scholarly journals Genetic and antigenic analysis of Babesia bigemina isolates from five geographical regions of Brazil

2002 ◽  
Vol 22 (4) ◽  
pp. 153-160 ◽  
Author(s):  
Claudio R. Madruga ◽  
Cássia R.B. Leal ◽  
Alda M.T. Ferreira ◽  
Flábio R. Araújo ◽  
Ana L.V. Bonato ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Genetic Analysis ◽  
Fluorescent Antibody Technique ◽  
Babesia Bigemina ◽  
Chain Reaction ◽  
Antigenic Analysis ◽  
Antibody Technique ◽  
Variable Surface ◽  
Geographical Regions ◽  
Polymerase Chain

A molecular epidemiological study was performed with Babesia bigemina isolates from five geographical regions of Brazil. The genetic analysis was done with random amplification of polymorphic DNA (RAPD), repetitive extragenic palindromic elements-polymerase chain reaction (REP-PCR) and enterobacterial repetitive intergenic consensus sequences-polymerase chain reaction (ERIC-PCR) that showed genetic polymorphism between these isolates and generated fingerprinting. In RAPD, ILO872 and ILO876 primers were able to detect at least one fingerprinting for each B. bigemina isolate. The amplification of B. bigemina DNA fragments by REP-PCR and ERIC-PCR gave evidence for the presence in this haemoprotozoan of the sequences described previously in microorganisms of the bacterial kingdom. For the first time it was demonstrated that both techniques can be used for genetic analysis of a protozoan parasite, although the ERIC-PCR was more discriminatory than REP-PCR. The dendogram with similarity coefficient among isolates showed two clusters and one subcluster. The Northeastern and Mid-Western isolates showed the greatest genetic diversity, while the Southeastern and Southern isolates were the closest. The antigenic analysis was done through indirect fluorescent antibody technique and Western blotting using a panel of monoclonal antibodies directed against epitopes on the merozoite membrane surface, rhoptries and membrane of infected erythrocytes. As expected, the merozoite variable surface antigens, major surface antigen (MSA)-1 and MSA-2 showed antigenic diversity. However, B cell epitopes on rhoptries and infected erythrocytes were conserved among all isolates studied. In this study it was possible to identify variable and conserved antigens, which had already been described as potential immunogens. Considering that an attenuated Babesia clone used as immunogen selected populations capable of evading the immunity induced by this vaccine, it is necessary to evaluate more deeply the cross-protection conferred by genetically more distant Brazilian B. bigemina isolates and make an evaluation of the polymorphism degree of variable antigens such as MSA-1 and MSA-2.

scholarly journals Cattle rabies: the effect of clinical evolution, viral genetic lineage, and viral load on the severity of histological lesions

2020 ◽  
Vol 40 (4) ◽  
pp. 227-233
Author(s):  
Claudia S. Wisser ◽  
André Thaler Neto ◽  
Helena B.C.R. Batista ◽  
Enio Mori ◽  
Maria E.R. Chierato ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcriptase ◽  
Fluorescent Antibody ◽  
Inflammatory Lesion ◽  
Genetic Lineage ◽  
Fluorescent Antibody Technique ◽  
Chain Reaction ◽  
Antibody Technique ◽  
Qrt Pcr ◽  
Polymerase Chain

ABSTRACT: Our objective was the characterization and staging of histological lesions in different anatomical sites of the central nervous system (CNS) of rabid cattle. The severity of the lesions was compared with the clinical stages of the disease, the variants of viral isolates, and with the load of virus. Thirty-one spontaneously affected rabid cattle the state of Santa Catarina underwent clinical follow-up and were eventually necropsied. CNS tissues were sampled and submitted to direct fluorescent antibody technique (DFAT), immunohistochemistry (IHC), routine histopathology with hematoxylin and eosin stain (HE), reverse transcriptase polymerase chain reaction (RT-PCR), and polymerase chain reaction in quantitative reverse transcriptase in real time (qRT-PCR). Affected cattle were allotted in four groups according to their clinical stage when euthanized: G1, euthanized while standing; G2, euthanized when in sternal recumbence; G3, euthanized when in lateral recumbence; and G4, affected cattle with natural death. In order to evaluate the degree of severity of the lesions and the presence of Negri bodies (NBs), the brain was sectioned at 9 sites. Additionally, spinal cord and trigeminal ganglion sections were examined. The intensity of the lesions was graded as either absent, mild, moderate, or marked, and the presence or absence of the NBs was noted. Histological lesions were characterized by lymphocytic and monocytic meningoencephalitis with NBs in 28 cases. In all analyzed groups, intensities of histological lesions ranging from mild to severe were observed. Brain regions with the highest inflammatory lesion intensity were the medulla at the level of obex, followed by the colliculus and thalamus. NBs were observed in a higher percentage in the cerebellum, followed by medulla at the obex level, striatum complex, and frontal telencephalon. The duration of the clinical course of the disease did not influence the intensity of the inflammatory lesion, but it did influence the presence of NBs, with a higher percentage of these inclusions in cattle that died naturally than in euthanized cattle. All isolated rhabdovirus included in this study were genetically compatible with samples from hematophagous bats Desmodus rotundus. The evaluation by qRT-PCR did not demonstrate a correlation between lesion intensity and the amount of virus.

scholarly journals EVALUASI KIT DETEKSI CEPAT TERHADAP SAMPEL OTAK ANJING TERINFEKSI VIRUS RABIES

2015 ◽  
Vol 9 (1) ◽  
Author(s):  
Michael Haryadi Wibowo ◽  
Tri Untari ◽  
Sidna Artanto ◽  
Surya Amanu ◽  
AETH. Wahyuni ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Fluorescent Antibody ◽  
Rapid Test ◽  
Canine Parvovirus ◽  
Fluorescent Antibody Technique ◽  
Chain Reaction ◽  
Antibody Technique ◽  
Test Kit ◽  
Polymerase Chain

Penelitian ini bertujuan mengetahui potensi kit deteksi cepat Anigen® rapid test kit rabies Ag dalam mendeteksi virus rabies pada sampel otakanjing yang diperoleh dari lapangan yang meliputi batas deteksi, kecepatan reaksi, uji reaksi silang, uji sensitivitas, dan spesifisitas. Batas deteksi ditentukan dengan pengenceran secara serial kontrol positif virus rabies dan selanjutnya diuji dengan rapid test kit sesuai petunjuk produsen. Uji reaksi silang dilakukan dengan canine parvovirus, Escherichia coli, dan Salmonella sp. Uji sensitivitas dan spesifisitas dilakukan terhadap sampel otak yang telah dikonfirmasi positif rabies dengan uji fluorescent antibody technique. Konfirmasi uji rapid test tersebut dilakukan dengan reverse transcriptase polymerase chain reaction. Berdasarkan data yang diperoleh dalam penelitian ini dapat disimpulkan bahwa Anigen® rapid test kit rabies Ag mampu mendeteksi sampel yang mengandung virus rabies dengan titer 0,5 x log 106,5/0,03 ml, dengan rata-rata kecepatan reaksi 1,8 menit 29,35 detik (kurang dari 2 menit). Di samping itu Anigen® rapid test kit rabies menunjukkan tidak terdapat reaksi silang dengan canine parvovirus, Escherichia coli, dan Salmonella sp. serta mempunyai sensitivitas 92,30% dan spesifisitas 97,90%

scholarly journals Application of Polymerase Chain Reaction for the Correlation of Salmonella Serovars Recovered from Greyhound Feces with Their Diet

1993 ◽  
Vol 5 (3) ◽  
pp. 378-385 ◽  
Author(s):  
Gregory G. Stone ◽  
M. M. Chengappa ◽  
Richard D. Oberst ◽  
Nathan H. Gabbert ◽  
Scott McVey ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Fecal Material ◽  
Chain Reaction ◽  
Antigenic Analysis ◽  
Fecal Samples ◽  
E Coli ◽  
Staphylococcus Intermedius ◽  
Standard Procedures ◽  
Polymerase Chain ◽  
Salmonella Serovars

The polymerase chain reaction was employed to correlate Salmonella serovars isolated from fecal material of greyhounds suffering from gastroenteritis with those isolated from the diet fed to the greyhounds prior to onset of diarrhea. Kennels around the Abilene, Kansas, area were contacted and supplied with materials needed to collect a portion of the diet each day. With t e onset of diarrhea, the kennels were instructed to ship the fecal material and diet from the previous 10 days to the laboratory for testing. Forty-one fecal samples and corresponding diets were screened for Salmonella, Clostridium perfringens, Campylobacterjejuni, Staphylococcus aureus, Staphylococcus intermedius, and pathogenic (piliated) Escherichia coli by direct culture using standard procedures. The fecal material was also screened for coronavirus and parvovirus using electron microscopy. Thirty-five “normal” fecal samples were screened for all of the above mentioned microorganisms as a control. In addition, the fecal material was screened for E. coli verotoxins I and II and clostridial enterotoxins. A total of 61 Salmonella isolates were recovered from the 41 samples of feces and diet submitted for testing; 31 were recovered from the feces and 30 from the diet. Four Salmonella isolates were recovered from the normal fecal samples. Results obtained by PCR, plasmid profiles, antigenic analysis, and antibiogram profiles indicated that 16 of the 31 isolates recovered from the fecal material were the same strain as that recovered from the diet.

scholarly journals Babesia bovis and B. bigemina DNA detected in cattle and ticks from Zimbabwe by polymerase chain reaction

2000 ◽  
Vol 71 (1) ◽  
Author(s):  
I. Smeenk ◽  
P.J. Kelly ◽  
K. Wray ◽  
G. Musuka ◽  
A.J. Trees ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Boophilus Microplus ◽  
Babesia Bovis ◽  
Pcr Analysis ◽  
Babesia Bigemina ◽  
Chain Reaction ◽  
Boophilus Decoloratus ◽  
Polymerase Chain

From blood collected from 94 cattle at 12 locations in the eastern and northeastern areas of Zimbabwe, DNA was extracted and analysed by polymerase chain reaction with primers previously reported to be specific for Babesia bigemina and Babesia bovis. Overall, DNA of Babesia bigemina was detected in the blood of 33/94 (35 %) cattle and DNA from B. bovis was detected in 27/58 (47 %) of cattle. The prevalence of DNA of B. bigemina was significantly higher in young animals (<2 years) (23/46) than in animals over 2 years of age (10/48; (chi)2 = 8.77; P < 0.01 %). Although tick sampling was not thorough, Boophilus decoloratus could be collected at 7/9 sites sampled and Boophilus microplus at 4/9 sites. Of the 20 B. decoloratus allowed to oviposit before PCR analysis, 1 (5 %) contained DNA that could be amplified with primers for B. bigemina while 12 (60 %) were positive with primers for B. bovis. Of the B. microplus allowed to oviposit, 11/16 (69 %) were positive for B. bovis DNAby PCR and 2/16 (12 %) were positive for B. bigemina.

Genetic analysis of nuclear DNA restriction fragment patterns

Genome ◽  
1991 ◽  
Vol 34 (5) ◽  
pp. 693-703 ◽  
Author(s):  
Elizabeth M. Gillet
Keyword(s):  
Polymerase Chain Reaction ◽  
Restriction Fragment Length Polymorphism ◽  
Genetic Analysis ◽  
Restriction Fragment ◽  
Length Polymorphism ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Restriction Fragment Length

Restriction fragment length polymorphism (RFLP) analysis in the broad sense is the analysis of differences in restriction fragment pattern produced by defined target segments within or between cell compartments, cell types, etc., in a single individual or in different individuals. Thus both molecular hybridization and DNA amplification by two-primer extension using the polymerase chain reaction can define target segments for RFLP analysis. The two techniques are outlined with special consideration of characteristics important for genetic analysis. The mode of inheritance of restriction fragment patterns as a prerequisite for their use as genetic markers in inheritance studies is explained, leading to criticism of common usage. The importance of internal restriction sites for the determination of allelic variation is stressed. It is shown that, if target segments are under the control of a single nuclear diploid restriction fragment locus, then complete reconstruction of all parental target segments requires controlled crosses between individuals of like restriction fragment pattern.Key words: genetic analysis, inheritance, restriction fragment length polymorphism, controlled cross, polymerase chain reaction.

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