Studies on Leaf Blight Disease of Sissoo (Dalbergia sissoo Roxb.) in Bangladesh

S. Chowdhury, H. Rashid, R. Ahmed and M. M. U. Haque

  • Page No:  162 - 164
  • Published online: 16 Oct 2020
  • DOI: HTTPS://DOI.ORG/10.23910/2/2020.0383

  • Abstract
  •  masum-fes@sust.edu

The outbreak of leaf blight disease in sissoo (Dalbergia sissoo Roxb.) was detected in different plantation areas of Sylhet, Bangladesh, during surveys conducted in November 2019. Isolates were consistently recovered from the necrotic region of the blight infected leaves. Isolates grown on potato-dextrose-agar (PDA) showed greyish-white cottony mycelia with a concentric zone of orange conidial masses. Average conidia length and width ranging from 13.5-17.7 µm and 3.5-5.3 µm, respectively. In the pathogenicity test, the pathogen was able to cause infection on detached healthy leaves and spots developed were similar to those observed on the leaves affected in nature. However, the pathogen produced disease symptoms in wounded leaves but did not produce any symptoms on the non-wound ones. Findings also suggested that the pathogen was equally virulent to three-leaf grades (young, middle and mature-aged). Based on the above morphological features, the pathogen was identified as Colletotrichum sp. Molecular identification is needed to determine the pathogen up to species. The observation of the pathogen causing leaf blight disease of sissoo in Bangladesh has severe implications regarding the management of plantations and nurseries. More surveys are needed to determine the distribution and extent of damage caused by the pathogen in other regions.

Keywords :   Conidia, lesions, morphology, pathogenicity, sissoo

  • Introduction

    Sissoo (Dalbergia sissoo Roxb.), a deciduous tree of family ‘Papilionaceae’, is an essential plant of incredible monetary value. The tree has higher timber value for its use in a variety of works including furniture, fodder and medicinal (Jalota and Sangha, 2000). D. sissoo is native to the Indian subcontinent. It is extensively planted as a multipurpose tree species in Bangladesh. The enormous beneficial contributions of the tree species to the socio-economic development of the country and the South Asian regions are broadly recognized.

    Massive mortality of this economically important species has been accounted for in Bangladesh since 1993 (Webb and Hossain, 2005). One report has asserted that two million sissoo trees have died because of the infection amid a seven-year time frame in Bangladesh (Kumar et al., 2018). Foresters and pathologists have related that the high mortality with sissoo may be a pathological, edaphic, entomological, silvicultural, age and soil factors. Among these, pathological and management solutions have gotten the most consideration when Fusarium solani was reported as the main agent of mortality of sissoo (Bakshi, 1995). Pests responsible are Plecoptera reflexa (a defoliator), Dichomeris eridans (leaf folio), Brachytrypes portentosus (causing nursery harm) and termites that attack the young trees. Previously, it was reported that microorganisms like bacteria is responsible for dieback disease of sissoo (Valdez et al., 2013). Yet, numerous vital fungal diseases like leaf blight, Leaf spot, leaf rust, powdery mildew are likewise revealed. However, information on the fungal pathogens responsible for the diseases of sissoo in Bangladesh is scanty. Therefore, an attempt has been made in the present study to identify the pathogen associated with the leaf blight disease of D. sissoo and to characterize the pathogen by analyzing its morphology and pathogenicity.


  • Materials and Methods

    2.1.  Collection of diseased samples

    Leaves having typical symptoms including long, elliptical, and necrotic lesions were collected for the isolation of casual organism associated with leaf blight disease of D. sissoo. The samples were collected separately in polythene bags and brought to the laboratory for isolation and identification of the causal organism. The samples were kept in the refrigerator for future study.

    2.2.  Isolation of pathogen

    The collected samples were washed with distilled water. Small pieces of infected leaves including healthy zone, transition zone and some advanced zone of infection were cut out from the infected leaves. The pieces were dipped in 70% alcohol for two minutes and then washed with distilled water three times. The samples were then kept inside laminar airflow for air drying to avoid excess water present in the sterilized inoculum (pieces of infected host leaves). The surface-sterilized pieces were placed in Petri-dishes containing PDA medium and kept in room temperature and examined regularly for any mycelial growth. The fungus that developed from the infected leaf tissues was transferred to fresh PDA to obtain a pure culture.

    2.3.  Morphological identification

    To examine morphological features of the pathogen, agar discs (5 mm) were cut from the leading edge of 6-day old colonies and placed upside down in the centre of Petri dishes containing PDA media (one plug per plate). Then the plates were incubated at 25°C. Colony growth patterns were examined after six days and recorded. Morphology of the conidia was examined at 40X magnification.

    2.4.  Pathogenicity

    Pathogenicity test was performed following the modified detached leaf assay technique (Shamsi and Saha, 2015) to test whether the pathogen can develop the same symptoms of leaf blight as the naturally developed symptoms or not. Fresh and healthy leaves of three categories (young, middle-aged and matured) were collected from the disease-free areas. At first, the leaves were washed with distilled water, surface sterilized with 70% alcohol for one minute and rinsed in sterilized distilled water three times. Ventral and dorsal sides of the leaflets with and without pricking with needles were inoculated with 2 mm diameter mycelial block of the isolated fungi previously grown on PDA medium for seven days. The inoculated leaflets were placed in Petri dishes containing a water-soaked filter paper. Water was sprayed carefully inside the chamber at every 24 hours. The plates were incubated at 25°C and observed daily for any infection.

    2.5.  Statistical analysis

    To observe the effects leafage and isolates on lesion length, analysis of variance (ANOVA) was carried out. Statistical analyses were done by the R software environment (R core team, 2014).


  • Results and Discussion

    3.1.  Symptom of the disease studied

    A preliminary study in the symptoms of the leaf blight disease of D. sissoo was made in this study. The symptoms were recorded on the basis of the samples collected from the field for isolation of the associated organism. Before isolation of the organism, the infected leaves with more or less similar type of symptoms were sorted out from a large number of infected leaves. The symptoms of the leaf blight disease of D. sissoo were reported to occur, especially in the month of November-December. Symptoms were found to start from the margin of the leaf blade and gradually cover the entire leaf blade both dorsal and ventral side. Reticulate veins, as well as the midrib of the leaves, turned into brown. Comparatively, the dorsal side is more vulnerable (Figure 1A).

    3.2.  Identification of the pathogen

    The standard technique was followed to identify the pathogen responsible for the leaf blight disease of D. sissoo. The isolation technique was repeated several times to obtain a pure culture. The isolates grown on PDA showed greyish-white cottony mycelia with a coaxial zone of orange conidial masses (Figure 1B). The yielded pure cultures of the pathogen were maintained in PDA slants and Petri plates at 25±1°C in the incubator for further investigation. Conidia were cylindrical with rounded ends and aseptate with average conidia length and width ranging from 13.5-17.7 µm and 3.5-5.3 µm respectively. All these features matched the published description of Collectotrichum species as described (Weir et al., 2012).  The pure culture of pathogen was sent to the Biotechnology laboratory of Shahjalal University of Science and Technology (SUST), Bangladesh for further identification and confirmation of the pathogen.


    3.3.  Pathogenicity

    The pathogen produced typical symptoms of blight in the pathogenicity test (Figure 2). The sign of infection was seen to start after six days of inoculation in the leaves which were wounded. No lesions were observed in the non-wound leaves. The infection of the pathogens was not influenced by leaf age. All type of leaves (young, middle age, matured) was found to be susceptible to the pathogen. A significant interaction between the isolate and leafage was observed for the lesions (F=3.59, p< 0.05). According to the isolate-leaf age interaction, the most virulent isolate was S-2 on the middle-aged leaf with a lesion length 2.8±.84) (Table 1). However, no significant difference in length noted among the lesion lengths on three leaf grades (ages).


    In pathogenicity test, all ages of leaves of D. sissoo were found susceptible to the pathogen. However, no significant differences were found in lesion lengths of the different leaf grades, which matched with the findings of Chen et al. (2017). Un-pricked leaves didn’t produce any symptoms on the inoculated leaves in the pathogenicity test. Such wounding effects is consistent with the previous findings (Chen et al., 2017).


  • Conclusion

    The pathogen associated with the leaf blight disease of Dalbergia sissoo was identified as Colletotrichum sp.All the leaf grades (young, middle, mature) were found to be susceptible to the pathogen in the pathogenicity test. However, only the wounded leaves developed the lesions which suggested that when the leaves are damaged by any external source (insects, birds, animals etc. or any other mechanical damages), then it becomes more susceptible to the pathogen infection. More studies are necessary to understand the host-pathogen interactions and for its control and management.


  • Future Research

    Detailed molecular and physiological characterization will be done in order to identify the pathogen up to species.


  • Acknowledgement

    The authors would like to thank the Research Centre of the Shahjalal University of Science and Technology (SUST), Bangladesh, for funding this work (grant no. FES/2018/2/05).


    Reference

  • Bakshi, B.K., 1995. Wilt diseases of shisham (Dalbergia sissoo Roxb.) II-Behaviour of Furarium solani, the wilt organism in soil. Indian Forester 81, 276−281.

    Chen, Y., Qiao, W., Zeng, L., Shen, D., Liu, Z., Wang, X., Tong, H., 2017. Characterization, Pathogenicity, and Phylogenetic analyses of Colletotrichum species associated with brown blight disease on Camellia sinensis in China. Plant Disease 101, 1022−1028.

    Jalota, R., Sangha, K., 2000. Comparative ecological-economic analysis of growth performance of exotic Eucalyptus tereticornis and indigenous Dalbergia sissoo in mono-culture plantations. Ecological Economics 33, 487−495.

    Kumar, V., Jain, K., Kumar, S., Kumhar, B., 2018. Impact of different pruning of Dalbergia sissoo and different date of planting of turmeric on growth and yield. International Journal of Forestry and Crop Improvement 9, 29−32.

    R-Development-Core-Team., 2008. R: A language and environment for statistical computing: R Foundation for Statistical Computing. Vienna, Austria

    Shamsi, S., Saha, T., 2015. Management of anthracnose and blight diseases of Houttuynia cordata Thunb. with fungicides. Journal of Bangladesh Academy of Sciences 39, 83−90.

    Valdez, N., Karlovsky, P., Dobrindt, L., Hoque, M., Sarker, R., Tantau, H., Muhlbach, H., 2013. Role of bacteria in dieback disease of Dalbergia sissoo Roxb. Bangladesh Journal of Botany 42(1), 1−16.

    Webb, E.L., Hossain, S.M.Y., 2005. Dalbergia sissoo mortality in Bangladesh plantations: Correlations with environmental and management parameters. Forest Ecology and Management 206, 61–69.

    Weir, B.S., Johnston, P.R., Damm, U., 2012. The Colletotrichum gloeosporioides species complex. Studies in Mycology73, 115−180.

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