Multiple Myeloma: A Review of the Literature and a Case Report Highlighting the Immunocompromised State of Myeloma Patients.

Multiple myeloma (MM), a malignancy involving plasma cells, disproportionately affects older adults with an average age of diagnosis of about 70 years. Oftentimes, the therapies used in the treatment of MM are associated with a risk for immunotoxicity, lowering the ability of the immune system to fight off opportunistic infections. This is an important relationship for clinicians to realize as the incidence of opportunistic infections in myeloma patients is increasing. As an example, we present a case of a patient with MM who subsequently developed a cryptococcal infection. Our paper will highlight the key details of the case as well as shed light on the importance of understanding the immunodeficiencies in this patient population. We highlight important aspects of the current literature related to MM and relate them to the associated case.

Brachioradialis Involvement in Hirayama's Disease: An Atypical Presentation of a Rare Cervical Myelopathy.

Juvenile monomelic amyotrophy (JMA), also known as Hirayama's disease, is a rare cervical myelopathy that predominantly affects young Asian males. It is characterized by degeneration of anterior horn cells due to compression by the redundant dural sac. This study presents an atypical case of a 23-year-old Indian male who exhibited uncommon symptoms of JMA. The patient displayed progressive weakness and atrophy in the left forearm, including the usually spared brachioradialis muscle. Electrophysiological tests and MRI scans solidified the diagnosis of Hirayama's disease. After wearing a cervical collar for one year, the patient's condition stabilized, reinforcing the diagnosis. Unlike most JMA cases, this instance highlights the involvement of the brachioradialis muscle, underlining the variability in JMA presentations. A precise diagnosis is contingent upon clinical criteria, dynamic MRI, and electrophysiological findings. Recognizing these variations is crucial for early detection and appropriate management of the disease.

Intractable hiccups: How can chest X-ray help?

We describe the case of a young male presenting with hiccups refractory to antacids. Due to high index of suspicion was evaluated with a chest X-ray which revealed pericardial effusion. The effusion was drained, and the hiccups resolved. It is a teaching moment, to consider simple imaging, i.e., chest X-ray in patients with refractory hiccups to rule out more serious underlying causes.

Precursors and elicitor induced enhancement of cell biomass and phenolic compounds in cell suspensions of Indian basil-Ocimum basilicum (CIM-Saumya).

CIM-Saumya is an improved, methyl chavicol rich variety of Ocimum basilicum (Family-Lamiaceae), developed by Council of Scientific and Industrial Research-Central Institute of Medicinal and Aromatic Plants. This plant possesses analgesic, anti-ulcerogenic, anti-inflammatory, anti-oxidant, cardiac stimulant, Central Nervous System depressant, hepatoprotective and immunomodulator activities due to the presence of various phytoconstituents. Among them rosmarinic acid, caffeic acid and ferulic acid are the three major phenolic compounds responsible for its therapeutic utility. These compounds are produced in very low amounts in the in vivo plants. Therefore, the present study has been conducted for establishment of cell suspensions, optimization of inoculums size, growth kinetics and screening of elicitor and precursors for the accumulation of cell biomass and the production of the three important phenolic compounds in cell suspension of O. basilicum (CIM-Saumya). Leaf derived friable callus was used for establishing the cell suspension in liquid Murashige and Skoog's medium fortified with 1 g/L casein hydrolysate + 2.26 µM 2,4-dichlorophenoxyacetic acid + 0.465 µM kinetin + 2.68 µM naphthalene acetic acid. The growth kinetic analysis pattern of cell suspension revealed the maximum biomass increments (% BI = 486.7) and production of RA 8.086 mg/g dry weight was found in 30th day harvested cells. Whereas, the other two phenolic compounds i.e. ferulic acid (0.0125 mg/g dry weight) and caffeic acid (0.38 mg/g dry weight) was recorded highest on 25th day of growth cycle. In the present study, one biotic elicitor i.e. yeast extract and three precursors [peptone, tryptone and lactalbumin hydrolysate] were tested, among them, lactalbumin hydrolysate (100 mg/L; added at 16th day) treated cells recorded highest estimated phenolic compounds yield (251.5 mg/L; 6.81 fold compared to the control) and biomass increments i.e. % BI = 1207 with 1.85 fold compared to the control. The highest rosmarinic acid content i.e. 25.47 mg/g DW (4.4 fold compared to the control) and 24.42 mg/g dry weight (4.1 folds compared to the control) was noticed in 30th day harvested cells treated with yeast extract (1 g/L on 0 day) and lactalbumin hydrolysate (100 mg/L added on 16th day), respectively. While caffeic acid content (0.91 mg/g dry weight) showed 2.9 folds higher compared to the control in cells treated with peptone 200 mg/L added on 16th day of culture cycle. All the treated cells showed enhanced phenylalanine ammonia-lyase enzyme activity with highest specific activity in lactalbumin hydrolysate followed by tryptone, peptone, and yeast extract. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01316-6.

Validation of a New Graphic Facial Nerve Grading System: FAME Scale.

Background and Objective: Most of the existing qualitative facial nerve grading systems are very subjective while the quantitative grading systems are more complex, require longer data input time and specific software. There is a need for having a scoring system with graphic criteria to improve the subjectivity, reliability and convenience. We aimed to develop and validate such a reliable graphic scale for use in Bell's palsy. Methods: Face videos of patients with unilateral facial paralysis were recorded using smartphones and analyzed for six items including five voluntary facial movements apart from complications of facial palsy (synkinesis, hyperkinesis, and contracture). 15 videos were used for pilot study, 75 for the development of scale and 110 for its validation. Each video was rated on two separate occasions by 3 independent raters, a score of 0-4 was assigned to each item using the graphic scoring criteria, and a composite score was obtained (range 0-24). Five disease severity categories: normal (score 0), mild (score 1-6), moderate (score 7-12), severe (score: 13-18) and profound facial weakness (score: 19-24). Results: The proposed scale and its component items had high inter-rater and intra-rater reliability (Kappa >0.7). Good correlation (Pearson co-efficient >0.7) was seen among the voluntary movements. The proposed scale is a valid tool to score motor deficits and complications of facial palsy. Conclusions: The proposed scale is a valid and reliable graphic scale to describe facial motor dysfunction and its secondary defects.

De Novo Shoot Bud Induction from the Catharanthus roseus Leaf Explants and Agrobacterium tumefaciens-Mediated Technique to Raise Transgenic Plants.

The regeneration of a whole plant from a single cell or organ explant was a valuable task for plant biotechnology. However, important medicinal plants such as Catharanthus roseus have shown recalcitrance to regeneration protocols, thus limiting investigations on MIA metabolism and metabolic engineering in this plant system. In this chapter, successful regeneration protocols were detailed for Catharanthus roseus, either by direct shoot bud induction from leaf explants and Agrobacterium-mediated genetic transformation.

A Study of Role of Ngal in Diagnosis and Staging the Severity of Diabetic Nephropathy in Type 2 Diabetes Mellitus Patients in Sms Medical College, Jaipur.

Present work was conducted to study of role of NGAL in diagnosis and staging the severity of diabetic nephropathy in type 2 diabetes mellitus patients to assess serum NGAL, urine albumin levels in diabetic patients with and without apparent nephropathy. MATERIAL: Comparative study conducted in the General Medicine department of SMS Hospital. INCLUSION CRITERIA: Patients of type 2 Diabetes mellitus. EXCLUSION CRITERIA: Patients consuming high protein diet, pregnancy, severe muscular exercises, orthostatic albuminuria, congestive heart failure, urinary tract infections, liver diseases. Patients with type 1 diabetes mellitus. OBSERVATION: The mean age of patient was 51.7 years. Here, HbA1c and NGAL were negatively correlated with each other with Pearson correlation -0.484 (p-value<0.05). In prediabetic patients mean NGAL was 407ng/ml and as HbA1c increases mean NGAL decreases 276.69 ng/ml (P-value <0.0012). As ACR increases mean NGAL increases significantly (P-value<0.05). CONCLUSION: Neutrophil gelatinase-associated lipocalin could be a renal function evaluation marker for patients with renal dysfunction and markers for detection of diabetic nephropathy in patients with type2 diabetes mellitus.

A Study of Adrenal Insufficiency in Hemodynamically Stable Patients with Cirrhosis.

Adrenal insufficiency (AI) is well entrenched in medical constraints like septic shock, critically ill and multi-morbid hemodynamically unstable patients but its exact prevalence or differences in the cases of chronic liver disease (CLD) at variable grades of severity has recently gained momentum. The eventuality of AI propounding in stable compensated and decompensated cirrhosis without sepsis or in early and late stages of liver desecration are the existing lacunae in popular literature that this study aims to address. MATERIAL: A prospective, analytical study was conducted from March 2021 to December 2021 encompassing 100 hemodynamically stable patients with cirrhosis without infection, admitted at SMS Medical College, Jaipur, who were assessed clinically, biochemically and for adrenal functions. Adrenal insufficiency was defined on multivariable approach including basal 8am cortisol levels, followed by giving 250mcg synthetic adrenocorticotrophic hormone IM injection and retaking serum cortisol levels post-hourly interval to delineate peak and delta cortisol variables. All samples were processed by chemoluminiscence based method on fully automatic immunoassay analyser. OBSERVATION: The study comprised 81 males and 19 females with the mean age being 45.4±12.92 years, with CLD etiology concentrating substantially around alcohol consumption (71%). Viral comorbidities viz. HBV, HCV, both viral and alcohol related and miscellaneous causes were documented in 23, 10, 14 and 12 patients respectively. AI surfaced in 38% patients with CLD being statistically significant with p< 0.001. Inclusively, 10.5% patients with Child-Turcotte-Pugh (CTP) class A, 57.89% with CTP class B and 31.57% cases with CTP class C developed adrenal insufficiency. No statistical differences were found in age, sex; mean arterial pressure, heart rate, HDL, cirrhosis etiology, degree of alcohol consumption and manifestations of portal hypertension between patients with or without AI. For prudence, serum albumin levels were lower (p<0.5) with INR raised (p<0.33) in patients with AI than their counterparts. However, multivariate analysis revealed no direct independent adrenal insufficiency predictor. ROC curve showed that the CTP score may be a good predictor for AI in liver cirrhosis patients as supplemented by significant negative correlations found between CTP score and peak cortisol levels (p=0.001). CONCLUSION: Adrenal insufficiency found frequent even in stable cirrhotic patients form an integral division of the CLD spectra and worsening glucocorticoid levels should be periodically assessed in such patients for preventing parallel comorbidities.

A Study of Left Ventricular Dysfunction in Normotensive Non Diabetic Patients with Non Alcoholic Fatty Liver Disease.

Non-Alcoholic Fatty Liver Disease is an emerging epidemic in the face of the new generation. It is considered the hepatic manifestation of the metabolic syndrome. With the increasing prevalence of obesity and metabolic syndrome, it has become a common sight in our outpatient department. We have investigated the echocardiographic parameters for systolic and diastolic dysfunction in the patients with NAFLD to evaluate its effects on the heart and hope our study could shine a light and provide us with a perspective into the various effects the metabolic syndrome has on our body. MATERIAL: We recruited 35 Normotensive, Non-Diabetic Non-Alcoholic Fatty Liver Disease Patients (NAFLD) of age ranging from 18 to 60 years of age diagnosed on the basis of ultrasound Abdomen and 35 Controls from both inpatient and outpatient department of Sawai Man Singh Medical College and Allied Hospitals from the month of August 2021 to October 2021 for the study. Every Patient underwent conventional transthoracic and Tissue Doppler Echocardiography along with their physical and metabolic parameters. All the patients of NAFLD were graded ultrasongraphically based on Hamaguchi et al criteria. OBSERVATION: NAFLD patients had higher Body Mass Indices, Abdominal circumferences, Systolic Blood pressures, Total Cholesterol levels and Low-Density Lipoprotein levels in comparison to their normal counterparts. On the Echocardiographic front, the patient underwent transthoracic 2D ECHO and we observed an increased interventricular septum thickness (0.99 ± 0.04 vs 0.78 ± 0.05, p <0.0001), posterior wall thickness (0.96±0.05 vs 0.75±0.04 p < 0.0001) and left ventricular Mass (173±22.6 vs 116±8.24 p < 0.0001). On Tissue Doppler Imaging (TDI) we observed a decreased E/A ratio (1.25 ± 0.17 vs 1.44 ± 0.22 p < 0.0001) which was suggestive of an increased Left ventricular Dysfunction. On correlational analysis, we had made an observation that there was a positive correlation of the grading of fatty liver with that of interventricular septum thickness (r= 0.5305 p = 0.001), posterior wall thickness (r = 0.4362 p= 0.088) and left ventricular mass (r = 0.6292, p = 0.0001) with the grade of fatty liver. CONCLUSION: From our study, it was imperative that NAFLD even in the absence of Hypertension or Diabetes has a role in the impairment of Systolic and Diastolic function of the left ventricle and its role in cardiovascular morbidity and mortality cannot be ignored.

A Study of Pulmonary Function Tests in Patients with Rheumatoid Arthritis and its Association with Disease Activity.

Rheumatoid arthritis is a chronic inflammatory disease of unknown etiology marked by a symmetrical peripheral polyarthritis. Because it is a systemic disease RA may result in a variety of extra articular manifestations, Pulmonary involvement is a well known extra-articular manifestation of RA which can have myriad presentations and might be the second cause of death after infection. Pulmonary function test(PFTs) are widely used to provide objective measure of lung function for detecting and quantifying impairment of pulmonary function in patients with cardiopulmonary diseases. Although lung disease secondary to pulmonary infection and/or drug toxicity are frequent complications, lung disease directly associated with the underlying RA is more common. Lung involvement in RA carries a worse prognosis. The early recognition of changes in PFT may be used as early marker of pulmonary involvement and help in limiting the mortality and morbidity caused by the disease. Hence need for the study. MATERIAL: Descriptive cross sectional study conducted on 100 cases. INCLUSION CRITERIA: Patients Aged 18-60 years diagnosed with RA according to American college of Rheumatology-European league against rheumatism classification criteria. EXCLUSION CRITERIA: Known smokers, Presence of prior pulmonary disease, Known case of other Collagen vascular diseases and History of pulmonary infection in the last 3 months. Disease activity assessed by both DAS28 and CDAI score. OBSERVATION: Mean age of cases was 43.57 years with 21% males and 79% females. 96% cases had normal FEV1/ FVC ratio with mean DAS28 of 5.12±0.8 and 4% cases had mild obstruction with mean DAS28 in mild restriction being 6.2±0.588. There was statistically significant difference in DAS28 in cases having mild restriction compared to normal cases (P-value=0.0066). Here, we also found that as severity of restrictive pattern increases there was no significant effect on DAS score (P-value=0.3221). But duration of disease and duration of methotrexate use increases as severity of restrictive pattern increases (P-value <0.0001). CONCLUSION: we conclude from our study that there was predominant prevalence of restrictive pattern of PFT in the patients with RA, pulmonary involvement had significant association with duration of disease and duration of methotrexate use, on the contrary disease activity does not have much influence on pulmonary involvement in the diseased individual. Hence PFT alone can't be used as lone modality for early detection of pulmonary involvement in rheumatoid arthritis, additional implementation of DLCO along with spirometry can be tried to assess early involvement of respiratory membrane.

ZCTs knockdown using antisense LNA GapmeR in specialized photomixotrophic cell suspensions of Catharanthus roseus: Rerouting the flux towards mono and dimeric indole alkaloids.

The present study was carried out to silence the transcription factor genes ZCT1, ZCT2 and ZCT3 via lipofectamine based antisense LNA GapmeRs transfection into the protoplasts of established photomixotrophic cell suspensions. The photomixotrophic cell suspensions with a threshold of 0.5% sucrose were raised and established using two-tiered CO2 providing flasks kept under high light intensity. The photomixotrophic cell suspensions showed morphologically different thick-walled cells under scanning electron microscopic analysis in comparison to the simple thin-walled parenchymatous control cell suspensions. The LC-MS analysis registered the vindoline production (0.0004 ± 0.0001 mg/g dry wt.) in photomixotrophic cell suspensions which was found to be absent in control cell suspensions. The protoplasts were isolated from the photomixotrophic cell suspensions and subjected to antisense LNA GapmeRs silencing. Three lines, viz. Z1A, Z2C and Z3G were obtained where complete silencing of ZCT1, ZCT2 and ZCT3 genes, respectively, was observed. The Z3G line was found to show maximum production of vindoline (0.038 ± 0.001 mg/g dry wt.), catharanthine (0.165 ± 0.008 mg/g dry wt.) and vinblastine (0.0036 ± 0.0003 mg/g dry wt.). This was supported by the multifold increment in the gene expression of TDC, SLS, STR, SGD, d4h, dat, CrT16H and Crprx. The present work indicates the master regulation of ZCT3 knockdown among all three ZCTs transcription factors in C. roseus to enhance the terpenoid indole alkaloids production. The successful silencing of transcription repressor genes has been achieved in C. roseus plant system by using photomixotrophic cell cultures through GapmeR based silencing. The present study is a step towards metabolic engineering of the TIAs pathway using protoplast transformation in C. roseus. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01017-y.

Confirmation of "pre-plasmolysis mediated ex-osmosis hypothesis" to obtain shoot bud morphogenesis in Catharanthus roseus.

The antineoplastic herb, Catharanthus roseus is a classified high-value low-volume medicinal herb which is in global attention of scientific research for modulation of its monoterpenoid indole alkaloids (MIA) pathway through genetic engineering. These secondary metabolites are generally stored in specific types of structures/compartments due to their cytotoxic nature and designated roles in plant defense response. However, their presence can hinder the genetic engineering process used to develop transgenic plants through de novo morphogenesis and regeneration of plants from cultured cells/tissues and hence, it always remained a critical impediment in transgenic research in C. roseus. The pre-plasmolysis treatment of leaf explants can help to tackle the recalcitrant nature of leaf explant and can support the direct regeneration response by ex-osmosis that minimizes the concentration of alkaloids. Therefore, this study was performed to chase the effect of osmotic conditions on recalcitrant leaves of C. roseus engaged in vitro plant regeneration and hypothesis of alkaloids ex-osmosis is confirmed by HPLC analysis.

TIAs pathway genes and associated miRNA identification in Vinca minor: supporting aspidosperma and eburnamine alkaloids linkage via transcriptomic analysis.

V. minor contains monomeric eburnamine-type of indole alkaloids having utilization as a neuro-medicinal plant. The biosynthetic pathway studies using miRNAs has been the focal point for plant genomic research in recent years and this technique is utilized to get an insight into a possible pathway level study in V. minor as understanding of genes in this prized medicinal plant is meagrely understood. The de novo transcriptomic analysis using Illumina Next gen sequencing has been performed in glasshouse shifted plant and transformed roots to elucidate the possible non confirmed steps of terpenoid indole alkaloids (TIAs) pathway in V. minor. A putative TIA pathway is elucidated in the study including twelve possible TIAs biosynthetic genes. The specific miRNA associated with TIAs pathway were identified and their roles were discussed for the first time in V. minor. The comparative analysis of transcriptomic data of glasshouse shifted plant and transformed roots showed that the raw reads of transformed roots were higher (83,740,316) compared to glasshouse shifted plant (67,733,538). The EST-SSR prediction showed the maximum common repeats among glasshouse shifted plant and transformed roots, although small variation was found in trinucleotide repeats restricted to glasshouse shifted plant. The study reveals overall 37 miRNAs which were observed to be true and can have a role in pathway as they can regulate the growth and alkaloid production. The identification of putative pathway genes plays an important role in establishing linkage between Aspidosperma and Eburnamine alkaloids.

Present status of Catharanthus roseus monoterpenoid indole alkaloids engineering in homo- and hetero-logous systems.

Catharanthus roseus synthesizes one of the most structurally, chemically and biologically active phytomolecules monoterpenoids indole alkaloids (MIAs) with having a wide range of pharmaceutical activities. Being the sole source of antineoplastic MIAs vinblastine and vincristine C. roseus has become one of the most valued plant. The low in planta availability of these MIAs and unavailability of alternative chemical synthesis system has enhanced their demand and equally let to the exorbitant market cost. To bridge this gap alternative production systems have been investigated using MIAs metabolic engineering (ME) in the homologous and heterologous systems. The availability of improved recombinant technologies along with genomics and metabolomics tools has opened the door of tremendous new potentials of ME. To encash these potentials of ME for MIAs pathway, efforts were made by expressing constitutive structure biosynthesis enzymes, transporters, and transcription factors of C. roseus MIAs biosynthesis in both homologous and heterologous systems. Here we review the knowledge of C. roseus MIAs pathway metabolic engineering in homologous and heterologous systems, gained in the past 35 years of C. roseus research.

Infectious Diseases Specialty Intervention Is Associated With Better Outcomes Among Privately Insured Individuals Receiving Outpatient Parenteral Antimicrobial Therapy.

BACKGROUND: Outpatient parenteral antimicrobial therapy (OPAT) can be managed by specialists in infectious diseases (ID) or by other physicians. Better management of OPAT can reduce the likelihood of readmission or emergency department (ED) use. The relative success of ID specialists and other physicians in managing OPAT has received little study. METHODS: We analyzed a national database of insurance claims for privately insured individuals under age 65, locating inpatient acute-care stays in 2013 and 2014 that were followed by OPAT. Through propensity scoring, patients who received outpatient ID intervention (ID-led OPAT) were matched 1-to-1 with those who did not (Other OPAT). We estimated regression models of hospital and ED admissions and of total healthcare payments over the first 30 days after discharge. RESULTS: The final analytic sample of 8200 observations was well balanced on clinical and demographic characteristics. Soft-tissue infection and osteomyelitis were the most common infections in the index event, each affecting more than 40% of individuals. Relative to those with Other OPAT, people with ID-led OPAT had lower odds of an ED admission (odds ratio [OR] 0.449, 95% confidence interval [CI] 0.311-0.645) or hospitalization (OR 0.661, 95% CI 0.557-0.791) over 30 days, and they accumulated $1488 less in total healthcare payments (95% CI -2 688.56--266.58). CONCLUSIONS: Among privately insured individuals below age 65, ID consultations during OPAT are associated with large and significant reductions in the rates of ED admission and hospital admission in the 30 days after index events, as well as lower total healthcare spending.

Overexpression of tryptophan decarboxylase and strictosidine synthase enhanced terpenoid indole alkaloid pathway activity and antineoplastic vinblastine biosynthesis in Catharanthus roseus.

Terpenoid indole alkaloid (TIA) biosynthetic pathway of Catharanthus roseus possesses the major attention in current metabolic engineering efforts being the sole source of highly expensive antineoplastic molecules vinblastine and vincristine. The entire TIA pathway is fairly known at biochemical and genetic levels except the pathway steps leading to biosynthesis of catharanthine and tabersonine. To increase the in-planta yield of these antineoplastic metabolites for the pharmaceutical and drug industry, extensive plant tissue culture-based studies were performed to provide alternative production systems. However, the strict spatiotemporal developmental regulation of TIA biosynthesis has restricted the utility of these cultures for large-scale production. Therefore, the present study was performed to enhance the metabolic flux of TIA pathway towards the biosynthesis of vinblastine by overexpressing two upstream TIA pathway genes, tryptophan decarboxylase (CrTDC) and strictosidine synthase (CrSTR), at whole plant levels in C. roseus. Whole plant transgenic of C. roseus was developed using Agrobacterium tumefaciens LBA1119 strain having CrTDC and CrSTR gene cassette. Developed transgenic lines demonstrated up to twofold enhanced total alkaloid production with maximum ninefold increase in vindoline and catharanthine, and fivefold increased vinblastine production. These lines recorded a maximum of 38-fold and 65-fold enhanced transcript levels of CrTDC and CrSTR genes, respectively.

Genetic engineering approach using early Vinca alkaloid biosynthesis genes led to increased tryptamine and terpenoid indole alkaloids biosynthesis in differentiating cultures of Catharanthus roseus.

Catharanthus roseus today occupies the central position in ongoing metabolic engineering efforts in medicinal plants. The entire multi-step biogenetic pathway of its very expensive anticancerous alkaloids vinblastine and vincristine is fairly very well dissected at biochemical and gene levels except the pathway steps leading to biosynthesis of monomeric alkaloid catharanthine and tabersonine. In order to enhance the plant-based productivity of these pharma molecules for the drug industry, cell and tissue cultures of C. roseus are being increasingly tested to provide their alternate production platforms. However, a rigid developmental regulation and involvement of different cell, tissues, and organelles in the synthesis of these alkaloids have restricted the utility of these cultures. Therefore, the present study was carried out with pushing the terpenoid indole alkaloid pathway metabolic flux towards dimeric alkaloids vinblastine and vincristine production by over-expressing the two upstream pathway genes tryptophan decarboxylase and strictosidine synthase at two different levels of cellular organization viz. callus and leaf tissues. The transformation experiments were carried out using Agrobacterium tumefaciens LBA1119 strain having tryptophan decarboxylase and strictosidine synthase gene cassette. The callus transformation reported a maximum of 0.027% dry wt vindoline and 0.053% dry wt catharanthine production, whereas, the transiently transformed leaves reported a maximum of 0.30% dry wt vindoline, 0.10% catharanthine, and 0.0027% dry wt vinblastine content.

Differential rubisco content and photosynthetic efficiency of rol gene integrated Vinca minor transgenic plant: Correlating factors associated with morpho-anatomical changes, gene expression and alkaloid productivity.

Transgenic plants obtained from a hairy root line (PVG) of Vinca minor were characterized in relation to terpenoid indole alkaloids (TIAs) pathway gene expression and vincamine production. The hairy roots formed callus with green nodular protuberances when transferred onto agar-gelled MS medium containing 3.0mg/l zeatin. These meristematic zones developed into shoot buds on medium with 1.0mg/l 2, 4-dichlorophenoxyacetic acid and 40mg/l ascorbic acid. These shoot buds subsequently formed rooted plants when shifted onto a hormone-free MS medium with 6% sucrose. Transgenic nature of the plants was confirmed by the presence of rol genes of the Ri plasmid in them. The transgenic plants (TP) had elongated internodes and a highly proliferating root system. During glass house cultivation TP consistently exhibited slower growth rate, low chlorophyll content (1.02±0.08mg/gm fr. wt.), reduced carbon exchange rate (2.67±0.16µmolm-2s-1), less transpiration rate (2.30±0.20mmolm-2 s-1) and poor stomatal conductance (2.21±0.04mmolm-2 s-1) when compared with non-transgenic population. The activity of rubisco enzyme in the leaves of TP was nearly two folds less in comparison to non-transgenic controls (1.80milliunitsml-1mgprotein-1 against 3.61milliunits ml-1mgprotein-1, respectively). Anatomically, the TP had a distinct tetarch arrangement of vascular bundles in their stem and roots against a typical ployarched pattern in the non-transgenic plants. Significantly, the transgenic plants accumulated 35% higher amount of total TIAs (3.10±0.21% dry wt.) along with a 0.03% dry wt. content of its vasodilatory and nootropic alkaloid vincamine in their leaves. Higher productivity of alkaloids in TP was corroborated with more than four (RQ=4.60±0.30) and five (RQ=5.20±0.70) times over-expression of TIAs pathway genes tryptophan decarboxylase (TDC) and strictosidine synthase (STR) that are responsible for pushing the metabolic flux towards TIAs synthesis in this medicinal herb.