Determining Theileria annulata parasitemia through qPCR in lactating cows of Odisha, - a theileriosis-endemic region of India.

Study was undertaken in a theileriosis-endemic region of India during May 2018 to April 2019 among milch cows. Blood samples collected from apparently healthy (n = 65) and Theileria-suspect cows (n = 65) were screened against T. annulata and T. orientalis infection by SYBR Green‒based real time PCR using primers designed from the isolates of study area. Cows having single infection with T. annulata with/without clinical signs of inappetence, low milk yield, pale mucous membranes, fever, enlarged prescapular lymph node, soil licking, panting, coughing, salivation and lachrymation were subjected to further investigation where parasitaemia and piroplasms per 1000 erythrocytes ranged from 1.6 × 107 to 1.2 × 108 parasites/mL of blood and 3-24 piroplasms in moderate group (16/65), 4.4 × 108 to 6.9 × 109 parasites/mL of blood and >88 piroplasms in severe group (30/65) and 1.6 × 104 to 5.5 × 106 parasites/mL of blood and 0-1 piroplasms in asymptomatic or carriers (17/65), respectively. Study unfolded significant difference in T. annulata parasitaemia among apparently healthy and ill cows. Phylogenetic analysis of our T. annulata isolates (NCBI accession numbers MN098316, MN098317 and MN098318) exhibited maximum similarity with the isolates detected in other parts of India.

Structural characterization of a novel full-length transcript promoter from Horseradish Latent Virus (HRLV) and its transcriptional regulation by multiple stress responsive transcription factors.

KEY MESSAGE: The promoter fragment described in this study can be employed for strong transgene expression under both biotic and abiotic stress conditions. Plant-infecting Caulimoviruses have evolved multiple regulatory mechanisms to address various environmental stimuli during the course of evolution. One such mechanism involves the retention of discrete stress responsive cis-elements which are required for their survival and host-specificity. Here we describe the characterization of a novel Caulimoviral promoter isolated from Horseradish Latent Virus (HRLV) and its regulation by multiple stress responsive Transcription factors (TFs) namely DREB1, AREB1 and TGA1a. The activity of full length transcript (Flt-) promoter from HRLV (- 677 to + 283) was investigated in both transient and transgenic assays where we identified H12 (- 427 to + 73) as the highest expressing fragment having ~ 2.5-fold stronger activity than the CaMV35S promoter. The H12 promoter was highly active and near-constitutive in the vegetative and reproductive parts of both Tobacco and Arabidopsis transgenic plants. Interestingly, H12 contains a distinct cluster of cis-elements like dehydration-responsive element (DRE-core; GCCGAC), an ABA-responsive element (ABRE; ACGTGTC) and as-1 element (TGACG) which are known to be induced by cold, drought and pathogen/SA respectively. The specific binding of DREB1, AREB1 and TGA1a to DRE, ABRE and as-1 elements respectively were confirmed by the gel-binding assays using H12 promoter-specific probes. Detailed mutational analysis of the H12 promoter suggested that the presence of DRE-core and as-1 element was indispensable for its activity which was further confirmed by the transactivation assays. Our studies imply that H12 could be a valuable genetic tool for regulated transgene expression under diverse environmental conditions.

Prevalence of Newcastle disease and associated risk factors in domestic chickens in the Indian state of Odisha.

Newcastle disease (ND), caused by Newcastle disease virus (NDV), is a contagious disease that affects a variety of domestic and wild avian species. Though ND is vaccine-preventable, it is a persistent threat to poultry industry across the globe. The disease represents a leading cause of morbidity and mortality in chickens. To better understand the epidemiology of NDV among commercial and backyard chickens of Odisha, where chicken farming is being prioritized to assist with poverty alleviation, a cross-sectional study was conducted in two distinct seasons during 2018. Choanal swabs (n = 1361) from live birds (commercial layers, broilers, and backyard chicken) and tracheal tissues from dead birds (n = 10) were collected and tested by real-time reverse transcription polymerase chain reaction (RT-PCR) for the presence of matrix (M) and fusion (F) genes of NDV. Risk factors at the flock and individual bird levels (health status, ND vaccination status, geographical zone, management system, and housing) were assessed using multivariable logistic regression analyses. Of the 1371 samples tested, 160 were positive for M gene amplification indicating an overall apparent prevalence of 11.7% (95% CI 10.1-13.5%). Circulation of virulent NDV strains was also evident with apparent prevalence of 8.1% (13/160; 95% CI: 4.8-13.4%). In addition, commercial birds had significantly higher odds (75%) of being infected with NDV as compared to backyard poultry (p = 0.01). This study helps fill a knowledge gap in the prevalence and distribution of NDV in apparently healthy birds in eastern India, and provides a framework for future longitudinal research of NDV risk and mitigation in targeted geographies-a step forward for effective control of ND in Odisha.

RETRACTED: WRKY71 and TGA1a physically interact and synergistically regulate the activity of a novel promoter isolated from Petunia vein-clearing virus.

Caulimoviral promoters have become excellent tools for efficient transgene expression in plants. However, the transcriptional framework controlling their systematic regulation is poorly understood. To understand this regulatory mechanism, we extensively studied a novel caulimoviral promoter, PV8 (-163 to +138, 301 bp), isolated from Petunia vein-clearing virus (PVCV). PVCV was found to be Salicylic acid (SA)-inducible and 2.5-3.0 times stronger than the widely used CaMV35S promoter. In silico analysis of the PV8 sequence revealed a unique clustering of two stress-responsive cis-elements, namely, as-11 and W-box1-2, located within a span of 31 bp (-74 to -47) that bound to the TGA1a and WRKY71 plant transcription factors (TFs), respectively. We found that as-1 (TTACG) and W-box (TGAC) elements occupied both TGA1a and WRKY71 on the PV8 backbone. Mutational studies demonstrated that the combinatorial influence of as-1 (-57) and W-box1-2 (-74 and -47) on the PV8 promoter sequence largely modulated its activity. TGA1a and WRKY71 physically interacted and cooperatively enhanced the transcriptional activity of the PV8 promoter. Biotic stress stimuli induced PV8 promoter activity by ~1.5 times. We also established the possible pathogen-elicitor function of AtWRKY71 and NtabWRKY71 TFs. Altogether, this study elucidates the interplay between TFs, biotic stress and caulimoviral promoter function.