scholarly journals Molecular diagnosis and biochemical studies of tick-borne diseases (anaplasmosis and babesiosis) in Aberdeen Angus Cattle in New Valley, Egypt

2020 ◽  
Vol 13 (9) ◽  
pp. 1884-1891
Author(s):  
Nani Nasreldin ◽  
Rania M. Ewida ◽  
Hatem Hamdon ◽  
Yasser F. Elnaker
Keyword(s):  
Oxidative Stress ◽  
Polymerase Chain Reaction ◽  
Molecular Techniques ◽  
Babesia Bovis ◽  
Blood Parasites ◽  
Chain Reaction ◽  
Infected Female ◽  
Angus Cattle ◽  
Polymerase Chain ◽  
Female Cattle

Background and Aim: Anaplasmosis and babesiosis are tick-borne diseases that threaten livestock production with subsequent considerable economic losses. This study was conducted to diagnose Anaplasma and Babesia infection using molecular techniques in imported Aberdeen Angus cattle imported from Uruguay to El-Kharga Oasis in New Valley, Egypt, and to investigate the effects of disease on some serum biochemical and oxidative stress parameters. Materials and Methods: Blood samples were collected from 31 cattle, 21 diseased and ten apparently normal, of varying ages and sex. The blood was used for the preparation of blood smears, polymerase chain reaction assay, and separation of serum for biochemical investigation. The experimental production farm at the Faculty of Agriculture, New Valley University, was infested with ticks and variable clinical manifestations during the period from December 2017 to March 2018. One calf died of a suspected blood parasite infection. Results: The blood film examination revealed infection by blood parasites in 21 samples. Anaplasma marginale and Babesia bovis were identified in 12 and 14 samples, respectively. A total of 14 samples were examined by polymerase chain reaction (PCR) to make these identifications. Biochemical parameters showed significantly elevated serum alanine aminotransferase, aspartate aminotransferase, total bilirubin (T. Bil), and urea in blood from parasite-infected female cattle and male calves compared with controls. Increased serum total protein, globulin, and creatinine were recorded only in infected female cattle. The blood glucose level was significantly decreased in infected female cattle and male calves compared with controls. Furthermore, albumin and albumin/globulin ratio was significantly reduced in the infected female cattle. Oxidative stress profiles of infected animals showed a significant increase in serum nitric oxide and malondialdehyde, and both total antioxidant capacity and reduced glutathione (GSH) were significantly reduced in comparison with control animals. Conclusion: The incidence of A. marginale and B. bovis infection is high in imported Aberdeen Angus cattle in New Valley Province. PCR methods provide a short-term assessment of disease. An extensive epidemiological survey, employing serology together with molecular genetic methods, monitoring of abundance and distribution of tick vectors, availability of vaccination programs, and tracking of animal transport is also needed for control of blood parasites.

Diagnostic Accuracy of a Direct Panfungal Polymerase Chain Reaction Assay Performed on Stained Cytology Slides

2021 ◽  
pp. 030098582199156
Author(s):  
Alexandra N. Myers ◽  
Unity Jeffery ◽  
Zachary G. Seyler ◽  
Sara D. Lawhon ◽  
Aline Rodrigues Hoffmann
Keyword(s):  
Polymerase Chain Reaction ◽  
Diagnostic Accuracy ◽  
Molecular Techniques ◽  
Fungal Culture ◽  
Chain Reaction ◽  
Identification Of Fungi ◽  
Fungal Dna ◽  
Polymerase Chain ◽  
Panfungal Pcr ◽  
Pcr Targeting

Molecular techniques are increasingly being applied to stained cytology slides for the diagnosis of neoplastic and infectious diseases. Such techniques for the identification of fungi from stained cytology slides have not yet been evaluated. This study aimed to assess the diagnostic accuracy of direct (without nucleic acid isolation) panfungal polymerase chain reaction (PCR) followed by sequencing for identification of fungi and oomycetes on stained cytology slides from dogs, cats, horses, and other species. Thirty-six cases were identified with cytologically identifiable fungi/oomycetes and concurrent identification via fungal culture or immunoassay. Twenty-nine controls were identified with no cytologically or histologically visible organisms and a concurrent negative fungal culture. Direct PCR targeting the internal transcribed spacer region followed by sequencing was performed on one cytology slide from each case and control, and the sensitivity and specificity of the assay were calculated. The sensitivity of the panfungal PCR assay performed on stained cytology slides was 67% overall, 73% excluding cases with oomycetes, and 86% when considering only slides with abundant fungi. The specificity was 62%, which was attributed to amplification of fungal DNA from control slides with no visible fungus and negative culture results. Direct panfungal PCR is capable of providing genus- or species-level identification of fungi from stained cytology slides. Given the potential of panfungal PCR to amplify contaminant fungal DNA, this assay should be performed on slides with visible fungi and interpreted in conjunction with morphologic assessment by a clinical pathologist.

scholarly journals Sensitive SYBR Green—Real Time PCR for the Detection and Quantitation of Avian Rotavirus A

2018 ◽  
Vol 6 (1) ◽  
pp. 2 ◽  
Author(s):  
David De la Torre ◽  
Claudete Astolfi-Ferreira ◽  
Ruy Chacon ◽  
Antonio Piantino Ferreira
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Sybr Green ◽  
Molecular Techniques ◽  
Economic Losses ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Rotavirus A ◽  
Avian Nephritis Virus ◽  
Polymerase Chain

Avian rotavirus A (ARtV-A) is a virus that affects young birds, causing acute diarrhea and economic losses in the poultry industry worldwide. The techniques used for the diagnosis of ARtV-A include electron microscopy, isolation in cell culture, and serology, as well as molecular techniques, such as the reverse transcription-polymerase chain reaction (RT-PCR). The objective of this work was to standardize a real-time RT-polymerase chain reaction (RT-qPCR) using SYBR Green chemistry for the rapid detection and quantification of ARtV-A from bird tissues and materials fixed on FTA cards on the basis of the nucleotide sequence of segment 6 (S6), which codes for the structural VP6 protein of ARtV-A. The results show the efficient amplification of the proposed target, with a limit of detection (LoD) of one copy gene (CG) per microliter of cDNA and a limit of quantification (LoQ) of 10 CGs per microliter. The efficiency of the primers was determined to be 95.66% using a standard curve, with an R2 value of 0.999 and a slope of −3.43. The specificity was determined using samples coinfected with ARtV-A, the chicken parvovirus, the chicken astrovirus, and the avian nephritis virus as positive controls and commercially available vaccines of the infectious bronchitis virus, infectious bursa disease virus, avian reovirus and healthy organs as negative controls. This technique, which lacks nonspecific PCR products and dimers, demonstrated greater sensitivity and specificity than conventional RT-PCR, and it reduced the analysis time by more than 50%.

Extraction of microsporidial DNA from modified trichrome-stained clinical slides and subsequent species identification using PCR sequencing

Parasitology ◽  
2008 ◽  
Vol 135 (6) ◽  
pp. 701-703 ◽  
Author(s):  
K. S. CHAN ◽  
T. H. KOH
Keyword(s):  
Polymerase Chain Reaction ◽  
Species Identification ◽  
Environmental Samples ◽  
Species Level ◽  
Molecular Techniques ◽  
Chain Reaction ◽  
Clinical Specimens ◽  
Polymerase Chain ◽  
Pcr Conditions ◽  
Hiv Negative

SUMMARYMolecular techniques involving polymerase chain reaction (PCR) and sequencing provide a relatively simple and objective means of identifying microsporidia to species level. We modified previously described methods of DNA extraction and PCR conditions for identification of microsporidia from museum slides, clinical specimens and environmental samples and successfully identifiedVittaforma corneaein 11 out of 13 cases of microsporidial infection from used trichrome-stained slides of corneal scrapings from HIV-negative patients with keratoconjunctivitis.

scholarly journals Portable sequencer in the fight against infectious disease

2019 ◽  
Vol 65 (1) ◽  
pp. 35-40 ◽  
Author(s):  
Arthur Elia Mongan ◽  
Josef Sem Berth Tuda ◽  
Lucky Ronald Runtuwene
Keyword(s):  
Infectious Disease ◽  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Disease Management ◽  
Acid Composition ◽  
Molecular Techniques ◽  
Chain Reaction ◽  
Major Threat ◽  
The World ◽  
Polymerase Chain

Abstract Infectious disease is still a major threat in the world today. Five decades ago, it was considered soon to be eradicated, but the adaptation of pathogens to environmental pressure, such as antimicrobials, encouraged the emergence and reemergence of infectious disease. The fight with infectious disease starts with prevention, diagnosis, and treatment. Diagnosis can be upheld by observing the cause of disease under the microscope or detecting the presence of nucleic acid and proteins of the pathogens. The molecular techniques span from classical polymerase chain reaction (PCR) to sequencing the nucleic acid composition. Here, we are reviewing the works have been undertaken to utilize a portable sequencer, MinION, in various aspects of infectious disease management.

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.

scholarly journals Real-Time Polymerase Chain Reaction for One-Hour On-Site Diagnosis of Pierce's Disease of Grape in Early Season Asymptomatic Vines

2002 ◽  
Vol 92 (7) ◽  
pp. 721-728 ◽  
Author(s):  
N. W. Schaad ◽  
D. Opgenorth ◽  
P. Gaush
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
Xylella Fastidiosa ◽  
Molecular Techniques ◽  
Plant Diseases ◽  
Pierce's Disease ◽  
Chain Reaction ◽  
Pierce’S Disease ◽  
Polymerase Chain

Molecular-based techniques, such as polymerase chain reaction (PCR), can reduce the time needed for diagnosis of plant diseases when compared with classical isolation and pathogenicity tests. However, molecular techniques still require 2 to 3 days to complete. To the best of our knowledge, we describe for the first time a real-time PCR technique using a portable Smart Cycler for one-hour on-site diagnosis of an asymptomatic plant disease. Pierce's disease (PD) of grape, caused by the fastidious bacterium Xylella fastidiosa, causes serious losses in grapes in California and the southeastern United States. The disease has been difficult to diagnose because typical leaf scorching symptoms do not appear until late (June and after) in the season and the organism is very difficult to isolate early in the season. Sap and samples of macerated chips of secondary xylem from trunks of vines were used in a direct real-time PCR without extraction of DNA. Using two different sets of primers and probe, we diagnosed PD in 7 of 27 vines (26%) from four of six vineyards sampled 10 to 12 days after bud break in Kern, Tulare, and Napa counties of California. The diagnosis was confirmed by isolation of Xylella fastidiosa from two of the original PCR positive samples and later from symptomatic leaf petioles of four out of four vines from one vineyard that were originally PCR positive.

scholarly journals Molecular detection of Babesia bovis in cattle in Al-Qadisiyah province

2017 ◽  
Vol 40 (2) ◽  
pp. 155-158
Author(s):  
Noaman N. A,aiz
Keyword(s):  
Polymerase Chain Reaction ◽  
Infection Rate ◽  
Molecular Detection ◽  
Babesia Bovis ◽  
Chain Reaction ◽  
Adult Cattle ◽  
Blood Samples ◽  
Significant Difference ◽  
Polymerase Chain ◽  
Age And Sex

     This study aim to determine Babesia bovis infection in cattle based on genetic methods. A total of 96 blood samples were collected from alive and slaughtered cattle from different areas in addition to the abattoir of Al-Qadisiyah province from December 2013 to August 2014. Real time polymerase chain reaction (RT.PCR) technique was used to detect the presence of the protozoan with the effect of animal's age and sex in the infection rate 47.91 % (46/96) of examined cattle were given positive result to B. bovis infection. The highest infections were shown among the adult cattle (≥1 year), while there was non-significant difference (P>0.05) in the infection rate according to the sex. So the most cattle in Al-Qadisiyah province appear to be bearing the infection predominantly as a carrier hosts.

scholarly journals MOLECULAR DIAGNOSIS AND PHYLOGENETIC ANALYSIS OF BABESIA SPECIES ISOLATED FROM TICKS OF INFESTED CATTLE IN WASIT GOVERNORATE, IRAQ

2021 ◽  
Vol 52 (1) ◽  
pp. 136-145
Author(s):  
Al-Abedi & Al-Amery
Keyword(s):  
Polymerase Chain Reaction ◽  
Phylogenetic Analysis ◽  
Molecular Diagnosis ◽  
Babesia Bovis ◽  
Babesia Species ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Pcr Products ◽  
Polymerase Chain

The aim of current study is to detect Babesia bovis, B. bigemina, and B. divergens in ticks using molecular polymerase chain reaction (PCR) assay. In a totally 180 cattle examined to collect of tick samples during December 2018 to August 2019, the findings were revealed on 63 (35%) cattle infested with ticks that classified morphologically to belong to the genus of Hyalomma and genus of Rhipicephalus. From 50 tick samples tested by PCR assay, 41 (82%) were infested by Babesia genus including 30 (68.18%) infested with B. bovis and 11 (31.82%) infested with B. bigemina; whereas, no tick samples were found to be infested with B. divergens. To document the local isolated strains, five PCR products of each B. bovis and B. bigemina positive strains were selected, sequenced and reported in the NCBI under the accession numbers of (MN727083.1, MN727084.1, MN727085.1, MN727086.1, and MN727087.1) and (MN741113.1, MN741114.1, MN741115.1,MN741116.1, and MN741117.1) respectively.

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