Development of multiplex RT-PCR assay for simultaneous detection of four viruses infecting apple (Malus domestica).

The major viruses infecting apple cultivars throughout the world including India are apple mosaic virus (ApMV), apple stem pitting virus (ASPV), apple stem grooving virus (ASGV), apple chlorotic leaf spot virus (ACLSV), and recently, a new virus, apple necrotic mosaic virus (ApNMV), was reported from mosaic-infected apple cultivars in India. The aim of this study was to detect the ApNMV virus along with the other three viruses (ApMV, ASPV and ASGV) simultaneously by multiplex RT-PCR. Four primer-pair-produced amplicons of 670, 550, 350 and 210 bp corresponding to ApNMV, ApMV, ASPV and ASGV, respectively, were found to be specific for these viruses when tested individually. The annealing temperature (55°C), primer concentration (0·8 µl) and other components of the master mix were standardized for the development of one-step m-RT-PCR assay. The m-RT-PCR protocol developed was further validated with 30 samples from seven symptomatic or asymptomatic apple cultivars, which revealed the presence of more than one virus in these cultivars. Most of the viruses were found to be present either alone or in mixed infection; however, ASPV was more common in tested cultivars. An easy, cost-effective and rapid multiplex RT-RCR protocol was developed to detect the four viruses, which infect apple plants either in individually or together in the field. This assay will help in the surveying and indexing of apple germplasm and the distribution of all four viruses in the apple growing regions of India.

Value Addition and Fortification in Non-Centrifugal Sugar (Jaggery): A Potential Source of Functional and Nutraceutical Foods.

Nutraceutical, functional or fortified food not only supplements the diet but also assists in developing immunity and preventing diseases. Therefore, it also provides medical benefits apart from nutrition. Individual health and diet are receiving a lot of attention these days. Food products that are healthy, safe, and easy to use are in high demand. A lot of emphases has been directed toward food products with added health benefits. Jaggery (non-centrifugal sugar), made from sugarcane juice, is one of them. It is known to mankind for the last 3000 years and is an essential part of the diet in the rural part of many countries. Jaggery is recognized as a nutraceutical due to the presence of a variety of essential amino acids, antioxidants, phenolics, minerals like calcium, phosphorus, iron, and vitamins. Jaggery has a better natural source and nutrients for health benefits and could be used as a healthier dietary substitute to white sugar. The nutraceutical value of jaggery can be enhanced with value addition and fortification of medicinally important herbs and spices. This would not only enhance the quality of nutritive jaggery but will also offer value-added products with exciting flavors for daily use sweeteners with several nutritional health benefits.

Molecular characterization of 'Clover proliferation' phytoplasma subgroup-D (16SrVI-D) associated with vegetables crops in India.

Nine vegetable plants species exhibiting phytoplasma suspected symptoms of white/purple leaf, little leaf, flat stem, witches' broom, phyllody and leaf yellowing were observed in experimental fields at Indian Agricultural Research Institute, New Delhi from December 2015 to July 2016. Total DNA extracted from the three healthy and three symptomatic leaves of all the nine vegetables were subjected to PCR assays using phytoplasma specific primers P1/P7 followed by R16F2n/R16R2 and 3Far/3Rev to amplify the 16S rDNA fragments. No amplifications of DNA were observed in first round PCR assays with primer pair P1/P7 from any of the symptomatic samples. However, phytoplasma DNA specific fragments of ~ 1.3 kb were amplified from Apium graveolens L. (two isolates), Brassica oleracea vr. capitata L. (one isolate) and Solanum melongena L. (one isolate) by using 3Far/3Rev primer pair and 1.2 kb fragment was amplified from Lactuca sativa L. (one isolate) by using R16F2n/R16R2 primer pair. No DNA amplification was seen in other symptomatic vegetable samples of tomato, carrot, cucurbit, bitter gourd and Amaranthus species utilizing either P1/P7 primer pair followed by 3Far/3Rev or R16F2n/R16R2 primer pairs. Out of three leafhopper species collected from the symptomatic vegetable fields, only Hishimonus phycitis was found positive for association of phytoplasma. No DNA amplifications were observed in healthy plant samples and insects collected from non-symptomatic fields. Comparative sequence comparison analyses of 16S rDNA of positive found vegetable phytoplasma strains revealed 100% sequence identities among each other and with phytoplasma strains of 'clover proliferation' (16SrVI) group. Phytoplasma sequences, virtual RFLPs and phylogenetic analyses of 16S rDNA sequence comparison confirmed the identification of 16SrVI subgroup D strain of phytoplasmas in four vegetables and one leafhopper (HP) species. Further virtual RFLP analysis of 16S rDNA sequence of the vegetables phytoplasma strains confirmed their taxonomic classification with strains of 'clover proliferation' subgroup D. Since, H. phycitis feeding on symptomatic vegetable species in the study was also tested positive for the 16SrVI phytoplasma subgroup-D as of vegetables; it may act as potent natural reservoir of 16SrVI-D subgroup of phytoplasmas infecting vegetable and other important agricultural crops.

First Report of Myrothecium roridum Causing Leaf Spot of Anubias barteri var. nana in Mainland China.

Anubias barteri var. nana (belonging to family Araceae) is one of the most popular ornamental plants for aquaria. It has slow growth and small attractive ornamental leaves. It originated from West Africa, was first described in 1860 by Heinrich Wilhelm Schott (1), and is now commercially planted widely in southern China. A new disease of A. barteri var. nana was observed on leaves. Approximately 30% of the plants in an ornamental nursery in Baiyun district, Gaungzhou, China (113°22'45.15″ E, 23°23'41.15″ N) in July 2013 were found to be infected. The pathogen caused necrotic spots that progressed to form holes on leaves of the infected plants, negatively affecting their ornamental market value. Eventually the diseased plant died. White fungal fruiting bodies and black sporodochia were observed on the surfaces of the symptomatic leaves. A single-spore isolate (myr5) was obtained from the concentric lesions and cultured on potato dextrose agar (PDA). The floccose fungal colonies were white to buff, and black conidiomata were often visible on the surface after 25 days of incubation. The conidia had rounded ends and the average size of conidia was 5.98 ± 0.15 × 2.24 ± 0.08 µm. The rDNA internal transcribed spacer (ITS) region of isolate myr5 was amplified by PCR using ITS1 and ITS4 primer pairs. The amplified product was sequenced and deposited in GenBank (Accession No. KJ572115) and showed 99% identity to Myrothecium roridum isolates BBA 71015 (AJ302001) and BBA 67679 (AJ301995) (unpublished), and 100% identity to the isolate myr2-2 from Dieffenbachia picta 'Camilla' in Taiwan (2). On the basis of morphological and molecular characterization, the fungus causing leaf spot on A. barteri var. nana plants was identified as M. roridum Tode ex Fr (2,3). To confirm the etiology of the disease, Koch's postulates was performed in a greenhouse (28 ± 2°C) using 3-day-old cultures of isolate myr5 and fungal spore suspensions of 1 × 105 conidia ml-1 containing 0.05% of Tween 20. Sixteen healthy leaves, two each from eight plants, were infiltrated on two different parts close to the midribs with 100 µl of the fungal spore using needleless syringes. Water infiltration was used as the control treatment. Water-soaked brown lesions appeared on leaves 7 days after inoculation, followed by the development of dark concentric rings within the necrotic areas on the surface of the inoculated leaves after 15 days of incubation. These symptoms were similar to those in the naturally infected aquarium plants. No symptoms were observed on any of the water micro-infiltrated plants. The fungus was re-isolated from the inoculated plants, but not from control plants. To our knowledge, this is the first report of Myrothecium leaf spot caused by M. roridum on A. barteri var. nana in mainland China. References: (1) W. Crusio. Primitiae Africanae XII 79 (14):1-48, 1979. (2) C. F. Hong et al. Plant Dis. 97:1253, 2013. (3) M. Tulloch. Mycol. Pap. 130:1-42, 1972.

First Report of Phoma sp. Causing Twisting and Curling of Crown Leaves of Sugarcane in Mainland of China.

Sugarcane is a major sugar and the leading energy crop worldwide and Guangxi is the largest sugarcane production area in China (2011 Sugar Annual China, www.gain.fas.usda.gov). During survey of sugarcane crops in September 2012 and June 2013, ~5 to 10% of sugarcane (cvs. FN-40 and ROC22) planted in Chongzuo and Laibing, Guangxi Zhuang Autonomous Region, P. R. China, had twisted and curling symptoms of crown leaves similar to sugarcane Pokkah boeng disease (caused by Fusarium moniliforme Sheldon). The symptoms started appearing as yellowing on midribs and leaf margins that spread further to the entire leaf, along with twisting and curling of crown leaves. The symptomatic leaf tissues (5 × 5 mm) were surface-sterilized by 0.1% HgCl2 solution for 30 s, followed by rinsing three times in sterile water, placed on potato dextrose agar (PDA), and then incubated in darkness at 28°C. After 3 days of incubation, the isolated fungal colony appeared as white villous, spherical, radial, and dense colorless mycelium from the top, while it was reddish-brown at the bottom and later became grayish. Chlamydospores were also observed with a diameter of 5 to 10 µm and were dark brown, unicellular, intercalary, and smooth. The binucleate hyphae were colorless and transparent. Pycnidia appeared on the colonies after 20 days, and were dark brown, subglobose, and 150 to 230 µm in diameter, and the conidia were ~3 to 7 × 2.5 to 6 µm, unicellular, colorless, and ovoid to oval. The fungal isolates from the symptomatic leaves were obtained and pathogenicity was evaluated. Conidial suspensions (107 CFU/ml) of the single isolate from FN-40 were micro-injected into 20 sugarcane seedlings of cultivar FN-40. Another 20 seedlings were injected with water without conidia as control. The inoculated plants were grown in a growth chamber at 28°C with a 16-h photoperiod. Twisted and curly symptoms similar to the field appeared on the inoculated leaves at 10 days after inoculation, while the control leaves remained asymptomatic. The fungus was re-isolated and identified. Genomic DNA from the cultured fungal isolate was extracted with a modified Fungal DNA Midi Kit (Omega Bio-Tek, Inc., Norcross, GA), and amplified using fungus-conserved primer sequences (ITS1: 5'-TCCGTAGGTGAACCTGCGG-3' and ITS4: 5'-TCCTCCGCTTATTGATATGC-3'). The consensus rDNA-internal transcribed spacer sequence (GenBank Accession No. KC524502) was 100% identical with 97% coverage to the ITS sequence from Phoma sp. 3. TMS-2011 (HQ631000.1) in GenBank (3). The fungus Phoma sp. was identified on the basis of morphological characteristics (2,4) and the ITS sequence of rDNA (1,3). Disease caused by Phoma sp. has been reported earlier on sugarcane from Pakistan, Hawaii, and Taiwan, causing leaf blight and curling (2,4). However, to the best of our knowledge, this is the first report of Phoma sp. causing twisting and curling of crown leaves of sugarcane in mainland of China. References: (1) M. M. Aveskamp et al. Stud. Mycol. 65:1, 2010. (2) A. Sanguino and H. Tokeski. ISSCT Proc. 17:1555, 1980. (3) P. Shrestha et al. Appl. Environ. Microbiol. 77:5490, 2011. (4) Z. N. Wang. Rep. Taiwan Sugar Res. Inst. 129:1, 1980.

Draft genome sequence of 'Candidatus Phytoplasma australasia', strain SS02 associated with sesame phyllody disease.

'Candidatus Phytoplasma' is an uncultivated, intracellular bacterial plant pathogen transmitted by phloem-feeding insect vectors. Among the group of phytoplasmas, the Peanut Witches' Broom or 16SrII group of phytoplasmas associated with various diseases cause severe crop losses every year in India. The 'Ca. Phytoplasma sp.' strain SS02 was associated with phyllody disease of sesame plants collected from New Delhi. The genome sequence of strain SS02 was obtained using its genomic DNA enrichment and hybrid assembly of sequences generated on Illumina and Oxford Nanopore Technologies MinION platforms. The hybrid assembly strategy generated a draft genome with 60 contigs totaling 553,228 bp of length with more than 400 × depth coverage and 95.21% of the estimated completeness. The SS02 genome draft sequence contains 465 protein-coding genes, 17 tRNA genes, and 3 rRNA genes. The availability of this draft genome also provided a foundation for genome-scale genotypic analyses.

Multilocus gene analysis reveals the presence of two phytoplasma groups in Impatiens balsamina showing flat stem and phyllody.

Rose balsam (Impatiens balsamina) is an important ornamental species grown worldwide for its attractive flowers and also having medicinal properties. Flat stem, little leaf, and phyllody symptoms were observed in I. balsamina nurseries in Uttar Pradesh and Tripura states of India during surveys from 2018 to 2020, with an incidence from 6 to 27%. Amplicons of ~ 1.2 kb were amplified in all the tested symptomatic samples of I. balsamina using universal phytoplasma primer pairs from different surveyed locations, but not from the asymptomatic plants. Pairwise sequence comparison, phylogeny, and virtual RFLP analysis of 16S rRNA gene sequences identified the phytoplasmas as 16SrI-B subgroup strain from Tripura (Lembucherra) and 16SrII-D subgroup strain from Uttar Pradesh (Gorakhpur and Faizabad). Phytoplasma presence and identity was further confirmed by amplifying secA, rp, secY, and tuf genes. This is the first report of 16SrI-B and 16SrII-D phytoplasmas detection in I. balsamina in the world. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02666-2.

Characterization and genetic diversity of phytoplasmas associated with elephant foot yam (Amorphophallus paeoniifolius) in India.

During the growing season of 2015 and 2016, leaf yellowing, stunting, and declining symptoms were observed on elephant foot yam in three states of India.The 1.3 kb 16S rDNA fragments were amplified from genomic DNA extracted from all the symptomatic elephant foot yam samples in nested PCR assays using primer pairs, P1/P7 followed by 3F/3R. Pair wise sequence comparison, virtual RFLP and phylogenetic analysis of 16S rRNA gene sequences confirmed association of 'Candidatus Phytoplasma trifolii' (16SrVI-D) and 'Candidatus Phytoplasma oryzae' (16SrXI-B) related strains in Uttar Pradesh, Uttarakhand and Tripura states, respectively. To our knowledge, this is the first report of association of 'Ca. P. trifolii' and 'Ca. P. oryzae' related strains in elephant foot yam in world. In the present study, we also reported Datura stramonium showing witches' broom as a natural weed host for 'Ca. P. trifolii' phytoplasma in Gorakhpur and Kushinagar districts of Uttar Pradesh state, India.

Genetic diversity of phytoplasma strains infecting chrysanthemum varieties in India and their possible natural reservoirs.

Symptoms typical of phytoplasma infection such as phyllody, virescence, witches' broom and yellowing were observed in 12 varieties of Chrysanthemum morifolium in floral nurseries and experimental fields at New Delhi, Karnataka, Maharashtra and Andhra Pradesh, India, during surveys made from 2015 to 2017. Disease incidence ranged from 15 to 30%. Phytoplasma presence was confirmed in all symptomatic chrysanthemum varieties by molecular identification assays. Sequence comparison, phylogenetic and in silico RFLP analyses of 16S rDNA sequences allowed the identification of the chrysanthemum infecting phytoplasma strains into different ribosomal groups and subgroups, namely 16SrI, 16SrII-D, 16SrVI-D and 16SrXIV. Detection of phytoplasma strains of 16SrII-D subgroup were also confirmed in symptomatic Chenopodium album and Parthenium hysterophorus plants grown in and around the surveyed chrysanthemum fields at New Delhi, whereas 16SrVI-D phytoplasma strains were detected in symptomatic Cannabis sativa weed and leafhopper Hishimonus phycitis individuals collected from the symptomatic chrysanthemum fields at New Delhi. This is the first report on the presence of 16SrVI and 16SrXIV groups of phytoplasmas in chrysanthemum plants. Studies on genetic diversity of phytoplasmas infecting the major chrysanthemum varieties in India and their epidemiological aspects had previously not been reported. The detection and identification of phytoplasmas in different chrysanthemum varieties could contribute to increase the awareness among farmers in the management of these diseases.

Development and utilization of gyrA and gyrB gene-based diagnostics for the phytoplasma classified under 16Sr I group in plants and insects.

In the present study, a new set of primers of gyrA and gyrB genes of the phytoplasma genome were designed and validated for the successful detection and taxonomic classification of the previously identified phytoplasma strains of 'Candidatus P. asteris' (16SrI-B subgroup) associated with Catharanthus leaf yellows, sesame phyllody and the leafhopper (Hishimonus phycitis). Our results suggested the ability and sensitivity of gyrA and gyrB genes as an alternative molecular marker to identify the Ca. P. asteris strain up to subgroup level associated both with plants and insects.

Multilocus gene-specific characterization of 'Candidatus Phytoplasma australasia' associated with shoot proliferation disease of small cardamom in India.

Symptoms of excessive shoot proliferation were observed in the Njallani cultivar of small cardamom accompanied by stunting of stalks with fewer degenerated capsules at Nedumkandam Panchayat of Idukki district of Kerala in 2017. Five symptomatic Elettaria cardamomum shoot proliferation (ECSP) plant samples were collected and processed for DNA extraction and PCR assays utilizing universal phytoplasma 16S ribosomal-specific primers pair, P1/P7 followed by R16F2n/R16R2. Sequence comparison analysis of the R16F2n/R16R2 region of 16SrRNA gene showed 100% sequence identity with the 'Candidatus Phytoplasma australasia'- related strain. Phylogeny and virtual RFLP analyses of 16S rRNA gene sequences confirmed the association of 'Ca. P. australasia' strain subgroup D with ECSP disease. The association of 16SrII group was further established and validated by amplifying phytoplasma-specific multilocus candidate genes by utilizing specific primers of secA, secY, SAP11, and tuf genes. The multilocus gene sequence comparison analysis again confirmed the association of 'Ca. P. australasia' with the ECSP phytoplasma isolate. This is the first report of phytoplasma association with small cardamom.

Activated carbon from Ceiba pentandra hulls, an agricultural waste, as an adsorbent in the removal of lead and zinc from aqueous solutions.

The ability of low-cost activated carbon prepared from Ceiba pentandra hulls, an agricultural waste material, for the removal of lead and zinc from aqueous solutions has been investigated. In the batch tests experimental parameters were studied, including solution pH, contact time, adsorbent dose and initial metal ions concentration. The adsorbent exhibited good sorption potential at pH 6.0. Maximum removal of lead (99.5%) and of zinc (99.1%) with 10 g/l of sorbent was observed at 50 mg/L sorbate concentration. Removals of about 60-70% occurred in 10 min, and equilibrium was attained at around 50 min for both metals. The functional groups (CO, SO,-OH) present on the carbon surface were responsible for the adsorption of metal ions. The adsorption parameters were analysed using both the Freundlich and Langmuir models. The data are better fitted by the Freundlich isotherm as compared to Langmuir model, and the adsorption capacities for lead and zinc were 25.5 and 24.1 mg/g, respectively. Kinetics of adsorption obeyed a second order rate equation and the rate constant was found to be 2.71 x 10(-2) and 2.08 x 10(-2) g/mg/min for lead and zinc, respectively. The desorption studies were carried out using dilute HCl, and the effect of HCl concentration on desorption was studied. Maximum desorptions of 85% for lead and 78% for zinc were attained with 0.15 M HCl.

Molecular characterization of phytoplasma associated with four important ornamental plant species in India and identification of natural potential spread sources.

Phytoplasma suspected symptoms of phyllody, witches' broom, leaf yellowing, stunting and little leaf were observed in Chrysanthemum morifolium, Bougainvillea glabra, Jasminum sambac and Callistephus chinensis during survey of flower nurseries and experimental ornamental fields at Delhi, Maharashtra, Tamil Nadu and Karnataka from 2014 to 2016. Pleomorphic bodies typical to phytoplasma structures were observed in the phloem sieve elements of ultrathin sections of all the four symptomatic ornamental plants (stem tissue) in transmission electron microscope. Amplification of 1.8 and 1.2 kb phytoplasma DNA products was observed in all the four test plants in PCR assays using universal primer pairs P1/P7 followed by nested primer pair R16F2n/R16R2, respectively. Pairwise sequence comparison, phylogeny and virtual RFLP analysis of 16S rDNA sequences confirmed the association of two phytoplasma subgroups (16SrI-B and 16SrII-D) in four ornamental plant species. 'Ca. P. aurantifolia' subgroup D (16SrII-D) was found associated with chrysanthemum phyllody and leaf yellowing at Delhi and Tamil Nadu, bougainvillea little leaf and yellowing at Delhi and Chinese aster phyllody at Bengaluru, Karnataka. However, jasmine little leaf and yellowing at Bengaluru, Karnataka and chrysanthemum stunting at Pune were found to be associated with 'Ca. P. asteris' subgroup B-related strains (16SrI-B). The identification of 16SrII-D subgroup phytoplasma infecting bougainvillea and 16SrI-B subgroup infecting jasmine are the new reports to the world. Besides weed species, Cannabis sativa showing witches' broom in jasmine fields at Bengaluru and Parthenium hysterophorus showing witches' broom symptoms in chrysanthemum fields at Delhi were identified to be caused by phytoplasma strains classified under subgroups 16SrI-B and 16SrII-D, respectively, by PCR assays and 16Sr DNA sequence comparison analysis. Among the three major leafhopper species identified, only Hishimonas phycitis was identified positive for 16SrI-B and 16SrII-D subgroups of phytoplasmas from chrysanthemum fields at Delhi and jasmine fields at Bengaluru, respectively. The identity of similar phytoplasma strains infecting ornamental species in leafhopper and the weed species in the present study suggested that H. phycitis and weeds may act as potential natural sources for secondary spread of the identified phytoplasma strains.

Molecular detection of 16SrI-B and 16SrII-D subgroups of phytoplasma associated with flat stem and witches' broom disease of Celosia argentea L.

Symptoms of stunting (shortening of internodes), twisting and flat stem (the fasciation of a stem), discoloration of petals, deformed flowers, and witches' broom were recorded on an ornamental plant, plumed cockscomb (Celosia argentea L., fam: Amaranthaceae). The survey conducted at Indian Agricultural Research Institute (IARI) campus, New Delhi and Karnal region, Haryana, India, during September 2014 to March 2015 revealed disease incidence of 40 and 10%, respectively. The 16S rRNA gene sequence comparison and phylogenetic relationships of Celosia phytoplasma strains under study confirmed that they were associated with two different phytoplasma groups ('Candidatus Phytoplasma australasia' and 'Ca. P. asteris'). Virtual RFLP analysis of 16S rRNA gene sequences allowed further classification of the Celosia phytoplasma strains into the 16SrI-B and 16SrII-D subgroups. Notably, the detection of 'Ca. P. asteris' phytoplasma was reported in seeds of C. argentea by nested PCR assays; however, no evidence of phytoplasma presence was detected in seedlings raised from these seeds. This observation is the first record of the association of 16SrI-B and 16SrII-D subgroups of phytoplasmas with flat stem and witches' broom disease of C. argentea anywhere from the world.

First report of association of potato virus X and potato virus Y and 'Candidatus Phytoplasma trifolii' in brinjal in India.

Symptoms of little leaf, leaf chlorosis and leaf malformations with mosaic mottling symptoms were observed in two brinjal varieties (Pusa Shyamla and Pusa Purple Cluster) in fields of IARI, New Delhi, India during 2014-2015. Electron microscopy, PCR and sequence analysis first time provided evidence of association of Candidatus Phytoplasma trifolii with potato virus X and potato virus Y in brinjal in India.

Molecular characterization of phytoplasmas of 'Clover proliferation' group associated with three ornamental plant species in India.

Suspected phytoplasma symptoms of little leaf, yellowing, chlorosis, phyllody, witches' broom, and stunting were observed on ten different ornamental plant species at New Delhi, Andhra Pradesh, Haryana, Bengaluru, and Pune, India, during March to July 2016. To investigate the possibility of phytoplasma etiology, PCR assays were performed using universal primer pairs (P1/P7 followed by 3Far/3Rev) specific to the phytoplasma 16Sr RNA gene. First round PCR amplification with primer pair P1/P7 did not yield expected 1.8 kb product of 16S rRNA region from any of the 17 symptomatic samples. However, 1.3 Kb amplicons were observed in nested PCR assays with 3Far/3Rev primer pair in symptomatic leaf samples of Hibiscus rosa-sinensis L. (Pune isolate), Saponaria officinalis L. (Pune isolate), and Allamanda cathartica L. (Delhi isolate). No amplifications were observed in any of the other tested symptomatic and non-symptomatic plant samples either in first round or second round of nested PCR assays with phytoplasma specific primer pairs. Pairwise sequence comparison of 16S rDNA sequences of the five positive phytoplasma strains of A. catharica, H. rosa-sinensis, and S. officinalis in the present study revealed 99-100% sequence identities with strains of 'clover proliferation' (16SrVI) group. Phylogenetic and virtual RFLP analysis of 16S rDNA sequences of the five identified phytoplasma strains belonging to three ornamental species further confirmed their clustering and grouping with member strains of 'clover proliferation' subgroup D. This is the first record of the phytoplasma association of 'clover proliferation' subgroup D with H. rosa-sinensis, S. officinalis, and A. cathartica in the world.

Ophthalmic manifestations of Rathke's cleft cysts.

PURPOSE: We studied the neuro-ophthalmic features of patients with symptomatic Rathke's cleft cysts, to distinguish features of Rathke's cleft cysts from those of craniopharyngioma. METHODS: Eleven patients who had undergone surgery for symptomatic Rathke's cleft cysts in the Liverpool University Department of Neurological Science were identified from the records. Histopathologic analyses, clinical notes, and radiologic investigations were reviewed. RESULTS: Eight of the 11 patients initially had visual problems. Reduced visual acuity, visual field defect, or both, were detected in nine patients. Optic atrophy was observed in eight patients. Other initial features included headaches, epilepsy, and endocrine disturbance. All patients recovered from surgery, and an improvement in the visual acuity or visual field defect was noted in six patients. CONCLUSIONS: Symptomatic Rathke's cleft cysts often manifest as visual disturbance. Every effort to distinguish Rathke's cleft cysts from craniopharyngiomas should be made preoperatively. Rathke's cleft cysts only require limited surgical intervention, and radiotherapy is not necessary. Early recognition and treatment can lead to improvement in visual function.

Spinal intramedullary tuberculous lesion: medical management. Report of four cases.

The author describes the successful medical management of intramedullary tuberculous lesions in four patients who received treatment between 1994 and 1997. The role of magnetic resonance imaging and the treatment protocol for intramedullary tuberculous lesions are also discussed.

Paradoxical progression of tuberculous lesions during chemotherapy of central nervous system tuberculosis. Report of four cases.

The peculiar phenomenon of paradoxical progression during the treatment of central nervous system tuberculosis is discussed. A few cases with this phenomenon were reported in the past, and the authors have treated four such cases. During the treatment for tuberculous meningitis, the four patients developed new lesions, mainly in the form of tuberculomas, which progressed for some time and later regressed. In all these cases the initial drug regimen was not changed, except for the addition of steroids for a short period at the time of deterioration. All four patients underwent ventriculoperitoneal shunt insertion during the course of treatment. The authors discuss the significance of the changes in the lesions and management of such cases, and review the literature.

Intracerebral polyposis. Case report.

A 25-year-old man presented with nontraumatic cerebrospinal fluid rhinorrhea and meningitis. On investigation, he was found to have a multiloculated intracerebral cystic lesion of the right frontal lobe with a bony lesion inside the cyst, just above the right cribriform plate. Surgery revealed multiple grape-like cystic pedunculated lesions with narrow stalks attached to a bony outgrowth which was adherent to the right cribriform plate. Macroscopically and microscopically, the excised lesions were similar to nasal polyps.