Extraction and Biochemical Characterization of Sulphated Glycosaminoglycans from Chicken Keel Cartilage

The present study was conducted to explore the potential and cheaper source of major and abundantly found sulphated glycosaminoglycans (GAGs) in chicken keel cartilage. Chicken is comparatively readily accessible to all the communities of Pakistan and its cartilages are the rich source of sulphated GAGs. The GAGs were extracted from prewashed and ground keel cartilages (n=3) of chicken using 3 M MgCl2, dialyzed, digested with papain, precipitated with three volumes of ethanol, and finally lyophilized to dry powder. The dry products were used for proximate analysis (carbohydrates 65.49±0.10, crude protein 12.82±0.26, ash 11.12±.56, moisture 9.88±0.32 and fat 0.69±0.14%). Dimethylmethylene blue binding (DMMB) assay was performed to determine the quantity of total GAGs in each group of product and protein contents were estimated by Bradford method. Identification of extracted samples of GAGs was performed with FTIR spectrometer using KBr disc and purity of the samples was determined by SDS-PAGE. Quantity of total GAGs in extracted samples was 70.77±2.27% and estimated amount of protein was 4.64±0.29%. FTIR spectra of standard and samples of CS showed identical and characteristic peaks in finger print region. Finger print region revealed the presence of C-O-S, S=O, -COO, -C-C, R-SO2–R, -CONH2 and R-SO2-NH2 molecules. SDS-PAGE analysis revealed the presence of 77.8 and 50.5 kDa proteins in all extracted samples of GAGs. It can be concluded that chicken keel cartilage is the potential and cheap source of GAGs. Analysis by SDS-PAGE revealed that most of the non-collagen protein can be removed by three volumes of solvent extraction and FTIR is an advance technique for identification of GAGs in mid infrared region (400-4000 cm-1)

Diabetes mellitus among tuberculosis patients: a cross sectional study from Pakistan

Background: The co-occurrence of diabetes mellitus (DM) and tuberculosis (TB) is largely associated with high frequency of morbidity. Objective: To determine the prevalence of DM among TB patients and describe the socio-demographic and behavioral factors associated with TB-DM co-occurrence . Methods: We enrolled 500 TB patients from September, 2014 to August 2015 at four major public sector hospitals of Lahore, Pakistan. A questionnaire was used to collect information regarding associated socio-demographic and behavioral factors of the patients. We monitored the fasting blood sugar of each patient by using a semi automated clinical chemistry analyzer followed by an HbA1c level check of all hyperglycemic patients. Results: The prevalence of TB-DM co-occurrence was 14.8%. The prevalence of TB-DM was higher (62.2%) among males. The >57 year age group had the highest proportion of patients (35.1%), with co-existent TB-DM. Most were illiterate (73.0%) and unemployed (48%). Moreover, among the 74 patients positive for TB-DM had a history of smoking. Age and education level were significantly associated with DM-TB while gender, occupation and smoking were not associated. Conclusion: The study revealed a 14.8% prevalence of DM among TB patients. This was associated with several socio-demographic factors, including age, unemployment, literacy and polluted environment. Thus, poor and unhealthy lifestyles were the factors associated with DM among immunologically compromised individuals due to TB

Production and characterization of monoclonal antibodies to fowl adenoviruses

A panel of 10 monoclonal antibodies to a hypervirulent fowl adenovirus (FAdV) strain 398A was produced. Monoclonal antibodies (mAbs) were characterized for their isotype, neutralizing activity, ability to capture viral antigens, immunoblotting of viral polypeptides, competitive inhibition with chicken antisera and their reaction pattern with other FAdVs in indirect immunoperoxidase. Eight out of 10 mAbs reacted strongly in indirect immunoperoxidase staining with most of the FAdVs isolated from inclusion body hepatitis in Australia. One of these mAbs (6E1) was found to specifically react with the strains presumably characterized as hypervirulent FAdVs. IgG2a was the predominant sub-isotype. Two out of 10 mAbs neutralized the homologous strain of virus and six captured their target antigen onto a plastic surface. Chicken anti-serum to FAdV strain 398A inhibited the reaction of the seven mAbs that bound with high affinity in a blocking competitive enzyme-linked immunosorbent assay. This panel of mAbs can be used to improve diagnostic assays, study pathogenesis and carry out strain identification

Relationship between Disease Activity and Circulating Level of Collagen II C-Telopeptide Fragments in Papain Induced Osteoarthritis Rat Model

Osteoarthritis (OA) is a progressive degeneration of articular cartilage leading to failure in functional mobility of joints. It is characterized by morphological, biochemical and molecular changes in histology of cartilage. Different biological markers are used as indicators to precisely predict the stage of cartilage destruction of joints in OA patients and to evaluate the therapeutic efficacy of drugs used for OA. The present research was chalked out to establish relationship between disease activity and serum level of C-terminal telopeptide of type II collagen (CTX-II) in experimentally induced OA rat model. Out of 30 male Wistar rats, 25 were used to induce OA by injecting papain (10mg/0.5mL of 0.05M sodium acetate) in right knee joints whereas five (control) were injected with sterile normal saline solution on day 0. Blood samples (5mL each) were collected on weekly basis up to 28th days of post papain injection. Sera were separated and subjected to perform ELISA for estimating CTX-II fragments as cartilage biomarker (CartiLaps ® ELISA kit) in experimental groups. Maximum level of CTX–II (pg/mL) (40.44±3.07) was observed in sera samples of day 14 post papain injection followed by days 21 (40.22±2.01), 28 (36.82±3.81), 7 (34.48±4.17), 1 (15.08±4.22) and day 0 (2.55±0.10). The early changes in serum CTX-II from day 0 to 14 showed significant association with cartilage damage. Later on, no significant difference was observed in CTX-II level on day 14, 21 and 28 post papain injection. It is concluded that elevation in serum CTX-II level was concomitant with the onset of disease and degradation of cartilage. Moreover, CTX-II is a sensitive diagnostic biomarker to monitor joint disorder severity in papain induced OA rat experimental model on different days. These findings may be used as base line for early diagnosis of disease and initiation of therapy for successful outcome

Prevalence and Molecular Diagnosis of Babesia ovis and Theileria ovis in Lohi Sheep at Livestock Experiment Station (LES), Bahadurnagar, Okara, Pakistan

  Background: Babesia ovis and Theileria ovis are among the important and main etiological agents causing ovine babesiosis and ovine theileriosis, causing severe economic losses among sheep and goats. The aim of the present study was to determine the prevalence and molecular diagnosis of B. ovis and T. ovis in Lohi sheep at Livestock Experiment Station Bahadurnagar, Okara, Pakistan. Methods: The prevalence of B. ovis and T. ovis was investigated in 200 Lohi sheep of mixed age and sex by PCR during 2011. The assay was employed us-ing primers Bbo-F & Bbo-R, specific for a 549-bp fragment in B. ovis genomic DNA and primers TSsr 170F & TSsr 670R, specific for a 520-bp fragment in T. ovis genomic DNA. The animals were also screened for both haemoparasites through stained thin blood smears. Results: Thirty two (16%), 48 (24%) and 26 (13%) were the number of ani-mals found positive for B. ovis, T. ovis and for mixed infection with both para-sites respectively, through microscopy. Sixty eight (34%), 73 (37%) and 42 (21%) were the number of animals found positive for B. ovis, T. ovis and for mixed infection with both parasites respectively, through PCR test. Conclusion: The results indicate the high sensitivity of PCR for surveying babesiosis and theileriosis and there is noteworthy prevalence of these diseases in sheep at an experimental station where environmental conditions are rela-tively controlled as compared to field conditions

Cross Sectional Survey of Live Bird Markets and Zoo Birds for Circulating Influenza Subtypes in Pakistan

In Pakistan, avian influenza surveillance has been both active and passive. Here, we present the results of a survey effort focusing solely on the live bird markets and wild bird species from different zoos and national parks to understand the impact of live bird markets on the spread of highly pathogenic avian influenza viruses. A cross sectional survey was conducted from Jan-Dec 2011 to identify and isolate the circulating avian influenza virus subtypes in live bird markets and wild birds from different localities in and around Islamabad Capital Territory. Swabs, tracheal tissues and sera samples were collected, screened and diagnosed by hemagglutination inhibition assay and RT-PCR. The highest seropositivity was recorded for H9 (100%) followed by H5 (89.4%) and H7 (72.3%). All 27 isolates were of the low pathogenic H9N2 subtypes and no viruses could be successfully isolated of subtype H5N1 or H7N7. The higher prevalence of H5N1 (89.4%) observed in the present study was an alarming threat; therefore, we suggested immediate control strategies against this emerging risk of H5N1 for human in live bird markets in Pakistan. The factors unveiled in this study will help in understanding the lapses in controlling persistent outbreaks of avian influenza in country.CC BY-NC 4.0</p