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Background  Tsetse flies and trypanosomiasis are among several factors that constrain livestock development in Tanzania. Over the years Rufiji District was excluded from livestock production owing to tsetse fly infestation, however, a few years ago there was an influx of livestock following evictions aimed at conserving the Usangu wetlands.    Methods  A study was conducted to determine the efficiency of available traps for catching tsetse flies,  Glossina  species infesting the area, their infection rates and  Trypanosoma  species circulating in the area. Trapping was conducted during the semi dry season for a total of 30 days (ten days each month) during the onset of the dry season of May - July 2009. Harvested flies after every 24 hours were dissected and examined under a light microscope for trypanosome infections and whole fly DNA was extracted from 82 flies and analyzed for trypanosomes by polymerase chain reaction (PCR) using different sets of primers.    Results  The proportions of total tsetse catches per trap were in the following decreasing order S3 (33%), H-Trap (27%), Pyramidal (19%), sticky panel (11%) and biconical trap (10%). Of the 1200 trapped flies, 75.6% were identified as  Glossina pallidipes , 11.7%  as G. brevipalpis , 9.6% as  G. austeni  and 3.0%  G. morsitans morsitans . Dissections revealed the overall infection rate of 6.6% (13/197). Whole DNA was extracted from 82 tsetse flies and the prevalence of trypanosomes circulating in the area in descending order was 92.7% (76/82) for  T. simiae ; 70.7% (58/82) for  T. brucei  types; 48.8% (40/82) for the  T. vivax  types and 32.9% (27/82) for the  T. congolense  types as determined by PCR. All trypanosome types were found in all tsetse species analysed except for the  T. congolense  types, which were absent in  G. m. morsitans . None of the  T. brucei  positive samples contained human infective trypanosomes by SRA - PCR test    Conclusion  All tsetse species found in Rufiji are biologically important in the transmission of animal trypanosomiasis and the absence of  T. congolense  in  G. m. morsitans  could be a matter of chance only. Therefore, plans for control should consider all tsetse species.

Multiple Trypanosoma infections are common amongst Glossina species in the new farming areas of Rufiji district, Tanzania