Efficacy of Caprini risk assessment model in predicting venous thromboembolism risks among Asian surgical patients.

INTRODUCTION: Caprini risk assessment model (RAM) has been validated in Caucasians but evidence of its suitability in Asian surgical patients is still unknown. This study aims to determine the efficacy of Caprini model in venous thromboembolism (VTE) risk assessment among Asian surgical patients. MATERIALS AND METHODS: Consecutive surgical patients with Asian ethnicities admitted to a tertiary public hospital between January 2013 and December 2014, were included. Their demographic details, VTE risk factors and scores based on Caprini RAM were recorded. Primary outcome of this study was symptomatic VTE within 90 days of hospitalisation. Fisher's exact test and Lasso regression were performed for statistical analysis. RESULTS: A total of 4206 patients were included in this study. Distribution of this study population by risk level was very low, 14.7%; low, 44.1%; moderate, 25.6% and high, 15.7%. The overall symptomatic VTE incidence within 90 days was 0.5%. The incidence of deep venous thrombosis (DVT), pulmonary embolism (PE) and both was 0.31%, 0.19% and 0.05% respectively. VTE incidence by risk category was very low, 0%; low, 0.16%; moderate, 0.37% and high, 2.12%. Obesity (BMI >25), history of prior major surgery, history of DVT/PE and high-risk category (scores ≥5) were significant VTE factors with odds ratio > 5.0. Following the Caprini RAM with ACCP preventive recommendations, an estimated 85% of surgical patients would need prophylaxis. CONCLUSION: The overall VTE incidence among Asian surgical patients is low. Prophylaxis using Caprini RAM may subject a low incidence patient population to over utilisation of thromboprophylaxis and therefore not cost-effective when applied to Asian patients.

A simplified strategy for studying the etiology of viral diseases: Apple stem grooving virus as a case study.

A simple method to amplify infective, complete genomes of single stranded RNA viruses by long distance PCR (LD PCR) from woody plant tissues is described in detail. The present protocol eliminates partial purification of viral particles and the amplification is achieved in three steps: (i) easy preparation of template RNA by incorporating a pre processing step before loading onto the column (ii) reverse transcription by AMV or Superscript reverse transcriptase and (iii) amplification of cDNA by LD PCR using LA or Protoscript Taq DNA polymerase. Incorporation of a preprocessing step helped to isolate consistent quality RNA from recalcitrant woody tissues such as apple, which was critical for efficient amplification of the complete genomes of Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV) and Apple chlorotic leaf spot virus (ACLSV). Complete genome of ASGV was cloned under T7 RNA polymerase promoter and was confirmed to be infectious through transcript inoculation producing symptoms similar to the wild type virus. This is the first report for the largest RNA virus genome amplified by PCR from total nucleic acid extracts of woody plant tissues.

Genomic sequence analysis of four new chrysanthemum virus B isolates: evidence of RNA recombination.

Chrysanthemums worldwide suffer from a high incidence of infection with chrysanthemum virus B (CVB), a member of the genus Carlavirus, family Betaflexiviridae. Three major lineages or strains of this virus have been found in India, but none have been characterized beyond the genetic variation they display in their coat protein genes. Here, we describe the analysis of four near-complete genome sequences (from the three lineages) representing the genetic diversity of these strains. Ranging in size from 8815 to 8855 nucleotides (excluding the polyA tail), these four isolates have a genome organization very similar to that of the recently reported Japanese isolate of CVB, with which they share between 70 and 73% genome-wide sequence identity. We present further evidence that recombination may feature quite prominently in the evolution of CVB.

In Vitro Expression and Production of Antibody Against Cymbidium mosaic virus Coat Protein.

Polyclonal rabbit antisera were produced using coat protein of Cymbidium mosaic virus (CymMV) Indian isolate expressed in E. coli as GST fusion. The expressed protein was purified by GST-fusion protein purification kit for use as an immunogen in rabbits. Antisera prepared in this manner reacted in double antibody sandwich enzyme-linked immunosorbent assay (DAS ELISA) with extract from CymMV-infected tissue. The results indicate that polyclonal antisera prepared from expressed CymMV coat proteins were useful for the detection of CymMV in an array of assays. The detection system developed is highly effective for detection of Indian strain of the virus in comparison to kits available in the international market.

Determination of major viral and sub viral pathogens incidence in apple orchards in himachal pradesh.

Apple is the major commercial horticulture crop in Himachal Pradesh and other hill states of Jammu & Kashmir, Uttarakhand and some parts of Northeastern states of India. In order to gather data on health status and incidence of virus and virus-like pathogens in apple orchards, survey was conducted in the month of June and September, 2010 in Hatkoti, Rohru, Kuthara, Jubbal and Khadapathar areas of major apple producing Shimla district of Himachal Pradesh. A total of 250 samples were collected and analyzed by DAS-ELISA, NASH and RT-PCR. NASH results indicated that a total of 117 samples were infected with Apple chlorotic leaf spot virus (ACLSV), Apple mosaic virus (ApMV), Apple stem grooving virus (ASGV), Apple stem pitting virus (ASPV) and Apple scar skin viroid (ASSVd). Results showed that ASSVd is predominant in these areas with highest infection rate of 27.6% followed by ASPV (17.2%), ACLSV (16.8%), ApMV (15.2%) and ASGV (12%). Mixed infection of these viruses and viroid was frequently detected in apple trees in Himachal Pradesh. The trees, which were positive for viruses and viroids, showed a variety of fruit deformation and rusting symptoms besides leaf deformation, mosaic and chlorosis.

Velvet bean severe mosaic virus: a distinct begomovirus species causing severe mosaic in Mucuna pruriens (L.) DC.

Velvet bean [Mucuna pruriens (L.) DC] is one of the most important medicinal plants. It is used to treat many ailments, but is widely used for the treatment especially for Parkinson's disease because of the presence of 3,4-dihydroxyphenylalanine (L-dopa) in it. It was noticed in last 5 years that the plants in the field showed severe mosaic, downward curling of the leaves, stunting, etc. This is consistently observed over the years in India. The disease was transmitted by whiteflies and by grafting and the causal agent was found to be a bipartite begomovirus. The whole genome was amplified by rolling circle amplification (RCA) using ϕ-29 DNA polymerase and characterized. DNA-A and DNA-B shared a 124-nucleotide (nt) long highly conserved (98%) common region (CR). Comparisons with other begomovirus showed that DNA-A sequence has highest identity (76%) with an isolate of Mungbean yellow mosaic India virus (MYMIV; AY937195) reported from India. This data suggested that the present isolate is a new species of genus Begomovirus for which the name "Velvet bean severe mosaic virus" (VbSMV) is proposed. DNA-B has a maximum sequence identity of 49% with an isolate of Horsegram yellow mosaic virus (HgYMV; AM932426) reported from India. Infectious clones consisting of a 1.7 mer partial tandem repeat of DNA-A and a dimer of DNB-B were constructed and agro-inoculated to Macuna pruriens (L.) DC plants, which showed field observed symptoms 24 days post-infiltration (dpi). In phylogenetic analysis, DNA-A and DNA-B of the present isolate grouped with DNA-A of different begomoviruses reported from fabaceous crops. The study presents first ever molecular evidence of any disease in velvet bean and whole genome analysis of the causative virus which is a distinct bipartite species of Begomovirus.

Diversity of Apple mosaic virus Isolates in India Based on Coat Protein and Movement Protein Genes.

Apple mosaic virus (ApMV), an Ilarvirus is one of the most common pathogens of apple worldwide. During field surveys in commercial plantations of Himachal Pradesh and Jammu & Kashmir, observations of bright chlorotic mosaic like symptoms on apple trees indicated probable infection by the virus, which was later detected by double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA). An incidence of 24 and 28% (based on ELISA) was obtained as 6/25 and 15/53 samples from HP and J&K were positive, respectively. An amplification of approximately 700 and 850 bp was obtained for coat and movement protein genes (CP and MP), respectively. The CP was 223 amino acids in length and showed 87-99% identity when compared to 21 ApMV isolates. Whereas, MP (286 amino acids) showed 91-95% identity with other isolates. However, the gene sequences were quite conserved among Indian isolates and grouped together phylogenetically. CP of the Indian isolates showed maximum identity of 95% with Korean isolate (AY 125977) in apple and in other host these showed a maximum identity of 98% to Czech Republic pear isolate. MP showed maximum identity with Chinese isolate i.e., 95%. The diversity study will also help in analyzing variability among the isolates and also to formulate diagnostic and resistance strategies.

Complete nucleotide sequence of cherry virus A (CVA) infecting sweet cherry in India.

Cherry virus A (CVA) is a graft-transmissible member of the genus Capillovirus that infects different stone fruits. Sweet cherry (Prunus avium L; family Rosaceae) is an important deciduous temperate fruit crop in the Western Himalayan region of India. In order to determine the health status of cherry plantations and the incidence of the virus in India, cherry orchards in the states of Jammu and Kashmir (J&K) and Himachal Pradesh (H.P.) were surveyed during the months of May and September 2009. The incidence of CVA was found to be 28 and 13% from J&K and H.P., respectively, by RT-PCR. In order to characterize the virus at the molecular level, the complete genome was amplified by RT-PCR using specific primers. The amplicon of about 7.4 kb was sequenced and was found to be 7,379 bp long, with sequence specificity to CVA. The genome organization was similar to that of isolates characterized earlier, coding for two ORFs, in which ORF 2 is nested in ORF1. The complete sequence was 81 and 84% similar to that of the type isolate at the nucleotide and amino acid level, respectively, with 5' and 3' UTRs of 54 and 299 nucleotides, respectively. This is the first report of the complete nucleotide sequence of cherry virus A infecting sweet cherry in India.

Molecular variability analyses of Apple chlorotic leaf spot virus capsid protein.

The complete sequences of the coat protein (CP) gene of 26 isolates of Apple chlorotic leaf spot virus (ACLSV) from India were determined. The isolates were obtained from various pome (apple, pear and quince) and stone (plum, peach, apricot, almond and wild Himalayan cherry) fruit trees. Other previously characterized ACLSV isolates and Trichoviruses were used for comparative analysis. Indian ACLSV isolates among themselves and with isolates from elsewhere in the world shared 91-100% and 70-98% sequence identities at the amino acid and nucleotide levels, respectively. The highest degree of variability was observed in the middle portion with 9 amino acid substitutions in contrast to the N-terminal and C-terminal ends, which were maximally conserved with only 4 amino acid substitutions. In phylogenetic analysis no reasonable correlation between host species and/or geographic origin of the isolates was observed. Alignment with capsid protein genes of other Trichoviruses revealed the TaTao ACLSV peach isolate to be phylogenetically closest to Peach mosaic virus, Apricot pseudo chlorotic leaf spot virus and Cherry mottle leaf virus. Recombination analysis (RDP3 ver.2.6) done for all the available ACLSV complete CP sequences of the world and Indian isolates indicate no significant evidence of recombination. However, one recombination event among Indian ACLSV-CP isolates was detected. To the best of our knowledge, this is the first report of complete CP sequence variability study from India and also the first evidence of homologous recombination in ACLSV.

An ecological analysis of sociodemographic factors associated with the incidence of salmonellosis, shigellosis, and E. coli O157:H7 infections in US counties.

Identifying county-level sociodemographic and economic factors associated with the incidence of enteric disease may provide new insights concerning the dynamics of community transmission of these diseases as well as opportunities for prevention. We used data from the National Notifiable Diseases Surveillance System, the U.S. Census Bureau, and the Health Resources and Services Administration to conduct an ecological analysis of 26 sociodemographic and economic factors associated with the incidence of salmonellosis, shigellosis, and E. coli O157:H7 infections in US counties for the period 1993 to 2002. Our study indicates that race, ethnicity, place of residence, age, educational attainment, and poverty may affect the risk of acquiring one of these enteric bacterial diseases. The lack of specificity of information regarding salmonellae and shigellae serotypes may have led to less specific associations between community-level determinants and reported incidence of those diseases. Future ecological analyses should use serotype-specific data on incidence, which may be available from laboratory-based surveillance systems.

Coat protein gene diversity among Chrysanthemum virus B isolates from India.

The complete coat protein (CP) sequences from 29 Indian isolates of Chrysanthemum virus B (CVB) were determined and analysed in relation to other previously characterized carlaviruses. The CP genes of the Indian CVB isolates were highly heterogeneous, sharing nucleotide sequence identities of 74-98%. Based on phylogenetic analyses, the isolates formed three groups potentially representing either two or three major CVB strain groupings. Recombination analysis revealed at least one definite recombination event involving the exchange of sequences between members of different groups. To our knowledge this is the first reported evidence of homologous recombination in carlaviruses.

Wild Himalayan Cherry (Prunus cerasoides) as a Natural Host of Prunus necrotic ringspot virus in India.

Prunus cerasoides, also known as wild Himalayan cherry, grows naturally in the Himalayas. A member of the Rosaceae family, the tree has medicinal (astringent) and other (beads, dye, wood) uses. During surveys in the northwestern Himalayan Region of India, necrotic spots were observed on leaves of P. cerasoides. Since the symptoms were typical of Prunus necrotic ringspot virus (PNRSV), preliminary detection was done by double-antibody sandwich (DAS)-ELISA (Agdia, Elkhart, IN). Positive results were obtained three times more than the negative control which is provided in the kit. To further confirm its presence, reverse transcription (RT)-PCR analysis was performed using a primer pair (upstream 5'-AACTGCAGATGGTTTGCCGAATTTGCAA-3'; downstream 5'-GCTCTAGACTAGATCTCAAGCAGGTC-3') specific for the coat protein gene (GenBank Accession Nos. AJ619984 and AJ619983). Amplification of the expected 675-bp fragment was obtained. The sequence of a cloned copy of the amplified product showed 99% similarity to the PNRSV coat protein gene (GenBank Accession No. AF170165), confirming the presence of PNRSV in P. cerasoides (sequence submitted as Accession No. AM493717). The cloned DNA has the potential for utilization as an additional tool, and an early PNRSV screening (both pollen and seed transmitted) will be highly useful to ensure healthy rootstocks are used for grafting purposes (1). PNRSV mainly infects members of the Rosaceae family, including stone fruits and ornamental plants such as peach, plum, cherry, apricot, nectarines, and roses, and was first reported in P. persica (1). Proper management of PNRSV at this level can prevent its transmission and disease development in grafted scions of commercial Prunus spp. Reference: (1) A. A. Brunt et al. Page 1047 in: Viruses of Plants. CAB International, Wallingford, UK. 1996.

Coat protein sequence shows that Cucumber mosaic virus isolate from geraniums (Pelargonium spp.) belongs to subgroup II.

A viral disease was identified on geraniums (Pelargonium spp.) grown in a greenhouse at the Institute of Himalayan Bioresource Technology (IHBT), Palampur, exhibiting mild mottling and stunting. The causal virus (Cucumber mosaic virus, CMV) was identified and characterized on the basis of host range, aphid transmission, enzyme linked immunosorbent assay (ELISA), DNA-RNA hybridization and reverse transcription polymerase chain reaction (RT-PCR). A complete coat protein (CP) gene was amplified using degenerate primers and sequenced. The CP gene showed nucleotide and amino acid homology up to 97%-98% and 96%-99%, respectively with the sequences of CMV subgroup II. The CP gene also showed homologies of 75%-97% in nucleotide and 77%-96% in amino acid with the CMV Indian isolates infecting various crops. On the basis of sequence homology, it was concluded that CMV-infecting geraniums in India belong to subgroup II.

Pneumatic retinopexy: success rate and complications.

AIM: To evaluate the success rate and complications of pneumatic retinopexy performed at a university hospital and to identify which patients are best suited for pneumatic retinopexy. METHODS: This was an interventional case series. Retrospective review of 61 patients who had pneumatic retinopexy performed by two retina surgeons at two University of California, San Francisco hospitals between 1998 and 2004. Patients who had been treated for rhegmatogenous retinal detachment (RRD) with pneumatic retinopexy were identified by reviewing operative reports and billing records. The primary outcome measure was anatomical reattachment of the retina with a single intervention. Secondary outcome measures included postoperative visual acuity and postoperative complications. RESULTS: 33 of 61 (54%) cases were successful with a single procedure. 40 of 61 (66%) cases were successful with repeat injection of gas or laser retinopexy alone. All cases had anatomical success at final follow up. Age, myopia, lens status, and number of breaks were not proved to be risk factors for failure. The average duration of follow up was 15 months. CONCLUSIONS: In this case series, pneumatic retinopexy was less effective for the repair of RRD than most large published reports. However, failure of pneumatic retinopexy followed by scleral buckle or pars plana vitrectomy did not negatively influence visual acuity at final follow up.

Plant parasitic and vector nematodes associated with Asiatic and Oriental hybrid lilies.

A survey for distribution and abundance of plant parasitic nematodes in fields grown to Lilium in Himachal Pradesh, India at four study sites viz. Nagrota (at 810 m a.s.l.), Palampur (at 1270 m a.s.l.), Sunder Nagar (at 1400 m a.s.l.) and Chail (at 2250 m a.s.l.) was carried out. Moderate (101-500/200 ml soil) to high (501-1000/200 ml soil) populations of phytonematodes including the vectors for plant viruses (Aphelenchoides avenae, Criconemoides spp., Hoplolaimus spp., Longidorus spp., Paratylenchus spp., Pratylenchus spp., Rhabditis spp., Trichodorus spp., Tylenchoryhnchus spp., Tylenchulus spp. and Xiphinema diversicaudatum) were recorded. Mean population of nematodes was positively correlated with pH in all the study sites, negatively correlated with electrical conductivity (EC), percent organic matter (OM%), available potassium (K) and positively correlated with percent carbon (C%), available nitrogen (N) and available phosphorus (P) in all but one study site. The highest incidence of virus-vector nematodes viz. X. diversicaudatum, Longidorus spp. and Trichodorus spp. was recorded at Palampur. Only Strawberry latent ringspot nepovirus (SLRSV) was detected in Lilium cvs. Star Gazer Max and Galeili by Enzyme Linked Immunosorbent Assay (ELISA) and reverse transcription polymerase chain reaction (RT-PCR) and in X. diversicaudatum associated with the cultivars by RT-PCR. Cucumis sativus used as bait plants showed SLRSV symptoms after 15 days of nematode inoculation and further SLRSV was again detected by ELISA and RT-PCR in C. sativus plants confirming the transmission of SLRSV by X. diversicaudatum in Lilium.

Natural Occurrence of a Potyvirus on Murraya koenigii in India.

Murraya koenigii (L.) Spreng., a small, strong-smelling umbrageous tree with subcampanulate white flowers belonging to the family Rutaceae, is native to India and southeastern Asia (2). It is distributed across the Indian subcontinent excluding the higher elevations of the Himalayas. In India, the leaves are mainly used for culinary purposes. The leaves are commonly known as curry leaves or 'sweet neem'. The whole plant including bark, root, leaves, fruits, and fruit pulp is used medicinally. This plant was reported to be a host of Citrus tristeza virus (1). In a survey of potyvirus incidence in the northwestern Himalaya foothills of the Kangra and Hamirpur districts in the state of Himachal Pradesh in 2004, M. koenigii plants showing mosaic symptoms on leaves, typical of a virus infection, were frequently observed. Symptomatic leaves were tested for the presence of several viruses using enzyme-linked immunosorbent assay with specific antibodies. Positive results were obtained with potyvirus group specific antibodies (Agdia, Elkhardt, IN) in triplicate analyses of 5 of 15 leaf samples tested. To further identify the infecting virus, RNA from plants was tested using universal potyvirus primer pair P9502 and CPUP (3) and reverse transcription-polymerase chain reaction to amplify a genome fragment encoding portions of the coat protein and the 3'UTR (3). An amplification product of the expected size (~800 bp) was obtained. The product was cloned into the pGem-T easy vector (Promega, Madison, WI), and three clones were sequenced. The sequence (GenBank Accession No. AJ852504) had 92% identity to Chili vein banding mottle virus, a potyvirus infecting pepper reported from Thailand (GenBank Accession No. U72193). To our knowledge, this is the first report of a potyvirus naturally occurring on a Murraya sp. References: (1) K. Balaram and K. Ramakrishnan. Curr. Sci. 48:453, 1979. (2) J. D. Hooker. Flora British India 1:502, 1875. (3) R. A. A. van der Vlugt et al. Phytopathology. 89:148, 1999.

Occurrence of Cucumber mosaic virus in Gerbera jamesonii in India.

Gerbera jamesonii (family Asteraceae) is a popular perennial ornamental cut flower and potted plant with considerable economic importance. In a survey of gerbera grown in floriculture fields at the Institute of Himalayan Bioresource Technology (IHBT), Palampur and nearby nurseries, color break symptoms on the petals, asymmetrical ray florets, and deformed flowers were observed during 2003-2004. The virus evoked chlorotic local lesions on Chenopodium album, C. amaranticolor, and C. quinoa, while systemic mosaic was observed on Cucumis sativus, Nicotiana benthamiana, N. clevelandii, N. glutinosa, and N. tabacum cv. Samsun. The virus was transmitted nonpersistently by Myzus persicae and Aphis gossypii and was identified as Cucumber mosaic virus (CMV) using enzyme-linked immunosorbent assay (ELISA) with CMV-specific antibodies (Agdia, Elkhart, IN). Polyhedral particles approximately 29 nm were observed with electron microscopy of leaf dips from symptomatic gerbera leaves. Total RNA was isolated from the infected gerbera plants and N. glutinosa by using RNAqueous (Ambion, Austin, TX). CMV-specific primers (1) were used to detect the virus with reverse transcription-polymerase chain reaction that produced an amplicon predicted size of approximately 540 bp, but the virus was not detected in healthy controls. Sequence alignment of the amplicons (533 bp) utilizing BLAST resulted in 91 to 99% homology with the partial intercistronic region and partial coat protein gene (1042-1574 bp) (gene sequence submitted to EMBL database with Accession no. AJ634532) of CMV RNA3 in subgroup I. To our knowledge, this is the first report of CMV on gerbera in India. Reference: (1) C. De Blas et al. J. Phytopathol. 141:323, 1994.

Strawberry latent ringspot virus Infecting Roses in India.

Rose is an economically important crop of India and the world. A survey of rose plantations in and near the Kangra Valley of Himachal Pradesh, India, showed virus-like symptoms, including yellow flecking in young leaves and reduction in leaflet size, while some were symptomless. These symptoms are similar to those for Strawberry latent ringspot virus (SLRSV) (1). Sap inoculation from symptomatic and some symptomless leaves to Chenopodium amaranticolor resulted in chlorotic local lesions followed by systemic chlorosis. SLRSV was detected in this indicator host and six rose cultivars (Happiness, Iceberg, First Prize, Ganga, Pink Panther, and Oklahoma) showing characteristic symptoms of SLRSV using enzyme-linked immunosorbent assay (ELISA) with ELISA kit (DSMZ, Braunschweig, Germany). Reverse transcription-polymerase chain reaction was performed with SLRSV-specific primers (2), and a product of the expected size of ˜181 bp was amplified. The authenticity of the fragment was confirmed by sequencing. Isolated SLRSV was also inoculated to seed-grown rose seedlings and after 20 days postinoculation the same symptoms (yellow flecking in young leaves) were observed. These results established the identity of the virus that caused yellow flecking on rose leaves in India as SLRSV. To our knowledge, this is the first report of SLRSV infecting rose in India. References: (1) A. F. Murant. Strawberry latent ringspot virus. No. 126 in: Description of Plant Viruses, CMI/AAB, Surrey, U.K., 1974. (2) E. Bertolini et al. J. Virol. Methods 96:33, 2001.