A. M. Anthony Johnson

Bacilliform DNA-containing plant viruses in the tropics: commonalities within a genetically diverse group.

UNLABELLED Plant viruses, possessing a bacilliform shape and containing double-stranded DNA, are emerging as important pathogens in a number of agricultural and horticultural crops in the tropics. They have been reported from a large number of countries in African and Asian continents, as well as from islands from the Pacific region. The viruses, belonging to two genera, Badnavirus and Tungrovirus, within the family Caulimoviridae, have genomes displaying a common plan, yet are highly variable, sometimes even between isolates of the same virus. In this article, we summarize the current knowledge with a view to revealing the common features embedded within the genetic diversity of this group of viruses. TAXONOMY Virus; order Unassigned; family Caulimoviridae; genera Badnavirus and Tungrovirus; species Banana streak viruses, Bougainvillea spectabilis chlorotic vein banding virus, Cacao swollen shoot virus, Citrus yellow mosaic badnavirus, Dioscorea bacilliform viruses, Rice tungro bacilliform virus, Sugarcane bacilliform viruses and Taro bacilliform virus. MICROBIOLOGICAL PROPERTIES Bacilliform in shape; length, 60-900 nm; width, 35-50 nm; circular double-stranded DNA of approximately 7.5 kbp with one or more single-stranded discontinuities. HOST RANGE Each virus generally limited to its own host, including banana, bougainvillea, black pepper, cacao, citrus species, Dioscorea alata, rice, sugarcane and taro. DISEASE SYMPTOMS Foliar streaking in banana and sugarcane, swelling of shoots in cacao, yellow mosaic in leaves and stems in citrus, brown spot in the tubers in yam and yellow-orange discoloration and stunting in rice. USEFUL WEBSITES http://www.dpvweb.net.

Sequencing and computational analysis of complete genome sequences of Citrus yellow mosaic badna virus from acid lime and pummelo

Citrus yellow mosaic badna virus (CMBV), a member of the Family Caulimoviridae, Genus Badnavirus, is the causative agent of Citrus mosaic disease in India. Although the virus has been detected in several citrus species, only two full-length genomes, one each from Sweet orange and Rangpur lime, are available in publicly accessible databases. In order to obtain a better understanding of the genetic variability of the virus in other citrus mosaic-affected citrus species, we performed the cloning and sequence analysis of complete genomes of CMBV from two additional citrus species, Acid lime and Pummelo. We show that CMBV genomes from the two hosts share high homology with previously reported CMBV sequences and hence conclude that the new isolates represent variants of the virus present in these species. Based on in silico sequence analysis, we predict the possible function of the protein encoded by one of the five ORFs.

A comparison of four DNA extraction methods for the detection of Citrus yellow mosaic badna virus from two species of citrus using PCR and dot-blot hybridization.

Nucleic acid preparations extracted using four procedures were assessed to determine the suitability of the procedure for PCR-based and DNA dot-blot-based detection of Citrus yellow mosaic badna virus (CMBV) from two citrus species, acid lime and pummelo. It was found that the success of PCR detection depended upon the procedure of DNA extraction whereas the dot-blot detection was successful with all extraction methods examined. CMBV DNA sequences amplified from two citrus species indicated high nucleotide sequence identity to the sequences reported previously from sweet orange. These results will help in choosing the correct DNA extraction procedure to be followed for efficient virus screening of citrus propagules.

Analysis of full-length sequences of two Citrus yellow mosaic badnavirus isolates infecting Citrus jambhiri (Rough Lemon) and Citrus sinensis L. Osbeck (Sweet Orange) from a nursery in India

Citrus yellow mosaic badna virus (CMBV), a member of the Family Caulimoviridae, Genus Badnavirus is the causative agent of mosaic disease among Citrus species in southern India. Despite its reported prevalence in several citrus species, complete information on clear functional genomics or functional information of full-length genomes from all the CMBV isolates infecting citrus species are not available in publicly accessible databases. CMBV isolates from Rough Lemon and Sweet Orange collected from a nursery were cloned and sequenced. The analysis revealed high sequence homology of the two CMBV isolates with previously reported CMBV sequences implying that they represent new variants. Based on computational analysis of the predicted secondary structures, the possible functions of some CMBV proteins have been analyzed.

Development of loop-mediated isothermal amplification and SYBR green real-time PCR methods for the detection of Citrus yellow mosaic badnavirus in citrus species

Citrus yellow mosaic badnavirus (CMBV) is an important pathogen in southern India spread by infected citrus propagules. One of the measures to arrest the spread of CMBV is to develop methods to screen and certify citrus propagules as CMBV-free. The methods loop-mediated isothermal amplification (LAMP) and SYBR green real-time PCR (SGRTPCR) have been developed for the efficient detection of CMBV in citrus propagules. This paper compares the sensitivities of LAMP and SGRTPCR with polymerase chain reaction (PCR) for the detection of CMBV. Whereas PCR and LAMP were able to detect CMBV from a minimum of 10 ng of total DNA of infected leaf samples, SGRTPCR could detect the same from 1 ng of total DNA. Using SGRTPCR, the viral titres were estimated to be the highest in rough lemon and lowest in Nagpur Mandarin of the five naturally infected citrus species tested. The results will help in designing suitable strategies for the sensitive detection of CMBV from citrus propagules.

Association of a potyvirus with mosaic disease of gherkin (Cucumis anguria L.) in India

A virus associated with severe mosaic disease of gherkin (Cucumis anguria L.) in south India was identified. The infected plants showed mosaic, vein banding, blistering on malformed leaves and fruits. Host range, transmission, serological and electron microscopic studies were carried out to identify the virus. The virus was readily transmitted by Sap inoculation and by aphids in a non-persistent manner. The host range of the virus was mainly limited to cucurbitaceous and chenopodium species. The virus showed positive serological relationships with members of potyvirus genus but not with cucumo, ilar and taspoviruses. Electron microscopy of leaf dip preparation of infected leaves revealed long flexuous filamentous virus particles measuring 750 × 12 nm. On the basis of symptomotology, host range, transmission, serology and particle morphology the virus associated with mosaic disease of gherkin might be the member of potyvirus genus.

First report of Zucchini yellow mosaic virus infecting Gherkin (Cucumis anguira) in India.

A field visit in September 2011 to the Cucumis anguira (Gherkin) growing regions of Kuppam, Chittoor district of Andhra Pradesh, India revealed occurrence of mosaic, blistering and fruit malformation leading to the crop losses. Analysis of field samples revealed association of Zucchini yellow mosaic virus (ZYMV) with the disease. This is the first confirmed report of natural occurrence of ZYMV on Gherkin in India.

Small RNA-based interactions between rice and the viruses which cause the tungro disease

Rice tungro disease is caused by a complex of two viruses, Rice tungro bacilliform virus (RTBV) and Rice tungro spherical virus (RTSV). To examine the RNAi-based defence response in rice during tungro disease, we characterized the virus-derived small RNAs and miRNAs by Deep Sequencing. We found that, while 21 nt/22 nt (nucleotide) siRNAs are predominantly produced in a continuous, overlapping and asymmetrical manner from RTBV, siRNA accumulation from RTSV were negligible. Additionally, 54 previously known miRNAs from rice, predicted to be regulating genes involved in plant defence, hormone signaling and developmental pathways were differentially expressed in the infected samples, compared to the healthy ones. This is the first study of sRNA profile of tungro virus complex from infected rice plants. The biased response of the host antiviral machinery against the two viruses and the differentially-expressed miRNAs are novel observations, which entail further studies.