Browsing by Author "Ilhan, Z."

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Comparison of Culture and Pcr for the Detection of Brucella Melitensis in Blood and Lymphoid Tissues of Serologically Positive and Negative Slaughtered Sheep
(Blackwell Publishing, 2008) Ilhan, Z.; Aksakal, A.; Ekin, I. H.; Gulhan, T.; Solmaz, H.; Erdenlig, S.
Aims: To compare the culture and PCR methods for detection of Brucella melitensis in blood and lymphoid tissue samples obtained from slaughtered sheep (n = 162) testing positive/negative in serological tests (Rose Bengal test and serum agglutination test). Methods and Results: Of 162 sheep examined, 45 were positive and 117 negative in serological tests. A PCR assay based on a pair of Br. melitensis-specific primers was used to detect DNA in blood and lymphoid tissue. Brucella melitensis was isolated from 1.2% (2/162) and 17.2% (28/162) of the blood and lymphoid tissue samples respectively. Positive PCR products with a molecular size of 731 bp were obtained from 27.7% (45/162) of blood and 29.0% (47/162) of lymphoid tissue samples. Conclusions: The species-specific PCR assay detected a higher number of Br. melitensis DNA both from serologically positive (P < 0.01 in blood PCR, P < 0.001 in tissue PCR) and serologically negative (P < 0.001 in both blood PCR and tissue PCR) sheep compared with classical bacteriological culture methods. Significance and Impact of the Study: The results emphasize the importance of using more than one type of diagnostic technique for the detection of animals positive for brucellosis, especially with epidemiological purposes.
Comparison of Pcr Assay and Bacteriological Culture Method for the Detection of Brucella Melitensis in Stomach Content Samples of Aborted Sheep Fetuses
(M H Schaper Gmbh Co Kg, 2007) Ilhan, Z.; Solmaz, H.; Aksakal, A.; Gulhan, T.; Ekin, I. H.; Boynukara, B.
The aim of this study was to evaluate the polymerase chain reaction (PCR) assay for detection of Brucella melitensis in stomach content samples of aborted sheep fetuses and to compare its performance with bacteriological culture method. It was also aimed to determine the agreement between PCR and Rose Bengal plate test (RBPT). Materials were collected from aborted sheep from 109 different sheep flocks in the region of Van during the lambing seasons of 2004-2005 and 2005-2006. Stomach contents from 135 aborted sheep fetuses were examined by bacteriological culture and PCR, and 135 sera from these aborting ewes were tested by RBPT Identification and typing of Brucella strains were performed using standard classification test. B. melitensis biovar 3 was isolated from 26 (19.2 %) of foetal stomach contents. B. melitensis was detected by PCR in 29 (21.4 %) stomach content samples. Twenty five sera (18.5 %) from aborting ewes tested positive by RBPT The detection limit of B. melitensis 16 M strain by PCR was 1.7 x 10(3) cfu (colony forming units) /ml in spiked stomach contents. Diagnostic sensitivity and specificity of the PCR were detected as 100 % and 97.2 %, respectively. The agreement between PCR and RBPT was found to be 97 %. In conclusion, PCR assay would have an advantage over conventional bacteriological culture method, but in particular for its ability to meet the specificity requirements for the detection of B. melitensis in stomach content samples of aborted sheep fetuses.
Comparison of the Dot-Immunobinding Assay With the Serum Agglutination Test, the Rose Bengal Plate Test and the Milk Ring Test for the Detection of Brucella Antibodies in Bovine Sera and Milk
(Blackwell Verlag GmbH Berlin, 1999) Gürtürk, K.; Boynukara, B.; Ilhan, Z.; Hakki Ekin, I.; Gülhan, T.
In this study, Brucella antibodies in bovine sera and milk were detected using the dot-immunobinding assay (DIA), the serum agglutination test (SAT), the Rose Bengal plate test (RBPT) and the milk ring test (MRT). For this purpose, a total of 116 paired blood and milk samples collected at the same time from 56 aborted and from 60 healthy dairy cows was examined. In DIA, a nitrocellulose membrane (NCM) was used as the solid phase. Antigen adsorbed on the NCM was extracted from Brucella abortus S99 by heat treatment. The results obtained by DIA were compared with those of SAT, RBPT and MRT. Of the 116 paired blood and milk samples, 24 were positive and 72 were negative by all tests used. Serum samples of six aborted cows were positive by DIA, SAT and RBPT but the milk samples were negative by DIA and MRT. Serum and milk samples of four aborted cows gave positive reaction only by DIA tests. The remaining six aborted cows were negative only by MRT and two of them were negative by both RBPT and MRT. Four sera of healthy cows were found to be positive only by SAT. © 1999 Blackwell Wissenschafts-Verlag.
Detection of Brucella Antibodies in Sheep Sera Using Dot-Immunobinding Assay and Rose Bengal Plate Agglutination Test
(Scientific and Technical Research Council of Turkey, 1997) Gürtürk, K.; Ilhan, Z.; Erganiş, O.
In this study, a dot-immunobinding assay (DIA) using nitrocellulose membrane was performed for the detection of Brucella antibodies in 206 sheep sera. Extract of Brucella abortus strain S99 obtained by heat treatment was used as antigen. The results were compared with those of Rose Bengal plate agglutination test (RBPAT). Of the 96 sera collected from the 8 aborted sheep flocks, 46 were positive by both tests and 17 were positive only in DIA. Of the 110 sera collected from slaughtered sheep, 13 were positive by both tests and 4 were positive only in DIA. Of the total 206 sera tested, 147 were negative by RBPAT and only 126 were negative in DIA. All the sera giving positive reactions in RBPAT were also positive in DIA. It is concluded that DIA would be useful to apply for the detection of Brucella antibodies in sheep sera.
Detection of Brucella Melitensis Dna in the Milk of Sheep After Abortion by Pcr Assay
(Univ Austral Chile, Fac Ciencias veterinarias, 2008) Ilhan, Z.; Solmaz, H.; Aksakal, A.; Gulhan, T.; Ekin, I. H.; Boynukara, B.
Laboratory diagnosis of brucellosis is generally performed by microbiological and serological methods. PCR assay is a specific and sensitive choice for the detection of different bacterial agents. An evaluation of this test was carried out for the detection of Brucella melitensis DNA in sheep milk. 102 milk samples from sheep after abortion were taken and studied using bacteriological culture, PCR and milk ring test (MRT). PCR found B. melitensis DNA in 24 (23.5%) out of 102 milk samples, while only 8 (7.8%) of the samples were positive to B. melitensis through direct culture. MRT found 28 (27.4%) positive milk samples. The detection limit for PCR in sheep milk inoculated with B. melitensis strain 16 M was 1.7 x 10(3) - 1.7 x 10(4) cfu/ml. PCR and MRT coincidence was 96%. The diagnostic sensitivity and specificity were determined as 100% and 81.3% respectively for PCR assay and 75% and 75% for MRT. PCR is a useful tool for a fast diagnosis of B. melitensis in sheep milk.
Detection of Corynebacterium Pseudotuberculosis From Sheep Lymph Nodes by Pcr
(Ecole Nationale veterinaire Toulouse, 2013) Ilhan, Z.
Corynebacterium pseudotuberculosis is a facultative intracellular bacterium that causes caseous lymphadenitis in sheep and goats. This study was designed to evaluate the validity of PCR assay protocol for the direct detection of C. pseudotuberculosis in 147 samples of lymph nodes (prescapular and mediastinal) from carcasses of naturally infected sheep and to compare its performance with the traditional bacteriological culture technique. C. pseudotuberculosis was isolated in 81 samples mainly from prescapular nodes and a specific 203 bp PCR amplified pld gene DNA fragment was detected in 85 samples. The total agreement score between the 2 methods was 95.92%, the relative PCR sensitivity and specificity to culture being 98.76% and 92.42%, respectively. The positive and negative predictive probabilities given by PCR were 98.4% and 94.1%, respectively. The C. pseudotuberculosis detection limit given by PCR was 2.1x10(4)-2.1x10(3) CFU/mL in sheep lymph tissue samples. In conclusion, the PCR assay proved to be a sensitive and rapid method for the detection of C. pseudotuberculosis in lymph node samples from naturally infected sheep.
Determination of Avian Influenza a Viruses in Some Avian Species in Van Lake Basin by Real Time-Pcr, Isolation and Subtyping
(2011) Boynukara, B.; Gulhan, T.; Adizel, O.; Ilhan, Z.; Aksakal, A.; Durmus, A.; Solmaz, H.
In this study, feces samples collected during 37 months from February 2006 to March 2009 from 2013 animals consisting of 47 avian species covering irregular vagrant, transit migrant, winter visitor, migratory and native birds in the Van Lake Basin Turkey were tested by Real-Time PCR (RT-PCR) with respect to Avian Influenza (AI) type A virus M2 gene. Of them, 59 samples (2.9%) were found to be positive. RT-PCR positive samples were examined with the same method with respect to H5N1 and 4 samples (6.8%) were found to be positive. RT-PCR positive 59 samples were inoculated in Embryonated Chicken Egg (ECE) and AI type A virus was isolated from 12 samples (20.3%). Of the isolates, 3 were typed as H1N7, 2 as H7N9, 2 as H11N9 and 1 as H8N4 with Hemagglutination Inhibition (HI) and Neuraminidase Inhibition (NI) tests. About 4 isolates obtained from winter visitor Anas cylpeata which had been determined as H5N1 by RT-PCR and agarose gel electrophoresis, gave positive reaction by HI test both with HI and H5 antisera and all were typed as Nl by Nl-test. Feces samples found to be positive by RT-PCR belonged to avian species Anseriformes, Charadriiformes, Pas seriformes, Gruiformes and Phoenicopteriformes orders. The highest positivity was determined in winter visitor Anas acuta (37.1%) and Anas penelope (22.5%) ducks. Of the RT-PCR positive 59 samples, 43 (72.9%) were determined in the samples collected during winter and spring of 2006-2009. Positivity was determined at a rate of 35.2% in respect of AI type A by RT-PCR in different species sharing the same time and place. With this study, the presence of AI type A viruses in various wild birds in the Van Lake Basin was determined for the first time in Turkey. © Medwell Journals, 2011.
Determination of Campylobacter Fetus Subsp. Fetus and Campylobacter Jejuni in Aborted Sheep Fetuses by Multiplex Pcr Assay
(Israel veterinary Medical Assoc, 2021) Ilhan, Z.; Ekin, I. H.; Gulaydin, O.
Campylobacteriosis is a contagious and zoonotic infection characterized by abortion and infertility in animals. Ovine campylobacteriosis is caused by Campylobacter (C.) fetus subsp. fetus or C. jejuni. As a result of the slow-growing bacterial agents and the high lability of Campylobacter species, laboratory diagnosis of these agents has always been problematic. Several publications on detection of Campylobacter spp. DNA from reference strains or pure culture have been presented. However, studies that have been carried out on field or clinical samples of sheep are quite limited. The purpose of the current study was to evaluate the utilization of multiplex-PCR (m-PCR) assay in the diagnosis of ovine campylobacteriosis from abomasal content samples of aborted sheep fetuses. A total of 116 aborted sheep fetuses were tested and C. fetus subsp. fetus was isolated from 8 (6.9%) samples. The m-PCR assay was conducted and amplified C. fetus subsp. fetus-specific DNA in 13 (11.2%) of the abomasum contents. Cohen's kappa coefficient revealed substantial (kappa: 0.74) agreement between bacteriological culture and m-PCR methods. The results of this study suggest that the m-PCR assay is more sensitive and reliable than conventional bacteriological culture for detection of Campylobacter species.
Immunohistochemical Detection of Mannheimia (Pasteurella) Haemolytica Antigens in Goats With Natural Pneumonia
(Springer, 2009) Yener, Z.; Ilhan, F.; Ilhan, Z.; Saglam, Y. S.
Pneumonia is a leading cause of loss to ruminants throughout the world. Mannheimia (Pasteurella) haemolytica is one of the most important etiological agent of pneumonia in cattle, sheep, and goats. This study was carried out to determine the incidence of M.haemolytica antigens using immunohistochemistry labelling of formalin-fixed, paraffin-embedded tissues in pneumonic lungs of goats slaughtered at abattoir, and then to compare these immunohistochemistry results with the results of bacterial isolation. For these objectives, a total of 1505 goat lungs slaughtered in slaughterhouse were grossly examined and pneumonia was detected in 74 cases (4.91%). Of these, with the exception of verminous pneumonia observed in 32 cases, on 42 pneumonic lungs immunohistochemical examinations were performed. Formalin-fixed and paraffin-embedded lung tissue samples were immunohistochemically stained by the avidin-biotin-peroxidase complex (ABC) procedure using polyclonal antibodies to detect M.haemolytica antigens. Pneumonic lesions were more frequently encountered in cranioventral lobes than caudal lobes, and characterized by irregular lobular foci of atelectasis or lobar pneumonia. The presence of M.haemolytica antigens was detected in 19 (45%) out of 42 pneumonic lungs. Bacterial antigens were found most frequently in the cytoplasm of bronchial and bronchiolar epithelial cells, in the swirling degenerating leukocytes in the alveoli, and in the degenerating leukocytes in the area of coagulation necrosis, less frequently in the epithelial cells of bronchial glands, and lymphoid cells. Conclusionly, immunohistochemical detection of M.haemolytica antigens in pneumonic lungs appear to be more reliable compared to bacterial isolation.
A Pcr Method With Internal Control for Detection of Brucella Spp. From Bovine Abortion Samples
(Ecole Nationale veterinaire Toulouse, 2014) Solmaz, H.; Cantekin, Z.; Altug, N.; Ilhan, Z.; Aslan, S.; Ergun, Y.
Brucella spp. are important infectious agents in bovine abortions worldwide. The bacteriological culture of Brucella spp. is fastidious and time consuming procedure as a classical laboratory method. Brucella spp. can be detected by using different molecular techniques. The aim of this study is to develop a PCR technique with an internal control for detection of Brucella spp. from bovine abortion samples. For this purpose, the sensitivities of three different primer pairs (BgF/BgR, B4/B5 and JP-R/JP-F) were compared. Bovine 12S gene specific primer pairs (12SM-FW/12SBT-REV2) were used as an internal control. The sensitivity of BgF/BgR primers was found higher than the other primer sets. A PCR assay was developed by combining BgF/BgR primer sets and primers for bovine 12S. This protocol was tested and validated by using abomasal contents of two Brucella-positive and eighteen Brucella-negative clinical samples. In conclusion, the developed PCR method with an internal positive control has a potential for use in direct detection and identification of the Brucella spp. from bovine abortion samples.