Emerging challenges in the management of Orthotospoviruses in Indian agriculture.

Orthotospoviruses, a genera of negative-sense ssRNA viruses transmitted by thrips, have gained significant attention in recent years due to their detrimental impact on diverse crops, causing substantial economic losses and posing threats to food security. Orthotospoviruses are characterised by a wide range of symptoms in plants, including chlorotic/necrotic spots, vein banding, and fruit deformation. Seven species, including four definite and three tentative species in the genus Orthotospovirus, have so far been documented on the crops of the Indian subcontinent. Management of Orthotospoviruses under field conditions is challenging since they have a wide host range, adaptation to versatile environmental conditions, a lack of promising resistance sources, and the ubiquitous nature of thrips and their transmission through a propagative manner. Our present review elucidates the significance, molecular biology and evolutionary relationship of Orthotospoviruses; vector population; and possible management strategies for Orthotospoviruses and their vectors in the scenario of the Indian subcontinent.

A distinct strain of tomato leaf curl New Delhi virus that causes mosaic disease in ash gourd and other cucurbitaceous crops.

Ash gourd (Benincasa hispida) is a cucurbitaceous crop cultivated as an edible vegetable rich in vitamins, minerals, dietary fibers and antioxidants. In a field survey conducted in the Udumalpet region of Tamil Nadu during 2019, the incidence of mosaic disease on ash gourd crop was observed to be 75%. The DNA-A and DNA-B components of begomovirus genome have been identified as associated with this disease. Both the cloned DNA-A and DNA-B genomic components shared highest pairwise sequence identities with the isolates of tomato leaf curl New Delhi virus (ToLCNDV), a bipartite begomovirus. Recombinant analysis showed that both the components are possibly evolved through intra-species recombination between ToLCNDV isolates. Tomato leaf curl Bangladesh betasatellite (ToLCBB) is not naturally associated with this sample. The results of infectivity studies on ash gourd and other cucurbitaceous crops demonstrates the Koch's postulates, when co-inoculation of DNA-A and DNA-B of ToLCNDV was undertaken. However, the inoculation of non-cognate ToLCBB along with DNA-A and DNA-B enhances the symptom expression and reduces the time taken for symptom development. Thus, Koch's postulates were proved for these virus complexes on cucurbitaceous crops. Furthermore, an enhanced accumulation of DNA-A component was detected in the cucurbits co-inoculated with ToLCNDV and ToLCBB. This report highlights the importance of investigating the spread of these disease complexes with other cucurbitaceous crops in India.

Production of polyclonal antibodies for Capsicum chlorosis virus (CaCV) infecting chilli in India through recombinant nucleocapsid protein expression and its application.

Bud necrosis and chlorotic spots causing virus affecting chilli crop in Tamil Nadu (India) was identified as Capsicum chlorosis virus (CaCV). Specific primers were used for amplification and sequencing of the nucleocapsid protein (NP) gene. Polyclonal antibody against the bacterially expressed NP from the CaCV-TN-CBE isolate was produced using recombinant DNA technology. NP gene was subcloned into the pET-28a (+) vector and expressed by transformation in BL21 (DE3) pLysS. The expressed protein was about ∼34 kDa and was confirmed through western blot analysis using Groundnut bud necrosis virus (GBNV) polyclonal antiserum from ICRISAT, India. The purified recombinant protein was used to immunize rabbits to generate CaCV-specific polyclonal antiserum. The sensitivity levels of polyclonal antiserum thus raised was assayed through indirect ELISA or direct antigen coating (DAC)-ELISA using the recombinant protein as antigen. The recombinant antiserum produced in this study successfully detected the natural infection of CaCV on chilli plants collected from the field as well as on cowpea plants artificially inoculated with CaCV by using DAC-ELISA, DIBA and western blotting.

First report of lily mottle virus on lily (Lilium sp.) in southern India.

Lily samples collected from commercial cut flower growing field at Nilgiris, Tamil Nadu, India were serologically tested through direct antigen coating-ELISA and dot immuno binding assay for the infection of potyvirus through potyvirus group specific polyclonal antibody. The RT-PCR using potyvirus universal degenerate primers (PNIbF1/PCPR1) showed the presence of a potyvirus and the sequencing results showed the association of lily mottle virus (LMoV). Further LMoV was confirmed by RT-PCR amplification using newly designed primer pair covering entire coat protein gene and sequence results showed 100% nucleotide identity with the LMoV reported from South Korean. To our knowledge, this is the first confirmed evidence for the occurrence of LMoV on lily in southern India.

Molecular evidence for the occurrence of tomato leaf curl New Delhi virus on chayote (Sechium edule) in southern India.

During 2012-2014, mosaic disease on chayote in the farmers field of Kodaikanal region (high altitude zone) of Tamil Nadu was observed. The disease was characterized with severe mosaic, cupping and enation on leaves with reduced fruit size. Disease was found to causes an yield loss of more than 60% with the maximum disease incidence of 100% for the past 5 years consecutively. Preliminary serological and molecular screening indicated the association of begomovirus with the disease. Complete nucleotide sequence and phylogenetic analysis of DNA A revealed the identity of the virus as tomato leaf curl New Delhi virus (ToLCNDV). In recombination analysis study, the major parent was identified as ToLCNDV from Pakistan infecting tomato. Thus the present finding confirms expansion of new geographical region and host for ToLCNDV causing mosaic disease on chayote from Tamil Nadu. To our knowledge this is the first confirmed report for the occurrence of ToLCNDV on chayote in southern India.

Molecular characterization of a distinct bipartite Begomovirus species infecting ivy gourd (Coccinia grandis L.) in Tamil Nadu, India.

A distinct bipartite begomovirus was found to be associated with the mosaic disease on ivy gourd (Coccinia grandis L.) in Tamil Nadu, India. The complete DNA A and DNA B components were cloned by rolling circle amplification. Genome organization of this virus is found to be typical of Old World bipartite begomovirus. The association of betasatellite component with this virus is absent. The closest nucleotide identity of 73.4 % was seen with the Loofa yellow mosaic virus (LYMV-[VN]-AF509739) suggesting that it is a new virus species Coccinia mosaic virus (CoMoV-Ivy gourd [TN TDV Coc1]) and distantly related to the other known begomoviruses. The DNA B component shared a maximum identity of 55 % with that of Tomato leaf curl New Delhi virus (ToLCNDV). In the phylogenetic analysis, CoMoV-Ivy gourd form cluster separate from other begomoviruses. Recombination analysis showed that there was no recombination event in the genome. This is the distinct begomovirus infecting ivy gourd.

Pathogenesis and recovery of tetraplegia after electrical injury.

The site of neurological damage causing paralysis after electrical trauma remains to be clarified. A patient is described who developed a flaccid tetraplegia after a high voltage electrical injury. The findings on initial examination and neurophysiological investigation showed a very severe generalised sensory-motor polyneuropathy. His subsequent follow up over 60 months showed a remarkable degree of reinnervation and the unmasking of a myelopathy. The degree of reinnervation noted suggests an axonopathy that left the other elements of the peripheral nerves relatively spared. These findings provide the most convincing evidence to date that a generalised polyneuropathy can follow electrical injury and that it results from non-thermal mechanisms such as electroporation.

Patterns of neurophysiological abnormality in prolonged critical illness.

OBJECTIVE: To describe the various patterns of neurophysiological abnormalities which may complicate prolonged critical illness and identify possible aetiological factors. DESIGN: Prospective case series of neurophysiological studies, severity of illness scores, organ failures, drug therapy and hospital outcome. Some patients also had muscle biopsies. SETTING: General intensive care unit (ICU) in a University Hospital. PATIENTS: Forty-four patients requiring intensive care unit stay of more than 7 days. The median age was 60 (range 27-84 years), APACHE II score 19 (range 8-33), organ failures 3 (range 1-6), and mortality was 23%. RESULTS: Seven patients had normal neurophysiology (group I), 4 had a predominantly sensory axonal neuropathy (group II), 11 had motor syndromes characterised by markedly reduced compound muscle action potentials and sensory action potentials in the normal range (group III) and 19 had combinations of motor and sensory abnormalities (group IV). Three patients had abnormal studies but could not be classified into the above groups (group V). All patients had normal nerve conduction velocities. Electromyography revealed evidence of denervation in five patients in group III and five in group IV. There was no obvious relationship between the pattern of neurophysiological abnormality and the APACHE II score, organ failure score, the presence of sepsis or the administration of muscle relaxants and steroids. A wide range of histological abnormalities was seen in the 24 patients who had a muscle biopsy; there was no clear relationship between these changes and the neurophysiological abnormalities, although histologically normal muscle was only found in patients with normal neurophysiology. Only three of the eight patients from group III in whom muscle biopsy was performed had histological changes compatible with myopathy. CONCLUSIONS: Neurophysiological abnormalities complicating critical illness can be broadly divided into three types -- sensory abnormalities alone, a pure motor syndrome and a mixed motor and sensory disturbance. The motor syndrome could be explained by an abnormality in the most distal portion of the motor axon, at the neuromuscular junction or the motor end plate and, in some cases, by inexcitable muscle membranes or extreme loss of muscle bulk. The mixed motor and sensory disturbance which is characteristic of 'critical illness polyneuropathy' could be explained by a combination of the pure motor syndrome and the mild sensory neuropathy. More precise identification of the various neurophysiological abnormalities and aetiological factors may lead to further insights into the causes of neuromuscular weakness in the critically ill and ultimately to measures for their prevention and treatment.

Isotypes of naturally acquired antibodies to a repetitive & non-repetitive epitope on Plasmodium falciparum surface proteins in an endemic area of Sri Lanka.

Isotypes of antibodies in adults and 7-15 yr old children living in a malaria endemic area of Sri Lanka were measured by radioimmunoassay against synthetic target antigens derived from two Plasmodium falciparum surface proteins. Greater than 50 per cent of the sample population possessed IgM antibodies while < or = 13 per cent developed each of the IgG antibody isotypes against a repetitive epitope present on the circumsporozoite protein (CS repeat). A more even distribution of IgM, IgG1, IgG2, IgG3 and IgG4 antibodies was seen against a non-repetitive epitope (P103) on a 45 kDa merozoite surface protein. This difference is attributed to a T-independent antibody response against the CS repeat.

Dynamics of natural antibody responses to malaria parasite surface proteins in the intermediate rainfall zone of Sri Lanka.

Antibodies against repetitive epitopes on Plasmodium falciparum and P. vivax circumsporozoite (CS) proteins and epitopes on the 45 kDa and 185-200 kDa P. falciparum merozoite surface proteins were measured by radioimmunoassay in a two year longitudinal study in Nikawehera village located in the intermediate rainfall zone of Sri Lanka. The prevalence and concentrations of specific antibodies were in many, but not all instances, greater in adults than in children who were aged 7-15 yr at the beginning of the study. The concentrations and prevalence of antibodies were associated with malaria transmission levels previously determined from entomological and hospital admission data in the area. Antibody responses to epitopes on different P. falciparum antigens, two different epitopes within the 185-200 kDa merozoite surface protein and between the P. falciparum and P. vivax CS repeats were significantly correlated. Antibody concentrations against a conserved epitope in the 185-200 kDa protein were significantly higher in P. falciparum infected individuals than in non-parasitaemic subjects. Antibody concentration and prevalences in Nikawehera were lower than at Weheragala, a site located 70 km away in the dry zone of Sri Lanka. It is postulated that lower levels of immunity in the population in areas such as Nikawehera, that are adjacent to more highly malaria endemic areas, may promote epidemics when conditions favour transmission.

Antibody and clinical responses in volunteers to immunization with malaria peptide-diptheria toxoid conjugates.

Twenty residue peptides from the 185-200-kD and 45-kD merozoite surface antigens of the malaria parasite Plasmodium falciparum were covalently linked to diphtheria toxoid as a carrier and used to immunize human volunteers with aluminium hydroxide as an adjuvant. Significant antibody levels were elicited by two boosting injections. The antibodies reacted with acetone-methanol fixed merozoite membranes in an immunofluorescence assay, but no inhibition of merozoite reinvasion could be detected in in vitro cultures containing the antibodies. Antibody levels against the immunizing peptides declined markedly within 77 days after the third injection. No hypersensitivity was observed against the peptides. However, the volunteers developed hypersensitivity against diphteria toxoid, and in particular a pronounced type III (Arthus) hypersensitivity after three injections with the toxoid. This effect might appear to limit the use of peptide-diphtheria toxoid conjugates for human immunization. Several biochemical, haematological and immunological tests done on the volunteers showed no other adverse effects from the immunizations.

Serial electroencephalograms in a patient with D-lactic acidosis.

A patient with previous bowel surgery was followed through the course of her recurrent encephalopathy with biochemically demonstrated D-lactic acidosis by detailed serial electroencephalography (EEG). Although the EEG changes were marked and parallelled the clinical and biochemical abnormalities closely, they were essentially non-specific. This diagnosis should be considered in encephalopathic patients with a history of bowel disease particularly after extensive small bowel resection.

Neurological illness following treatment with fludarabine.

Fludarabine is a comparatively new drug for the treatment of low-grade lymphoid malignancy. This report describes five cases of unusual neurological illnesses occurring after treatment with fludarabine. These suggest that caution should be exercised in patients receiving fludarabine who develop neurological abnormalities, with prompt investigation and if necessary cessation of the drug.

Antibodies to epitopes on merozoite and sporozoite surface antigens as serologic markers of malaria transmission: studies at a site in the dry zone of Sri Lanka.

Antibodies against repetitive epitopes on Plasmodium falciparum and P. vivax circumsporozoite (CS) proteins and epitopes on the 45-kD and 185-200-kD P. falciparum merozoite surface antigens were measured by radioimmunoassay in Weheragala, a malaria-endemic site in the dry zone of Sri Lanka. Antibodies were measured in sera collected in February at the end of the main malaria transmission season and three months later in May during the low transmission period. Ninety-seven percent of the sample population had antibodies to the P. falciparum CS repeat in February and a significant proportion possessed antibodies directed against all epitopes tested. Concentrations and prevalence of antibodies to the CS repeats decreased with time after the end of malaria transmission in adults and children. Similar temporal changes were observed with antibodies to the epitopes on merozoite surface antigens. Children 7-15 years of age had lower antibody concentrations against most epitopes than adults. Antibody concentrations to two different epitopes within the same merozoite surface antigen showed significant association as did antibody levels against the P. falciparum CS repeat and the predominant P. vivax CS repeat. However, antibody concentrations did not correlate with the presence of blood-stage malaria infections.