Design and development of mechanical test bench for testing and calibration of multiple blood pressure measuring devices.

Blood pressure (BP) measurement is an important physiological parameter for human health monitoring, which plays a significant role in the diagnosis of many incurable diseases. However, due to inaccuracies in the different types of BP measuring devices, the calibration of these BP measuring instruments is a major concern for a medical practitioner. Currently, these devices' calibration, testing, and validation are performed using rigorous methods with complex clinical trials and following the available documentary standards. This article describes the design and development of an indigenous mechanical test bench (MTB) system for the testing and calibration of multiple BP devices, as per International Organization of Legal Metrology (OIML) recommended documents e.g., OIML R 16-1 and OIML R 16-2. The developed system can test and calibrate 20 BP devices, simultaneously. The traceability of the developed MTB is established by performing its calibration against the Air Piston Gauge, a national primary vacuum standard. The estimated expanded measurement uncertainty evaluated is found to be ±0.11 mmHg, which is almost one order better than the measurement uncertainty required for the test and calibration of BP measuring instruments as per standard. The MTB has successfully been used to test and calibrate several BP measuring instruments. The data of one such device is reported herein as an indicator of the performance process. The calibration of these BP measuring instruments was performed in the static mode, and the estimated expanded measurement uncertainty was found to be ±1.25 mmHg. The developed MTB system would prove to be an excellent instrument for calibration laboratories, hospitals, regulatory agencies, and other users to test and calibrate 20 BP measuring devices simultaneously and cost-effectively.

Clonal Expansion of Stem/Progenitor Cells in Cancer, Fibrotic Diseases, and Atherosclerosis, and CD47 Protection of Pathogenic Cells.

We proposed and demonstrated that myelogenous leukemia has a preleukemic phase. In the premalignant phase, normal hematopoietic stem cells (HSCs) gradually accumulate mutations leading to HSC clonal expansion, resulting in the emergence of leukemic stem cells (LSCs). Here, we show that preleukemic HSCs are the basis of clonal hematopoiesis, as well as late-onset blood diseases (chronic-phase chronic myeloid leukemia, myeloproliferative neoplasms, and myelodysplastic disease). The clones at some point each trigger surface expression of "eat me" signals for macrophages, and in the clones and their LSC progeny, this is countered by upregulation of "don't eat me" signals for macrophages such as CD47,opening the possibility of CD47-based therapies. We include evidence that similar processes result in fibroblast expansion in a variety of fibrotic diseases, and arterial smooth muscle clonal expansion is a basis of atherosclerosis, including upregulation of both "eat me" and "don't eat me" molecules on the pathogenic cells.

A KHz frequency cold atmospheric pressure argon plasma jet for the emission of O(1S) auroral lines in ambient air.

Creation of the "auroral" green line, a fascinating occurrence commonly observed in the upper atmosphere, has long been a difficult endeavor, especially at atmospheric pressure. Here we report strong emission of the "auroral" green line for the first time in a kHz frequency, linear field atmospheric pressure plasma jet system. The device used 99.999% pure argon as a working gas for the plasma generation. Optical emission spectroscopy measurements of the after discharge region show the existence of 557.7 nm emission which corresponds to the transition O([Formula: see text]S)-O([Formula: see text]D). The intensity of the produced green line is strong enough that the entire plasma plume in the ambient air is visible as a green plasma. We provide the chemical reactions of O([Formula: see text]S) production in the plasma and the estimation of the density of the O([Formula: see text]S) metastable state using the kinetic reactions. Further, the O([Formula: see text]S) emission is characterized by changing the flow rate of argon, applied voltage and electrode gap. The adequate plasma length ([Formula: see text]) along with the production of a variety of reactive components viz; OH, [Formula: see text] and oxygen (777 nm) make this configuration useful for applications such as: blood coagulation, cancer treatment, sterilization, and waste treatment. Moreover, this setup can be potentially used as a test bed for the in-depth understanding of plasma chemistry relevant to the aurora and comet tails using a laboratory setting.

Genome-Wide Association Study and Post-genome-Wide Association Study Analysis for Spike Fertility and Yield Related Traits in Bread Wheat.

Spike fertility and associated traits are key factors in deciding the grain yield potential of wheat. Genome-wide association study (GWAS) interwoven with advanced post-GWAS analysis such as a genotype-phenotype network (geno-pheno network) for spike fertility, grain yield, and associated traits allow to identify of novel genomic regions and represents attractive targets for future marker-assisted wheat improvement programs. In this study, GWAS was performed on 200 diverse wheat genotypes using Breeders' 35K Axiom array that led to the identification of 255 significant marker-trait associations (MTAs) (-log10P ≥ 3) for 15 metric traits phenotyped over three consecutive years. MTAs detected on chromosomes 3A, 3D, 5B, and 6A were most promising for spike fertility, grain yield, and associated traits. Furthermore, the geno-pheno network prioritised 11 significant MTAs that can be utilised as a minimal marker system for improving spike fertility and yield traits. In total, 119 MTAs were linked to 81 candidate genes encoding different types of functional proteins involved in various key pathways that affect the studied traits either way. Twenty-two novel loci were identified in present GWAS, twelve of which overlapped by candidate genes. These results were further validated by the gene expression analysis, Knetminer, and protein modelling. MTAs identified from this study hold promise for improving yield and related traits in wheat for continued genetic gain and in rapidly evolving artificial intelligence (AI) tools to apply in the breeding program.

Sex-specific aortic root anatomy in patients with bicuspid aortic valve undergoing TAVR in a Chinese cohort.

OBJECTIVES: The aim of this study is to investigate the sex-specific aortic root anatomy in patients with bicuspid aortic valve (BAV). PATIENTS AND METHODS: This retrospective study includes 73 consecutive patients with BAV who underwent CT evaluation before transcatheter aortic valve replacement (TAVR) between July 2013 and April 2017 in the Second Affiliated Hospital of Zhejiang University. RESULT: The size of the annulus, diameter and height of the sinotubular junction (STJ), height of the coronary artery ostia, and dimension of the aorta were measured. Women had significantly smaller annulus parameters (mean diameter: 23.4 ± 1.8 vs. 26.1 ± 2.1 mm; area: 425.3 ± 59.4 vs. 527.4 ± 84.6 mm2; perimeter: 74.3 ± 5.2 vs. 83.2 ± 6.4 mm), and STJ diameter (29.7 ± 3.1 vs. 32.6 ± 4.5 mm) than men (p < 0.01 for all), even after adjustment for their smaller body surface area (BSA). Dimension of aorta and height of right coronary artery were also significantly smaller in women, although not when indexing for the BSA. The left ventricular ejection fraction of women is significantly higher than that of men before discharge (60.2 ± 9.7% vs. 53.7 ± 13.6%, p = 0.01). There were no differences between women and men in the all-cause 30-day and 1­year mortality. CONCLUSION: Women with BAV had smaller annulus and STJ diameter after indexing for BSA, reflecting a sex-specific difference. There were no differences in 30-day and 1­year mortality between the two groups.

New piperidine derivative DTPEP acts as dual-acting anti-breast cancer agent by targeting ERα and downregulating PI3K/Akt-PKCα leading to caspase-dependent apoptosis.

OBJECTIVES: In our ongoing studies to develop ER targeting agents, we screened for dual-acting molecules with a hypothesis that a single molecule can also target both ER positive and negative groups of breast cancer. MATERIALS AND METHODS: 1-(2-(4-(Dibenzo[b,f]thiepin-10-yl)phenoxy)ethyl)piperidine (DTPEP) was synthesized and screened in both MCF-7 (ER+ve) and MDA-MB-231 (ER-ve) cells. Assays for analysis of cell cycle, ROS, apoptosis and MMP loss were carried out using flow cytometry. Its target was investigated using western blot, transactivation assay and RT-PCR. In vivo efficacy of DTPEP was validated in LA-7 syngeneic rat mammary tumour model. RESULTS: Here, we report identification of dual-acting molecule DTPEP that downregualtes PI3K/Akt and PKCα expression, induces ROS and ROS-dependent apoptosis, loss of mitochondrial membrane potential, induces expression of caspase indicative of both intrinsic and extrinsic apoptosis in MCF-7 and MDA-MB-231 cells. In MCF-7 cells, DTPEP downregulates ERα expression and activation. In MDA-MB-231 cells, primary cellular target of DTPEP is not clearly known, but it downregualtes PI3K/Akt and PKCα expression. In vivo study showed regression of LA-7 syngeneic mammary tumour in SD rat. CONCLUSIONS: We identified a new dual-acting anti-breast cancer molecules as a proof of concept which is capable of targeting both ER-positive and ER-negative breast cancer.

Frontal Theta Asymmetry as a Biomarker of Depression.

INTRODUCTION: Electroencephalography (EEG) has been used extensively to study affective disorders. Quantitative spectral analysis of an EEG scan has been used to assess the biological basis of emotional disorders such as depression as well as to investigate biomarkers of affective disorders. Inter-hemispheric asymmetries in both baseline and stimulus-evoked frequencies (alpha, beta, theta, and delta) are potential biomarkers of depression. The role of frontal alpha asymmetry has been established, but other spectral frequencies such as frontal theta remain elusive. We compared the hemispheric differences in frontal theta power in depressed patients and controls before and during listening to music to study the correlation of frontal theta asymmetry with depression. METHODS: To determine whether stimulus-evoked frontal theta asymmetry is a biomarker of depression, we compared 23 patients with mild depression (based on the Hamilton Depression Rating Scale) with 17 age- and sex-matched controls by conducting EEG at rest and after listening to Indian classical music. RESULTS: In controls without depression, the mean frontal theta power of the left hemisphere and frontal theta asymmetry increased significantly during music listening. In depressed patients, frontal theta asymmetry was reversed during music listening. CONCLUSION: Frontal theta asymmetry is a potential biomarker of depression.

Acquired Constricting Band Syndrome.

The superstitious beliefs of gaining a good fortune led to wearing rubber bands around the hand leading to the development of the constricting band syndrome. Acute compartment syndrome is a limb-threatening condition characterized by elevated interstitial pressure in a confined fascial compartment and if left untreated, it results in tissue necrosis, irreversible nerve and muscle injury and permanent functional impairment. We report a 46 years old male with history of wearing rubber hand on all his fingers, who presented late with gangrene and autoamputation.

45° sign switching of effective exchange bias due to competing anisotropies in fully epitaxial Co3FeN/MnN bilayers.

We report an unusual angular-dependent exchange bias effect in ferromagnet/antiferromagnet bilayers, where both ferromagnet and antiferromagnet are epitaxially grown. Numerical model calculations predict an approximately 45° period for the sign switching of the exchange-bias field, depending on the ratio between magnetocrystalline anisotropy and exchange-coupling constant. The switching of the sign is indicative of a competition between a fourfold magnetocrystalline anisotropy of the ferromagnet and a unidirectional anisotropy field of the exchange coupling. This predicted unusual angular-dependent exchange bias and its magnetization switching process are confirmed by measurements on fully epitaxial Co3FeN/MnN bilayers by longitudinal and transverse magneto-optic Kerr effect magnetometry. These results provide a deeper understanding of the exchange coupling phenomena in fully epitaxial bilayers with tailored materials and open up a complex switching energy landscape engineering by anisotropies.

Curcumin mediated attenuation of carbofuran induced toxicity in the heart of Wistar rats.

Carbofuran is used to improve the agricultural productivity as well as to protect the house hold and industrial products, but due to accumulation in the biological system, it causes serious side effects in many non-targets mammalian systems. The aim of present study is to evaluate the carbofuran induced oxidative stress in rat heart and its attenuation by using herbal product curcumin. Rats were divided into four groups; one group received 20 % LD50 of carbofuran another group of rats received same doses of carbofuran was  pretreated with curcumin (100 mg kg-1 body weight) and remaining two other groups served as control and curcumin treated animals. The activity of lactate dehydrogenase (LDH) in the heart tissues and serum was evaluated and the activity of enzymatic antioxidants superoxide dismutase (SOD) and catalase (CAT) was estimated in the heart tissues. The level of malondialdehyde (MDA) in heart tissues was also measured. The Total cholesterol (TC) and high density lipoprotein (HDL) was measured in the serum of the entire animals group. The results of present study showed that the activity of LDH in heart tissues were decreased and in serum was elevated. The MDA level was significantly elevated due to exposure of carbofuran. The enzymatic antioxidants, SOD and CAT activities were also inhibited. The ratio of pro-oxidant (P)/antioxidant (A) was also found to be sharply increased in the rat heart tissues of carbofuran exposed animals. The alterations in all the parameter were recovered by the pretreatment of curcumin (100 mg kg-1 body weight).

Point prevalence & risk factor assessment for hospital-acquired infections in a tertiary care hospital in Pune, India.

BACKGROUND & OBJECTIVES: Hospital-acquired infections (HAIs) are a major challenge to patient safety and have serious public health implications by changing the quality of life of patients and sometimes causing disability or even death. The true burden of HAI remains unknown, particularly in developing countries. The objective of this study was to estimate point prevalence of HAI and study the associated risk factors in a tertiary care hospital in Pune, India. METHODS: A series of four cross-sectional point prevalence surveys were carried out between March and August 2014. Data of each patient admitted were collected using a structured data entry form. Centers for Disease Control and Prevention guidelines were used to identify and diagnose patients with HAI. RESULTS: Overall prevalence of HAI was 3.76 per cent. Surgical Intensive Care Unit (ICU) (25%), medical ICU (20%), burns ward (20%) and paediatric ward (12.17%) were identified to have significant association with HAI. Prolonged hospital stay [odds ratio (OR=2.81), mechanical ventilation (OR=18.57), use of urinary catheter (OR=7.89) and exposure to central air-conditioning (OR=8.59) had higher odds of acquiring HAI (P<0.05). INTERPRETATION & CONCLUSIONS: HAI prevalence showed a progressive reduction over successive rounds of survey. Conscious effort needs to be taken by all concerned to reduce the duration of hospital stay. Use of medical devices should be minimized and used judiciously. Healthcare infection control should be a priority of every healthcare provider. Such surveys should be done in different healthcare settings to plan a response to reducing HAI.

Probing ultrafast changes of spin and charge density profiles with resonant XUV magnetic reflectivity at the free-electron laser FERMI.

We report the results of resonant magnetic XUV reflectivity experiments performed at the XUV free-electron laser FERMI. Circularly polarized XUV light with the photon energy tuned to the Fe M2,3 edge is used to measure resonant magnetic reflectivities and the corresponding Q-resolved asymmetry of a Permalloy/Ta/Permalloy trilayer film. The asymmetry exhibits ultrafast changes on 240 fs time scales upon pumping with ultrashort IR laser pulses. Depending on the value of the wavevector transfer Qz , we observe both decreasing and increasing values of the asymmetry parameter, which is attributed to ultrafast changes in the vertical spin and charge density profiles of the trilayer film.

Choline metabolic pathway gene polymorphisms and risk for Down syndrome: An association study in a population with folate-homocysteine metabolic impairment.

BACKGROUND/OBJECTIVES: Choline is an essential nutrient involved in one-carbon metabolism, but its role in mechanisms underlying meiotic non-disjunction is poorly known. The relationship between folate-homocysteine metabolic pathway gene polymorphism and Down syndrome (DS) risk has been widely analyzed, but there are limited reports on its correlation with choline metabolism. In the present case-control association study, we investigated the relationship of three single-nucleotide polymorphisms (SNPs) (phosphatidylethanolamine N-methyltransferase (PEMT) rs12325817, choline dehydrogenase (CHDH) rs12676 and homocysteine methyltransferase (BHMT) rs3733890) of choline metabolism with risk for DS. SUBJECT/METHODS: Genotyping of 228 mothers of a down syndrome child (DSM) and 200 control mothers (CMs) for all SNPs was performed by PCR coupled with restriction fragment length polymorphism method. RESULTS: A significantly increased risk for BHMT +742AA genotype with an odds ratio of 4.96 (95% confidence interval (CI): 1.66-14.88, P=0.0036) was observed. For PEMT rs12325817 and CHDH rs12676, no significant difference in allelic and genotypic frequencies was observed. In genotypic combination analysis considering PEMT -744GG/CHDH +432GG/BHMT +742GG as the reference combination, PEMT -744GC/CHDH +432GG/BHMT +742GG genotypic combination was significantly higher in DSM compared with that in CMs with an odds ratio of 2.061 (95% CI: 1.10-3.86, P=0.0342). We also observed an epistatic interaction between methylenetetrahydrofolate reductase (MTHFR) rs1801133 and choline metabolic pathway gene variants. CONCLUSIONS: Our findings indicate impaired choline metabolism showing a greater risk for DS, especially in a population associated with homocysteine-folate impairment. Further studies are required to confirm our findings.

Stressful life events, hopelessness, and suicidal intent in patients admitted with attempted suicide in a tertiary care general hospital.

BACKGROUND: Suicide is a psychiatric emergency. Stressors in life and social variables (like marital status, family, and social support) are among the determinants of suicide. Hopelessness and suicidal intent are among the psychological variables that have shown promise in the prediction of suicide. AIMS AND OBJECTIVES: To assess stressful life events, hopelessness, suicidal intent, and sociodemographic variables in patients of attempted suicide. MATERIALS AND METHODS: Fifty consecutive patients admitted with attempted suicide were interviewed. Presumptive Stressful Life Event Scale, Beck Hopelessness Scale, and Beck Suicidal Intent Scale were used along with a semistructured pro forma for interview. Data were analyzed with statistical tests. RESULTS: Sixty-six percent of the participants were females, 72% were less than 30 years of age. Sixty-six percent of the patients had stressful life event score between 101 and 200 with the mean score of 127. The stressful life event score in those who considered they are in need of psychiatric help was significantly high. Most of the patients had mild (34%) and moderate (40%) degrees of hopelessness, and the mean score was 9.64. The mean suicidal intent in the participants was 25.14, when correlated with hopelessness score significant positive correlation was found. CONCLUSION: Lethality of the attempt increases with the increase in hopelessness.

Dusty Plasma Experimental (DPEx) device for complex plasma experiments with flow.

A versatile table-top dusty plasma experimental device to study flow induced excitations of linear and nonlinear waves/structures in a complex plasma is presented. In this Π-shaped apparatus, a DC glow discharge plasma is produced between a disc shaped anode and a grounded long cathode tray by applying a high voltage DC in the background of a neutral gas (argon) and subsequently a dusty plasma is created by introducing micron sized dust particles that get charged and levitated in the sheath region. A flow of the dust particles is induced in a controlled manner by adjusting the pumping speed and the gas flow rate into the device. A full characterisation of the plasma, using Langmuir and emissive probe data, and that of the dusty plasma using particle tracking data with the help of an idl based (super) Particle Identification and Tracking (sPIT) code is reported. Experimental results on the variation of the dust flow velocity as a function of the neutral pressure and the gas flow rate are given. The neutral drag force acting on the particles and the Epstein coefficient are estimated from the initial acceleration of the particles. The potential experimental capabilities of the device for conducting fundamental studies of flow induced instabilities are discussed.

Leishman and Giemsa stain: a new reliable staining technique for blood/bone marrow smears.

INTRODUCTION: Peripheral blood and bone marrow smear examination is an important basic tool for the diagnosis of different haematological conditions including haematological malignancies. We created a newer modification of the conventional Leishman and Giemsa stains as Leishman and Giemsa (L&G) stain and compared the efficacy and reliability of this stain with conventional stains. The study was performed to evaluate the staining efficacy, feasibility, time and cost of L&G stain over the conventional Leishman and Giemsa stains. METHODS: A pilot study was carried out in the Department of Haematology of our hospital from October 2013 to December 2013. Hundred selected cases, each with peripheral blood and bone marrow smears were taken, and three sets of the smears were prepared from each sample--one for L&G stain and other two--one each for conventional Leishman and Giemsa stains. This staining is further incorporated in our routine standard operating protocols for staining of all the peripheral blood smears in automated stainer, Sysmex SP10. RESULT: The average grading score from each staining methods from all the three experts was compiled. The average grading score of L&G staining method was noted to be significantly higher than the other two methods (analysis of variance test, P value < 0.05). When modified L&G stain (C) was compared with stain conventional stains (A and B), a P value of <0.001 was noted in all parameters except between Leishman stain and L&G stain in mature RBC and WBC nucleus and RBC inclusions (P value between 0.05 and 0.001). CONCLUSION: L&G staining is a newer staining technique of immense help in high-throughput haematology laboratories by offering a time-saving, cost-effective and better staining option to conventional staining methods. It gives a better nuclear and cytoplasmic differential staining and can also be used in automated blood counters/stainer.

Role of folate-homocysteine pathway gene polymorphisms and nutritional cofactors in Down syndrome: A triad study.

STUDY QUESTION: Do gene-gene and gene-environment interactions in folate-homocysteine (Hcy) pathway have a predisposing role for Down syndrome (DS)? SUMMARY ANSWER: The study provides evidence that in addition to advanced age, maternal genotype, micronutrient deficiency and elevated Hcy levels, individually and in combination, are risk factors for Down syndrome. WHAT IS KNOWN ALREADY: Polymorphisms in certain folate-Hcy-pathway genes (especially the T allele of MTHFR C677T), elevated Hcy and poor folate levels in mothers during pregnancy have been shown to be risk factors for Down syndrome in certain Asian populations (including the eastern region of India), while the same SNPs are not a risk factor in European populations. This conflicting situation alludes to differential gene-environment (nutrition) interactions in different populations which needs to be explored. STUDY DESIGN, SIZE, DURATION: Between 2008 and 2012, 151 Down syndrome triads and 200 age-matched controls (Control mothers n = 186) were included in the study. Seven polymorphisms in six genes of folate-Hcy metabolic pathway, along with Hcy, cysteine (Cys), vitamin B12 (vit-B12) and folate levels, were analysed and compared among the case and control groups. PARTICIPANTS/MATERIALS, SETTING, METHODS: Genotyping was performed by the PCR-RFLP technique. Levels of homocysteine and cysteine were measured by HPLC while vitamin B12 and folate were estimated by chemiluminescence. MAIN RESULTS AND THE ROLE OF CHANCE: We demonstrate that polymorphisms in the folate-Hcy pathway genes in mothers collectively constitute a genotypic risk for DS which is effectively modified by interactions among genes and by the environment affecting folate, Hcy and vitamin B12 levels. The study also supports the idea that these maternal risk factors provide an adaptive advantage during pregnancy supporting live birth of the DS child. LIMITATIONS AND REASONS FOR CAUTION: Our inability to obtain genotype and nutritional assessments of unaffected siblings of the DS children was an important limitation of the study. Also, its confinement to a specific geographic region (the eastern part) of India, and relatively small sample size is a limitation. A parallel investigation on another population could add greater authenticity to the data. WIDER IMPLICATIONS OF THE FINDINGS: For mothers genetically susceptible to deliver a DS child (particularly in South Asia), peri-conceptional nutritional supplementation and antenatal care could potentially reduce the risk of a DS child. Additionally, nutritional strategies could possibly be used for better management of the symptoms of DS children. STUDY FUNDING/COMPETING INTERESTS: The work is funded through Programme support for Genetic disorders by Department of Biotechnology, Government of India to R.R. The authors declare no conflict of interest.

Amoeboma-lest we forget!!!

Amoebiasis is an infection with the parasitic intestinal protozoan Entamoeba histolytica (E. histolytica). Most infections are usually asymptomatic, but the disease spectrum can range from dysentery to extraintestinal infections, including liver abscesses. The management of a 68-year-old male with amoeboma, a rare manifestation of intestinal amoebiasis, is described along with a review of literature.

Antimicrobial use and antimicrobial resistance in nosocomial pathogens at a tertiary care hospital in Pune.

BACKGROUND: Resistance to antimicrobial agents is emerging in wide variety of nosocomial and community acquired pathogens. Widespread and often inappropriate use of broad spectrum antimicrobial agents is recognized as a significant contributing factor to the development and spread of bacterial resistance. This study was conducted to gain insight into the prevalent antimicrobial prescribing practices, and antimicrobial resistance pattern in nosocomial pathogens at a tertiary care hospital in Pune, India. METHODS: Series of one day cross sectional point prevalence surveys were carried out on four days between March and August 2014. All eligible in patients were included in the study. A structured data entry form was used to collect the data for each patient. Relevant samples were collected for microbiological examination from all the clinically identified hospital acquired infection cases. RESULTS: 41.73% of the eligible patients (95% CI: 39.52-43.97) had been prescribed at least one antimicrobial during their stay in the hospital. Beta-lactams (38%) were the most prescribed antimicrobials, followed by Protein synthesis inhibitors (24%). Majority of the organisms isolated from Hospital acquired infection (HAI cases) were found to be resistant to the commonly used antimicrobials viz: Cefotaxime, Ceftriaxone, Amikacin, Gentamicin and Monobactams. CONCLUSION: There is need to have regular antimicrobial susceptibility surveillance and dissemination of this information to the clinicians. In addition, emphasis on the rational use of antimicrobials, antimicrobial rotation and strict adherence to the standard treatment guidelines is very essential.