A study of salivary opiorphin levels using different anesthetic drugs and techniques - A randomized controlled clinical study.

PURPOSE: This study aimed at assessing the change in salivary opiorphin levels before and after administration of local anesthesia, with the use of three different local anesthetic agents, and different anaesthetic techniques. METHODS: The investigators implemented a randomized controlled clinical study in 144 patients who required tooth extraction after administration of local anaesthesia. A total of 288 samples were collected in sterile containers before and after administration of local anesthetics. The salivary samples were centrifuged and salivary opiorphin levels were estimated using ELISA testing and spectrophotometric analysis. Statistical analysis was done using one way ANOVA and unpaired t test. RESULTS: There was a mean decrease in salivary opiorphin levels after administration of local anesthesia. There was no significant difference in the change in salivary opiorphin levels across different anesthetic techniques and different drug subgroups. CONCLUSION: The present study did not show much association between various local anesthetic agents and techniques and change in salivary opiorphin levels. The role of opiorphin as a biomarker for pain control and its effect on various pain control methods including local anesthesia must be evaluated in detail. Institutional review board number SRMDC/IRB/2014/MDS/No. 405.

Totally drug-resistant tuberculosis and adjunct therapies.

The first cases of totally drug-resistant (TDR) tuberculosis (TB) were reported in Italy 10 years ago; more recently, cases have also been reported in Iran, India and South Africa. Although there is no consensus on terminology, it is most commonly described as 'resistance to all first- and second-line drugs used to treat TB'. Mycobacterium tuberculosis (M.tb) acquires drug resistance mutations in a sequential fashion under suboptimal drug pressure due to monotherapy, inadequate dosing, treatment interruptions and drug interactions. The treatment of TDR-TB includes antibiotics with disputed or minimal effectiveness against M.tb, and the fatality rate is high. Comorbidities such as diabetes and infection with human immunodeficiency virus further impact on TB treatment options and survival rates. Several new drug candidates with novel modes of action are under late-stage clinical evaluation (e.g., delamanid, bedaquiline, SQ109 and sutezolid). 'Repurposed' antibiotics have also recently been included in the treatment of extensively drug resistant TB. However, because of mutations in M.tb, drugs will not provide a cure for TB in the long term. Adjunct TB therapies, including therapeutic vaccines, vitamin supplementation and/or repurposing of drugs targeting biologically and clinically relevant molecular pathways, may achieve better clinical outcomes in combination with standard chemotherapy. Here, we review broader perspectives of drug resistance in TB and potential adjunct treatment options.

Inflammation and tuberculosis: host-directed therapies.

Tuberculosis (TB) is an airborne infectious disease that kills almost two million individuals every year. Multidrug-resistant (MDR) TB is caused by strains of Mycobacterium tuberculosis (M. tb) resistant to isoniazid and rifampin, the backbone of first-line antitubercular treatment. MDR TB affects an estimated 500,000 new patients annually. Genetic analysis of drug-resistant MDR-TB showed that airborne transmission of undetected and untreated strains played a major role in disease outbreaks. The need for new TB vaccines and faster diagnostics, as well as the development of new drugs, has recently been highlighted. The major problem in terms of current TB research and clinical demands is the increasing number of cases of extensively drug-resistant and 'treatment-refractory' TB. An emerging scenario of adjunct host-directed therapies is intended to target pulmonary TB where inflammatory processes can be deleterious and lead to immune exhaustion. 'Target-organ-saving' strategies may be warranted to prevent damage to infected tissues and achieve focused, clinically relevant and long-lasting anti-M. tb cellular immune responses. Candidates for such interventions may be biological agents or already approved drugs that can be 're-purposed' to interfere with biologically relevant cellular checkpoints. Here, we review current concepts of inflammation in TB disease and discuss candidate pathways for host-directed therapies to achieve better clinical outcomes.

True facets of TB diagnosis in 2012: Hypes and realities.

Tuberculosis takes a heavy toll of ~5000 lives every day from the disease; responsible for the 86% of DALY burden. Despite having drugs to treat TB efficiently, we have failed to control the disease. Mycobacterium tuberculosis has exploited it to their advantage evolving with multiple mutations making it resistant to first-line and second-line drugs. Most of the high-burden countries are low-medium income countries, their national TB program (NTP) still use sputum smear microscopy as the tool of diagnosis. Many new molecular tools are emerging, but confuse the larger TB clinical scientific community at the NTPs. Coherent information need to be disseminated, encouraging TB scientific community to generate evidences within NTPs assessing new tools through critical analyses in terms of value addition and cost benefit before considering rolling out in the program. It is also imperative that the scientific community need to have an open mind to use different tools in the right permutation and combination than being exclusive of one another. This article portrays an overview of the diagnostics landscape in 2012 with pros and cons of different tools to be able to generate a step-wise algorithm for optimal exploitation of the tools within available resources in each of the settings.

Effect of construction of an irrigation canal on malaria situation in two primary health centres of Dhenkanal district of Orissa, India.

To assess the impact of irrigation canals on malaria transmission, a study was conducted in Dhenkanal district of Orissa, India. The district is situated in the central part of Orissa and hyperendemic area for malaria. A canal system is being constructed for irrigation in the district, which passes through Parjang and Analabereni Primary Health Centres (PHC), endemic for malaria. The water has been released only up to Parjang (Canal with water -CWW) area during the end of 2004 and construction work is still going on in Analabereni PHC (Canal under construction-CUC). Retrospective clinical data (2001-2008) collected from health services from two study sites showed average Slide Positivity Rate (SPR) before release of water (2001-2004) was 9.25% and 18.04% in CWW and CUC areas, respectively. After release of water (2005-2008) the SPR was 5.77% and 10.19%, in CWW and CUC areas, respectively. The average Annual Parasite Incidence (API) was 7.66 and 22.67 in CWW and CUC areas before the release of water and 5.32 and 12.28 after release of water, respectively. A point fever survey was conducted in 2009 which revealed the presence of Plasmodium falciparum (Pf) and P. vivax (Pv) in both study areas. The survey found SPR of 18.82% and 24.54%, and Pf percentages of 75% and 85%, in CWW and CUC areas, respectively. The present study revealed the presence of two malaria vectors, Anopheles culicifacies and Anopheles annularis in the area. Vector Per Man Hour Density was 2.38 in CWW and 2.69 in CUC for An. culicifacies and 1.46 and 1.54 for An. annularis respectively. The sporozoites rates were found to be 3.6 and 3.8 for CWW and CUC, respectively. The present study reveals that, the construction of canal system did not increase the malaria prevalence during post water release period - implying that the malaria control programme was effective although still more intensive situation specific vectors control programme need to be continued simultaneously so that malaria transmission can be curtailed.

Rapid spread of chikungunya virus infection in Orissa: India.

BACKGROUND & OBJECTIVES: A large number of cases of undiagnosed fever and joint pain were reported from different parts of the State of Orissa since February 2006. Epidemiological and laboratory investigation were carried out to confirm the cause of emerging illness, which was provisionally suspected as Chikungunya (CHIK) fever. METHODS: Upon getting the reports of suspected CHIK like illness in different parts of the State, epidemic investigations were carried out in the outbreak affected villages. Case history was recorded, clinical examination undertaken and blood samples collected for seroconfirmation for CHIK IgM antibody using ELISA based kit. Simultaneously vector survey was also carried out. RESULTS: With no previous record of CHIK infection in the State, the first outbreak was confirmed during February 2006. Subsequently, the infection spread to 13 of 30 districts in different episodes covering 79 villages till November 2007. Attack rate was 9-43 per cent in the different outbreaks with average seropositivity of 24 per cent to CHIK specific IgM. Morbidity was high though no deaths were recorded. Aedes aegypti and Ae. albopictus were identified as the possible vectors for transmission. INTERPRETATION & CONCLUSIONS: The report confirmed emergence of CHIK infection in the State of Orissa, India, and its spread to a larger geographic zone in a short period which warrants public health measures to control further spread.

Human gene expression profiles of susceptibility and resistance in tuberculosis.

Tuberculosis (TB) still poses a profound burden on global health, owing to significant morbidity and mortality worldwide. Although a fully functional immune system is essential for the control of Mycobacterium tuberculosis infection, the underlying mechanisms and reasons for failure in part of the infected population remain enigmatic. Here, whole-blood microarray gene expression analyses were performed in TB patients and in latently as well as uninfected healthy controls to define biomarkers predictive of susceptibility and resistance. Fc gamma receptor 1B (FCGRIB)was identified as the most differentially expressed gene, and, in combination with four other markers, produced a high degree of accuracy in discriminating TB patients and latently infected donors. We determined differentially expressed genes unique for active disease and identified profiles that correlated with susceptibility and resistance to TB. Elevated expression of innate immune-related genes in active TB and higher expression of particular gene clusters involved in apoptosis and natural killer cell activity in latently infected donors are likely to be the major distinctive factors determining failure or success in controlling M. tuberculosis infection. The gene expression profiles defined in this study provide valuable clues for better understanding of progression from latent infection to active disease and pave the way for defining predictive correlates of protection in TB.

The development and evaluation of a single step multiplex PCR for simultaneous detection of Anopheles annularis group mosquitoes, human host preference and Plasmodium falciparum sporozoite presence.

The Anopheles annularis group mosquitoes, subgenus Cellia Theobald (Diptera: Culicidae), includes five recognized species: An. annularis Van der Wulp, An. nivipes Theobald, An. pallidus Theobald, An. philippinensis Ludlow and An. schueffneri Stanton. From these five, the three most common species found in Orissa were considered for this study because of their remarkable vectorial and behavioral variation and the important role they play in malaria transmission. To identify and understand their role in malaria transmission we developed a single multiplex PCR-based assay. This assay included the detection of human blood feeding habit and Plasmodium falciparum sporozoite presence. Of the 186 An. annularis mosquitoes collected, morphological character-based identification showed that 94 were An. annularis, 54 were An. philippinensis and 38 were An. pallidus. However, the multiplex PCR assay confirmed that 91 were An. annularis, 56 were An. philippinensis and 39 were An. pallidus individuals after adjustments were made for misidentified specimens in the morphological method. Anopheles annularis and An. philippinensis were found positive for human blood, and two samples of An. annularis species were positive for P. falciparum sporozoites. This one-step PCR-based method constitutes a very powerful tool in large surveys of anopheline populations.

Host feeding patterns of malaria vectors of Orissa, India.

Blood meals of 1,491 Anopheles fluviatilis sensu lato (s.l.), 1,690 An. culicifacies s.l., 719 An. annularis s.l., and 358 An. varuna sensu stricto were examined by gel diffusion method. The overall anthropophilic index (AI) was 78.9%, 1.6%, 3.2%, and 6.7% for An. fluviatilis, An. culicifacies, An. annularis, and An. varuna, respectively. Out of 4 anopheline species studied, only 0.2% of An. culicifacies blood meals contained blood from humans and cattle. Anopheles fluviatilis and An. culicifacies revealed seasonality in their anthropophilic index. An. fluviatilis showed a human forage ratio of more than 1, whereas An. culicifacies, An. annularis, and An. varuna had forage ratios of 2.6, 2.5, and 2.4, respectively, for bovine. There was a correlation between the Al of An. fluviatilis and the malaria slide positivity rate. This study suggests that the use of repellent, insecticide-treated nets will be effective for controlling biting mosquitoes inside houses in Orissa.

Immunoprophylactic effects of the anti-leprosy Mw vaccine in household contacts of leprosy patients: clinical field trials with a follow up of 8-10 years.

We report here a large scale, double blind immunoprophylactic trial of a leprosy vaccine based on Mycobacterium w (Mw) in an endemic area of Kanpur Dehat, Uttar Pradesh, India. A population of 420,823 spread over 272 villages was screened where 1226 multibacillary (MB) and 3757 paucibacillary (PB) cases of leprosy were detected. A total of 29,420 household contacts (HHC) of these patients were screened for evidence of active or inactive leprosy. After exclusion of 1622 contacts for any of the different exclusion criteria, a total of 24,060 HHC could be vaccinated for vaccine or placebo under coding (20,194 administered two doses and 3866 received single dose). The vaccine consisted of 1 x 10(9) heat killed bacilli (Mw) in normal saline for the first dose and half of the first dose, i.e. 5 x 10(8) bacilli for the second dose, given 6 months after the first dose. The placebo consisted of 1/8th dose of the normal dose of tetanous toxoid. Both placebo and vaccine were given under double-blind coding, The contacts were followed up during three surveys at 3, 6 and 9 years after the initial vaccination, for detection of post-vaccination cases (PVCs) and observing any side-effects caused as a result of vaccination. The codes were opened on 24th January 2001, after the analysis of the data following completion of the third and final follow-up survey. When only contacts received the vaccine, Mw vaccine showed a protective efficacy (PE) of 68-6% at the end of first, 59% at the end of the second and 39.3% at the end of the third follow-up survey. When both patients and contacts received the vaccine, the protective efficacy observed was 68%, 60% and 28% at the end of the first, second and third surveys, respectively. When patients, and not the contacts, received the vaccine, a PE of 42.9% in the first, 31% in the second and 3% in the third survey was shown. These results suggest that the vaccination of the contacts is more valuable in achieving the objective of immunoprophylaxis than that of patients, and the vaccine effects are noted maximally in children (as compared to adolescents and adults) who constitute the most responsive group The effect of vaccine is sustained for a period of about 7-8 years, following which there is a need to provide a booster vaccination for the sustained protection.

Two distinct phospholipases C of Listeria monocytogenes induce ceramide generation, nuclear factor-kappa B activation, and E-selectin expression in human endothelial cells.

Infection of endothelial cells by Listeria monocytogenes is an essential step in the pathogenesis of listeriosis. We recently reported that L. monocytogenes induces up-regulation of E-selectin and other endothelial adhesion molecules and subsequent polymorphonuclear leukocyte (PMN) adhesion into cultured human endothelial cells. In the present study, we characterized the mechanisms of enhanced E-selectin expression using L. monocytogenes wild type (EGD), the isogenic in-frame deletion mutants for phosphatidylcholine (PC)- and phosphatidylinositol (PI)-specific phospholipases EGD delta plcA and EGD delta plcB, as well as the nonvirulent control strain Listeria innocua. Infection of endothelial cells with EGD delta plcA or EGD delta plcB for 6 h induced, as compared with EGD wild type, intermediate levels of E-selectin mRNA and protein as well as PMN rolling and adhesion at a shear rate of 1 dyne/cm2, indicating that both bacterial phospholipases are required for a maximal effect. Similarly, ceramide content and NF-kappa B activity were increased in L. monocytogenes-exposed endothelial cells, but only to intermediate levels for PC- or PI-phospholipase C (PLC)-deficient listerial mutants. Phospholipase effects could be mimicked by exogenously added ceramides or bacterial sphingomyelinase. The data presented indicate that PI-PLC and PC-PLC are important virulence factors for L. monocytogenes infections that induce accumulation of ceramides that in turn may act as second messengers to control host cell signal-transduction pathways leading to persistent NF-kappa B activation, increased E-selectin expression, and enhanced PMN rolling/adhesion. The ability of L. monocytogenes to stimulate PMN adhesion to endothelial cells may be an important mechanism in the pathogenesis of severe listeriosis.

Internalin B is essential for adhesion and mediates the invasion of Listeria monocytogenes into human endothelial cells.

Listeria monocytogenes causes rhombencephalitis in humans and animals and also affects the fetus in utero, causing disseminated sepsis. In both instances, the infection occurs by the crossing of endothelial cells lining a physiological barrier, the blood-brain barrier or the transplacental barrier. In this study, the ability of L. monocytogenes wild-type EGD to invade human umbilical vein endothelial cells (HUVECs) was evaluated using wild-type bacteria and isogenic Listeria mutants. Here, we show that invasion of HUVECs by L. monocytogenes is dependent on the expression of the internalin B gene product. This was demonstrated in several ways. First, L. monocytogenes strains lacking the inlB gene did not invade HUVECs. Secondly, avid invasion was obtained when a strain deleted for inlAB was complemented with a plasmid harbouring inlB only, whereas strains expressing inlA did not enter HUVECs. Thirdly, entry of wild-type EGD could be blocked effectively with antibodies to InlB. Fourthly, cell binding assays and flow cytometry with HUVECs showed binding of purified InlB, but not InlA, suggesting a tropism of InlB for this cell type. Finally, physical association of purified native InlB with the surface of non-invasive mutants dramatically increased their ability to invade HUVECs. In laser-scanning confocal microscopy, binding of InlB was observed as focal and localized patches on the cell surface of HUVECs. Qualitative examination of the entry process by scanning electron microscopy revealed that both wild-type EGD and a recombinant strain overexpressing only InlB enter HUVECs in a similar fashion. The entry process was polarized, involved single bacteria and occurred over the entire surface of endothelial cells.

Elevated circulating calcitonin gene-related peptide in umbilical cord and infant blood associated with maternal and neonatal sepsis and shock.

The role of the sensory neuropeptide calcitonin gene-related peptide (CGRP) was studied in preterm and term neonates with sepsis and shock. CGRP levels in blood were measured by RIA. The identity of immunoreactive CGRP (irCGRP) in adult and infant human blood was confirmed by reverse phase-HPLC. CGRP levels were analyzed in a total of 189 samples (95 from cord blood and 94 from neonates). The gestational ages ranged from 24 to 43 wk, and the birth weights ranged from 520 to 4445 g. Cord samples were collected immediately after delivery and infant blood samples were collected within 12 h of birth. Samples were coded, and the data were assigned to groups after determination of CGRP levels. There was a weight- and gestation-dependent increase in irCGRP in the newborn population. The direct correlation of circulating CGRP with ascending birth weight and gestation may have significance in the development of the fetus. Infants with and without certain complications were grouped in 500-g intervals. CGRP levels in cord blood were significantly elevated when certain stressful situations existed in the mother. These included culture-positive chorioamnionitis, placental abruption, and severe preeclampsia. There was a similar elevation in CGRP in patient blood in infants with culture-positive sepsis and/or shock with blood pressure <2 SD from the mean for corresponding gestation. CGRP levels did not differ between male and female infants and did not appear to be influenced by type of delivery (vaginal versus cesarean section). There was no significant difference in CGRP level between cord and patient blood in preterm neonates, but at term gestation cord blood levels were slightly higher than those in the patient blood. These results suggest that inflammation and hemodynamic imbalance (e.g. shock) are associated with increased in CGRP levels in the circulation in neonates. Future studies will focus on the biologic effects of elevated CGRP during neonatal complications and will examine the utility of CGRP measurement for diagnosis and treatment of disease in preterm infants.

Hiatus/paraesophageal hernias in neonatal Marfan syndrome.

We report on an infant with neonatal Marfan syndrome (NMS) and hiatus/paraesophageal hernia who presented to a university hospital with an unusual early complication of this connective tissue disorder. An abnormal course of the nasogastric tube was noted on the first day of life by a radiograph of the chest and abdomen performed for bloody gastric drainage. The question of esophageal perforation was raised. Subsequent contrast study demonstrated a large hiatus/paraesophageal hernia with pronounced gastroesophageal reflux (GER). A part of the hernia was positioned posterior and to the right of the gastroesophageal junction (GEJ), presumably the location of the nasogastric tube as noted on the initial films. Although characterized by cardiac/aortic abnormalities, NMS can be a difficult diagnosis and should be considered in any infant with hiatus/ paraesophageal hernia with or without GER.

Endotracheal tolazoline administration in neonates with persistent pulmonary hypertension.

OBJECTIVES: Our purpose was to study the effectiveness of endotracheal tolazoline (ET-Tz) in the treatment of neonatal persistent pulmonary hypertension (PPHN). STUDY DESIGN: ET-Tz was administered to 12 neonates with a clinical diagnosis of PPHN. The gestational age ranged from 25 to 42 weeks, and the birth weights from 850 to 3612 gm. The dose of tolazoline ranged from 1 to 2.5 mg/kg. RESULTS: There was a significant increase (p < 0.005) in the mean levels of oxygen saturation and the arterial oxygen tension, and a significant decrease (p < 0.005) in the oxygenation index, between the pretolazoline and the posttolazoline groups, but arterial carbon dioxide tension did not change. After the initial analysis, the groups were subdivided into preterm and term subgroups, because we secondarily observed that the average changes from predose to postdose levels in the above parameters were significantly different (p < 0.001) in the two subgroups by Student's paired t test. CONCLUSIONS: The data indicate that ET-Tz is effective in improving oxygenation in neonates with PPHN, particularly sick preterm infants. The endotracheal route is preferred because it is devoid of significant side effects (e.g., hypotension and flushing). A randomized, controlled, double-blinded, multicenter trial for the use of ET-Tz in PPHN is warranted.

Penoscrotal transposition and associated anomalies: report of five new cases and review of the literature.

We present the largest single series of cases (n = 5) of penoscrotal transposition (PST) with carefully documented nongenitourinary/anal anomalies, none of which fell into categories of known syndromes, associations, sequences or chromosome disorders. Several unexpected anomalies were observed including coloboma of the iris and retina, hydrocephalus, microcephaly, diaphragmatic hernia, tracheo-esophageal fistula/esophageal atresia and cleft palate. The most frequent anomalies other than PST were renal defects (100%) such as renal agenesis and dysplasia, imperforate anus (60%), central nervous system anomalies (60%) and preaxial upper limb defects (40%). Cardiovascular defects (atrial septal defect, double aortic arch with vascular ring) were noted in only one case. The surviving patients (3/5) had postnatal growth failure and mental retardation. Our 5 PST patients are compared to 16 well-documented cases from the literature. The overall incidence of various extragenital abnormalities were: renal (90%), mental retardation (60%), imperforate anus (33%), central nervous system (CNS) anomalies (29%), vertebral defects (29%), preaxial limb defects (24%) and congenital heart disease (19%). PST is a rare heterogenous anomaly, the detection of which should warrant careful clinical evaluation to rule out other anomalies, especially of the urinary system, gastrointestinal tract, upper limbs, craniofacial region and central nervous system. PST may be a localized field defect involving the genitourinary system; however, the wide variety of more distant defects noted in our series and the literature would raise doubt about that assumption. The high frequency of growth deficiency and mental retardation has also not been given due respect as accompanying problems associated with PST.

Observations on nocturnal activity and man biting habits of malaria vectors, Anopheles fluviatilis, An. annularis and An. culicifacies in the hill tracts of Koraput District, Orissa, India.

Biting and feeding behavior of malaria vectors were studied in nine villages (5 from Jeypore zone and 4 from Malkangiri zone) of Koraput District. Man biting catches comprised of 16 anopheline species including the incriminated vectors of this area: An. fluviatilis, An. annularis and An. culicifacies. An. fluviatilis was predominant and biting of this species recorded indoors throughout the year in both the zones. The period, during which the biting activity peaked, was different between the two zones and consequently the time of peak transmission was also different between the zones. The biting activity was at its peak between 21.00 and 03.00 hours in both the zones. However, in cold season the biting activity peaked in the first quarter of the night in Jeypore zone. The anthropophilic index (AI) of An. fluviatilis was 26.2% in Jeypore and 83.7% in Malkangiri and of An. culicifacies the AI was 0.4% in Jeypore and 7% in Malkangiri. Analysis of gonotrophic stages of night resting females indicated that in Jeypore zone, the majority of An. fluviatilis female left indoors for outdoor resting before the completion of gonotrophic cycle, but in Malkangiri, the females remained indoors till the end of the gonotrophic cycle. The presence of full-fed females in night resting catches in Jeypore village further suggested that the females do not leave the house immediately after taking blood meal but rest for some time.

Field trials on the use of Mycobacterium w vaccine in conjunction with multidrug therapy in leprosy patients for immunotherapeutic and immunoprophylactic purposes.

A double blind field trial was started with a candidate anti-leprosy vaccine, Mycobacterium w as an immunotherapeutic and immunoprophylactic agent against leprosy in a highly endemic region with a prevalence rate of over 18 per 1000 population. By 31 August 1992, 224 villages have been surveyed, covering a population of 307,981 (1981 census). A total of 979 MB patients and 2801 PB patients have been registered. A total of 19,453 household contacts of leprosy patients have been examined for clinical signs of disease, of which 16,519 have received the initial dose while 10,434 have also received the booster dose of vaccine/placebo. The aims and objectives, study design of the trial, present status as well as the socio-cultural aspect involved are highlighted in this paper.