Characterization of plaque variants and the involvement of Quasi-Species in a population of EV-A71

Viral plaque morphologies in human cell lines are markers for growth capability and they have been used to assess the viral fitness and selection of attenuated mutants for live-attenuated vaccine development. In this study, we investigate whether the naturally occurring plaque size variation reflects the virulence of the variants of EV-A71. Variants of two different plaque sizes (big and small) from EV-A71 sub-genotype B4 strain 41 were characterized. The plaque variants displayed different in vitro growth kinetics compared to the parental wild type. The plaque variants showed specific mutations being present in each variant strain. The big plaque variants showed four mutations I97L, N104S, S246P and N282D in the VP1 while the small plaque variants showed I97T, N237T and T292A in the VP1. No other mutations were detected in the whole genome of the two variants. The variants showed stable homogenous small plaques and big plaques, respectively, when re-infected in rhabdomyosarcoma (RD) and Vero cells. The parental strain showed faster growth kinetics and had higher viral RNA copy number than both the big and small plaque variants. Homology modelling shows that both plaque variants have differences in the structure of the VP1 protein due to the presence of unique spontaneous mutations found in each plaque variant This study suggests that the EV-A71 sub-genotype B4 strain 41 has at least two variants with different plaque morphologies. These differences were likely due to the presence of spontaneous mutations that are unique to each of the plaque variants. The ability to maintain the respective plaque morphology upon passaging indicates the presence of quasi-species in the parental population

Microbial biodegradation of paraffin wax in Malaysian crude oil mediated by degradative enzymes

The deposition of paraffin wax in crude oil is a problem faced by the oil and gas industry during extraction, transportation, and refining of crude oil. Most of the commercialized chemical additives to prevent wax are expensive and toxic. As an environmentally friendly alternative, this study aims to find a novel thermophilic bacterial strain capable of degrading paraffin wax in crude oil to control wax deposition. To achieve this, the biodegradation of crude oil paraffin wax by 11 bacteria isolated from seawater and oil-contaminated soil samples was investigated at 70°C. The bacteria were identified as Geobacillus kaustophilus N3A7, NFA23, DFY1, Geobacillus jurassicus MK7, Geobacillus thermocatenulatus T7, Parageobacillus caldoxylosilyticus DFY3 and AZ72, Anoxybacillus geothermalis D9, Geobacillus stearothermophilus SA36, AD11, and AD24. The GCMS analysis showed that strains N3A7, MK7, DFY1, AD11, and AD24 achieved more than 70% biodegradation efficiency of crude oil in a short period (3 days). Notably, most of the strains could completely degrade C37–C40 and increase the ratio of C14–C18, especially during the initial 2 days incubation. In addition, the degradation of crude oil also resulted in changes in the pH of the medium. The degradation of crude oil is associated with the production of degradative enzymes such as alkane monooxygenase, alcohol dehydrogenase, lipase, and esterase. Among the 11 strains, the highest activities of alkane monooxygenase were recorded in strain AD24. A comparatively higher overall alcohol dehydrogenase, lipase, and esterase activities were observed in strains N3A7, MK7, DFY1, AD11, and AD24. Thus, there is a potential to use these strains in oil reservoirs, crude oil processing, and recovery to control wax deposition. Their ability to withstand high temperature and produce degradative enzymes for long-chain hydrocarbon degradation led to an increase in the short-chain hydrocarbon ratio, and subsequently, improving the quality of the oi

Feasibility study on ornamental fish production in Tabriz great park

Study was conducted during 2007 to 2010 for feasibility study and planning of ornamental fish bisines farms in Tabriz great park . All aspects of study support of 16 production stors in the plane. Each store has 450 squer neter and can support more than 168000 fry and mature fish for sailing . some extended supporting systems were suggested

Newly isolated alkane hydroxylase and lipase producing Geobacillus and Anoxybacillus species involved in crude oil degradation

Isolation and studies of novel, crude oil biodegrading thermophilic strains may provide a wider knowledge in understanding their role in petroleum degradation. In this study, the screening of ten new thermophilic strains revealed that all strains were alkane hydroxylase producers and seven of them produced lipase concurrently. Three best strains were characterized and identified through 16S rRNA sequence analysis as Geobacillus sp. D4, Geobacillus sp. D7, and Anoxybacillus geothermalis D9 with GenBank accession numbers MK615934.1, MK615935.1, and MK615936.1, respectively. Gas chromatography (GC) analysis showed that all three strains were able to breakdown various compounds in crude oil such as alkanes, toxic poly-aromatic hydrocarbons (PAHs), organosulfur, carboxylic acids, alkene, resins, organosilicon, alcohol, organochlorine, and ester. For the first time, alkane hydroxylase and lipase activity as well as crude oil degradation by A. geothermalis species were reported. Geobacillus sp. D7 is the best alkane degrader followed by A. geothermalis D9 and Geobacillus sp. D4 with 17.3%, 13.1%, and 12.1% biodegradation efficiency (BE%), respectively. The potential of thermophiles isolated can be explored further for bioremediation of sites polluted by petroleum and oil spills

Main structural targets for engineering lipase substrate specificity

Microbial lipases represent one of the most important groups of biotechnological biocatalysts. However, the high-level production of lipases requires an understanding of the molecular mechanisms of gene expression, folding, and secretion processes. Stable, selective, and productive lipase is essential for modern chemical industries, as most lipases cannot work in different process conditions. However, the screening and isolation of a new lipase with desired and specific properties would be time consuming, and costly, so researchers typically modify an available lipase with a certain potential for minimizing cost. Improving enzyme properties is associated with altering the enzymatic structure by changing one or several amino acids in the protein sequence. This review detailed the main sources, classification, structural properties, and mutagenic approaches, such as rational design (site direct mutagenesis, iterative saturation mutagenesis) and direct evolution (error prone PCR, DNA shuffling), for achieving modification goals. Here, both techniques were reviewed, with different results for lipase engineering, with a particular focus on improving or changing lipase specificity. Changing the amino acid sequences of the binding pocket or lid region of the lipase led to remarkable enzyme substrate specificity and enantioselectivity improvement. Site-directed mutagenesis is one of the appropriate methods to alter the enzyme sequence, as compared to random mutagenesis, such as error-prone PCR. This contribution has summarized and evaluated several experimental studies on modifying the substrate specificity of lipase

A survey on health status of coldwater rearing and hatcheries fish farms in Iran (Mazandarn, Gilan, Ardebil, West Azerbaiejan, East Azerbaiejan and Kordestan provinces)

The national research plan with title of "Study on health status of Iran Coldwater hatchery and rearing farms" was done in order to identification and tracking of main causative agents of recent mortality in Iran Coldwater hatchery and rearing farms and recognize of infected zones and design of landscape distribution of Epizootic Coldwater fish diseases in the country. This plan was conducted parallel and in same time in some polar provinces of Coldwater fish production such as Mazandaran, Gilan, Ardebil, West Azerbaijan, East Azerbaijan and Kurdistan in three years from 2006-2008. In this regard, about 23,21 and 20 farms in Mazandaran province, 30 farms in West and East Azerbaijan and Kurdistan provinces and 19 farms in Gilan and Ardebil were selected as randomly and tracking were done yearly from 2006-2008. This research was conducted according to regular planning consists of farms visit that was done according to statistical plan and completion of Questionnaires and sampling. The findings in Mazandaran province revealed that fish infections background and infectious diseases were increased in period of 2006-2008. In fact, 56% of all fish farms in 2006, 71% in 2007 and 85% in 2008 were recorded as infected farms. In other side, average weight of fish final products was more 500gr in consume market. So, regarding to long period of fish culture in mentioned fish occurrence of more morbidity could be expected. The streptococcus infections were most important fish bacterial diseases that have more incidence and pathogenicity in collected questionnaires. Also it revealed more occurrences in summer season in above 15oC temperature in affected fish farms. Also, Enteric Redmouth Disease (ERM) and Saprolegniasis were reported as second and third degree in examined farms. Meanwhile, findings of control, prevention and treatment of our survey revealed that using of antibiotics and detergent materials were increased in mentioned province. Indeed, 34.7% of all fish farms in 2006, 71.4% in 2007 and 75% in 2008 have applied treatment operation and using of detergent materials in affected farms. In other side, frequent using of Erythromycin antibiotic was reported several times in 2008 so fish farmer faced to Bacterial resistance and should be using from Florfenicol as alternative antibacterial. Also, in feed sampling from all fish farms about 3 fish farms in 2006 and 5 fish farms in 2008 were faced with food poisoning that originated from high TVN and peroxidase in fish food consumed. These farms revealed mass mortality that stopped after change feeding regime. In conclusion absence of fish health management could be considered as main agent of mentioned mortality in Mazandaran province. The findings of our survey in West and East Azerbaijan and Kurdistan revealed that absence of sufficient experiences in fish farmers and their neglect from water quality concepts, farm cleanness, using of suitable detergent materials for fish ponds and instruments and disability in continuous monitoring of physic-chemical factors of consumed water could be considered as most important problems. So, fish morbidity and mortality, growth decrement and low production rate were expected. Unfortunately, despite the occurrence of infectious and non- infectious diseases in examined fish farms, there were no documents and information about diseases history, clinical signs in dead fish, feeding regime, fish mortality (rate of daily mortality, age and weight of moribund fish), treatment operation, drugs usage, growth rate, physico-chemical factors and fish density. So planning for control and prevention of mentioned diseases were unsuccessful in affected fish farms. Also, occurrence of some epizootic fish viral diseases such as VHS, IPN and IHN were observed frequently with mass mortality about 40100% in some examined fish farms that produced some tragic economic lost in the mentioned provinces. Similarly, some bacterial, fungal and parasitic diseases were observed repeatedly in some inspected farms such as Flavobacterium, streptococcus infections. Also, some unknown causative agents in parasitic diseases were observed several times such as Dactylogyrus, Gyrodactylus, Ichthyophthirius multifiliis, Trichodina, Chilodonella, Diplostomum and fungal diseases such as Saprolegniasis. In conclusion it seems that absence of fish health management, inadequate rate of hygienic technical services, absence of quarantine programs for transfer of new eyed-eggs, juvenile and live broodstocks packages without health certificates from authorized organizations, uncontrolled entry of foreign eyed-eggs (France, Denmark, Australia and Armenia) and neglect in using of suitable detergent materials for fish ponds, instruments, consumed water and infected eggs, not screening of broodstocks in hatcheries could be considered as most important causative agents in occurrence of infectious diseases and main problems in affected farms in mentioned provinces. Similarly, our findings in Gilan and Ardebil provinces revealed that their problems were similar to other provinces. In fact, neglect in establish of necessary Infrastructures in fish farms and dereliction in health management concepts could be considered as main reasons of occurrence of fish infectious and non-infectious diseases in studied fish farms in mentioned provinces. In fact, often fish farms visited hadn't suitable structures without control and prevention approach. There were not observed detergent using, quarantine programs and prevention methods. Entrance of unknown persons, birds and wild animals were ordinary in mentioned farms. Unfortunately level of farmer’s knowledge was low and no training courses and extension programs were planned by authorized organizations. Meanwhile, food storages were unsuitable and food packages were stored in inappropriate situation so poisoning conditions were increased in examined farms. In conclusion it could be finalized that Coldwater fish farms in Gilan and Ardebil provinces situated in insufficient position and correction of current situation are needed urgently. In final elicitation, it would be mentioned that lack of basic infrastructures could be introduced as most important reasonfor spread of diseases, mortality and related economic losses in studied provinces. Therefore, attention to environmental affairs and access to Sustainable Development are recommended. Also consideration of biosecurity regulations and health management concepts would be important requirements for modification and reformation of Coldwater fish farms in examined provinces towards A Better Tomorrow

PROTOZOANS; (COCCIDIA AND MYXOSPOREA;) INFECTIONS IN SOME FISHES OF ARAS AND MAHABAD DAMS (NORTHWEST OF IRAN)

During a protozoological study on the fishes of West-Azarbayejan province (Northwest of Iran) in 1998, three different protozoans were revealed. Myxobolus musayevi from Capoeta capoeta, Myxobolus dispar from Aspius aspius taeniatus and Goussia carpelli from Cyprinus carpio. Altogether 13 different fish specimen were examined. The fishes including: Aspius aspius taeniatus (2), Rutilus ruti/us (3), Capoeta cpoeta (2) and Cyprinus carpio (6). They were caugth from Aras and Mahabad Dams and transported alive to the laboratory where they were wieghted and measured. Then, they were examined for protozoan infections when their spinal cord was cut. Myxobolus musayevi had already reported from Capoeta capoeta from Taj an in Mazandaran province. This is the first record of Myxobolus dispar from Aspius aspius taeniatus from Iranian freshwaters. Goussia carpelli also had already recorded (unpublished) from the Cyprinus carpio from Sepeedroud River in Guilan province. In this study, three of C. carpio which were caught from Aras Dam had been infected by the oocyst of the Goussia carpelli in the mocus of the intestine

Parasitological investigations on Rutilus rutilus caspicus from south-east of the Caspian Sea (Iran)

A parasitological investigations on Rutilus rutilus caspicus from south-cast of the Caspian Sea (coast of the Mazandaran and Golestan Provinces, Iran} was carried out in 1997. During this study a total of 140 fish specimen were examined. The list of parasites and their prevalence of infections are as follows: Anisakis larvae, in live fish (37%), Aspidogaster limacoides in the intestine or only one fish (0.7%). Bothriocephalus gowkongensis in the intestine of two fishes, Dactylogyrus turaliensis the gills of 83 fishes (67%) and finally Diplostomum spathaceum in the eyes of 10 fishes (12%). This is the record of Anisakis larvae, Diplostomum spathaceum, Aspidogaster limacoides from the rutilus caspicus in Iran, under the certain condition, these parasites may produce mortality in the farms

Survey on Myxosporean Parasites in the Mazandaran and Guilan Provinces (Southern Part of Caspian Sea)

A parasitological survey was carried out during 1994-1996 in Mazandaran and Guilan Provinces (Caspian Sea). 159 samples from 17 different species were examined. Samples were collected from Tajan River, Anzali Lagoon and some fish farms. Immediately after caught live fish were transported to laboratory and kept in aquarium for further examination. During this study ten species of Myxosporean parasites were separated, identified and classified. This is the first report on infestation of inland fishes of Iran by these ten parasite species. Morphology of spores, development stages of plasmodia, tissue specifications and cyst production of these parasite species were investigated in the present study. Four of the Myxosporeans were identified as potential pathological hazards for three fish species. These are Myxobolus pavlovski for Silver Carp (Hypophthalmichthys molitrix) and Big Head (Hypophthalmichthys nobilis), M. bramae for Kutum (Rutilus frisii kutum). M.muelleri and M. ellipsoides f....