PENAPISAN AKTIVITAS ANTI-QUORUM SENSING DAN PROBIOTIK DARI BAKTERI ASAM LAKTAT TERHADAP PATOGEN Aeromonas hydrophila

Abstract

Produksi daging ayam merupakan sektor penting dalam industri pangan global, khususnya di Asia. Sektor ini berkontribusi besar terhadap pemenuhan kebutuhan protein hewani dan perekonomian melalui distribusi pasar domestik maupun internasional. Namun, peningkatan produksi juga diikuti oleh tantangan kesehatan ternak, terutama wabah penyakit yang disebabkan oleh bakteri patogen dari genus Aeromonas, khususnya Aeromonas hydrophila. Infeksi bakteri ini dapat menyebabkan penyakit sistemik dengan tingkat mortalitas tinggi, sehingga menimbulkan kerugian ekonomi bagi peternak dan mengganggu stabilitas pasokan daging ayam di pasar. Ketergantungan pada antibiotik dalam sistem produksi intensif memicu resistensi antimikroba serta menimbulkan implikasi kesehatan dan lingkungan. Oleh karena itu, diperlukan pendekatan alternatif yang lebih berkelanjutan, termasuk penghambatan quorum sensing (QS), yaitu mekanisme komunikasi bakteri berbasis sinyal Acyl-homoserine lactones (AHL) yang mengatur virulensi A. hydrophila. Penelitian ini bertujuan mengeksplorasi potensi bakteri asam laktat (BAL) dalam menghambat quorum sensing (QS) pada A. hydrophila sebagai pendekatan pengendalian hayati yang inovatif, melalui karakterisasi profil genomik dan metabolomik isolat yang berpotensi tinggi serta evaluasi aktivitas anti-QS dan kapasitas probiotiknya. Pendekatan penelitian meliputi skrining aktivitas anti-QS dan antibiofilm secara in vitro, analisis ekspresi gen virulensi dan fenotipe yang diregulasi QS, pendekatan genomik dan metabolomik berbasis whole genome sequencing (WGS) dan GC-MS, analisis in silico melalui molecular docking, serta evaluasi karakterisasi probiotik dan aspek keamanan isolat. Isolat Pediococcus acidilactici AZMS2, Limosilactobacillus fermentum AZMFK2, dan Pediococcus acidilactici AZMDH7 terbukti menunjukkan aktivitas anti-QS yang kuat serta karakteristik probiotik yang menjanjikan. Ketiga isolat menghasilkan zona non-ungu 4.80–6.00 mm pada uji reporter Chromobacterium violaceum, mengindikasikan aktivitas penghambatan QS. Identifikasi molekuler menegaskan spesies masing-masing isolat, dan analisis Whole Genome Sequencing (WGS) menunjukkan ukuran genom berkisar 2,009–2,010 Mb. Meskipun tidak terdeteksi gen degradasi AHL khusus, analisis Biosynthetic Gene Cluster (BGC) mengidentifikasi keberadaan jalur biosintesis RiPPs dan poliketida yang berpotensi menghasilkan metabolit bioaktif. Metabolit BAL pada konsentrasi 2–10 mg/mL mampu mendegradasi AHL hingga 99 %, menghambat pembentukan biofilm A. hydrophila sebesar 77–79 %, serta menekan motilitas dan ekspresi gen virulensi utama (termasuk ahyR, ahyI, ahhI, aerA, dan lip). Evaluasi sifat probiotik menunjukkan bahwa seluruh isolat memiliki ketahanan terhadap kondisi saluran pencernaan, termasuk toleransi pH 3, garam empedu 0.3 %, suhu 25–42 °C, NaCl 8 %, dan glukosa 10 %. Ketiga isolat menunjukkan aktivitas proteolitik ringan, tidak memproduksi H2S, serta tidak memiliki aktivitas hemolitik, sehingga memenuhi kriteria keamanan dasar. Profil resistensi antibiotik menunjukkan resistensi terhadap streptomisin, sensitivitas terhadap ampisilin, dan tingkat resistensi menengah terhadap tetrasiklin. Analisis genom mengonfirmasi keberadaan gen fungsional terkait probiotik, seperti groEL, groES, bsh, opuA, clpL, dan clpP, serta ketiadaan gen virulensi dan jalur biosintesis amina biogenik. Selain itu, tidak ditemukan potensi transfer gen melalui sex pili maupun profag pada AZMS2 dan AZMDH7, meskipun sekuens profag pada AZMFK2 memerlukan analisis lebih lanjut. Penelitian ini menunjukkan potensi AZMS2, AZMFK2, dan AZMDH7 sebagai agen anti-quorum sensing sekaligus kandidat probiotik untuk mitigasi infeksi Aeromonas hydrophila. Eksplorasi mikrobiota sebagai sumber agen anti-QS dan probiotik memberikan peluang strategis untuk pengembangan pendekatan pengendalian penyakit yang lebih ramah lingkungan dan berkelanjutan dalam industri perunggasan.

Chicken meat production represents a critical sector in the global food industry, particularly in Asia, where it contributes substantially to animal protein supply and economic growth through domestic and international distribution networks. However, increased production intensity has been accompanied by major health challenges, especially outbreaks caused by pathogenic bacteria of the genus Aeromonas, notably Aeromonas hydrophila. This pathogen can induce severe systemic infections with high mortality, leading to economic losses for farmers and destabilizing poultry meat supply chains. Reliance on antibiotics in intensive production systems has further exacerbated antimicrobial resistance and raised health and environmental concerns. Therefore, more sustainable alternative approaches are required, including the inhibition of quorum sensing (QS), a bacterial communication mechanism mediated by Acyl-homoserine lactone (AHL) signaling molecules that regulate the virulence of A. hydrophila. This study aims to explore the potential of lactic acid bacteria (LAB) to inhibit quorum sensing (QS) in A. hydrophila as an innovative biocontrol strategy, through comprehensive genomic and metabolomic characterization of high-potential isolates, along with evaluation of their anti-QS activity and probiotic capacity. The research approach includes in vitro screening of anti-QS and antibiofilm activities; analysis of QS-regulated virulence gene expression and phenotypes; genome- and metabolome-based approaches using whole genome sequencing (WGS) and GC–MS; in silico molecular docking; and evaluation of probiotic characteristics and safety aspects of the isolates. The isolates Pediococcus acidilactici AZMS2, Limosilactobacillus fermentum AZMFK2, and Pediococcus acidilactici AZMDH7 demonstrated strong anti-QS activity and promising probiotic characteristics. All isolates produced non-purple zones of 4.80–6.00 mm in the Chromobacterium violaceum reporter assay, indicating QS inhibition. Molecular identification confirmed the species identity of each isolate, and Whole Genome Sequencing (WGS) revealed genome sizes of approximately 2.009–2.010 Mb. Although specific AHL-degrading genes were not detected, Biosynthetic Gene Cluster (BGC) analysis identified RiPP and polyketide biosynthetic pathways with potential to produce bioactive metabolites. LAB metabolites at 2–10 mg/mL degraded AHL up to 99 %, inhibited A. hydrophila biofilm formation by 77–79 %, and suppressed motility as well as the expression of key virulence genes (ahyR, ahyI, ahhI, aerA, and lip). Assessment of probiotic traits showed that all isolates were tolerant to gastrointestinal conditions, including pH 3, 0.3% bile salts, temperatures of 25–42 °C, 8 % NaCl, and 10 % glucose. The isolates exhibited mild proteolytic activity, did not produce H2S, and showed no hemolytic activity, fulfilling basic safety criteria. Antibiotic susceptibility profiling revealed resistance to streptomycin, sensitivity to ampicillin, and intermediate resistance to tetracycline. Genomic analysis confirmed the presence of probiotic-associated genes such as groEL, groES, bsh, opuA, clpL, and clpP, along with the absence of virulence genes and biogenic amine biosynthesis pathways. No evidence of gene transfer potential via sex pili or prophages was detected in AZMS2 and AZMDH7, although the prophage sequence in AZMFK2 warrants further attention. This study demonstrates the potential of AZMS2, AZMFK2, and AZMDH7 as anti–quorum sensing agents as well as probiotic candidates for the mitigation of Aeromonas hydrophila infections. The exploration of microbiota as a source of anti-QS agents and probiotics offers a strategic opportunity for the development of more environmentally friendly and sustainable disease control approaches in the poultry industry.