Respiratory tract viral infections associated sepsis in patients with underlying liver disease: Viral sepsis an entity to look forward!

BACKGROUND: Sepsis remains a global health burden associated with significant morbidity and mortality. Bacteria are known to be the predominant pathogens in sepsis; however, viral etiologies in sepsis are still under diagnosed. Respiratory viral pathogens have been previously linked to sepsis, but the knowledge of incidence, disease burden and mortality of viral-induced sepsis remains limited. This study aimed at understanding the role of respiratory viral infections in the causation of sepsis in liver disease patients. METHODS: In this retrospective study, the clinical records of liver disease patients with influenza-like illness, whose requests for respiratory viral testing were received from January 2019 to December 2022, were reviewed. Respiratory viruses were identified using FilmArray 2.0 respiratory panel (BioFire Diagnostics, Utah, USA). RESULTS: Of 1391 patients tested, a respiratory viral etiology was detected in 23%. The occurrence of sepsis was seen in 35%. Among these, isolated viral etiology with no other bacterial/fungal coinfection was found in 55% of patients. Rhinovirus/Enterovirus was found as the most common underlying viral etiology (23.4%). The sepsis prevalence was higher among patients with associated comorbidities (45%) and decompensated cirrhosis (84%). On multi-variable analysis, no factor was found independently associated with sepsis-related mortality. CONCLUSION: This study underlines the importance of isolated viral etiology in causation of sepsis among liver disease patients. Patients with comorbidities, older age and decompensated cirrhosis are at an increased risk of developing sepsis and are associated with poorer outcomes. Accurate and timely identification of the viral etiology in sepsis would prevent the misuse of antibiotics and improve overall patient care.

Eco-hydrological modeling of soil wetting pattern dimensions under drip irrigation systems.

Reliable information on the horizontal and vertical dimensions of the wetted soil beneath a point source is critical for designing accurate, cost-effective, and efficient surface and subsurface drip irrigation systems. Several factors, including soil properties, initial soil conditions, dripper flow rate, number of drippers, spacing between drippers, irrigation management, plant root characteristics, and evapotranspiration, influence the dimensions and shape of wetting patterns. The objective of this study was to briefly review previous studies, collect the analytical, numerical, and empirical models developed, and evaluate the effectiveness of the most common empirical method for predicting the dimensions of soil wetted around drippers using measured data from field surveys. With this review study, we aim to promote a better understanding of soil water dynamics under point-source drip irrigation systems, help improve soil water dynamics under point-source drip irrigation systems, and identify issues that should be better addressed in future modeling efforts. A drip irrigation system was configured with three different emitters with different capacities (2, 4, and 8 l h-1) in the point source to determine the soil wetting front under the point source. The five most selected empirical equations (Al-Ogaidi, Malek and Peters, Amin and Ekhmaj, Li and Schwartzman and Zur) were statistically analyzed to test the efficiency in sandy loam soil. According to the results of the field investigation, statistical comparisons of the empirical models with the field investigation data were performed using the mean absolute error (MAE), root mean square error (RMSE), Nash-Sutcliffe model efficiency (CE), and coefficients of determination (R2). The advanced simulation of the wetting front was used based on the best accuracy of the selected empirical model. In general, the Li model (MAE, RMSE, EF, and R2 were 0.698 cm, 0.894 cm, 0.970 cm2 cm-2, and 0.970, respectively, for the wetted soil width and 1.800 cm, 1.974 cm, 0.927 cm2 cm-2, and 0.986, for the vertical advance) proved to be the best after statistical analysis with field data.

Hepatopulmonary syndrome is associated with low sphingosine-1-phosphate levels and can be ameliorated by the functional agonist fingolimod.

BACKGROUND & AIMS: Hepatopulmonary syndrome (HPS) is characterised by a defect in arterial oxygenation induced by pulmonary vascular dilatation in patients with liver disease. Fingolimod, a sphingosine-1-phosphate (S1P) receptor modulator, suppresses vasodilation by reducing nitric oxide (NO) production. We investigated the role of S1P in patients with HPS and the role of fingolimod as a therapeutic option in an experimental model of HPS. METHODS: Patients with cirrhosis with HPS (n = 44) and without HPS (n = 89) and 25 healthy controls were studied. Plasma levels of S1P, NO, and markers of systemic inflammation were studied. In a murine model of common bile duct ligation (CBDL), variations in pulmonary vasculature, arterial oxygenation, liver fibrosis, and inflammation were estimated before and after administration of S1P and fingolimod. RESULTS: Log of plasma S1P levels was significantly lower in patients with HPS than in those without HPS (3.1 ± 1.4 vs. 4.6 ± 0.2; p <0.001) and more so in severe intrapulmonary shunting than in mild and moderate intrapulmonary shunting (p <0.001). Plasma tumour necrosis factor-α (76.5 [30.3-91.6] vs. 52.9 [25.2-82.8]; p = 0.02) and NO (152.9 ± 41.2 vs. 79.2 ± 29.2; p = 0.001) levels were higher in patients with HPS than in those without HPS. An increase in Th17 (p <0.001) and T regulatory cells (p <0.001) was observed; the latter inversely correlated with plasma S1P levels. In the CBDL HPS model, fingolimod restored pulmonary vascular injury by increasing the arterial blood gas exchange and reducing systemic and pulmonary inflammation, resulting in improved survival (p = 0.02). Compared with vehicle treatment, fingolimod reduced portal pressure (p <0.05) and hepatic fibrosis and improved hepatocyte proliferation. It also induced apoptotic death in hepatic stellate cells and reduced collagen formation. CONCLUSIONS: Plasma S1P levels are low in patients with HPS and even more so in severe cases. Fingolimod, by improving pulmonary vascular tone and oxygenation, improves survival in a murine CBDL HPS model. IMPACT AND IMPLICATIONS: A low level of plasma sphingosine-1-phosphate (S1P) is associated with severe pulmonary vascular shunting, and hence, it can serve as a marker of disease severity in patients with hepatopulmonary syndrome (HPS). Fingolimod, a functional agonist of S1P, reduces hepatic inflammation, improves vascular tone, and thus retards the progression of fibrosis in a preclinical animal model of HPS. Fingolimod is being proposed as a potential novel therapy for management of patients with HPS.

Prevalence and Predictors for Respiratory Viral Infections among Liver Disease Patients.

Aim and background: Respiratory viral infections (RVIs) cause significant hospitalizations every year. Also, RVIs caused by either influenza or noninfluenza group of viruses can have adverse outcomes, especially among immunosuppressed patients. Regular and timely supervision is needed for accurate etiological identification, to prevent inappropriate use of antibiotics in patients with nonbacterial etiology. This study aimed to identify the spectrum of RVIs and clinical characteristics among liver disease patients with influenza-like illness (ILI). Materials and methods: In this study, medical records of patients with ILI, whose requests for respiratory viral testing came from September 2016 to December 2022 were retrospectively reviewed. Respiratory viruses were identified using FilmArray 2.0 respiratory panel (BioFire Diagnostics, USA). Results: Of the 1,577 liver disease patients with ILI, the overall prevalence of RVI was 28% (n = 449). Infection by noninfluenza viruses (NIVs) was detected in 329 patients (73%), higher than those infected with influenza viruses. In multivariable logistic regression analysis, female gender [odds ratio (OR): 2.5, 95% confidence interval (CI): 1.5-4.2], infection with influenza B (OR: 3.3, 95% CI: 1.09-9.9) and decompensated cirrhosis (OR: 3.9, 95% CI: 1.7-8.5) were independent risk factors for mortality. Regarding seasonality, influenza peaked in monsoons and winters, whereas NIVs circulated throughout the year. Conclusion: Overall, this study adds new knowledge regarding the incidence of RVI and the distribution of respiratory viral etiologies among liver disease patients with ILI. The findings highlight that female gender, decompensated cirrhosis, and influenza B infection are independently associated with poor clinical outcomes. Early etiological identification of viral causes of ILI could aid in an enhanced understanding of the prevalence of ILI and the timely management of the patients. Clinical significance: Respiratory viral infections can cause severe illness in individuals with underlying liver disease. Accurate diagnosis and risk stratification is crucial in mitigating the adverse health effects. How to cite this article: Samal J, Prabhakar T, Prasad M, et al. Prevalence and Predictors for Respiratory Viral Infections among Liver Disease Patients. Euroasian J Hepato-Gastroenterol 2023;13(2):108-114.

Association of Obstructive Sleep Apnea with Nocturnal Hypoxemia in Metabolic-Associated Fatty Liver Disease Patients: A Cross-sectional Analysis of Record-based Data.

RATIONALE: Obstructive sleep apnea (OSA) is often seen among obese individuals and the obesity has a linear association with MAFLD. The contribution of chronic intermittent nocturnal hypoxia of OSA and association of MAFLD with OSA is an unmet need. The present study aimed to determine the etiology, impact and association of OSA severity and nocturnal hypoxemia among patients ofChronic liver disease (CLD). METHODS: In this study, analysis of the medical records and clinical details of the patients of CLD who had undergone polysomnography were analyzed after appropriate inclusion in study as per inclusion and exclusion criteria. After assessing the eligibility criteria, a total of 78 patients were included in the final analysis. Nocturnal hypoxemia was gauged from the baseline oxygen saturation record of study. Presence and severity of OSA were graded as per American Academy of Sleep Medicine (AASM) criteria. The primary objective of the study was to determine the association between OSA severity and nocturnal hypoxemia to the presence of Non-alcoholic Fatty Liver Disease (NAFLD). Secondary objectives were to assess the association of OSA severity and extent of nocturnal hypoxemia to the BMI and to determine the proportions of NAFLD subjects with OSA. RESULTS: A total of 78 patients were screened, of which only 11 (14.1%) were female. Out of these, 56 (71.8%) were classified to MAFLD group while 22 (28.2%) were to the non-MAFLD group. The patients in MAFLD group with mean age of 56.02 years were older as compared to non-MAFLD with mean age of 51.05 years but that was not statistically different. Patients were categorized into MAFLD (n = 56) and non-MAFLD, representing other etiologies of CLD (n = 22; ethanol, chronic Hepatitis B virus (HBV), chronic Hepatitis C virus (HCV), cryptogenic, Non-cirrhosis portal fibrosis (NCPF), Primary sclerosing cholangitis (PSC), Autoimmune hepatitis (AIH), sarcoidosis, Wilson's disease). The mean BMI was significantly higher in MAFLD in comparison to non-MAFLD (34.51 ± 8.79 vs. 25.47 ± 5.75; P = 0.000) and also the median AHI of MAFLD group was significantly higher than the non-MAFLD 4.95 {(1.85, 25.47) vs. 0.85 (0.30, 2.72) (P value < 0.000)} [Table 1]. Among the desaturation indices, the number of desaturations >3% {median of 122.50 (75.00, 241.25) vs. 63.00 (13.75, 158.00), P value 0.009} and average desaturation {mean of (5.04 ± 2.16) vs. (3.78 ± 1.226)%, P value 0.002} were significantly higher in MAFLD versus non-MAFLD group [Table 2]. The AHI and all desaturation parameters, although not statistically significant, were worst in Child B [Table 3]. CONCLUSION: MAFLD patients have higher prevalence and greater severity of OSA and worse nocturnal desaturation parameters as compared to non-MAFLD patients. OSA is independent of obesity among patients of CLD, but prevalent among NAFLD group. Further prospective studies are needed among MAFLD and OSA patients to elucidate the mechanism linking pathophysiology of OSA-MAFLD and guide therapy.

Elevated plasma ICAM1 levels predict 28-day mortality in cirrhotic patients with COVID-19 or bacterial sepsis.

BACKGROUND & AIMS: Endothelial injury and dysfunction play a detrimental role in the pathogenesis of infections. Endothelium-related molecules have been reported as potential diagnostic and/or prognostic biomarkers of infection. The prognostic value of these biomarkers in patients with cirrhosis and infections remains elusive. METHODS: In this study, we investigated the performance of key soluble endothelial injury biomarkers, including intercellular adhesion molecule 1 (ICAM1), von Willebrand factor (vWF), vascular endothelial growth factor receptor 1 (VEGFR1), and angiopoietin 1 and 2 (Ang1, 2) as mortality predictors in patients with cirrhosis and severe COVID-19 or bacterial sepsis. RESULTS: A total of 66 hospitalized patients (admitted to the COVID-19 ward or liver intensive care unit [ICU]) were included. Twenty-two patients had COVID-19 alone, while 20 patients had cirrhosis plus COVID-19. Twenty-four patients had cirrhosis plus bacterial sepsis. Among patients with cirrhosis, the most common aetiology of liver disease was alcohol. ICAM1 was increased (p = 0.003) while VEGFR1 (p <0.0001) and Ang1 (p <0.0001) were reduced in patients with COVID-19 and cirrhosis, compared to patients with COVID-19 alone. Endothelial biomarker levels did not differ significantly between patients with cirrhosis and severe COVID-19 or bacterial sepsis in the ICU. In these patients, ICAM1 levels significantly and independently predicted mortality (hazard ratio 3.24; 95% CI 1.19-8.86) along with model for end-stage liver disease (MELD) score, renal and coagulation failures. The AUC for ICAM1 was 0.74, MELD was 0.60 and combined ICAM1 and MELD was 0.70. ICAM1 also positively correlated with the composite organ failure scores recorded 3-5 days post ICU admission (CLIF-OF and SOFA) in this subgroup of patients. CONCLUSION: The study indicates that in patients with cirrhosis, elevated plasma ICAM1 serves as an independent predictor of severe COVID-19- or sepsis-associated 28-day mortality. LAY SUMMARY: Bacterial sepsis and COVID-19 lead to increased mortality in patients with cirrhosis. In this study, we demonstrate that high plasma levels of ICAM1, an endothelial injury biomarker, is one of the important factors predicting mortality in critically ill cirrhotic patients with severe COVID-19 or bacterial sepsis.

Impact of temperature and sunshine duration on daily new cases and death due to COVID-19.

CONTEXT: Control of COVID-19 has now become a critical issue for public health. Many ecological factors are proven to influence the transmission and survival of the virus. However, the association between different climatic factors and spread and mortality due to COVID-19 is unknown. AIM: To determine the association of different climatic factors with the spread and mortality due to COVID-19 during January 2020 to May 2020. METHODS AND MATERIAL: The climatic indicators included in the study were duration of sunshine, average minimum temperature, and average maximum temperature, with cumulative confirmed cases, deceased, and recovered cases. The data was performed for 138 different countries of the world, from January 2020 to May 2020. STATISTICAL ANALYSIS USED: Spearman's correlation analysis was used to assess the correlation between temperature and the spread and mortality of COVID-19 cases. Both univariate and multivariate analysis was performed for cumulative and month-wise analysis, using SPSS software. RESULTS: Average maximum temperature and sunshine duration were significantly associated with COVID-19 confirmed cases, deceased, and recovered. For every 1° increase in average temperature, the confirmed, deceased, and recovered cases decreased by 2047 (P = 0.03), 157 (P = 0.016), and 743 (P = 0.005) individuals. The association remained significant even after adjusting for environmental as well as non-environmental variables. Average sunshine duration was inversely correlated with an increase in daily new cases (r = - 2261) and deaths (r = - 0.2985). CONCLUSION: Higher average temperature and longer sunshine duration are strongly associated with COVID-19 cases and deaths in 138 countries.

COVID-19 in Liver Transplant Recipients - A Series with Successful Recovery.

The severe acute respiratory syndrome corona virus-2 (referred to as SARS CoV2) pandemic had a great impact on public life in general as well as on populations with pre-existing disease and co-morbidities. Liver transplant and immunosuppressant medication predisposes to more severe disease and is often associated with poor outcome. The clinical features, disease course, treatment and process of modulating the immunosuppression is challenging. Here, we describe the clinical presentation, treatment and outcomes in six liver transplant recipients. Out of those six patients, three had mild, one had moderate and one had severe COVID-19, and one was asymptomatic. The immunosuppression minimization or withdrawal was done based upon the clinical severity. Consideration of tocilizumab and/o convalescent plasma as well as antivirals i.e. remdesvir done in severe cases. The routine practice of prophylactic anticoagulation, consideration of repurposed drugs (i.e. teicoplanin and doxycycline), and watchful monitoring of asymptomatic recipients helped to achieve an uneventful recovery.

Polygenic study of endurance-associated genetic markers ACE I/D, ACTN3 Arg(R)577Ter(X), CKMM A/G NcoI and eNOS Glu(G)298Asp(T) in male Gorkha soldiers.

BACKGROUND: Gorkhas, a sub-mountainous population of the Himalayan region, are known for strength and bravery. In the present study when "Gorkha" is used without brackets, we are mentioning Gorkhas of Tibeto-Burman origin. Physical capability, strength and endurance are important components of fitness associated with genetic traits. The aim of this study was to examine the endurance potential of male Gorkha soldiers, based on endurance-related genetic markers ACE I/D, ACTN3 Arg (R)577Ter(X), CKMM A/G NcoI and eNOS Glu(G)298Asp(T). METHODS: Genotypic and allelic frequencies were determined in 374 male Gorkha soldiers (Tibeto-Burman). These frequencies were compared with frequencies obtained from Gorkha (Indo-Aryan), high-altitude natives (Tibeto-Burman) and Indian lowlanders (Indo-Aryan). "Total genotype score" (TGS) was calculated from accumulated combination of polymorphisms with maximum value "100" for theoretically "optimal" polygenic score. Probability of occurrence of "optimal" endurance profile was also determined. RESULTS: ACE II genotypic frequency was highest in Tamangs followed by Gurungs, Rais, Limbus and Magars. No statistical difference in genotypic and allelic frequency of ACTN3 Arg(R)577Ter(X) was noted within the groups. Rais showed the highest CKMM A allele frequency (0.908) compared to other Gorkha (Tibeto-Burman) groups. Limbus and Tamangs showed the highest eNOS G allele frequency (0.938 and 0.915, respectively) compared to that of other groups. Probability of male Gorkha soldiers possessing a theoretically optimal polygenic endurance profile for four candidate polymorphisms was ~3.35% (1 in 30). Four percent of the population of male Gorkha soldiers (15 in 374) exhibited an optimal TGS 100, and 16% exhibited TGS 87 for endurance compared to male Indian soldiers belonging to the lowland (Indo-Aryan) and Gorkha (Indo-Aryan) populations suggesting an overall more "favourable" polygenic profile in the male Gorkha soldier (Tibeto-Burman) population. CONCLUSIONS: This study presents evidence of higher frequency of endurance-associated genes in the Gorkhas implying thereby that such genetically endowed individuals from the population may be selected and trained for achieving excellence in endurance-related elite sports activities.

Polymorphism profiling of nine high altitude relevant candidate gene loci in acclimatized sojourners and adapted natives.

BACKGROUND: Sea level sojourners, on ascent to high altitude, undergo acclimatization through integrated physiological processes for defending the body against oxygen deprivation while the high altitude natives (resident population) are adapted to the prevailing hypobaric hypoxic condition through natural selection. Separating the acclimatization processes from adaptive changes and identifying genetic markers in lowlanders that may be beneficial for offsetting the high altitude hypoxic stress, although challenging, is worth investigating. We genotyped nine candidate gene polymorphisms, suggested to be relevant in high altitude environment, in sea level acclimatized sojourners and adapted natives for understanding differences/commonality between the acclimatized and the adapted cohorts at the genetic level. RESULTS: Statistically similar genotypic and allelic frequencies were observed between the sea level sojourners (acclimatized) and the high altitude natives (adapted) in six loci viz., EDN1 (endothelin 1) -3A/-4A VNTR, ADRB2 (beta-2 adrenergic receptor, surface) Arg16Gly (rs1042713:A > G), ADRB3 (beta-3 adrenergic receptor) Trp64Arg (rs4994:T > C), eNOS (nitric oxide synthase, endothelial) Glu298Asp (rs1799983:T > G), TH (tyrosine hydroxylase) Val81Met (rs6356:G > A) and VEGF (vascular endothelial growth factor) 963C > T (rs3025039:C > T) while SCNN1B (amiloride-sensitive sodium channel, subunit beta) Thr594Met (rs1799979:C > T) was monomorphic. Genotypic and allelic frequencies in EDN1 9465G > A (rs2071942:G > A) and ADRB2 Gln27Glu (rs1042714:G > C) were significantly different between the acclimatized sojourners and the high altitude natives with higher frequency of GG and GA genotypes of EDN1 rs2071942 and CC genotype of ADRB2 rs1042714 being observed in Ladakh natives. Mutated A allele (AA genotype) of rs2071942 and carriers of G allele (GG + GC genotypes) of rs1042714 were less favorable during acclimatization under recessive and dominant genetic models of inheritance respectively indicating thereby that GG genotype and G allele of EDN1 rs2071942 and CC genotype of ADRB2 rs1042714 conferred acclimatization benefit. CONCLUSION: Sea level acclimatized individuals shared similarity with the adapted natives in certain high altitude relevant genetically based trait variation suggesting advantageous consequence as well as commonality in gene regulatory pathways in which these gene products function both during process of acclimatization and adaptation in high altitude environment.

Positive Association of D Allele of ACE Gene With High Altitude Pulmonary Edema in Indian Population.

OBJECTIVE: High altitude pulmonary edema (HAPE) is a potentially fatal high altitude illness occurring as a result of hypobaric hypoxia with an unknown underlying genetic mechanism. Recent studies have shown a possible association between HAPE and polymorphisms in genes of the renin-angiotensin-aldosterone system (RAAS), which play a key role in sensitivity of an individual toward HAPE. METHODS: For the present investigation, study groups consisted of HAPE patients (HAPE) and acclimatized control subjects (rCON). Four single-nucleotide polymorphisms (SNPs) were genotyped using restriction fragment length polymorphism (RFLP) analysis in genes of the RAAS pathway, specifically, renin (REN) C(-4063)T (rs41317140) and RENi8-83 (rs2368564), angiotensin (AGT) M(235)T (rs699), and angiotensin-converting enzyme (ACE) insertion/deletion (I/D) (rs1799752). RESULTS: Only the I/D polymorphism of the ACE gene showed a significant difference between the HAPE and rCON groups. The frequency of the D allele was found to be significantly higher in the HAPE group. Arterial oxygen saturation levels were significantly lower in the HAPE group compared with the rCON group and also decreased in the I/D and D/D genotypes compared with the I/I genotype in these groups. The other polymorphisms occurring in the REN and AGT genes were not significantly different between the 2 groups. CONCLUSIONS: These findings demonstrate a possible association of the I/D polymorphism of the ACE gene with the development of HAPE, with D/D being the at-risk genotype.

Identification of risk factors for malaria control by focused interventions in Ranchi district, Jharkhand, India.

BACKGROUND & OBJECTIVES: Ranchi, the capital of Jharkhand state is endemic for malaria, particularly the Bundu Primary Health Centre (PHC) is the worst affected. Therefore, a study was initiated during 2009 using remote sensing (RS) and geographical information system (GIS) to identify risk factors responsible for high endemicity in this PHC. METHODS: Bundu and Angara in Ranchi district were identified as high and low malaria endemic PHCs based on epidemiological data of three years (2007-09). The habitation, streams, other water body, landform, PHC and village boundary thematic maps were prepared using IRS-P6/LISS III-IV imageries and macro level breeding sites were identified. Digital elevation model (DEM) of the PHCs was generated using Cartosat Stereo Pair images and from DEM, slope map was derived to calculate flat area. From slope, aspect map was derived to indicate direction of water flow. Length of perennial streams, area under rocky terrain and buffer zones of 250, 500 and 750 m were constructed around streams. High resolution remote sensing imageries were used to identify micro level breeding sites. Based on macro-micro breeding sites, six villages from each PHC were selected randomly having combination of different parameters representing all ecotypes. Entomological data were collected during 2010-11 in pre- and post-monsoon seasons following standard techniques and analyzed statistically. Differential analysis was attempted to comprehend socioeconomic and other determinants associated with malaria transmission. RESULTS: The study identified eight risk factors responsible for higher malaria endemicity in Bundu in comparison to Angara PHC based on ecological, entomological, socioeconomic and other local parameters. CONCLUSION: Focused interventions in integrated vector management (IVM) mode are required to be carried out in the district for better management and control of disease.

Impact of deforestation on known malaria vectors in Sonitpur district of Assam, India.

BACKGROUND & OBJECTIVES: An alarming rate of deforestation has been reported from Sonitpur district of Assam, India therefore, a study was initiated during 2009 using remote sensing (RS) to assess deforested areas in the district and to study the impact on malaria vectors in order to formulate appropriate control strategy. METHODS: RS imageries of 2000 and 2009 were used to assess deforested areas in the selected district. Entomological data were collected in four surveys during 2009-2011. The data were analyzed statistically using test of single proportions (χ 2 ) and pair-wise comparison. Vector incrimination was done using enzyme-linked immunosorbent assay (ELISA) and entomological inoculation rate (EIR) was calculated to estimate transmission intensity. RESULTS: The deforested areas were identified in north-western parts of Sonitpur district falling in Dhekiajuli Primary Health Centre (PHC). The forest cover of the PHC decreased >50% during 2000-2009. Five species of anopheline vectors were collected. Anopheles minimus sensu lato (s.l.) was collected least abundantly while An. culicifacies s.l. prevailed most abundantly and significant difference was observed between proportions of the collected vector species. Pair-wise comparison between An. culicifacies s.l. and An. minimus s.l. was also found statistically significant indicating that An. culicifacies s.l. is establishing its population in deforested areas. An. culicifacies s.l. was found ELISA positive and EIR was measured as 4.8 during transmission season. CONCLUSION: An. culicifacies s.l. replaced An. minimus s.l., the vector of malaria in northeast India and was found ELISA positive, therefore could have possible role in malaria transmission in the deforested areas of the district.

Deletion mutant comprising 198 residues of BoNT/A toxin receptor binding domain retained gt1b binding property but failed to induce protective antibody response in a mouse model.

The most effective protection against toxin is inducing protective immune response through vaccination that will produce neutralizing antibodies. Antibodies will bind to and clear toxin from the circulation before it can enter nerve cells and block neurotransmission and can also be used for development of detection system. In the present study we constructed a deletion mutant of the binding domain (1098-1296) to produce smallest toxin fragment as vaccine candidate against BoNT/A. The BoNT/A-HCC protein was highly expressed in Escherichia coli SG13009 and found to form inclusion bodies. The purified inclusion bodies were solubilized in 6 M guanidine-HCl containing 10 mM ß-mercaptoethanol and 20 mM imidazole and the rBoNT/A-HCC was purified and refolded in a single step on Ni2+ affinity column. The purified protein was ∼98 % pure as assessed by SDS-polyacrylamide gel with the yield of 8 mg/L and showed binding to polysialoganglioside (GT1b). The rBoNT/A-HCC at dose of 40 µg/mouse generated high IgG antibody titre with predominance of IgG1 subtype, but failed to protect animals against BoNT/A challenge. Antibody titre in serum was determined by enzyme linked immunosorbent assay and specific binding to rBoNT/A-HCC was demonstrated by surface plasmon resonance (SPR), with a dissociation constant of 0.8 pM.

Expression, purification and development of neutralizing antibodies from synthetic BoNT/B LC and its application in detection of botulinum toxin serotype B.

Botulism is a neuroparalytic disease caused by Clostridium botulinum, which produces seven (A-G) neurotoxins (BoNTs). The mouse bioassay is the gold standard for the detection of botulinum neurotoxins, however it requires at least 3-4 days for completion. Most of the studies were carried out in botulinum toxin A and less on type B. Attempts have been made to develop an ELISA based detection system, which is potentially an easier and more rapid method of botulinum neurotoxin detection. In the present study, the synthetic BoNT/B LC gene was constructed using PCR overlapping primers, cloned in a pET28a+ vector and expressed in E. coli BL21DE3. The maximum yield of recombinant proteins was optimized after 16 hrs of post induction at 21°C and purified the recombinant protein in soluble form. Antibodies were raised in Mice and Rabbit. The IgG antibody titer in the case of Mice was 1: 1,024,000 and Rabbit was 1: 512,000 with alum as adjuvant via intramascular route. The biological activity of the recombinant protein was confirmed by in-vitro studies using PC12 cells by the synaptobrevin cleavage, the rBoNT/B LC protein showed the maximum blockage of acetylcholine release at a concentration of 150nM rBoNT/B LC in comparison to the control cells. When the cells were incubated with rBoNT/B LC neutralized by the antisera raised against it, the acetylcholine release was equivalent to the control. IgG specific to rBoNT/B LC was purified from raised antibodies. The results showed that the developed antibody against rBoNT/B LC protein were able to detect botulinum toxin type B approximately up to 1 ng/ml. These developed high titer antibodies may prove useful for the detection of botulinum neurotoxins in food and clinical samples.

Comparative genomic analysis of a neurotoxigenic Clostridium species using partial genome sequence: Phylogenetic analysis of a few conserved proteins involved in cellular processes and metabolism.

Clostridial organisms produce neurotoxins, which are generally regarded as the most potent toxic substances of biological origin and potential biological warfare agents. Clostridium tetani produces tetanus neurotoxin and is responsible for the fatal tetanus disease. In spite of the extensive immunization regimen, the disease is an important cause of death especially among neonates. Strains of C. tetani have not been genetically characterized except the complete genome sequencing of strain E88. The present study reports the genetic makeup and phylogenetic affiliations of an environmental strain of this bacterium with respect to C. tetani E88 and other clostridia. A shot gun library was constructed from the genomic DNA of C. tetani drde, isolated from decaying fish sample. Unique clones were sequenced and sequences compared with its closest relative C. tetani E88. A total of 275 clones were obtained and 32,457 bases of non-redundant sequence were generated. A total of 150 base changes were observed over the entire length of sequence obtained, including, additions, deletions and base substitutions. Of the total 120 ORFs detected, 48 exhibited closest similarity to E88 proteins of which three are hypothetical proteins. Eight of the ORFs exhibited similarity with hypothetical proteins from other organisms and 10 aligned with other proteins from unrelated organisms. There is an overall conservation of protein sequences among the two strains of C. tetani and. Selected ORFs involved in cellular processes and metabolism were subjected to phylogenetic analysis.

Immobilization of anaerobic bacteria on rubberized-coir for psychrophilic digestion of night soil.

Low-ambient temperatures, <30 degrees C, are known to cause drastic reduction in the efficiency of anaerobic biodigesters due to low-growth rate of the constituent bacterial consortium. Immobilization of anaerobic bacteria has been attempted in the biodigester operating at 10 degrees C. Various matrices were screened and evaluated for the immobilization of bacteria in digesters. Anaerobic digestion of night soil was carried out with hydraulic retention time in the range of 9-18 days. Among the tested matrices, rubberized-coir was found to be the most useful at 10 degrees C with optimum hydraulic retention time of 15 days. Optimum amount of coir was found as 25 g/L of the working volume of biodigesters. Immobilization of bacteria on the coir was observed by scanning electron microscopy and fluorescent microscopy. The study indicates that rubberized-coir can be utilized to increase biodegradation of night soil at higher organic loading. Another advantage of using this matrix is that it is renewable and easily available in comparison to other synthetic polymeric matrices.