Keragaman Universal Stress Protein pada Enterobacteriales dan Penurunan Ekspresi Gen usp pada Pectobacterium brasiliense dengan Perlakuan Madu Manuka

CHOTIMAH, Dr. Tri Joko, S.P., M.Sc.; Alan Soffan, S.P., M.Sc., Ph.D

2023 | Tesis | MAGISTER FITOPATOLOGI

Universal stress protein (usp) mendukung kelangsungan hidup suatu organisme dengan meningkatkan ekspresinya dalam kondisi cekaman, meliputi kekurangan nutrisi, heat shock, dan antibiotik. Namun, ekspresi gen uspA pada meticilin-resistant Staphylococcus aureus (MRSA) dilaporkan menurun setelah terpapar madu Manuka. Madu manuka juga diketahui menghambat pertumbuhan Pectobacterium brasiliense, meskipun pengaruhnya terhadap ekspresi gen usp dari patogen busuk lunak ini belum diteliti. Penelitian ini bertujuan untuk mengetahui kekerabatan Usp P. brasiliense dan ortolognya dari Enterobacteriales dengan menggunakan analisis filogenetik serta Multiple Sequence Alignment (MSA) dan pengaruh madu Manuka terhadap ekspresi gen usp diamati menggunakan quantitative reverse transcription polymerase chain reaction (qRT-PCR) dan uji patogenisitas P. brasiliense pada sawi putih. Hasil penelitian menunjukkan bahwa UspA, UspB, dan UspE dari P. brasiliense memiliki kekerabatan paling dekat dengan P. carotovorum, sedangkan UspG memiliki kekerabatan paling dekat dengan P. polaris. Ekspresi keempat usp tersebut menurun masing-masing sebesar 0,05 kali, 0,16 kali, 0,1 kali, dan 0,62 kali dalam perlakuan madu Manuka 5% (w/v). P. brasiliense dengan perlakuan madu Manuka menimbulkan lesi yang lebih kecil dibandingkan dengan kontrol saat diuji pada sawi putih. Penurunan virulensi P. brasiliense ini mungkin berhubungan dengan penurunan ekspresi gen usp yang dipicu oleh madu Manuka.

Universal stress protein (usp) supports the survival of an organism by increasing its expression under stress conditions. These stress conditions include nutritional deficiencies, heat shock, and antibiotics. However. uspA gene expression in meticilin-resistant Staphylococcus aureus was reported to decrease after Manuka honey exposure. Manuka honey is also known to inhibit the growth of Pectobacterium brasiliense, although its effect on the usp gene of this soft rot pathogen has never been studied. This study aimed to determine the diversity of the Usp of P. brasiliense and its ortholog from Enterobacteriales using phylogenetic analysis and multiple sequence alignment (MSA) and the effect of Manuka honey on usp gene expression was investigated using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and pathogenicity assay of P. brasiliense on Chinese cabbage. The results revealed that the UspA, UspB, and UspE of P. brasiliense had the closest similarity to those of P. carotovorum, while the UspG had the closest similarity to that of P. polaris. Their expressions were down-regulated by 0.05 fold, 0,16 fold, 0,1 fold, and 0,62 fold, respectively, in 5% (w/v) Manuka honey treatment. P. brasiliense treated with Manuka honey caused smaller lesion symptoms compared to that control treatment when tested on Chinese cabbage. This reduced virulence of P. brasiliense might be related to reduced expression of the usp genes triggered by Manuka honey.

Kata Kunci : madu manuka, Pectobacterium brasiliense, qRT-PCR, busuk lunak, universal stress protein