Development of shotgun metabolomic profile analysis for detecting canine visceral leishmaniasis using flow-through pinhole paper spray mass spectrometry

Visceral leishmaniasis (<b>VL</b>) is a neglected infectious disease that can be transmitted between dogs and humans. In this work, a novel flow-through pinhole paper spray mass spectrometry (<b>MS</b>) technique is described to detect metabolites that might be up- or down-regulated during canine VL infection. The flow-through pinhole paper spray MS allows direct analysis of dried samples prepared in paper substrates, without prior sample pre-treatment. Here, clinical canine serum samples from ten dogs positives for VL were prepared in embossed hydrophobic paper substrates in the lab. Similar dried serum samples from uninfected control dogs were prepared in embossed paper. Partial least squares discriminant analysis was performed on positive-ion mode full mass spectra recorded from the 20 samples, which showed significant separation between VL positive and negative samples. Volcano plots confirmed the presence of several up- and down-regulated species in the infected dog samples. Results from this work demonstrate detectable metabolomic changes can be measured during canine VL from dried serum samples without sample preparation.