IL-10 and WNV RNA levels in the skin and draining lymph nodes following intradermal inoculation with 10⁴ pfu of WNV in 10 µl of PBS.

<p>RNA levels were assessed by real-time RT-PCR at 24 and 72 h post-infection and normalized to those of the GAPDH gene. The means ± standard deviation are shown, and an asterisk signifies a statistically significant difference (p<0.05) as compared to the group infected with WNV alone as determined by Mann-Whitney test. Control groups were mock infected with PBS alone. n = 5/group; displayed results are representative of two experimental replicates.</p

Leukocytes present in the dermal site of inoculation and the draining lymph nodes at 24 and 48 h after intradermal inoculation of WNV.

<p>Mice were inoculated with 10⁴ pfu of WNV in 10 µl of PBS with (n = 5) or without (n = 5) mosquito exposure, exposed to mosquitoes in the absence of WNV (n = 5), or mock infected with PBS alone (n = 3). Cells obtained from three ears per group were pooled, while lymph nodes were assessed individually. The populations of leukocytes were identified by staining and flow cytometry as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0011704#s2" target="_blank">Materials and Methods</a>. The data shown for lymph nodes are the mean ± standard deviation (representative of 3 experimental replicates). The data for the skin are displayed as the cell numbers from the pooled samples, and the differences between experimental groups are representative of three independent replicates of this experiment. The skin T cells are represented as the mean ± standard deviation of three separate experiments; analysis of the other skin populations in this manner (across replicates) was not possible due to relatively large, yet proportional, differences that existed between replicates. Nonetheless, the differences between groups maintained a similar pattern in individual experiments, regardless of the variation in levels of particular cell types. An asterisk indicates values significantly different (p<0.05) than the values in the comparable cell populations of the group exposed to WNV alone as determined by Mann-Whitney test.</p

Cytokine expression and WNV titers in resident peritoneal macrophages following <i>in vitro</i> infection in the presence mosquito salivary gland extract (SGE).

<p>IFN-β, IL-10, IL-12, and iNOS levels were assessed by real-time RT-PCR at 24 and 48 h post-infection (MOI = 5). Their mRNA expression levels were normalized to those of the GAPDH gene. WNV production was quantified by titration on vero cells. The means ± standard deviation are shown, and an asterisk signifies a statistically significant difference (p<0.05) as compared to the group infected with WNV alone as determined by Mann-Whitney test. IFN-γ and IL-1β mRNA expression were found not to vary among groups (data not shown). Control groups were mock infected with PBS alone. Results were similar in three independent replicates.</p

<i>In vitro</i> expression of cytokines in SINV-infected resident peritoneal macrophages.

<p>IFN-β, IL-10, and iNOS levels were assessed by real-time RT-PCR at 24 and 48 h post-infection (MOI  = 1). Their mRNA expression levels were normalized to those of the GAPDH gene. The means ± standard deviation are shown, and an asterisk signifies a statistically significant difference (p<0.05) as compared to the group infected with SINV alone as determined by Mann-Whitney test. IFN-γ, IL-12, and IL-1β mRNA expression, as well as SINV titers (assessed by titration on cell culture) were found not to vary among groups (data not shown). Control groups were mock infected with PBS alone. These results are representative of two independent replicates.</p

Serologic and behavioral risk survey of workers with wildlife contact in China

<div><p>We report on a study conducted in Guangdong Province, China, to characterize behaviors and perceptions associated with transmission of pathogens with pandemic potential in highly exposed human populations at the animal-human interface. A risk factor/exposure survey was administered to individuals with high levels of exposure to wildlife. Serological testing was performed to evaluate prior infection with several wildlife viral pathogens. Follow up serology was performed on a subset of the cohort as well as close contacts of individuals. 1,312 individuals were enrolled in the study. Contact with a wide range of wildlife species was reported in both occupational and occasional contexts. The overall proportion of individuals seropositive to any of the tested wildlife pathogens was approximately 4.0%. However, persons employed as butchers demonstrated a seropositivity of 9.0% to at least one pathogen of interest. By contrast, individuals working as hunters had lower rates of seropositivity. Among the study population, a number of other behaviors showed correlation with seropositivity, including contact with particular wildlife species such as field rats. These results demonstrate the need to further explore zoonotic risks of particular activities regarding wildlife contact, and to better understand risks of persons working as butchers with wildlife species.</p></div

Multivariate models for seropositivity–common activities, animal specific.

<p>Multivariate models for seropositivity–common activities, animal specific.</p

Reported exposures to wild animals.

<p>Reported exposures to wild animals.</p

Serological results, by primary occupation and reported activities.

<p>Serological results, by primary occupation and reported activities.</p

Flow diagram of recruitment and enrollment.

<p>12 prefectures were selected after primary investigation by visiting over 20 prefectures. Totally 1,267 individuals were enrolled, and serological tests and behavioral interviews were conducted. In the follow up study, 43 respondents showing seropositive results to SARS virus, hantavirus, and/or bunyavirus in the enrollment phase and 45 of their close contacts were enrolled. Additionally, 52 respondents showing indeterminate or positive results were excluded in the follow up study since they moved to other locations between the two study phases. Another round of serological analysis and behavioral investigation were conducted to 88 respondents in the follow up study.</p

Beliefs and reported behaviors.

<p>Beliefs and reported behaviors.</p