Search results for: endemic%20species

Authors: Monika Soni, Shovonlal Bhowmick, Chandra Bhattacharya, Jitendra Sharma, Prafulla Dutta, Jagadish Mahanta

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Aedes aegypti and Aedes albopictus are considered as two major vectors to transmit dengue virus. In North-east India, two states viz. Assam and Arunachal Pradesh are known to be high endemic zone for dengue and Chikungunya viral infection. The taxonomical classification of medically important vectors are important for mapping of actual evolutionary trends and epidemiological studies. However, misidentification of mosquito species in field-collected mosquito specimens could have a negative impact which may affect vector-borne disease control policy. DNA barcoding is a prominent method to record available species, differentiate from new addition and change of population structure. In this study, a combined approach of a morphological and molecular technique of DNA barcoding was adopted to explore sequence variation in mitochondrial cytochrome c oxidase subunit I (COI) gene within dengue vectors. The study has revealed the map distribution of the dengue vector from two states i.e. Assam and Arunachal Pradesh, India. Approximate five hundred mosquito specimens were collected from different parts of two states, and their morphological features were compared with the taxonomic keys. The analysis of detailed taxonomic study revealed identification of two species Aedes aegypti and Aedes albopictus. The species aegypti comprised of 66.6% of the specimen and represented as dominant dengue vector species. The sequences obtained through standard DNA barcoding protocol were compared with public databases, viz. GenBank and BOLD. The sequences of all Aedes albopictus have shown 100% similarity whereas sequence of Aedes aegypti has shown 99.77 - 100% similarity of COI gene with that of different geographically located same species based on BOLD database search. From dengue prevalent different geographical regions fifty-nine sequences were retrieved from NCBI and BOLD databases of the same and related taxa to determine the evolutionary distance model based on the phylogenetic analysis. Neighbor-Joining (NJ) and Maximum Likelihood (ML) phylogenetic tree was constructed in MEGA6.06 software with 1000 bootstrap replicates using Kimura-2-Parameter model. Data were analyzed for sequence divergence and found that intraspecific divergence ranged from 0.0 to 2.0% and interspecific divergence ranged from 11.0 to 12.0%. The transitional and transversional substitutions were tested individually. The sequences were deposited in NCBI: GenBank database. This observation claimed the first DNA barcoding analysis of Aedes mosquitoes from North-eastern states in India and also confirmed the range expansion of two important mosquito species. Overall, this study insight into the molecular ecology of the dengue vectors from North-eastern India which will enhance the understanding to improve the existing entomological surveillance and vector incrimination program.

Keywords: COI, dengue vectors, DNA barcoding, molecular identification, North-east India, phylogenetics

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Authors: Yichao Liu, Peter Fransson, Julian Heidecke, Jonas Wallin, Joacim Rockloev

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Dengue, a mosquito-borne viral disease, poses a significant public health challenge in endemic tropical or subtropical countries, including Sri Lanka. To reveal insights into the complexity of the dynamics of this disease and study the drivers, a comprehensive model capable of both robust forecasting and insightful inference of drivers while capturing the co-circulating of several virus strains is essential. However, existing studies mostly focus on only one aspect at a time and do not integrate and carry insights across the siloed approach. While mechanistic models are developed to capture immunity dynamics, they are often oversimplified and lack integration of all the diverse drivers of disease transmission. On the other hand, purely data-driven methods lack constraints imposed by immuno-epidemiological processes, making them prone to overfitting and inference bias. This research presents a hybrid model that combines machine learning techniques with mechanistic modelling to overcome the limitations of existing approaches. Leveraging eight years of newly reported dengue case data, along with socioeconomic factors, such as human mobility, weekly climate data from 2011 to 2018, genetic data detecting the introduction and presence of new strains, and estimates of seropositivity for different districts in Sri Lanka, we derive a data-driven vector (SEI) to human (SEIR) model across 16 regions in Sri Lanka at the weekly time scale. By conducting ablation studies, the lag effects allowing delays up to 12 weeks of time-varying climate factors were determined. The model demonstrates superior predictive performance over a pure machine learning approach when considering lead times of 5 and 10 weeks on data withheld from model fitting. It further reveals several interesting interpretable findings of drivers while adjusting for the dynamics and influences of immunity and introduction of a new strain. The study uncovers strong influences of socioeconomic variables: population density, mobility, household income and rural vs. urban population. The study reveals substantial sensitivity to the diurnal temperature range and precipitation, while mean temperature and humidity appear less important in the study location. Additionally, the model indicated sensitivity to vegetation index, both max and average. Predictions on testing data reveal high model accuracy. Overall, this study advances the knowledge of dengue transmission in Sri Lanka and demonstrates the importance of incorporating hybrid modelling techniques to use biologically informed model structures with flexible data-driven estimates of model parameters. The findings show the potential to both inference of drivers in situations of complex disease dynamics and robust forecasting models.

Keywords: compartmental model, climate, dengue, machine learning, social-economic

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Authors: Etibar Guliyev

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The breakdown of the Soviet Union in 1991 has led to a considerable rise in the religious self-consciousness of Muslim population of the Central Asia. Additionally, huge amount of money spent by various states further facilitated the spread of religious ideas. According to some sources, Saudi Arabia spent 87 billion dollars to propagate Wahhabism abroad during two decades, whereas the Communist Party of the Soviet Union spent just over 7 billion dollars to spread its ideology worldwide between 1921 and 1991. As the result, today once a remote area from international politics has turned into third major source of recruitment of fighters for global terrorist organizations. In order to illustrate to scope of the involvement of the Central Asian residents in international terrorist networks it is enough to mention the name of Colonel Gulmorod Khalimov, the former head of the Tajik special police forces who served as ISIS war minister between 2016 and 2017. The importance of the topic stems from the fact that the above-mentioned republics with a territory of 4 million square km and the population of around 80 million people borders Russia, Iran Afghanistan and China. Moreover, the fact that political and military activities motivated with religious feelings in those countries have implications not only for domestic but also for regional and global political relations and all of them has root in politics of religions adds value to the research. This research aims to provide an in-depth analyses of the marked features of the state policies to regulate religious activities and approach this question both from individual, domestic, regional and global levels of analyses. The research will enable us to better understand what implications have the state of religious freedom in post-Soviet Muslim republics for international relations and the rise of global jihadi terrorism. The paper tries to find a linkage between the mentioned terror attacks and underground rise of religious extremism in Central Asia. This research is based on multiple research methods, mainly on qualitative one. The process tracing method is also employed to review religious policies implemented from 1918-1991 and after the collapse of the Soviet Union in a chronological way. In terms of the quantitative method, it chiefly will be used in a bid to process various statistics disseminated in academic and official sources. The research mostly explored constructivist, securitization and social movement theories. Findings of the research suggests that the endemic problems peculiar to authoritarian regimes of Central Asia such as crackdown on the expression of religious believe and any kind of opposition, economic decline, instrumental use of religion and corruption and tribalism further accelerated the recruitment problem. Paper also concludes that the Central Asian states in some cases misused counter-terrorism campaign as a pretext to further restrict freedom of faith in their respective countries.

Keywords: identity, political Islam, religious extremism, security, terrorism

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Authors: Arsyam Mawardi, Sony Suhandono, Azzania Fibriani, Fifi Fitriyah Masduki

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Malaria is an infectious disease caused by Plasmodium sp. This disease has a high prevalence in Indonesia, especially in Jayapura. The vaccine that is currently being developed has not been effective in overcoming malaria. This is due to the high polymorphism in the Plasmodium genome especially in areas that encode Plasmodium surface proteins. Merozoite Surface Protein 1 (MSP1) Plasmodium falciparum is a surface protein that plays a role in the invasion process in human erythrocytes through the interaction of Glycophorin A protein receptors and sialic acid in erythrocytes with Reticulocyte Binding Proteins (RBP) and Duffy Adhesion Protein (DAP) ligands in merozoites. MSP1 can be targeted to be a specific antigen and predicted epitope area which will be used for the development of diagnostic and malaria vaccine therapy. MSP1 consists of 17 blocks, each block is dimorphic, and has been marked as the K1 and MAD20 alleles. Exceptions only in block 2, because it has 3 alleles, among others K1, MAD20 and RO33. These polymorphisms cause allelic variations and implicate the severity of patients infected P. falciparum. In addition, polymorphism of MSP1 in Jayapura isolates has not been reported so it is interesting to be further identified and projected as a specific antigen. Therefore, in this study, we analyzed the allele polymorphism as well as detected the MSP1 epitope antigen candidate on block 2 P. falciparum. Clinical samples of selected malaria patients followed the consecutive sampling method, examining malaria parasites with blood preparations on glass objects observed through a microscope. Plasmodium DNA was isolated from the blood of malarial positive patients. The block 2 MSP1 gene was amplified using PCR method and cloned using the pGEM-T easy vector then transformed to TOP'10 E.coli. Positive colonies selection was performed with blue-white screening. The existence of target DNA was confirmed by PCR colonies and DNA sequencing methods. Furthermore, DNA sequence analysis was done through alignment and formation of a phylogenetic tree using MEGA 6 software and insilico analysis using IEDB software to predict epitope candidate for P. falciparum. A total of 15 patient samples have been isolated from Plasmodium DNA. PCR amplification results show the target gene size about ± 1049 bp. The results of MSP1 nucleotide alignment analysis reveal that block 2 MSP1 genes derived from the sample of malarial patients were distributed in four different allele family groups, K1 (7), MAD20 (1), RO33 (0) and MSP1_Jayapura (10) alleles. The most commonly appears of the detected allele is MSP1_Jayapura single allele. There was no significant association between sex variables, age, the density of parasitemia and alel variation (Mann Whitney, U > 0.05), while symptomatic signs have a significant difference as a trigger of detectable allele variation (U < 0.05). In this research, insilico study shows that there is a new epitope antigen candidate from the MSP1_Jayapura allele and it is predicted to be recognized by B cells with 17 amino acid lengths in the amino acid sequence 187 to 203.

Keywords: epitope candidate, insilico analysis, MSP1 P. falciparum, polymorphism

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Authors: Al. Tsertsvadze. L. Ebralidze, I. Matchutadze. D. Berashvili, A. Bakuridze

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Peatlands are landscape elements, they are formed over a very long period by physical, chemical, biologic, and geologic processes. In the moderate zone of Caucasus, the Kolkheti lowlands are distinguished by the diversity of relictual plants, a high degree of endemism, orographic, climate, landscape, and other characteristics of high levels of biodiversity. The unique properties of the Kolkheti region lead to the formation of special, so-called, endemic peat peloids. The composition and properties of peloids strongly depend on peat-forming plants. Peat is considered a unique complex of raw materials, which can be used in different fields of the industry: agriculture, metallurgy, energy, biotechnology, chemical industry, health care. They are formed in permanent wetland areas. As a result of decay, higher plants remain in the anaerobic area, with the participation of microorganisms. Peat mass absorbs soil and groundwater. Peloids are predominantly rich with humic substances, which are characterized by high biological activity. Humic acids stimulate enzymatic activity, regenerative processes, and have anti-inflammatory activity. Objects of the research were Kolkheti peat peloids (Ispani, Anaklia, Churia, Chirukhi, Peranga) possessing different formation phases. Due to specific physical and chemical properties of research objects, the aim of the research was to develop analytical methods in order to study the chemical composition of the objects. The research was held using modern instrumental methods of analysis: Ultraviolet-visible spectroscopy and Infrared spectroscopy, Scanning Electron Microscopy, Centrifuge, dry oven, Ultraturax, pH meter, fluorescence spectrometer, Gas chromatography-mass spectrometry (GC-MS/MS), Gas chromatography. Based on the research ration between organic and inorganic substances, the spectrum of micro and macro elements, also the content of minerals was determined. The content of organic nitrogen was determined using the Kjeldahl method. The total composition of amino acids was studied by a spectrophotometric method using standard solutions of glutamic and aspartic acids. Fatty acid was determined using GC (Gas chromatography). Based on the obtained results, we can conclude that the method is valid to identify fatty acids in the research objects. The content of organic substances in the research objects was held using GC-MS. Using modern instrumental methods of analysis, the chemical composition of research objects was studied. Each research object is predominantly reached with a broad spectrum of organic (fatty acids, amino acids, carbocyclic and heterocyclic compounds, organic acids and their esters, steroids) and inorganic (micro and macro elements, minerals) substances. Modified methods used in the presented research may be utilized for the evaluation of cosmetological balneological and pharmaceutical means prepared on the base of Kolkheti's Sphagnum Peat Peloids.

Keywords: modern analytical methods, natural resources, peat, chemistry

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Authors: Olga Smilevska Spasova, Katerina Boshkovska, Gorica Popova, Mirjana Popovska

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Introduction: Pneumonia is an infection of the pulmonary parenchyma. HPIV 3 is one of four viruses that is a member of the Paramyxoviridae family designated types 1-4 that have a nonsegmented, single-stranded RNA genome with a lipid-containing envelope. They are spread from the respiratory tract by aerosolized secretions or by direct contact with secretions. Type 3 is endemic and can cause serious illness in immunocompromised patients. Illness caused by parainfluenza occurs shortly after inoculation with the virus. The level of immunoglobulin A antibody in serum is the best predictor of susceptibility to infection. Streptococcus pneumonia or pneumococcus is a Gram-positive, spherical bacteria, usually found in pairs and it is a member of the genus Streptococcus. Streptococcus pneumonia resides asymptomatically in healthy carriers typically colonizing the respiratory tract, sinuses, and nasal cavity. In individuals with weaker immune systems like young infants, pneumococcal bacterium is the most common cause of community-acquired pneumonia in the world. Case Report: The aim is to present a case of lower respiratory tract infection in an infant caused by parainfluenza virus 3, S. pneumonia and undifferentiated gram-negative bacteria that was successfully treated. The infant is with a history of recurrent episodes of wheezing in the past 3mounts.Infant of 10months presents 2weeks before admittance with high fever, runny nose, and cough. The primary pediatrician prescribed oral cefpodoxime for 10days and inhaled salbutamol. Two days before admittance in hospital the infant with high fever, cough, and difficulty breathing. At admittance, infant is pale, anxious with rapid respirations, cough, wheezing and tachycardia. On auscultation: vesicular breathing sounds with high pitched wheezing and on the right coarse crackles. Investigations: Blood analysis: RBC: 4, 7 x1012L, WBC: 8,3x109L: Neut: 42.73% Lym: 41.57%, Hgb: 9.38 g/dl MCV: 62.7fl, MCH: 20.0pg MCHC: 31.8 g/dl RDW: 18.7% Plt-307.9 x109LCRP: 2,5mg/l, serum iron-7.92umol/l, O2sat-97% on blood gas analysis, puls-125/min.X-ray of chest with hyperinflationand right pericardial consolidation. Microbiological analysis of sputum sample is positive for undifferentiated gram-negative bacteria (colonizer)–resistant to cefotaxime, ampicillin, cefoxitin, sulfamet.+trimetoprim and sensitive to amikacin, gentamicin, and ciprofloxacin. Molecular multiplex RT-PCR for 19 viruses and multiplex PCR for 7 bacteria test for respiratory pathogens positive for Parainfluenza virus 3(Ct=22.73), Streptococcus pneumonia (Ct=26.75).IED: IgG-9.31g/l, IgA-0.351g/l, IgM-0.86g/l. Therapy: Treatment was started with inhaled salbutamol, intravenous antibiotic cefotaxime as well as systemic corticosteroids. On day 7 because of slow clinical resolution of chest auscultation findings and an etiologic clue with a positive sputum sample for resistant undifferentiated gram negative bacteria, a second intravenous antibiotic was administered amikacin. The infant is discharged on day 14 with resolution of clinical findings. Conclusion: Mixed co-infections with respiratory viruses and bacteria in immunocompromised infants are likely to lead to a more severe form of community acquired pneumonia that will need hospitalization.

Keywords: HPIV- 3, infant, pneumonia, S. pneumonia, x-ray chest

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Authors: Raham El-Kahlout, Hadi Yassin, Asmaa Athani, Marwan Abou Madi, Gheyath Nasrallah

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Background: Since its first isolation in September 2012, Middle East respiratory syndrome coronavirus (MERS-CoV) has diffused across 27 countries infecting more than two thousand individuals with a high case fatality rate. MERS-CoV–specific antibodies are widely found in Dromedary camel along with viral shedding of similar viruses detected in human at same region, suggesting that MERS epidemiology may be central role by camel. Interestingly, MERS-CoV has also been also reported to be asymptomatic or to cause influenza-like mild illnesses. Therefore, in a country like Qatar (bordered Saudi Arabia), where camels are widely spread, serological surveys are important to explore the role of camels in MERS-CoV transmission. However, widespread strategic serological surveillances of MERS-CoV among populations, particularly in endemic country, are infrequent. In the absence of clear epidemiological view, cross-sectional MERS antibody surveillances in human populations are of global concern. Method: We performed a comparative serological screening of 4719 healthy blood donors, 135 baseline case contacts (high risk individual), and four MERS confirmed patients (by PCR) for the presence of anti-MERS IgG. Initially, samples were screened using Euroimmune anti- MERS-CoV IgG ELISA kit, the only commercial kit available in the market and recommended by the CDC as a screening kit. To confirm ELISA test results, farther serological testing was performed for all borderline and positive samples using two assays; the anti MERS-CoV IgG and IgM Euroimmune indirect immunofluorescent test (IIFT) and pseudoviral particle neutralizing assay (PPNA). Additionally, to test cross reactivity of anti-MERS-CoV antibody with other family members of coronavirus, borderline and positive samples were tested for the presence of the of IgG antibody of the following viruses; SARS, HCoV-229E, HKU1 using the Euroimmune IIFT for SARS and HCoV-229E and ELISA for HKU1. Results: In all of 4858 screened 15 samples [10 donors (0.21%, 10/4719), 1 case contact (0.77 %, 1/130), 3 patients (75%, 3/4)] anti-MERS IgG reactive/borderline samples were seen in ELISA. However, only 7 (0.14%) of them gave positive with in IIFT and only 3 (0.06%) was confirmed by the specific anti-MERS PPNA. One of the interesting findings was, a donor, who was selected in the control group as a negative anti-MERS IgG ELISA, yield reactive for anti-MERS IgM IIFT and was confirmed with the PPNA. Further, our preliminary results showed that there was a strong cross reactivity between anti- MERS-COV IgG with both HCoV-229E or anti-HKU1 IgG, yet, no cross reactivity of SARS were found. Conclusions: Our findings suggest that MERS-CoV is not heavily circulated among the population of Qatar and this is also indicated by low number of confirmed cases (only 18) since 2012. Additionally, the presence of antibody of other pathogenic human coronavirus may cause false positive results of both ELISA and IIFT, which stress the need for more evaluation studies for the available serological assays. Conclusion: this study provides an insight about the epidemiological view for MERS-CoV in Qatar population. It also provides a performance evaluation for the available serologic tests for MERS-CoV in a view of serologic status to other human coronaviruses.

Keywords: seroprevalence, MERS-CoV, healthy individuals, Qatar

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Authors: Eunice Oluwakemi Chukwuma-Nwuba

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Nigeria is a developing country with an increasing rate of graduate unemployment. This has triggered successive government administrations to promote the variety of programmes to address the situation. However, none of these efforts yielded the desired outcome. Accordingly, in 2006 the government included entrepreneurship module in the curriculum of universities as a compulsory general programme for all undergraduate courses. This is in the hope that the programme will help to promote entrepreneurial mind-set and new venture creation among graduates and as a result reduce the rate of graduate unemployment. The study explores the effectiveness of entrepreneurship education in promoting entrepreneurship. This study is significant in view of the endemic graduate unemployment in Nigeria and the social consequences such as youth restiveness and militancy. It is guided by the theory of planned behaviour. It employed the two-stage structural equation modelling (AMOS) to model entrepreneurial intentions as a function of innovative teaching methods, traditional teaching methods and culture Personal attitude and subjective norm are proposed to mediate the relationships between the exogenous and the endogenous variables. The first stage was tested using multi-group confirmatory factor analysis (MGCFA) framework to confirm that the two groups assign the same meaning to the scale items and to obtain goodness-of-fit indices. The multi-group confirmatory factor analysis included the tests of configural, metric and scalar invariance. With the attainment of full configural invariance and partial metric and scalar invariance, the second stage – the structural model was applied hypothesising that, the entrepreneurial intentions of graduates (respondents who have participated in the compulsory entrepreneurship programme) will be higher than those of undergraduates (respondents who are yet to participate in the programme). The study uses the quasi-experimental design. The samples comprised 409 graduates (experimental group) and 402 undergraduates (control group) from six federal universities in Nigeria. Our findings suggest that personal attitude is positively related with entrepreneurial intentions, largely confirming prior literature. However, unlike previous studies, our results indicate that subjective norm has significant direct and indirect impact on entrepreneurial intentions indicating that reference people of the participants have important roles to play in their decision to be entrepreneurial. Furthermore, unlike the assertions in prior studies, the result suggests that traditional teaching methods have indirect effect on entrepreneurial intentions supporting that since personal characteristics can change in an educational situation, an education purposively directed at entrepreneurship might achieve similar results if not better. This study has implication for practice and theory. The research extends to the theoretical understanding of the formation of entrepreneurial intentions and explains the role of the reference others in relation to how graduates perceive entrepreneurship. Further, the study adds to the body of knowledge on entrepreneurship education in Nigeria universities and provides a developing country perspective. It proposes further research in the exploration of entrepreneurship education and entrepreneurial intentions of graduates from across the country’s universities as necessary and imperative.

Keywords: entrepreneurship education, entrepreneurial intention, structural equation modeling, theory of planned behaviour

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Authors: Igor N. Kosenko

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The present contribution is an analysis of more than 300 specimens of Upper Jurassic to Lower Cretaceous oysters collected by V.A. Zakharov during the 1960s and currently stored in the Trofimuk Institute of Geology and Geophysics SB RAS (Novosibirsk, Russia). They were sampled in the northwestern bounder of Western Siberia (Yatriya, Maurynia, Tol’ya and Lopsiya rivers) and the north of Eastern Siberia (Boyarka, Bolshaya Romanikha and Dyabaka-Tari rivers). During the last five years, they were examined with taxonomical and palaeoecological purposes. Based on carbonate material of oyster’s shells were performed isotopic analyses and associated palaeotemperatures. Taxonomical study consists on classical morphofunctional and biometrical analyses. It is completed by another large amount of Cretaceous oysters from Crimea as well as modern Pacific oyster - Crassostrea gigas. Those were studied to understand the range of modification variability between different species. Oysters previously identified as Liostrea are attributed now to four genera: Praeexogyra and Helvetostrea (Flemingostreidae), Pernostrea (Gryphaeidae) and one new genus (Gryphaeidae), including one species “Liostrea” roemeri (Quenstedt). This last is characterized by peculiar ethology, being attached to floating ammonites and morphology, outlined by a beak-shaped umbo on the right (!) valve. Endemic Siberian species from the Pernostrea genus have been included into the subgenus Boreiodeltoideum subgen. nov. Pernostrea and Deltoideum genera have been included into the tribe Pernostreini n. trib. from the Gryphaeinae subfamily. Model of phylogenetic relationships between species of this tribe has been proposed. Siberian oyster complexes were compared with complexes from Western Europe, Poland and East European Platform. In western Boreal and Subboreal Realm (England, northern France and Poland) two stages of oyster’s development were recognized: Jurassic-type and Cretaceous-type. In Siberia, Jurassic and Lower Cretaceous oysters formed a unique complex. It may be due to the isolation of the Siberian Basin toward the West during the Early Cretaceous. Seven oyster’s shells of Pernostrea (Pernostrea) uralensis (Zakharov) from the Jurassic/Cretaceous Boundary Interval (Upper Volgian – Lower Ryazanian) of Maurynia river were used to perform δ13C and δ18O isotopic analyses. The preservation of the carbonate material was controlled by: cathodoluminescence analyses; content of Fe, Mn, Sr; absence of correlation between δ13C and δ18O and content of Fe and Mn. The obtained δ13C and δ18O data were compared with isotopic data based on belemnites from the same stratigraphical interval of the same section and were used to trace palaeotemperatures. A general trend towards negative δ18O values is recorded in the Maurynia section, from the lower part of the Upper Volgian to the middle part of the Ryazanian Chetaites sibiricus ammonite zone. This trend was previously recorded in the Nordvik section. The higher palaeotemperatures (2°C in average) determined from oyster’s shells indicate that belemnites likely migrated laterally and lived part of their lives in cooler waters. This work financially supported by the Russian Foundation for Basic Researches (grant no. 16-35-00003).

Keywords: isotopes, oysters, Siberia, taxonomy

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Authors: Hosein I. Hosein, Ragab Azzam, Ahmed M. S. Menshawy, Sherin Rouby, Khaled Hendy, Ayman Mahrous, Hany Hussien

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Brucellosis is a highly contagious bacterial zoonotic disease of a worldwide spread and has different names; Infectious or enzootic abortion and Bang's disease in animals; and Mediterranean or Malta fever, Undulant Fever and Rock fever in humans. It is caused by the different species of genus Brucella which is a Gram-negative, aerobic, non-spore forming, facultative intracellular bacterium. Brucella affects a wide range of mammals including bovines, small ruminants, pigs, equines, rodents, marine mammals as well as human resulting in serious economic losses in animal populations. In human, Brucella causes a severe illness representing a great public health problem. The disease was reported in Egypt for the first time in 1939; since then the disease remained endemic at high levels among cattle, buffalo, sheep and goat and is still representing a public health hazard. The annual economic losses due to brucellosis were estimated to be about 60 million Egyptian pounds yearly, but actual estimates are still missing despite almost 30 years of implementation of the Egyptian control programme. Despite being the gold standard, bacterial isolation has been reported to show poor sensitivity for samples with low-level of Brucella and is impractical for regular screening of large populations. Thus, serological tests still remain the corner stone for routine diagnosis of brucellosis, especially in developing countries. In the present study, a total of 1533 cows (256 from Beni-Suef Governorate, 445 from Al-Fayoum Governorate and 832 from Damietta Governorate), were employed for estimation of relative sensitivity, relative specificity, positive predictive value and negative predictive value of buffered acidified plate antigen test (BPAT), rose bengal test (RBT) and complement fixation test (CFT). The overall seroprevalence of brucellosis revealed (19.63%). Relative sensitivity, relative specificity, positive predictive value and negative predictive value of BPAT,RBT and CFT were estimated as, (96.27 %, 96.76 %, 87.65 % and 99.10 %), (93.42 %, 96.27 %, 90.16 % and 98.35%) and (89.30 %, 98.60 %, 94.35 %and 97.24 %) respectively. BPAT showed the highest sensitivity among the three employed serological tests. RBT was less specific than BPAT. CFT showed the least sensitivity 89.30 % among the three employed serological tests but showed the highest specificity. Different tissues specimens of 22 seropositive cows (spleen, retropharyngeal udder, and supra-mammary lymph nodes) were subjected for bacteriological studies for isolation and identification of Brucella organisms. Brucella melitensis biovar 3 could be recovered from 12 (54.55%) cows. Bacteriological examinations failed to classify 10 cases (45.45%) and were culture negative. Bruce-ladder PCR was carried out for molecular identification of the 12 Brucella isolates at the species level. Three fragments of 587 bp, 1071 bp and 1682 bp sizes were amplified indicating Brucella melitensis. The results indicated the importance of using several procedures to overcome the problem of escaping of some infected animals from diagnosis.Bruce-ladder PCR is an important tool for diagnosis and epidemiologic studies, providing relevant information for identification of Brucella spp.

Keywords: brucellosis, relative sensitivity, relative specificity, Bruce-ladder, Egypt

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Authors: Asim Abbasi, Muhammad Sufyan, Iqra, Hafiza Javaria Ashraf

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The share of vector-borne diseases (VBDs) in the global burden of infectious diseases is almost 17%. The advent of new drugs and latest research in medical science helped mankind to compete with these lethal diseases but still diseases transmitted by different mosquito species, including filariasis, malaria, viral encephalitis and dengue are serious threats for people living in disease endemic areas. Injudicious and repeated use of pesticides posed selection pressure on mosquitoes leading to development of resistance. Hence biological control agents are under serious consideration of scientific community to be used in vector control programmes. Fish have a history of predating immature stages of different aquatic insects including mosquitoes. The noteworthy examples in Africa and Asia includes, Aphanius discolour and a fish in the Panchax group. Moreover, common mosquito fish, Gambusia affinis predates mostly on temporary water mosquitoes like anopheline as compared to permanent water breeders like culicines. Mosquitoes belonging to genus Toxorhynchites have a worldwide distribution and are mostly associated with the predation of other mosquito larvae habituating with them in natural and artificial water containers. These species are harmless to humans as their adults do not suck human blood but feeds on floral nectar. However, their activity is mostly temperature dependent as Toxorhynchites brevipalpis consume 359 Aedes aegypti larvae at 30-32 ºC in contrast to 154 larvae at 20-26 ºC. Although many bacterial species were isolated from mosquito cadavers but those belonging to genus Bacillus are found highly pathogenic against them. The successful species of this genus include Bacillus thuringiensis and Bacillus sphaericus. The prime targets of B. thuringiensis are mostly the immatures of genus Aedes, Culex, Anopheles and Psorophora while B. sphaericus is specifically toxic against species of Culex, Psorophora and Culiseta. The entomopathogenic nematodes belonging to family, mermithidae are also pathogenic to different mosquito species. Eighty different species of mosquitoes including Anopheles, Aedes and Culex proved to be highly vulnerable to the attack of two mermithid species, Romanomermis culicivorax and R. iyengari. Cytoplasmic polyhedrosis virus was the first described pathogenic virus, isolated from the cadavers of mosquito specie, Culex tarsalis. Other viruses which are pathogenic to culicine includes, iridoviruses, cytopolyhedrosis viruses, entomopoxviruses and parvoviruses. Protozoa species belonging to division microsporidia are the common pathogenic protozoans in mosquito populations which kill their host by the chronic effects of parasitism. Moreover, due to their wide prevalence in anopheline mosquitoes and transversal and horizontal transmission from infected to healthy host, microsporidia of the genera Nosema and Amblyospora have received much attention in various mosquito control programmes. Fungal based mycopesticides are used in biological control of insect pests with 47 species reported virulent against different stages of mosquitoes. These include both aquatic fungi i.e. species of Coelomomyces, Lagenidium giganteum and Culicinomyces clavosporus, and the terrestrial fungi Metarhizium anisopliae and Beauveria bassiana. Hence, it was concluded that the integrated use of all these biological control agents can be a healthy contribution in mosquito control programmes and become a dire need of the time to avoid repeated use of pesticides.

Keywords: entomopathogenic nematodes, protozoa, Toxorhynchites, vector-borne

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Authors: Viviana Manzulli, Luigina Serrecchia, Adelia Donatiello, Valeria Rondinone, Sabine Zange, Alina Tscherne, Antonio Parisi, Antonio Fasanella

Abstract:

Anthrax is a zoonotic disease that affects a wide range of animal species (primarily ruminant herbivores), and can be transmitted to humans through consumption or handling of contaminated animal products. The etiological agent B.anthracis is able to survive in unfavorable environmental conditions by forming endospore which remain viable in the soil for many decades. Furthermore, B.anthracis is considered as one of the most feared agents to be potentially misused as a biological weapon and the importance of the disease and its treatment in humans has been underscored before the bioterrorism events in the United States in 2001. Due to the often fatal outcome of human cases, antimicrobial susceptibility testing plays especially in the management of anthrax infections an important role. In Italy, animal anthrax is endemic (predominantly found in the southern regions and on islands) and is characterized by sporadic outbreaks occurring mainly during summer. Between 2012 and 2017 single human cases of cutaneous anthrax occurred. In this study, 90 diverse strains of B.anthracis, isolated in Italy from 2001 to 2017, were screened to their susceptibility to sixteen clinically relevant antimicrobial agents by using the broth microdilution method. B.anthracis strains selected for this study belong to the strain collection stored at the Anthrax Reference Institute of Italy located inside the Istituto Zooprofilattico Sperimentale of Puglia and Basilicata. The strains were isolated at different time points and places from various matrices (human, animal and environmental). All strains are a representative of over fifty distinct MLVA 31 genotypes. The following antibiotics were used for testing: gentamicin, ceftriaxone, streptomycin, penicillin G, clindamycin, chloramphenicol, vancomycin, linezolid, cefotaxime, tetracycline, erythromycin, rifampin, amoxicillin, ciprofloxacin, doxycycline and trimethoprim. A standard concentration of each antibiotic was prepared in a specific diluent, which were then twofold serial diluted. Therefore, each wells contained: bacterial suspension of 1–5x104 CFU/mL in Mueller-Hinton Broth (MHB), the antibiotic to be tested at known concentration and resazurin, an indicator of cell growth. After incubation overnight at 37°C, the wells were screened for color changes caused by the resazurin: a change from purple to pink/colorless indicated cell growth. The lowest concentration of antibiotic that prevented growth represented the minimal inhibitory concentration (MIC). This study suggests that B.anthracis remains susceptible in vitro to many antibiotics, in addition to doxycycline (MICs ≤ 0,03 µg/ml), ciprofloxacin (MICs ≤ 0,03 µg/ml) and penicillin G (MICs ≤ 0,06 µg/ml), recommend by CDC for the treatment of human cases and for prophylactic use after exposure to the spores. In fact, the good activity of gentamicin (MICs ≤ 0,25 µg/ml), streptomycin (MICs ≤ 1 µg/ml), clindamycin (MICs ≤ 0,125 µg/ml), chloramphenicol(MICs ≤ 4 µg/ml), vancomycin (MICs ≤ 2 µg/ml), linezolid (MICs ≤ 2 µg/ml), tetracycline (MICs ≤ 0,125 µg/ml), erythromycin (MICs ≤ 0,25 µg/ml), rifampin (MICs ≤ 0,25 µg/ml), amoxicillin (MICs ≤ 0,06 µg/ml), towards all tested B.anthracis strains demonstrates an appropriate alternative choice for prophylaxis and/or treatment. All tested B.anthracis strains showed intermediate susceptibility to the cephalosporins (MICs ≥ 16 µg/ml) and resistance to trimethoprim (MICs ≥ 128 µg/ml).

Keywords: Bacillus anthracis, antibiotic susceptibility, treatment, minimum inhibitory concentration

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Authors: Ipsita Pujari, Abitha Thomas, Vidhu S. Babu, K. Satyamoorthy

Abstract:

Orchids are esteemed as celebrities in cut flower industry globally, due to their long-lasting fragrance and freshness. Apart from splendor, the unique metabolites endowed with pharmaceutical potency have made them one of the most hunted in plant kingdom. This had led to their trafficking, resulting in habitat loss, subsequently making them occupiers of IUCN red list as RET species. Many of the orchids especially wild varieties still remain undiscovered. In view to protect and conserve the wild germplasm, researchers have been inventing novel micropropagation protocols; thereby conserving Orchids. India is overflowing with exclusive wild cultivars of Orchids, whose pharmaceutical properties remain untapped and are not marketed owing to relatively small flowers. However, their germplasm is quite pertinent to be preserved for making unusual hybrids. Dendrobium genus is the second largest among Orchids exists in India and has highest demand attributable to enduring cut flowers and significant therapeutic uses in traditional medicinal system. Though the genus is quite endemic in Western Ghat regions of the country, many species are still anonymous with their unknown curative properties. A standard breeding cycle in Orchids usually takes five to seven years (Dendrobium hybrids taking a long juvenile phase of two to five years reaching maturity and flowering stage) and this extensive life cycle has always hindered the development of Dendrobium breeding. Dendrobium is reported with essential therapeutic plant bio-chemicals and ‘Moscatilin’ is one, found exclusive to this famous Dendrobium genus. Moscatilin is reported to have anti-mutagenic and anti-cancer properties, whose positive action has very recently been demonstrated against a range of cancers. Our preliminary study here established a simple and economic small-scale propagation protocol of Dendrobium ovatum describing in vitro production of Moscatilin. Subsequently for enhancing the content of Moscatilin, an efficient experimental related to the organization of transgenic (hairy) D. ovatum root cultures through infection of Agrobacterium rhizogenes 2364 strain on MS basal medium is being reported in the present study. Hairy roots generated on almost half of the explants used (spherules, in vitro plantlets and calli) maintained through suspension cultures, after 8 weeks of co-cultivation with Agrobacterium rhizogenes. GFP assay performed with isolated hairy roots has confirmed the integrative transformation which was further positively confirmed by PCR using rolB gene specific primers. Reverse phase-high performance liquid chromatography and mass spectrometry techniques were used for quantification and accurate identification of Moscatilin respectively from transgenic systems. A noticeable ~3 fold increase in contents were observed in transformed D. ovatum root cultures as compared to the simple in vitro culture, callus culture and callus regeneration plantlets. Role of elicitors e.g., Methyl jasmonate, Salicylic acid, Yeast extract and Chitosan were tested for elevating the Moscatilin content to obtain a comprehensive optimized protocol facilitating the in vitro production of valuable Moscatilin with larger yield. This study would provide evidence towards the in vitro assembly of Moscatilin within a short time-period through not a so-expensive technology for the first time. It also serves as an appropriate basis for bioreactor scale-up resulting in commercial bioproduction of Moscatilin.

Keywords: bioproduction, Dendrobium ovatum, hairy root culture, moscatilin

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Authors: F. Varga, Z. Liber, J. Jakše, A. Turudić, Z. Šatović, I. Radosavljević, N. Jeran, M. Grdiša

Abstract:

Dalmatian pyrethrum (Tanacetum cinerariifolium (Trevir.) Sch. Bip.; Asteraceae), a source of the commercially dominant plant insecticide pyrethrin, is a species endemic to the eastern Adriatic. Genetic diversity of T. cinerariifolium was previously studied using amplified fragment length polymorphism (AFLP) markers. However, microsatellite markers (single sequence repeats - SSRs) are more informative because they are codominant, highly polymorphic, locus-specific, and more reproducible, and thus are most often used to assess the genetic diversity of plant species. Dalmatian pyrethrum is an outcrossing diploid (2n = 18) whose large genome size and high repeatability have prevented the success of the traditional approach to SSR markers development. The advent of next-generation sequencing combined with the specifically developed method recently enabled the development of, to the author's best knowledge, the first set of SSRs for genomic characterization of Dalmatian pyrethrum, which is essential from the perspective of plant genetic resources conservation. To evaluate the effectiveness of the developed SSR markers in genetic differentiation of Dalmatian pyrethrum populations, a preliminary genetic diversity study was conducted on 30 individuals from three geographically distinct natural populations in Croatia (northern Adriatic island of Mali Lošinj, southern Adriatic island of Čiovo, and Mount Biokovo) based on 12 SSR loci. Analysis of molecular variance (AMOVA) by randomization test with 10,000 permutations was performed in Arlequin 3.5. The average number of alleles per locus, observed and expected heterozygosity, probability of deviations from Hardy-Weinberg equilibrium, and inbreeding coefficient was calculated using GENEPOP 4.4. Genetic distance based on the proportion of common alleles (DPSA) was calculated using MICROSAT. Cluster analysis using the neighbor-joining method with 1,000 bootstraps was performed with PHYLIP to generate a dendrogram. The results of the AMOVA analysis showed that the total SSR diversity was 23% within and 77% between the three populations. A slight deviation from Hardy-Weinberg equilibrium was observed in the Mali Lošinj population. Allele richness ranged from 2.92 to 3.92, with the highest number of private alleles observed in the Mali Lošinj population (17). The average observed DPSA between 30 individuals was 0.557. The highest DPSA (0.875) was observed between several pairs of Dalmatian pyrethrum individuals from the Mali Lošinj and Mt. Biokovo populations, and the lowest between two individuals from the Čiovo population. Neighbor-joining trees, based on DPSA, grouped individuals into clusters according to their population affiliation. The separation of Mt. Biokovo clade was supported (bootstrap value 58%), which is consistent with the previous study on AFLP markers, where isolated populations from Mt. Biokovo differed from the rest of the populations. The developed SSR markers are an effective tool for assessing the genetic diversity and structure of natural Dalmatian pyrethrum populations. These preliminary results are encouraging for a future comprehensive study with a larger sample size across the species' range. Combined with the biochemical data, these highly informative markers could help identify potential genotypes of interest for future development of breeding lines and cultivars that are both resistant to environmental stress and high in pyrethrins. Acknowledgment: This work has been supported by the Croatian Science Foundation under the project ‘Genetic background of Dalmatian pyrethrum (Tanacetum cinerariifolium /Trevir./ Sch. Bip.) insecticidal potential’- (PyrDiv) (IP-06-2016-9034) and by project KK.01.1.1.01.0005, Biodiversity and Molecular Plant Breeding, at the Centre of Excellence for Biodiversity and Molecular Plant Breeding (CoE CroP-BioDiv), Zagreb, Croatia.

Keywords: Asteraceae, genetic diversity, genomic SSRs, NGS, pyrethrum, Tanacetum cinerariifolium

Procedia PDF Downloads 87