Tritrichomonas foetus isolates from cats and cattle show minor genetic differences in unrelated loci ITS-2 and EF-1alpha

The protozoan parasite Tritrichomonas foetus is well known as an important causative agent of infertility and abortion in cattle (bovine trichomonosis). This World Organisation for Animal Health (O.I.E.) notifiable disease is thought to be under control in many countries including Switzerland. In recent studies, however, T. foetus has also been identified as an intestinal parasite that causes chronic large-bowel diarrhoea in cats. Since the feline isolates were considered indistinguishable from bovine isolates, the possibility and risk of parasite transmission from cats to cattle and vice versa has been intensively discussed in current literature. Therefore, we investigated if cat and cattle isolates are genetically distinct from each other or in fact represent identical genotypes. For this purpose, two independent genetic loci were selected that turned out to be well-suited for a PCR sequencing-based genotyping of trichomonad isolates: (i) previously published internal transcribed spacer region 2 (ITS-2) and (ii) a semi-conserved sequence stretch of the elongation factor-1 alpha (EF-1alpha) gene used for the first time in the present study. Respective comparative analyses revealed that both loci were sufficiently variable to allow unambiguous genetic discrimination between different trichomonad species. Comparison of both genetic loci confirmed that T. suis and T. mobilensis are phylogenetically very close to T. foetus. Moreover, these two genetic markers were suited to define host-specific genotypes of T. foetus. Both loci showed single base differences between cat and cattle isolates but showed full sequence identity within strains from either cat or cattle isolates. Furthermore, an additional PCR with a forward primer designed to specifically amplify the bovine sequence of EF-1alpha was able to discriminate bovine isolates of T. foetus from feline isolates and also from other trichomonads. The implications these minor genetic differences may have on the biological properties of the distinct isolates remain to be investigated.

Unexpected sensitivity of sst2 antagonists to N-terminal radiometal modifications

Chelated somatostatin agonists have been shown to be sensitive to N-terminal radiometal modifications, with Ga-DOTA agonists having significantly higher binding affinity than their Lu-, In-, and Y-DOTA correlates. Recently, somatostatin antagonists have been successfully developed as alternative tracers to agonists. The aim of this study was to evaluate whether chelated somatostatin antagonists are also sensitive to radiometal modifications and how. We have synthesized 3 different somatostatin antagonists, DOTA-p-NO(2)-Phe-c[D-Cys-Tyr-D-Aph(Cbm)-Lys-Thr-Cys]-D-Tyr-NH(2), DOTA-Cpa-c[D-Cys-Aph(Hor)-D-Aph(Cbm)-Lys-Thr-Cys]-D-Tyr-NH(2) (DOTA-JR11), and DOTA-p-Cl-Phe-c[D-Cys-Tyr-D-Aph(Cbm)-Lys-Thr-Cys]-D-Tyr-NH(2), and added various radiometals including In(III), Y(III), Lu(III), Cu(II), and Ga(III). We also replaced DOTA with 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA) and added Ga(III). The binding affinity of somatostatin receptors 1 through 5 was evaluated in all cases. In all 3 resulting antagonists, the Ga-DOTA analogs were the lowest-affinity radioligands, with a somatostatin receptor 2 binding affinity up to 60 times lower than the respective Y-DOTA, Lu-DOTA, or In-DOTA compounds. Interestingly, however, substitution of DOTA by the NODAGA chelator was able to increase massively its binding affinity in contrast to the Ga-DOTA analog. The 3 NODAGA analogs are antagonists in functional tests. In vivo biodistribution studies comparing (68)Ga-DOTATATE agonist with (68)Ga-DOTA-JR11 and (68)Ga-NODAGA-JR11 showed not only that the JR11 antagonist radioligands were superior to the agonist ligands but also that (68)Ga-NODAGA-JR11 was the tracer of choice and preferable to (68)Ga-DOTA-JR11 in transplantable HEK293-hsst(2) tumors in mice. One may therefore generalize that somatostatin receptor 2 antagonists are sensitive to radiometal modifications and may preferably be coupled with a (68)Ga-NODAGA chelator-radiometal complex.

Point-of-care prothrombin time testing in paediatric intensive care: an observational study of the ease of use of two devices

Two major difficulties arise when taking blood samples in children: the challenge of venous access and the comparatively large amount of blood required.