Italy had to come from behind to seal their place with two games to spare after former Lazio frontman Libor Kozak fired Czech Republic ahead after 19 minutes in Turin.
Giorgio Chellini headed the Azzurri level on 52 minutes, when Petr Cech failed to deal with a corner, before Mario Balotelli stroked home the winning penalty moments later.
Theodore Gebre Selassie recklessly gave away the penalty and a bad night for the Czech defence was rounded off when Daniel Kolar saw red with a minute left.
Robin van Persie sent Holland through to Brazil with a second-half double in a 2-0 win over Andorra.
After Turkey beat Romania 2-0 in Bucharest earlier in the evening, the Dutch knew victory would be enough to confirm their place in the World Cup finals.
Louis van Gaal's side made heavy work of it but Van Persie's goals on 50 and 55 minutes were enough.
Algeria rounded off the Group H campaign with a 1-0 win over Mali in Blida.
El Arabi Soudani's 51st-minute goal ensured Vahid Halilhodzic's team finished seven points clear at the top of the group - and they too will be hoping to win their play-off and make sure of a presence at Rio de Janeiro.
England edged to the top of Group H with a goalless draw against Ukraine, who joined the inactive Montenegro a point behind Roy Hodgson's men.
Poland are two points further back after they eased past San Marino 5-1 - Piotr Zielinski netting twice for the visitors.
Bosnia and Greece remain locked on 19 points at the top of Group G after they both collected three points.
Bosnia came from behind to beat Slovakia 2-1 - Ermin Bicakcic and Izet Hajrovic scoring in the final 20 minutes - while Dimitris Salpigidis (58) grabbed the only goal as Greece beat 10-man Latvia.
Lithuania beat Liechtenstein 2-0 to round out the group.
The battle for second place behind Holland in Group D tightened further with Hungary's 5-1 win over Estonia giving them the advantage.
The eastern Europeans are a point ahead of both Romania and Turkey, whose qualification hopes have improved vastly following back-to-back wins under the re-installed Fatih Terim.
Bulgaria held on to second place in Group B behind Italy with a 2-1 win in Malta but Denmark are just a point further back after skipper Daniel Agger converted a second-half penalty against Armenia.
The 1-0 win was revenge for the Danes after Armenia beat them 4-0 in Copenhagen earlier in qualifying.
Group A's top two Belgium and Croatia were not in competitive action, and in their absence it was left to Scotland to climb off the foot of the section with a 2-1 win in Macedonia.
Wales moved bottom - despite the presence of 100million euro winger Gareth Bale off the bench - as they were thumped 3-0 by Serbia in Cardiff.