An Infinite Needle in a Finite Haystack: Finding Infinite Counter-Models in Deductive Verification

First-order logic, and quantifiers in particular, are widely used in deductive verification. Quantifiers are essential for describing systems with unbounded domains, but prove difficult for automated solvers. Significant effort has been dedicated to finding quantifier instantiations that establish unsatisfiability, thus ensuring validity of a system's verification conditions. However, in many cases the formulas are satisfiable: this is often the case in intermediate steps of the verification process. For such cases, existing tools are limited to finding finite models as counterexamples. Yet, some quantified formulas are satisfiable but only have infinite models. Such infinite counter-models are especially typical when first-order logic is used to approximate inductive definitions such as linked lists or the natural numbers. The inability of solvers to find infinite models makes them diverge in these cases. In this paper, we tackle the problem of finding such infinite models. These models allow the user to identify and fix bugs in the modeling of the system and its properties. Our approach consists of three parts. First, we introduce symbolic structures as a way to represent certain infinite models. Second, we describe an effective model finding procedure that symbolically explores a given family of symbolic structures. Finally, we identify a new decidable fragment of first-order logic that extends and subsumes the many-sorted variant of EPR, where satisfiable formulas always have a model representable by a symbolic structure within a known family. We evaluate our approach on examples from the domains of distributed consensus protocols and of heap-manipulating programs. Our implementation quickly finds infinite counter-models that demonstrate the source of verification failures in a simple way, while SMT solvers and theorem provers such as Z3, cvc5, and Vampire diverge

Transcatheter edge-to-edge mitral valve repair in patients with acute decompensated heart failure due to severe mitral regurgitation

Background: Transcatheter edge-to-edge mitral valve repair (TEER) has been established as a therapy for severe symptomatic mitral regurgitation (MR) in stable patients, and it has recently emerged as a reasonable option for acutely ill patients. The aim of this study was to evaluate the safety and efficacy of TEER in hospitalized patients with acute decompensated heart failure (ADHF) and severe MR that was deemed to play a major role in their deterioration. Methods: We included 31 patients who underwent emergent TEER for MR ≥ 3+ from 2012 to 2022 at Sheba Medical Center. Outcomes included procedural safety, procedural success, all-cause mortality, heart failure readmission, and functional improvement. Outcomes were evaluated at 3 months and at 1 year. Data were obtained retrospectively by chart review. Results: Implantation of a TEER device was achieved in 97% of patients, and reduction in MR severity of at least two grades and final MR ≤ 2+ at discharge was achieved in 74%. No intra-procedural mortality or life-threatening complications were noted. Mortality at 30 days was 23%. No excess mortality occurred beyond 6 months, with a total mortality of 41%. At 1 year all survivors had MR ≤ 2+, all were free of heart failure hospitalizations, and 88% were at New York Heart Association class ≤ II. Conclusions: Mitral valve TEER for patients with ADHF and significant MR is safe, feasible, and achieves substantial reduction in MR severity. Despite high early mortality, procedural success is associated with good long-term clinical outcomes for patients surviving longer than 6 months

Tumor-Targeted Fluorescent Proteinoid Nanocapsules Encapsulating Synergistic Drugs for Personalized Cancer Therapy

Personalized cancer treatment based on specific mutations offers targeted therapy and is preferred over “standard” chemotherapy. Proteinoid polymers produced by thermal step-growth polymerization of amino acids may form nanocapsules (NCs) that encapsulate drugs overcoming miscibility problems and allowing passive targeted delivery with reduced side effects. The arginine-glycine-glutamic acid (RGD) sequence is known for its preferential attraction to αvβ3 integrin, which is highly expressed on neovascular endothelial cells that support tumor growth. Here, tumor-targeted RGD-based proteinoid NCs entrapping a synergistic combination of Palbociclib (Pal) and Alpelisib (Alp) were synthesized by self-assembly to induce the reduction of tumor cell growth in different types of cancers. The diameters of the hollow and drug encapsulating poly(RGD) NCs were 34 ± 5 and 22 ± 3 nm, respectively; thereby, their drug targeted efficiency is due to both passive and active targeting. The encapsulation yield of Pal and Alp was 70 and 90%, respectively. In vitro experiments with A549, MCF7 and HCT116 human cancer cells demonstrate a synergistic effect of Pal and Alp, controlled release and dose dependence. Preliminary results in a 3D tumor spheroid model with cells derived from patient-derived xenografts of colon cancer illustrate disassembly of spheroids, indicating that the NCs have therapeutic potential

Measurement of apoptosis by SCAN©, a system for counting and analysis of fluorescently labelled nuclei

Apoptosis-like programmed cell death (A-PCD) is a universal process common to all types of eukaryotic organisms. Because A-PCD-associated processes are conserved, it is possible to define A-PCD by a standard set of markers. Many of the popular methods to measure A-PCD make use of fluorescent ligands that change in intensity or cellular localization during A-PCD. In single cell organisms, it is possible to quantify levels of A-PCD by scoring the number of apoptotic cells using flow cytometry instruments. In a multicellular organism, quantification of A-PCD is more problematic due to the complex nature of the tissue. The situation is further complicated in filamentous fungi, in which nuclei are divided between compartments, each containing a number of nuclei, which can also migrate between the compartments. We developed SCAN©, a System for Counting and Analysis of Nuclei, and used it to measure A-PCD according to two markers – chromatin condensation and DNA strand breaks. The package includes three modules designed for counting the number of nuclei in multi-nucleated domains, scoring the relative number of nuclei with condensed chromatin, and calculating the relative number of nuclei with DNA strand breaks. The method provides equal or better results compared with manual counting, the analysis is fast and can be applied on large data sets. While we demonstrated the utility of the software for measurement of A-PCD in fungi, the method is readily adopted for measurement of A-PCD in other types of multicellular specimens

The bumblebee Bombus terrestris carries a primary inoculum of Tomato brown rugose fruit virus contributing to disease spread in tomatoes.

The bumblebee Bombus terrestris is a beneficial pollinator extensively used in tomato production. Our hypothesis was that bumblebee hives collected from a Tomato brown rugose fruit virus (ToBRFV) infected tomato greenhouse, preserve an infectious primary inoculum. Placing a bumblebee hive collected from a ToBRFV contaminated greenhouse, in a glass-/net-house containing only uninfected healthy tomato plants, spread ToBRFV disease. Control uninfected tomato plants grown in a glass-/net-house devoid of any beehive remained uninfected. ToBRFV-contaminated hives carried infectious viral particles as demonstrated in a biological assay on laboratory test plants of virus extracted from hive components. Viral particles isolated from a contaminated hive had a typical tobamovirus morphology observed in transmission electron microscopy. Assembly of ToBRFV genome was achieved by next generation sequencing analysis of RNA adhering to the bumblebee body. Bumblebee dissection showed that ToBRFV was mostly present in the abdomen suggesting viral disease spread via buzz pollination. These results demonstrate that bumblebee hives collected from ToBRFV-contaminated greenhouses carry a primary inoculum that reflects the status of viruses in the growing area. This new mode of ToBRFV spread by pollinators opens an avenue for detection of viruses in a growing area through analysis of the pollinators, as well as emphasizes the need to reevaluate the appropriate disease management protocols

Genomic characterization of viruses associated with the parasitoid Anagyrus vladimiri (Hymenoptera: Encyrtidae)

International audienceKnowledge on symbiotic microorganisms of insects has increased dramatically in recent years, yet relatively little data are available regarding non-pathogenic viruses. Here we studied the virome of the parasitoid wasp Anagyrus vladimiri Triapitsyn (Hymenoptera: Encyrtidae), a biocontrol agent of mealybugs. By high-throughput sequencing of viral nucleic acids, we revealed three novel viruses, belonging to the families Reoviridae [provisionally termed AnvRV (Anagyrus vladimiri reovirus)], Iflaviridae (AnvIFV) and Dicistroviridae (AnvDV). Phylogenetic analysis further classified AnvRV in the genus Idnoreovirus , and AnvDV in the genus Triatovirus . The genome of AnvRV comprises 10 distinct genomic segments ranging in length from 1.5 to 4.2 kb, but only two out of the 10 ORFs have a known function. AnvIFV and AnvDV each have one polypeptide ORF, which is typical of iflaviruses but very un-common among dicistroviruses. Five conserved domains were found along both the ORFs of those two viruses. AnvRV was found to be fixed in an A. vladimiri population that was obtained from a mass rearing facility, whereas its prevalence in field-collected A. vladimiri was ~15 %. Similarly, the prevalence of AnvIFV and AnvDV was much higher in the mass rearing population than in the field population. The presence of AnvDV was positively correlated with the presence of Wolbachia in the same individuals. Transmission electron micrographs of females’ ovaries revealed clusters and viroplasms of reovirus-like particles in follicle cells, suggesting that AnvRV is vertically transmitted from mother to offspring. AnvRV was not detected in the mealybugs, supporting the assumption that this virus is truly associated with the wasps. The possible effects of these viruses on A. vladimiri ’s biology, and on biocontrol agents in general, are discussed. Our findings identify RNA viruses as potentially involved in the multitrophic system of mealybugs, their parasitoids and other members of the holobiont

Identification and Localization of a Rickettsia sp. in Bemisia tabaci (Homoptera: Aleyrodidae)

Whiteflies (Homoptera: Aleyrodidae) are sap-sucking insects that harbor “Candidatus Portiera aleyrodidarum,” an obligatory symbiotic bacterium which is housed in a special organ called the bacteriome. These insects are also home for a diverse facultative microbial community which may include Hamiltonella, Arsenophonus, Fritchea, Wolbachia, and Cardinium spp. In this study, the bacteria associated with a B biotype of the sweet potato whitefly Bemisia tabaci were characterized using molecular fingerprinting techniques, and a Rickettsia sp. was detected for the first time in this insect family. Rickettsia sp. distribution, transmission and localization were studied using PCR and fluorescence in situ hybridizations (FISH). Rickettsia was found in all 20 Israeli B. tabaci populations screened but not in all individuals within each population. A FISH analysis of B. tabaci eggs, nymphs, and adults revealed a unique concentration of Rickettsia around the gut and follicle cells, as well as a random distribution in the hemolymph. We postulate that the Rickettsia enters the oocyte together with the bacteriocytes, leaves these symbiont-housing cells when the egg is laid, multiplies and spreads throughout the egg during embryogenesis and, subsequently, disperses throughout the body of the hatching nymph, excluding the bacteriomes. Although the role Rickettsia plays in the biology of the whitefly is currently unknown, the vertical transmission on the one hand and the partial within-population infection on the other suggest a phenotype that is advantageous under certain conditions but may be deleterious enough to prevent fixation under others

A New Israeli <i>Tobamovirus</i> Isolate Infects Tomato Plants Harboring <i>Tm-2²</i> Resistance Genes

<div><p>An outbreak of a new disease infecting tomatoes occurred in October-November 2014 at the Ohad village in Southern Israel. Symptomatic plants showed a mosaic pattern on leaves accompanied occasionally by narrowing of leaves and yellow spotted fruit. The disease spread mechanically and rapidly reminiscent of tobamovirus infection. Epidemiological studies showed the spread of the disease in various growing areas, in the South and towards the Southeast and Northern parts of the country within a year. Transmission electron microscope (TEM) analysis showed a single rod-like form characteristic to the <i>Tobamovirus</i> genus. We confirmed Koch’s postulates for the disease followed by partial host range determination and revealed that tomato cultivars certified to harbor the <i>Tm-2</i>^<i>2</i> resistance gene are susceptible to the new viral disease. We further characterized the viral source of the disease using a range of antisera for serological detection and analyzed various virus genera and families for cross-reactivity with the virus. In addition, next generation sequencing of total small RNA was performed on two cultivars grown in two different locations. In samples collected from commercial cultivars across Israel, we found a single virus that caused the disease. The complete genome sequence of the new Israeli tobamovirus showed high sequence identity to the Jordanian isolate of tomato brown rugose fruit virus.</p></div