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. 2022 Aug 3;11(8):879.
doi: 10.3390/pathogens11080879.

AG129 Mice as a Comprehensive Model for the Experimental Assessment of Mosquito Vector Competence for Arboviruses

Lívia V R Baldon  1 , Silvana F de Mendonça  1 , Flávia V Ferreira  1 , Fernanda O Rezende  1 , Siad C G Amadou  2 , Thiago H J F Leite  2 , Marcele N Rocha  1 , João T Marques  2   3 , Luciano A Moreira  1 , Alvaro G A Ferreira  1
Affiliations

AG129 Mice as a Comprehensive Model for the Experimental Assessment of Mosquito Vector Competence for Arboviruses

Lívia V R Baldon et al. Pathogens. .

Abstract

Arboviruses (an acronym for "arthropod-borne virus"), such as dengue, yellow fever, Zika, and Chikungunya, are important human pathogens transmitted by mosquitoes. These viruses impose a growing burden on public health. Despite laboratory mice having been used for decades for understanding the basic biological phenomena of these viruses, it was only recently that researchers started to develop immunocompromised animals to study the pathogenesis of arboviruses and their transmission in a way that parallels natural cycles. Here, we show that the AG129 mouse (IFN α/β/γ R-/-) is a suitable and comprehensive vertebrate model for studying the mosquito vector competence for the major arboviruses of medical importance, namely the dengue virus (DENV), yellow fever virus (YFV), Zika virus (ZIKV), Mayaro virus (MAYV), and Chikungunya virus (CHIKV). We found that, after intraperitoneal injection, AG129 mice developed a transient viremia lasting several days, peaking on day two or three post infection, for all five arboviruses tested in this study. Furthermore, we found that the observed viremia was ample enough to infect Aedes aegypti during a blood meal from the AG129 infected mice. Finally, we demonstrated that infected mosquitoes could transmit each of the tested arboviruses back to naïve AG129 mice, completing a full transmission cycle of these vector-borne viruses. Together, our data show that A129 mice are a simple and comprehensive vertebrate model for studies of vector competence, as well as investigations into other aspects of mosquito biology that can affect virus-host interactions.

Keywords: AG129; arbovirus; mice model; vector competence; vertebrate transmission model; virus transmission.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Medically important mosquito-borne arboviruses tested in the AG129 mice model. The global distribution includes the present the contemporary range, and both present and past outbreaks. The transmission cycles include the major vectors and the major reservoirs.
Figure 2
Figure 2
Mortality of immunocompromised AG129 (IFNα/β/γR−/−) mice after inoculation with arbovirus. (A) Scheme of the experimental design. Three or eight weeks-old AG129 mice were inoculated with arbovirus intraperitoneal (IP) injection performed in the lower right quadrant. (B–F) Survival probability (Kaplan–Meier plot) of AG129 mice inoculated with mock or viruses. Mice were monitored daily for seven days. For each virus seven mice were used. (B) Three-week-old mice inoculated with 106 p.f.u. of DENV-1. (C) Three-week-old mice inoculated with 105 p.f.u. of ZIKV. (D) Three-week-old mice inoculated with 106 p.f.u. of YFV. (E) Eight-week-old mice inoculated with 105 p.f.u. of CHIKV. (F) Eight-week-old mice inoculated with 105 p.f.u. of MAYV.
Figure 3
Figure 3
Arbovirus viremia in AG129 mice. (A) Scheme of the experimental design. Three or eight week-old AG129 mice were inoculated with arbovirus intraperitoneal (IP) injection performed in the lower right quadrant. Blood was collected every 24 h during 7 days for viral RNA quantification. Samples were tested individually by RT-qPCR. Each dot represents a blood sample from an individual mouse. For each time point, three different mice were sampled. Mice were monitored daily for seven days. Mice were euthanized after a single blood collection. (B–F) Virus RNA levels of blood samples from AG129 mice inoculated with mock or viruses. (B) Three-week-old mice inoculated with 106 p.f.u. of DENV-1. (C) Three-week-old mice inoculated with 105 p.f.u. of ZIKV. (D) Three-week-old mice inoculated with 106 p.f.u. of YFV. (E) Eight-week-old mice inoculated with 105 p.f.u. of CHIKV. (F) Eight-week-old mice inoculated with 105 p.f.u. of MAYV.
Figure 4
Figure 4
Viremic AG129 mice can transmit arbovirus to vector mosquitoes. (A) Scheme of the experimental design. Three or eight week-old AG129 mice were inoculated with arbovirus intraperitoneal (IP) injection performed in the lower right quadrant. After two or three days, mice were anaesthetized and then Ae. aegypti mosquitoes (five to seven-day-old females) were allowed to feed on virus-infected mice. Eight days post blood meal, mosquitoes were collected and tested individually for the presence of virus. (B) Three-week-old mice were inoculated with 106 p.f.u. of DENV-1 and three days later, were exposed to mosquito blood-feeding. (C) Three-week-old mice were inoculated with 105 p.f.u. of ZIKV and three days later, were exposed to mosquito blood-feeding. (D) Three-week-old mice were inoculated with 106 p.f.u. of YFV and three days later, were exposed to mosquito blood-feeding. (E) Eight-week-old mice were inoculated with 105 p.f.u. of CHIKV and two days later, were exposed to mosquito blood-feeding. (F) Eight-week-old mice were inoculated with 105 p.f.u. of MAYV and two days later, were exposed to mosquito blood-feeding. In the infection prevalence bar graphs, “n” refers to the number of infected mosquitoes out of the total of mosquitoes tested.
Figure 5
Figure 5
The AG129 mouse is a valid model to study mosquitoes-to-vertebrate transmission of arbovirus. (A) Scheme of the experimental design to test arbovirus transmission from mosquitoes to AG129 mice. Infected mosquitoes (that had taken an infectious blood meal from viremic AG129 mice 14 days earlier) were allowed to take blood meals in naïve AG129 mice for a second blood feeding. Three days later, the presence of arbovirus in the AG129 mice from which the mosquitos took the second blood meal, was detected by RT-qPCR. Each AG129 mouse was exposed to 5 infected mosquitoes for 30 min. Between two and five mosquitoes were able to complete a blood meal. Fully engorged mosquitoes were counted and collected to confirm the presence of viruses using RT-qPCR. Samples were tested individually by RT-qPCR. Each dot represents a blood sample from an individual mouse. (B) DENV-1 RNA levels in the mice after exposure to infected mosquitoes. (C) ZIKV RNA levels in the mice after exposure to infected mosquitoes. (D) YFV RNA levels in the mice after exposure to infected mosquitoes. (E) CHIKV RNA levels in the mice after exposure to infected mosquitoes. (F) MAYV RNA levels in the mice after exposure to infected mosquitoes.
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