It is the first time since the 2012 Stuttgart Open that all four top seeds reached the final four in a WTA 500 event.
Pegula, the top overall seed and the only remaining American, used her commanding serving game to overwhelm 12th seed Paula Badosa 6-3 7-6 (8-6).
She ended up winning 80 per cent of her accurate first serves – compared to 57 per cent for Badosa – and it resulted in the Spaniard producing just one break point opportunity in the match, which she could not take.
Pegula will meet fourth seed and Olympic gold medallist Belinda Bencic next after her relatively comfortable 6-3 6-3 triumph against Russia's Ekaterina Alexandrova.
Switzerland's Bencic needed just 80 minutes to get the job done, creating 12 break point chances compared to Alexandrova's two, while winning the first three games of each set.
After coming into the tournament without consecutive wins since early January, reigning Wimbledon and US Open finalist Ons Jabeur looked back to her best in a dominant 6-0 4-1 (retired) drubbing of Anna Kalinskaya.
Jabeur is yet to lose a set in Charleston, and she will try to keep that the case when she faces Daria Kasatkina in her semi-final.
Kasatkina earned her spot in the final four with Friday's only three-setter, emerging victorious 6-7 (5-7) 6-4 6-2 against Madison Keys in just over two and a half hours.
Pegula, who is the second-highest seed remaining in the field after Iga Swiatek's withdrawal, had no issue dispatching Canada's Katherine Sebov 6-3 6-1 in just 66 minutes.
Gauff, the six seed, followed suit as she raced to a 6-4 6-3 win over Canadian Rebecca Marino in just 73 minutes.
Florida's own Danielle Collins thrilled her hometown crowd with a 7-6 (7-3) 6-2 triumph against Bulgaria's Viktoriya Tomova, and rising 22-year-old talent Clare Liu made it a perfect 4-0 sweep for the USA representatives by defeating Julia Grabher 6-4 6-3.
Last week's Indian Wells Open champion Elena Rybakina was made to work in her 7-5 4-6 6-3 win over Anna Kalinskaya, while an even more gruelling effort was required in Paula Badosa's two-hour-and-52-minute 7-6 (7-2) 4-6 6-2 result against Laura Siegemund.
The top seed to fall on Thursday was eighth seed Daria Kasatkina, as she won the first set before going down 4-6 6-2 6-2 against Elise Mertens.
Liudmila Samsonova, the 12th seed, made light work of Viktorija Golubic in a 6-1 6-1 drubbing, and 22nd seed Jelena Ostapenko prevailed 6-3 6-4 against Mirjam Bjorklund.
Ostapenko will next play 13th seed Beatriz Haddad Maia after she won a strange back-and-forth contest 7-6 (7-4) 0-6 6-0 over Tereza Martincova.
Pegula, the top-ranked American and the tournament's overall top seed, may have thought it was going to be smooth sailing after collecting a break-to-love in the match's opening game, but it was just the beginning of a two-hour-and-32-minute battle.
Begu played terrific tennis against a world-class opponent, winning 47 per cent of the match's total points, but she shot herself in the foot with eight double faults while Pegula had just one.
With the victory, Pegula booked her place in the quarter-final against Spain's Paula Badosa after her stylish 6-1 6-3 result over Russian 19-year-old Diana Shnaider.
Badosa now has a 4-2 record since the beginning of March, but both of those losses came to the in-form Elena Rybakina in eliminations from both the Indian Wells Open and the Miami Open.
Meanwhile, reigning Wimbledon and US Open finalist Ons Jabeur needed just 82 minutes to dispatch Caroline Dolehide 6-3 7-5, and combined with her opening victory against Lesia Tsurenko it is Jabeur's first consecutive wins since the Adelaide International in early January.
Jabeur will play Anna Kalinskaya in her quarter-final after the Russian upset two-time Australian Open champion Victoria Azarenka 6-4 7-6 (7-5).
It was a strong day for the Russians, as Ekaterina Alexandrova got the better of Julia Grabher 6-4 6-2, and world number eight Daria Kasatkina knocked out Bernarda Pera 6-3 7-6 (7-3).
Olympic gold medalist Belinda Bencic won the longest match of the day in a two-hour-and-45-minute war of attrition against Shelby Rogers 4-6 7-5 6-2, but the Americans had something to cheer for in the late window as Madison Keys advanced past Magda Linette 6-2 3-6 6-1.
Badosa, the former world number two, is back in the WTA's top 50 following Sunday's 6-1 4-6 6-4 defeat of Marie Bouzkova.
It marks the Spaniard's fourth Tour-level title but her first since she triumphed in Sydney at the start of the 2022 season.
Last year was a difficult one for Badosa, who sustained a stress fracture in her back and subsequently dropped out of the top 100.
But now she is back to feeling like her old self.
"What I went through the last year, for me being back, winning big titles like a [WTA] 500, [beating] the best players in the world, being competing against them again, for me it means a lot," said Badosa, who downed Emma Raducanu en route to clinching the trophy.
"That's why I had this mix of emotions. I was really, really nervous because I really wanted it really badly.
"One year ago I was on the couch, so it's a big difference now. Now I'm an athlete again.
"Even before the tournament I was feeling good, I was feeling confident.
"I think I needed to win something, a big title for me. Because of my personality, I'm not happy always with finals and semi-finals. I want to win titles.
"I'm [a] perfectionist. Of course, that sometimes brings me down, but also in important moments I think it gives me [something] extra. I felt I needed this moment very much."
Kazakhstan's Rybakina, who is also the reigning Wimbledon Champion and Australian Open finalist, got the better of Badosa for the second time during her current run, also eliminating her from Indian Wells.
The 23-year-old has to be considered one of the favourites to go all the way, having knocked off world number one Iga Swiatek and world number two Aryna Sabalenka to lift the trophy in California.
Rybakina will meet Belgium's Elise Mertens in the fourth round after she beat Croatia's Petra Martic 6-4 6-3.
Meanwhile, the biggest upset of the day was delivered by Russia's Anastasia Potapova, bouncing sixth seed Coco Gauff 6-7 (8-10) 7-5 6-2.
Potapova, who has never won a tournament above the WTA 250 level, will face China's Zheng Qinwen for a spot in the quarter-finals after her three-hour 5-7 7-6 (7-5) 6-3 triumph over Liudmila Samsonova.
Latvia's Jelena Ostapenko, the 24th seed, eliminated Brazilian 13th seed Beatriz Haddad Maia 6-2 4-6 6-3, while 20th seed Magda Linette of Poland knocked out Belarusian 14th seed Victoria Azarenka 7-6 (7-3) 2-6 6-4.
Unseeded American Clare Liu went down 4-6 7-5 6-4 against Italy's Martina Trevisan, but third seed Jessica Pegula will continue to fly the flag for the United States after advancing 6-1 7-6 (7-0) in her all-American showdown with Danielle Collins.
It was an ominous start for Cirstea, being broken to love in the opening game, but she stuck with second seed Sabalenka and broke back, before ultimately being pipped to the first set by the impressive Belarusian.
The key was on break points, with Sabalenka claiming all five that she won against the Cirstea serve, while saving six of eight on her own as she ultimately eased to victory.
Sabalenka now has 26 wins on clay in the WTA since 2021, with only Ons Jabeur (37), Iga Swiatek (34), Paula Badosa (31) and Coco Gauff (28) having more during this time.
Gauff also advanced after a routine 6-4 6-1 win over Irene Burillo Escorihuela, making the sixth seed in Madrid the first player to win 35 WTA-1000 main draw matches as a teenager since 2009.
Ninth seed Maria Sakkari defeated Arantxa Rus 6-4 6-4 and fifth seed Caroline Garcia also had few problems against Yulia Putintseva, winning 6-3 6-4.
However, it was not a good day for 10th seed Petra Kvitova, who was beaten 7-6 (11-9) 6-1 by Jule Niemeier, while 13th seed Beatriz Haddad Maia also lost, 7-6 (8-6) 6-3 to Mirra Andreeva and 20th seed Donna Vekic was eliminated in straight-sets by Rebeka Masarova.
Jelena Ostapenko took just 61 minutes to get past Linda Fruhvirtova 6-0 6-3, and will face 14th seed Liudmila Samsonova next, who did not take much longer to see off Maryna Zanevska 6-2 6-3.
There were also wins for Elise Mertens, Badosa, Camila Osorio, Shelby Rogers, Mayar Sherif, Magda Linette and Irina-Camelia Begu.
Losing her opening matches at the sunshine double of Miami and Indian Wells, after only taking five games off Iga Swiatek at the quarter-final in Doha, the world number five improved her head-to-head record with Riske to 5-0.
Sabalenka by no means cruised to victory, though, failing to close the match out at 5-2 in the second set. Riske had four break points to level at 5-5, but the 23-year-old powered through when she needed to, eventually closing out after an hour and 49 minutes.
Second-seeded Paula Badosa of Spain and ninth-seeded American Madison Keys also advanced to the third round at Charleston, with respective wins over Anna Bondar and Ulrikke Eikeri.
Badosa, who made the semi-finals in Charleston last year, overcame a mid-match rain delay to eventually win in an hour and 35 minutes.
Jessica Pegula continued her positive run of form after making the semi-finals in Miami, accounting for Jasmine Paolini with a 6-2 6-1 victory.
Other seeded winners included Belinda Bencic and Alize Cornet, while Elina Rybakina, Ajla Tomljanovic and Shuai Zhang were among seeds who lost on Wednesday.
The weather in South Carolina continued to be a problem at the first WTA clay-court event this season, however, as the match between Ons Jabeur and Emma Navarro was suspended due to heavy rain.
Meanwhile at the Copa Colsanitas in Bogota, Rebecca Peterson was the only seeded player in action on Wednesday, and lost 6-1 6-4 to Germany's Tatjana Maria.
The pair have endured contrasting fortunes in their opening two matches at Roland-Garros this week.
Sabalenka has dropped just seven games on the way to straight-set victories over Erika Andreeva and Moyuka Uchijima, while Badosa has come from behind to beat Katie Boulter and Yulia Putintseva.
The reigning Australian Open champion - fresh from reaching successive clay-court finals in Madrid and Rome - cannot wait to face the Spaniard, but acknowledged their friendship will be put aside when they collide.
"She's my favourite," Sabalenka said. "I love her so much. I love to see her play, and she's really a great fighter, so it's always great matches.
"It's always tough to play your friend, really your best friend on tour, but we know how to manage that. We know how to separate court and life. So, it's always a great battle, a great fight against her. I always enjoy playing against her."
Sabalenka has won each of the last four meetings between the players, though the most recent in Stuttgart earlier this year ended prematurely with an emotional Badosa forced to retire injured.
The former world number two has endured a seemingly endless battle with injuries in recent times, with a back problem forcing her to miss the second half of last season and seeing her drop out of the WTA's top 100.
"She has a big personality," Badosa said of her opponent. "She always brings this good energy, even on court. She's a very active, very intense player.
"I'm really looking forward to that match. I think the last time [in Stuttgart], it finished in a sad way. I learned a lot from that match. I think it was a very good one from both sides.
"Sharing the court with her - after all these results she's getting - is a pleasure for me, because this past year hasn't been easy. Playing this kind of match makes it all worth it."
The second seed trailed 4-2 and was a break down in the opening set, but went on to win 11 of the next 13 games to seal a fourth-round clash with either Madison Keys or Emma Navarro.
Sabalenka has dropped just seven games in her opening two matches at Roland-Garros.
Although, it looked like she may lose the opening set when a fortuitous net shot gave Badosa a second break and a 4-2 lead on Court Philippe-Chatrier.
However, she dug deep from 5-3 adrift, winning the last four games to draw first blood.
The reigning Australian Open champion broke her opponent a further two times on the way to a commanding 4-0 advantage in the second.
Badosa did well to save three match points from 0-40 in game six, but the Spaniard could not rescue a fourth as her opponent continued her surge into the second week.
Data Debrief: Sabalenka completes top four in last 16
Following on from her victories when they met in Miami and Stuttgart earlier this season, Sabalenka became the first player to beat Badosa three times at WTA events in a calendar year.
Joining Iga Swiatek, Coco Gauff and Elena Rybakina, this marks the first time since 2013 that the top four seeds are through to the last 16 in the women's singles at Roland-Garros.
Two-time Wimbledon champion Kvitova, the 12th favourite in the United Arab Emirates, eased past Italy's Martina Trevisan 6-2 6-1 to make a dominant start.
World number 15 Samsonova was made to work to beat Paula Badosa, winning 6-7 (3-7) 7-6 (7-5) 6-4 to tee up a clash with Qinwen Zheng, who overcame compatriot Zhang Shuai in straight sets.
Samsonova's triumph took three hours and 22 minutes, the longest match of the WTA Tour season thus far, in a thrilling first-round encounter between two top-20 players.
"Playing against Paula is always tough," 14th seed Samsonova said in her on-court interview. "She's doing unbelievable, so I'm really proud that I stayed on the court until the end.
"I think I'm growing match after match, day by day. I know it's a long journey, and I hope to continue like that."
Barbora Krejcikova, the 2021 French Open champion, cruised into the last 32 with a comfortable 6-4 6-2 victory over Irina-Camelia Begu as she aims to go one better than her runners-up finish two years ago in Dubai.
Another routine victory saw American Madison Keys ease past Jasmine Paolini 6-1 6-1, but there was no such luck for 2020 Australian Open champion Sofia Kenin.
She was dispatched 6-1 6-1 by world number 26 Marie Bouzkova, while Amanda Anisimova claimed a 6-3 6-2 win over veteran two-time grand slam finalist Vera Zvonareva.
Viktoriya Tomova had too much for Kaia Kanepi in a 6-3 6-1 success, with her reward a second-round clash against third seed Jessica Pegula.
World number four Pegula lost to the in-form Iga Swiatek in the Qatar Ladies Open final on Saturday, and Leylah Fernandez will face the Pole next after beating Julia Grabher 6-4 6-2.
Williams, playing for the first time since declaring on Tuesday her intention to retire after this month's US Open, was no match for Tokyo Olympics gold medalist Bencic.
The 23-time major winner was unable to claim back-to-back WTA singles wins for the first time since last year's French Open, having defeated Nuria Parrizas-Diaz for her first victory in 430 days on Monday.
Bencic triumphed in one hour and 17 minutes, winning 84.2 per cent of first-serve points and converted five of eight break points throughout the match.
Williams' power was on show with 13 winners, but Bencic was physically more capable and decisively managed 25 winners with only 13 unforced errors.
Elsewhere, 2019 US Open winner and local hope Bianca Andreescu edged Alize Cornet in a see-sawing clash in the evening, winning 6-3 4-6 6-3 in two hours and 26 minutes.
Fourth seed Paula Badosa and fifth seed Ons Jabeur, who was last month's Wimbledon runner-up, were forced to retire due to injury.
Spanish 24-year-old Badosa withdrew against Yulia Putintseva 7-5 1-0 due to muscle cramping, while Zheng Qinwen had a walkover against Jabeur 6-1 2-1 due to abdominal pain.
Top seed Iga Swiatek brushed aside Ajla Tomljanovic 6-1 6-2 in 64 minutes. The win means Swiatek is the first player to win 15 WTA 1000 matches in straight sets in a row since 2009.
Canadian 13th seed Leylah Fernandez also bowed out, going down 7-6 (7-4) 6-1 to Brazil's Beatriz Haddad Maia, who will face Swiatek in the third round.
Second seed Anett Kontaveit lost 6-4 6-4 to Jil Teichmann in one hour and 27 minutes. Teichmann will next face Simona Halep who won in 71 minutes against Zhang Shuai 6-4 6-2.
Sixth seed Aryna Sabalenka got past Sara Sorribes Tormo 6-4 6-3 to set up a third-round meeting with Coco Gauff after she defeated Wimbledon champion Elena Rybakina 6-4 6-7 (8-10) 7-6 (7-3) in an epic that lasted two hours and 49 minutes.
Jessica Pegula won 6-2 7-5 over American qualifier Asia Muhammad to progress through to face Camila Giorgi after she knocked off Elise Mertens 7-3 7-5.
Third seed Maria Sakkari triumphed in three sets 6-2 4-6 6-2 over Sloane Stephens and will face Karolina Pliskova next after the Czech beat Amanda Anisimova 6-1 6-1.
Eighth seed Garbine Muguruza won 6-4 6-4 against Kaia Kanepi and Alison Riske toppled 16th seed Jelena Ostapenko 7-6 (7-2) 0-6 7-5.
Swiatek made a disastrous start, going 4-0 down, and though she broke back once, was unable to find the second as Pliskova took the opening frame 6-4.
The determined Pole stepped up a level though to take the second 6-1, before breaking early in the third.
It was still an improvement for the Czech player, who lost 6-0 6-0 to Swiatek in their only previous meeting in Rome in 2021.
Pliskova kept her opponent honest on her own serve, but Swiatek had too much for the world number 17 as she clinched the decider 6-2.
Aryna Sabalenka also showcased her powers of recovery as she beat Paula Badosa despite going a set down.
The second seed won 4-6 6-4 6-4 in an even contest that saw nine breaks of serve, before the Belarusian eventually got over the line.
Sabalenka is now the player with the joint-most comeback wins over the last two seasons (13) level with Caroline Garcia, though the French star was actually the victim of a comeback herself on Friday.
Garcia won the first set against Anastasia Potapova, before the Russian came back to take it 4-6 6-3 6-3 to set up a semi against Sabalenka.
There was no need for such drama from Ons Jabeur, who eased to a 6-3 6-0 victory against Beatriz Haddad Maia in just 68 minutes to confirm a clash with Swiatek.
World number one Swiatek landed a 21st consecutive victory as she edged out US Open winner Raducanu 6-4 6-4 in an hour and 45 minutes on the German clay.
There was plenty to admire from both players, but in the end it was another straight-sets success for Swiatek, who dropped only two games in her previous round against German Eva Lys.
It makes the 20-year-old Pole the first woman to win 28 consecutive sets on tour since Serena Williams, who did so from the 2012 US Open to the 2013 Australian Open.
Swiatek broke early in the first set to take charge, and with 19-year-old Raducanu battling a back problem the rankings leader soon got ahead in the second too.
This was Raducanu's first-ever match against a player ranked inside the WTA top 10, a peculiar statistic given she is already a grand slam champion.
Swiatek, like her opponent, knows how it feels to win a grand slam as a teenage surprise package, having triumphed as a 19-year-old at the 2020 French Open when ranked only 54th in the world.
At 4-3 in the second set of this contest, Swiatek saved two break points with clinical forehand winners out of the reach of Raducanu, shouting out in satisfaction moments later as she held serve to move a game away.
Raducanu had two more break chances in Swiatek's next service game but again could not convert as her opponent sealed victory.
Swiatek said: "I'm pretty happy that today's match was longer. Not for now, but for the future it's going to give me a lot of experience.
"Right now I want to play really aggressively, and I think this game style is going to fit the surface, and it fit the hardcourts as well."
She will face unseeded Liudmila Samsonova next after the Russian, playing as a neutral, beat Laura Siegemund 7-5 6-3.
The other semi-final in Stuttgart will see second seed Paula Badosa take on third seed Aryna Sabalenka.
Badosa was a 7-6 (11-9) 1-6 6-3 winner against Ons Jabeur, while Sabalenka fended off Anett Kontaveit 6-4 3-6 6-1.
At the Istanbul Cup, Friday saw quarter-final wins for Veronika Kudermetova and Anastasia Potapova, along with Sorana Cirstea and Yulia Putintseva.
Those results set up a semi-final on Saturday between second seed Cirstea and third seed Kudermetova, with Putintseva and Potapova also facing off.
Swiatek was irrepressible against the 18-year-old American, extending her head-to-head dominance to 4-0 with a 6-0 6-3 victory in 66 minutes.
The Pole won the first eight games of the match and always appeared in control with her reliable forehand a key feature.
Swiatek laced 13 winners with only nine unforced errors, breaking Gauff five times. The American committed 26 unforced errors for the match.
The victory is Swiatek's 62nd of the season, moving her closer to Angelique Kerber's 2016 mark of 63.
Swiatek will face fourth seed Jessica Pegula in the semi-finals as the Pole aims for her eighth WTA title of the season.
Pegula secured her berth in the final four with a 6-4 7-5 victory over compatriot Madison Keys in their first-ever meeting in one hour and 34 minutes.
Third seed Aryna Sabalenka was a surprise casualty as Croatian qualifier Donna Vekic continued her excellent week with a 6-4 6-7 (5-7) 6-1 in two hours and 37 minutes.
World number 77 Vekic has beaten former world number three Maria Sakkari and two-time major finalist Karolina Pliskova this week.
Second seed Paula Badosa was also bundled out by American Danielle Collins 7-6 (7-5) 6-4. Collins will meet Vekic in the last four.
The 2022 French Open and US Open champion triumphed 6-4 4-6 6-1 in two hours and two minutes, bouncing back strongly after losing the second set.
Zheng's power and forehand top spin proved challenging for the Pole, who was solid from the baseline to secure her 11st tour quarter-finals appearance this season.
The Chinese had broken Swiatek to lead 1-0 in the third set, but the top seed responded in trademark style, improving her record in the United States this season to 21-1.
Swiatek will take on eighth seed Coco Gauff in the quarter-finals, with the match-up marking a re-match of this year's French Open final.
Gauff worked her way into the last eight with a 6-4 4-6 6-3 victory over 2019 US Open champion Bianca Andreescu.
The 18-year-old American also had to fight back from a set down in the third set to win in a match that lasted just over two-and-a-half hours.
Third seed Aryna Sabalenka fought back from a lopsided opening set to knock out American Sloane Stephens 1-6 6-3 6-2 in two hours and one minute. The Belarussian will take on Donna Vekic in the next round.
Second seed Paula Badosa eased into the last eight, winning in 53 minutes against American qualifier Louisa Chirico 6-0 6-3. Badosa will face Danielle Collins in the quarter-finals.
Madison Keys triumphed over eighth seed Daria Kasatkina 6-4 6-3 in one hour and 27 minutes to secure a quarter-final date with fourth seed Jessica Pegula.
Swiatek, who became the first female player to win 25 consecutive matches since Serena Williams in 2015, will face Bianca Andreescu in the last eight after she eased past Petra Martic 6-4 6-4.
"I didn't start well, and everybody could see that," Swiatek told reporters. "I'm really happy with the way I reacted and how I improved in the first set.
"Also how different the second set looked to the first one because I could really reset and really change the way I played. That's the most positive thing for me."
There was a shock in the final game of the day, though, as Daria Kasatkina dumped out number two seed Paula Badosa 6-4 6-4.
Third seed Aryna Sabalenka sealed a third straight win over Jessica Pegula, easing to a 6-1 6-4 victory, while fourth seed Maria Sakkari defeated Coco Gauff 6-4 7-5.
Seventh seed Danielle Collins was knocked out by unseeded compatriot Amanda Anisimova, the 20-year-old cruising past the Australian Open finalist 6-2 6-2.
There were also victories for Jil Teichmann, who overcame Elena Rybakina 6-7 (3-7) 6-3 7-5, and ninth seed Ons Jabeur, who beat Yulia Putintseva 6-3 6-2.
Swiatek had not played since sustaining the issue in the semi-finals of Indian Wells over a month ago, but advanced on Thursday despite stating that she felt "rusty".
The world number one won 84 per cent of points after landing her first serve in as she made up for lost time.
Swiatek also forced eight break points, winning four of them as she took just and hour and 26 minutes to set up a quarter-final with Karolina Pliskova, who bested Donna Vekic in a thriller.
Pliskova looked to be on her way to a routine win as she claimed the first set 6-2, only for the Croatian to take the second via a tie-break.
The decider also went the distance, with Pliskova able to finally put Vekic away 6-2 6-7 (5-7) 7-6 (7-5) to book her clash with Swiatek.
Coco Gauff perhaps paid the price for taking almost three hours to beat Veronika Kudermetova on Wednesday, as the fifth seed fell to a straight-sets loss to Anastasia Potapova on Thursday, going down 6-2 6-3.
Fourth seed Caroline Garcia will go up against Potapova next after she defeated Tatjana Maria 7-6 (7-5) 6-4, while Paula Badosa had few problems seeing off fellow Spaniard Cristina Bucsa 6-1 6-2 and will face second seed Aryna Sabalenka in the last eight.
The world number one produced a 22nd consecutive victory as she scraped a 6-7 (4-7) 6-4 7-5 win in a contest that lasted more than three hours.
The Pole will face third seed Aryna Sabalenka in Sunday's final on the German clay.
Swiatek broke early to race out to a 3-0 lead, but was pegged back by her Russian opponent, who fought hard to claim the first set on a tie-break.
The 20-year-old was looking to break the record of Serena Williams, equalling a feat of winning 28 sets in a row, but Samsonova prevented her from doing so.
It was the first time Swiatek had dropped a set since her Indian Wells Open last 16 match against Angelique Kerber in March, but she soon got back into her rhythm and clinched the second set 6-4.
She broke early again in the decider, but was once more broken back by a determined Samsonova, and Swiatek showed frustration with herself as she struggled to put away her opponent.
However, an unusually sloppy service game from Samsonova gave Swiatek another break in the 11th game of the set, which she closed out to seal her place in the final.
The other semi-final in Stuttgart saw Sabalenka overcome second seed Paula Badosa 7-6 (7-5) 6-4.
There were 14 double faults (seven each) in the match, but it was Sabalenka's big serve that ultimately led her to victory, hitting nine aces and winning 76.9 per cent of points on her first serve.
The Belarusian also saved six of eight break points faced as she ultimately eased past her Spanish opponent.
At the Istanbul Cup, third seed Veronika Kudermetova will play Anastasia Potapova in the final after seeing off second seed Sorana Cirstea in straight sets, 6-3 6-3.
Potapova had earlier come from a set down to beat Yulia Putintseva 2-6 6-2 6-2 in the other semi-final.
Top seed Switaek, seeking a fifth consecutive title, breezed past Elena-Gabriela Ruse 6-3 6-0 as she became the first female player to win 24 consecutive matches since Serena Williams in 2015.
Badosa followed suit by dispatching Aliaksandra Sasnovich 6-2 6-1 in just an hour and 17 minutes, teeing up a third-round clash with Daria Kasatkina, who beat Leylah Fernandez 4-6 6-3 7-6 (7-2).
Aryna Sabalenka, the third favourite in Italy, also made light work of Zhang Shuai to triumph 6-2 6-0, while fourth seed Maria Sakkari cruised past Madrid Open semi-finalist Ekaterina Alexandrova 6-3 6-2.
Sakkari will next face a Rome rematch with Coco Gauff, who defeated compatriot Madison Brengle 6-2 6-4. Teenager Gauff's sole win against Sakkari came at this tournament in the second round last year.
Yulia Putintseva came from behind to beat Garbine Muguruza 3-7 7-6 (7-4) 6-1, and will next meet Ons Jabeur, who made it eight successive main-draw wins by defeating Ajla Tomljanovic 7-5 6-2.
Seventh seed Danielle Collins also battled past former world number one Simona Halep 7-6 (7-1) 6-3.
There was no such joy for fifth seed Anett Kontaveit as she crashed out to Petra Martic in straight sets, while Jil Teichmann upset former Rome champion Karolina Pliskova in a three-set thriller.
Belinda Bencic, who was ranked as 12th seed, was another early casualty as she fell to a 7-6 (7-4) 6-1 defeat against American Amanda Anisimova.
Meanwhile, Jessica Pegula secured her third-round spot after Anhelina Kalinina withdrew due to injury, with Victoria Azarenka earning a straightforward 6-2 6-4 victory over Camila Osorio to get a shot at favourite Swiatek in the next round.
The world number one had won the first set 7-5, but her Australian opponent was forced to concede at 2-2 in the second.
Swiatek is now the female player with the most wins in a single year (58) in the last five seasons. Former world number one Ash Barty recorded 57 victories in 2019.
The second seed in Ostrava is out after Paula Badosa was beaten 7-6 (7-4) 6-4 by Petra Kvitova in front of a delighted Czech crowd, while Elena Rybakina is also through to the last eight after coming from behind to win against Aliaksandra Sasnovich 6-7 (5-7) 6-4 7-5.
Wednesday's two other matches saw more success for Czech participants as Karolina Muchova beat seventh seed Beatriz Haddad Maia 6-4 6-4, while Barbora Krejcikova defeated Shelby Rogers 6-2 6-2.
There was less Czech joy at the Jasmin Open in Tunisia, where Katerina Siniakova was beaten 7-5 6-2 by Claire Liu.
Elsewhere, number three seed Alize Cornet eased past Harriet Dart 6-3 6-4, Elise Mertens came from behind to beat Despina Papamichail and Diane Parry went through against French compatriot Lucrezia Stefanini after the latter retired hurt in the second set.
