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Lala Lajpat Rai University of Veterinary & Animal Sciences, Hisar
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ThesisItem Open Access Effect of pellet diets with or without phytogenic feed additives on performance of broilers(LUVAS Hisar, 2023-04) Devika; Sihag,Zile SinghA trial of 42 days was carried out to study the effect of pellet diets with or without phytogenic feed additives on performance of broilers. A total of 240, day-old commercial broiler chicks were randomly distributed into eight dietary treatments with three replicates per treatment and each replicate had ten birds. Ashwagandha and shatavari were used as herbal feed additives. The basal diet was formulated as per BIS (2007) specifications. The treatments included T1: Basal diet + antibiotics (mash form), T2: Basal diet + antibiotics (pellet form), T3: Basal diet + 1% Ashwagandha powder (mash form), T4: Basal diet + 0.5% Ashwagandha powder (pellet form), T5: Basal diet + 1% Ashwagandha powder (pellet form), T6: Basal diet + 1% Shatavari powder (mash form), T7: Basal diet + 0.5% Shatavari powder (pellet form) and T8: Basal diet + 1% Shatavari powder (pellet form). Body weight (g/bird), Feed intake (g/bird) and FCR improved significantly (P<0.05) after supplementing the diet with phytogenic feed additives and the best results were observed in 1% Ashwagandha supplemented group in pellet form. In between the control groups, the pelleted diet fed group had better growth performance than mash diet fed group. Dietary supplementation of 1% Ashwagandha in pellet form resulted in significantly (P<0.05) higher DM metabolizability and gross energy metabolizability as compared to control. Nitrogen retention was found significantly (P<0.05) higher in groups T3, T5, T6 and T8. The carcass traits were found to be significantly (P<0.05) better in 1% Ashwagandha supplemented group in pellet form. The Hb levels and TLC count were found to be significantly (P<0.05) higher in all Shatavari supplemented groups while decreased % of heterophils and increased % of lymphocytes thereby, reducing heterophil: lymphocyte ratio in T6, T7 and T8 groups compared to control. No significant effect of pelleting was observed on haematological parameters. Significant reduction in serum cholesterol and triglycerides levels was observed in 1% Ashwagandha and 1% Shatavari supplemented groups. Inclusion of 0.5% and 1% Ashwagandha and Shatavari powder resulted in significant improvement in HDL and reduction in LDL as compared with control. But no effect of pelleting was observed on serum parameters. A significant reduction in E.colicount was recorded in the T3 and T5 groups compared to control. Higher values of Lactobacilli were observed in all herbal additives supplemented groups. Highest villi height andVillus height: Villi depth of intestinal crypts was recorded in 1% Ashwagandha supplemented group in pellet form. The activity of antioxidant enzymes; SOD and Catalase was observed in groups T3, T5, T6 and T8. Thus, the dietary supplementation of Ashwagandha root powder and Shatavari root powder leads to significant improvement in the growth performance, nutrient metabolizability and hematological indices; possess potent hypocholestrolemic, and gut microbiota stabilizing effects, thus help in achieving a profitable and feasible poultry production. The feeding of pelleted ration also improved the growth performance of birds and this will help in establishing profitable poultry farming.ThesisItem Open Access Comparative evaluation of interlocking nailing alone and in combination with Beta–Tricalcium Phosphate (β-TCP) for management of long bone fracture in dogs(LUVAS Hisar, 2023-04) Manikant; Sandeep SaharanThe objective of this research was to determine whether beta-tricalcium phosphate (β-TCP) might be used to enhance fracture healing following interlocking nail stabilisation. Twelve dogs with long bone fractures, regardless of age, breed, or sex, were divided into two groups at random. Six randomly chosen animals from group I underwent interlocking nailing, whereas six animals from group II underwent interlocking nailing as well as the application of β-TCP to the fracture site. On the day of presentation, a detailed clinical history was taken, and radiographic, haematological, biochemical, and clinical examinations were performed before surgery as well as on the 15th, 30th, and 60th postoperative days. At different intervals, group II performed better than group I in terms of weight bearing when walking and standing. The higher bone union and bone formation scores (BFS and BUS, respectively) were seen in group II compared to group I. In group II, the stage of bone union and functional limb usage was achieved earlier as compare to group I. The haemato-biochemical and physiological parameters did not differ significantly between the groups. Also, the complications identified in this analysis included suture dehiscence, delayed healing, screw loosening, and suture tract infection. According to the findings of the current study, interlocking nailing combined with the application of βTCP was found to be more effective than interlocking nailing alone in the management of long bone fracture in dogsThesisItem Open Access Effect of inclusion of Moringa leaf meal and pelleting of the ration on the performance of broilers(LUVAS Hisar, 2023-04-10) Shabnam; Tewatia,B.S.An experiment was conducted to study the effects of pelleting and Moringa oleifera leaf meal supplementation on the performance of broiler chicken. One hundred sixty, one day old chicks, were randomly divided into 4 treatment groups having 4 replicates each, and each replicate had 10 birds. The control group (T1) was offered basal diet with antibiotics in mash form; T2 group was fed basal diet with antibiotics in pellet form; while birds in T3 group were offered ration having 5% Moringa oleifera leaf meal (MOLM) without antibiotics in mash form and T4 group ration had 5% Moringa oleifera leaf meal without antibiotics in pellet form. During the feeding trial of 42 days, feed intake and body weight changes were recorded. At the end of feeding trial, a metabolism trial of five days duration was conducted during which samples of feed offered, residue left and faeces voided were collected. For carcass evaluations, one bird from each dietary treatment was sacrificed. The average feed intake (g/bird) during overall period (0 to 42 day) did not differ significantly among groups. Average body weight gain (g/bird) in all the three growth phases differed significantly (P<0.05) and maximum body weight gain (2281.32) was recorded in group (T4) supplemented with 5% Moringa oleifera leaf meal in pellet form. Similar trend was observed in respect of feed conversion ratio, carcass traits, DM metabolizability, nitrogen metabolizability and gross energy metabolizability. Quality of the breast and thigh muscles in terms of crude protein (21.97, 17.11) and ether extract (5.52, 6.76) also improved significantly (P<0.05) in T4 group. Hematological parameters (Hb, TEC and lymphocyte) increased significantly (P<0.05), while TLC and heterophils decreased (P<0.05) in T4, T3 and T2 as compared to the control group. A significant (P<0.05) reduction in serum total cholesterol, LDL and triglycerides levels and increase in HDL was also observed in Moringa supplemented groups (T4 and T3) as compared to the antibiotic supplemented groups (T1 and T2). MOLM supplemented groups (T4 and T3) resulted in significant (P<0.05) decrease in E.coli count and increase in Lactobacilli count as compared to T1 and T2 groups. Antioxidant enzymes (SOD and Catalase) activity was also significantly higher (P<0.05) in MOLM supplemented group in pelleted form (T4) followed by T3, T2 and T1 group. Villus height: intestinal crypts depth of the experimental birds ranged from 4.47µm (T1) to 6.30 µm (T4) and significantly (P<0.05) higher effect was observes in the groups supplemented with MOLM. Thus, it can be inferred from the results of this study that inclusion of 5% MOLM in pellet form had better effect on body weight gain, FCR, carcass traits and antioxidant enzyme activity besides being better in terms of feeding cost per kg gain in broilers.ThesisItem Open Access Effect of feeding pelleted ration with or without herbal feed additives on the performance of broiler chickens(LUVAS Hisar, 2023-04) Radha Bai; R.S. BerwalThis research was conducted to investigate the effect of feeding pelleted ration with or without herbal feed additives on the growth performance, nutrient utilization, carcass traits, hematological, serological parameters and to economise the feeding of broiler chickens. Study was conducted on two hundred and forty, day-old broiler chicks which were randomly divided into eight dietary treatment groups with three replicates of each treatment. Each treatment had different feeding regimen: T1 received a basal diet with antibiotics in mash form; T2: basal diet with antibiotics in pellet form; T3: basal diet with 1% aloe vera in mash, T4: basal diet with 0.5% aloe vera in pellet form, T5: basal diet with 1% aloe vera in pellet form, T6: basal diet with 1% amla in mash form, T7: basal diet with 0.5% amla in pellet form and T8: basal diet with 1% amla in pellet form. In the 42-day feeding trial, Feed intake and body weight were recorded at every week and residual feed and faecal samples were collected and analyzed during a five-day metabolic trial during the end of growth trial. The results showed that the average overall body weight differ significantly (p<0.05) in amla and aloe vera supplemented groups in comparison to control groups (T1 and T2 ), with the highest average body weight was recorded in the group supplemented with 1% amla in pellet form, followed by 1% amla in mash form. Better feed conversion ratio was observed in 1% aloe- vera supplemented group in pellet form (T5 ) followed by T8 , T4 , and T3 . Significantly (p<0.05) higher nutrient metabolizability was observed in 1% aloe vera supplemented group followed by 1% amla supplemented groups. Significantly (p<0.05) higher percentage of carcass traits were observed in 1% amla supplemented groups in pellet and mash form as compared to the control groups. Hematological parameters (Hb, TEC, TLC and lymphocyte) were significantly (p<0.05) improved, while heterophils were significantly (p<0.05) decreased in the 1% amla supplemented groups compared to the control group. The herbal feed supplemented groups also showed a significant reduction in serum cholesterol, LDL, and triglyceride levels, as well as an increase in HDL levels, compared to the antibiotic-added groups. But better results were obtained in 1% aloe vera supplemented groups. The groups supplemented with 1% aloe vera in pellet and mash form had a significant decrease in E. coli count and an increase in Lactobacilli count compared to the T1 and T2 groups. The antioxidant activity of SOD and catalase enzyme were significantly (p<0.05) increased in all herbal feed supplemented groups with the highest improvement observed in T8 followed by T6 , T5 , and T3 . The villus height and intestinal crypts depth of the experimental birds ranged from 898.41µm (T1 ) to 1035.40 µm (T5 ) and 3.15 µm (T1 ) to 4.24 µm (T5 ) under different dietary treatments, and herbal feed supplemented group had significantly better effect on gut morphometry higher villus height was observed in the groups with inclusion of 1% aloe-vera in pellet as well in mash form. Better economics was found in 1% amla supplemented group in pellet form as compared to that of other herbal feed supplemented and control groups. In conclusion, the study suggested that inclusion of amla and aloe-vera supplementation at 1% level improved the growth performance, nutrient digestibility, carcass traits, hematological and serological parameters of broiler chickens.ThesisItem Open Access Studies on the ameliorating effect of Eugenol (clove essential oil) on Aflatoxicosis in broiler chicken(LUVAS Hisar, 2023-07) Nehra,Neetu; DeepikaAn experimental study was conducted on 132 broiler chicks to assess the hepatoprotective and antioxidative potential of eugenol (clove essential oil) in broiler chicken and to evaluate the ameliorative effect of eugenol supplementation on haemato-biochemical, oxidative stress, pathological changes and immune response in aflatoxin administered broiler chicken. Aspergillus parasiticus MTCC 411 spores were revived on potato dextrose agar and Cezpak yeast extract agar and used for aflatoxin production in maize and final experimental feed was estimated to contain 184 ppb of aflatoxin. One hundred and thirty-two (132) broiler chicks at an age of 7 days were randomly divided into 2 groups viz, Group A and Group B having 66 chicks in each group. All chicks were vaccinated against NCDV as per standard schedule. Group A chicks were supplemented with eugenol (@ 5ml/kg) of feed throughout the experiment. Group B chicks were kept as control and fed with basal control diet (without eugenol). At the age of 14 days, the chicks of groups A and B were further divided into two subgroups {group A into A1(AF+E) and A2(E); group B into B1(AF) and B2(C)} of 33 chicks each. All the chicks of group A1 and B1 were given aflatoxin (AF) @184ppb mixed with feed till the end of experiment and chicks of group B2 were fed on basal feed and act as control i.e., without any treatment. Blood samples were collected from six chicks of each subgroup directly from heart on 0, 7th, 14th 21st and 28th day post aflatoxin administration (DPAA) for assessing different parameters like haematological and oxidative stress parameters in blood while biochemical parameters and humoral immune response in serum. After collection of blood, chicks were sacrificed and detailed postmortem examination was conducted. Representative tissues of the different organs were preserved with 10 per cent neutral buffered formalin for histopathological examination. The aflatoxin administered chicks (group B1 and A1) exhibited varying degree of clinical signs such as dullness, ruffled feathers and reduced feed intake at different time intervals. Mean body weight was significantly decreased in groups B1 and A1 as compared to control group (B2) at different time intervals. Haematological studies showed significant decrease in mean Hb, PCV and TEC and non-significant decrease in MCV and MCHC in groups B1 and A1 at different time intervals. Mean TLC was also found significantly decreased in group B1 and A1. Biochemical studies showed significant decrease in mean total serum protein and albumin concentration and significant increase in liver function parameters like serum AST, ALT, GGT in aflatoxin fed groups (B1 and A1) at different time interval as compared to control group B2. Kidney function parameters like uric acid and creatinine concentration were also significantly increased in group B1 and A1. Blood catalase and SOD activities were significantly decreased and blood LPO (nMDA) was significantly increased in toxin fed groups as compared to control group at different time intervals. Eugenol supplementation significantly improved the body weight and ameliorated the adverse effect of aflatoxin on hemato-biochemical parameters and oxidative stress parameters. Immunological studies revealed significant decrease in humoral immune response in aflatoxin administered groups as observed by reduced mean HI titre (log2) against NDV in Group B1 and A1. Delayed type hypersensitivity study to assess CMI also revealed significant decreased skin thickness 24 hour after DNCB application in aflatoxin administered group B1(AF) as compared to control groups B2(C) and A2(E) while slight reduction with no significant difference was observed in group A1(AF+E). Pathomorphological lesions characteristic of aflatoxicosis were prominent in liver, kidneys and lymphoid organs and eugenol supplementation reduced the severity of lesions. In case of histopathology the lesions were vacuolar, vascular, fatty, degenerative, necrotic, infiltrative and proliferative in aflatoxin intoxicated group in different organs and were of mild nature in eugenol supplemented group A1(AF+E). Results of the present study indicated the hepatoprotective and antioxidative potential of eugenol (@ 0.5% in feed) along with its ameliorative effects against aflatoxin induced pathological alterations in broiler chickenThesisItem Open Access Patho-biological and Immunological studies on Monocrotophos intoxicated broiler chicken fed on Curcumin supplemented feed(LUVAS Hisar, 2023-07) Chahal,Ashmita; Nehra,VikasThe present experimental study was undertaken in broiler chicken to study the patho-biological changes produced due to Monocrotophos toxicity and to elucidate the ameliorating effect of curcumin supplementation on clinical signs, mortality, body weight, biochemical changes, oxidative stress parameters, acetyl choline esterase activity, pathological lesions and immune responses in relation to the toxicity of monocrotophos. 132 broiler chicks at the age of 7 days were divided randomly into two groups (group A and B) of 66 chicks in each group. All the chicks of group A were given feed supplemented with curcumin (@ 200 mg/kg of feed) throughout the experiment. All the chicks of group B were given normal feed without any supplementation. At the age of 14 days, the chicks of groups A and B were divided into two subgroups (group A into A1 and A2, group B into B1 and B2) of 33 chicks each. All the chicks of group A1 and B1 were given monocrotophos @ 5 ppm [as used earlier by Babulal (2007) in his experiments] per kg of feed up to the end of experiment. The chicks of group B2 served as normal control group i.e. without any curcumin and monocrotophos. Blood samples were collected from six chicks of each subgroup directly from heart on 0th, 7th, 14th, 21st and 28th day post initiation of monocrotophos feeding in sterile ethylene diamine tetra acetate (EDTA) coated vials for assessment of haematological and oxidative stress parameters and in sterile tubes for serum separation. Serum samples were stored in aliquots at -20oC until its use for assessing different biochemical parameters and humoral immune response. After collection of blood, these chicks were sacrificed and they were subjected to thorough post-mortem examination. Representative tissues of the different organs such as liver, heart, lungs, brain, kidneys, intestine, spleen, thymus and bursa of fabricius were collected in 10 % buffered formalin for histopathological studies. Brain tissue samples were collected from each bird after sacrifice and stored at -20oC for estimation of acetylcholine esterase activity. The chicks of both the monocrotophos administered groups (A1 and B1) exhibited varying degree of clinical signs such as dullness, ruffled feathers, torticollis, head held down near the chest and reduced feed intake at different time intervals. Mean body weight was found significantly decreased in groups A1 and B1 as compared control group B2 at different time intervals. Haematological studies showed significant decrease in mean Hb and TEC and significant increase in PCV in groups A1 and B1 as compared to control group B2. MCV and MCHC values indicated that there was presence of macrocytic hypochromic anemia in both the monocrotophos administered groups (A1 and B1) but the severity of the anemia was less in curcumin supplemented group A1. Biochemical studies showed significant decrease in mean total serum protein and albumin concentration and significant increase in serum AST, ALT, GGT, creatinine and uric acid concentrations in groups A1 and B1 as compared to control group B2 at different time intervals. Blood Catalase activity and SOD were significantly decreased in group A1 and B1 as compared to control group B2. The values of mean LPO activity were found higher in both the monocrotophos administered groups (A1 and B1) as compared to control group B2 throughout the experiment on all days post monocrotophos administration. Humoral mediated immune (HMI) response (assessed through serum antibody titre via haemagglutination inhibition test/HI test) and cell mediated immune (CMI) response (assessed through delayed type hypesensitivity/DTH test) were found significantly lowered in monocrotophos administered groups (A1 and B1) as compared to control group B2. Acetylcholine esterase activity (AChE) in brain was found significantly decreased in both monocrotophos administered groups (A1 and B1) as compared to control group B2. On gross examination, chicks of groups A1 and B1 showed varying degrees of atrophy, congestion, haemorrhages and leucoytic cells infiltration in liver, lungs, heart, kidneys, bursa of Fabricius, spleen, thymus, intestine and brain. Histopathological lesions in groups A1 and B1 revealed varying degrees of congestion, degenerative changes in hepatocytes, nephritis, goblet cell hyperplasia in intestinal villi and depletion of lymphocytes in spleen, thymus and bursa of Fabricius. In case of histopathology the lesions were inflammatory, vascular, degenerative, necrotic, proliferative and mild infiltrative type in the monocrotophos administered group (B1) in different organs and were of lesser severity in curcumin supplemented group A1. On the basis of results of the present study it is concluded that curcumin supplementation (@ 200 mg/kg feed) in monocrotophos administered (@ 5 ppm/kg in feed) broiler chicken showed significant improvement in clinical signs, body weight, haemato-biochemical alterations showing its ameliorative effect; significant reduction in oxidative stress, and pathological alterations in different organs which showed protective effect of curcumin; showed significant improvement in humoral and cellular immune response along with acetyl choline esterase activity in brain due to its ameliorative effect.ThesisItem Open Access Development of reverse transcription recombinase polymerase amplification assay for rapid detection of porcine sapelovirus(LUVAS Hisar, 2023-04) Kaur,Ramandeep; Sushila MaanPorcine Sapelovirus (PSV) A, a member of newly classified genus Sapelovirus, belongs to family Picornaviridae. PSV is non-enveloped virus with icosahedral symmetry with diameter of around ~35nm and the genome is a linear, non-segmented, single-stranded positive-sense RNA. PSV infection in pigs is mostly asymptomatic, but it can also cause severe neurologic, enteric, and respiratory symptoms or reproductive failure. Sapelovirus infections have been reported worldwide in pigs. PSV is considered as the emerging pathogen of porcine species. For the virus detection, isothermal Recombinase polymerase amplification (RPA) were developed. The RT-RPA assay was developed using commercially synthesized positive control and primers based on 5´conserved region of PSV genome. The assay was optimized at low and constant temperature of 35°C for 20 minutes with forward primer concentration 20pm, reverse primer concentration 5pm and MgOAc concentration of 14mM. The analytical sensitivity of assay was determined using serial dilutions of positive control. The limit of detection was up to 283 copies using agarose gel electrophoresis. The assay was specific to PSV and no cross reactivity was observed with other related viruses. The assay was validated using field samples followed by visual assay for detection. A total 85 samples were collected from different organized and unorganized piggery farms of Haryana. Out of 85 samples 21 samples were found positive for PSV. The positivity rate of PSV was 24.7%. This is the first report of PSV from Haryana suggesting the virus circulation among pig population. Therefore, the above study concludes that the developed RT-RPA assay is rapid, highly sensitive and specific and works at low, constant temperature and do not require any special instrument. Visual dye-based detection makes the developed RPA assay a potential candidate for pointof-care testing (POCT)ThesisItem Open Access Studies on ameliorative potentials of quercetin nanoparticles against subacute toxicity of imidacloprid with special reference to genotoxicity in Swiss albino mice(LUVAS Hisar, 2023-07) Vipin; Preeti BagriThe human population is increasing day by day, which leads to increase in food demand and use of pesticides. The imidacloprid (IMI) is a neonicotinoid, which act as agonist on nicotinic acetylcholine receptors (nAchRs), specifically at the α-subunits of the nicotinic receptor and use of imidacloprid is increasing. Quercetin (Que) is a bioflavonoid, mainly found in onions, grapes and apple. Quercetin is well known for its pharmacological properties like antioxidant, anti-cancerous, neuroprotective, anti-inflammatory etc. The pharmacological application of quercetin is limited due to its low oral bioavailability (less than 17% in rats and even 1% in humans), low brain permeability and its hydrophobic nature, short biological half-life. However, nanoparticles of quercetin have attracted particular attention in therapeutics due to their excellent stability, high bioavailability and ready ability of crossing the blood brain barrier (BBB). In present study, quercetin nanoparticles were synthesized and characterization was done by hydrodynamic diameter, UV-visible spectra, FT-IR, Zeta potential, SEM, TEM and encapsulation efficiency analysis. In vitro analysis of biocompatibity of nanoparticles was done by hemolytic assay. For in vivo subacute toxicity study for 28 days, total 48 mice were taken and divided in eight groups with six animals in each group. Group 1, 2, 3 and 4 received 3% gum acacia, 22 mg/kg b.wt. imidacloprid, 25 mg/kg b.wt. quercetin and 25 mg/kg b.wt. quercetin nanoparticles high dose [QNPs (HD)], respectively. Group 5, 6, 7 and 8 received 22 mg/kg b.wt. imidacloprid + 25 mg/kg b.wt. quercetin, 22 mg/kg b.wt. imidaclopid + 25 mg/kg b.wt. QNPs (HD), 22 mg/kg b.wt. imidacloprid + 12.5 mg/kg b.wt. quercetin nanoparticle medium dose [QNPs (MD)] and 22 mg/kg b.wt. imidacloprid + 6.25 mg/kg b.wt. quercetin nanoparticles low dose [QNPs (LD)], respectively. Test for Anti-inflammatory effect (Carrageenan induced paw edema), neurobehavioural effect (SMA, Forced locomotor activity and Heat induced algesia), Mutagenic effects (MNT, Comet assay and SHA), oxidative stress in brain and liver (TP, LPO, SA and GSH), acetylcholinesterase activity in brain and histopathology of bone marrow, cerebrum, cerebellum and testes were performed. Imidacloprid predisposed animal to inflammation and both quercetin and quercetin nanoparticles decreased this inflammation. In MNT, there was significant increase in MnPCE, MnNCE and decrease P/N ratio in imidacloprid group as compared to control whereas both quercetin and quercetin nanoparticles decreased occurrence of micronuclei and increase P/N ratio. In comet assay a significant increase in comet length, tail length, tail DNA and tail moment was noticed in imidacloprid group as compared to control whereas both quercetin and quercetin nanoparticle decreased these parameters when given in combination with imidacloprid. In SHA, there was significant increase in abnormal sperm in imidacloprid group as compared to control whereas highest reduction in sperm head abnormality in imidacloprid + quercetin nanoparticles high dose was noticed. Both quercetin and quercetin nanoparticles protect brain and liver tissue from imidacloprid induced oxidative stress. There was no significant change in acetylcholinesterase activity in any group compared to control. In histopathological findings, imidacloprid produced degenerative changes in bone marrow, cerebrum, cerebellum and testes. Both quercetin and quercetin nanoparticles showed ameliorative effect markedly by causing restoration of degenerative changes produced by imidacloprid. On the basis of above results, it may be concluded that both quercetin and quercetin nanoparticles have ameliorative effect against imidacloprid induced subacute toxicity.ThesisItem Open Access Detection of cyromazine and melamine residues in poultry eggs(LUVAS Hisar, 2023-02) Soni, Deepak; Vijay J. JadhavCyromazine (CYR) is an insect growth regulator for fly control at animal farms. It is used as a feed additive in layer farms so that it gets excreted in the droppings of the birds and exert its larvicidal activity in the manure. However, due to this practice, residues of CYR and its metabolite melamine (MEL) also appear in the eggs. A new instrumental method was developed for separation and detection of CYR and MEL on HPLC-UV system. The mobile phase comprised of ACN and 0.05% TFA at pH 2.5 was applied in gradient mode at a flow rate of 1 mL/min. A new method using anhydrous magnesium sulfate, calcium silicate, celite 545 as adsorbents and acidified acetonitrile as extraction solvent was developed for extraction and clean up of CYR and MEL residues in eggs which was found to be quick, easy, cheap, effective, rugged and safe (QuEChERS). The LOD and LOQ of the method was found to be 5.3 and 16.0 µg/kg for CYR and 11.4 and 34.7 µg/kg for MEL, respectively. The accuracy of the method ranged between 84-103 % and precision of the method remained less than 5 % RSD. Linearity studies of spiked sample showed the consistent performance of the method at various concentrations (R²>0.996). A total of 110 egg samples were collected from 52 farms located in 5 different districts of Haryana (Panchkula, Ambala, Panipat, Sirsa and Hisar) and analyzed for the presence of CYR and MEL by applying newly developed QuEChERS method. The mean concentration of the CYR was found to be 42.71 µg/kg with a range of 16.88 -112.61 µg/kg. MEL was not detected in any of the sample analyzed. Estimates of dietary exposure of CYR through consumption of eggs for population in Haryana was found to be 33.40 µg/kg b.w./day which contributed 0.55 % of ADI of CYR. A cross-sectional study was carried out in 52 layer farms situated in 5 districts (Panchkula, Ambala, Panipat, Sirsa and Hisar) of Haryana, India to identify the risk factors associated with the usage of CYR revealed that, factors such as cleanliness practices, number of birds kept at the farm and fly density in the farm were significantly associated with the CYR usage in feed for the purpose of fly control. The findings of the present study suggested that 57.69 % farmers had knowledge of CYR and are adding it in the feed of layers as larvicidal agent. Further it was observed that 66.67 % farmers (n=30) use CYR above the recommended levels. Data analysis revealed that factors such as cleanliness practices at the farm (OR: 5.143, 95% CI: 1.403-18.858), number of birds kept at the farm (OR: 17.417, 95% CI: 4.307-75.146) and fly density in the farm (OR: 6.22, 95% CI: 1.836-21.090) were significantly associated with CYR usage in the layer farms.
