Kajian Keragaman Fenotipik dan Total Genom Mitokondria Kerbau Jambi Sebagai Dasar Strategi Pelestariannya

Abstract

Kerbau merupakan salah satu ternak lokal yang memiliki peran strategis dalam sektor peternakan, pertanian, berperan penting dalam berbagai seremonial keagamaan, dan upacara adat masyarakat Jambi. Namun, populasi kerbau Jambi telah mengalami penurunan drastis sebesar 34,13% dalam dua dekade terakhir, sehingga mengancam keberlanjutan populasi dan berpotensi menyebabkan hilangnya identitas genetik ternak lokal. Kondisi tersebut memerlukan upaya mitigasi melalui program pelestarian dan perbaikan mutu genetik yang berlandaskan kajian ilmiah. Sayangnya, hingga saat ini data dasar mengenai kajian struktur populasi, keragaman fenotipik, serta profil genetik pada tingkat DNA mitokondria masih sangat terbatas. Padahal, informasi tersebut merupakan landasan utama bagi pengembangan strategi pelestarian serta peningkatan mutu genetik yang terukur dan berkelanjutan. Penelitian ini bertujuan untuk mengkarakterisasi struktur populasi, keragaman fenotipik, dan genetik pada tingkat DNA kerbau Jambi. Penelitian terdiri atas empat tahap: (1) analisis struktur populasi, estimasi ukuran populasi efektif, dan laju inbreeding berbasis data sekunder; (2) karakterisasi fenotipik menggunakan analisis multivariat; (3) analisis keragaman genetik menggunakan penanda genetik ND1 (NADH dehydrogenase subunit 1) mitokondria; dan (4) karakterisasi dan analisis total genom mitokondria (whole mtDNA). Data populasi ternak kerbau sebanyak 3.149 ekor diperoleh dari Sistem Informasi Kesehatan Hewan Nasional Terintegrasi (iSIKHNAS) Kabupaten Batanghari tahun 2023. Data populasi mencakup kerbau anak (523 ekor: 230 jantan, 293 betina), muda (932 ekor: 346 jantan, 586 betina), dan kerbau dewasa (1.694 ekor: 164 jantan, 1.530 betina). Hasil analisis struktur populasi menunjukkan laju pertumbuhan alami sebesar 14,74%, tingkat penggantian bersih sebesar 279,51%, estimasi ukuran populasi efektif sebesar 592 ekor, dan laju inbreeding per generasi sebesar 0,08%. Analisis dinamika populasi menunjukkan penurunan populasi rata-rata 8,94% per tahun, dengan proyeksi mencapai 4.112 ekor pada tahun 2028, sehingga diperlukan intervensi manajemen pemeliharaan yang meliputi peningkatan ketersediaan jantan dewasa unggul, manajemen penyakit, dan pemberian pakan tambahan untuk meningkatkan produktivitas. Pengamatan fenotipik dilakukan pada 168 sampel kerbau dari lima kabupaten di provinsi Jambi yaitu kabupaten Tebo, Tanjung Jabung Barat, Batanghari, Kerinci, dan Merangin. Karakterisasi fenotipik meliputi sifat kualitatif, pengukuran morfometrik, dan indeks morfometrik. Data sifat kualitatif dianalisis menggunakan metode statistik deskriptif, selanjutnya data morfometrik dan indeks morfometrik dianalisis menggunakan General linear model (GLM), pearson korelasi, analisis komponen utama (PCA), canonical discriminant analysis (CDA), dan hierarchical cluster analysis (HCA). Hasil penelitian menunjukkan bahwa warna kulit abu gelap mendominasi semua populasi (44,7–62,5%), dengan variasi warna hitam yang tertinggi di Batanghari (42,1%). Bentuk tanduk menunjukkan keragaman tertinggi dengan lima tipe, didominasi bentuk Siki (26,9–62,5%). Analisis morfometrik mengungkapkan perbedaan signifikan (p < 0,05) antar populasi, dengan lebar pinggul (LPU) dan lingkar dada (LDD) menjadi penciri pembeda kerbau Jantan. Berdasarkan analisis indeks morfometrik menunjukkan populasi dikategorikan sebagai tipe pendek-kompak dengan kedalaman dada yang baik, mengindikasikan potensi sebagai penghasil daging. Analisis komponen utama (PCA) mengklasifikasikan pemisahaan yang lebih jelas pada jantan (PC1: 58,9%) dibanding betina (PC1: 54,9%), dengan lebar pinggul dan lingkar dada sebagai pembeda utama. Canonical discriminant analysis (CDA) mencapai akurasi 87,43% dalam membedakan kelima populasi. Hierarchical cluster analysis (HCA) mengidentifikasi tiga klaster berdasarkan tipe tubuh. Analisis keragaman genetik berdasarkan gen ND1 menggunakan sampel DNA yang dikoleksi dari 93 individu kerbau yang terdistribusi di lima sub populasi Jambi (Tebo: 23; Merangin: 9; Tanjung Jabung Barat: 9; Kerinci: 13; Batanghari: 21) dan tiga populasi pembanding (Banten: 4; Sumatera Utara: 6; Riau: 8). Gen ND1 diamplifikasi menggunakan metode polymerase chain reaction (PCR) dengan desain primer spesifik berbasis sekuen referensi GenBank Bubalus bubalis. Sekuensing dilakukan terhadap seluruh 93 sampel, dan analisis sekuen gen ND1 dilakukan menggunakan perangkat lunak MEGA 12, DnaSP6, dan NETWORK. Hasil analisis keragaman genetik menunjukan keragaman genetik yang sangat rendah pada kerbau Jambi dengan keragaman haplotipe (Hd) sebesar 0,128 dan keragaman nukleotida (p) sebesar 0,0002. Sebanyak 93,33% terdistribusi dalam satu klaster haplotipe nenek moyang dominan (Haplotipe 2) dan ditemukan haplotipe spesifik (H1, H3, dan H4) yang terdeteksi pada sub populasi Merangin dan Tanjung Jabung Barat. Nilai Fu's negatif (-2,146) menunjukan inbreeding tinggi dengan indikasi populasi telah mengalami peristiwa bottleneck yang parah. Analisis filogenetik menunjukkan bahwa kerbau Jambi berada dalam kelompok kerbau lumpur Asia Tenggara dan menjadi sub populasi berbeda dengan kerbau Asia maupun kerbau lokal lainnya. Analisis total genom mitokondria dilakukan menggunakan empat sampel selanjutnya disekuensing menggunakan platform Illumina NextSeq 2000 dengan pembacaan 300 siklus. Analisis sekuen total genom mtDNA dilakukan menggunakan software Geneious Prime. Sekuensing dan anotasi total genom mitokondria kerbau Jambi menghasilkan sekuen sepanjang 16.360 pasang basa (bp) yang mengkode 13 gen penyandi protein (PCGs), 22 gen transfer RNA (tRNA), 2 gen ribosomal RNA (rRNA) serta satu daerah D-loop. Analisis komposisi nukleotida mengungkapkan adanya bias AT yang dominan (69,81%), konsisten dengan karakter umum mitokondria kerbau, 12 PCGs dikodekan pada untai berat dan ND6 dikodekan pada untai ringan. Start kodon yang umum ditemukan adalah (ATG dan ATA) dan start kodon tidak lengkap TA-. Analisis filogenetik berdasarkan total genom mtDNA menunjukkan bahwa kerbau Jambi berbeda (sub populasi) dengan kerbau lain khususnya di Asia Tenggara sedangkan analisis filogenetik berdasarkan region d-loop mtDNA menunjukkan pemisahan yang jelas antara kerbau Batanghari, Tebo dan kerinci menjadi dua kelompok yaitu kerbau dataran tinggi dan dataran rendah mengindikasikan perbedaan yang spesifik antar populasi tersebut. Berdasarkan hasil penelitian dapat disimpulkan bahwa kerbau Jambi memiliki karakteristik struktural, fenotipik, dan genetik yang spesifik dan memerlukan program konservasi prioritas. Data populasi menunjukkan tren penurunan rata-rata 8,94% per tahun dengan proyeksi menurun menjadi 4.112 ekor pada tahun 2028 dengan ukuran populasi efektif rendah (592 ekor). Karakterisasi fenotipik menunjukkan tipe pendek-kompak berpotensi penghasil daging dengan lebar pinggul dan lingkar dada sebagai penciri pembeda berdasarkan analisis komponen utama (PCA). Analisis keragaman genetik gen ND1 menunjukkan keragaman sangat rendah (Hd: 0,128; p: 0,0002) dengan 93,33% konsentrasi pada haplotipe dominan, indikasi bottleneck parah dan inbreeding tinggi (Fu's: -2,146). Ditemukan haplotipe (H1, H3, H4) spesifik pada Merangin dan Tanjung Jabung Barat sebagai penyimpan keragaman genetik. Analisis filogenetik D-loop mtDNA mengungkapkan pemisahan jelas antara kerbau dataran tinggi (Kerinci) dan dataran rendah (Batanghari dan Tebo), menunjukkan adaptasi genetik spesifik terhadap lingkungan. Disimpulkan bahwa kerbau Jambi memiliki struktur genetik kompleks dengan dua kelompok ekologis berbeda, haplotipe ND1 langka pada Merangin dan Tanjung Jabung Barat, dan region D-loop sebagai penciri pembeda antar populasi. Kerbau Jambi layak dipertahankan sebagai sumber daya genetik ternak dengan program konservasi terukur yang mempertimbangkan diferensiasi ekologis dan penanda genetik (ND1 dan D-loop) untuk identifikasi dan pelestarian. Kata kunci: Genom mitokondria, keragaman fenotipik, keragaman ND1, strategi pelestarian, struktur populasi

Buffaloes are local livestock with a strategic role in the livestock and agricultural sectors, playing an important part in various religious ceremonies and traditional rituals of the Jambi community. However, the buffalo population in Jambi has declined drastically by 34.13% over the past two decades, threatening the sustainability of the population and potentially resulting in the loss of the local livestock genetic identity. These conditions require mitigation efforts through conservation programs and genetic quality improvement based on scientific research. Unfortunately, to date, baseline data on population structure studies, phenotypic diversity, and genetic profiles at the mitochondrial DNA level remain limited. Such information is fundamental for developing measurable and sustainable conservation strategies and improving genetic quality. This study aimed to characterize the population structure, phenotypic diversity, and genetic diversity at the DNA level of Jambi buffalo. The research consists of four stages: (1) analysis of population structure, estimation of effective population size, and inbreeding rate based on secondary data; (2) phenotypic characterization using multivariate analysis; (3) analysis of genetic diversity using the ND1 (NADH dehydrogenase subunit 1) mitochondrial genetic marker; and (4) characterization and analysis of the whole mitochondrial genome (mtDNA). The buffalo livestock population, totaling 3,149 heads, was obtained from the Integrated National Animal Health Information System (iSIKHNAS) of Batanghari Regency in 2023. The population data included calves (523 heads: 230 males and 293 females), young buffaloes (932 heads: 346 males and 586 females), and adult buffaloes (1,694 heads: 164 males and 1,530 females). Analysis of the population structure showed a natural growth rate of 14.74%, net replacement rate of 279.51%, estimated effective population size of 592 individuals, and inbreeding rate per generation of 0.08%. Population dynamics analysis indicated an average annual population decline of 8.94%, with a projected population of 4,112 heads by 2028. Therefore, management interventions are needed, including the increased availability of superior adult males, optimal disease management, and the provision of supplementary feed to enhance productivity. Phenotypic observations were performed on 168 buffalo samples from five regencies in Jambi Province: Tebo, Tanjung Jabung Barat, Batanghari, Kerinci, and Merangin. Phenotypic characterization included qualitative traits, morphometric measurements and morphometric indices. Qualitative trait data were analyzed using descriptive statistical methods, whereas morphometric and morphometric index data were analyzed using the General Linear Model (GLM), Pearson correlation, principal component analysis (PCA), canonical discriminant analysis (CDA), and hierarchical cluster analysis (HCA). The results showed that dark gray skin color dominated all populations (44.7–62.5%), with the highest black color variation found in Batanghari cattle (42.1%). Horn shape exhibited the highest diversity, with five types, dominated by the Siki shape (26.9–62.5%). Morphometric analysis revealed significant differences (p < 0.05) between populations, with hip width (LPU) and chest girth (LDD) distinguishing the male buffaloes. Based on morphometric index analysis, the populations were categorized as short-compact types with good chest depth, indicating their potential as meat producers. Principal component analysis (PCA) revealed a clearer separation among males (PC1:58.9%) than among females (PC1:54.9%), with hip width and chest girth as the main distinguishing factors. Canonical discriminant analysis (CDA) achieved an accuracy of 87.43% in differentiating the five populations studied. Hierarchical cluster analysis (HCA) identified three clusters based on the body type. Genetic diversity analysis based on the ND1 gene was conducted using DNA samples collected from 93 individual buffaloes distributed across five Jambi subpopulations (Tebo: 23; Merangin: 9; Tanjung Jabung Barat: 9; Kerinci: 13; Batanghari: 21) and three comparison populations (Banten: 4; North Sumatra: 6; Riau: 8). The ND1 gene was amplified using the polymerase chain reaction (PCR) method with a specific primer design based on the Bubalus bubalis reference sequence from the GenBank database. Sequencing was performed on all 93 samples, and ND1 gene sequence analysis was conducted using MEGA 12, DnaSP6, and NETWORK software programs. The results of the genetic diversity analysis showed very low genetic diversity in Jambi buffalo, with a haplotype diversity (Hd) of 0.128 and nucleotide diversity (p) of 0.0002. A total of 93.33% were distributed in a single dominant ancestral haplotype cluster (Haplotype 2), and rare haplotypes (H1, H3, and H4) were detected in the Merangin and Tanjung Jabung Barat subpopulations. The negative Fu’s value (-2.146) indicates high inbreeding, suggesting that the population has experienced a severe bottleneck. Phylogenetic analysis showed that the Jambi buffalo belong to the Southeast Asian swamp buffalo group and form a distinct subpopulation from other Asian buffaloes and local buffalo breeds. Total mitochondrial genome analysis was performed using two liver tissue samples, which were sequenced using the Illumina NextSeq 2000 platform with 300-cycle reads. Total mtDNA genome sequence analysis was conducted using the Geneious Prime software. Sequencing and annotation of the total mitochondrial genome of the Jambi buffalo produced a sequence of 16,360 base pairs (bp) encoding 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes, and one D-loop region. Nucleotide composition analysis revealed a predominant AT bias (69.81%), consistent with the general characteristics of buffalo mitochondria; 12 PCGs are encoded on the heavy strand, and ND6 is encoded on the light strand. The common start codons identified were (ATG and ATA) and an incomplete start codon, TA-. Phylogenetic analysis based on the total mtDNA genome showed that Jambi buffalo are distinct (a sub-population) from other buffalo, particularly in Southeast Asia, while phylogenetic analysis based on the mtDNA D-loop region revealed a clear separation between Batanghari, Tebo, and Kerinci buffalo into two groups highland and lowland buffalo indicating specific differences between these populations. Based on the research results, it can be concluded that Jambi buffalo possess specific structural, phenotypic, and genetic characteristics and require a priority conservation program. Population data show a declining trend at an average rate of 8.94% per year, with projections dropping to 4,112 individuals by 2028 and a low effective population size (592 individuals). Phenotypic characterization indicated a short-compact type with potential for meat production, with hip width and chest girth serving as distinguishing features based on principal component analysis (PCA). Analysis of genetic diversity in the ND1 gene showed very low diversity (Hd: 0.128; p: 0.0002), with 93.33% concentrated in a dominant haplotype, indicating a severe bottleneck and high inbreeding (Fu's: -2.146). Rare haplotypes (H1, H3, and H4) specific to Merangin and Tanjung Jabung Barat were found, serving as reservoirs of genetic diversity. Phylogenetic analysis of the mtDNA D-loop revealed a clear separation between highland buffaloes (Kerinci) and lowland buffaloes (Batanghari and Tebo), indicating specific genetic adaptation to their environment. In conclusion, the Jambi buffalo has a complex genetic structure with two distinct ecological groups, rare ND1 haplotypes in Merangin and Tanjung Jabung Barat, and the D-loop region serves as a distinguishing marker between populations. The Jambi buffalo is worthy of preservation as a genetic livestock resource through a measurable conservation program that considers ecological differentiation and genetic markers (ND1 and D-loop) for identification and preservation. Keywords: conservation strategy, mitochondrial genome, ND1 diversity, phenotypic diversity, population structure