Johnson left the track immediately after the race and later pulled out of Sunday's grand prix meeting in Sheffield. The 29-year-old is to head back to America for a couple of weeks and is unlikely to race again until Stuttgart on July 13.
The Olympic 200m and 400m champion, who said he had recovered from the injury which caused him to miss the US trials, finished in 45.76sec in a race won by one of the Americans who did qualify for this year's World Championships, the 1991 champion, Antonio Pettigrew.
"He is going home for two weeks," said his manager, Brad Hunt. "It is not an injury but a fitness thing. All we can do is work on it."
Pettigrew recorded 44.86sec, with the Olympic bronze medallist, Davis Kamoga, second in 45.19sec after four runners passed a flagging Johnson near the line.
Johnson, whose winning streak began on April Fools' Day, 1989, injured his quadricep muscle in the self-styled One To One Challenge against Donovan Bailey over 150m on 1 June. The American gained the historic double in the 1996 Atlanta Olympics, setting a world record of 19.32 in the 200m, but the last time he finished a race was on 25 May in Oregon. The only 400m he ran before yesterday was on 19 April when he won in 43.75, the fastest early-season time this year.
Bailey prepared for Sunday's pounds 50,000 challenge against Linford Christie with an easy 100m victory. Despite racing on a track left damp by the day-long downpour, the world and Olympic champion won in a time of 10.07sec to see off a field containing all of his leading American rivals.
The Canadian won by 500ths of a second from Tim Montgomerie, second in the recent US trials. Another American - Brian Lewis - was third in 10.19, while Maurice Greene, the US trials winner, could only finish fourth in 10.23.
Bailey is clearly in shape to give Christie a severe test on Sunday, though he has failed to beat Britain's top sprinter in their two previous meetings over the rarely run distance in Sheffield in recent years.
Hicham El Guerrouj, whose fall in last summer's Olympic 1500m final prevented him making his expected challenge against the world champion, Noureddine Morceli, earned a measure of revenge last night as he beat his Algerian rival with a winning time of 3:31.87.
Sonia O'Sullivan, Ireland's reigning world 5,000m champion, had to settle for fourth place in the 3,000m. The 27-year-old Teddington-based runner, in her first serious outing over the distance this year, was never on the same pace as the leaders as she finished in 8min 53.53sec.
O'Sullivan was nearly 13 seconds down on the winner Gabriela Szabo, the Romanian who was beaten into second place by Britain's Kelly Holmes in the European Cup 1500m in Munich last weekend. "I have no explanation to what has happened," said O'Sullivan after losing out to one of her likely chief rivals at this summer's World Championships. "If I did I wouldn't have finished where I did. I am very disappointed."
In the 200m, Frankie Fredericks, the last man to beat Johnson at the distance, won last night's event in 20.38, with Trinidad's Ato Boldon - the fastest man this year over 100m - way back in sixth in 20.75.
The local favourite Marie-Jose Perec, the women's 200 and 400 metres Olympic champion, disappointed her fans by coming in a poor seventh in the 200m. It was Perec's first race of the season and she explained afterwards that she was only just getting over a stress fracture of the shin. Inger Miller, of the United States, won the race with a time of 22.48, equalling the year's best. Perec clocked 23.17.
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