Low guanine content and biased nucleotide distribution in vertebrate mtDNA can cause overestimation of non-CpG methylation

ABSTRACT Mitochondrial DNA (mtDNA) methylation in vertebrates has been hotly debated for over 40 years. Most contrasting results have been reported following bisulfite sequencing (BS-seq) analyses. We addressed whether BS-seq experimental and analysis conditions influenced the estimation of the levels of methylation in specific mtDNA sequences. We found false positive non-CpG methylation in the CHH context (fpCHH) using unmethylated Sus scrofa mtDNA BS-seq data. fpCHH methylation was detected on the top/plus strand of mtDNA within low guanine content regions. These top/plus strand sequences of fpCHH regions would become extremely AT-rich sequences after BS-conversion, whilst bottom/minus strand sequences remained almost unchanged. These unique sequences caused BS-seq aligners to falsely assign the origin of each strand in fpCHH regions, resulting in false methylation calls. fpCHH methylation detection was enhanced by short sequence reads, short library inserts, skewed top/bottom read ratios and non-directional read mapping modes. We confirmed no detectable CHH methylation in fpCHH regions by BS-amplicon sequencing. The fpCHH peaks were located in the D-loop, ATP6, ND2, ND4L, ND5 and ND6 regions and identified in our S. scrofa ovary and oocyte data and human BS-seq data sets. We conclude that non-CpG methylation could potentially be overestimated in specific sequence regions by BS-seq analysis.

Identifying Blunt Force Traumatic Injury on Thermally Altered Remains: A Pilot Study Using Sus scrofa

In forensic scenarios involving homicide, human remains are often exposed to fire as a means of disposal and/or obscuring identity. Burning human remains can result in the concealment of traumatic injury, the creation of artifacts resembling injury, or the destruction of preexisting trauma. Since fire exposure can greatly influence trauma preservation, methods to differentiate trauma signatures from burning artifacts are necessary to conduct forensic analyses. Specifically, in the field of forensic anthropology, criteria to distinguish trauma from fire signatures on bone is inconsistent and sparse. This study aims to supplement current forensic anthropological literature by identifying criteria found to be the most diagnostic of fire damage or blunt force trauma. Using the skulls of 11 adult pigs (Sus scrofa), blunt force trauma was manually produced using a crowbar and flat-faced hammer. Three specimens received no impacts and were utilized as controls. All skulls were relocated to an outdoor, open-air fire where they were burned until a calcined state was achieved across all samples. Results from this experiment found that blunt force trauma signatures remained after burning and were identifiable in all samples where reassociation of fragments was possible. This study concludes that distinct patterns attributed to thermal fractures and blunt force fractures are identifiable, allowing for diagnostic criteria to be narrowed down for future analyses.

First report of Eimeria sp. from Eurasian wild boar (Sus scrofa) in Iran

In Iran, wild boar is distributed throughout the country, except the Central Desert. One of the important habitats of this animal is the northern mountainous area. They may pass infected faeces in these areas, which can lead to zoonotic disease, though information about parasitic infections in wild boar is rare in Iran and further study is necessary. In February 2019, a wild boar carcass was submitted to the Veterinary Parasitology Department of the Science and Research University of Tehran by the Department of Environment of Mazandaran province in order to study its parasitic infections. Faeces and the gastrointestinal tract were separated and their contents were tested using two flotation methods (Willis and Clayton-Lane), and the samples were referred to the Parasitology Laboratory of Urmia University for further study. The sample was infected with Eimeria protozoa; Eimeria porci and Eimeria neodebliecki were diagnosed based on the size and morphology factors of the isolated oocysts. This is the first report of wild boar infection by Eimeria sp. protozoa in Iran.

Genetic diversity of the Central European wild boar (Sus scrofa scrofa) population and domestic pig (Sus scrofa domesticus) breeds based on a microsatellite DNA locus

The results of studies of the genetic structure of the Central European wild boar (Sus scrofa scrofa) population and four breeds of domestic pigs (Duroc, Yorkshire, Large White and Landrace) bred in the Central Black Earth region of Russia are presented in this work. Based on 12 microsatellite loci, a significant ( p <0.05) decrease in the level of genetic variability in bred breeds was shown. The expected heterozygosity and Shannon index were as follows: in the wild boar, Ho = 0.763 ± 0.026, I = 1.717 ± 0.091; in the Duroc breed, Ho = 0.569 ± 0.068, I = 1.191 ± 0.157; in the Landrace, Ho = 0.618 ± 0.062, I = 1.201 ± 0.147; in the Large White, Ho = 0.680 ± 0.029, I = 1.362 ± 0.074; and in the Yorkshire, Ho = 0.642 ± 0.065, I = 1.287 ± 0.156. The results of checking genotypic Hardy–Weinberg equilibrium based on the G-test of maximum likelihood demonstrated that the overwhelming majority of loci in the wild boar population were in the state of said equilibrium. By contrast, in pig breed populations, some loci demonstrated a significant deviation from the indicated equilibrium. In addition, the Yorkshire, Large White, and Landrace populations had loci, for which the hypothesis of neutrality was reliably rejected based on the results of the Ewens–Watterson test. The revealed private alleles, characteristic of the wild boar and breeds, can later be used to identify them. The ordination of the centroids of different herds in the space of the first two principal coordinates based on the matrix of pairwise estimates of Nei’s genetic distances showed that the most distant populations are the Duroc and Boar breeds, and the most genetically close are the Yorkshire and Landrace breeds. The closest to the wild boar population was the Large White breed. The assessment of the effective size, carried out using the method based on the linkage disequilibrium and the molecular coancestry method, showed that in all studied groups, including the wild boar population, the effective size was less than 100 individuals. The low effective size of the wild boar population (Ne = 21.8, Neb = 4.0) is probably caused by the death and shooting of animals due to Pestis africana suum.

Priorización semi-cuantitativa basada en evidencia de enfermedades bacterianas zoonóticas presentes en la interfaz cerdos silvestres (Sus scrofa), cerdos domésticos y humanos en Colombia

El objetivo del estudio fue priorizar las enfermedades bacterianas zoonóticas presentes en la interfaz cerdos silvestres (Sus scrofa), cerdos domésticos y humanos en Colombia. Se realizó una revisión de sistemas de indexación y resúmenes (SIR) y de datos del Sistema Mundial de Información Zoosanitaria (WAHID/WAHIS). Se empleó el método de priorización semi-cuantitativo basado en la evidencia con 27 criterios en cinco categorías: epidemiología, prevención/control, economía/comercio, salud pública y sociedad, con valores entre 0 (sin riesgo) y 7 (riesgo severo). La salmonelosis (148 puntos) y la tuberculosis (148) presentaron potencial zoonótico alto, con evidencia de mayor puntuación en los criterios de epidemiología, salud pública y sociedad; seguido de yersiniosis (140) y brucelosis (127) con potencial zoonótico medio, y finalmente leptospirosis (119) y pasteurelosis (114) con potencial zoonótico bajo. Objetivo. Priorizar las enfermedades bacterianas zoonóticas presentes en la interfaz cerdos silvestres (Sus scrofa), cerdos domésticos y humanos en Colombia. Materiales y métodos. Se realizó una revisión de sistemas de indexación y resúmenes (SIRes), de datos del Sistema Mundial de Información Zoosanitaria (WAHID/WAHIS) y se empleó el método de priorización semi-cuantitativo basado en la evidencia con 27 criterios en 5 categorías: epidemiología, prevención/control, economía/comercio, salud pública y sociedad, valorados de 0-7. Resultados. Se encontró que salmonelosis (148) y tuberculosis (148) presentaron potencial zoonótico alto, con evidencia de mayor puntuación en los criterios de epidemiología, salud pública y sociedad; seguido de yersiniosis (140) y brucelosis (127) con potencial zoonótico medio; finalmente leptospirosis (119) y pasteurelosis (114) con potencial bajo. Conclusión. Aunque la salmonelosis y la tuberculosis fueron las enfermedades con mayor puntuación, las cuales tienen grandes efectos económicos en la industria porcícola; falta una mayor vigilancia epidemiológica especialmente en la región 3 y 4 de Colombia donde departamentos, como Bolívar, cumplen un papel importante en la transmisión de la enfermedad al ser áreas de mayor riesgo epidemiológico y presencia de interacción entre cerdos silvestres (Sus scrofa) y cerdos domésticos.