Molecular Characterization of the Functional Genes Associated with Silk Assembly, Transport, and Protection in the Silk Glands of Popular Multivoltine Breeds of Silkworm Bombyx mori. L.

Bombyx mori is an agriculturally important insect used extensively for silk production. India, especially the eastern regions, is mostly dependent on the multivoltine breeds of silkworm Bombyx mori and their hybrids/crossbreeds. The multivoltine breeds are indigenous and superior in survival and hardiness but are relatively inferior in terms of qualitative traits, typically the silk quality. Therefore, it is highly relevant to understand the mechanism of silk production in the multivoltine breeds to decipher the reasons for the inferior quality of silk produced by the multivoltine breeds and thus gain leads to improve the quality of silk production in multivoltine breeds. With this background, study was carried to identify differential expression of the major genes associated with silk proteins in the silk gland region of the popular multivoltine breeds. Our results indicated that although fib-L, fib-H, Sericins, and P25 are the major genes associated with silk filament, a few other genes associated with silk assembly, transport, and protection in the silk glands are the ones that largely contribute towards efficient silk production. The differential expression of these genes had a major effect on the movement of silk proteins within the silk gland and the efficiency of silk production as well. The Pearson correlation revealed a positive correlation amongst the genes dealt with in this study, indicating that the concurrent increase in expression of both the types of genes in the silk glands, significantly improves the silk production.

Targeting genes involved in nucleopolyhedrovirus DNA multiplication through RNA interference technology to induce resistance against the virus in silkworms.

RNA interference (RNAi) has become an efficient tool for inducing resistance to viruses in many organisms. In this study, Escherichia coli cells were engineered to produce stable double-stranded RNA (dsRNA) against the nucleopolyhedrosis virus to elicit RNAi in silkworms. The immediate-early-1 (ie-1) and late expression factor-1 (lef-1) genes of the Bombyx mori nucleopolyhedrovirus (BmNPV) involved in viral DNA multiplication were cloned in the plasmid L4440 under the influence of the double T7 promoter and transformed to E. coli HT115 DE3 host cells. On induction with isopropyl ß-D-thiogalactopyranoside, these cells efficiently produced dsRNA of the cloned genes. The B. mori larvae were fed with 50 µL of E. coli cells expressing ie-1 and lef-1 dsRNAs (each approximately 25 µg) to elicit RNAi. The semi-quantitative and quantitative PCR analysis of RNA from the midgut of the dsRNA-fed larvae revealed a significant reduction in the expression of the target genes involved in BmNPV multiplication, which restricted virus copy numbers to 100 compared with 1.9 × 105 in the infected controls. Furthermore, the dsRNA-fed infected larvae showed > 50% increased survivability compared with the infected controls. The study revealed the successful use of bacteria as vectors for efficiently delivering dsRNA to elicit RNAi against BmNPV in silkworms.

First Report of Tobacco streak virus Infecting Pigeon Pea (Cajanus cajan) in India.

Pigeon pea is an important pulse crop grown in diversified cropping systems in India. In the rainy season of August 2011 and September 2012, pigeon pea cv. LRG 30 plants with leaf necrosis having wrinkled margin on one side were observed in Kadiri mandal of the Anantapuram district of Andhra Pradesh (A.P.), India. Symptoms included necrotic spots on young leaves followed by wilting of leaves, petiole and branch/axillary shoot proliferation, with small leaves having mosaic symptoms. Symptomatic leaves were sap-inoculated onto 10 seedlings of cowpea (cv. Pusa Komal) using 0.01 M phosphate buffer (pH 7.0). Localized necrotic lesions developed in all the inoculated plants after 2 days post inoculation. Field symptoms were reproduced on healthy pigeon pea upon back inoculation using single lesions of infected cowpea leaves. In direct antigen coating (DAC)-ELISA, all the infected pigeon pea and cowpea leaf samples were positive to a polyclonal antiserum specific to Tobacco streak virus (TSV) supplied by ICRISAT, India. Total RNA was extracted using infected pigeon pea and healthy leaf samples by TRI Reagent (Sigma). Reverse transcription (RT)-PCR was carried out using primers specific to the coat protein (CP) gene of TSV (1). A product of the 700-bp DNA fragment was obtained in field-infected pigeon pea samples but not in healthy controls. The amplicon was cloned into PTZ57R/T using the Ins TA clone PCR kit (Fermentas). Recombinant clone was sequenced in both directions and the CP gene sequence obtained was deposited in GenBank (KF220492). Sequence analysis of the CP gene of TSV from pigeon pea shared 98 to 100% identity with Indian TSV isolates originating from different hosts including groundnut (FJ355948), mung bean (FJ749259), and sunflower (DQ864448), and 88 to 92% similarity with TSV type isolate (white clover: X00435) both at nucleotide and amino acid levels. TSV belongs to the genus Ilarvirus of family Bromoviride and has a wide host range. TSV is pollen borne, assisted by thrips causing mechanical injury (2). To our knowledge, this is the first report of TSV on pigeon pea in India and was widespread in Anantapuram, Kadapa, Kurnool, and Mahbubnagar districts of A.P. Yield loss depends on the stage of infection as early infection resulted in complete failure of the crop. TSV was prevalent on many legume crops such as black gram, green gram, and groundnut in A.P, Tamil Nadu, Karnataka, and Maharashtra states (3). TSV infection of pigeon pea may pose a serious implication for pulse production. References: (1). A. I. Bhat et al. Arch. Virol. 147:651, 2002. (2). M. Sharman et al. Australian Plant Dis. 6:54, 2011. (3). K. Vemana and R. K. Jain. Indian J. Virol. 21:117, 2010.

The post-cystectomy pseudotumour sign: MRI appearances of a modified chronic pelvic haematoma due to retained haemostatic gauze.

The magnetic resonance imaging (MRI) findings of five men who had incidental similar pelvic masses identified after radical cystectomy are presented. In all patients the haemostatic agent Kaltostat had been used. In one patient, surgical resection of the mass was performed and histological evaluation showed a foreign body inflammatory reaction within a chronic haematoma. The differentiation between this lesion and other post-operative collections or tumours is discussed.

Hepatic lesion detection: comparison of MR imaging after the administration of superparamagnetic iron oxide with dual-phase CT by using alternative-free response receiver operating characteristic analysis.

PURPOSE: To compare the performance of magnetic resonance (MR) imaging after the administration of superparamagnetic iron oxide (SPIO) and dual-phase computed tomography (CT) in the depiction of liver metastases. MATERIALS AND METHODS: Fifty-one hepatic resection candidates with known colorectal metastases were examined. MR imaging comprised fast spin-echo (SE) T2-weighted imaging, T1-weighted gradient-echo (GRE) fast low-angle shot imaging before SPIO enhancement, dual-echo SE imaging, T2-weighted fast low-angle shot imaging, and T1-weighted GRE imaging after SPIO enhancement. CT was performed with 8-mm collimation and 1:1 pitch; imaging commenced 20 seconds and 65-70 seconds after injection of 150 mL of contrast medium. All images were reviewed independently by four blinded observers. The alternative-free response receiver operating characteristic (ROC) method was used to analyze the results, which were correlated with findings from surgery, intraoperative ultrasonography, and histopathologic studies in 31 patients and with consensus review together with all other imaging and clinical follow-up in 20 patients. Sensitivities were also calculated. RESULTS: The mean sensitivity of MR was significantly higher than that of CT (p < .02): 79.8% for MR and 75.3% for CT for all lesions, and 80.6% for MR and 73.5% for CT for malignant lesions. The mean areas under the alternative-free response ROC curves were 0.83 for MR and 0.78 for CT (difference not significant). CONCLUSION: SPIO-enhanced MR imaging was more sensitive than dual-phase CT in the depiction of colorectal metastases.

The transrectal ultrasound and MRI appearances of granulomatous prostatitis and its differentiation from carcinoma.

AIMS AND METHODS: Granulomatous prostatitis is a benign inflammatory condition of the prostate which can be mistaken for prostatic carcinoma both clinically and on ultrasound, but is distinguishable histologically. The transrectal ultrasound (TRUS) and magnetic resonance imaging (MRI) appearances of 10 patients with histologically confirmed granulomatous prostatitis were evaluated to try and identify any correlation between the two techniques or any specific features on MRI to help distinguish it from carcinoma. Clinical findings and serum prostatic specific antigen (PSA) levels were also evaluated. RESULTS: In five patients, both TRUS and MRI were concordant, showing only changes of benign prostatic hypertrophy (three patients) or showing no abnormality (two patients). In a further three patients, both TRUS and MRI were abnormal, with appearances suggestive of carcinoma. One of these patients had tuberculous prostatitis and had a past history of tuberculosis. In the remaining two patients, there was a discrepancy between TRUS and MRI findings, carcinoma being suspected on TRUS in one with a normal MRI, and carcinoma suspected on MRI in the other with a normal TRUS. CONCLUSION: There is no pattern of clinical, biochemical, ultrasound or MRI findings that allows a specific diagnosis of granulomatous prostatitis to be made, or differentiation from prostatic carcinoma.

Imaging the thyroid.

Thyroid imaging has historically relied heavily on scintigraphy, although, not surprisingly in view of the superficial position of the gland, ultrasound has assumed an increasingly prominent role in recent years. The other cross-sectional imaging modalities can also be useful, and the emergence of new radiopharmaceuticals and the increasingly central role of fine needle aspiration cytology have further added to the range of diagnostic techniques available. This review attempts to summarize the current state of knowledge, and makes some suggestions for the most efficient use of imaging resources in the investigation of thyroid disease.

Staging lymphoma with CT: comparison of contiguous and alternate 10 mm slice techniques.

PURPOSE: To compare two computed tomography (CT) techniques, contiguous 10 mm and alternate 10 mm slices of the chest, abdomen and pelvis, for initial staging of lymphoma. SUBJECTS AND METHODS: Fifty-two consecutive patients referred at initial diagnosis for lymphoma staging by CT were examined with contiguous 10 mm slices of the chest, abdomen and pelvis. Oral contrast was administered, but no intravenous contrast. Two sets of films for each examination were printed, one with 10 mm contiguous slices and one with 10 mm alternate slices. The two sets of films for each patient were reviewed separately in a randomized order and blinded fashion by two independent observers. Discrepancies were reviewed by a third observer and subsequently resolved by consensus. RESULTS: Staging assessments by both techniques were concordant in 51 of 52 cases: one patient was recorded as stage II on the alternate slice technique, but stage III with contiguous slices but this difference did not affect management. Inter-technique agreement was very good (kappa=0.97). The staging assessments showed discrepancies between the two observers in 12 of the 52 cases which were independent of technique (kappa=0.71; good agreement). Consensus review showed them to result from differences in perception in seven cases and differences in interpretation of abnormalities in five cases. CONCLUSION: Our findings support the use of an alternate 10 mm slice technique in the staging of lymphoma at initial diagnosis. Observer variation was greater than the difference between techniques.

Magnetic resonance cholangiopancreatography accurately predicts the presence or absence of choledocholithiasis.

Accurate common bile duct (CBD) imaging in patients with biliary calculi is an important determinant of specific therapy. Noninvasive methods to evaluate calculi in the CBD have limited accuracy and rely mainly on ultrasonography and computed tomography. Magnetic resonance cholangiopancreatography (MRCP) is a new noninvasive modality available to evaluate the biliary system. This study was undertaken to assess the accuracy of MRCP in predicting the presence or absence of CBD stones in patients at increased risk for choledocholithiasis. The medical records of 48 patients with a final diagnosis of biliary calculous disease undergoing MRCP between November 1995 and April 1997 were retrospectively reviewed. Three groups were identified: choledocholithiasis (n = 19), gallstone pancreatitis (n 5 11), and uncomplicated cholelithiasis (n = 18). In all patients the presence or absence of CBD calculi, as determined by MRCP, was correlated with the final diagnosis obtained from endoscopic retrograde cholangiopancreatography (ERCP) (n = 19), intraoperative cholangiography (n = 6), CBD exploration (n = 13), or clinical follow-up (n = 10). Sensitivity, specificity, and accuracy of MRCP were determined. The major clinical indications for MRCP in the 48 patients ware abnormal liver function tests followed by hyperamylasemia. Twenty patients were diagnosed with CBD stones and 28 were not. MRCP correctly predicted the presence of CBD stones in 19 of 20 patients and failed to detect CBD stones in one patient with gallstone pancreatitis. MRCP incorrectly predicted the presence of CBD stones in 3 of 28 patients ultimately found to have gallstones and no CBD stones. MRCP correctly predicted the absence of CBD stones in the other 25 patients including 10 patients with gallstone pancreatitis. Overall, MRCP had a sensitivity of 95%, a specificity of 89%, and an accuracy of 92%. MRCP is an accurate, noninvasive test for evaluating the CBD duct for the presence or absence of calculi in patients suspected of having CBD stones. Our data support the use of MRCP in the preoperative evaluation of these patients as findings may influence therapeutic decisions.

Comparison of dynamic contrast enhanced MRI and Doppler ultrasound in the pre-operative assessment of the portal venous system.

The purpose of this study was to compare dynamic contrast enhanced MRI (DCEMR) with Doppler ultrasound (US) in the assessment of portal venous anatomy and to analyse the causes of discrepancy. Over a 1 year period, 97 patients undergoing assessment prior to hepatic surgery underwent imaging of the liver and portal venous system using US with colour and spectral Doppler and MRI with axial T2 weighted spin echo (SE) and coronal oblique T1 weighted rapid gradient echo (GRE) imaging before and immediately after bolus injection of Gd-DTPA (0.1 mmol kg-1). When the US and MRI findings were discrepant, the images were reviewed by two observers and compared with surgical findings. US and DCEMR were concordant in 90 patients (portal vein patent in 80, occluded in 10). In three patients with cirrhosis and gross ascites the portal vein was reported as occluded on US and patent on MRI; surgery confirmed the MRI findings. In one patient the portal vein was patient on US but not on MRI, but there was a 3 week interval between the examinations. In three patients the portal vein was patent on US, but MRI detected occlusion of intrahepatic portal vein branches in two, and encasement of an intrahepatic branch in the third case. Spontaneous splenorenal shunts were seen in 15 patients only on MRI; varices were seen in 39 patients on MRI and in 22 patients on US. Both US and DCEMR contribute to the pre-operative assessment of the portal venous system. MRI provides additional information over US in assessing intrahepatic portal branches and detecting varices and splenorenal shunts, and is recommended for all surgical candidates and in patients with abnormal portal venous anatomy and equivocal US findings.

Is computed tomography of the body overused? An audit of 2068 attendances in a large acute hospital.

Computed tomography (CT) delivers a substantial proportion of the medical radiation dose to the population. The National Radiological Protection Board (NRPB) recommends that "CT examinations should only be carried out after there has been proper clinical justification for the examination of each individual by an experienced radiologist". The aim of this study was to review the indications for body CT in a large acute hospital in order to identify groups of patients in whom CT might not be "clinically justified". We reviewed the age, sex and diagnosis of all patients undergoing CT of the abdomen and pelvis (including lumbar spine) in our department in 1992. All requests had been checked by a senior radiologist before booking. CT was performed in 1078 male patients and in 990 females. Patients examined for benign disease were younger (mean age 52 years) than patients with malignant disease (mean age 57 years). Of 915 examinations performed for benign disease, 293 (32%) were orthopaedic in nature and included 255 (87%) for back pain. CT was carried out for benign non-orthopaedic disease in 97 women under 50 years old, usually following other tests with inconclusive or normal results. 49 of these examinations were reported as normal. Of 191 patients who had more than one CT examination, 158 (83%) were undergoing follow-up for malignant disease. All 39 patients who underwent CT on three or more occasions had life-threatening conditions, either malignancy or complicated acute pancreatitis. We have been unable to find substantial numbers of patients in whom CT was performed inappropriately. Substitution of magnetic resonance imaging (MRI) for CT of the spine will help population dose reduction. The benign condition responsible for the largest radiation dose from CT is complicated acute pancreatitis, and it may be possible to substitute MRI for CT in these patients. Women under 50 years old with normal CT form an important group for clinical audit.

Multispiral three-dimensional computed tomography in the investigation of craniosynostosis: technique optimization.

21 infants with craniosynostosis were studied with a new three-dimensional (3D) computed tomography (CT) methodology. We describe technique optimization using multiple spiral data acquisitions with low dose (85 mAs) technique. One caudal volume of 3 mm slice thickness was obtained with a further two volumes of 1 mm slice thickness at the vertex. Image reconstruction of spiral raw data allowed overlapping 3 mm sections to be generated without the dose increase that would result from conventional axial CT scanning. We illustrate common technical artefacts of 3D CT and explain their cause and solution. A dramatic dose reduction to the lens was achieved with no loss in 3D image quality. Lens dose was 8.91 mSv compared with 24.6 mSv using the standard paediatric head technique.

Corticosteroid therapy and intercurrent illness: the need for continuing patient education.

In patients receiving long-term therapeutic or replacement corticosteroids, delayed or inappropriate adjustment of steroid dosage during intercurrent illness may be fatal. We used a questionnaire to assess current levels of patient knowledge, awareness of the need for action during intercurrent illness and the frequency with which steroid warning cards and Medic Alert pendants were carried, in 61 patients on long-term replacement corticosteroids and in 40 patients receiving long-term therapeutic corticosteroids. Only 67 of the 101 patients taking corticosteroids were carrying a steroid warning card. Eleven of the 21 Medic Alert owners wore their pendants. Only 18 of the 41 patients in the therapeutic group and 41 of the 60 patients in the replacement group would take appropriate action during an intercurrent illness (P < 0.001). Lack of patient knowledge in this important area emphasizes the need for continuing and effective education of these groups of patients during follow-up. An information sheet detailing the exact changes to be made during intercurrent illness may help reinforce verbal advice.

Is antral gastrin important in the resistance of duodenal ulcers to H2 receptor antagonists or in recurrent ulceration after highly selective vagotomy?

Basal serum gastrin, integrated gastrin response to a meal, and integrated gastrin response to insulin induced hypoglycaemia were measured in 60 patients with duodenal ulcer before and after elective highly selective vagotomy to determine whether antral gastrin has a role in resistance to H2 receptor antagonist treatment which the patients had received before surgery or in the development of recurrent ulceration after vagotomy. The basal gastrin, integrated gastrin response to a meal, and the integrated gastrin response to insulin were similar in patients whose ulcers healed after H2 receptor agonist treatment or were refractory to at least three months of this treatment. The same parameters measured before or after highly selective vagotomy were similar in patients who eventually developed recurrent ulceration compared with those who did not. As expected the basal and meal stimulated (but not insulin stimulated) serum gastrin values increased after highly selective vagotomy. Ulcer patients with particularly high gastrin values (whether basal or stimulated) were not more resistant to H2 receptor antagonist treatment or prone to develop ulcer recurrence after highly selective vagotomy. This study suggests that antral gastrin is not important in 'resistance' of duodenal ulceration either to H2 receptor antagonist treatment or to highly selective vagotomy.