Bone morphogenetic protein (BMP) signalling regulates embryonic dorsal-ventral cell fate decisions in flies, frogs and fish. BMP activity is controlled by several secreted factors including the antagonists chordin and short gastrulation (SOG). Here we show that a second secreted protein, Twisted gastrulation (Tsg), enhances the antagonistic activity of Sog/chordin. In Drosophila, visualization of BMP signalling using anti-phospho-Smad staining shows that the tsg and sog loss-of-function phenotypes are very similar. In S2 cells and imaginal discs, TSG and SOG together make a more effective inhibitor of BMP signalling than either of them alone. Blocking Tsg function in zebrafish with morpholino oligonucleotides causes ventralization similar to that produced by chordin mutants. Co-injection of sub-inhibitory levels of morpholines directed against both Tsg and chordin synergistically enhances the penetrance of the ventralized phenotype. We show that Tsgs from different species are functionally equivalent, and conclude that Tsg is a conserved protein that functions with SOG/chordin to antagonize BMP signalling.

Background

There have been various efforts to identify less costly but still accurate methods for monitoring the response to HIV treatment. We evaluated a pooling method to determine if this could improve screening efficiency and reduce costs while maintaining accuracy in Seoul, South Korea.

Methods

We conducted the first prospective study of pooled nucleic acid testing (NAT) using a 5 minipool + algorithm strategy versus individual viral load testing for patients receiving antiretroviral therapy (ART) between November 2011 and August 2012 at an urban hospital in Seoul, South Korea. The viral load assay used has a lower level of detection of 20 HIV RNA copies/ml, and the cost per assay is US$ 136. The 5 minipool +algorithm strategy was applied and 43 pooled samples were evaluated. The relative efficiency and accuracy of the pooled NAT were compared with those of individual testing.

Results

Using the individual viral load assay, 15 of 215 (7%) plasma samples had more than 200 HIV RNA copies/ml. The pooled NAT using the 5 minipool + algorithm strategy was applied to 43 pooled samples; 111 tests were needed to test all samples when virologic failure was defined at HIV RNA ≥ 200 copies/ml. Therefore, 104 tests were saved over individual testing, with a relative efficiency of 0.48. When evaluating costs, a total of US$ 14,144 was saved for 215 individual samples during 10 months. The negative predictive value was 99.5% for all samples with HIV RNA ≥ 200 copies/ml.

Conclusions

The pooled NAT with 5 minipool + algorithm strategy seems to be a very promising approach to effectively monitor patients receiving ART and to save resources.

Less costly but still accurate methods for monitoring HIV treatment response are needed. We prospectively evaluated if a qualitative polymerase chain reaction (PCR) amplification assay for virologic monitoring could maintain accuracy while reducing costs in Seoul, South Korea. We conducted the first prospective study comparing a qualitative PCR amplification of HIV-1 reverse transcriptase (RT) versus a commercial real time PCR assay (i.e., viral load) for virologic monitoring of 150 patients receiving antiretroviral therapy (ART) between November 2011 and August 2012 at an urban hospital in Seoul, South Korea. A total of 215 blood plasma samples from 150 patients receiving ART for more than 6 months were evaluated. Using the individual viral load assay, 12 of 215 (5.6%) plasma samples had more than 500 HIV RNA copies/ml. The qualitative PCR amplification assay detected individual samples with ≥500 HIV RNA copies/ml with 100% sensitivity. The specificities of the qualitative PCR amplification of the HIV-1 RT assay were 94.1%, 93.6%, and 93.2% compared to the real time PCR at 500, 1,000, and 5,000 threshold of HIV RNA copies/ml, respectively, and $24,940 USD would have been saved for 150 patients during 10 months. The qualitative PCR amplification of the HIV-1 RT assay might be a useful approach to effectively monitor patients receiving ART and save resources.