Detection and molecular identification of Hepatozoon canis and Babesia vogeli from domestic dogs in Palestine

SUMMARYDogs serve as hosts for a great number of parasites, which may affect their health and wellbeing. This study aimed to observe tick borne pathogens in dogs from Palestine including Hepatozoon canis and Babesia species. The prevalence of both H. canis and Babesia species infections in apparently healthy dogs, from ten districts of the West Bank was surveyed. DNA was extracted from blood samples obtained from dogs (n = 362) and ticks (n = 213) collected from dogs (n = 77). A primer set that amplifies a partial sequence of the Babesia and Hepatozoon 18S rRNA gene was used for PCR and the DNA sequences of the PCR products of all samples were determined. Twenty-nine (8·0%) of the dogs were found infected including 20 with H. canis (5·5%), seven with Babesia vogeli (1·9%) and two with undefined Babesia spp. (0·6%). Twelve Rhipicephalus sanguineus s.l ticks were pathogen-positive, including ten with H. canis (4·7%), one with B. vogeli (0·5%), and one with Hepatozoon felis (0·5%). The results indicated that a wide range of tick borne pathogens is circulating in the canine population in the surveyed region. This study is the first report on the prevalence of H. canis, B. vogeli and Babesia spp. in dogs in Palestine and its results will assist in the management of diseases associated with these blood parasites.

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Species-specific duplex PCR for the detection of Pratylenchus penetrans

Nematology ◽

10.1163/156854109x428016 ◽

2009 ◽

Vol 11 (6) ◽

pp. 847-857 ◽

Cited By ~ 21

Author(s):

Lieven Waeyenberge ◽

Nicole Viaene ◽

Maurice Moens

Keyword(s):

Internal Control ◽

Dna Sequences ◽

Rrna Gene ◽

Duplex Pcr ◽

Specific Primers ◽

5.8S Rrna Gene ◽

5.8S Rrna ◽

Wide Range ◽

Pcr Products ◽

Species Specific

Abstract ITS1, the 5.8S rRNA gene and ITS2 of the rDNA region were sequenced from 20 different Pratylenchus species. Additionally, the same region was sequenced from seven populations of P. penetrans. After purifying, cloning and sequencing the PCR products, all sequences were aligned in order to find unique sites suitable for the design of species-specific primers for P. penetrans. Since ITS regions showed variability between and even within populations of P. penetrans, only three small DNA sequences were suitable for the construction of three potentially useful species-specific primers. New species-specific primers were paired with existing universal ITS primers and tested in all possible primer combinations. The best performing primer set, supplemented with a universal 28S rDNA primer set that served as an internal control, was tested in duplex PCR. The ideal annealing temperature, Mg2+ concentration and primer ratios were then determined for the most promising primer set. The optimised duplex PCR was subsequently tested on a wide range of different Pratylenchus spp. and 25 P. penetrans populations originating from all over the world. To test the sensitivity, the duplex PCR was conducted on DNA extracted from a single P. penetrans nematode mixed with varying amounts of nematodes belonging to another Pratylenchus species. Results showed that a reliable and sensitive P. penetrans species-specific duplex PCR was constructed.

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Emergence of Babesia Conradae Infection in Coyote-hunting Greyhounds in Oklahoma, USA

10.21203/rs.3.rs-154040/v1 ◽

2021 ◽

Author(s):

Erin M Stayton ◽

Megan Lineberry ◽

Jennifer Thomas ◽

Tina Bass ◽

Kelly Allen ◽

...

Keyword(s):

18S Rrna ◽

Clinical Signs ◽

18S Rrna Gene ◽

Post Treatment ◽

Rrna Gene ◽

Babesia Gibsoni ◽

Wide Range ◽

Pcr Products ◽

Life Threatening ◽

Tick Vectors

Abstract Background: Babesia species are intraerythrocytic Apicomplexan parasites that infect a wide range of vertebrate hosts. These pathogens are typically transmitted either by tick vectors or by direct blood-to-blood contact, and may cause life-threatening clinical disease such as thrombocytopenia, hemolytic anemia, and acute renal failure in canine hosts. While Babesia vogeli and Babesia gibsoni infections have both been reported in Oklahoma, reports of B. conradae infections have been limited to California. Methods: Whole blood samples were collected in EDTA tubes from all dogs in four separate kennels in Oklahoma. DNA was extracted from each blood sample and a nested PCR was performed using general Apicomplexan primers for the partial 18S rRNA gene. PCR products were electrophoresed in agarose matrix and appropriately sized amplicons were sequenced. Sequences were compared to reference 18S rRNA sequences available in GenBank, and samples with >98% homology to B. conradae (GenBank MK256976) were considered positive. B. conradae positive dogs were then treated with atovaquone (13.5 mg/kg TID) and azithromycin (10 mg/kg SID) for 10 days and retested at 30 and 60 days post treatment by PCR. Results: Fifteen of 40 dogs tested positive for B. conradae with 98–100% sequence homology to B. conradae from California. All positive cases were coyote-hunting Greyhounds. Treatment of clinically ill dogs with atovaquone and azithromycin resulted in complete clinical recovery in clinically ill dogs and all treated dogs had negative follow-up PCR at 30 and 60 days post treatment. Conclusions: Collectively, this study (i) documents the occurrence of B. conradae in Oklahoma, (ii) highlights this pathogen as a differential to be considered when clinical signs are present, and (iii) supports the use of atovaquone and azithromycin as effective treatment in these cases.

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Confirmation of occurrence of Babesia vogeli in a dog in Windhoek, central Namibia

Journal of the South African Veterinary Association ◽

10.4102/jsava.v87i1.1427 ◽

2016 ◽

Vol 87 (1) ◽

Cited By ~ 2

Author(s):

Barend L. Penzhorn ◽

Ilse Vorster ◽

Gernot Redecker ◽

Marinda C. Oosthuizen

Keyword(s):

Rhipicephalus Sanguineus ◽

18S Rrna Gene ◽

Rrna Gene ◽

Babesia Vogeli ◽

Rhipicephalus Sanguineus Sensu Lato ◽

Babesia Rossi ◽

Haemaphysalis Elliptica ◽

Blot Hybridisation ◽

Polymerase Chain ◽

Babesia Spp

Although there is evidence of high seroprevalence of antibodies to Babesia spp. in dogs in central Namibia, clinical babesiosis is rarely diagnosed. Rhipicephalus sanguineus sensu lato, the vector of Babesia vogeli, is common in Namibia while Haemaphysalis elliptica, the vector of the highly virulent but morphologically indistinguishable Babesia rossi, has rarely been recorded, mainly in northern Namibia. On the basis of vector occurrence, clinical cases of canine babesiosis in Windhoek, central Namibia, have been ascribed to B. vogeli. DNA extracted from a blood smear made from a sick dog was subjected to the reverse line blot hybridisation assay. The polymerase chain reaction amplicons hybridised with the B. vogeli–specific probe, but not with the Babesia canis– and B. rossi–specific probes. Although attempts at cloning and sequencing of the full-length 18S rRNA gene were unsuccessful, we can confirm that B. vogeli occurs in central Namibia.

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Stenamoeba dejonckheerei sp. nov., a Free-Living Amoeba Isolated from a Thermal Spring

Pathogens ◽

10.3390/pathogens9070586 ◽

2020 ◽

Vol 9 (7) ◽

pp. 586

Author(s):

Manuel Alejandro Borquez-Román ◽

Luis Fernando Lares-Jiménez ◽

Libia Zulema Rodriguez-Anaya ◽

Jose Reyes Gonzalez-Galaviz ◽

Paul A. Fuerst ◽

...

Keyword(s):

Thermal Spring ◽

Small Subunit ◽

18S Rrna Gene ◽

Rrna Gene ◽

Likelihood Analysis ◽

Ribosomal Ribonucleic Acid ◽

Pcr Products ◽

Northwestern Mexico ◽

Rrna Gene Sequencing ◽

A New Species

Two amoeboid organisms were obtained from water samples taken from a thermal spring, "Agua Caliente", in Northwestern Mexico. The isolates were obtained when samples were cultivated at 37 °C on non-nutrient agar coated with Escherichia coli. The initial identification of the isolates was performed morphologically using light microscopy. The samples were found to have trophozoite morphology consistent with members of the genus Stenamoeba, a genus derived in 2007 from within the abolished polyphyletic genus Platyamoeba. Further analysis was performed by sequencing PCR products obtained using universal eukaryotic primers for the small subunit ribosomal ribonucleic acid (SSU rRNA) gene. Sequencing primers were designed to allow the comparison of the 18S rRNA gene sequences of the new isolates with previous sequences reported for Stenamoeba. Phylogenetic relationships among sequences from Stenamoeba were determined using Maximum Likelihood analysis. The results showed the two "Agua Caliente" sequences to be closely related, while clearly separating them from those of other Stenamoeba taxa. The degrees of sequence differentiation from other taxa were considered sufficient to allow us to propose that the Mexican isolates represent a new species.

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Molecular survey of Babesia vogeli and Hepatozoon species in dogs from urban area of Midwestern Brazil

Semina Ciências Agrárias ◽

10.5433/1679-0359.2019v40n3p1357 ◽

2019 ◽

Vol 40 (3) ◽

pp. 1357

Author(s):

Maerle Oliveira Maia ◽

André Luís Santos de Freitas ◽

Jamila Guimarães Santos ◽

Thábata dos Anjos Pacheco ◽

Dirceu Guilherme de Souza Ramos ◽

...

Keyword(s):

Dna Sequences ◽

Urban Areas ◽

Ixodid Ticks ◽

Hepatozoon Canis ◽

Molecular Survey ◽

Rdna Sequences ◽

Babesia Vogeli ◽

18S Rdna Sequences ◽

Polymerase Chain ◽

Hepatozoon Species

In Brazil, the most important tickborne pathogens affecting dogs include Babesia vogeli, Ehrlichia canis, Anaplasma platys, Hepatozoon canis, and Mycoplasma haemocanis. Babesia spp. and Hepatozoon spp., transmitted by ixodid ticks, have been reported to naturally infect dogs and are widespread. The authors aimed to investigate the incidence of B. vogeli and Hepatozoon spp. infection using molecular methods to identify factors associated with the infection in dogs from urban areas of Cuiabá municipality, Midwestern Brazil. Polymerase chain reaction (PCR) assay revealed a prevalence of 9.36% (Confidence Interval-CI 95%; 2.72%; 6.79%) and 9.61% (CI 95%; 7.0%; 13.0%) among dogs for B. vogeli and Hepatozoon, respectively. DNA sequences obtained from 10 Hepatozoon PCR positive samples were sequenced and were identical to one another and, moreover, were 100% (541/541 base of pairs-bp) homologous to the corresponding 18S rDNA sequences of H. canis. Twenty-five dogs (6.02%) generated amplicons using PCR protocols for both organisms, indicating co-infection by these two protozoans. To the best of our knowledge, our study was the first molecular survey to consider the entire population of dogs from the study area. Moreover, young dogs (0-12 months of age), as well as animals living in walled houses?without access to the street?were more susceptible to infection with B. vogeli and H. canis, respectively.

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Confirmatory Evidence from 18S rRNA Gene Analysis for In Vivo Development of Propamidine Resistance in a Temporal Series ofAcanthamoeba Ocular Isolates from a Patient

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.8.2144 ◽

1998 ◽

Vol 42 (8) ◽

pp. 2144-2145 ◽

Cited By ~ 10

Author(s):

Dolena R. Ledee ◽

David V. Seal ◽

Thomas J. Byers

Keyword(s):

Mixed Infection ◽

Dna Sequences ◽

18S Rrna ◽

18S Rrna Gene ◽

Gene Analysis ◽

Rrna Gene ◽

Temporal Series ◽

Before And After

ABSTRACT DNA sequences of three 18S rRNA gene alleles present in trophozoites obtained before and after therapy forAcanthamoeba keratitis substantiate a previous report that the infection was due to a single Acanthamoeba strain. Thus, the possibility that propamidine resistance which developed during therapy was due to a mixed infection was ruled out.

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Molecular identification ofSarcocystisspp. helped to define the origin of green pythons (Morelia viridis) confiscated in Germany

Parasitology ◽

10.1017/s0031182013001960 ◽

2013 ◽

Vol 141 (5) ◽

pp. 646-651 ◽

Cited By ~ 9

Author(s):

GASTÓN MORÉ ◽

NIKOLA PANTCHEV ◽

DALAND C. HERRMANN ◽

MAJDA GLOBOKAR VRHOVEC ◽

SABINE ÖFNER ◽

...

Keyword(s):

18S Rrna ◽

Sequence Data ◽

18S Rrna Gene ◽

Rrna Genes ◽

Rrna Gene ◽

Intermediate Hosts ◽

Apicomplexan Parasites ◽

Large Numbers ◽

Wide Range ◽

18S Rrna Genes

SUMMARYSarcocystisspp. represent apicomplexan parasites. They usually have a heteroxenous life cycle. Around 200 species have been described, affecting a wide range of animals worldwide, including reptiles. In recent years, large numbers of reptiles have been imported into Europe as pets and, as a consequence, animal welfare and species protection issues emerged. A sample of pooled feces from four confiscated green pythons (Morelia viridis) containingSarcocystisspp. sporocysts was investigated. These snakes were imported for the pet trade and declared as being captive-bred. Full length 18S rRNA genes were amplified, cloned into plasmids and sequenced. Two differentSarcocystisspp. sequences were identified and registered asSarcocystissp. fromM. viridisin GenBank. Both showed a 95–97% sequence identity with the 18S rRNA gene ofSarcocystis singaporensis.Phylogenetic analysis positioned these sequences together with otherSarcocystisspp. from snakes and rodents as definitive and intermediate hosts (IH), respectively. Sequence data and also the results of clinical and parasitological examinations suggest that the snakes were definitive hosts forSarcocystisspp. that circulate in wild IH. Thus, it seems unlikely that the infected snakes had been legally bred. Our research shows that information on the infection of snakes withSarcocystisspp. may be used to assess compliance with regulations on the trade with wildlife species.

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

Revista Brasileira de Parasitologia Veterinária ◽

10.1590/s1984-29612016065 ◽

2016 ◽

Vol 25 (4) ◽

pp. 450-458 ◽

Cited By ~ 6

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.

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Specific PCR detection of Peptostreptococcus magnus

Journal of Medical Microbiology ◽

10.1099/jmm.0.05004-0 ◽

2003 ◽

Vol 52 (4) ◽

pp. 309-313 ◽

Cited By ~ 9

Author(s):

M.P. Riggio ◽

A. Lennon

Keyword(s):

Pcr Primers ◽

Oral Bacteria ◽

Rrna Gene ◽

Subgingival Plaque ◽

Pcr Assay ◽

Clinical Specimens ◽

Specific Pcr ◽

Wide Range ◽

Pcr Products ◽

Peptostreptococcus Magnus

Peptostreptococcus magnus is the most pathogenic and one of the most common Gram-positive anaerobic cocci found in human clinical specimens. The organism has been isolated in pure culture from a range of serious infections, including meningitis and endocarditis. However, isolation of Peptostreptococcus magnus from the oral cavity has rarely been attempted. Identification of Peptostreptococcus magnus in clinical specimens is reliant upon microbiological culture and biochemical methods, which often give ambiguous results. The aim of this study was to develop a PCR assay for the specific detection of Peptostreptococcus magnus in oral clinical specimens. PCR primers specific for Peptostreptococcus magnus DNA were derived by comparison of 16S rRNA gene sequences and selection of primers that demonstrated specificity at their 3′ ends for Peptostreptococcus magnus. PCR positivity for Peptostreptococcus magnus DNA was indicated by the amplification of a 553 bp product. The PCR assay was then used to attempt detection of Peptostreptococcus magnus DNA in subgingival plaque samples from adult periodontitis patients and pus aspirates from subjects with acute dento-alveolar abscesses. The PCR assay was demonstrated to be highly specific for Peptostreptococcus magnus DNA, since no PCR products were obtained when genomic DNA from a wide range of other oral bacteria, including closely related Peptostreptococcus species, was used in the PCR assay. Confirmation of specific amplification of Peptostreptococcus magnus DNA was obtained by digestion of PCR products with the restriction endonuclease RsaI, which gives a unique restriction profile for Peptostreptococcus magnus. Of the 33 subgingival plaque samples analysed, 2 (6 %) were positive for Peptostreptococcus magnus DNA. None of the 60 pus aspirates analysed was positive for Peptostreptococcus magnus DNA. It is concluded that Peptostreptococcus magnus is not a major pathogen in adult periodontitis or dento-alveolar abscesses. The PCR assay provides a more rapid, specific and sensitive alternative to conventional methods for identification of Peptostreptococcus magnus in clinical specimens.

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Phylogenetic analysis based on 18S rRNA gene sequences ofSchellackiaparasites (Apicomplexa: Lankesterellidae) reveals their close relationship to the genusEimeria

Parasitology ◽

10.1017/s0031182013000553 ◽

2013 ◽

Vol 140 (9) ◽

pp. 1149-1157 ◽

Cited By ~ 20

Author(s):

R. MEGÍA-PALMA ◽

J. MARTÍNEZ ◽

S. MERINO

Keyword(s):

Phylogenetic Analysis ◽

Blood Cells ◽

18S Rrna ◽

Taxonomic Status ◽

Refractile Body ◽

18S Rrna Gene ◽

Blood Parasites ◽

Recent Common Ancestor ◽

Rrna Gene ◽

Amphibian Species

SUMMARYIn the present study we detectedSchellackiahaemoparasites infecting the blood cells ofLacerta schreiberiandPodarcis hispanica, two species of lacertid lizards from central Spain. The parasite morphometry, the presence of a refractile body, the type of infected blood cells, the kind of host species, and the lack of oocysts in the fecal samples clearly indicated these blood parasites belong to the genusSchellackia. Until now, the species of this genus have never been genetically characterized and its taxonomic position under the Lankesterellidae family is based on the lack of the exogenous oocyst stage. However, the phylogenetic analysis performed on the basis of the 18S rRNA gene sequence revealed that species of the genusSchellackiaare clustered withEimeriaspecies isolated from a snake and an amphibian species but not withLankesterellaspecies. The phylogenetic analysis rejects that both genera share a recent common ancestor. Based on these results we suggest a revision of the taxonomic status of the family Lankesterellidae.

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