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1. Isolation And Characterization Of Phytase Producing Microrganism From Soil

by Ghazal Aziz | Dr. Aftab Ahmad Anjum | Prof. Dr | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2013Dissertation note: Phytase is an enzyme of great importance because it is added as a biofertilizer to soil and added in animal feed to increase the uptake of inorganic phosphorous. Phytase production is the property of plant growth promoting rhizobacteria (PGPR) that harbor in rhizosphere part of the soil. These phytase producing bacteria can be utilized as biofertilizers as and can increase the soil fertility and crop production. Soil samples were collected and screened for the production of phytase (an extracellular) enzyme on phytase screening media (PSM). Six bacterial isolates (PHY02, PHY03, PHY06, PHY07, PHY12, and PHY30) showed distinguished clear zones (> 6mm) on PSM. Isolates were identified as Lactobacillus casei PHY02, Enterobactor intermedius PHY03, Bacillus badius PHY06, Escherichia coli PHY07, Shigella sonnei PHY12, and Klebsiella pneumonia PHY30. Effect of physical parameters (temperatures, pH and osmotic pressure) on growth and enzyme production by selected isolates was determined. Optimum growth and production of phytae by PHY02, PHY03, PHY06, PHY07, PHY12, and PHY30 (27, 9, 19, 40, 32, and 19 IU, respectively) was at 37°C. PHY07 showed highest enzyme production, followed by PHY30 and PHY02. Isolate PHY06 showed similar growth and enzyme activity at 37°C and 42°C but it was significantly reduced at low temperature. Effect of pH on phytase production on selected isolates indicates that all isolates produces maximum amount of phytase at pH 6.5. At pH 6.5 enzyme units released by PHY02, PHY03, PHY06, PHY07, PHY12, and PHY30, were 26, 15, 19, 41, 19, and 32 IU, respectively. Production of enzyme decreased with the increase in osmotic pressure. PHY02, PHY03, PHY06, PHY07, PHY12, and PHY30 showed optimum enzyme production (27, 15, 17, 41, 18, and 32 IU, respectively) at 1 % NaCl in PSM (Figure 1C). Effects of carbon source on both growth and phytase production of isolates showed that PHY03, PHY06, PHY07, PHY12 had significantly higher (P<0.05) cell densities and enzyme production in glucose, while PHY02 and PHY30 had higher enzyme activity at 0.3% lactose. Nitrogen source in growing media also effects the growth and production of enzyme. PHY02 and PHY12 had better growth and production at 0.1% peptone, while PHY07 and PHY30 had significantly higher phytase level in media modified with peptone but at higher concentration (0.3%). Addition of tryptone in growth medium significantly enhanced the growth and enzyme production by PHY03, and PHY06. Availability: Items available for loan: UVAS Library [Call number: 1685,T] (1).

2. Seroprevalence Of Bluetongue In Domestic Animals

by Farid ahmed khan | Prof..Dr. Khushi Muhammad | Ms. Sehrish | Prof. Dr Tahir yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2014Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1925,T] (1).

3. Efficacy Of Commercial Disinfectants Against The Water Contaminating Bacteria At Commercial Broiler Farms

by Mian Muhammad Salman | Dr. Aftab Ahmad Anjum | Pfor. Dr | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print Publisher: 2012Dissertation note: Water is an important constituent for poultry. Poor hygienic conditions of water are health hazard for poultry. Many outbreaks are caused by consuming poor quality water.. Fifty water samples from different broiler farms in and around Lahore were collected from drinkers in sterile containers. Bacterial load was assessed using total viable count and coliform count. The counts were above the threshold level (50cfu/ml for coliform and 100cfu/ml for total viable count) showing that water used at poultry farms was of low microbiological quality. Five Disinfectants PHMB20% (.75ml/lit, 1.5ml/lit, 3.0ml/lit), PHMB11% (1.5ml/lit, 3.0ml/lit, 6ml/lit), 0.2% chlorine dioxide (0.1ml/lit , 0.2ml/lit, 0.4ml/lit) Glutral 9.8%(1.5ml/lit, 3.0ml/lit, 6ml/lit), organic acid(1.5ml/lit, 3.0ml/lit, 6ml/lit) were used and they resulted in log reduction of TVC by PHMB20% (5.83±4.36, 6.14±3.98, 9.35± 0.68), PHMB11% (9.42±0.21), 0.2% chlorine dioxide (2.45±0.97, 3.19±0.73, 6.33±0.80 ) Glutral 9.8%(6.87±1.00, 9.73±1.00,9.73±1.00), organic acid(4.75±1.21, 6.62±1.26, 6.90±1.15).PHMB20%,PHMB11%, Glutral 9.8% and organic acid were effective at normal dose, while 0.25 chlorine dioxide was effective at normal dose against at normal dose. Log reduction in Coliform count at half, normal and double dose by PHMB20% (6.52±3.33, 6.96±2.46, 7.96±0.98), PHMB11% (7.89±1.01), 0.2% chlorine dioxide (3.65±0.73, 5.08±0.98, 6.27±0.97) Glutral 9.8%(8.48±0.99), organic acid(5.18±1.21, 5.93±1.26,6.46±1.15±) . PHMB20%, PHMB11%, 9.8% Glutral, organic acid were effective against coliform bacteria at half dose while 0.2% chlorine dioxide was effective at normal dose. Glutraldehyde was effective at normal dose amongst all disinfectants against Total viable bacteria and coli form bacteria. Availability: Items available for loan: UVAS Library [Call number: 1482,T] (1).

4. Detection Of Soulfonamide Residues With Associated Histopathological Findings In The Tissues Of Cattle An Buffalo

by Mujahid Iqbal | r. Muhammad Yasin Tipu | Dr. Asim Aslam | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print Publisher: 2012Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1483,T] (1).

5. Characterization Of Mycoplasma Gallisepticum Isolates And Their Use In The Production Of Indigenous

by Mushtaq Ahmad | Prof. Dr. Masood Rabbani | Prof. Dr | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print ; Literary form: drama Publisher: 2012Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1564,T] (1).

6. Molecular Epidemiology Of Subclinical Tuberculosis In Peri-Urban Human Population Of Lahore.

by Sadeem Shahzad | Dr. Muhammad Yasir Zahoor | Dr. Muhammad | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print ; Literary form: drama Publisher: 2013Dissertation note: Tuberculosis (TB) is known to be a major health problem worldwide causing disease among millions of people every year. Major cause of tuberculosis in human is the infection with M.tuberculosiswhich usually causes pulmonary or lungs TB but an unknown number of patients are also infected with M.bovis which causes tuberculosis in humans as zoonotic agent along with its major hosts like cattle and deer. In developing countries where raw milk is used without pasteurisation there is a heavy risk of tuberculosis infection with M.bovis. TB infection with M.bovis mainly appears as extra pulmonary tuberculosis with and without specific symptoms of the disease.Diagnosis of subclinical asymptomatic tuberculosis and that of extra pulmonary tuberculosis is a difficult task and most of the time disease remains undiagnosed or misdiagnosed due to the unavailability of specific and sensitive diagnostic tool to diagnose the disease at early stage. Moreover prevalence of M.bovisinfection is not properly known. This study was designed to measure the diagnostic value of Interferon gamma release assay (IGRA) for early and reliable diagnosis of subclinical extra pulmonary TB along with the molecular epidemiology of subclinical extra pulmonary TB to check the prevalence of M.bovisinfection. IGRA is a latest blood test with high specificity and sensitivity based on the principle of Interferon gamma released by effector T-Cell when exposed to M.tuberculosis antigens like ESAT-6 and CFP-10 in controlled in-vitro conditions. Eighty patients were selected for the study on the bases of the history of having day to day cattle contact along with feelings of sickness. Biopsy tissue samples of all the patients which were positive with IGRA were requested, however 24 out of 27 positive samples were collected and were first examined histologically. Twenty seven samples out of eighty were found positive with IGRA while 22 out of 24 samples were confirmed by histological examination as infected with MTB. Both IGRA and histological examination are unable todifferentiate between the specie specific infection with M.tuberculosis orM.bovis for which differential amplification of specific fragments of bothof the species was done by running a multiplex PCR using M.tuberculosis specific 185 bp pncA product and M.bovis specific 500 bp segment. Genomic DNA was extracted from previously formalin fixed paraffin embedded (FFPE) tissues which requires pretreatment for deparaffinization. Xylene was used as deparaffinization agent. All of the twenty two samples positive with IGRA and histological study were found positive for M.tuberculosis infection and none of the sample was found positive for M.bovis infection. Results showeda close correlation among all three techniques with their specific benefits and limitations. Study concluded that T.Spot TB (IGRA) is a potentially reliable test for the diagnosis of subclinical, extrapulmonary TB.Formalin Fixed Paraffin Embedded (FFPE) tissues may be used for TB diagnosis and other DNA based researches. Duplex PCR is a reliable technique for differential diagnosis of infection with different species of MTB complex, though none of the sample was found positive for M.boviswhich is may be due to small sample size of the study and it may further be studied in future researches. The research findings will help the clinicians to depend on IGRA testing for timely and reliable diagnosis of extrapulmonary subclinical tuberculosis and potential use of FFPE tissue samples as appropriate specimen for molecular based diagnosis of TB. Further studies are however, required to check the prevalence of M.bovis infection byincreasing sample size. Availability: Items available for loan: UVAS Library [Call number: 1621,T] (1).

7. Isolation And Characterization Of Auxin Producing Bacterial Strains From Plant Rhizosphere

by Kanwal Aziz | Dr. Jawad Nazir | Dr | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print ; Literary form: drama Publisher: 2013Dissertation note: Auxins are a class of plant hormones or plant growth substances. Auxins have a cardinal role in the regulation of many growth and developmental processes in the plant. Soil samples were serially diluted and screened for auxin production by Salkowski method. Isolates that have the ability to produce auxin were identified by culture characters, morphology, and bio-chemical profile. From 150 isolates, 04 bacteria were selected (AUX-36, AUX-53, AUX-137, and AUX-142). The bacteria were identified by following the flowcharts described in “Berges Mannual of Determinative Bacteriology”, 9th addition. These isolates were identified as Bacillus megaterium, Escherichia coli, Klebsiella pneumonia and Bacillus marinus respectively. Next, different physical and chemical parameters for growth of bacteria and auxin biosynthesis were optimized. For the optimization of bacterial growth OD values of the culture broth (at wavelength 600nm) was taken by spectrophotometer. To estimate the amount of auxin (ìg/ml) produced in the culture broth; a standard curve (concentration of auxin ìg/ml at x-axis and OD value at y-axis) was prepared by using commercially available auxin. The optimum conditions for growth and auxin production by AUX-36 was found to be pH 7, 0.98% osmotic pressure at 37 °C after 72 hours of incubation. If the medium is supplemented with 0.1 and 1.0% glucose, sucrose and peptone then it increased the bacterial growth which ultimately increased the auxin concentration in the broth medium. The growth and auxin Production by AUX-53 was 0.98% NaCl concentration at 37 °C after 72 hours of incubation. The optimum pH was found to be 7 but it showed good growth at acidic as well as alkaline pH. The addition of glucose and sucrose in the growth medium increased the growth as well as auxin production. The optimum conditions for the growth of AUX-137 were as follows: pH=7, 0.98% osmotic pressure, temperature 37 °C. However the isolate had good growth at 28 °C and 2% NaCl concentration as well. The bacterial cell density and auxin increased with incubation time up to 72 hours. The isolate produced highest concentration of auxin under the same conditions. Similarly, the cell density and auxin increased with the increasing concentration of glucose in the growth medium. Sucrose increased the auxin only in the culture filtrate. While the bacteria AUX-142 showed highest growth as well as auxin production at 42 °C after 72 hours of incubation. The optimum pH and osmotic pressure was found to be 7 and 2% respectively. The cell density and concentration of auxin increased with the increasing concentration of peptone in the growth medium. Addition of tryptophan (1-2%) increased the auxin concentration in the culture supernatant of all isolates. Next, the seed germination test and plant pot experiment were performed of selected isolates to observe the effect of bacterial inoculation on wheat plants. In seed germination test treatment of seeds with AUX-36, AUX-53 and AUX-142 significantly increased the root length and number of root hairs as compared to non-treated seeds. In plant pot experiment comparison of various growth parameters of inoculated plants with non-inoculated plants revealed the improvement in plant growth by bacterial inoculation. Availability: Items available for loan: UVAS Library [Call number: 1684,T] (1).

8. To Investigate The Morphology Of Lip Prints And Their Effectiveness In Individualization And Sex Determination

by Makhdoom Saad Waseem Ghouri | Dr. Muhammad Wasim | Dr. Abu Saeed | Prof. Dr. Tahir Yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2013Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1720,T] (1).

9. Genetic And Evolutionary Characterization Of Pakistani Pigeons And Parrots Through Mitochondrial D-

by Sehrish firyal | Dr. Ali raza awan | Prof, Dr. Aftab | Prof, Dr. Tahir yaqub.

Material type: book Book; Format: print Publisher: 2013Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1873,T] (1).

10. Cytochrome B Gene Amplification A Novel Approach For Diagnosis Of Theileriosis In Cattle Under Field

by Muhammad Faiz rasool | Prof. Dr.Kamran ashraf | Dr.Nisar ahmed | prof. Dr. Tahir yaqub.

Material type: book Book; Format: print Publisher: 2014Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1893,T] (1).

11. Isolation And Molecular Identification Of H9N2 Avian Influena Virus From Human In Punjab Province Pakistan

by Abdul ahad | Prof .Dr, Masood rabbani | Prof. Dr. Rana | Prof. Dr. Tahir yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2012Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1926,T] (1).

12. Polymorphism Identification In Bovine Growth Hormone Gene And Association With Milk Production Trait

by Aasma Manzoor | Dr.Asif Nadeem | Ms. Faiza | Prof.Dr. Tahir yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2014Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1968,T] (1).

13. Genetic Evaluation Of Cyp19A1 As A Candidate Gene For Silent Estrus Behavior In Nili-Ravi Buffalo

by Sana Imran | Miss. Maryam javed | Miss.Asma | Prof. Dr. Tahir yaqub.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2014Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1969,T] (1).

14. Antibody Resoinse Of Broilers To Oil Based Combined Mycoplasma Gallisepticum And Avian Influenza H9

by Sadi Sarfaraz | Prof. Dr. Khushi Muhammad | Prof.Dr. Asim | Prof.Dr.Tahir yaqub.

Material type: book Book; Format: print Publisher: 2014Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1976,T] (1).

15. Production Of Laccase By Immobilized White Rot Fungi And Its Application For The Decolorization Of Textile Effluent Dyes

by Iqra Ghulam Rasool (2012-VA-579) | Ms. Faiza Masood | Dr. Muhammad Tayyab | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2014Dissertation note: Textile wastewater effluent contains several types of dyes that are toxic, carcinogenic, and dangerous for environment (Nyanhongo et al. 2002). More than 10,000 different kinds of dyes and pigments are used in dyeing and textile industries. Approximately 8, 00, 000 tons colorant is produced annually and 10% of used dyes are enters the environment in the form of wastes. There are different types of textile dyes such as direct dyes, disperse dyes, reactive dyes, acid dyes, and basic dyes. Wastewater effluents discharge from textile industries contain more than 10-15% of these dyes (Kunamneni et al. 2007). Such wastewater effluents are being discharged into water stream without or after only partial treatments, causing water pollution and negatively affecting the aquatic life. The treatment of textile wastewater effluents are of major environment concerns (Nyanhongo et al. 2002). White rot fungi (WRF) is a wide class of fungi and it is mostly comprised of basidiomycetes, ascomycetes and lignin-decomposing fungi (Wesenberg et al. 2003). WRF are the most abundant wood degraders, and are so named because they leave a bleached appearance of the wood fibers following their attack. WRF has the ability to degrade contaminants by virtue of the nonspecific nature of its extracellular ligninolytic enzyme system (Nyanhongo et al. 2002) The white rot fungus is also known as lignin degraders because it degrades lignin effectively due to some enzymes present in it. The important enzymes involves in degradation of lignin are following: (i) lignin peroxidase: It oxidizes both phenolics and non pheolics compounds, (ii) manganese-dependent peroxidase, (iii) laccase: It oxidises phenolic compounds and produce phenoxy radicals and quinones; (iv) glucose oxidase and glyoxal oxidase used for H2O2 production, and (v) celloulobiose quinone oxidoreductase for quinone reduction (Kunamneni et al. 2007). Laccase (oxidoreductase, EC belongs to polyphenol oxidases group of enzymes. Copper atoms are present in the catalytic center of enzyme so it is also known as multicopper oxidases (Baldrain et al. 2006). The molecular mass of laccase is 50–100 kDa (Couto and Toca 2006). According to the mechanism of laccase, it carries out the reduction of oxygen to water along with the oxidation of its substrate. Laccases oxidize wide range of compounds such as polyphenols, methoxy substituted phenols, aromatic diamines, and other compounds (Baldrain et al. 2006). The substrate specificity of laccase is very wide and broad. In ortho and para substituted mono and polyphenolics substrate, it carries out reduction by removing hydrogen atom from hydroxyl group. In aromatic amines, it removes one electron and produces free radicals. These radical are able of many other reactions such as depolymerization, repolymerization, demethylation, or quinone formation. During lignin degradation, oxidation of methoxyhydroquinones followed by auto-oxidation of the methoxysemiquinones. Furthermore, formation of superoxide anion radicals undergoes more chemical reactions. The activity of laccase may be increased by using different kind of activators, such as ABTS (2, 2-azinobis (3-ethylbenzthiazoline- sulfonic acid), 1-hydroxybenzotriazole, or compounds secreted by fungi (Abadulla et al. 2000). In the presence of ABTS, the decolorization efficiency increases up to 45% (Tong et al. 2007). Laccases have been produced from different kind of sources such as some species of fungus like white rot fungi, different kinds of bacteria, and some insects (Heinzkill et al. 1998; Diamantidis et al. 2000; Dittmer and Kanost 2010). This enzyme is widely distributed in Ascomycetes, Deuteromycetes, and Basidiomycetes, WRF is the major source for the production of laccase enzyme because this fungi is involved in metabolism of lignin (Bourbonnais et al. 1995). There are many applications of fugal laccases such as effluent decolorization discharged from industries, degradation of pulp released from paper and pulp industries, removal of phenolics compounds from alcohols, synthesis of organic compounds, biosensors, pharmaceutical sector (Yaver et al. 2001). This enzyme can also improve animal performance, increase nutrient digestibility when added to animal feed (Sharma et al. 2013). Fungal laccases have higher redox potential of +800mV as compared to plants or bacterial laccases that’s why there are several applications of laccase in biotechnology field especially in the decolorization of dyes. Enzymes can be produce in larger amount so that laccase based decolorization techniques are advantageous to bioremediation technologies (Devi et al. 2012). Pleurotus is a species of WRF and few laccases have been isolated, purified and cloned from Pleurotus species. However, the physiological significance of laccase produced by the white rot fungi is not known. Literature reports that mycelia culture of Pleurotus florida produces at least two laccases (L1 and L2), one of which appears to be linked with the mycelia growth of the fungus (Das et al. 1997). The L1 isoenzyme is dominantly involved in the dye decolorization process. Submerged fermentation (SmF) is a type of fermentation in which microorganism is grow in liquid broth and enzymes and other compounds are released in the broth. This technique used free liquid substrates such as nutrients etc. The substrates are utilized quite rapidly and constantly supplemented with nutrients. In fermentation broth, microorganisms are provided with appropriate nutrients and conditions such as high oxygen concentration for the production of microorganism in order to get desired products. In this technique, mycelium formation is takes place. Mycelium formation can lead to pellet formation which hinders the diffusion of oxygen and nutrients in the medium. In recent times, wide variety of secondary metabolites has been produced commercially by fungal fermentation. Fungi are complex microorganism that is different morphologically and structurally at different phases of their life cycles form others. It is also differ in form between surface and submerged growth in fermentation media. Nature of liquid media also effect on the growth of fungi. Different culture conditions such as temperature, pH and mechanical forces are important for fungi growth but these parameters are different for different fungi (Kossen et al. 2000). Enzymes act like catalyst and they speed up any chemical reaction without being used up in the reaction. The uses of enzymes are advantageous due to its several characteristics and features as compared to conventional chemical catalyst. However, there are some problems that can reduce the operational life time of any enzymes. These problems includes; non-reusability of enzyme, the instability of their structure, high cost of isolation, purification and characterization and their sensitivity to harsh condition of reaction. These objectionable limitations of enzymes may be reduced by the use of immobilized enzymes. There are mainly four procedures present for immobilization of any cell (Kunamneni et al. 2007). These procedures are following: adsorption, gels entrapment or polymer entrapment, covalent coupling, and cross-linking to insoluble matrices (Brouers et al. 1989). For immobilization different kinds of matrices, such as agar, calcium alginate beads, polyacrylamide gel, etc have been used. In order to select suitable matrix and immobilization procedure, type of the cell, type of the substrate, medium conditions and products are major factors (Prasad et al. 2005). During immobilization, enzyme is fixed to or within solid matrix in order to get heterogeneous immobilized enzyme system. Naturally enzymes are bounded to cellular membrane in living cells for most cases so in order to get the natural form of enzyme, immobilization of the cell is done. This immobilized system stabilized the structure and activity of the enzyme for longer period of time. When enzymes are immobilized, they are stronger and more challenging to harsh environment changes. Immobilization also allows easy recovery of enzyme and also it’s multiple re-use in processes. The Michaelis constant of immobilized enzymes increased and their activity usually lowered when compared to free enzyme. When immobilization procedure applied, different structural changes introduced to an enzyme which leads to these alterations. Immobilization helps to maintain the structure, stability and activity of enzyme for longer time without being de-activated (Kunamneni et al. 2007). Immobilization represents an attractive option to obtain enzymatic catalyst for dyes treatment. This technique provides different advantages: (i) it prevents enzyme leakage even under harsh conditions; (ii) it facilitates enzyme use in different types of reactors like packed bed, stirred tank and continuous bed; (iii) it causes stabilization of the enzyme tertiary structure, usually as a consequence of multipoint attachment of the enzyme to the support, providing enzyme rigidity. The stabilization provided by covalent bonding is usually counter balanced by partial enzyme deactivation. This negative effect can be mitigated by carefully optimizing the immobilization conditions in order to maximize the ratio between immobilized enzyme activity and activity of the primary enzyme solution (Pezzella et al. 2014). Immobilization of laccase was extensively investigated with broad range of different techniques and substrates. Inactivation of enzyme occurs when oxidized products are absorbed on the surface of the immobilization matrix support (Kunamneni et al. 2007). Textile industries discharged wastewater effluents comprised of toxic dyes are dangerous for aquatic life and have harmful impacts on the environment. There are different methods including physical and chemical methods which are use previously to decolorized dyes. These physical and chemical methods are quite costly, prolonged, ineffective and insecure (Shang and Chi 1996; Mechichi et al. 2006). In comparison to these methods, biological processes are quite suitable and helpful. Biological processes are less expensive, safe and take less time and effective. Biological processes used microorganisms to decolorize dyes. Laccase as an extracellular oxidative enzyme produced by white rot fungi are eco-friendly and cheap. In order to decolorize dye, three day old fermentation media is used and dyes is added in the broth. To get 70-75% decolorization in fungal culture, more than 48 hours are required. Pleurotus Species produced laccase efficiently and this laccase could decolorize malachite green dye upto 70% within 24 hours (Yan et al. 2009). Laccases can degrade several dye structures such as phenol, polyphenols and diamines (Abadulla. et al. 2000) to degrade harmful compounds into less toxic compounds and may be helpful to reduce environmental pollution (Gianfreda et al. 1999). The specific features and mechanism of laccase helps to make it a versatile biocatalyst. Due to its versatility, it is suitable for several applications such as biopulping, biobleaching, and industrial wastewater treatment. Due to the severe environment legislation, the textile industry is trying to introduce new innovative technologies for the treatment of wastewater effluents discharged from textile industries. Laccase has potential to degrade dyes of various chemical structures so that development of techniques based on laccase seems an attractive and suitable solution in decolorizing dyes (Madhavi and Lele 2009). The decolorization and detoxification of the wastewater effluent would help to use it again and again in dying process in textile wet processing. The major purpose of this research is to decolorize the textile effluents dyes discharged by industries after partial treatment and cause water pollution and have negative effect on aquatic life and ecosystem. It is necessary to established most effective and efficient method to produce sufficient amount of laccase enzyme through immobilized white rot fungus and then utilized it in the process of bioremediation. Availability: Items available for loan: UVAS Library [Call number: 2208,T] (1).

16. DNA Based Characterization Of Protease Gene From Geobacillussp.Sbs-4s

by Anam Shabbir (2012-VA-608) | Dr. Muhammad Tayyab | Ms. Huma Mujahid | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2014Dissertation note: Proteases are hydrolytic enzymes responsible for the hydrolysis of proteins(Qadar et al.2004).These enzymes contribute major role in textile and leather industry,accounting 60% of the world wide enzyme market(Nascimento et al.2004).These enzymes are also being used in food ,pharmaceutical ,detergent, brewage sweet industry and as digestive additives in human and animal feed (Wilson, 2012). Proteases are produced by microbes,animal and plants but microbial proteases are preferred due to ease in production and cheaper cost (Ningthoujam et al.2010).Microbes produce a variety of proteases according to their requirement that are specific in their function (Neurath 1999).Microbes might be involved in the production of intra or extracellular proteases.Extracellular proteases help the organism to absorb and utilize hydrolytic products from proteinious substrates in order to get energy by catabolism or to synthesize the biomolecules through anabolism reactions(Ningthoujamet al.2010). Proteases can be classified in different ways.On the basis of cutting preferences these can be divided in to two groups:endopeptidases and exopeptidases (Barret and Mcdonald 1985).Exopeptidases are involved in hydrolysis of the peptide bond near N or C terminal whereas endopeptidases are responsible for the hydrolysis of peptide bond, with the chain, distant from the peptide ends(Motyan et al .2013).On the basis of catalytic residues in active site the proteases can be divided into six groups including glutamate,serine, therionine cysteine,aspartate and metalloproteases(Li et al.2013). Microorganisms occupy all possible environments including habitats that provides appropriate conditions for growth(Sharma et al.2009).Thermophiles have ability to grow at highertemperature whereas other microbes fail to survive.There has been increasing interest in thermophilic bacteria because of their thermostable enzyme(Obeidat et al.2012).Hyperthermophiles can survive in extremely hot environment. Hyperthermophiles occupy the most basal positions of the phylogenetic tree of life(Bouzas et al. 2006). About 70 species of hyperthermophilic bacteria and archea has been isolated from different terrestrial, marine and thermal areas in the world.Hyperthermophiles are very divergent in their phylogeny and physiological properties.Proteolytic enzymes from hyperthermophiles are catalytically active at high temperature and they can alsoretain their catalytic activity in the presence of detergent and other denaturing substances (Stetter et al.1993). Geobacillusis widely distributed thermophiles isolated from geothermal areas (Chalopagorn et al.2014).On the basis of16SrRNA gene sequences, Geobacillus belongs to Bacillus genetic group 5. It is phenotypically and phylogeneticallyconsistent group of thermophilicbacilli (Rahman et al. 2007).Bacillus and Geobacillus species are the dominant workhorses in industrial biotechnology. These bacteria produce a variety of extracellular enzymes, such as amylases, xylanases, proteases, phytases, carbonic anhydrases, catalases, pectinases. Bacillus and Geobacillus species hasability to grow at acidic, alkaline, neutral pH and at elevated temperature has positioned them among the most important industrial enzyme producers(Satyanarayana et al. 2012). Geobacillus are gram-positive, rod-shaped, aerobic,endospore-forming obligate thermophiles.The growth temperature for various Geobacillus species ranges from 37 to 75 °C and pH range of 6.0 to 8.5.The members of Geobacillusare homologus to each other and share homology 99% among them(Tayyab et al.2011). The genus Geobacillusthermophilicstrains, produce a variety of thermostable hydrolytic extracellular enzymes, such as proteases, amylases, and lipases used in various industrial applications (Wiegand et al. 2013) GeobacillusSBS-4S was isolated from a hot spring located in Gilgit, Northern areas of Pakistan.Geobacillus SBS-4S strain is Gram positive, rod-shaped bacteria and occurs in chains. That could grow at a wide range of temperature (45 to 75˚C) and pH ranging 5.5 to 9.5.Geobacillus SBS-4S produced several extracellular enzymes including amylase, protease and lipase.The comparison of the strain SBS-4S with the already reported species of genus Geobacillus showed that SBS-4S is resistant to antibiotics such as streptomycine, spectinomycin and rifampicin(Tayyab et al.2011). Availability: Items available for loan: UVAS Library [Call number: 2242-T] (1).

17. Seroprevalence And Molecular Detection Of Brucellosis In Hospitalized Patients With Clinical Manifestations Of Brucellosis

by Riffat Yousaf (2008-VA-342) | Dr. Wasim Shehzad | Prof. Dr.Tahir Yaqub | Dr. Haroon Akbar.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: Brucella species are host specific facultative intracellular pathogens which cause brucellosis in both animals and humans. Brucellosis is one of the most common zoonotic diseases worldwide. In Pakistan, the incidence of brucellosis is increasing day by day due to lack of awareness of this deadly malady. It is transmitted from infected animals to humans who are in close contact with infected vaginal secretions, feces, blood, aborted fetus, or by consumption of unpasteurized milk and dairy products. Infection due to B. melitensis and B. abortus are mostly prevalent for brucellosis in human. Total 218 blood samples were collected in gel vacutainer tubes from hospitalized patients who were clinically manifested with brucellosis. Out of 218, 12 RBPT positive blood samples were collected in EDTA containing vacutainer tubes separately. Serum was isolated from all blood samples (without EDTA). These serum samples were first screened by Rose Bengal Plate Test (RBPT). DNA was extracted from all positive RBPT blood and serum samples and randomly selected negative RBPT serum samples. All extracted DNA (≤10ng/µL) were subjected to Brucella genus and two species specific (B. abortus and B. melitensis) Quantitative Real Time Polymerase Chain Reaction (qRT-PCR) assay. Furthermore, few selected extracted DNA (≥20ng/µL) from blood and serum samples were examined by genus and Multiplex specie specific PCR. The PCR products were electrophoresed on 2.5% agarose gel. Then selected products were sequenced by ABI 3130 XL sequencer. The data were analyzed by SPSS software using Chi square test. The present study helped to diagnose accurately and precisely brucellosis in clinical manifested patients, which is further helpful for devising the strategies to control this disease. Availability: Items available for loan: UVAS Library [Call number: 2286-T] (1).

18. Efficacy Of Surface Disinfectants Against Bacterial Pathogens On Experimentally Contaminated Pathogens

by Sana Ahmed (2009-VA-241) | Dr. Jawad Nazir | Dr. Amir Ghafoor | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: Surface disinfection plays a key role in the prevention, control and reduction of many communicable infections as contaminated surfaces are the major source for transmission of microorganisms. Five types of surface cleaning products (Astonish Germ Clear, Cif Floor Cleaner, Dettol Surface Cleaner, Finis Floor Cleaner and Lysol Surface Cleaner) were evaluated for antibacterial activity against three bacterial cultures (E. coli, K. pneumonia, S. aureus) through Quantitative non-porous Surface Carrier Test derived from EN 13697. Efficacy testing was performed through surface contamination techniques. Glass slides surfaces were artificially contaminated followed by recovery of the bacteria through vortex method both before and after the application of product for 5 minutes. Each of the experiment was repeated thrice and microbicidal effect (ME) values after the application of each product were calculated. Residual antimicrobial activity of the surface cleaning products was measured by applying the working solution of disinfectant on contaminated glass surfaces. Exposure time was given to the test surfaces, after each set exposure time the surfaces were treated to recover the microorganisms. Viable count from the eluant was calculated by serial dilution spread plate method. Each of the experiment was repeated thrice to find out the residual antimicrobial effect of the disinfectant products. ME values of the Finis Floor Cleaner ranged from 4.03 to 5.14, which was the maximum value among all surface cleaning products used against E. coli. Highest ME value against S. aureus and K. pneumonia was shown by Astonish Germ Clear. The ME values ranged from 4.99 to 5.10 against S. aureus and 5.34 to 6.99 against K. pneumoniae. Finis Floor Cleaner was proved to be of maximum efficiency against E. coli where as Astonish Germ Clear was most effective against Staph. aureus and K. pneumonia. The mean log10 CFU values recovered from disinfectant treated Summary 49 surfaces when they were exposed to the environment for different time periods of five minutes, six hours and 24 hours are 5.62 to 7.94, 4.17 to 6.35 and 7.16 to 10.25 respectively. The results indicated that the microbial count was reduced significantly at interaction time of five minutes, then at six hours the count was further reduced by Astonish Germ Clear, Cif Floor Cleaner and Dettol Surface Cleaner i.e. these surface cleaners were able to maintain their antimicrobial activity upto six hours. When the exposure to environment further increased to 24 hours, the microbial count started to increase, hence none of the disinfectants has shown antimicrobial activity upto 24 hours. This indicates that significant microbial count can be achieved within the interaction time of five minutes to six hours. Loss of antimicrobial activity upto 24 hours is probably because the active ingredients of cleaning agents get degraded during long interaction time. Present study emphasizes that the surface disinfection process must be repeated at regular intervals. Regular and timely use of surface cleaning agents must be considered as a crucial measure in controlling disease transmission rates. Availability: Items available for loan: UVAS Library [Call number: 2294-T] (1).

19. Study On The Pathogenesis Of Co-Infection Of Infectious Bronchitis (Ibv) And Escherichia Coli (E. Coli) In Experimental Chickens

by Sohail Khan (2013-VA-606) | Prof. Dr. Asim Aslam | Dr. Muti Ur Rehman Khan | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: Infectious bronchitis and colibacillosis are infectious diseases affecting chicken of all ages and breeds. They are of major economic importance in commercial chicken flocks, causing huge losses. As both in humans and animals it is well documented that preceding respiratory infection of virus predispose individual to bacterial infection. Moreover, mix infection in poultry which occur when different organisms simultaneously invade birds is a major threat to poultry industry causes highly epidemic debilitating disease with high mortality which eventually leads to economic catastrophe. In recent past prevalence studies of field, E. coli had been reported with high prevalence and exaggerated disease along with other respiratory pathogens, additionally IBV had also isolated from same flocks in same season. Although a plethora of pioneering work had been done on IBV and E. coli in the previous decades but still a window in time exist in revealing there co-infection. Looking to field scenario in our country, the present study was designed to study an ideal challenge model for IBV and E. coli, by reproducing the natural infection. 80 SPF day old broiler birds were arranged into four groups, (A, B, C and D). Each group was comprised of 20 birds. Group D served as uninoculated control while, Group A and B were challenged with IBV on 23rd day of trails, and Group B and C were inoculated with E. coli infection on day 26th. Birds, (n=3) from each group were slaughtered on various days post infection, gross and histopathological lesions were observed and serum samples for HI were taken, throughout experiment. Variable clinical signs were recorded in various groups. In IBV infected group, respiratory distress i.e., tracheal rales, coughing, sneezing and gasping were noted during early stages, later up to 10 days post infection watery diarrhea with ruffled feathers were observed. In mix infected group clinical signs manifested rapidly and were persistent with high severity. Gross lesions in mixed infection were more profound, Summary 56 including; airsacullitis, tracheitis with catarrhal exudation throughout respiratory tract; severe sepicemic lesions i.e. perihepaitis, pericarditis, pneumonia and polyserositis with swollen and pale kidneys distended by urates. 5 birds died in mix infected group revealed ascites with asphyxiation of trachea with caseous exudate. While in IBV infected group lesions were mild and confined to trachea, airsac and kidneys. Mortality was high in mix infected followed by IBV in which two birds died. While in E. coli and control group mortality were not noted. Histopathological lesions in mix infected group were aggravated markedly tracheal epithelium degeneration, deciliation and sloughing; congestion, interstitial nephritis, leukocytes infiltration, tubular degeneration and necrosis while were observed. In lungs, pneumonia of peribronchiolar area and interstitium with lymphocyte and macrophages infiltration, additionally degeneration and vacuolization of hepatocytes with focal necrotic areas were also noted. In IBV and E. coli group microscopic lesions were of mild degree. GMT of both IBV and mix infected birds were high but were not significant different (P>0.05). Among the groups, statistically significant increase in FCR of birds in mix infected group was observed followed by E. coli, with IBV infected came third in the row. On the bases of these findings we might conclude that mix infection of IBV and E. coli causes severe lesions with high morbidity and mortality. Availability: Items available for loan: UVAS Library [Call number: 2306-T] (1).

20. Genetic Characterization Of Pakistani Wild Quails Using Mitochondrial Coi Gene

by Wajiha Shakil (2012-VA-817) | Dr. Ali Raza Awan | Dr. Muhammad Yasir Zahoor | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: A latest taxonomic tool termed as DNA barcoding is being used to genetically characterize animals. DNA barcoding is helpful in determining evolutionary relationship among species. Being a small sized genome and agile enough to show rapid mutation, mtDNA has been used as a pertinent marker of molecular biodiversity. DNA barcode works as an efficient tool in food manufacturing industry, diet investigation, forensics analysis, preventing unlawful trade and felonious poaching. The aim of this study was to develop DNA barcode for genetic characterization of Pakistani wild quail. Pakistani wild quail is important due to its demand for eggs, meat production, experimental purposes and gaming as well. Japanese quail was also included in this study because this quail is excessively produced in Pakistani farms. Japanese quail is present throughout the year and is comparatively bigger in size than wild quail. It has longer lifespan; farmers can easily breed this species in farms. It is suitable in poultry due to better meat yield. COI gene (500bp) was used as a molecular marker for identification at species level. DNA was extracted from blood samples of ten wild quails (Coturnix coturnix and fifteen japanese quails (Coturnix japonica). Reported bird universal primers were used to amplify COI region from the extracted mtDNA samples using PCR. Amplicon were then sequenced by Sanger sequencing method (Sanger et al. 1977). Forward and reverse DNA Sequences were aligned with the reference sequence using nucleotide BLAST on NCBI to observe the dissimilarity among the sequences. Consensus sequences generated were used to construct their phylogenetic tree to see their evolutionary relationship with other bird species. Japanese quail which is thought to be domesticated from Japan, its Pakistani population showed close relation with sequences Summary 90 generated in Japan for this particular species. Pakistani wild quail species showed its closest linkage with C. coturnix. In conclusion, COI barcode proved as an authentic tool for species identification and phylogenetic inference of Pakistani wild and farm grown quails. Wild quail species has been characterized using partial COI gene sequences. This study has provided a specific genetic marker which can differentiate Japanese quail from wild quail at molecular level as most of the time both species are confused with each other. It can be helpful to the farmers and bird fanciers because they can select the birds of their choice correctly. This is the first study reporting DNA barcode of this Pakistani quail species. It would help researchers to study about phylogenetic and taxonomic status and also assist quail fanciers and quail farmers to unaffectedly identify their species of interest in farming. Identification of quail species is also important for conservation of biodiversity as it helps in preservation and identification of endangered species by generating their barcodes from even minimal evidence available. Availability: Items available for loan: UVAS Library [Call number: 2311-T] (1).

21. Mutation Analysis Of Alpha-Synuclein Gene In Patients With Parkinson Disease

by Iffat Aleem (2009-VA-566) | Dr. Asif Nadeem | Prof. Dr. Tahir Yaqub | Ms. Huma Mujahid.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: Parkinson disease is a complex, heterogeneous and chronic neurodegenerative disorder with a cumulative prevalence of greater than one per thousand, caused by neuronal loss, mainly affecting dopaminergic neurons of the substantia nigra. Parkinson disease is an idiopathic disorder of the extra pyramidal system characterized by tremors. Genetic factors contribute to its complex pathogenesis. A functional repeat polymorphism in the α-synuclein (SNCA) gene promoter conveys susceptibility for Parkinson disease. The α-synuclein (SNCA) has been implicated in rare autosomal dominant forms of Parkinson disease. The mutations in α-synuclein were associated with severe disease progression and a typical physical signs, indicative of neuro degeneration extending beyond the substantia nigra. Mutation in α-synuclein gene may have association with dopaminergic neuronal loss in Parkinson disease. Blood samples were collected from Parkinson disease patients. DNA was extracted by organic method. Primers were designed using Primer3 software. Amplification of gene was done by Polymerase Chain Reaction. PCR products were sequenced bi-directionally on ABI 3130XL Genetic analyzer. Sequence alignment was performed for polymorphism identification. The analysis of identified polymorphism has been done by CHROMAS software. Sequences were aligned by BLAST tool of NCBI. The results of analysis showed that no mutation found in exonic region of α-synuclein (SNCA) gene in Pakistani individuals selected for this study. Any change in exonic region of α-synuclein (SNCA) gene is a rare cause of sporadic and familial Parkinson disease in different populations. Availability: Items available for loan: UVAS Library [Call number: 2325-T] (1).

22. Identification Of Genetic Variations In Toll Like Receptor 1(Tlr-1) Gene To Evaluate Its Potential For Enhanced Resistance To Bovine Tuberculosis

by Shehar Bano (2013-VA-09) | Dr. Maryam Javed | Prof. Dr. Tahir Yaqub | Miss Huma Mujahid.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2015Dissertation note: Bovine tuberculosis is a disease caused by the species included in the Mycobacterium tuberculosis complex. Toll-like receptors (TLRs) are a family of conserved innate immune recognition receptors that trigger adaptive immune responses. TLR1 play an important role in host defense against mycobacteria, especially by mediating the response to mycobacterial triacylated lipopeptides. The objective of this study is the identification of single nucleotide polymorphisms within the coding region of TLR1 gene to evaluate its potential for enhanced the resistance to bovine tuberculosis in Nili-Ravi buffalo breed. Fifty blood samples of Nili-Ravi breed were collected from UVAS Pattoki Campus, Research Farm B and Buffalo Research Institute (BRI) Pattoki. Inorganic method was used for DNA extraction, for amplification of the coding region of TLR1 gene PCR (Polymerase Chain Reaction) was used using specially designed primers and the PCR products were sequenced through Sanger’s Chain Termination method. For the analysis and alignment of sequencing the results obtained after sequencing were analyzed and aligned using the CLUSTAL W and BLAST software. After all these analysis Ten SNPs were identified in the coding region of TLR1 mentioned in table. The Ten SNPs identified in the coding region of TOLL LIKE RECEPTOR 1 were in this order P1 C>T, P2 T>C, P3 T>C, P4 T>C, P5 T>C, P6 C >T, P7 T>C, P8 C >T, P9 A>G and P10 A>G. The one SNP found in the current research is in compliance with the (Sun et al. 2012) research on TOLL LIKE RECEPTOR 1 hence Nine SNPs found in the current research are novel in Nili Ravi buffalo. The SNPs in the exonic region that is P1 C>T, P2 T>C, P3 T>C, P4 T>C, P5 T>C, P6 C >T, P7 T>C, P8 C >T, P9 A>G and P10 A>G were all transitions i.e. the conversion of purines to purines. Population genetic analysis and allelic distribution at all loci was analyzed using POPGENE 32 software indicated that at [P3=0.243009> 0.05] followed the assumptions of the Hardy-Weinberg equilibrium indicating that the alleles were randomly distributed throughout the population, no migration had occurred, no bottlenecks happened and population remained large in numbers.This Non-significant and obeying HWE, so can be potential marker for genetic selection.At [P1= 0.040418< 0.05], [P2=0.033603< 0.05], [P4=0.000649< 0.05], [P5=0.000262< 0.05], [P7=0.015112< 0.05] and [P9=0.000111< 0.05] the probability value below 0.05 indicated that population at these polymorphic sites was not obeying Hardy-Weinberg equilibrium. This indicated that at these positions alleles were not equally distributed in population. It can be concluded from my research that the SNPs identified in the current research may also hold potential for marker-assisted breeding programs, with the aim of breeding more BTB-resistant animals and herds within both the national farms and the private sector. Availability: Items available for loan: UVAS Library [Call number: 2335-T] (1).

23. Pathological Investigations Of Different Isolates Of H9n2 Prevalent In Broiler Chicken

by M. Furqan Shahid (2014-VA-322) | Dr. Yasin Tipu | Prof. Dr. Asim Aslam | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2016Dissertation note: In recent years, H9N2 virus has attained a great importance as its infection has reached panzootic proportions. AIV H9 has different antigenic variants that has made it problematic to diagnose and thus to understand the pathogenicity of this virus is also very difficult. Detection of AI H9 antibodies can be used as a complementary method for sero-epidemiological studies as an indicator of AI H9 infection. The haemagglutination inhibition (HI) assay is used routinely for subtyping and detecting an increase of antibodies to AI viruses. Surveillance and early diagnosis of AI virus is essential for poultry. It demands rapid, sensitive and inexpensive diagnostic tests. Thus, it is important to identify different antigenic variants of H9. In this study a total of seven H9 virus samples were isolated out of total 100 collected sample from field outbreaks. These isolates were confirmed by molecular methods like PCR. Then four isolates from these seven isolates were used to infect the experimental broiler chicken. Clinical signs were recorded after the inoculation of H9N2 virus to the broiler. The results of this study showed that clinical signs were more sever upto 5 DPI. The severity of signs was proved by observing the gross pathology and histopathology of organs (Lung, Kidney, Trachea and Liver) of infected birds which were collected on 5 and 9 DPI. Serum of infected birds was also collected on 7 and 14 DPI to analyze the antibody level of infected birds against experimentally used isolate of H9N2. Then cross reactivity of different isolates of H9N2 was also checked against pannel of hyperimmune sera raised against different isolates of H9N2 and their results showed different antibody level against different isolates of H9N2. The sero-biochemical study of serum of infected birds revealed that H9N2 virus has pathogenic potential on kidney and liver. Availability: Items available for loan: UVAS Library [Call number: 2459-T] (1).

24. Studies on Pathogenesis and Molecular Diagnosis of Infectious Bursal Disease Virus In Broiler Chicken

by Beenish Zahid (2003-VA-134) | Prof. Dr. Asim Aslam | Dr. Yasin Tipu | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2015Dissertation note: Blank CD. Availability: Items available for loan: UVAS Library [Call number: 2510-T] (1).

25. Molecular Exploration Of Zbed6 Gene For Growth Trait In Lohi Sheep

by Usman Sagheer (2014-VA-03) | Dr. Maryam Javed | Dr. Akhtar Ali | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2016Dissertation note: ZBED6 gene is a central transcription factor. It is as a repressor of IGF2 (insulin-like growth factor II) interpretation in skeletal muscle myogenesis and development. It is essentially included in organism development, signaling, cell to cell collaboration, hepatic fibrosis, clathrin intervened endocytosis and tight intersection signaling falls. Chromatin immune precipitation (ChIP) sequencing utilizing C2C12 cells recognized around 2,500 ZBED6 binding locations in the genome, and the derived accord theme gave an immaculate match with the set up tying site in IGF2. Silencing of ZBED6 in myoblast cells influences IGF2 expression, wound healing, cell proliferation and myotube arrangement. Genes connected with ZBED6 binding sites demonstrated a very huge advancement for certain Gene Ontology groupings, including improvement and transcriptional regulation. Forty two blood samples were collected. DNA extraction was done by using organic extraction method. Primers for PCR amplification designed using Primer3 software. PCR products were sequenced and then analyzed by using BioEdit software. Expasy translational tool for translation and POPGENE 32 software for analysis of population genetics at all the loci were used. Using this software the overall allele frequency, heterozygosity, probability using Chi-square test and Likelihood ratio test and Hardy-Weinberg equilibrium, genotype distribution at all SNP position, summary of genetic variation statistics for all loci and association were calculated. After this, for the association one way ANOVA was performed. Single nucleotide polymorphism within ZBED6 could be potential candidate gene to be serving as genetic marker for the selection of animals with higher tendencies towards weight gain. Availability: Items available for loan: UVAS Library [Call number: 2539-T] (1).

26. Molecular Investigation Of Low Density Lipoprotein Receptor Gene Causing Familial Hypercholesterolemia And Its Evolutionary Relationship With Pan Troglodytes

by Rida Zainab (2014-VA-808) | Dr. Maryam Javed | Dr. Asif Nadeem | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2016Dissertation note: Familial Hypercholesterolemia (FH) phenotype is related to improper metabolism of low density lipoproteins due to mutations in Low-density lipoprotein receptor (LDLR) gene with increased risk of ischemic heart disease. Genetic variants in LDLR gene are associated with defective catabolism of cholesterol effecting lipid metabolism which results in familial hypercholesterolemia. It occurs in both forms: Homozygous Familial Hypercholesterolemia and Heterozygous Familial Hypercholesterolemia. Patients having high cholesterol were identified by observing the values of their serum lipid profile test reports. Their detailed history was taken and blood samples from the identified patients of familial hypercholesterolemia were collected. DNA extraction was done by Organic method. Primers were synthesized and PCR was conducted using optimized recipe and conditions. PCR products were sequenced. Sequenced data was analyzed using Chromas or BioEdit software. BLAST was performed and sequences were aligned individually by comparing it to the reference sequence. This showed difference in any specific position of a mutated sequence against the reference sequence. CLUSTALW aligned all the sequences together in one time. Sequences were compared with reference sequence to detect the presence of any mutation or SNPs. SNPs were identified manually and the peaks were observed in order to determine if the genotype is heterozygous or homozygous. Statistical Analysis was done and any amino acid change due to the observed SNPs was determined by using Expasy Translate Tool. It was found that both the SNPs showed amino acid changes. In the end, homology analysis was done which showed that Homo sapiens had their LDLR gene closest to that of Gorilla gorilla gorilla. Availability: Items available for loan: UVAS Library [Call number: 2551-T] (1).

27. Pathological Investigation And Molecular Detection Of Avian Pathogenic E.Coli Serogroups In Broiler Birds

by Muhammad Azeem Riaz (2008-VA-132) | Prof. Dr. Asim Aslam | Dr. Muti Ur Rehman Khan | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2016Dissertation note: The present study was designed to identify the serogroups present in field and to study their pathological effects in experimentally infected broiler chicks. The present study was attempted to scan the rfb gene clusters in APEC predominant serotypes O1, O2 and O78 strains and to develop Multiplex PCR method for serotyping of the O-antigens. The Multiplex PCR method was used for the identification of serotypes of APEC. The second part of the study was to study the pathological lesions caused by most prevalent serogroup in experimentally infected broiler chicks. A total of 100 tissue samples (lungs and livers) were collected from colibacillosis suspected broiler birds. Streaking was done from these samples on three different media and it was found that 80% isolates were positive on MacConkey media, 60% were positive on EMB media and 40% were found pathogenic for E.coli on Congo red media. The colonies which were of pink color on congo red media were considered as pathogenic. DNA was extracted from these colonies by boiling method by picking single colony from each petri plate. Extracted DNA was further used for PCR to confirm the three serogroups i.e O1, O2 and O78. The PCR results showed that 8% isolated samples were found as pathogenic as O2 strain was found dominant among all. Only two genomic DNA samples were found of O1 serogroup After confirmation of serogroups inoculum of Avian pathogenic E.coli O2 strain was prepared to experimentally infect the broiler birds. Birds were infected at the age of day 7 via intratracheal route. Following the experimental infection of birds, they were monitored for any pathological lesions which were not present significantly while some birds were off feed, reluctant to move, head down posture and were keeping themselves in isolation. Summary 46 Postmortem of dead birds was performed and pathological lesions were noted. Livers were found to be congested, enlarged and white fibrinous layer over liver was present. Lungs were also affected with the disease and white layer was present on lungs too. Lungs were consolidated and congested. Histopathology of lungs and livers was performed. It was noted that there was mononuclear cells infiltration and thin fibrinous layer over liver. Thickening of the liver capsule was noted due to infiltration of mononuclear cells and there was marked congestion in hepatic portal areas and the central vein. There was atrophy of adjoining hepatic cords due to greatly distended and congested sinusoids. Besides these changes, hepatocytes in various stages of degeneration along with hemorrhages, areas of congestion and fatty changes in a few places could be seen. There was infiltration of heterophils, severe congestion, lymphocytes and macrophages in the wall of the bronchus as well as in the peribronchial alveoli. There was marked presence of granuloma in lungs. Some birds displayed thickening of the pleura and consolidated areas covered with yellowish fibrin in lungs. The experimental infection of avian pathogenic E.coli confirmed the hypothesis that it causes pronounced histopathological lesions in broiler birds. Availability: Items available for loan: UVAS Library [Call number: 2591-T] (1).

28. Molecular Epidemiology Of Mycobacterium At The Animal Human Interface And Its Co-Morbidity With Diabetes Mellitus

by Zarfishan Tahir (2011-VA-624) | Prof. Dr. Mansur-ud-Din Ahmad | Dr. Abdul Majeed Akhtar | Dr. Muhammad Hassan Mushtaq | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2016Dissertation note: Tuberculosis (TB) is a common and fatal infectious disease which has afflicted mankind for several millennia. At the moment, TB is positioned at number five when it comes to the most common causes of fatality worldwide. TB is curable if it is properly diagnosed and treated. In 2015, it was estimated that 1.5 million deaths (an equivalent of 4,000 deaths per day) and 9 million new TB cases have been reported. Diabetes Mellitus is also widely distributed and estimated to affect 366 million people by 2030. The co-morbidity of DM and TB is re-emerging because of the progressive epidemiology of both diseases especially in the developing countries. Endemicity of TB and DM is growing in developing countries because of low socio-economic status and poor living conditions. In this study, a total of 500 tuberculosis positive patients were selected under TB DOTS program from five tertiary care hospitals of Lahore. Sputum samples were collected from all the enrolled patients and smear microscopy was performed for TB confirmation. Blood samples were collected from the same patients for screening of diabetes mellitus. Sputum samples were also processed for culture and drug sensitivity on LJ medium. Molecular identification by PCR technique was carried out on all positive cultured strains and results were compared with reference strain H37RV. For DNA sequencing, PCR products were sent to Singapore where sequencing was performed by Sanger method. Data was compiled and variables including gender, age, drug resistance and treatment history and correlation among different variables was analyzed using chi-square test and Fischer’s exact test method at P-value of ≤0.05. SPSS (Statistical Package for Social Sciences, Version 20.0) was used for statistical analysis. The count data was statistically analyzed using SUMMARY 124 descriptive statistical tools. On screening for fasting blood sugar level, 74 (14.8%) patients were recorded as diabetics as well i.e. blood sugar level ≥ 126 mg/dl. Out of these 74 patients, 22 patients had previous history of diabetes whereas remaining 52 patients were newly diagnosed at the time of screening. The maximum distribution of TB-DM patients was found in age group > 57 years. Mean age of the group without DM was 39 years and with DM was 48 years. Coexistence of DM in TB patients was higher in males (62.2%) as compared to female study subjects. However, the gender difference is statistically non-significant (p value 0.243). The distribution of education level revealed that out of the total participants, maximum number of patients (n=220) were illiterate and similar trend was observed in diabetic patients with 54 (73%) individuals belonging to the illiterate group of the subjects. There is statistically significant difference between existence of DM and literacy level in tuberculosis patients. Among social and behavioral risk factors in tuberculosis patients, majority of the patients were unemployed (24%) in TB-DM group. Significant correlation p value ≤ 0.05 was found between coexistence of TB-DM and tobacco use. TB cases with diabetes were known to have history of smoking with 73% (n=54) while non-smokers were 27% (n=20). On sputum smear microscopy frequency of 3+ results showing high bacterial load, was profoundly higher i.e. 67.6% in diabetic tuberculosis patients as compared to non-diabetics which was 4.9% only. Total culture yield was 363 out of 500 sputum samples. There were 193 samples that were sensitive to all drugs, 9.4% were MDR strains (resistant to Isoniazid and Rifampicin). MDR-TB is significantly higher in TB-DM patients i.e. 13.5% as compared to 8.7% in TB only patients. In our study, DNA sequence data for drug resistance was studied by the sequence of rpoB gene of the wild type MTB strain. Sequencing results showed mutations at various spots of rpoB gene. SUMMARY 125 Most common mutational sites identified were at codon 531, 526 and 516 with frequency of 70%, 15% and 7.5%, respectively. Moreover, mutation sites at 512 and 574 codon had also been reported. In this study, predominantly two phylogenetic variants were identified. Majority of the isolated strains were Central Asia Strain (CAS) with a prevalence of 88.2% and rest were Beijing strain. However, attempts to find zoonosis could not be established. A total of 900 raw milk samples were also screened for M. bovis and no positive sample could be detected. The present study emphasizes the importance of screening for DM in TB patients, which had not been done in routine. This practice may prove to be helpful in reducing the disease burden of TB patients as well as DM patients. Thus it is recommended that the screening for DM should be implemented in TB/DOTS clinics. Emergence of Multi drug resistant Mycobacterium tuberculosis is also a serious challenge for clinicians. A very large financial implication in terms of treatment, duration of chemotherapy and spread of MDR TB strains is being faced. Treating MDR TB is more complicated than treating drug sensitive TB. Patients with MDR TB require longer, much more costly treatment and experience higher mortality rates. Such a long time to initiate the treatment is not affordable, thus there is a dire need for some rapid technique like molecular based diagnostics for MDR detection, which can provide quick results and making it possible to start treatment at earlier to minimize transmission, morbidity and mortality. Availability: Items available for loan: UVAS Library [Call number: 2710-T] (1).

29. Comparative Genomic Study of Motor Neuron Disease in Horses and Human

by Shakeela Daud (2011-VA-534) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Dr. Abu Saeed Hashmi.

Material type: book Book; Literary form: not fiction Publisher: 2017Dissertation note: cd not submitted Availability: Items available for loan: UVAS Library [Call number: 2810-T] (1).

30. Study Of Tyrosine Hydroxylase (Th) Gene Sequence Variations In Association With Δ9-Tetrahydrocannabinol (Thc) Dependence

by Ali Raza (2015-VA-446) | Dr. Maryam Javed | Dr. Asif Nadeem | Prof. Dr. Tahir Yaqub.

Material type: book Book; Literary form: not fiction Publisher: 2017Dissertation note: Anti-Social Personality Disorder (ASPD) is ability of an individual to adopt social norms. These ASPDs are driving force in majority of criminal activities. Dopamine being important neurotranmiter of nervous system controls major behavioral traits. Low level of dopamine can be causative factor for an individual to start drug abuse to restore it because majority of drug are proven to enhance dopamine production. In this study genetic exploration of genes coding for dopamine producing enzymes. Tyrosine Hydroxylase (TH) gene was selected and its two regions (Intron 1 and exon 3) were amplified and analyzed through Sanger’s sequencing method followed by statistical analysis. Total five SNP were recorded at locus TH1, TH2, TH3, TH4, and TH5. One insertion, three transversion and only one mutation was transition. No exonic mutation was recorded hence no change in protein structure was found. Mutations at TH1 and TH4 were found to be highly associated with addiction. Mutant “B” allele were also present but still wild “A” was most common allele in our population. TH1, TH2, TH4 have positive correlation with addiction, TH3 is correlated with Nicotine and TH5 shown protective role against nicotine. There are few genetic changes in our population that can be associated with drug addiction statistically but still there prevalence in our gene pool is very low. We can conclude on the basis of these findings that drug addiction in our population is more likely a social issue rather than genetical. Limitations of our research was sample size. There are further possibilities for this project to investigate on mRNA level. Advanced neurobiological techniques can also be applied on subjects to analyze dopamine level in different brain regions. For genetic studies epistatic role of few other genes can also be considered for validation. Availability: Items available for loan: UVAS Library [Call number: 2858-T] (1).

31. Genetic Evolution And Development Of Recombinant Vaccine Against Newcastle Disease For Chicken In Pakistan

by Abdul Wajid (2009-VA-705) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Dr. Muhammad Tayyab.

Material type: book Book; Format: print ; Literary form: not fiction Publisher: 2017Dissertation note: Newcastle disease (ND) is one of the most contagious diseases of poultry worldwide. The disease is endemic in Pakistan and recurrent outbreaks have been reported in commercial poultry flocks, domestic pet and migratory birds since 1963 an inception of commercial poultry farming in the country. Disease surveillance is necessary to determine the incidence of the disease as well as to identify the etiological agent of the disease status in the region. The analysis of the field data provides a clue for the higher authorities to take steps for the remedy of the devastating outbreak. A virulent form of Newcastle disease virus caused an outbreak in the northern region of Pakistan during the mid of 2011. The virus was identified as a virulent viscerotropic vvNDV and characterize, belonging to the sub genotype VIIi. However, the virus of this genotype is still circulating in the field though the intensity of the strain to succumb the chickens to cause mortality does not exist. The particular thing in this genotype was its susceptibility to other avian species like pheasants, peafowls, ducks turkeys, peacocks, sparrows and parakeets. As this genotype is circulating since 2011 2016 and still spill over in these avian species. Thus for the last five years (2011-16), 3500 healthy, diseased and dead chickens, pheasants, peacocks, turkeys, peafowls, ducks, sparrows, exotic parakeets, rosy-faced parrots, pigeons, and partridges from 750 different locations s were monitored. Samples were collected from the Northern region of the country Punjab, Khyber Pakhtoonkhawa, Azad Kashmir, including Gilgit,Baltitssan and from Southern region, Karachi, Hyderabad , Mirpursakro and other small cities where the poultry farms are located. The samples were collected by the local veterinarians, poultry Assistants and Animal health practitioners who assist during the surveillance program. Samples were also collected from the farmers who brought their birds for inspection in the lab with the details of the 141 farm. Mostly sampling was done where there was reports of NDV outbreak, tissues were collected usually the trachea, spleen and brain, moreover, the pharyngeal and cloacal swabs not only from the infected birds but also from the healthy birds were collected to assess the virus shedding in the flock. Blood samples were also collected (1% of the birds at farm), for serum collection to assess the immune status of the flock using Haemagglutination Inhibition (HI) test and Enzyme linked immunosorbant assay (ELISA). The Survey Form meet the international standard was filled for each farm for recording the information required to find the diagnostic clue as well as the molecular characterization of the isolates. Pool of five pharyngeal swabs were processed after the passage into 9-day old chicken embryonated eggs and confirming the positive HA test and then confirmed by real time PCR (RT-PCR). In addition, sera were tested against NDV by HI and ELISA tests. The targeted samples were sequenced by complete fusion gene and whole genome using 22 pairs of overlapping primers. The observations indicated that the commercial broiler industry is highly susceptible to virulent NDV and confirmed by data available in the laboratory in the survey form. Contrary to that a little is known regarding the maintenance and enzootic trends of vNDV infection level in domestic and wild birds. Poor strategy of the use of vaccines and vaccination as well as the existence of virulent form of NDV in the domestic and pet birds indicate a possibility of the root cause of the ND eruption in the developing countries. A continuous isolation of virulent viruses of the panzootic Newcastle disease virus of sub-genotype VIIi since (2011-2016 from commercial chickens and from various other avian species in the country provide evidence for the existence of epidemiological links intermingling of the strain among them. Therefore, to avoid the huge economical losses in the commercial poultry the second largest industry in Pakistan, their close proximity should be strictly avoided. The mass vaccination of the poultry flocks is in practice in all commercial 142 poultry farms in Pakistan. However, the use and availability of a reliable and standard vaccine, as well as the correct usage of vaccine dose of the live attenuated LaSota vaccine are the key factors to improve their efficacy in the field. Minor outbreaks have been occurring in the field even though a severe outbreak was occurred in 2011-12 collapsed the poultry industry with other pet and wild birds. To minimize the continuity of these minor outbreaks in the field for long time there is a need for more effective vaccine to control the particular genotype of the ND virus. In the present study, DNA vaccine was developed using the SFR-55 NDV strain antigens, in the form of fusion (F) and hemagglutinin-neuroaminidase (HN), namely pcDNA3.1-F and pcDNA3.1-HN. In vitro expression of both genes construct was assessed by reversetranscriptase- PCR (RT-PCR) and western blotting. In the trial an inactivated oil-based emulsion vaccine was prepared using the field strain SFR-55 and compare with the commercial vaccine LaSota strain commonly used by the poultry industry. Birds were divided into six groups, the first two groups were immunized with pcDNA3.1-F and pcDNA3.1-HN alone respectively and third group with was vaccinated with both antigens pcDNA3.1-F+HN. The other two groups were immunized with inactivated (wvSFR-55) and LaSota vaccines as described above, the last group was injected with empty vector as control. The birds were immunized twice at 14 and 21 days of age intramuscularly (DNA vaccine), subcutaneous and eye-drop by inactivated and LaSota vaccines respectively. The birds were challenged with live virulent NDV strain using a dose of 10,000 ELD50/0.1ml per chicken. Results indicate that Inactivated and LaSota vaccines provided high protection (>80%), as compared to pcDNA3.1-F, pcDNA3.1-HN, pcDNA3.1- F+HN gave 70%, 75% and 20% respectively. There was 100% mortality in control chickens. The administration of two vectors expressing F and HN antigens induced high immune response, and provide protection than when used separately. However, the groups immunized with 143 pcDNA3.1-F, pcDNA3.1-F+HN and inactivated vaccine resulted in lower amount of virulent virus shed after challenge when compared to the group immunized with standard LaSota. In summary, the co-administration of both NDV glycoprotein antigens increased protection than use separately. DNA-based vaccine can be used safely to reduce mortality and most importantly lower the risk of virus transmission due to decreased level of virulent virus shedding. Availability: Items available for loan: UVAS Library [Call number: 2910-T] (1).

32. Analysis Of Tp53 Gene Isolated From Oral Cancer Patients

by Amir Saleem (2013-VA-897) | Dr. Muhammad Wasim | Prof. Dr. Tahir Yaqub | Prof. Dr. Habib-ur-Rehman.

Material type: book Book; Literary form: not fiction Publisher: 2017Dissertation note: Cancer is a term used for disease in which abnormal cells divide without control and are able to invade other tissues. Cancer cells can spread to other parts of the body through blood and lymph system. TP53 is one of the most important tumor suppressor genes, mutated in more than 50% of human malignancies. It controls DNA repair, cell cycle and apoptosis and therefore plays an essential role in keeping genetic constancy. TP53 gene is present on the short arm of chromosome number 17. In human it extends 19,200 bp in 11 exons. Various parameters are used in the present study was aimed to investigate coding regions of TP53 gene for analyzing the mutations involved in oral cancer. Human OSCC samples (15) and normal tissue samples (15) were collected from Fatima Memorial Hospital Lahore. Samples were collected in tubes by oncologist containing ethanol and then brought to Molecular Biology and Biotechnology Lab of Institute of Biochemistry and Biotechnology, UVAS Lahore and frozen at -20°C before DNA extraction. Samples have been processed for research purpose. DNA was extracted from tissue by using Trizol Method and quantity was checked by nanodrop spectrophotometer. Two Primer sets were designed to amplify protein coding region of TP53 gene. After amplification through PCR, DNA Sequencing was done. Data interpretation was done by using several softwares like BLAST alignment tool, Bioedit, Clustal W2. In this study it was tried to find out the mutations in TP53 but no any kind of mutations were identified. Because I conducted my research only on 15 samples. So in future to use this gene as a potentional biomarker we can increase our number of samples. 46 The need for today is to develop valid biomarkers, which can be incorporated in ongoing in vivo and in vitro clinical mechanistic and improve the diagnosis and prognosis of this dreadful disease. In the conclusion, we must say that further study, with much larger sample number, is required to revise the effects of this polymorphism and accurately identifying the associated factors. There is a need to explore the other gene mutations causing oral cancer in local population of Punjab and Pakistan that will ultimately help to develop genetic counseling strategies, gene therapies and prenatal diagnostic procedures for the population of Pakistan. Availability: Items available for loan: UVAS Library [Call number: 2937-T] (1).

33. Bacterial Profiling And Development Of Molecular Diagnostic Assays For Detection Of Bacterial Pathogens Associated With Bovine Mastitis

by Aqeela Ashraf (2012-VA-388) | Dr. Muhammad Imran | Prof. Dr. Tahir Yaqub | Dr. Muhammad Tayyab.

Material type: book Book; Literary form: not fiction Publisher: 2017Dissertation note: The livestock sector plays a critical role in strengthening the economy of Pakistan. Control of livestock diseases is the primary objective of government livestock departments. Bovine mastitis is among the most significant diseases of livestock as reported by various field surveys in Pakistan. Despite considerable knowledge about mastitis and its etiology, this disease is still prevalent in many dairy herds; it remains most difficult to eradicate or control. It is an inflammation of mammary gland resulting in decreased milk production, veterinary care costs and culling losses. In animal health improvement, there is a paradigm shift from treatment of clinical illness to disease prevention. Recognition of disease is the foundation of disease control and prevention. California mastitis test and somatic cell counting are the most commonly used methods for diagnosis of bovine mastitis. These methods are unable to identify the causative agent. Detection of pathogen is critically important for better control of mastitis. Microbial culturing and biochemical tests are traditionally used methods for pathogen identification. But, these methods are very time consuming and can only detect viable bacteria from the sample and can lead to false negative results. The progress in molecular methods based on PCR has improved the veterinary diagnostics. For the identification of bovine mastitis pathogens, an economical, rapid and sensitive molecular diagnostic assay was developed using multiplex PCR, detecting the pathogenic species-specific DNA. The target species areS. aureus,E. coli, S. uberis, S. agalactiae, S. dysagalactia, S. haemolyticus, S. epidermidis, S. chromogenes andM. bovis. Multiplex PCR assay was developed for the detection of these significantly important bacterial pathogen causing bovine mastitis. Species specific primers were designed which have the ability to specifically amplify the particular gene in the target species. For this purpose various gene regions were selected for different bacterial species which included 16S rRNA, cpn60, phoA and rdr. Initially monoplex PCRs were optimized individually for each target species. For optimizing multiplex PCR assay, various combinations of individual PCRs with varying concentrations of primers and template DNA were used. The final protocol included all the nine sets of primer pairs, every set targeting a unique mastitic pathogen. Multiplex PCR assay was checked for its specificity and analytic sensitivity was calculated. Mastitic milk samples were collected aseptically from various farms. Initial screening was based on Surf field mastitis test and California mastitis test. Milk samples were cultured on nutrient agar, blood sheep agar and MacConkey’s agar. The bacterial isolates were identified and further sub-cultured in nutrient broth. All the isolates were identified on the basis of 16S rRNA sequencing analysis. The developed multiplex PCR assay was used to detect the target bacterial pathogens from the collected milk samples. Limit of detection of developed assay was up to 50 pg for DNA isolated from pure cultures and 104 CFU/ml for spiked milk samples. The results obtained by 16S rRNA sequencing, bacterial culture based identification and multiplex PCR assay were compared. Sensitivity and specificity were calculated using latent class analysis, specificity was up to 88% and sensitivity was up to 98% for targeted mastitic pathogens. The developed multiplex PCR detected nine bacterial species in a single reaction. Multiplex PCR assay has also detected the bacterial pathogens in a few culture-negative mastitis milk samples. This is the first multiplex PCR assay which can efficiently detect nine important mastitic bacterial pathogens in a single reaction. The development of multiplex PCR assay is useful in early diagnosis and prevention & control of bovine mastitis. Mycoplasma is often ignored as a major mastitis-causing pathogen due to lack of rapid and accurate diagnostic tools. In this study a LAMP assay was developed for the identification of M. bovis from clinical mastitic milk samples. LAMP primers were designed from three gene regions including uvrC, 16S rRNA and gyrB. Bacterial reference strains and mastitic milk samples positive for M. bovis were collected from Quality Milk Production Services, Cornell University, Ithaca, NY. Bacterial strains were further cultured on Hayflick medium containing 15% horse serum and incubated for up to 7 d at 37°C with CO2 enrichment. DNA was isolated from mastitic milk samples and bacterial culture using Qiagen DNeasy Blood and Tissue Kit (Life Technologies, Carlsbad, CA) according to the manufacturer’s instructions with few modifications. PCR and LAMP assay was performed for all the samples obtained. Analytic sensitivity was calculated and the limit of detection was up to 50pg/reaction for LAMP assay which is higher as compared to PCR. Sensitivity and specificity was calculated for each of the three tests. Cohen’s kappa values obtained were 0.940 for uvrC, 0.970 for gyrB and 0.807 for 16S rRNA. All three tests showed a high level of agreement between test results and the true mastitis status, indicated by the receiver operating characteristic (ROC) curve. A robust, sensitive and specific LAMP assay has been developed for the detection of M. bovis from mastitic milk. These novel molecular assays could be helpful for correct and timely identification of bovine mastitic pathogens, which is crucial for the control and treatment of the disease.Molecular diagnostic assayshave been developed in the current study based on multiplex PCR assay and loop-mediated isothermal amplification assay. Availability: Items available for loan: UVAS Library [Call number: 2930-T] (1).

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