Study sites

Study sites: Site name (abbreviation), location (latitude and longitude) and altitude (metres above sea level)

Temperature data

Temperature data: Air temperature data collected every two hours, by site. Date, Time: when air temperature was recorded. Temperature in degrees centigrade by site

Freeze tolerance

Freeze tolerance: Individual ID, Mountain of origin (abbreviation), Altitude of origin (Low or High), Weight (g) and Survival (1=survived, 0=died)

Routine metabolic rate

Routine metabolic rate: Individual ID, Family, Mountain of origin (abbreviation), Altitude of origin (Low or High) and Routine_metabolic_rate (ml O2 g-1 h-1)

Spatial Spread of ZEBOV

<div><p>(A) Relationship between date and longitude of outbreaks in Gabon-Congo border area. Blue squares, human outbreaks [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b02" target="_blank">2</a>] and 2003 outbreak at Mbandza village; red circles, animal carcasses testing positive for Ebola [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b18" target="_blank">18</a>]; gray diamonds, ape die-off at Ibounji\Lokoue clearing. Regression line is for pooled data. Analyzed separately, human outbreaks and Ebola+ animal carcasses both show significant correlations between longitude and date (human outbreaks <i>n</i> = 12, R² = 0.48, <i>p</i> = 0.01; animal carcasses <i>n</i> = 13, R² = 0.91, <i>p</i> < 0.001).</p> <p>(B) Added are 1996 human outbreak at Booué and Ebola+ chimpanzee carcass from nearby Lope [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b01" target="_blank">1</a>]. The lack of reported human outbreaks between 1996 and 2001 may simply reflect the extremely low village density between Booué and Mendemba (<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-g001" target="_blank">Figure 1</a>).</p> <p>(C) Time after Yambuku versus straight line distance from Yambuku to all subsequent human outbreaks, including [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b02" target="_blank">2</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b14" target="_blank">14</a>] and Mbandza village (R² = 0.42, <i>n</i> = 17, <i>p</i> = 0.005).</p> <p>(D) Same as (C) but with distance from Yambuku to the recent Gabon-Congo border outbreaks measured as passing through Booué (R² = 0.97,<i>n</i> =17, <i>p</i> < 0.001). All figures include outbreak sites cited [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b02" target="_blank">2</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b14" target="_blank">14</a>] for which no ZEBOV-GP sequences were publicly available.</p></div

Epizootic Pivot Point

<p>Shading of each grid cell indicates the strength of correlation (R²) between geographic distance and patristic genetic distance when that grid cell (rather than Booué) is used as the epizootic pivot point. The position of the best fitting pivot point (shown with a white X) along the Ogooue River (blue line) is consistent with a river crossing near Booué, with subsequent movement east toward the Mendemba area. It is not consistent with gene flow directly between the other mid-1990s outbreak localities (Mekouka and Mayibout) and Mendemba.</p

Maps of Ebola Zaire Outbreaks

<div><p>(A) Human outbreak locations in Gabon and Congo as reported [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b02" target="_blank">2</a>]. Also shown are October 2003 human outbreak at Mbandza village and April 2004 ape die-off around Iboundji (Lokoué) Clearing in Odzala National Park. Yellow arrows represent epizootic path suggested by phylogenetic analyses.</p> <p>(B) Sites of all primary outbreaks of Ebola Zaire in humans documented [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b02" target="_blank">2</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#pbio-0030371-b14" target="_blank">14</a>] and the epizootic path suggested by the spatio-temporal pattern of outbreaks (yellow arrows). Best fitting origin found through ML search for the spatial location that produced the strongest correlation between outbreak date and geographic distance from the origin. ML search based on the correlation between patristic genetic distance and spatial separation between outbreaks places the epizootic pivot point just southeast of Booué. In both figures, shading of circles is proportional to time after first outbreak in series.</p></div

Spatial Spread from 2001–2005

<p>Distance of each human outbreak site from the initial outbreak at Mendemba village, Gabon, plotted as a function of time after the Mendemba outbreak. Dashed regression line uses only outbreaks from 2001–2003 (R² = 0.43, <i>p</i> = 0.04). Solid regression line includes May 2005 outbreak at Etoumbi village (R² = 0.73, <i>p</i> < 0.001).</p

Correlation between Geographic Distance and Patristic Genetic Distance

<div><p>(A) ML genetic distances (substitutions per nucleotide site) plotted as function of geographic distance separating pairs of outbreak sites for the six full-length, georeferenced sequences sampled by Leroy et al. (R² = 0.70, Mantel test <i>p</i> = 0.002). Makokou and Yembelengoye sequences excluded because of unknown spatial origin of case and partial sequence, respectively (<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0030371#sd003" target="_blank">Protocol S3</a>).</p> <p>(B) Correlation between straight line distance from the initial ZEBOV outbreak site at Yambuku and patristic genetic distance to Yambuku for all available georeferenced, full-length sequences (R² = 0.38 , <i>n</i> = 11, <i>p</i> = 0.040).</p> <p>(C) Same as (B) but with geographic distances to the recent Gabon-Congo border outbreaks measured as passing through Booué (R² = 0.92 , <i>n</i> = 11, <i>p</i> < 0.001).</p></div

Environmental parameters

pH, conductivity (μS), total dissolved solids (ppm) and dissolved oxygen content (mg/l) measurements by site