Infection dynamics of western equine encephalomyelitis virus (Togaviridae: Alphavirus) in four strains of Culex tarsalis (Diptera: Culicidae): an immunocytochemical study

Figures & data

Figures 1–8 Evidence of WEEV infection in Cx. tarsalis tissues. 1a) Infected cardia and anterior midgut, HVP strain, 5-log group, 21 days postinfective blood meal. 1b) Cardia and anterior midgut, negative control. 2a) Infected brain and subesophageal ganglion, HVP strain, 5-log group, 21 days postinfective blood meal. 2b) Brain and subesophageal ganglion, negative control. 3a) Infected thoracic ganglion, HVP strain, 5-log group, 21 days postinfective blood meal. 3b) Thoracic ganglion, negative control. 4a) Infected salivary glands, HVP strain, 5-log group, 21 days postinfective blood meal. 4b) Salivary glands, negative control. 5a) Infected eggs, COAV strain, 5-log group, 21 days postinfective blood meal. 5b) Eggs, negative control. 6a) Infected abdominal ganglion, HVP strain, 3-log group, 7 days postinfective blood meal. 6b) Abdominal ganglion, negative control. 7a) Infected retrocerebral complex, HVP strain, 5-log group, 7 days postinfective blood meal. 7b) Retrocerebral complex, negative control. 8a) Infected trachea, KNWR strain, 5-log group, 14 days postinfective blood meal. 8b) Trachea, negative control. In all figures, rusty-brown staining in internal tissues corresponds to positive ABC reactions, indicating the presence of WEE viral antigen. Notice that the cuticle (exoskeleton) often displays a similar brown coloring (eg, in Figures 2b, 3b, 6b, 7b, and 8b), which is not indicative of positive ABC reactions.

Abbreviations: ABC, avidin-biotin-peroxidase complex; AG, abdominal ganglion; AM, anterior midgut; BR, brain; C, cuticle; CE, cardial epithelium; FB, fat body; FT, neural fiber tract; HVP, high viremia producer strain; IE, infected egg; M, muscle (skeletal); NC, nerve cord; NE, noninfected egg; PP, pharyngeal pump lumen; RCC, retrocerebral complex; SEG, subesophageal ganglion; SG, salivary gland; T, trachea; TG, thoracic ganglion; VD, ventral diverticulum; WEEV, western equine encephalomyelitis virus.

Figure 9 Infection, dissemination, and salivary gland infection rates among experimental individuals.

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.

Table 1 Overall rates of infection, dissemination, and salivary gland infection

	HVP		WR		COAV		KNWR
	3 log (n = 35)	5 log (n = 35)	3 log (n = 35)	5 log (n = 35)	3 log (n = 35)	5 log (n = 34)	3 log (n = 34)	5 log (n = 35)
Infectiona	66%	100%	31%	83%	69%	94%	50%	86%
Disseminationb	31%	51%	6%	11%	34%	53%	6%	29%
Salivary gland infectionc	29%	37%	6%	9%	31%	44%	6%	26%

Notes:

a Infection rate = the number of females with midgut infection/total females that ingested an infectious blood meal

b Dissemination rate = the number of females with infection beyond the midgut/total number of females infected

c Salivary gland infection rate = the number of females with salivary gland infection/total number of females with disseminated infections.

Abbreviations: COAV, Coachella Valley strain; HVP, high viremia producer strain; KNWR, Kern National Wildlife Refuge strain; WEEV, western equine encephalomyelitis virus; WR, WEEV resistant strain.

Figure 10 Rates of dissemination and cardia infection among infected individuals. A significant correlation (P < 0.05) between these variables was found in the HVP strain, 5-log group. No significant correlations were found in the 3-log group. Asterisks mark time points where no infected specimens were found.

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.

Figure 11 Infection rates of neural tissue and salivary glands among infected individuals. A significant correlation (P < 0.05) between these variables was found in the 5-log group in all strains except the HVP. No significant correlations were found in the 3-log group. Asterisks mark time points where no infected specimens were found.

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.

Figure 12 Midgut infection barrier (MIB) rates. Ingestion of 5-log of virus is associated with a significant decrease (P < 0.05) in the percentage of individuals displaying a MIB in all strains. In the HVP strain, MIB was completely overwhelmed in the 5-log group.

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.

Table 2 Overall rates of individuals displaying a midgut infection barrier, midgut escape barrier, and salivary gland infection barrier

	HVP		WR		COAV		KNWR
	3 log	5 log	3 log	5 log	3 log	5 log	3 log	5 log
MIBR (N/T)	34% (12/35)	0% (0/35)	69% (24/35)	17% (6/35)	31% (11/35)	6% (2/34)	50% (17/34)	14% (5/35)
MEBR (N/I)	52% (12/23)	49% (17/35)	82% (9/11)	86% (25/29)	50% (12/24)	44% (14/32)	88% (15/17)	67% (20/30)
SIBR (N/D)	9% (1/11)	28% (5/18)	0% (0/2)	25% (1/4)	8% (1/12)	17% (3/18)	0% (0/2)	10% (1/10)

Abbreviations: COAV, Coachella Valley strain; D, number of specimens with a disseminated infection; HVP, high viremia producer strain; I, number of infected specimens; KNWR, Kern National Wildlife Refuge strain; MEBR, midgut escape barrier; MIBR, midgut infection barrier; N, number of individuals presenting the corresponding barrier; SIBR, salivary gland infection barrier; T, total number of experimental specimens; WEEV, western equine encephalomyelitis virus; WR, WEEV resistant strain.

Figure 13 Midgut escape barrier (MEB) rates. No significant difference between dose groups was found in overall MEB rates for any strain. Asterisks mark time points where MEB rates could not be estimated because no indication of gut infection was seen (MIB rate = 100%).

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.

Figure 14 Salivary gland infection barrier (SIB) rates. No significant differences were found either between dose groups, or between strains. Asterisks mark time points where rates could not be estimated due to the absence of individuals with disseminated infections.

Abbreviations: HVP, high viremia producer strain; WR, WEEV resistant strain; COAV, Coachella Valley strain; KNWR, Kern National Wildlife Refuge strain.