Displacement Loop Region Research Articles

Mitogenome based adaptations and phylogeny of Beetal goats in India

Beetal goats, the second largest Indian goat breed, are known for their adaptation to hot arid tropical climates. Beyond their phylogenetic significance, recent evidence suggests that mitogenome modifications play a crucial role in the adaptation of animals to environmental stress factors. This study aims to characterize the mitogenome of Beetal goats at the molecular level, elucidate their mitogenomic adaptations and determine the maternal phylogenetic status of Indian Beetal goats. Whole genome sequencing of pooled DNA samples from five unrelated Beetal goats was carried out to obtain mitogenome sequence. The Beetal goat mitogenome comprised of a coding region with 37 genes: 22 transfer RNAs (tRNAs), 13 protein-coding genes (PCGs), and two ribosomal RNAs (rRNAs), as well as a non-coding hypervariable displacement loop (D-loop) region. We identified nine SNPs inclusive of seven synonymous and two nonsynonymous ones located in PCGs that too in respiratory complex I subunit genes, five of which were novel. Two nonsynonymous SNPs (10,229 A > G in ND4 and 13,964 G > A in ND6) were homoplasmic ones with marked influence on their mRNA and protein structures. Synonymous SNPs influenced the minimum free energy (MFE) and topology of predicted mRNA secondary structures, thereby affecting mRNA stability and translation efficiency. Phylogenetic analysis of the D-loop region classified Beetal goats into haplogroup B. Mitogenome analysis of Beetal goats, in comparison with other goat populations revealed the greatest genetic similarity to Southeast Asian goats. The comprehensive analysis of mitochondrial DNA in Beetal goats reveals significant genetic adaptations crucial for their survival in hot, arid environments. This study also highlights the complex matrilineal origins of Beetal goats.

Mitochondrial displacement loop region single nucleotide polymorphisms and mitochondrial DNA copy number associated with risk of ankylosing spondylitis.

The pathogenesis of ankylosing spondylitis (AS) seems to be associated with genetics, the environment, heredity, and oxidative stress. Single nucleotide polymorphisms (SNPs) in the displacement loop (D-loop) region of mitochondrial DNA (mtDNA) and mtDNA copy number were investigated for their correlation with AS patients. This study included 83 AS patients and 100 healthy controls from the Second Hospital of Hebei Medical University. DNAs were extracted from blood samples for polymerase chain reaction analysis and quantitative real-time polymerase chain reaction analysis. Plasma reactive oxygen species (ROS) levels were measured by fluorescent probe technology. The distribution frequencies of the minor alleles of nucleotides 16304C (p = .037), 16311C (p = .027), and 152C (p = .034) were remarkably higher in AS patients than in healthy controls, which indicated that the16304C, 16311C, and 152C alleles were correlated with an increased risk of AS. Simultaneously, mtDNA copy number was statistically higher in patients with AS compared with controls (1.450 ± 0.876 versus 0.835 ± 0.626, p < .001). We also observed an increased ROS generation in AS patients compared with controls (27 066.169 ± 18 364.819 versus 14 758.330 ± 5854.946, p < .001) subsequently. In addition, the AS susceptible SNP 16311C is associated with high ROS levels (35 065.177 ± 26 999.934 vs. 25 005.818 ± 14 999.495, p = .043). Our study demonstrated that SNPs in the mtDNA D-loop could be AS risk biomarkers with the potential to promote oxidative stress levels; mtDNA copy number-induced mitochondrial dysfunction may also be involved in AS pathogenesis.

The Complete Mitochondrial Genome and Phylogenetic Analyses of To Chicken in Vietnam.

Indigenous chicken breeds have both cultural significance and economic value since they possess unique genetic characteristics that enable them to adapt to the local environment and contribute to biodiversity, food security, and sustainable agriculture in Vietnam. To (Tò in Vietnamese) chicken, a Vietnamese indigenous chicken breed, is popularly raised in Thai Binh province; however, little known is about the genetic diversity of this breed. In this study, we sequenced the complete mitochondrial genome of To chicken for a better understanding of the diversity and origin of the breed. The results of sequencing showed that the mitochondrial genome of To chicken spans a total length of 16,784 base pairs and comprises one non-coding control region (known as the displacement-loop (D-loop) region), two ribosomal RNA genes, 13 protein-coding genes, and 22 transfer RNA genes. The phylogenetic tree analyses and estimated genetic distances based on 31 complete mitochondrial genome sequences indicated that To chicken has a close genetic distance with the Laotian native chicken breed, Lv'erwu breed in China, and Nicobari black and Kadaknath breeds in India. The result of the current study might be important for conservation, breeding, and further genetic studies of To chicken.

Characterization of the Mitochondrial Genetic Landscape in Abdominal Aortic Aneurysm.

Background Abdominal aortic aneurysm (AAA) is a vascular disease with a mortality rate of >80% if ruptured. Mitochondrial dysfunction has been previously implicated in AAA pathogenesis. In this study, we aimed to characterize the mitochondrial genetic landscape in AAA. Methods and Results Whole mitochondrial genome sequencing and bioinformatics analysis were performed in comorbidity matched 48 cases without AAA and 48 cases with AAA, objectively diagnosed, and selected from a cohort of 65-year-old men recruited for a screening program. We identified differential mutational landscapes in men with and without AAA, with errors in mitochondrial DNA replication or repair as potential sources. Heteroplasmic insertions and overall heteroplasmy of structural rearrangements were significantly elevated in AAA cases. Three heteroplasmic variants were associated with risk factors of AAA: leukocyte concentration, plasma glucose, and cholesterol levels, respectively. Interestingly, mutations were more prevalent in regulatory part of the mitochondria, the displacement loop region, in AAA as compared with controls (P value <0.05), especially in the conserved and critical mitochondrial extended termination-associated sequenceregion. Moreover, we report a novel 24 bp mitochondrial DNA duplication present exclusively in cases with AAA (4%) and 75% of the unmatched AAA biopsies. Finally, the haplogroup cluster JTU was overrepresented in AAA and significantly associated with a positive family history of AAA (odds ratio, 2.9 [95% CI, 1.1-8.1]). Conclusions This is the first study investigating the mitochondrial genome in AAA, where important genetic alterations and haplogroups associated with AAA and clinical risk factors were identified. Our findings have the potential to fill in gaps in the missing genetic information on AAA.

The Non-Invasive Assessment of Circulating D-Loop and mt-ccf Levels Opens an Intriguing Spyhole into Novel Approaches for the Tricky Diagnosis of NASH.

Nonalcoholic fatty liver disease (NAFLD) is the commonest liver disease worldwide affecting both adults and children. Nowadays, no therapeutic strategies have been approved for NAFLD management, and hepatic biopsy remains the gold standard procedure for its diagnosis. NAFLD is a multifactorial disease whose pathogenesis is affected by environmental and genetic factors, and it covers a spectrum of conditions ranging from simple steatosis up to nonalcoholic steatohepatitis (NASH), fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). Several studies underlined the urgent need to develop an NAFLD risk prediction model based on genetics, biochemical indicators, and metabolic disorders. The loss of mitochondrial dynamics represents a typical feature of progressive NAFLD. The imbalance of mitochondrial lifecycle together with the impairment of mitochondrial biomass and function trigger oxidative stress, which in turn damages mitochondrial DNA (mtDNA). We recently demonstrated that the main genetic predictors of NAFLD led to mitochondrial dysfunction. Moreover, emerging evidence shows that variations in the displacement loop (D-loop) region impair mtDNA replication, and they have been associated with advanced NAFLD. Finally, lower levels of mitophagy foster the overload of damaged mitochondria, resulting in the release of cell-free circulating mitochondrial DNA (mt-ccf) that exacerbates liver injury. Thus, in this review we summarized what is known about D-loop region alterations and mt-ccf content during NAFLD to propose them as novel non-invasive biomarkers.

Mitochondrial DNA diversity of D-loop region in three native Turkish cattle breeds.

This study aimed to reveal the genetic variability of mitochondrial DNA (mtDNA) displacement-loop (D-loop) region in 62 animals belonging to three native Turkish cattle breeds, namely Anatolian Black (AB), East Anatolian Red (EAR) and Zavot (ZAV), and to conduct phylogenetic relationship analyses to obtain deeper information on their genetic origin and breeding history by comparison of 6 taurine and 11 indicine breeds, together with 66 polymorphic sites, a total of 31 haplotypes, of which 15, 10 and 6 were detected in AB, EAR and ZAV, respectively. Mean nucleotide and haplotype diversity were 0.01 and 0.891, respectively, whereas the genetic differentiation derived from Wright's index was 0.174 across the breeds. A significant level of total variation (17.42 %) was observed among breeds in molecular variance analysis. Six main haplogroups (T, T1, T2, T3, Q and I2) were detected in Anatolian cattle populations, where T3 was the most frequent among breeds (43.55 %), whereas I2, an indicine specific haplogroup, was observed only in ZAV. At the breed level, phylogenetic analyses supported by 198 sequences of 17 cattle breeds and 3 outgroup species retrieved from the GenBank clustered native Turkish cattle breeds with the taurine group rather than the indicine one, as expected. However, indicine admixture at low frequency (8.89 %) was detected in the ZAV breed for the first time due to more likely gene flow from indicine cattle breeds raised in neighbour countries, particularly Iran. This finding should be further investigated in all native Turkish and indicine cattle breeds from nearby countries to clarify gene flow and indicine admixture in Anatolian cattle.

Comparative mitochondrial genomics and phylogenetics for species of the snakehead genus Channa Scopoli, 1777 (Perciformes: Channidae)

The family Channidae, members of which are commonly known as snakehead fish, includes 53 Channa species and three Parachanna species. In this study, we characterized mitochondrial genomes (mitogenomes) of five Channa species (C. andrao, C. bleheri, C. ornatipinnis, C. pulchra, and C. stewartii) for the first time. We compared the mitogenomes with the mitogenomes of 11 other Channidae fish. The newly sequenced mitogenomes were 16,714 – 16,895 bp in length and contained 37 typical genes [13 protein-coding genes (PCGs), two ribosomal RNA genes (rRNAs) and 22 transfer RNA genes (tRNAs)]. Positive AT-skews and negative GC-skews were found in the mitogenomes of Channidae. Most PCGs started with the conventional start codon, ATN; however, the sequence of the stop codon was variable. There was no essential difference in relative synonymous codon usage (RSCU) among the Channidae mitogenomes. The fastest-evolving gene atp8 and slowest-evolving gene cox1 were identified using Ka/Ks and pairwise relative genetic distance (p-distance). The displacement loop (D-loop) regions showed great variability, which affected the size of Channa mitogenomes. One origin of replication on the light strand (OL) region was found between trnN and trnC in the mitogenomes of Channidae. Phylogenetic analysis revealed three new sister pairs (C. andrao + C. bleheri, C. ornatipinnis + C. pulchra, and C. stewartii + C. gachua). Phylogenetic relationships established between the five Channa species based on mitogenomes were also supported by their morphological characteristics and geographical distribution. The novel information we obtained about these mitogenomes will contribute to elucidating the complex relationships among Channa species.

Phylogenetic Analysis of Philippine Native Pigs (Sus scrofa L.) from Batanes, Quezon, and Marinduque Based on Mitochondrial DNA D-loop Markers

By analyzing the mitochondrial DNA (mtDNA) displacement loop (D-loop) region, we aimed to assess the genetic diversity of Philippine native pigs from Batanes, Quezon, and Marinduque and to determine the phylogenetic relationship between these subpopulations and other Asian and European pig breeds. Ear tissue samples were collected from the native pigs in Batanes (n = 17), Quezon (n = 37), and Marinduque (n = 40). Genomic DNA was extracted, and the D-loop sequences were amplified by PCR. The D-loop sequences of 94 Philippine samples were initially analyzed using the Kimura 2-parameter model and neighbor-joining method in MEGA7, identifying a total of 17 haplotypes designated as H1–H17. Notably, only eight haplotypes were specific to individuals coming from a particular province, whereas the remaining haplotypes were detected in samples from different provinces. Additional qualitative trait analysis that the majority of Philippine native pigs had black coat color, a plain coat pattern, short cylindrical snouts, straight head profiles, erect and forward-oriented ears, curly tails, and straight backlines. Phylogenetic analysis of the Philippine samples and 16 exotic pig breeds revealed that Philippine native pigs with H1 and H2 haplotypes formed a distinct clade with Taiwanese Type I Lanyu pig, whereas those with H3–H17 haplotypes clustered with other Asian and European pigs. Therefore, our findings suggest a close and unique relationship between Taiwanese Type I Lanyu and Philippine native pigs. The presence of several haplotypes also implies high genetic variation among Philippine native pigs. Specifically, the native pigs from Quezon and Marinduque are the most closely related, whereas those from Quezon and Batanes are the least closely related.

Increase in Mitochondrial D-Loop Region Methylation Levels in Mild Cognitive Impairment Individuals

Methylation levels of the mitochondrial displacement loop (D-loop) region have been reported to be altered in the brain and blood of Alzheimer’s disease (AD) patients. Moreover, a dynamic D-loop methylation pattern was observed in the brain of transgenic AD mice along with disease progression. However, investigations on the blood cells of AD patients in the prodromal phases of the disease have not been performed so far. The aim of this study was to analyze D-loop methylation levels by means of the MS-HRM technique in the peripheral blood cells of 14 mild cognitive impairment (MCI) patients, 18 early stage AD patients, 70 advanced stage AD patients, and 105 healthy control subjects. We found higher D-loop methylation levels in MCI patients than in control subjects and AD patients. Moreover, higher D-loop methylation levels were observed in control subjects than in AD patients in advanced stages of the disease, but not in those at early stages. The present pilot study shows that peripheral D-loop methylation levels differ in patients at different stages of AD pathology, suggesting that further studies deserve to be performed in order to validate the usefulness of D-loop methylation analysis as a peripheral biomarker for the early detection of AD.

Complete mitochondrial genome of Takydromus kuehnei (Squamata: Takydromus) and its phylogenetic analysis

We sequenced and annotated the complete mitochondrial genome (mitogenome) of Takydromus kuehnei Van Denburgh, 1909 (Squamata: Takydromus). This mitogenome was 17,224 bp long and encoded 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, one non-coding regions of an L-strand replication origin and a displacement loop region. The overall nucleotide composition was 32.8% of A, 13.8% of G, 24.8% of T, and 30.5% of C. Phylogenetic analysis using maximum likelihood method validated the taxonomic status of T. kuehnei, exhibiting the close relationship with the species from the genus Takydromus.

The SNPs of mitochondrial DNA displacement loop region and mitochondrial DNA copy number associated with risk of polymyositis and dermatomyositis

Oxidative damage-induced mitochondrial dysfunction may activate muscle catabolism and autophagy pathways to initiate muscle weakening in idiopathic inflammatory myopathies (IIMs). In this study, Single nucleotide polymorphisms (SNPs) in the mitochondrial displacement loop (D-loop) and mitochondrial DNA (mtDNA) copy number were assessed and their association with the risk of polymyositis and dermatomyositis (PM/DM) was evaluated. Excessive D-loop SNPs (8.779 ± 1.912 vs. 7.972 ± 1.903, p = 0.004) correlated positively with mtDNA copy number (0.602 ± 0.457 vs. 0.300 ± 0.118, p < 0.001). Compared with that of the controls, the mtDNA of PM/DM patients showed D-loop SNP accumulation. In addition, the distribution frequencies of 16304C (p = 0.047) and 16519C (p = 0.043) were significantly higher in the patients with PM/DM. Subsequent analysis showed that reactive oxygen species (ROS) generation was increased in PM/DM patients compared with that in the controls (18,477.756 ± 13,574.916 vs. 14,484.191 ± 5703.097, p = 0.012). Further analysis showed that the PM/DM risk-related allele 16304C was significantly associated with lower IL-4 levels (p = 0.021), while 16519C had a trend to be associated with higher IL-2 expression (p = 0.064). The allele 16519C was associated with a positive antinuclear antibody (ANA) status in PM/DM patients (p = 0.011). Our findings suggest that mitochondrial D-loop SNPs could be potential biomarkers for PM/DM risk and these SNPs associated with cytokine expression may be involved in the development of PM/DM. Further, mtDNA copy number-mediated mitochondrial dysfunction may precede the onset of PM/DM.

Characterization of the complete mitochondrial genome and phylogenetic analysis of Opsarius pulchellus (Cypriniformes, Danionidae, Chedrinae)

This study aimed to sequence and annotate the complete mitochondrial DNA genome sequence of Opsarius pulchellus. Complete mitochondrial genome is a circular molecule of 16,552 bp in length, including 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a displacement loop region. Proportion of nucleotides in mitochondrial genome is 26.8% T, 26.9% C, 27.6% A, and 18.8% G, with an AT bias of 54.4%. Maximum-likelihood phylogenetic tree was reconstructed using the connected protein sequences of 13 PCGs of O. pulchellus and other 18 fishes. Result of phylogenetic analysis supports a close relationship between O. pulchellus and Barilius bendelisis (Hamilton, 1807). Fundamental genetic data of O. pulchellus would be beneficial for further studies on genetic diversity and phylogeny.

Mitochondrial Displacement Loop Region SNPs Modify Sjögren's Syndrome Development by Regulating Cytokines Expression in Female Patients.

Mitochondrial dysfunction could induce innate immune response with cytokines releasing to initiate Sjögren’s syndrome (SS) onset. Single nucleotide polymorphisms (SNPs) in the mitochondrial displacement loop (D-loop) and mitochondrial DNA (mtDNA) copy number of female SS patients were evaluated for their association with SS in female patients. At the nucleotide site of 152, 16304, 16311 and 16362 in the D-loop, the frequencies for the minor alleles of 152C (p = 0.040, odds ratio [OR] = 0.504), 16304C (p = 0.045, OR = 0.406), 16311C (p = 0.045, OR = 0.406) and 16362C (p = 0.028, OR = 0.519) were significantly higher in the SS patients than those in the female controls, which indicated that 152,C, 16304C, 16311C, and 16362C allele in the D-loop of mtDNA were associated with the risk of SS. Meanwhile, the excessive SNPs were accumulated in D-loop region of SS patients (8.955 ± 2.028 versus 7.898 ± 1.987, p < 0.001, 95% confidence interval [CI]: 0.477–1.637) and mtDNA copy number increased in SS patients (1.509 ± 0.836 versus 1.221 ± 0.506, p = 0.006, 95% CI: 0.086–0.490) by a case-control analysis. The subsequent analysis showed that SS risk-related allele 16311C was associated with higher IL-2 levels (p = 0.010) at significantly statistical level whereas 152C associated with lower IL-10 levels (p = 0.058) at a borderline statistical levels. Our findings suggest that mitochondrial D-loop SNPs are predictors for SS risk, it might modify the SS development by regulating cytokine expression.

The Mitochondrial Epigenome: An Unexplored Avenue to Explain Unexplained Myopathies?

Mutations in either mitochondrial DNA (mtDNA) or nuclear genes that encode mitochondrial proteins may lead to dysfunctional mitochondria, giving rise to mitochondrial diseases. Some mitochondrial myopathies, however, present without a known underlying cause. Interestingly, methylation of mtDNA has been associated with various clinical pathologies. The present study set out to assess whether mtDNA methylation could explain impaired mitochondrial function in patients diagnosed with myopathy without known underlying genetic mutations. Enhanced mtDNA methylation was indicated by pyrosequencing for muscle biopsies of 14 myopathy patients compared to four healthy controls, at selected cytosines in the Cytochrome B (CYTB) gene, but not within the displacement loop (D-loop) region. The mtDNA methylation patterns of the four healthy muscle biopsies were highly consistent and showed intriguing tissue-specific differences at particular cytosines with control skin fibroblasts cultured in vitro. Within individual myopathy patients, the overall mtDNA methylation pattern correlated well between muscle and skin fibroblasts. Despite this correlation, a pilot analysis of four myopathy and five healthy fibroblast samples did not reveal a disease-associated difference in mtDNA methylation. We did, however, detect increased expression of solute carrier family 25A26 (SLC25A26), encoding the importer of S-adenosylmethionine, together with enhanced mtDNA copy numbers in myopathy fibroblasts compared to healthy controls. To confirm that pyrosequencing indeed reflected DNA methylation and not bisulfite accessibility, mass spectrometry was employed. Although no myopathy-related differences in total amount of methylated cytosines were detected at this stage, a significant contribution of contaminating nuclear DNA (nDNA) was revealed, and steps to improve enrichment for mtDNA are reported. In conclusion, in this explorative study we show that analyzing the mitochondrial genome beyond its sequence opens novel avenues to identify potential molecular biomarkers assisting in the diagnosis of unexplained myopathies.

Complete mitochondrial genome and phylogenetic analysis of Danio roseus (Teleostei, Cypriniformes, Danionidae)

Danio roseus, collected from Gadu River in Yingjiang area, Yunnan Province, China, had been sequenced on Illumina HiSeq platform with 16523 bp in length, which included 37 genes encoding 13 proteins-coding genes (PCGs), 22 tRNAs, 2 rRNAs and a displacement loop region. The proportion of nucleotides in mitochondrial genome was T (28.7%), C (23.2%), A (32.3%), G (15.8%) with an AT bias of 61%. Maximum-Likelihood phylogenetic tree was reconstructed using concatenated mitochondrial protein-coding genes of D. roseus and other 12 fishes. The result of phylogenetic analysis supported the consanguineous relationship among D. roseus, D. rerio, D. nigrofasciatus and D. dangila.

Application of molecular methods in forensic analyses of wild animals

In European countries, hunting of wildlife is regulated by various directives. However, illegal hunting activities are becoming more common. To deal with wildlife crime cases, it becomes necessary to select the best molecular markers for the analysis of animal sex, species, and relationships identification. This study investigated potential markers within chromosomal and mitochondrial DNA (mtDNA) for animal forensics. By applying markers targeting nuclear DNA and mitochondrial DNA (the cytochrome b gene, hypervariable displacement loop (D-Loop) region, SRY and AMEL genes), the suspected illegal hunting cases were solved by the identification of wild species and sex. Currently, molecular forensic techniques are necessary before conclusions of its validation in forensic investigations.

Transcutaneous electrical stimulation therapy and genetic analysis in Dercum's disease: A pilot study.

Dercum's disease (DD), or adiposis dolorosa, is a rare condition of unknown etiology characterized by growth of painful subcutaneous adipose tissue. No specific treatment exists. Pain is often invalidating and resistant to analgesic drugs. We tested the efficacy of Frequency Rhythmic Electrical Modulation System (FREMS) therapy on pain relief. Subcutaneous biopsies were performed for genetic analysis.Nine DD patients were enrolled. Five cycles of FREMS at 3-month intervals during 1 year were administered. Visual analogue scale (VAS), Bartel Index Questionnaire and Short Form 36 questionnaire were used to measure pain and general health status at baseline, 6 and 12 months. Dual-energy X-ray absorptiometry (DEXA) quantified fat mass. Next-Generation Sequencing (NGS) was performed on adipose tissue biopsies and peripheral blood sample to search for somatic variants and specific protein pathway mutation.Seven patients were included in the final analysis. FREMS induced a reduction in VAS score (from 92 to 52.5, P = .0597) and a significant improvement in SF-36 domains (Physical functioning, Role limitation due to physical health, Body pain, Vitality, Social functioning, P < .05). No modification in anthropometrics and DEXA values was observed. The analysis of the mitochondrial Displacement loop (D-loop) region confirmed the clonality of all lipomatous lesions. The presence of the mitochondrially encoded tRNA-Lysine (MT-TK) m.8344A>G variant, occasionally identified in patients with multiple symmetric lipomatosis, was excluded in all subjects. On the other hand, we observed variants in genes belonging to signaling pathways involved in cell cycle and proliferation (Phosphoinositide 3-kinase/AKT/mTOR, MAPK/ERK, and Hippo).FREMS can be a useful tool to alleviate pain and improve overall quality of life in patients with DD. Genetic analysis highlighted the molecular heterogeneity of lipomas.

Mitochondrial phylogenetic and diversity analysis in Azi-Kheli buffalo.

The present study was conducted for the first time in Pakistan to investigate Cytochrome C Oxidase Subunit 1 (CO1) gene and full-length Displacement Loop (D-loop) region of mitochondrial DNA in Azi-Kheli buffalo breed native to northern hilly areas of Khyber Pakhtunkhwa Province of Pakistan. The present study was designed to investigate phylogeny and diversity in Azi-Kheli buffalo, through two mitochondrial DNA regions, i.e., Cytochrome C Oxidase Subunit-I (CO1) and Displacement Loop (D-loop) region. Thirty (30) blood samples were taken from Azi-Kheli pure breed animals from original breeding tract, i.e., Khwazakhela, Swat. Polymerase chain reactions using gene-specific primers were carried out for amplifying 709-bp region of CO1 gene and 1159-bp region of D-Loop for identification, phylogeny, and diversity in Azi-Kheli buffalo, respectively. The sequences of CO1 gene revealed four (04) haplotypes, whereas D-loop sequences revealed five (05) haplotypes. Mean interspecific diversity with related species was 2.56%, and mean intraspecific diversity within Azi-Kheli buffalo was 0.25%, estimated via Kimura-2 parameter. Phylogenetic tree (maximum likelihood) revealed clustering of Azi-Kheli haplotypes with river buffalo and is distinct from swamp buffalo and other related species of genus Bubalus. Mean haplotype and nucleotide diversity of D-loop were Hd = 0.9601 ± SD = 0.096 and π = 0.01208 ± SD = 0.00182, respectively. Phylogenetic tree (neighbor-joining) revealed two main clades, i.e., river buffalo and swamp buffalo clade. The haplotypes of Azi-Kheli clustered with haplotypes of different river buffalo breeds at different positions. The current study suggests that Azi-Kheli has common origin with other river buffalo breeds; hence, it is river buffalo which harbors high genetic diversity.

Spectrum of mitochondrial genomic variation in parathyroid neoplasms.

Mitochondrial DNA (mtDNA) variations have been implicated in various cancer types. Several attempts have been made in benign parathyroid adenoma (PA); however, mtDNA variants and copy number alterations have not been investigated in parathyroid carcinoma (PC). The present study aimed to explore the variations in the mitochondrial genome in parathyroid neoplasms. In this study, ultradeep targeted sequencing of mtDNA was performed in 12 cases with PC and 25 cases with PA. Germline and somatic variants in the mitochondrial genome were analysed according to clinical features. The mtDNA copy number relative to nuclear DNA was measured. A total of 821 germline variants and 23 somatic mutations were identified. All 160 germline nonsynonymous variants in the coding sequence (CDS) were predicted to be likely benign, and germline variants frequently occurred (26.3%) in the displacement loop region. Tumour somatic mutation in a CDS with heteroplasmic factor (HF) >0.8 showed a higher mtDNA copy number (p = 0.039). Using Spearman's rank test, tumour mtDNA copy number revealed a positive correlation with serum levels of intact parathyroid hormone and calcium. Our results demonstrate that null recurrent mtDNA mutational hotspots were PC specific. An increased mtDNA copy number in parathyroid neoplasms was associated with somatic CDS mutations with a high HF and major clinical features. Larger cohort investigations should be performed in future analyses.

Placental mitochondrial DNA mutational load and perinatal outcomes: Findings from a multi-ethnic pregnancy cohort

Mitochondria fuel placental activity, with mitochondrial dysfunction implicated in several perinatal complications. We investigated placental mtDNA mutational load using NextGen sequencing in relation to birthweight and gestational length among 358 mother-newborn pairs. We found that higher heteroplasmy, especially in the hypervariable displacement loop region, was associated with shorter gestational length. Results were similar among male and female pregnancies, but stronger in magnitude among females. With regard to growth, we observed that higher mutational load was associated with lower birthweight-for-gestational age (BWGA) among females, but higher BWGA among males. These findings support potential sex-differential fetal biological strategies for coping with increased heteroplasmies.