appropriate sialic acid linkages ( http://www.functionalglycomics.org ) . it is also possible that this study was able to detect lower levels of receptor expression due to the methodology used ( confocal microscopy with fluorescent detection compared with immunohistochemistry ) . we found that the observed predominance of sa2,6-gal in chicken trachea was consistent in different ages and breed of chicken . we also performed lectin staining on sialidase treated sections , which abrogated all staining confirming that the lectins used in the present study did not bind to non - sialic acid residues ( data not shown ) . the difference in the predominant receptor across the tracheal epithelial lining in chicken and ducks could be an important contributing factor to influenza virus entry via the upper respiratory tract . in particular , such differences could impact on the susceptibility of each species to avian h5n1 influenza with its preferential tropism for infection of the respiratory tract rather than the intestines . the predominant receptor in the human upper airway is sa2,6-gal ( shinya et al , 2006 ; yao et al , 2007 ) . the high levels of expression of sa2,6-gal receptors in chicken tracheal epithelium suggests that this species may be more able to support the evolution of viruses with higher affinity for human sa2,6-gal receptors . this supports the suggestion that chickens may the source of emerging h9n2 viruses with a human - virus like receptor specificity ( gambaryan et al , 2002 ) . widespread presence of sa2,6 gal ( sna lectin ) and sa2,3 gal ( maa ii lectin ) receptors in many of organs of chickens ( a ) and ducks ( b ) . composite confocal images show distribution of sa2,6 gal receptors ( green ) and sa2,3 gal receptors ( red ) with nuclear staining ( blue ) . differential expression of sa2,6 gal and sa2,3gal receptors in tracheae and small intestine ( si ) of chickens and ducks . ( a ) for both avian species , the epithelial lining of the si shows predominant expression of sa2,3gal receptor ( red ) , with little evidence of sa2,6 gal ( green ) expression . host receptor distribution is similar for both si and large intestines ( data not shown ) . ( b ) sa2,6gal receptor is the more dominant type in chicken tracheal epithelium whereas sa2,3gal receptor is the main type in duck tracheal epithelium . we further distinguished sa2,3-gal receptor subtypes , based on the third sugar residue , in chicken and duck trachea and intestines , with the use of maa i ( sa2,3-gal (1 - 4)glcnac specificity ) and maa ii ( sa2,3-gal (1 - 3)galnac specificity ) . in chicken trachea , both sa2,3-gal receptor subtypes were detected in the sub - epithelial region ( figure 3a ) . however , along the chicken tracheal epithelium , sa2,3-gal (1 - 4)glc nac receptor ( maa i lectin ) was more dominant than sa2,3-gal (1 - 3 ) galnac receptor ( maa ii lectin ) . in duck trachea , in contrast , minimal maa i lectin binding was observed along the epithelium ; only sub - epithelial mucous glands were maa i positive . in duck trachea , sa2,3-gal (1 - 3 ) galnac receptor ( maa ii lectin ) was the main subtype detected , with distribution along the epithelial lining and in the mucosa ( figure 3a ) . in chicken large intestine , both sa2,3-gal 1 - 4 gal nac receptor ( maa i lectin ) and sa2,3-gal 1 - 3 glc nac receptor ( maa ii lectin ) expression was observed . in duck large intestine , sa2,3-gal 1 - 3 glc nac receptor ( maa ii lectin ) was the main subtype detected , while the goblet cells were positive for sa2,3-gal (1 - 4 ) glc nac receptor subtype ( maa i lectin ) ( figure 3b ) . the binding pattern of maa i and maa ii in the small intestines of both avian species was very similar to the large intestine ( data not shown ) . in humans , maa i shows more widespread binding throughout the upper and lower respiratory tract compared to maa ii ( nicholls et al , 2007 ) . avian influenza viruses isolated from chicken and ducks have been shown to preferentially bind to sa 2,3-gal (1 - 4 ) glc nac ( recognized by maa i ) and sa(2,3 ) gal (1 - 3 ) gal nac ( recognized by maa ii ) respectively ( gambaryan et al , 2003 ) . this reported virus tropism correlates with the observed receptor distribution in chicken and duck trachea and intestinal tissues . the presence of both sa2 - 3-gal receptor subtypes in chickens suggests that they may be susceptible to infection with wider range of avian influenza viruses with broader receptor specificity . maa i ( sa(2,3)-gal (1 - 4 ) glc nac detection ; green ) and maa ii ( sa(2,3)-gal (1 - 3 ) gal nac detection ; red ) were used to distinguish sa(2,3)-gal receptor subtypes at the third sugar residue position . ( a ) chicken trachea expresses sa(2,3)-gal (1 - 4 ) glc nac receptor ( maa i lectin ) more strongly along the epithelial lining than sa(2,3 ) gal (1 - 3 ) gal nac ( maa ii lectin ) . both sa(2,3)-gal receptor subtypes are clearly present in the sub - epithelial region ( see also maa ii lectin staining in fig . minimal sa (2,3)-gal (1 - 4 ) glc nac receptor subtype ( maa i lectin ) is detected along the epithelium . sa (2,3)-gal (1 - 3 ) gal nac receptor ( maa ii lectin ) is the main subtype with distribution along the epithelial lining and in the mucosa ( see also maa ii lectin staining in fig . ( b ) chicken large intestine expresses similar levels of both sa(2,3)-gal 1 - 4 gal nac receptor ( maa i lectin ) and sa(2,3)-gal (1 - 3 ) gal nac ( maa ii lectin ) along the epithelial lining . in duck large intestine , sa (2,3)-gal (1 - 3 ) gal nac receptor ( maa ii lectin ) is the main subtype with distribution along the epithelial lining and in the mucosa . similar observations were made in small intestines of chicken and duck ( data not shown ) . to relate the observed receptor distribution with the ability to bind viruses of avian or mammalian origin , we performed virus binding assays with avian h2n3 and swine h1n1 influenza viruses on tracheal and digestive tract sections . this showed the predicted preferential binding of the avian virus for sa2,3-gal receptor and the swine virus for sa2,6-gal receptors ( figure 4 ) . the main sa2,3-gal receptor type in duck tracheal epithelium showed greater affinity for the avian h2n3 virus . in contrast , the dominant receptor type of sa2,6-gal in chicken trachea showed preferential binding of the swine h1n1 virus . the predominant expression of sa2,3-gal receptors type along the small and large intestinal epithelia of chickens and ducks showed preferential affinity for the avian h2n3 virus with no significant attachment of the swine virus . virus - binding specificity was therefore consistent with host receptor type , as determined by lectin staining . the sa2,6-gal receptor type expressed by the intestinal goblet cells did not appear seem to be functionally significant as no virus binding was observed with swine h1n1 virus ( figure 4 ) . virus binding assays with avian h2n3 and swine h1n1 influenza viruses on tracheal and intestinal tract sections reflect the observed receptor distribution . the main sa(2,3)-gal receptor type in duck tracheal epithelium shows greater affinity for the avian virus than the swine virus . by contrast , the dominant sa(2,6)-gal receptor type in chicken trachea shows preferential binding of the swine virus . the primary sa(2,3)-gal receptor type along the small intestine ( si ) epithelia of both chickens and ducks is associated with preferential affinity for the avian virus , with no significant binding of swine virus . the differences in receptor expression reported in this study suggest that they may be responsible , at least in part , for some of the differences between ducks and chickens in the pattern of disease following influenza infection . while the presence of a virus receptor is clearly not sufficient to confirm that cells or tissue support efficient virus replication or transmission , the widespread replication of influenza virus in multiple organs has been reported in both chickens ( swayne , 1997 ) and ducks ( londt et al , 2008 ) following infection with highly pathogenic viruses . genetic studies have revealed that previous pandemic influenza strains were partially or entirely derived from the viruses of avian origin ( webster et al , 1992 and taubenberger et al , 1997 , horimoto and kawawoka 2005 ) . this study suggests that some chicken and duck tissues may facilitate entry of both human and avian viruses , with the ensuing danger of virus reassortment . however , further work is required to confirm that the tissues expressing both receptor types are able to support virus replication . the dominant presence of sa(2,6)gal receptor along the chicken tracheal epithelium shows some similarities to the prevalence of the receptor in mammals such as human and pig . this suggests that chickens may be important intermediate hosts for the transmission of influenza to humans , in particular for influenza viruses such as h5n1 , which show a respiratory tropism in birds . whilst much attention has been placed on the role of pigs as  mixing vessels  , the potential importance of chickens for the evolution of humanised influenza viruses should not be overlooked and , as such , warrants further study . both sa2,3-gal ( avian ) and sa2,6-gal ( human ) receptors are expressed in many tissues of chickens and ducks . sa2,6-gal receptor is the dominant receptor type in chicken tracheal epithelium , whereas in ducks the sa2,3-gal receptor is dominant . chicken trachea and intestinal tissues showed positive binding with maa i to maa ii whereas positive maa ii binding alone was noted across the epithelial lining in ducks . this suggests that chicken may be susceptible to infection with avian influenza viruses with broader receptor specificity . the host receptor distribution pattern in the chicken upper respiratory tract may be functionally significant for the evolution of viruses with a human like receptor specificity and thus for the transmission of influenza from birds and mammals .
avian influenza viruses are considered to be key contributors to the emergence of human influenza pandemics . 
 a major determinant of infection is the presence of virus receptors on susceptible cells to which the viral haemagglutinin is able to bind . 
 avian viruses preferentially bind to sialic acid 2,3-galactose ( sa2,3-gal ) linked receptors , whereas human strains bind to sialic acid 2,6-galactose ( sa2,6-gal ) linked receptors . 
 while ducks are the major reservoir for influenza viruses , they are typically resistant to the effects of viral infection , in contrast to the frequently severe disease observed in chickens . in order to understand whether differences in receptors might contribute to this observation 
 , we studied the distribution of influenza receptors in organs of ducks and chickens using lectin histochemistry with linkage specific lectins and receptor binding assays with swine and avian influenza viruses . 
 although the intestinal epithelial cells of both species expressed only sa2,3-gal receptors , we found widespread presence of both sa2,6-gal and sa2,3-gal receptors in many organs of both chickens and ducks . 
 co - expression of both receptors may allow infection of cells with both avian and human viruses and so present a route to genetic reassortment . 
 there was a marked difference in the primary receptor type in the trachea of chickens and ducks . in chicken trachea , 
 sa2,6-gal was the dominant receptor type whereas in ducks sa2,3-gal receptors were most abundant . 
 this suggests that chickens could be more important as an intermediate host for the generation of influenza viruses with increased ability to bind to sa2,6-gal receptors and thus greater potential for infection of humans . 
 chicken tracheal and intestinal epithelial cells also expressed a broader range of sa2,3-gal receptors ( both (1 - 4)glcnac and (1 - 3)galnac subtypes ) in contrast to ducks , which suggests that they may be able to support infection with a broader range of avian influenza viruses .