Isolation and characterization of different forms of Bordetella pertussis adenylate cyclase

This thesis describes the different forms of AC produced by B. pertussis, B. parapertussis, B. bronchiseptica and those expressed in E. coli. The work was done with both crude and purified preparations of AC and also with whole cells of B. pertussis. Immunoblot analysis with various antisera, including anti-AC monoclonal, monospecific, and polyclonal antibodies, revealed the production of two different forms of AC with different electrophoretic mobilities i.e., 210 and 200 kDa as the major cell-associated forms in B. pertussis and B. bronchiseptica. In 3 B. parapertussis and E. coli, the predominant form was 210 kDa. These forms may represent the toxic and non-toxic forms observed by Rogel et al. (1989). Production of the holotoxic AC was not observed in culture supernates under the growth conditions used. Only low MW forms of 45-50 kDa were observed and these forms also appeared to be produced by proteolysis of AC during storage and other experimental manipulations. The breakdown was partially prevented by using a protease inhibitor. The enzymic activity of AC was found to be unstable in purified and crude forms of AC in the absence of urea. Thus, urea seemed to be beneficial in protecting loss of activity of AC during storage. A noteworthy observation of the present study was that the different Bordetellae, including B. pertussis Tn5-insertion mutants deficient in individual virulence factors, and a recombinant B. pertussis strain, produced different amounts of AC enzymic and toxic activities under the same set of condition which indicate that the production varied from strain to strain . The highest yields of enzymically-active and toxic AC were obtained from recombinant strain B. pertussis BP348 (pRMBl) and from B. pertussis Tn5-insertion mutant BP357 (deficient in a functional PT gene). On the other hand, B. pertussis Tohama and Tn5 mutant BP353 showed the lowest AC enzymic and toxic activities of the B. pertussis strains examined. B. bronchiseptica produced an intermediate level of AC enzymic and toxic activities compared to those of the B. pertussis strains. Enzymically and biologically-active AC was obtained from the E. coli constructs. However, the toxic activity was approximately half than that of native AC. E. coli strains harbouring the cloned B. pertussis cyaA, B, D genes together with C genes either from P. vulgaris, E. coli, OTB. pertussis on separate plasmids, showed production of CyaA (AC), as assessed by immunoblotting and the Salomon assay,and its activation by C genes, as assessed by inhibition of chemiluminescence. Various attempts were made to develop more convenient, economical, and sensitive methods for detection of AC and determination of its enzymic and toxic activities. An alternative method for detection of AC by calmodulin-affinity probing of a Western blot was found to be reproducible and suitable for use when specific anti-AC antibodies were not available. This is the first report of such a use for calmodulin. A microplate assay was developed for detection of AC enzymic activity and this proved economical, reproducible, and convenient for preliminary screening of AC-producing strains and as a semi-quantitative assay. The method has some advantages over conventional assays, such as the Salomon assay and the Binding protein assay, in that it does not require radioisotope and it is rapid. Further development of this assay as a diagnostic test for pertussis may be possible. A comparative study was made of different assays for determination of toxic effects of AC on a variety of target cells. These included : haemolysis of sheep and rabbit RBCs, CHO cell elongation, arborization of BHK cells, inhibition of the nitroblue tetrazolium reduction capacity of rabbit neutrophils, and inhibition of the chemiluminescence response of neutrophils to various stimuli. The study indicated that CL inhibition by AC toxin was a sensitive assay and could detect ACT in crude preparations at the level of 1mug protein /10

COMPARATIVE PRODUCTIVE PERFORMANCE AND EGG CHARACTERISTICS OF PULLETS AND SPENT LAYERS

Production performance and egg quality characteristics of pullets and spent layers were compared in this study. Forty birds, each from the flocks of pullets (24 weeks old) and spent layers (76 weeks old) were selected as experimental birds. The birds from each age group were divided into five replicates, each comprising of eight birds. All the experimental birds were fed a commercial layer ration @ 110g/bird/day for 12 weeks. The data on egg production, feed consumption, egg weight and egg quality characteristics viz. shell thickness, shell weight, breaking strength, albumen diameter, albumen weight and yolk weight were recorded. The data thus collected were utilized for calculation of FCR, Haugh unit and yolk index values. The results revealed that pullets produced more eggs and utilized their feed more efficiently than spent layers. However, egg weight in spent layers was higher than in their counterparts. Pullets also produced eggs with thicker shell and higher Haugh unit values when compared to the spent layers. Feed consumption and yolk index values remained unaffected due to the age. Pullets also had better egg quality characteristics than those of spent layers

The Effect of Ursolic Acid and Hyperinsulinemia on the Liver and Kidney Function of Alloxan-Induced Diabetic Male Rabbits

The study aimed to examine the effect of extracted and standard ursolic acid in addition to hyperinsulinemia compared to vitamin B complex on the liver and kidney function of alloxan-induced diabetic male rabbits. The liver and kidney functions were determined and analyzed by one–way of a NOVA at variance software at a P-Value of 0.05. The result showed a significant decrease in aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and uric acid values in the groups treated with standard and extraction ursolic acid(UA) compared to the diabetic group. The rabbit group that was treated with vitamin B complex recorded a significant decrease in the three liver enzymes, uric acid, and creatinine values than the diabetic group. In addition, the rabbit group that was treated with hyperinsulinemia recorded a significant increase in aspartate aminotransferase (AST) and creatinine values than the diabetic rabbit group. While the histological results in the groups treated with standard and extraction ursolic acid had a clear ameliorative effect in liver and kidney function tests better than hyperinsulinemia, also vitamin B complex caused some good effects

Characterization of oligomers from methylglyoxal under dark conditions : a pathway to produce secondary organic aerosol through cloud processing during nighttime

Aqueous-phase oligomer formation from methylglyoxal, a major atmospheric photooxidation product, has been investigated in a simulated cloud matrix under dark conditions. The aim of this study was to explore an additional pathway producing secondary organic aerosol (SOA) through cloud processes without participation of photochemistry during nighttime. Indeed, atmospheric models still underestimate SOA formation, as field measurements have revealed more SOA than predicted. Soluble oligomers (n = 1-8) formed in the course of acid-catalyzed aldol condensation and acid-catalyzed hydration followed by acetal formation have been detected and characterized by positive and negative ion electrospray ionization mass spectrometry. Aldol condensation proved to be a favorable mechanism under simulated cloud conditions, while hydration/acetal formation was found to strongly depend on the pH of the system and only occurred at a pH < 3.5. No evidence was found for formation of organosulfates. The aldol oligomer series starts with a beta-hydroxy ketone via aldol condensation, where oligomers are formed by multiple additions of C3H4O2 units (72 Da) to the parent beta-hydroxy ketone. Ion trap mass spectrometry experiments were performed to structurally characterize the major oligomer species. A mechanistic pathway for the growth of oligomers under cloud conditions and in the absence of UV-light and OH radicals, which could substantially enhance in-cloud SOA yields, is proposed here for the first time

Terpenylic acid and related compounds: precursors for dimers in secondary organic aerosol from the ozonolysis of α- and β-pinene

In the present study, we have characterized the structure of a higher-molecular weight (MW) 358 α- and β-pinene dimeric secondary organic aerosol (SOA) product that received ample attention in previous molecular characterization studies and has been elusive. Based on mass spectrometric evidence for deprotonated molecules formed by electrospray ionization in the negative ion mode and chemical considerations, it is suggested that diaterpenylic acid is a key monomeric intermediate for dimers of the ester type. It is proposed that cis-pinic acid is esterified with the hydroxyl-containing diaterpenylic acid, which can be explained through acid-catalyzed hydrolysis of the recently elucidated lactone-containing terpenylic acid and/or diaterpenylic acid acetate, both first-generation oxidation products. To a minor extent, higher-MW 358 and 344 diester products are formed containing other terpenoic acids as monomeric units, i.e., diaterpenylic acid instead of cis-pinic acid, and diaterebic acid instead of diaterpenylic acid. It is shown that the MW 358 diester and related MW 344 compounds, which can be regarded as processed SOA products, also occur in ambient fine (PM2.5) rural aerosol collected at night during the warm period of the 2006 summer field campaign conducted at K-puszta, Hungary, a rural site with coniferous vegetation. This indicates that, under ambient conditions, the higher-MW diesters are formed in the particle phase over a longer time-scale than that required for gas-to-particle partitioning of their monomeric precursors in laboratory α-/β-pinene ozonolysis experiments

Estimation of particulate matter and gaseous concentrations using low-cost sensors from broiler houses

Particulate and gaseous emissions from intensive poultry facilities are major public and environmental health concern. The present study was aimed at exploratively monitoring particulate matter (PM) and gaseous concentrations in controlled-environment facilities using low-cost sensors in Lahore, Pakistan. The indoors and outdoors of 18 broiler houses, grouped into three categories based on the age of birds: group I (1–20 days), group II (21–30 days) and group III (31–40 days), were examined. Low-cost sensors Dylos 1700 and Aeroqual 500 series with different gas sensor heads were used to monitor PM and different gases such as nitrogen dioxide (NO2), hydrogen sulphide (H2S), carbon dioxide (CO2) and methane (CH4), respectively. Overall, the mean PM and gaseous concentrations increased with the age and activity of birds as compared with the non-activity time of birds. Statistically significant differences were observed in all measured parameters among the groups. The negative correlation between indoor and outdoor environments for PM and gas concentrations at some broiler houses demonstrates the contribution of additional sources to emissions in outdoor environments. The findings contribute to our knowledge of temporal characteristics of particulate and gaseous concentrations from poultry facilities particularly in Pakistan and generally to the capability of using low-cost sensors to evaluate emissions from such facilities

Assessment of respiratory problems in workers associated with intensive poultry facilities in Pakistan

Background The poultry industry in Pakistan has flourished since the 1960s; however, there are scarce data regarding the impact of occupational exposure on the pulmonary health of farm workers in terms of years working in the industry. The objective of the present study was to assess the effect of poultry environment on the health of occupationally exposed poultry farmers in countries of warm climatic regions, such as Pakistan. This study will also show the effect of exposure to poultry facilities on the health of poultry farmers in the context of low-income countries with a relatively inadequate occupational exposure risk management. Materials and methods The lung function capacity of 79 poultry workers was measured using a spirometer. Along with spirometry, a structured questionnaire was also administrated to obtain information about age, height, weight, smokers/nonsmokers, years of working experience, and pulmonary health of farm workers. The workers who were directly involved in the care and handling of birds in these intensive facilities were considered and divided into four groups based on their years of working experience: Group I (3-10 months), Group II (1-5 years), Group III (6-10 years), and Group IV (more than 11 years). The forced vital capacity (FVC), forced expiratory volume in one second (FEV1) and the FEV1/FVC ratio were considered to identify lung function abnormalities. Statistical analysis was carried out using independent sample t test, Chi-square test, Pearson's correlation, and linear regression. Results Based on the performed spirometry, 68 (86 %) of workers were found normal and healthy, whereas 11 (14 %) had a mild obstruction. Of the 11 workers with mild obstruction, the highest number with respect to the total was in Group IV (more than 11 years of working experience) followed by Group III and Group II. Most of the workers were found healthy, which seems to be because of the healthy survivor effect. For the independent sample t test, a significant difference was noticed between healthy and nonhealthy farmers, whereas Chi-square test showed a significant association with height, drugs, and working experience. Linear regression that was stratified by respiratory symptoms showed for workers with symptoms, regression models for all spirometric parameters (FVC, FEV1, and FEV1/FVC) have better predictive power or R square value than those of workers without symptoms. Conclusion These findings suggest that lung function capacity was directly related to years of working experience. With increasing number of working years, symptoms of various respiratory problems enhanced in the poultry workers. It should be noted that most of the poultry workers were healthy and young, the rationale being that there is a high turnover rate in this profession. The mobility in this job and our finding of 86% of the healthy workers in the present study also proposed healthy worker survivor effec

Xylem anatomical responses of Larix Gmelinii and Pinus Sylvestris influenced by the climate of Daxing'an mountains in Northeastern China

Wood anatomy and plant hydraulics play a significant role in understanding species-specific responses and their ability to manage rapid environmental changes. This study used the dendro-anatomical approach to assess the anatomical characteristics and their relation to local climate variability in the boreal coniferous tree species Larix gmelinii (Dahurian larch) and Pinus sylvestris var. mongolica (Scots pine) at an altitude range of 660-m to 842- m. We measured the xylem anatomical traits (lumen area (LA), cell wall thickness (CWt), cell counts per ring (CN), ring width (RW), and cell sizes in rings) of both species at four different sites Mangui (MG), Wuerqihan (WEQH), Moredagha (MEDG) and Alihe (ALH) and investigated their relationship with temperature and precipitation of those sites along a latitude gradient. Results showed that all chronologies have strong summer temperature correlations. LA extremes were mostly associated with climatic variation than CWt and RWt. MEDG site species showed an inverse correlation in different growing seasons. The correlation coefficient with temperature indicated significant variations in the May-September months at MG, WEQH, and ALH sites. These results suggest that climatic seasonality changes in the selected sites positively affect hydraulic efficiency (increase in the diameter of the earlywood cells) and the width of the latewood produced in P. sylvestris. In contrast, L. gmelinii showed the opposite response to warm temperatures. It is concluded that xylem anatomical responses of L. gmelinii and P. sylvestris showed varied responses to different climatic factors at different sites. These differences between the two species responses to climate are due to the change of site condition on a large spatial and temporal scale

Depression, sleeping pattern, and suicidal ideation among medical students in Bangladesh: a cross-sectional pilot study

© 2020, Springer-Verlag GmbH Germany, part of Springer Nature. Background: Depression is a major morbidity and the most common mental disorder among the medical students in medical schools globally. Undergraduate students suffer stress more due to their academic curriculum than the students of other faculties. In low-resource settings like Bangladesh, there is a dearth in research on the mental health of undergraduate medical students. This pilot study was conducted to add to the existing limited evidence by reporting the prevalence of depression and describing sleeping pattern and suicidal tendencies among medical students. Relevantly, we have investigated the overall mental health status among the medical students in Bangladesh. Methods: This cross-sectional study was conducted in two medical colleges of Dhaka between July 2013 and December 2013, among 221 Bangladeshi medical students from first to fifth year. By the convenience sampling technique, data were collected by a pretested, structured, self-administered questionnaire and analysis was done by SPSS version 18.0. Depression was assessed by the validated Patient Health Questionnaire-9 (PHQ-9) tool among the respondents. Goldberg’s General Health Questionnaire (GHQ-28) was used for assessing overall mental health status. Results: Depression was found in 38.9% of participants, with 3.6%, 14.5%, and 20.8% being severe, moderate, and mild depression, respectively. 17.6% of medical students had suicidal tendency or attempted suicide at least once after attending medical school. The sleeping hours were inadequate and altered after starting this stressful academic course. 33.5% of medical students had poor mental health status. There was a statistically significant association between poor mental health status in the age group less than 22 years old and initial academic study year (1st to 3rd of MBBS). Conclusion: The findings are suggestive of a higher prevalence of depression among early-year medical students and marginal predominance in males. Suicidal tendency is also higher. This calls for further investigation with situation analysis, qualitative explorations, and surveys to explore the burden of such disorders in Bangladesh

Papaya Maturity Classifications using Deep Convolutional Neural Networks

Papaya is a tropical fruit with a green cover, yellow pulp, and a taste between mango and cantaloupe, having commercial importance because of its high nutritive and medicinal value. The process of sorting papaya fruit based on maturely is one of the processes that greatly determine the mature of papaya fruit that will be sold to consumers. The manual grading of papaya fruit based on human visual perception is time-consuming and destructive. The objective of this paper is to the status classification of papaya fruits if it's mature or partially matured or unmatured. A deep learning technique that was extensively applied to image recognition was used. The trained model achieved an accuracy of 100% on a held-out test set, demonstrating the feasibility of this approach. Classification model of VGG16 achieved a 100% accuracy and 112 seconds of training time