In the space of a week, the Royals have lost to both Fareham Town and Michael Birmingham's side.
It is also a third successive loss for Portchester who were knocked out of the FA Cup at two divisions higher Wimborne Town at the weekend.
Royals manager Mick Catlin was disappointed with the result.
'I thought after we went 2-1 we should have managed things a lot better,' he said.
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'Fair play to Horndean, they were a young energetic side and we ran out of steam towards the end.
'The youngsters we put in did well but I expected a bit more from some of my more experienced players.’
The home side gave goalkeeper Cameron Scott his first game since returning to the club from Baffins Rovers last month.
They also had to make a late change when Ellis Martin picked up an injury in the warm up to be replaced by Luke Dempsey.
The first chance of the game fell to the Royals after seven minutes.
Steve Ramsey lofted a free-kick into the area and Dempsey sent a free header over the bar from six yards.
A mistake by Pete Castle gave Horndean their first opportunity in the 17th minute but the shot was dragged wide.
The visitors had a let off two minutes later after a brilliant run by Patrik Hnath was ended with a foul on the edge of the area.
When the ball was delivered into the box, Craig Harding's header bounced out off the post.
At the other end Scott kept the scores level with an excellent save, pushing a powerful shot onto the upright.
It was the visitors who struck the first blow on the half hour.
A long ball over the top was controlled well by Connor Duffin who went on to place his shot beyond Scott.
Looking for a quick response, Ramsey went close twice in the space of as many minutes.
His powerful curling shot was saved by Lloyd Thomas and then he sent a header narrowly over the top.
The hosts turned the tie on its head with two goals in five minutes before the break.
First, Casey Grieves did brilliantly on the right and, from his cross, Joe Johnson calmly volleyed past the visiting 'keeper.
On the stroke of half-time Castle flicked on a corner and Harding completed the finish.
Horndean hauled themselves level on 67 minutes when a ball into the box left 17-year-old Ben Anderson with an easy finish.
Zak Brownlie put Horndean in front nine minutes from time after a good through ball split the home defence.
Horndean boss Birmingham was proud of all his players.
'My youngsters were immense out there,' he remarked.
'I thought they were phenomenal.
'What was most pleasing was the way the more experienced players trusted them and gave them the ball.
'The kids refused to be overawed and played with no fear.
'It was a joy to watch and they were all a credit.'
AFC Portchester: Scott; Searle O, Cripps, Castle, Dempsey, Clark (Evans), Grieves, Ramsey, Harding (Wort 46), Hnath, Johnson (Aston 65).
Horndean: Thomas; Bush, Clark, Grant, Morgan, Brownlie, Hookey, Anderson, Adams, Duffin, Hall. Substitutes used: Tierney, Taw, Harris.