The B and Q 'biotypes' of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) have been invading many parts of the world and causing severe damage to a range of crops. Recent phylogenetic analyses indicate that B and Q are cryptic species within the B. tabaci species complex. Although various attempts have been made to examine the reproductive compatibility between B and Q, few studies have tested the fertility of the F1 females and so the extent of possible gene flow remains unclear. In this study, we conducted a series of crossing experiments and behavioural observations to examine in detail the reproductive compatibility between the B and Q biotypes collected from Zhejiang, China, a region recently invaded by these whiteflies. Crossing experiments between the two biotypes using either single-pairs or small groups demonstrated that proportions of females in the F1 progeny were only 0-2% in the inter-biotype crosses compared to 58-68% in the intra-biotype treatments. Furthermore, all inter-biotype F1 females were sterile. Continuous video observations showed that B and Q adults very rarely copulated, and copulation occurred only when adults of opposite sex from different biotypes were enclosed in dense cohorts for a relatively long period of time. These data show that the B and Q biotypes examined in this study are completely isolated in reproduction. The isolation was due to mainly a copulation barrier, but post-copulation barriers were also involved.