Race leader Rodriguez of Katusha posted the seventh fastest time on the hilly 39.5km time trial in West Galicia, which was unexpectedly won by the Swede Frederik Kessiakoff of Astana.
Kessiakoff completed the demanding course in a time of 52 minutes and 36 seconds, 17 seconds quicker than second-place Contador of Saxo Bank-Tinkoff.
Contador rises to second on GC, one second behind Rodriguez, while Britain's Chris Froome drops to third place, 16 seconds in arrears, after the Team Sky rider could only post the third-best time in the race against the clock, 39 seconds slower than Kessiakoff.
"I'm very relieved," said Kessiakoff, who started his ITT in the town of Cambados well over an hour before the race favourites. "I knew I had a good time when I finished and I knew it would be close to the times of the favourites like Contador and Froome."
Contador was two seconds ahead of the Swede at the first time check after 13.5km but the pendulum swung on the middle section of the course, which featured a testing 10km Cat.3 climb, leaving Kessiakoff two seconds up at the 23.5km mark.
Aware that a strong ride could put him in the red jersey, Contador kept things together on the technical descent, but could not match Kessiakoff's fast pace over the final flat section to the town of Pontevedra.
"I'm really happy and relieved that I pulled it off," Kessiakoff admitted after a nervous hour watching events pan out from the finish zone. "I was very focused this morning and last night. I came here ready to suffer and I'm very happy to have won."
If Kessiakoff's stage win was unexpected then so too was Rodriguez's success in holding on to the race lead. Not a renowned time triallist, the Spaniard was expected to lose his red jersey to one of the favourites for the ITT: either Froome, 53 seconds down overnight, or Contador, a further seven seconds down before stage 11.
Froome was four seconds slower than Contador at the first check and then 16 seconds behind after the Alto Monte Castrove, which boasted an average gradient of 4.4 per cent. Clearly suffering from the accumulative efforts of an active summer, the Tour de France runner-up and Olympic time trial bronze medallist faded further in the second half of the race but is still within striking distance of the top of overall standings ahead of the main mountain stages.
Omega Pharma-QuickStep's Tony Martin, the world time trial champion from Germany, could only manage the 11th best time, 1:39 behind Kessiakoff, while Australians Richie Porte (Team Sky) and Cameron Meyer (Orica-GreenEdge) held the top two spots for a long period before finally being ousted by the Swede. Porte finished sixth, one place behind Spain's Benat Intxausti (Movistar) while eighth-place Meyer was one second slower than Rodriguez and a few seconds faster than the American Andrew Talansky (Garmin-Sharp)
Another big surprise on Wednesday came from double stage winner Alejandro Valverde who, like Rodriguez, was expected to concede a large slice of time to both Contador and Froome.
Wearing the blue polka dot jersey as best climber, the Movistar rider limited his losses to post the fourth-best time, just 1:08 down on the stage winner to hold on to his fourth place on GC, 59 seconds in arrears.
With the Vuelta's four best climbers all within less than a minute of each other at the top of the standings, the scene is perfectly set for an explosive second half of the race. Thursday's 190.5km flat stage from Vilagarcia de Arousa concludes with a punchy Cat.3 ramp at Dumbria - a finish ideally suited to the explosive attacking strengths of Rodriguez.
A largely flat stage on Friday is then followed by three consecutive mountain-top finishes ahead of the second rest day next Tuesday.