First Report of <i>Pectobacterium brasiliense</i> Causing Soft Rot and Blackleg on Potato ( <i>Solanum tuberosum</i> ) in Montenegro

During June and July 2023, soft rot and blackleg outbreak was observed on potato (Solanum tuberosum L.) cultivars Carrera and Rudolph, covering approximately 0.8 ha in two localities near Podgorica, Montenegro, resulting in high yield losses. Water-soaked lesions appeared at the stem base and spread toward upper parts, followed by stem softening, blackleg symptoms and collapse of the entire plant. Incidence of stem infection exceeded 60% of all the plants. Underground, tubers of these plants were soft-rotten. The tuber decay was extended over time and favored by increased soil moisture. Weather conditions significantly contributed to disease development, with average daily temperatures ranging from 24°C to 31°C and relative humidity around 70%. Several isolations from the diseased surface-sterilized stem tissue, made on nutrient agar (NA) medium, repeatedly yielded in shiny, white-gray, round colonies, 2-4 mm in diameter. Pathogenic strains were selected by testing pectolytic activity on potato slices as a preliminary screening test. The three strains (KFB 1539, 1540 and 1558), out of seven isolated, were further characterized as Gram and oxidase negative, catalase positive, nonfluorescent and grew at 37°C. They were positive in acid production from L-arabinose, lactose, sucrose and melibiose and negative for erythritol, sorbitol and inositol utilization. The strains did not utilize arginine nor lysine, showing characteristics of Pectobacterium brasiliense. PCR assay, using primers BR1f/L1r (Duarte et al. 2004), specific for P. brasiliense, amplified a gene fragment of 322bp in all tested strains. Our strain KFB 1540, 16S rRNA gene sequence (1354 bp, GenBank PQ388327) showed 100% identity to P. brasiliense isolated from potato in Serbia (GenBank MT240621) and China (GenBank CP136134). The tested strain recA (GenBank PQ399780) and gapA (GenBank PQ399799) genes showed 99.43% and 99.80% identity with previously deposited sequences of P. brasiliense isolated in South Korea (CP113504) and Brazil (MW046308), respectively. Pathogenicity test was conducted on 2-weeks-old potato plants grown in a commercial potting mix in a greenhouse. Two plants per each of the three strains were inoculated by applying a mass of 24-h-old bacteria, grown on KB medium, with a sterile toothpick to three aseptically made one-centimeter long wounds on the stems. The inoculation site was covered with moist sterile cotton pad and wrapped in parafilm for 3 days to provide high humidity. The plants were maintained in the greenhouse at 28 ± 2°C until symptom development. Seven days after inoculation, inoculated stems developed dark, necrotic lesions, resembling the natural infection. Bacteria were re-isolated and their identity was confirmed by specific PCR assay (Duarte et al. 2004). Negative control plants remained symptomless. Based on all characteristics, the strains isolated from diseased potato stems were identified as P. brasiliense, the causal agent of potato soft rot and blackleg disease. This is the first report of P. brasiliense affecting potato in Montenegro. This pathogen, first described in Brazil, followed by reports of its occurrence worldwide, emerged as economically very important due to significant losses causing in potato production. It spreads over long distances by infected seed and may persist in the soil for few years. Therefore, certified seed and good agricultural practices are essential in disease prevention.