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Replication and transmission of H9N2 influenza viruses in ferrets: evaluation of pandemic potential.

Wan H, Sorrell EM, Song H, Hossain MJ, Ramirez-Nieto G, Monne I, Stevens J, Cattoli G, Capua I, Chen LM, Donis RO, Busch J, Paulson JC, Brockwell C, Webby R, Blanco J, Al-Natour MQ, Perez DR - PLoS ONE (2008)

Bottom Line: Five wild-type (WT) H9N2 viruses, isolated from different avian species from 1988 through 2003, were tested in vivo and found to replicate in ferrets.A leucine (Leu) residue at amino acid position 226 in the hemagglutinin (HA) receptor-binding site (RBS), responsible for human virus-like receptor specificity, was found to be important for the transmission of the H9N2 viruses in ferrets.Our results suggest that the establishment and prevalence of H9N2 viruses in poultry pose a significant threat for humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Medicine, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
H9N2 avian influenza A viruses are endemic in poultry of many Eurasian countries and have caused repeated human infections in Asia since 1998. To evaluate the potential threat of H9N2 viruses to humans, we investigated the replication and transmission efficiency of H9N2 viruses in the ferret model. Five wild-type (WT) H9N2 viruses, isolated from different avian species from 1988 through 2003, were tested in vivo and found to replicate in ferrets. However these viruses achieved mild peak viral titers in nasal washes when compared to those observed with a human H3N2 virus. Two of these H9N2 viruses transmitted to direct contact ferrets, however no aerosol transmission was detected in the virus displaying the most efficient direct contact transmission. A leucine (Leu) residue at amino acid position 226 in the hemagglutinin (HA) receptor-binding site (RBS), responsible for human virus-like receptor specificity, was found to be important for the transmission of the H9N2 viruses in ferrets. In addition, an H9N2 avian-human reassortant virus, which contains the surface glycoprotein genes from an H9N2 virus and the six internal genes of a human H3N2 virus, showed enhanced replication and efficient transmission to direct contacts. Although no aerosol transmission was observed, the virus replicated in multiple respiratory tissues and induced clinical signs similar to those observed with the parental human H3N2 virus. Our results suggest that the establishment and prevalence of H9N2 viruses in poultry pose a significant threat for humans.

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Replication and direct contact transmission of H9N2 viruses.Ferrets were inoculated intranasally (i.n.) with 106 TCID50 of H9N2 viruses RGWF10 (A), Dk/HK/Y280/97 (B), RGQa88 (C), Ck/HK/SF3/99 (D), and Ck/Jordan/554/03 (E). Twenty-four hours later, one naïve ferret (direct contact) was added to the same cage as each of the infected ferrets. Nasal washes were collected daily and were titrated in MDCK cells. Black, white and gray bars represent individual ferrets sampled and the amount of viral shedding at different days pi. The titers are expressed as log10 values of TCID50/ml with the limit of detection at 0.699 log10TCID50/ml. The dotted line was arbitrarily set at <0.3 log10TCID50/ml in order to represent samples below the detection limit. L and Q correspond to Leu226 and Gln226, respectively in the HA RBS.
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pone-0002923-g001: Replication and direct contact transmission of H9N2 viruses.Ferrets were inoculated intranasally (i.n.) with 106 TCID50 of H9N2 viruses RGWF10 (A), Dk/HK/Y280/97 (B), RGQa88 (C), Ck/HK/SF3/99 (D), and Ck/Jordan/554/03 (E). Twenty-four hours later, one naïve ferret (direct contact) was added to the same cage as each of the infected ferrets. Nasal washes were collected daily and were titrated in MDCK cells. Black, white and gray bars represent individual ferrets sampled and the amount of viral shedding at different days pi. The titers are expressed as log10 values of TCID50/ml with the limit of detection at 0.699 log10TCID50/ml. The dotted line was arbitrarily set at <0.3 log10TCID50/ml in order to represent samples below the detection limit. L and Q correspond to Leu226 and Gln226, respectively in the HA RBS.

Mentions: Virus was detected in the nasal washes from all of the inoculated ferrets, with peak titers ranging from 2.7 to 5.2 log10TCID50/ml (Fig. 1). The highest peak titer was achieved in the Dk/HK/Y280/97 group while the lowest was in the RGQa88 group. For the RGWF10 group, virus was detected in all the inoculated animals from 1 to 5 days pi and was transmitted to all direct contact ferrets, as demonstrated by the detection of virus in nasal washes using FLU DETECT™ Antigen Capture Test Strip (data not shown), and viral titration (Fig. 1A). The direct contact animals began to shed detectable levels of virus by day 4 and 5 post-contact (pc), and each shed virus for 4 to 6 days, with peak titers comparable to those found in the inoculated ferrets (Fig. 1A). Anti-H9 antibodies were detected in all ferrets, with hemagglutination inhibition (HI) titers of 1280 in the inoculated ferrets, and 320 to 640 in the contact ferrets. Virus was detected in both Dk/HK/Y280/97-inoculated ferrets for 6 days and transmitted to 1 of the 2 direct contacts, which shed virus for 5 days (Fig. 1B). In the RGWF10 and Dk/HK/Y280/97 groups, the viral positive contacts exhibited lethargy, anorexia, weight loss and elevated temperature similar to the inoculated ferrets. The RGQa88, Ck/HK/SF3/99 and Ck/Jordan/554/03 groups shed viruses for up to 7 days (Figs. 1C, D, and E), and developed high titers (640–1280) of H9 antibodies (Table 2). However, neither virus shedding nor seroconversion was detected in any of the contact ferrets (Figs. 1C, D, E and Table 3), reflecting the lack of direct contact transmission. These results suggest that the ferret model is able to recapitulate the infection of H9N2 viruses as observed in humans and pigs. Our findings suggest that the ferret represents a good animal model to study the potential changes that could lead to efficient transmission of avian H9N2 viruses in humans.


Replication and transmission of H9N2 influenza viruses in ferrets: evaluation of pandemic potential.

Wan H, Sorrell EM, Song H, Hossain MJ, Ramirez-Nieto G, Monne I, Stevens J, Cattoli G, Capua I, Chen LM, Donis RO, Busch J, Paulson JC, Brockwell C, Webby R, Blanco J, Al-Natour MQ, Perez DR - PLoS ONE (2008)

Replication and direct contact transmission of H9N2 viruses.Ferrets were inoculated intranasally (i.n.) with 106 TCID50 of H9N2 viruses RGWF10 (A), Dk/HK/Y280/97 (B), RGQa88 (C), Ck/HK/SF3/99 (D), and Ck/Jordan/554/03 (E). Twenty-four hours later, one naïve ferret (direct contact) was added to the same cage as each of the infected ferrets. Nasal washes were collected daily and were titrated in MDCK cells. Black, white and gray bars represent individual ferrets sampled and the amount of viral shedding at different days pi. The titers are expressed as log10 values of TCID50/ml with the limit of detection at 0.699 log10TCID50/ml. The dotted line was arbitrarily set at <0.3 log10TCID50/ml in order to represent samples below the detection limit. L and Q correspond to Leu226 and Gln226, respectively in the HA RBS.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2500216&req=5

pone-0002923-g001: Replication and direct contact transmission of H9N2 viruses.Ferrets were inoculated intranasally (i.n.) with 106 TCID50 of H9N2 viruses RGWF10 (A), Dk/HK/Y280/97 (B), RGQa88 (C), Ck/HK/SF3/99 (D), and Ck/Jordan/554/03 (E). Twenty-four hours later, one naïve ferret (direct contact) was added to the same cage as each of the infected ferrets. Nasal washes were collected daily and were titrated in MDCK cells. Black, white and gray bars represent individual ferrets sampled and the amount of viral shedding at different days pi. The titers are expressed as log10 values of TCID50/ml with the limit of detection at 0.699 log10TCID50/ml. The dotted line was arbitrarily set at <0.3 log10TCID50/ml in order to represent samples below the detection limit. L and Q correspond to Leu226 and Gln226, respectively in the HA RBS.
Mentions: Virus was detected in the nasal washes from all of the inoculated ferrets, with peak titers ranging from 2.7 to 5.2 log10TCID50/ml (Fig. 1). The highest peak titer was achieved in the Dk/HK/Y280/97 group while the lowest was in the RGQa88 group. For the RGWF10 group, virus was detected in all the inoculated animals from 1 to 5 days pi and was transmitted to all direct contact ferrets, as demonstrated by the detection of virus in nasal washes using FLU DETECT™ Antigen Capture Test Strip (data not shown), and viral titration (Fig. 1A). The direct contact animals began to shed detectable levels of virus by day 4 and 5 post-contact (pc), and each shed virus for 4 to 6 days, with peak titers comparable to those found in the inoculated ferrets (Fig. 1A). Anti-H9 antibodies were detected in all ferrets, with hemagglutination inhibition (HI) titers of 1280 in the inoculated ferrets, and 320 to 640 in the contact ferrets. Virus was detected in both Dk/HK/Y280/97-inoculated ferrets for 6 days and transmitted to 1 of the 2 direct contacts, which shed virus for 5 days (Fig. 1B). In the RGWF10 and Dk/HK/Y280/97 groups, the viral positive contacts exhibited lethargy, anorexia, weight loss and elevated temperature similar to the inoculated ferrets. The RGQa88, Ck/HK/SF3/99 and Ck/Jordan/554/03 groups shed viruses for up to 7 days (Figs. 1C, D, and E), and developed high titers (640–1280) of H9 antibodies (Table 2). However, neither virus shedding nor seroconversion was detected in any of the contact ferrets (Figs. 1C, D, E and Table 3), reflecting the lack of direct contact transmission. These results suggest that the ferret model is able to recapitulate the infection of H9N2 viruses as observed in humans and pigs. Our findings suggest that the ferret represents a good animal model to study the potential changes that could lead to efficient transmission of avian H9N2 viruses in humans.

Bottom Line: Five wild-type (WT) H9N2 viruses, isolated from different avian species from 1988 through 2003, were tested in vivo and found to replicate in ferrets.A leucine (Leu) residue at amino acid position 226 in the hemagglutinin (HA) receptor-binding site (RBS), responsible for human virus-like receptor specificity, was found to be important for the transmission of the H9N2 viruses in ferrets.Our results suggest that the establishment and prevalence of H9N2 viruses in poultry pose a significant threat for humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Medicine, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
H9N2 avian influenza A viruses are endemic in poultry of many Eurasian countries and have caused repeated human infections in Asia since 1998. To evaluate the potential threat of H9N2 viruses to humans, we investigated the replication and transmission efficiency of H9N2 viruses in the ferret model. Five wild-type (WT) H9N2 viruses, isolated from different avian species from 1988 through 2003, were tested in vivo and found to replicate in ferrets. However these viruses achieved mild peak viral titers in nasal washes when compared to those observed with a human H3N2 virus. Two of these H9N2 viruses transmitted to direct contact ferrets, however no aerosol transmission was detected in the virus displaying the most efficient direct contact transmission. A leucine (Leu) residue at amino acid position 226 in the hemagglutinin (HA) receptor-binding site (RBS), responsible for human virus-like receptor specificity, was found to be important for the transmission of the H9N2 viruses in ferrets. In addition, an H9N2 avian-human reassortant virus, which contains the surface glycoprotein genes from an H9N2 virus and the six internal genes of a human H3N2 virus, showed enhanced replication and efficient transmission to direct contacts. Although no aerosol transmission was observed, the virus replicated in multiple respiratory tissues and induced clinical signs similar to those observed with the parental human H3N2 virus. Our results suggest that the establishment and prevalence of H9N2 viruses in poultry pose a significant threat for humans.

Show MeSH
Related in: MedlinePlus