scholarly journals Hepatozoon canis and Leishmania spp. coinfection in dogs diagnosed with visceral leishmaniasis

2016 ◽  
Vol 25 (4) ◽  
pp. 450-458 ◽  
Author(s):  
Fernanda Nazaré Morgado ◽  
Amanda dos Santos Cavalcanti ◽  
Luisa Helena de Miranda ◽  
Lúcia Helena O’Dwyer ◽  
Maria Regina Lucas da Silva ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Dna Isolation ◽  
Parasite Burden ◽  
Leishmania Species ◽  
18S Rrna Gene ◽  
Hepatozoon Canis ◽  
Histopathological Analysis ◽  
Rrna Gene ◽  
Pcr Rflp ◽  
Leishmania Spp

Abstract This study describes the occurrence of dogs naturally co-infected with Hepatozoon canis and two Leishmania species: L. infantum or L. braziliensis. Four dogs serologically diagnosed with Visceral Leishmaniasis were euthanized. Liver and spleen samples were collected for histopathological analysis and DNA isolation. H. canis meronts were observed in tissues from all four dogs. H. canis infection was confirmed by PCR followed by sequencing of a fragment of 18S rRNA gene. Leishmania detection and typing was confirmed by ITS1' PCR-RFLP and parasite burden was calculated using ssrRNA quantitative qPCR. A DPP - Dual Path platform test was performed. One out (Dog #2) of four animals was asymptomatic. Dogs #1 and #4 were infected by L. infantum and were DPP test positive. Dogs #2 and #3 were infected by L. braziliensis and were DPP test negative. Furthermore, visceral dissemination was observed in Dogs #2 and #3, since L. braziliensis was detected in liver and spleen samples. The visceral dissemination of L. braziliensis associated with systemic signs suggested that this co-infection could influence the parasite burden and disease progression.

scholarly journals Molecular Characterization ofCryptosporidiumspp. in Wild Rodents of Southwestern Iran Using 18s rRNA Gene Nested-PCR-RFLP and Sequencing Techniques

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Jasem Saki ◽  
Masoud Foroutan-Rad ◽  
Reza Asadpouri
Keyword(s):  
Molecular Characterization ◽  
Nested Pcr ◽  
18S Rrna ◽  
Restriction Enzymes ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Wild Rodents ◽  
Nested Polymerase Chain Reaction ◽  
Pcr Rflp ◽  
Southwest Of Iran

Background. Rodents could act as reservoir forCryptosporidiumspp. speciallyC. parvum, a zoonotic agent responsible for human infections. Since there is no information aboutCryptosporidiuminfection in rodents of Ahvaz city, southwest of Iran, hence, this survey was performed to determine the prevalence and molecular characterization ofCryptosporidiumspp. in this region.Materials and Methods. One hundred rodents were trapped from different regions of Ahvaz city. Intestine contents and fecal specimens of rodents were studied using both microscopy examination to identify oocyst and nested-polymerase chain reaction (PCR) technique for 18s rRNA gene detection. Eventually restriction fragment length polymorphism (RFLP) method usingSspIandVspIrestriction enzymes was carried out to genotype the species and then obtained results were sequenced.Results. Three out of 100 samples were diagnosed as positive and overall prevalence ofCryptosporidiumspp. was 3% using both modified Ziehl-Neelsen staining under light microscope and nested-PCR (830 bp) methods. Afterwards, PCR-RFLP was performed on positive samples andC. parvumpattern was identified. Finally PCR-RFLP findings were sequenced and presence ofC. parvumwas confirmed again.Conclusions. Our study showed rodents could be potential reservoir forC. parvum. So an integrated program for control and combat with them should be adopted and continued.

Bench-scale experiments for the development of a unified loop-mediated isothermal amplification (LAMP) assay for the in vitro diagnosis of Leishmania species' promastigotes

2013 ◽  
Vol 142 (8) ◽  
pp. 1671-1677 ◽  
Author(s):  
M. KARANI ◽  
I. SOTIRIADOU ◽  
J. PLUTZER ◽  
P. KARANIS
Keyword(s):  
Lamp Assay ◽  
High Sensitivity ◽  
Leishmania Species ◽  
18S Rrna Gene ◽  
Isothermal Amplification ◽  
Rrna Gene ◽  
Bench Scale ◽  
Loop Mediated Isothermal Amplification ◽  
Wide Range

SUMMARYWe developed, in bench-scale experiments, a unified loop-mediated isothermal amplification (LAMP) assay for the detection of cutaneous, mucocutaneous and visceral leishmaniasis using DNA of cultivated promastigotes. Two primer sets for the LAMP assay were designed based on the 18S rRNA gene, and their sensitivity and specificity were tested and compared. Both of them were specific for Leishmania as the DNA of all ten Leishmania species tested was amplified, whereas the DNA of other parasites, including that of Trypanosoma, was not. The detection limit for primer set 1 ranged between 30 pg and 3·6 fg, depending on which Leishmania species tested. Primer set 2 showed high sensitivity, but was less sensitive than primer set 1. Our findings lead to the conclusion that the LAMP assay with primer set 1 is a promising and effective assay for the successful detection of a wide range of Leishmania infections using only a unified multiplex LAMP test.

scholarly journals Natural canine infection by Leishmania infantum and Leishmania amazonensis and their implications for disease control

2016 ◽  
Vol 25 (4) ◽  
pp. 465-469 ◽  
Author(s):  
Letícia da Cruz Sanches ◽  
Cleber Costa de Martini ◽  
Alex Akira Nakamura ◽  
Maria Emília Bodini Santiago ◽  
Beatriz Dolabela de Lima ◽  
...  
Keyword(s):  
Public Health Problem ◽  
Leishmania Species ◽  
Canine Leishmaniasis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rrna Gene ◽  
Major Public Health Problem ◽  
Pcr Rflp ◽  
New Hosts ◽  
Canine Infection ◽  
Polymerase Chain

Abstract Leishmaniasis is a major public health problem worldwide. Because Leishmania can adapt to new hosts or vectors, knowledge concerning the current etiological agent in dogs is important in endemic areas. This study aimed to identify the Leishmania species detected in 103 samples of peripheral blood from dogs that were naturally infected with these protozoa. The diagnosis of leishmaniasis was determined through parasitological examination, the indirect enzyme-linked immunosorbent assay (ELISA) and the polymerase chain reaction (PCR). The Leishmania species were identified by means of PCR-restriction fragment length polymorphism (PCR-RFLP). The samples were subjected to PCR using oligonucleotide primers that amplify the intergenic region ITS1 of the rRNA gene in order to identify the species. The amplified DNA was digested using the restriction enzyme HaeIII. A restriction profile identical to L. amazonensis was shown in 77/103 samples and the profile was similar to L. infantum in 17/103. However, a mixed profile was shown in 9/103 samples, which impeded species identification. In conclusion, the infection in these dogs was predominantly due to L. amazonensis, thus indicating that diagnosing of cases of canine leishmaniasis needs to be reexamined, since the causative agent identified is not restricted to L. infantum.

scholarly journals PCR-RFLP: a targeted method to reveal host specific malacosporean infection profiles (Cnidaria: Myxozoa: Malacosporea)

2020 ◽  
Vol 141 ◽  
pp. 91-101
Author(s):  
P Ruggeri ◽  
J Naldoni ◽  
H Hartikainen ◽  
B Okamura
Keyword(s):  
Sanger Sequencing ◽  
Sequence Data ◽  
Pcr Amplification ◽  
18S Rrna Gene ◽  
Single Step ◽  
Rrna Gene ◽  
Mixed Infections ◽  
Intermediate Hosts ◽  
Tissue Samples ◽  
Pcr Rflp

Malacosporeans are a group of endoparasitic cnidarians (Myxozoa) that use freshwater bryozoans and fish as final and intermediate hosts, respectively. The malacosporean Tetracapsuloides bryosalmonae causes proliferative kidney disease (PKD), an emerging disease in aquaculture and wild fish populations, including threatened salmonids in Europe and the USA. Mixed infections of malacosporeans are often encountered, and a monitoring tool for screening of multiple malacosporean species in either their fish or bryozoan hosts is therefore desirable. We describe an inexpensive method that combines PCR amplification of the partial 18S rRNA gene (~260 bp) and a single-step restriction fragment length polymorphism (RFLP) method for identification of 10 malacosporean lineages and species. We demonstrate and test this methodology on a set of DNA extracted from malacosporeans infecting fish kidney and tissues sampled from bryozoan colonies and compare the results with Sanger sequencing of the same parasite DNA isolates. The PCR-RFLP and Sanger sequencing methods agreed in 100% of cases. The PCR-RFLP method offers a number of opportunities, including screening large panels of host tissue samples to gain insights into infection patterns, characterizing mixed infections, and confirming highly pathogenic T. bryosalmonae infections. The method can also be further refined as new sequence data become available for malacosporeans.

scholarly journals Genetic and Antigenic Evidence Supports the Separation of Hepatozoon canis and Hepatozoon americanum at the Species Level

2000 ◽  
Vol 38 (3) ◽  
pp. 1298-1301 ◽  
Author(s):  
Gad Baneth ◽  
John R. Barta ◽  
Varda Shkap ◽  
Donald S. Martin ◽  
Douglass K. Macintire ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
18S Rrna ◽  
Distinct Species ◽  
18S Rrna Gene ◽  
Species Level ◽  
Hepatozoon Canis ◽  
Rrna Gene ◽  
Western Immunoblotting ◽  
Hepatozoon Americanum

Recognition of Hepatozoon canis and Hepatozoon americanum as distinct species was supported by the results of Western immunoblotting of canine anti-H. canis and anti-H. americanum sera against H. canisgamonts. Sequence analysis of 368 bases near the 3′ end of the 18S rRNA gene from each species revealed a pairwise difference of 13.59%.

scholarly journals 6 Molecular typing of sand fly species by PCR-RFLP of 18S rRNA gene(Proceedings of the 63rd Annual Meeting of Western Region)

2009 ◽  
Vol 60 (2) ◽  
pp. 157
Author(s):  
H. Kato ◽  
Y. Terayama ◽  
A. Gomez Eduardo ◽  
H. Uezato ◽  
Calvopina Manuel ◽  
...  
Keyword(s):  
Annual Meeting ◽  
Molecular Typing ◽  
18S Rrna ◽  
18S Rrna Gene ◽  
Sand Fly ◽  
Rrna Gene ◽  
Western Region ◽  
Pcr Rflp
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