Citrus tristeza virus (CTV), a member of the aphid-transmitted closterovirus group, is the causal agent of the notorious tristeza disease in several citrus species worldwide. The codon usage patterns of viruses reflect the evolutionary changes for optimization of their survival and adaptation in their fitness to the external environment and the hosts. The codon usage adaptation of CTV to specific citrus hosts remains to be studied; thus, its role in CTV evolution is not clearly comprehended. Therefore, to better explain the host–virus interaction and evolutionary history of CTV, the codon usage patterns of the coat protein (CP) genes of 122 CTV isolates originating from three economically important citrus hosts (55 isolate from Citrus sinensis, 38 from C. reticulata, and 29 from C. aurantifolia) were studied using several codon usage indices and multivariate statistical methods. The present study shows that CTV displays low codon usage bias (CUB) and higher genomic stability. Neutrality plot and relative synonymous codon usage analyses revealed that the overall influence of natural selection was more profound than that of mutation pressure in shaping the CUB of CTV. The contribution of high-frequency codon analysis and codon adaptation index value show that CTV has host-specific codon usage patterns, resulting in higheradaptability of CTV isolates originating from C. reticulata (Cr-CTV), and low adaptability in the isolates originating from C. aurantifolia (Ca-CTV) and C. sinensis (Cs-CTV). The combination of codon analysis of CTV with citrus genealogy suggests that CTV evolved in C. reticulata or other Citrus progenitors. The outcome of the study enhances the understanding of the factors involved in viral adaptation, evolution, and fitness toward their hosts. This information will definitely help devise better management strategies of CTV.

Chilli leaf curl virus (ChiLCV; genus: Begomovirus), transmitted by Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) in a persistent-circulative manner, is a major constraint in chilli production. The present study demonstrates for the first time that a topical spray of naked double-stranded RNA (dsRNA) on chilli plants causes mortality and inability to acquire and transmit ChiLCV in B. tabaci. dsRNA targeting heat shock protein 70 (hsp70) and fasciclin 2 (fas2) of B. tabaci Asia II 1 was first assessed under controlled conditions through oral delivery. Hsp70 and fas2 dsRNA resulted in up to 82.22% and 72% mortality of B. tabaci and around 12.4- and 8.5-fold decreases in mRNA levels, respectively, 24 h post-ingestion. ChiLCV copies in hsp70 dsRNA-fed B. tabaci steadily decreased with an increase in dsRNA concentration and were undetectable at a higher concentration of dsRNA. However, ChiLCV copies significantly increased in fas2 dsRNA-fed B. tabaci. Transmission of ChiLCV by B. tabaci was completely inhibited post-24 h feeding on hsp70 dsRNA at 3 μg/mL. Naked hsp70 dsRNA was topically sprayed on ChiLCV-infected chilli plants like an insecticide. 67.77% mortality of B. tabaci, 4.6-fold downregulation of hsp70 mRNA, and 1.34 × 10¹⁵-fold decreased ChiLCV copies in B. tabaci were recorded when adults were exposed to the dsRNA-treated plants under semi-field conditions. Foliar application of naked dsRNA reduced the ChiLCV transmission by 75% without any visible symptoms in the inoculated plants. A total of 2 consecutive sprays of dsRNA provided significant protection to B. tabaci for up to 20 days under semi-field conditions.

About 25 samples of infected bottle gourd were collected from an area of 5 km radius of Cooch behar district of West Bengal. All the samples were subjected to DAC-ELISA and RT-PCR. Out of which, amplicon of one positive sample was sequenced. A single ORF of 435 nucleotides (GenBank Accession No. KJ481802) of the isolated virus showed highest identity (94%) with SMV, which led us to conclude that bottle gourd is a new host of the virus which was not reported earlier.

‘Pointed gourd’ or ‘parwal’ (Trichosanthes dioica) is a member of the cucurbit family and an important vegetable crop in India which is also widely grown in West Bengal. Fungal diseases are a major constraint to cultivation of pointed gourd. Yellow to dark brown spots were observed on leaves of pointed gourd in different fields in West Bengal during June–August 2015. In extreme cases, plants showed necrosis during early stages of growth and did not bear any fruit. Symptomatic leaves were collected for fungal isolation and colonies that produced an olive-grey mycelial mat on potato dextrose agar were obtained. Ascendant or erect conidiophores with dark brown pigmentation and light brown, oblong to cylindrical conidia with 2–3 transverse septa were visible using a light microscope. The fungus was identified as Curvularia spicifera (anamorph: Bipolaris spicifera) on the basis of morphological characteristics. Pathogenicity tests on detached leaves and whole plants confirmed that the isolates were pathogenic. The ITS and 28S rRNA large subunit regions of ribosomal DNA were amplified with two different primer sets for molecular identification of the fungus. The amplicons were cloned and the sequences submitted to GenBank (accession nos. KX910098 and KY411823). Sequence similarities confirmed the organism as C. spicifera. This is the first report of C. spicifera as a pathogen of pointed gourd.

Pointed gourd (Trichosanthes dioica Roxb.), of the Cucurbitaceae family, is widely cultivated as a vegetable in many countries such as Bangladesh, India, Pakistan, Myanmar, Nepal and Sri Lanka. Over 800,000 metric tons of pointed gourds are produced annually in India, where cultivation is estimated to occupy over 33,000 hectares of land (MoA & FW, Government of India). In summer 2018, significant losses (approximately 15-20%) occurred in the sub-Himalayan region in West Bengal state of India (21.14-21.30° N, 78.82-79.02°E) due to a disease with typical anthracnose-like symptoms on the fruits. Light yellowish, small sized round to irregular spots were also apparent on the leaves. These spots gradually increased in size and turned into light brown and were surrounded by yellow halo. The lesions on the fruits were circular, yellow-brown, necrotic and sunken. A survey of four fields (1.5 ha) was conducted and a disease incidence of 30-40% was observed. Necrotic tissues from fruit as well as leaves were cut into approximately 5 mm2, surface sterilized with 0.1% HgCl2, plated in potato dextrose agar and incubated at 28ºC for 7 days in the dark. A total of 50 morphologically similar colonies were obtained from 20 sampled fruits and 10 sampled leaves. Fungal colonies were initially white, becoming gray as the cultures aged on PDA. The cultures developed black acervuli around the center of the colony. Setae were brown in colour, 1-5 septate, 40-100 µm long. Conidia were also observed through light and scanning electron microscopy and exhibit as (4-6 ×13-19 µm) hyaline, aseptate, cylindrical to oblong, with one end round and other truncate. The morphological characteristics were found similar to Colletotrichum orbiculare Damm, P.F. Cannon & Crous as reported by Damm et al. (2013). Ten isolates were obtained by transferring hyphal tips to new PDA plates and incubating under the same conditions. To confirm the identity of the pathogen, genomic DNA was extracted from five pure isolates (PG-Pha, PG-Pha-2, PG-Pha-3, PG-Pha-4, PG-Pha-5) with the cetyltrimethylammonium bromide (CTAB). Further, the ITS1-5.8S-ITS2 region, D1/D2 region of the 28S rRNA large subunit (LSU), Actin (ACT) gene and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene were amplified using specific primers, ITS1/ITS4 (White et al. 1990), NL1/NL4, ACT1/ACT2 and GDF1/GDR1 respectively and PCR conditions described in Damm et al. (2012). A GenBank BLAST search showed 99-100% identity to the Colletotrichum orbiculare (Acc. Nos. KP898988 for ITS1-5.8S-ITS2, Z18997 for 28S rRNA, AB778553 for ACT gene, and KF178482 for GAPDH). All obtained sequences were submitted to the GenBank (Acc. Nos. MN006616, OP811046-OP811049, [ITS1-5.8-ITS2], MN006684, PP391616-PP391619 [28S rRNA], MN168524, PP400822-PP400825 [ACT gene], OP627091, PP400826-PP400829 [GAPDH]). For phylogenetic analysis, MEGA version 11 (Tamura et al. 2021) was used to construct a maximum likelihood tree with 1000 bootstrap replicates, based on a concatenation alignment of three gene sequences (ITS, Actin and GAPDH) of the all the five C. orbiculare isolates as well as sequences of other Colletotrichum species obtained from GenBank. The cluster analysis revealed that, isolate PG-Pha form a cluster with other C. orbiculare isolates. Pathogenicity tests were conducted to confirm Koch's postulates. Pathogenicity tests were performed in mature fruits by inoculating them (n=8) with 10 µl of a 1×106 conidia/ml suspension at needle puncture wound sites. In control set up sterile distilled water was pipetted on fruits. Fruits were placed on sterile trays covered with glasses and incubated at humid chambers at 28±2ºC with 12 h of light. Healthy one-month old potted pointed gourd plants (n=15) were sprayed with conidial suspension until run-off. A set of 15 plants were sprayed with sterile distilled water and maintained as control. The plants were kept in a greenhouse at 25ºC, >75% relative humidity, and a 16/8 h day/night cycle for 15 days. Sterile distilled water was sprayed on the plants at one day interval to maintain the humidity. Inoculated fruits started showing yellowing symptoms one day post inoculation and gradually yellow-brown sunken spots became visible at the place of puncture, whereas control fruits remain symptomless even after 7 days of inoculation. Inoculated leaves showed disease symptoms similar to those observed in the field whereas leaves of control sets were symptomless even after 15 days. The pathogenicity test was repeated thrice under the same conditions mentioned before. C. orbiculare was successfully re-isolated from all the symptomatic tissues of leaves as well as fruits, completing Koch's postulates. Previously, the pathogen has been reported as an important anthracnose pathogen of Cucurbitaceae, especially of cucumber (Cucumis sativus), melons (Cucumis melo), watermelon (Citrullus lanatus), pumpkin (Cucurbita pepo) and squash (Cucurbita maxima) (Farr and Rossman 2020). To our knowledge, this is the first report of C. orbiculare causing anthracnose of pointed gourd. This disease represents a threat to producers in India and central Asia. Further research may contribute to the development of management strategies for this disease.

Pointed gourd (Trichosanthes dioica Roxb.) is a popular summer vegetable in India. During April 2017, severe leaf blight symptoms were observed in many fields located in the sub-Himalayan region in West Bengal state of India (21.14–21.30° N, 78.82–79.02°E). Lesions initially appeared as yellow spots (4-5 mm diameter) on leaf margins that gradually expanded to cover almost the whole leaf between the veins. As lesions aged, they turned brown and dry ultimately leading to defoliation. Tiny black pycnidia were scattered throughout the dried lesions. Symptomatic leaves were collected, cut into small square pieces approximately 5mm2, surface sterilized with 1% NaOCl, and transferred to potato dextrose agar. The agar plates were incubated at 28ºC for 7 days. After 5 days of incubation, compressed, white velvety mycelia appeared on the plates and the colonies gradually turned grey after sporulation. Conidia were hyaline, 2-3 septate, 4-8 μm long and 1.5-3 μm wide, thin and smooth walled, ellipsoidal to cylindrica...