The present study deals with the cytological investigations on the meristematic root cells of carnation (Dianthus caryophyllus Linn.) grown in vivo and in vitro. Cellular parameters including the mitotic index (MI), chromosome count, ploidy level (nuclear DNA content), mean cell and nuclear areas, and cell doubling time (Cdt) were determined from the 2 mm root tip segments of this species. The MI value decreased when cells were transferred from in vivo to in vitro conditions, perhaps due to early adaptations of the cells to the in vitro environment. The mean chromosome number was generally stable (2n = 2x = 30) throughout the 6-month culture period, indicating no occurrence of early somaclonal variation. Following the transfer to the in vitro environment, a significant increase was recorded for mean cell and nuclear areas, from 26.59 ± 0.09 μm² to 35.66 ± 0.10 μm² and 142.90 ± 0.59 μm² to 165.05 ± 0.58 μm², respectively. However, the mean cell and nuclear areas of in vitro grown D. caryophyllus were unstable and fluctuated throughout the tissue culture period, possibly due to organogenesis or rhizogenesis. Ploidy level analysis revealed that D. caryophyllus root cells contained high percentage of polyploid cells when grown in vivo and maintained high throughout the 6-month culture period.
The effect of leaf extracts of Psidium guajava and Piper betle on prolonging vase life of cut carnation flowers was studied. "Carola" and "Pallas Orange" carnation flowers, at bud stage, were pulsed 24 hours with a floral preservative. Then, flowers were placed in a vase solution containing sprite and a "germicide" (leaf extracts of P. guajava and P. betle, 8-HQC, or a copper coin). Flowers treated with 8-HQC, copper coin, and leaf extracts had longer vase life, larger flower diameter, and higher rate of water uptake compared to control (tap water). The leaf extracts of P. guajava and P. betle showed highest antibacterial and antifungal activities compared to the other treatments. Both showed similar effects on flower quality as the synthetic germicide, 8-HQC. Therefore, these extracts are likely natural germicides to prolong vase life of cut flowers.
Rambutan (Nephelium lappaceum L.) is among the tropical fruit grown in Malaysia and the demand for export rose in 2011. A fruit rot was observed between August and December 2011 from several areas in the states of Pulau Pinang and Perak, Malaysia. The symptoms initially appeared as light brown, water-soaked lesions that developed first in the pericarp and pulp, later enlarging and becoming dark brown. Greyish brown mycelia were observed on infected areas that turned yellowish at later stages of infection. Gliocephalotrichum bacillisporum was isolated from infected fruit by surface sterilization techniques. Conidia were mass-transferred onto potato dexstrose agar (PDA) plates and incubated at 27 ± 1°C. Tissue pieces (5 × 5 mm) excised from the margins between infected and healthy areas were then surface sterilized in 1% sodium hypochlorite for 3 to 5 min before being rinsed with distilled water, plated on PDA, and incubated at 27 ± 1°C for 7 days. Ten isolates of G. bacillisporum were obtained. Colonies on PDA were initially white before turning yellow with a feathery appearance. Microscopic characteristics on carnation leaf agar (CLA) consisted of hyaline conidia that were slightly ellipsoid to bacilliform with rounded apex ranging from 6.0 to 8.5 μm long and 2.0 to 2.5 μm wide. Conidiophores (70 to 130 μm long) were mostly single arising from large hypha approximately 13 to 16 μm. The conidiogenous structures were mostly quadriverticillate with dense, short, penicillate branches. The phialides were cylindrical and finger-like. Chlamydospores were present singly, in groups of 2 to 4, or in occasionally branched short chains and were brown in color with thick walls ranging from 11 to 13 μm. The cultural and morphological characteristics of G. bacillisporum isolates in the present study were very similar to previously published descriptions (1) except the conidiophores formed without sterile stipe extensions. All the G. bacillisporum isolates were deposited in culture collection at the Plant Pathology Lab, University Sains Malaysia, Penang. Molecular identification was accomplished from the ITS regions using ITS1 and ITS2 primers, and the β-tubulin gene using Bt2a and Bt2b primers (2). BLAST results from the ITS regions showed a 98 to 99% similarity with sequences of G. bacillisporum isolates reported in GenBank. Accession numbers of G. bacillisporum ITS regions: JX484850, JX484852, JX484853, JX484856, JX484858, JX484860, JX484862, JX484866, JX484867, and JX484868. The identity of G. bacillisporum isolates infecting rambutan was further confirmed by β-tubulin sequences (KC683909, KC683911, KC683912, KC683916, KC683919, KC683920, KC683923, KC683926, and KC683927), which showed 92 to 95% similarity with sequences of G. bacillisporum. Pathogenicity tests were also performed using mycelial plug (5 mm) and sprayed conidial suspensions (20 μl suspension of 106 conidia/ml) prepared from 7-day-old cultures. Inoculated fruits were incubated at 27 ± 1°C and after 10 days, similar rotting symptoms appeared on the fruit surface. The pathogen was reisolated from fruit rot lesions, thus fulfilling Koch's postulates, and tests were repeated twice. To our knowledge, this is the first report of G. bacillisporum causing fruit rot of rambutan (N. lappaceum L.) in Malaysia. References: (1) C. Decock et al. Mycologia 98:488, 2006. (2) N. L. Glass and G. C. Donaldson. Appl. Environ Microbiol. 61:1323, 1995.
Fusarium species is one of the common pathogens of post-harvest disease to cause rot on tomato and other perishable vegetable fruits. The objectives of this study were to determine the diversity of Fusarium isolated species from post-harvest diseases of tomato fruit, to identify the causal organisms by using phenotype characteristics and to verify the pathogens of Fusarium fruit of tomato based on pathogenicity test. Carnation leaf-piece agar (CLA) and potato dextrose agar (PDA) media were used for phenotype-based identification of the Fusarium isolates with emphasis for characterizations of the shapes and sizes of the macroconidia and microconidia, colony features, growth rates, conidiogenous cells and chlamydospores. A total of 180 Fusarium isolates were obtained from 13 locations throughout Selangor. Fusarium solani was most abundantly isolated (34%) followed by F. semitectum (31%) and F. oxysporum (31%), F. subglutinans (3%) while the least was F. equiseti (1%). Twenty seven isolates were tested for pathogenicity test by injecting 1 mL of the conidial suspension onto healthy tomatoes. All the tested Fusarium isolates were pathogenic on tomato with different severity levels. The non-inoculated controls showed no symptoms of fruit rot. The most virulent was F. oxysporum isolate B711T with DSI 93.75%, while the least were isolates of F. solani (B647T) and F. oxysporum (B727T) with DSI 37.5%. Majority of the isolated Fusarium species can potentially produce mycotoxins as their secondary metabolites. The potential production of mycotoxins by pathogenic isolates of Fusarium species in contaminated tomato fruits could pose health hazards when consumed.
Fusarium nygamai Burgess & Trimboli was first described in 1986 in Australia (1) and subsequently reported in Africa, China, Malaysia, Thailand, Puerto Rico, and the United States. F. nygamai has been reported on sorghum, millet, bean, cotton, and in soil where it exists as a colonizer of living plants or plant debris. F. nygamai was also reported as a pathogen of the witch-weed Striga hermonthica (Del.) Benth. To our knowledge, no reports are available on its pathogenicity on crops of economic importance. In a survey of species of Fusarium causing seedling blight and foot rot of rice (Oryza sativa L.) carried out in Sardinia (Oristano, S. Lucia), F. nygamai was isolated in association with other Fusarium species-F. moniliforme, F. proliferatum, F. oxysporum, F. solani, F. compactum, and F. equiseti. Infected seedlings exhibited a reddish brown cortical discoloration, which was more intense in older plants. The identification of F. nygamai was based on monoconidial cultures grown on carnation leaf-piece agar (CLA) (2). The shape of macroconidia, the formation of microconidia in short chains and false heads, and the presence of chlamydospores were used as the criteria for identification. Two pathogenicity tests comparing one isolate of F. nygamai with one isolate of F. moniliforme were conducted on rice cv. Arborio sown in artificially infested soil in a greenhouse at 22 to 25°C. The inoculum was prepared by growing both Fusarium species in cornmeal sand (1:30 wt/wt) at 25°C for 3 weeks. This inoculum was added to soil at 20 g per 500 ml of soil. Pre- and post-emergence damping-off was assessed. Both F. nygamai and F. moniliforme reduced the emergence of seedlings (33 to 59% and 25 to 50%, respectively, compared to uninoculated control). After 25 days, the seedlings in infested soil exhibited a browning of the basal leaf sheaths, which progressed to a leaf and stem necrosis. Foot rot symptoms caused by F. nygamai and F. moniliforme were similar, but seedlings infected by F. nygamai exhibited a more intense browning on the stem base and a significant reduction of plant height at the end of the experiment. Either F. nygamai or F. moniliforme were consistently isolated from symptomatic tissue from the respective treatments. References: (1) L. W. Burgess and D. Trimboli. Mycologia 78:223,1986. (2) N. L. Fisher et al. Phytopathology 72:151,1982.