Genomic Medicine in Canine Periodontal Disease: A Systematic Review.

Genomic medicine has become a growing reality; however, it is still taking its first steps in veterinary medicine. Through this approach, it will be possible to trace the genetic profile of a given individual and thus know their susceptibility to certain diseases, namely periodontal disease. This condition is one of the most frequently diagnosed in companion animal clinics, especially in dogs. Due to the limited existing information and the lack of comprehensive studies, the objective of the present study was to systematically review the existing scientific literature regarding genomic medicine in canine periodontal disease and determine which genes have already been studied and their probable potential. This study followed the recommendations of the PRISMA 2020 methodology. Canine periodontal disease allied to genomic medicine were the subjects of this systematic review. Only six articles met all of the inclusion criteria, and these were analyzed in detail. These studies described genetic variations in the following genes: interleukin-6, interleukin-10, interleukin-1, lactotransferrin, toll-like receptor 9, and receptor activator of nuclear factor-kappa B. Only in two of them, namely interleukin-1 and toll-like receptor 9 genes, may the identified genetic variations explain the susceptibility that certain individuals have to the development of periodontal disease. It is necessary to expand the studies on the existing polymorphic variations in genes and their relationship with the development of periodontal disease. Only then will it be possible to fully understand the biological mechanisms that are involved in this disease and that determine the susceptibility to its development.

Estimating sequence diversity of prion protein gene (PRNP) in Portuguese populations of two cervid species: red deer and fallow deer.

Among the transmissible spongiform encephalopathies (TSEs), chronic wasting disease (CWD) in cervids is now a rising concern in wildlife within Europe, after the detection of the first case in Norway in 2016, in a wild reindeer and until June 2022 a total of 34 cases were described in Norway, Sweden and Finland. The definite diagnosis is post-mortem, performed in target areas of the brain and lymph nodes. Samples are first screened using a rapid test and, if positive, confirmed by immunohistochemistry and Western immunoblotting. The study of the genetics of the prion protein gene, PRNP, has been proved to be a valuable tool for determining the relative susceptibility to TSEs. In the present study, the exon 3 of PRNP gene of 143 samples from red deer (Cervus elaphus) and fallow deer (Dama dama) of Portugal was analysed. Three single nucleotide polymorphisms (SNPs) were found in red deer - codon A136A, codon T98A, codon Q226E - and no sequence variation was detected in fallow deer. The low genetic diversity found in our samples is compatible with previous studies in Europe. The comparison with results from North America suggests that the free-ranging deer from our study may present susceptibility to CWD, although lack of experimental data and the necessity of continuous survey are necessary to evaluate these populations.

Genomic Medicine in Periodontal Disease: Old Issue, New Insights.

Genetic variability is the main cause of phenotypic variation. Some variants may be associated with several diseases and can be used as risk biomarkers, identifying animals with higher susceptibility to develop the pathology. Genomic medicine uses this genetic information for risk calculation, clinical diagnosis and prognosis, allowing the implementation of more effective preventive strategies and/or personalized therapies. Periodontal disease (PD) is the inflammation of the periodontium induced mainly by bacterial plaque and is the leading cause of tooth loss. Microbial factors are responsible for the PD initiation; however, several studies support the genetic influence on the PD progression. The main purpose of the present publication is to highlight the main steps involved in the genomic medicine applied to veterinary patients, describing the flowchart from the characterization of the genetic variants to the identification of potential associations with specific clinical data. After investigating which genes might potentially be implicated in canine PD, the RANK gene, involved in the regulation of osteoclastogenesis, was selected to illustrate this approach. A case-control study was performed using DNA samples from a population of 90 dogs - 50 being healthy and 40 with PD. This analysis allowed for the discovery of four new intronic variations that were banked in GenBank (g.85A>G, g.151G>T, g.268A>G and g.492T>C). The results of this study are not intended to be applied exclusively to PD. On the contrary, this genetic information is intended to be used by other researchers as a foundation for the development of multiple applications in the veterinary clinical field.

A Case of XX Disorder of Sexual Development in a Female-Phenotype Roe Deer (Capreolus capreolus L.) Associated with Antlers Growth with Retained Velvet.

A 3-to-4-year-old roe deer (Capreolus capreolus L.) was admitted to the Veterinary Hospital. Although it showed well-developed antlers with retained velvet, an external female appearance and genitalia were evident. External biometrical measurements were taken for the antlers, and a computed tomography was performed. Molecular studies targeting the SRY gene were performed, and a PIS (polled intersex syndrome) mutation diagnosis was implemented. The gonads consisted of a right testicle paired with a left ovotestis. Histologically, the ovary-like structures in the ovotestis were functional, but the testis, as the testis-like structure in the ovotestis, did not show active spermatogenesis. No evidence of SRY gene was detected by PCR, suggesting an XX-chromosome constitution. Additionally, polled intersex syndrome (PIS) deletion was not detected in the case under study. The clinical and histopathological findings confirmed the DSD with the presence of a testicle and a contralateral ovotestis.

Scrapie at Abattoir: Monitoring, Control, and Differential Diagnosis of Wasting Conditions during Meat Inspection.

Wasting disease in small ruminants is frequently detected at slaughterhouses. The wasting disorder is manifested by the deterioration of the nutritional and physiological state of the animal indicated by thinness, emaciation, and cachexia. Evidence of emaciation and cachexia, alone, are pathological conditions leading to carcass condemnation during an inspection. Several diseases are associated with a wasting condition, including scrapie, pseudotuberculosis, tuberculosis, paratuberculosis, Maedi Visna, and tumor diseases. On the other hand, parasitic diseases, nutrition disorders, exposure or ingestion of toxins, metabolic conditions, inadequate nutrition due to poor teeth, or poor alimentary diet are conditions contributing to poor body condition. Classical and atypical scrapie is naturally occurring transmissible spongiform encephalopathies in small ruminants. The etiological agent for each one is prions. However, each of these scrapie types is epidemiologically, pathologically, and biochemically different. Though atypical scrapie occurs at low incidence, it is consistently prevalent in the small ruminant population. Hence, it is advisable to include differential diagnosis of this disease, from other possibilities, as a cause of wasting conditions detected during meat inspection at the abattoir. This manuscript is a review of the measures in force at the abattoir for scrapie control, focusing on the differential diagnosis of gross lesions related to wasting conditions detected in small ruminants during meat inspection.

Neuropathology of Animal Prion Diseases.

Transmissible Spongiform Encephalopathies (TSEs) or prion diseases are a fatal group of infectious, inherited and spontaneous neurodegenerative diseases affecting human and animals. They are caused by the conversion of cellular prion protein (PrPC) into a misfolded pathological isoform (PrPSc or prion- proteinaceous infectious particle) that self-propagates by conformational conversion of PrPC. Yet by an unknown mechanism, PrPC can fold into different PrPSc conformers that may result in different prion strains that display specific disease phenotype (incubation time, clinical signs and lesion profile). Although the pathways for neurodegeneration as well as the involvement of brain inflammation in these diseases are not well understood, the spongiform changes, neuronal loss, gliosis and accumulation of PrPSc are the characteristic neuropathological lesions. Scrapie affecting small ruminants was the first identified TSE and has been considered the archetype of prion diseases, though atypical and new animal prion diseases continue to emerge highlighting the importance to investigate the lesion profile in naturally affected animals. In this report, we review the neuropathology and the neuroinflammation of animal prion diseases in natural hosts from scrapie, going through the zoonotic bovine spongiform encephalopathy (BSE), the chronic wasting disease (CWD) to the newly identified camel prion disease (CPD).

Toll-like receptor 9 gene in Periodontal Disease - A promising biomarker.

Periodontal Disease is an infectious and inflammatory disorder triggered mainly by periodontopathogenic bacteria, however, as a multifactorial disease, several factors modulate its progression, namely, genetic factors. Toll-like receptors (TLR) recognize molecular patterns present in pathogens and trigger an immune response against them. Thus, sequences variants in TLR genes seem to have the potential to modify the predisposition to Periodontal Disease and its progression. Based on this fact, TLR9 gene were analysed in a case-control study. DNA was obtained from 90 dogs (50 control and 40 cases) and a fragment of TLR9 gene was amplified by PCR and sequenced. The variants were identified by comparison with the dog wild type sequences. Our results suggest that rs375556098 and rs201959275 polymorphisms in the TLR9 gene are good candidates to become biomarkers of the canine predisposition to Periodontal Disease. It's important to notice that these polymorphic sites exist in Human exactly in the same codon. Since the dog is the best animal model to replicate the pathophysiological mechanisms of human Periodontal Disease, these results can potentially be extrapolated to humans.

Evaluation of biomarker canine-prostate specific arginine esterase (CPSE) for the diagnosis of benign prostatic hyperplasia.

BACKGROUND: Benign prostatic hyperplasia (BPH) is the most common canine prostatic disorder. Although most or even all intact male dogs may develop BPH by 5-8 years of age, many show no clinical signs. Taking into account the non-specific character of clinical and ultrasonographic findings, a new diagnostic approach has recently been proposed based on the augmentation of blood canine prostate-specific arginine esterase (CPSE) in hyperplasic dogs. The aim of the present study was to verify CPSE levels in negative controls and hyperplasic dogs, considering cytological findings as the reference method and taking into account the fact that controls were middle-aged intact dogs (median of 5.0 years), contrarily to previous studies carried out with very young control dogs. RESULTS: Significant differences of median CPSE levels were found between controls and hyperplasic dogs (29.1 versus 160.7 ng/mL, respectively); and significant positive correlations were found between median CPSE levels and age or prostatic volume (r = 0.549 and 0.448, respectively; p < 0.001). Sensitivity, specificity, positive and negative likelihood ratios put into evidence the good performance of the test. The agreement between methods was found to be very high, notably between CPSE levels and cytological results (Cohen's kappa coefficients above 0.8). CONCLUSIONS: Considering the results all together, measurement of CPSE is confirmed as a useful and accurate method and should be considered as an alternative or complementary tool to conventional methods for the diagnosis of BPH in middle-aged dogs.

Extreme genetic structure in a social bird species despite high dispersal capacity.

Social barriers have been shown to reduce gene flow and contribute to genetic structure among populations in species with high cognitive capacity and complex societies, such as cetaceans, apes and humans. In birds, high dispersal capacity is thought to prevent population divergence unless major geographical or habitat barriers induce isolation patterns by dispersal, colonization or adaptation limitation. We report that Iberian populations of the red-billed chough, a social, gregarious corvid with high dispersal capacity, show a striking degree of genetic structure composed of at least 15 distinct genetic units. Monitoring of marked individuals over 30 years revealed that long-distance movements over hundreds of kilometres are common, yet recruitment into breeding populations is infrequent and highly philopatric. Genetic differentiation is weakly related to geographical distance, and habitat types used are overall qualitatively similar among regions and regularly shared by individuals of different populations, so that genetic structure is unlikely to be due solely to isolation by distance or isolation by adaptation. Moreover, most population nuclei showed relatively high levels of genetic diversity, suggesting a limited role for genetic drift in significantly differentiating populations. We propose that social mechanisms may underlie this unprecedented level of genetic structure in birds through a pattern of isolation by social barriers not yet described, which may have driven this remarkable population divergence in the absence of geographical and environmental barriers.

Next-generation sequencing and comparative analysis of Pyrrhocorax pyrrhocorax and Pyrrhocorax graculus (Passeriformes: Corvidae) mitochondrial genomes.

The complete mitochondrial genomes of Red-billed Chough (Pyrrhocorax pyrrhocorax) and Yellow-billed Chough (Pyrrhocorax graculus) were sequenced using the Ion Torrent PGM platform. These mitogenomes contain 16,889 bp (Red-billed Chough) and 16,905 bp (Yellow-billed Chough), including 13 protein-coding genes (PCGs), two ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and a control region (D-loop). The gene content, orientation, and structure are similar to a wide range of other vertebrate species and the nucleotide composition is very similar to other Passeriformes. All PCGs start with ATG, except for COX1 that starts with GTG, and four stop codons and one incomplete stop codon are used (TAA, TAG, AGG, AGA, and T-). The size of PCGs is the same in both mitogenomes, except for ND6 that has one codon less in the Yellow-billed Chough. All the tRNAs can fold into a typical cloverleaf secondary structure. These mitogenomic data can be of great value in complementing forthcoming approaches on molecular ecology, comparative and functional genomics.

A case-control study between interleukin-10 gene variants and periodontal disease in dogs.

Periodontal disease (PD) refers to a group of inflammatory diseases that affect the periodontium, the organ which surrounds and supports the teeth. PD is a highly prevalent disease with a multifactorial etiology and, in humans the individual susceptibility is known to be strongly determined by genetic factors. Several candidate genes have been studied, namely genes related with molecules involved in the inflammatory response. Interleukin-10 (IL-10) is a cytokine with important anti-inflammatory and immunomodulatory roles, and several studies indicate an association between IL10 polymorphisms and PD. In dogs, an important animal model in periodontology, PD is also a highly prevalent naturally occurring disease, and only now are emerging the first studies evaluating the genetic predisposition. In this case-control study, a population of 90 dogs (40 dogs with PD and 50 healthy dogs) was used to study the IL10 gene, and seven new genetic variations in this gene were identified. No statistically significant differences were detected in genotype and allele frequencies of these variations between the PD cases and control groups. Nevertheless, one of the variations (IL10/2_g.285G>A) leads to an amino acid change (glycine to arginine) in the putative signal peptide, being predicted a potential influence on IL-10 protein functionality. Further investigations are important to clarify the biological importance of these new findings. The knowledge of these genetic determinants can help to understand properly the complex causal pathways of PD, with important clinical implications.

The influence of ACE ID and ACTN3 R577X polymorphisms on lower-extremity function in older women in response to high-speed power training.

BACKGROUND: We studied the influence of the ACE I/D and ACTN3 R577X polymorphisms (single or combined) on lower-extremity function in older women in response to high-speed power training. METHODS: One hundred and thirty-nine healthy older Caucasian women participated in this study (age: 65.5 ± 8.2 years, body mass: 67.0 ± 10.0 kg and height: 1.57 ± 0.06 m). Walking speed (S10) performance and functional capacity assessed by the "get-up and go" (GUG) mobility test were measured at baseline (T1) and after a consecutive 12-week period of high-speed power training (40-75% of one repetition maximum in arm and leg extensor exercises; 3 sets 4-12 reps, and two power exercises for upper and lower extremity). Genomic DNA was extracted from blood samples, and genotyping analyses were performed by PCR methods. Genotype distributions between groups were compared by Chi-Square test and the gains in physical performance were analyzed by two-way, repeated-measures ANOVA. RESULTS: There were no significant differences between genotype groups in men or women for adjusted baseline phenotypes (P > 0.05). ACE I/D and ACTN3 polymorphisms showed a significant interaction genotype-training only in S10 (P = 0.012 and P = 0.044, respectively) and not in the GUG test (P = 0.311 and P = 0.477, respectively). Analyses of the combined effects between genotypes showed no other significant differences in all phenotypes (P < 0.05) at baseline. However, in response to high-speed power training, a significant interaction on walking speed (P = 0.048) was observed between the "power" (ACTN3 RR + RX & ACE DD) versus "non-power" muscularity-oriented genotypes (ACTN3 XX & ACE II + ID)]. CONCLUSIONS: Thus, ACE I/D and ACTN3 R577X polymorphisms are likely candidates in the modulation of exercise-related gait speed phenotype in older women but not a significant influence in mobility traits.

Percutaneous approach for sialolith removal in a donkey.

Salivary duct lithiasis is a condition characterized by the partial or total obstruction ofa salivary gland or its excretory duct due to the formation of sialoliths. A 9-year-old female donkey, belonging to the unique and endangered indigenous breed of donkey in Portugal, was diagnosed with a sialolith in the rostral portion of the right parotid duct based on clinical, oral, dental, and radiographic examination results. Surgical removal of the sialolith was done through a percutaneous approach.

Muscle tissue changes with aging.

Sarcopenia is characterized by a progressive generalized decrease of skeletal muscle mass, strength and function with aging. Recently, the genetic determination has been associated with muscle mass and muscle strength in elderly. These two phenotypes of risk are the most commonly recognized and studied for sarcopenia, with heritability ranging from 30 to 85% for muscle strength and 45-90% for muscle mass. It is well known that the development and maintenance of muscle mass in early adulthood reduces the risk of developing sarcopenia and leads to a healthy aging. For that reason it seems important to identify which genetic factors interact with aging and in particular with the musculoskeletal response to exercise in such individuals. This review is designed to summarize the most important and representative studies about the possible association between certain genetic polymorphisms and muscle phenotypes in older populations. Also we will focuses on nutrition and some concerns associated with aging, including the role that exercise can have on reducing the negative effects of this phenomenon. Some results are inconsistent between studies and more replication studies underlying sarcopenia are needed, with larger samples and with different life cycles, particularly in the type and level of physical activity throughout life. In future we believe that further progress in understanding the genetic etiology and the metabolic pathways will provide valuable information on important biological mechanisms underlying the muscle physiology. This will enable better recognition of individuals at higher risk and the ability to more adequately address this debilitating condition.

Sarcopenia é caracterizada por uma diminuição generalizada e progressiva da força, massa e função muscular com o envelhecimento. Recentemente, a determinação genética tem sido associada com a massa muscular e força muscular em idosos. Estes dois fenótipos de risco são os mais comumente reconhecidos e estudados em relação à sarcopenia, com hereditariedade variando de 30 a 85% para a força muscular e 45-90% para a massa muscular. É bem conhecido que o desenvolvimento e manutenção da massa muscular na idade adulta reduz o risco de desenvolver sarcopenia e conduz a um envelhecimento saudável. Por isso, é importante identificar quais os fatores genéticos que interagem com o envelhecimento e, em particular, com a resposta músculo-esquelética ao exercício. Esta revisão destina-se a resumir os estudos mais importantes e representativos sobre a possível associação entre determinados polimorfismos genéticos e fenótipos musculares nas populações mais velhas. Os aspetos nutricionais serão discutidos, incluindo o papel que o exercício pode ter sobre a redução dos efeitos negativos deste fenômeno. Alguns resultados são inconsistentes e desta forma é necessária uma maior replicação subjacente à sarcopenia, com amostras maiores e em diferentes ciclos da vida, especialmente no tipo e nível de atividade física. No futuro, acreditamos que mais progressos na compreensão da etiologia genética e as vias metabólicas vai fornecer informações valiosas sobre importantes mecanismos biológicos envolvidos na fisiologia muscular. Isto irá permitir um melhor reconhecimento dos indivíduos com maior risco e uma maior capacidade de enfrentar adequadamente essa condição debilitante.

Analysis of new Matrilin-1 gene variants in a case-control study related to dental malocclusions in Equus asinus.

Prognathism and brachygnathism are craniofacial deformities that severely affect the health of human and vertebrates, such as donkeys. The multifactorial etiology of this disease makes the genetic analysis a powerful tool for its understanding and prevention of spreading these deformities. This study aims to contribute to the characterization of the genetic basis of prognathism and brachygnathism in donkeys, using the Zamorano-Leonés donkey, an endangered Spanish breed, as a model. Matrilin-1 (MATN1) polymorphisms have been previously described as markers for mandibular prognathism in Korean and Japanese human populations. Genetic variations in MATN1 gene were sought, in order to verify its association in a case-control study, including 30 donkeys presenting brachygnathism, 30 donkeys presenting prognathism and 30 donkeys with normal occlusion phenotypes. One genetic variation (g503G > A) located in an intronic region of MATN1 gene was identified and characterized. Statistically significant differences were detected between the control group and prognathism cases, but no statistical significant results were found between the control group and the brachygnathism cases. These results support evidence for an important role of MATN1 on prognathism in the analyzed population with MATN1 genetic variation - 503G>A - having a protective effect. Further studies should be developed in order to understand the whole role of MATN1 and the mechanisms affected by its genetic variations.

High-resolution melting analysis for bird sexing: a successful approach to molecular sex identification using different biological samples.

High-resolution melting (HRM) analysis is a very attractive and flexible advanced post-PCR method with high sensitivity/specificity for simple, fast and cost-effective genotyping based on the detection of specific melting profiles of PCR products. Next generation real-time PCR systems, along with improved saturating DNA-binding dyes, enable the direct acquisition of HRM data after quantitative PCR. Melting behaviour is particularly influenced by the length, nucleotide sequence and GC content of the amplicons. This method is expanding rapidly in several research areas such as human genetics, reproductive biology, microbiology and ecology/conservation of wild populations. Here we have developed a successful HRM protocol for avian sex identification based on the amplification of sex-specific CHD1 fragments. The melting curve patterns allowed efficient sexual differentiation of 111 samples analysed (plucked feathers, muscle tissues, blood and oral cavity epithelial cells) of 14 bird species. In addition, we sequenced the amplified regions of the CHD1 gene and demonstrated the usefulness of this strategy for the genotype discrimination of various amplicons (CHD1Z and CHD1W), which have small size differences, ranging from 2 bp to 44 bp. The established methodology clearly revealed the advantages (e.g. closed-tube system, high sensitivity and rapidity) of a simple HRM assay for accurate sex differentiation of the species under study. The requirements, strengths and limitations of the method are addressed to provide a simple guide for its application in the field of molecular sexing of birds. The high sensitivity and resolution relative to previous real-time PCR methods makes HRM analysis an excellent approach for improving advanced molecular methods for bird sexing.

ACE I/D and ACTN3 R/X polymorphisms as potential factors in modulating exercise-related phenotypes in older women in response to a muscle power training stimuli.

Genetic variation of the human ACE I/D and ACTN3 R577X polymorphisms subsequent to 12 weeks of high-speed power training on maximal strength (1RM) of the arm and leg muscles, muscle power performance (counter-movement jump), and functional capacity (sit-to-stand test) was examined in older Caucasian women [n = 139; mean age 65.5 (8.2) years; 67.0 (10.0) kg and 1.57 (0.06) m]. Chelex 100 was used for DNA extraction, and genotype was determined by PCR-RFLP methods. Muscular strength, power, and functional testing were conducted at baseline (T1) and after 12 weeks (T2) of high-speed power training. At baseline, the ACE I/D and ACTN3 R/X polymorphisms were not associated with muscle function or muscularity phenotypes in older Caucasian women. After the 12-week high-speed training program, subjects significantly increased their muscular and functional capacity performance (p < 0.05). For both polymorphisms, significant genotype-training interaction (p < 0.05) was found in all muscular performance indices, except for 1RM leg extension in the ACE I/D (p = 0.187). Analyses of the combined effects between genotypes showed significant differences in all parameters (p < 0.05) in response to high-speed power training between the power (ACTN3 RR + RX & ACE DD) versus "non-power" muscularity-oriented genotypes (ACTN3 XX & ACE II + ID)]. Our data suggest that the ACE and ACTN3 genotypes (single or combined) exert a significant influence in the muscle phenotypes of older Caucasian women in response to high-speed power training. Thus, the ACE I/D and ACTN3 R/X polymorphisms are likely factors in modulating exercise-related phenotypes in older women, particularly in response to a resistance training stimuli.