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Experimental transmission of Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) to SCID-hu Thy/Liv mice.

Dittmer D, Stoddart C, Renne R, Linquist-Stepps V, Moreno ME, Bare C, McCune JM, Ganem D - J. Exp. Med. (1999)

Bottom Line: Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) is a novel human lymphotropic herpesvirus linked to several human neoplasms.KSHV virions were inoculated directly into the implants, and viral DNA and mRNA production was assayed using real-time quantitative polymerase chain reaction.Viral gene expression was most abundant in CD19(+) B lymphocytes, suggesting that this model faithfully mimics the natural tropism of this virus.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of California, San Francisco, California 94143, USA.

ABSTRACT
Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) is a novel human lymphotropic herpesvirus linked to several human neoplasms. To date, no animal model for infection by this virus has been described. We have examined the susceptibility of C.B-17 scid/scid mice implanted with human fetal thymus and liver grafts (SCID-hu Thy/Liv mice) to KSHV infection. KSHV virions were inoculated directly into the implants, and viral DNA and mRNA production was assayed using real-time quantitative polymerase chain reaction. This revealed a biphasic infection, with an early phase of lytic replication accompanied and followed by sustained latency. Ultraviolet irradiation of the inoculum abolished all DNA- and mRNA-derived signals, and infection was inhibited by ganciclovir. Viral gene expression was most abundant in CD19(+) B lymphocytes, suggesting that this model faithfully mimics the natural tropism of this virus. Short-term coinfection with HIV-1 did not alter the course of KSHV replication, nor did KSHV alter levels of HIV-1 p24 during the acute phase of the infection. Although no disease was evident in infected animals, SCID-hu Thy/Liv mice should allow the detailed study of KSHV tropism, latency, and drug susceptibility.

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Analysis of transcript levels in infected implants. A and B show latent and lytic transcript levels expressed as the number of latent or lytic cells per 106 cells (logarithmic scale) for implants at day 14 after inoculation that were infected with KSHV (KSHV), infected with KSHV and treated with ganciclovir (KSHV ganc.), or infected with UV-inactivated virus (KSHV UV); BCBL-1 cells are shown as control. The median of each group is indicated by the dashed line. P values were calculated using the nonparametric Mann-Whitney test.
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Figure 5: Analysis of transcript levels in infected implants. A and B show latent and lytic transcript levels expressed as the number of latent or lytic cells per 106 cells (logarithmic scale) for implants at day 14 after inoculation that were infected with KSHV (KSHV), infected with KSHV and treated with ganciclovir (KSHV ganc.), or infected with UV-inactivated virus (KSHV UV); BCBL-1 cells are shown as control. The median of each group is indicated by the dashed line. P values were calculated using the nonparametric Mann-Whitney test.

Mentions: Fig. 5 shows relative transcript levels in infected implants at day 14 after inoculation. Total RNA was isolated from individual implants in each group (implants injected with KSHV, implants injected with UV-inactivated KSHV, and implants injected with KSHV and treated with ganciclovir) and assayed in duplicate. To normalize for the amount of RNA in each reaction, rRNA was coamplified and quantified using a differently labeled probe (primers riboF, riboR, and riboP; Table ). Ct values for KSHV-specific probes were subtracted from corresponding Ct values for rRNA and expressed as the number of latent or lytic cells per 106 cells (based on BCBL-1 control). As expected, implants injected with live virus exhibited significant levels of spliced latent (P < 0.005) and lytic (P < 0.01) transcripts compared with implants injected with UV-inactivated virus. Many animals displayed transcript levels that would correspond to 1 infected cell in 1,000 (assuming that transcript levels per cell are similar to those in BCBL-1 cells). Animals treated with ganciclovir showed a reduction in both latent (P < 0.1) and lytic (P < 0.05) transcript levels compared with untreated implants. RNA derived from latent BCBL-1 cells served as positive control. These data suggest that ganciclovir affected both the initial replication and subsequent spread of KSHV as well as establishment of latent infection, resulting in a decrease for both messages.


Experimental transmission of Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) to SCID-hu Thy/Liv mice.

Dittmer D, Stoddart C, Renne R, Linquist-Stepps V, Moreno ME, Bare C, McCune JM, Ganem D - J. Exp. Med. (1999)

Analysis of transcript levels in infected implants. A and B show latent and lytic transcript levels expressed as the number of latent or lytic cells per 106 cells (logarithmic scale) for implants at day 14 after inoculation that were infected with KSHV (KSHV), infected with KSHV and treated with ganciclovir (KSHV ganc.), or infected with UV-inactivated virus (KSHV UV); BCBL-1 cells are shown as control. The median of each group is indicated by the dashed line. P values were calculated using the nonparametric Mann-Whitney test.
© Copyright Policy
Related In: Results  -  Collection

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Figure 5: Analysis of transcript levels in infected implants. A and B show latent and lytic transcript levels expressed as the number of latent or lytic cells per 106 cells (logarithmic scale) for implants at day 14 after inoculation that were infected with KSHV (KSHV), infected with KSHV and treated with ganciclovir (KSHV ganc.), or infected with UV-inactivated virus (KSHV UV); BCBL-1 cells are shown as control. The median of each group is indicated by the dashed line. P values were calculated using the nonparametric Mann-Whitney test.
Mentions: Fig. 5 shows relative transcript levels in infected implants at day 14 after inoculation. Total RNA was isolated from individual implants in each group (implants injected with KSHV, implants injected with UV-inactivated KSHV, and implants injected with KSHV and treated with ganciclovir) and assayed in duplicate. To normalize for the amount of RNA in each reaction, rRNA was coamplified and quantified using a differently labeled probe (primers riboF, riboR, and riboP; Table ). Ct values for KSHV-specific probes were subtracted from corresponding Ct values for rRNA and expressed as the number of latent or lytic cells per 106 cells (based on BCBL-1 control). As expected, implants injected with live virus exhibited significant levels of spliced latent (P < 0.005) and lytic (P < 0.01) transcripts compared with implants injected with UV-inactivated virus. Many animals displayed transcript levels that would correspond to 1 infected cell in 1,000 (assuming that transcript levels per cell are similar to those in BCBL-1 cells). Animals treated with ganciclovir showed a reduction in both latent (P < 0.1) and lytic (P < 0.05) transcript levels compared with untreated implants. RNA derived from latent BCBL-1 cells served as positive control. These data suggest that ganciclovir affected both the initial replication and subsequent spread of KSHV as well as establishment of latent infection, resulting in a decrease for both messages.

Bottom Line: Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) is a novel human lymphotropic herpesvirus linked to several human neoplasms.KSHV virions were inoculated directly into the implants, and viral DNA and mRNA production was assayed using real-time quantitative polymerase chain reaction.Viral gene expression was most abundant in CD19(+) B lymphocytes, suggesting that this model faithfully mimics the natural tropism of this virus.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of California, San Francisco, California 94143, USA.

ABSTRACT
Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) is a novel human lymphotropic herpesvirus linked to several human neoplasms. To date, no animal model for infection by this virus has been described. We have examined the susceptibility of C.B-17 scid/scid mice implanted with human fetal thymus and liver grafts (SCID-hu Thy/Liv mice) to KSHV infection. KSHV virions were inoculated directly into the implants, and viral DNA and mRNA production was assayed using real-time quantitative polymerase chain reaction. This revealed a biphasic infection, with an early phase of lytic replication accompanied and followed by sustained latency. Ultraviolet irradiation of the inoculum abolished all DNA- and mRNA-derived signals, and infection was inhibited by ganciclovir. Viral gene expression was most abundant in CD19(+) B lymphocytes, suggesting that this model faithfully mimics the natural tropism of this virus. Short-term coinfection with HIV-1 did not alter the course of KSHV replication, nor did KSHV alter levels of HIV-1 p24 during the acute phase of the infection. Although no disease was evident in infected animals, SCID-hu Thy/Liv mice should allow the detailed study of KSHV tropism, latency, and drug susceptibility.

Show MeSH
Related in: MedlinePlus