Association of Acinetobacter baumannii with Soft Rot Disease of Carrot in India

Research Article

J Bacteriol Mycol. 2022; 9(1): 1194.

Association of Acinetobacter baumannii with Soft Rot Disease of Carrot in India

Chandrashekar BS¹, PrasannaKumar MK¹*, Buela Parivallal P¹, Pramesh D², Swathi SP¹, Sahana NB¹, Mahesh HB³ and Puneeth ME¹

1Department of Plant Pathology, University of Agricultural Sciences, Bangalore, India

2Rice Pathology Laboratory, All India Coordinated Rice Improvement Programme, Gangavathi, University of Agricultural Sciences, Raichur, India

3Department of Genetics and Plant Breeding, College of Agriculture, V. C. Farm, Mandya, India

*Corresponding author: MK Prasanna Kumar, Department of Plant Pathology, University of Agricultural Sciences, Bangalore, India

Received: December 24, 2021; Accepted: January 31, 2022; Published: February 07, 2022


Soft rot disease of carrots is an important limiting factor of carrot production. In this study, carrot roots showing typical soft rot symptoms were identified in the fields, and diseased and healthy root samples were collected for pathogen identification. The pathogen was isolated using an enriched bell pepper method. The bell pepper developed a water-soaked lesion around the pricking region when it was pricked after stabbing the diseased root whereas, no symptoms were produced when bell pepper was pricked after stabbing a healthy carrot root. From samples of the infected roots, circular, whitish, smooth, mucoid, round, convex, and medium-sized colonies were formed on the nutrient agar medium and were morphologically identified as Acinetobacter spp. Pure culture for four isolates was obtained, and one of the isolates (AB1) was further subjected to 16S rDNA sequencing. The BLAST analysis of the 16S rDNA confirmed the identity of AB1 as Acinetobacter baumannii. Pathogenicity test using whole-root assay and slice assay proved AB1 as pathogenic on carrot by producing water-soaked lesion, maceration, and rotting symptoms, whereas water inoculated roots remain healthy. The rotting symptoms on the artificially diseased carrot roots were similar to those caused by Pectobacterium caratovorum and Klebsiella variicola on the carrot. Based on the colony morphology, biochemical tests, and 16S rDNA sequence identity followed by pathogenicity assays, it is evident that A. baumannii causes soft rot disease in carrots. This report is essential for developing specific diagnostics and management against this newly emerging bacterial pathogen of carrot.

Keywords: Soft rot; Acinetobacter baumannii; 16S rDNA; Water-soaked lesion


Cross-kingdom pathogens, and their potential plant reservoirs, have important implications for the emergence of infectious human/ plant diseases. However, the directionality of host association (plant to human or human to plant) is difficult to determine in cross-kingdom pathogenicity [1]. Surprisingly, some of these plant pathogens cause disease in humans and are frequently isolated from human infections in the nosocomial environment. Despite this specialization in plants, species of Pantoea have also been discovered to be pathogenic to humans. Now classified as an opportunistic human pathogen, P. agglomerans has been reported to be associated with septicaemia in humans [2]. Interestingly, Burkholderia cepacia, which causes onion rot, can also cause life-threatening pulmonary infections in humans [3]. Several recent studies have reported that these human-pathogenic bacterial species are also capable of colonizing and causing disease in many plant hosts [4]. Notably, many of these studies have been conducted under laboratory conditions, providing evidence for the phytopathogenic potentials of these cross-kingdom bacterial pathogens; but, the incidence of plant disease caused by many of these human pathogens in the natural environment remains unknown. Similarly, Enterobacter cloacae have evolved to colonize the human host, and it has also been identified as the causal agent of grey kernel disease of macadamia plants. The onset of grey kernel disease affects not only the quality of the kernels produced by the tree but results in grey discolouration and a foul smell [5]. E. cloacae also causes bacterial soft rot disease in dragon fruit [6], and bacterial leaf rots in Odontioda orchids [7]. Bacteria that cause disease in both plants and animals may have genes required to infect both hosts. For example, Pseudomonas aeruginosa causes persistent lung infections in humans also can infect plants and other hosts [8].

Bacterial soft rots are a group of diseases that cause more crop loss worldwide than any other bacterial disease. Bacterial soft rots damage succulent plant parts such as fruits, tubers, stems, and bulbs of plants in nearly every plant family [9]. Soft rot bacteria degrade pectate molecules that bind plant cells together, causing plant structure to fall apart eventually. Soft rots commonly affect vegetables such as potato, carrot, tomato, cucurbits (e.g., cucumbers, melons, squash, pumpkins), and cruciferous crops (e.g., cabbage, cauliflower, Boy Choy) [10-13]. These diseases can occur on crops in the field and on harvested crops in storage. Rot can occur over a wide temperature range, with the worst decay between 21°C and 80°C, particularly when oxygen is limited [14].

Carrot (Daucus carota subsp. sativus) is an important vegetable in India. Soft rot is a serious disease of carrots in the field that causes total loss, and rotting can also be observed in the storage. This disease is caused by Erwinia carotovora [15] and Pectobacterium carotovorum [9]. Recently, Klebsiella variicola has also been reported to cause rotting disease in carrots in India [16]. Our initial studies on soft rot infected carrot samples indicated negative for the presence of E. caratovora, P. carotovorum and K. variicola, Pseudomonas viridiflava, and P. marginalis pathogens, and the isolated bacteria was shown similarities with the Acinetobacter spp. Considering the above facts, the present study was undertaken to identify and characterize the bacterial pathogens associated with the soft rot disease of carrots. We identified an isolate of A. baumannii as the causal organism of soft rot in carrots.

Acinetobacter baumannii is a ubiquitous bacterium that exists under a wide variety of environmental conditions [17] and is found as a food contaminant [18]. A. baumannii has been reported to be associated with hospital-acquired nosocomial infections in humans [19]. A. baumannii is an opportunistic organism and showed high resistance to carbapenem throughout Asia and America [20] Pneumonia, bloodstream infections, urinal tract infections, and meningitis are the most common clinical manifestations [21]. In plants, A. baumannii has been in diverse association forms from endophytic beneficial to the disease-causing pathogen. In chilli and pearl millet, A. baumannii has been reported as beneficial to plants by producing plant hormones [19,22]. Multidrug-resistant strains of A. baumannii have also been frequently isolated from crop fields [23]. Most importantly, A. baumannii has been reported as a plant pathogen causing the top rot phase of the sugarcane red stripe disease in India [24] and dieback disease of mango in Pakistan [25]. A study also indicated the competitive nature of A. baumannii in the Ralstonia infected tomato plants for in-planta multiplication [26].

In this study, we report the isolation, identification, and characterization of a cross-kingdom infecting bacteria A. baumannii associated with the soft rot disease of carrot in India. One of the strains, AB1, was taxonomically identified through sequencing 16S rDNA, followed by biochemical and molecular characterization.

Materials and Methods

Collection of diseased samples and pathogen isolation

The carrot sample exhibiting water-soaked lesions, rotting of taproot (Figure 1) were collected from Chintamani, Karnataka, India (N 130 24' 5.4'' E0 780 03' 20.1'') during 2020. About six samples were collected along with two healthy roots. Both healthy and diseased samples were pre-cleaned in the field and brought to the laboratory for further analysis. As the rotting phase of the disease is associated with many saprophytes, an enrichment technique was followed using a healthy bell pepper. Healthy and green bell pepper was washed with running tap water, then surface sterilized with 0.5% sodium hypochlorite for 1min, followed by a wash with sterile distilled water (SDW) and air-dried. A sterile toothpick was stubbed into the diseased tissue of carrot and then stubbed into the surface-sterilized bell pepper. Inoculated peppers were then placed in a plastic bag, and the moistened wet cotton was then incubated at 30°C for 24-48 hours. The diseased/symptomatic tissue around the toothpick pricking bell pepper was aseptically removed and surface sterilized with 0.5% sodium hypochlorite solution for a few seconds and then washed with SDW. The portion of the infected region was macerated in a sterile saline solution (0.85%) using a sterile pestle and mortar under aseptic conditions. The resulting suspension was left undisturbed for a few minutes for the bacterium to release from the tissue. This suspension was then streaked on Nutrient Agar (NA) plates, and the plates were incubated at 28°C for 24hr for colony emergence.