Evaluating the competence of the primary vector, Culex tritaeniorhynchus, and the invasive mosquito species, Aedes japonicus japonicus, in transmitting three Japanese encephalitis virus genotypes

Abstract

Japanese encephalitis virus (JEV) is maintained in an enzootic cycle between swine, water birds, and mosquitoes. JEV has circulated indigenously in Asia, with Culex tritaeniorhynchus as the primary vector. In some areas where the primary vector is scarce or absent, sporadic cases of Japanese encephalitis have been reported, with Aedes japonicus japonicus presumed to have the potential as a secondary vector. As one of the world’s most invasive culicid species, Ae. j. japonicus carries a considerable health risk for spreading diseases to wider areas, including Europe and North America. Thus, evaluation of its competency as a JEV vector, particularly in a native population, will be essential in preventing potential disease spread. In this study, the two mosquito species’ vector competence in transmitting three JEV genotypes (I, III, and V) was assessed, with Cx. tritaeniorhynchus serving as a point of reference. The mosquitoes were virus-fed and the infection rate (IR), dissemination rate (DR), and transmission rate (TR) evaluated individually by either RT-qPCR or focus forming assay. Results showed striking differences between the two species, with IR of 95% (261/274) and 9% (16/177) in Cx. tritaeniorhynchus and Ae. j. japonicus, respectively. Both mosquitoes were susceptible to all three JEV genotypes with significant differences in IR and mean viral titer. Results confirm the primary vector’s competence, but the fact that JEV was able to establish in Ae. j. japonicus is of public health significance, and with 2%–16% transmission rate it has the potential to successfully transmit JEV to the next host. This may explain the human cases and infrequent detection in primary vector-free areas. Importantly, Ae. j. japonicus could be a relevant vector spreading the disease into new areas, indicating the need for security measures in areas where the mosquito is distributed or where it may be introduced.

Mosquito Vector Competence for Japanese Encephalitis Virus

“…This technique was used to demonstrate the vector competence of Cx . tritaeniorhynchus from Japan [ 23 , 153 , 157 ], Singapore [ 149 ] and Taiwan [ 92 , 155 ]. Another route for infection is the direct injection of JEV into the thorax of a mosquito.…”

“…quinquefasciatus [ 114 ], JEV genotype I had a shorter EIP in Cx . tritaeniorhynchus compared to genotypes III and V, whereas the transmission rates where similar for all three tested genotypes [ 157 ].…”

“…A recent study with a laboratory colony of Ae . japonicus from Japan demonstrated the successful infection, dissemination and transmission of three different JEV genotypes [ 157 ].…”

“…Finally, survival rate upon JEV infection should also be carefully monitored, as a certain amount of mosquitoes dies from JEV infection [ 157 ]. This effect of JEV on the lifespan of the mosquitoes is important to consider when studying the vector competence and estimating the risk of transmission.…”

Identification and Isolation of Japanese Encephalitis Virus Genotype IV from Culex vishnui Collected in Bali, Indonesia in 2019

“…The library preparation, next-generation sequencing (NGS) procedure, and de novo assembly were conducted using the Neb next ultra RNA first-strand synthesis module (New England BioLabs, Ipswich, MA), MiniSeq (Illumina, San Diego, CA) and CLC Genomics Workbench version 11 (QIAGEN, Venlo, Limburg, Netherlands), respectively, as previously described. 9 To confirm the JEV pool source, viral RNA was extracted from the final supernatant of the virus isolation process using the rapid viral RNA screening method described in Faizah et al 10 Detergent mixture was used for extraction and reverse transcription-polymerase chain reaction (RT-PCR) using a PrimeScript One-Step RT-PCR Kit version 2 (Takara Bio, Shiga, Japan) was used for sequence amplification. A primer set targeting the JEV envelope gene (JEV955f: 5 9 -TGYTGGTCGCTCCGGCTTA-3 9 ; JEV1606R: 3'-GACY-TYGAMCCACGGTCAT-5 9 ) was used.…”