Probabilistic reasoning with an enzyme-driven DNA device

We present a biomolecular probabilistic model driven by the action of a DNA toolbox made of a set of DNA templates and enzymes that is able to perform Bayesian inference. The model will take single-stranded DNA as input data, representing the presence or absence of a specific molecular signal (the evidence). The program logic uses different DNA templates and their relative concentration ratios to encode the prior probability of a disease and the conditional probability of a signal given the disease. When the input and program molecules interact, an enzyme-driven cascade of reactions (DNA polymerase extension, nicking and degradation) is triggered, producing a different pair of single-stranded DNA species. Once the system reaches equilibrium, the ratio between the output species will represent the application of Bayes? law: the conditional probability of the disease given the signal. In other words, a qualitative diagnosis plus a quantitative degree of belief in that diagno- sis. Thanks to the inherent amplification capability of this DNA toolbox, the resulting system will be able to to scale up (with longer cascades and thus more input signals) a Bayesian biosensor that we designed previously.

Translocation and Functional Analysis of Pseudomonas savastanoi pv. savastanoi NCPPB 3335 Type III Secretion System Effectors Reveals Two Novel Effector Families of the Pseudomonas syringae Complex

Pseudomonas savastanoi pv. savastanoi NCPPB 3335 causes olive knot disease and is a model pathogen for exploring bacterial infection of woody hosts. The type III secretion system (T3SS) effector repertoire of this strain includes 31 effector candidates plus two novel candidates identified in this study which have not been reported to translocate into plant cells. In this work, we demonstrate the delivery of seven NCPPB 3335 effectors into Nicotiana tabacum leaves, including three proteins from two novel families of the P. syringae complex effector super-repertoire (HopBK and HopBL), one of which comprises two proteins (HopBL1 and HopBL2) that harbor a SUMO protease domain. When delivered by P. fluorescens heterologously expressing a P. syringae T3SS, all seven effectors were found to suppress the production of defense-associated reactive oxygen species. Moreover, six of these effectors, including the truncated versions of HopAA1 and HopAZ1 encoded by NCPPB 3335, suppressed callose deposition. The expression of HopAZ1 and HopBL1 by functionally effectorless P. syringae pv. tomato DC3000D28E inhibited the hypersensitive response in tobacco and, additionally, expression of HopBL2 by this strain significantly increased its competitiveness in N. benthamiana. DNA sequences encoding HopBL1 and HopBL2 were uniquely detected in a collection of 31 P. savastanoi pv. savastanoi strains and other P. syringae strains isolated from woody hosts, suggesting a relevant role of these two effectors in bacterial interactions with olive and other woody plants.