The Experts below are selected from a list of 57 Experts worldwide ranked by ideXlab platform
Selwyn Arlington Headley - One of the best experts on this subject based on the ideXlab platform.
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Neorickettsia helminthoeca associated lymphoid, enteric, and pulmonary lesions in dogs from Southern Brazil: An immunohistochemical study
Transboundary and Emerging Diseases, 2019Co-Authors: Selwyn Arlington Headley, Nayara Emily Viana, Mariana De Mello Zanim Michelazzo, Ana Aparecida Correa Xavier, Cícero Júlio Silva Costa, Felipe Pinho, Marcelo Diniz Dos SantosAbstract:Neorickettsia helminthoeca (NH), the agent of Salmon Poisoning Disease or canine neorickettiosis (CN), is a bacterial endosymbiont of the nematode Nanophyetus salmincola, and infections are spreading among specific fish-eating mammalians. This article describes the pathologic and immunohistochemical findings associated with spontaneous NH-induced infections in dogs from Southern Brazil. The principal pathologic findings were hypertrophy of Peyer patches and lymphadenopathy with lymphocytic proliferation, chronic interstitial pneumonia, and chronic enteritis associated with positive intralesional immunoreactivity to antigens of NH within macrophages and histiocytes. Positive immunoreactivity against canine parvovirus-2 (CPV-2) or/and canine distemper virus was not detected in the evaluated intestinal segments or in the samples from the cerebellum and lungs, respectively, from the dogs evaluated. These findings demonstrated that NH was involved in the enteric, pulmonary, and lymphoid lesions herein described, and provide additional information to confirm the occurrence of this bacterial endosymbiont within this geographical location. It is proposed that chronic pneumonia should be considered as a pathologic manifestation of NH-induced infections. Additionally, our results show that the occurrences of CN seem to be underdiagnosed in Southern Brazil due to the confusion with the incidence of CPV-2.
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Neorickettsia helminthoeca and Salmon Poisoning Disease: A review
Veterinary journal (London England : 1997), 2009Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen DumlerAbstract:Neorickettsia helminthoeca is an obligate intra-cytoplasmic bacterium that causes Salmon Poisoning Disease (SPD), an acute, febrile, fatal Disease of dogs. The complex life-cycle of this pathogen involves stages in an intestinal fluke (Nanophyetus salmincola), a river snail (Oxytrema silicula), in fish, and in fish-eating mammals. This complexity has created confusion with respect to the various bacterial and parasitic infections associated with the Disease and its significance in dogs in specific geographical locations has likely to have previously been under-estimated. This paper addresses the history, taxonomy, microbiology of N. helminthoeca and summarises the pathogenesis, clinical signs and pathological features associated with infection. Furthermore, the biological cycles, treatment, control, and both public and veterinary health impacts associated with this pathogen and the intestinal fluke N. salmincola are discussed.
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Neorickettsia helminthoeca in Brazilian dogs: a cytopathological, histopathological and immunohistochemical study.
Clinical Microbiology and Infection, 2009Co-Authors: Selwyn Arlington Headley, Ana Paula Frederico Rodrigues Loureiro Bracarense, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Flora Satiko Kano, Katia Tamekuni, J S DumlerAbstract:Pathology, The Johns Hopkins University School of Medicine, Baltimore, MD,USAINTRODUCTIONNeorickettsia helminthoeca is an intracellular bacte-rium of reticuloendothelial cells of dogs thatcauses Salmon Poisoning Disease (SPD), whichwas considered endemic only to specific geo-2graphical locations of North America [1–3].However, 20 mature dogs demonstrated lesionsconsistent with this infection in Southern Braziland intracytoplasmic and intralesional neorickett-sial bodies were identified within reticuloendo-thelial cells by Giemsa stain [4]. Additionally, bymolecular biology methods, it was shown that themesenteric lymph node of one dog containedDNA that was similar to N. helminthoeca as knownin the USA [5].This article describes the cytological, histopath-ological and immunohistochemical findings asso-ciated with N. helminthoeca in additional casesfrom Brazil.MATERIALS AND METHODS
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Neorickettsia helminthoeca in dog, Brazil.
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, J. Stephen DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (
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neorickettsia helminthoeca in dog brazil
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Stephen J. DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (<380/1,000 iterations) below the genus level for the short rrs region examined. However, both the short rpoB and groESL regions examined had high bootstrap values (941/1,000 and 995/1,000 iterations, respectively). This finding allowed differentiation of N. helminthoeca and the Brazilian dog strain from N. sennetsu, N. risticii, and other related Anaplasmataceae and provided a high degree of confidence in the identification. More work is being implemented to obtain longer sequences to confirm and extend these genotypic comparisons. We propose further study to isolate the pathogen from other dogs for comparative biologic analyses. Although SPD is caused by N. helminthoeca, infections by other Neorickettsia spp., including N. risticii (Potomac horse fever) and N. sennetsu (sennetsu fever), illustrate the potential of these widely distributed species to infect and cause Disease in mammals and humans. Detection of N. helminthoeca in Brazilian dogs extends the range of this species and warrants a broad search for infections and spectrum of Disease of Neorickettsia in animals and humans.
Diana G. Scorpio - One of the best experts on this subject based on the ideXlab platform.
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Neorickettsia helminthoeca and Salmon Poisoning Disease: A review
Veterinary journal (London England : 1997), 2009Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen DumlerAbstract:Neorickettsia helminthoeca is an obligate intra-cytoplasmic bacterium that causes Salmon Poisoning Disease (SPD), an acute, febrile, fatal Disease of dogs. The complex life-cycle of this pathogen involves stages in an intestinal fluke (Nanophyetus salmincola), a river snail (Oxytrema silicula), in fish, and in fish-eating mammals. This complexity has created confusion with respect to the various bacterial and parasitic infections associated with the Disease and its significance in dogs in specific geographical locations has likely to have previously been under-estimated. This paper addresses the history, taxonomy, microbiology of N. helminthoeca and summarises the pathogenesis, clinical signs and pathological features associated with infection. Furthermore, the biological cycles, treatment, control, and both public and veterinary health impacts associated with this pathogen and the intestinal fluke N. salmincola are discussed.
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Neorickettsia helminthoeca in Brazilian dogs: a cytopathological, histopathological and immunohistochemical study.
Clinical Microbiology and Infection, 2009Co-Authors: Selwyn Arlington Headley, Ana Paula Frederico Rodrigues Loureiro Bracarense, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Flora Satiko Kano, Katia Tamekuni, J S DumlerAbstract:Pathology, The Johns Hopkins University School of Medicine, Baltimore, MD,USAINTRODUCTIONNeorickettsia helminthoeca is an intracellular bacte-rium of reticuloendothelial cells of dogs thatcauses Salmon Poisoning Disease (SPD), whichwas considered endemic only to specific geo-2graphical locations of North America [1–3].However, 20 mature dogs demonstrated lesionsconsistent with this infection in Southern Braziland intracytoplasmic and intralesional neorickett-sial bodies were identified within reticuloendo-thelial cells by Giemsa stain [4]. Additionally, bymolecular biology methods, it was shown that themesenteric lymph node of one dog containedDNA that was similar to N. helminthoeca as knownin the USA [5].This article describes the cytological, histopath-ological and immunohistochemical findings asso-ciated with N. helminthoeca in additional casesfrom Brazil.MATERIALS AND METHODS
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neorickettsia helminthoeca in dog brazil
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Stephen J. DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (<380/1,000 iterations) below the genus level for the short rrs region examined. However, both the short rpoB and groESL regions examined had high bootstrap values (941/1,000 and 995/1,000 iterations, respectively). This finding allowed differentiation of N. helminthoeca and the Brazilian dog strain from N. sennetsu, N. risticii, and other related Anaplasmataceae and provided a high degree of confidence in the identification. More work is being implemented to obtain longer sequences to confirm and extend these genotypic comparisons. We propose further study to isolate the pathogen from other dogs for comparative biologic analyses. Although SPD is caused by N. helminthoeca, infections by other Neorickettsia spp., including N. risticii (Potomac horse fever) and N. sennetsu (sennetsu fever), illustrate the potential of these widely distributed species to infect and cause Disease in mammals and humans. Detection of N. helminthoeca in Brazilian dogs extends the range of this species and warrants a broad search for infections and spectrum of Disease of Neorickettsia in animals and humans.
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Neorickettsia helminthoeca in dog, Brazil.
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, J. Stephen DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (
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Suspected cases of Neorickettsia-like organisms in Brazilian dogs.
Annals of the New York Academy of Sciences, 2004Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen Dumler, Joseph L. MankowskiAbstract:Preliminary findings of gross and histopathological lesions consistent with Salmon Poisoning Disease in 10 dogs from southern Brazil are described. Lesions were restricted to the spleen, lymph nodes, and intestinal lymphoid tissues. Grossly, there was marked hyperplasia of mesenteric lymph nodes and Peyer's patches. Microscopic alterations were characterized by diffuse hyperplasia of intestinal lymphoid tissues and Peyer's patches. Intracytoplasmic organisms consistent with Neorickettsia helminthoeca were demonstrated by Giemsa stain in reticuloendothelial cells of the intestine, spleen, Peyer's patches, and lymph nodes. We have named this organism Neorickettsia helminthoeca-like because of its marked similarity with the agent described in the United States.
Odilon Vidotto - One of the best experts on this subject based on the ideXlab platform.
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Neorickettsia helminthoeca and Salmon Poisoning Disease: A review
Veterinary journal (London England : 1997), 2009Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen DumlerAbstract:Neorickettsia helminthoeca is an obligate intra-cytoplasmic bacterium that causes Salmon Poisoning Disease (SPD), an acute, febrile, fatal Disease of dogs. The complex life-cycle of this pathogen involves stages in an intestinal fluke (Nanophyetus salmincola), a river snail (Oxytrema silicula), in fish, and in fish-eating mammals. This complexity has created confusion with respect to the various bacterial and parasitic infections associated with the Disease and its significance in dogs in specific geographical locations has likely to have previously been under-estimated. This paper addresses the history, taxonomy, microbiology of N. helminthoeca and summarises the pathogenesis, clinical signs and pathological features associated with infection. Furthermore, the biological cycles, treatment, control, and both public and veterinary health impacts associated with this pathogen and the intestinal fluke N. salmincola are discussed.
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Neorickettsia helminthoeca in Brazilian dogs: a cytopathological, histopathological and immunohistochemical study.
Clinical Microbiology and Infection, 2009Co-Authors: Selwyn Arlington Headley, Ana Paula Frederico Rodrigues Loureiro Bracarense, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Flora Satiko Kano, Katia Tamekuni, J S DumlerAbstract:Pathology, The Johns Hopkins University School of Medicine, Baltimore, MD,USAINTRODUCTIONNeorickettsia helminthoeca is an intracellular bacte-rium of reticuloendothelial cells of dogs thatcauses Salmon Poisoning Disease (SPD), whichwas considered endemic only to specific geo-2graphical locations of North America [1–3].However, 20 mature dogs demonstrated lesionsconsistent with this infection in Southern Braziland intracytoplasmic and intralesional neorickett-sial bodies were identified within reticuloendo-thelial cells by Giemsa stain [4]. Additionally, bymolecular biology methods, it was shown that themesenteric lymph node of one dog containedDNA that was similar to N. helminthoeca as knownin the USA [5].This article describes the cytological, histopath-ological and immunohistochemical findings asso-ciated with N. helminthoeca in additional casesfrom Brazil.MATERIALS AND METHODS
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neorickettsia helminthoeca in dog brazil
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, Stephen J. DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (<380/1,000 iterations) below the genus level for the short rrs region examined. However, both the short rpoB and groESL regions examined had high bootstrap values (941/1,000 and 995/1,000 iterations, respectively). This finding allowed differentiation of N. helminthoeca and the Brazilian dog strain from N. sennetsu, N. risticii, and other related Anaplasmataceae and provided a high degree of confidence in the identification. More work is being implemented to obtain longer sequences to confirm and extend these genotypic comparisons. We propose further study to isolate the pathogen from other dogs for comparative biologic analyses. Although SPD is caused by N. helminthoeca, infections by other Neorickettsia spp., including N. risticii (Potomac horse fever) and N. sennetsu (sennetsu fever), illustrate the potential of these widely distributed species to infect and cause Disease in mammals and humans. Detection of N. helminthoeca in Brazilian dogs extends the range of this species and warrants a broad search for infections and spectrum of Disease of Neorickettsia in animals and humans.
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Neorickettsia helminthoeca in dog, Brazil.
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, J. Stephen DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (
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Suspected cases of Neorickettsia-like organisms in Brazilian dogs.
Annals of the New York Academy of Sciences, 2004Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen Dumler, Joseph L. MankowskiAbstract:Preliminary findings of gross and histopathological lesions consistent with Salmon Poisoning Disease in 10 dogs from southern Brazil are described. Lesions were restricted to the spleen, lymph nodes, and intestinal lymphoid tissues. Grossly, there was marked hyperplasia of mesenteric lymph nodes and Peyer's patches. Microscopic alterations were characterized by diffuse hyperplasia of intestinal lymphoid tissues and Peyer's patches. Intracytoplasmic organisms consistent with Neorickettsia helminthoeca were demonstrated by Giemsa stain in reticuloendothelial cells of the intestine, spleen, Peyer's patches, and lymph nodes. We have named this organism Neorickettsia helminthoeca-like because of its marked similarity with the agent described in the United States.
Yasuko Rikihisa - One of the best experts on this subject based on the ideXlab platform.
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Characterization of the SF agent, an Ehrlichia sp. isolated from the fluke Stellantchasmus falcatus, by 16S rRNA base sequence, serological, and morphological analyses.
International Journal of Systematic and Evolutionary Microbiology, 1996Co-Authors: Bohai Wen, Yasuko Rikihisa, Sheigo Yamamoto, Norihiko Kawabata, Paul A. FuerstAbstract:The organism designated the SF agent was originally isolated in Japan in 1962 from Stellantchasmus falcatus metacercaria parasitic on gray mullet fish. The SF agent resembles members of the genus Ehrlichia morphologically and exhibits weak antigenic cross-reactivity with Ehrlichia sennetsu. This organism causes mild clinical signs in dogs, but severe splenomegaly and lymphadenopathy in mice. This suggests that the SF agent may be similar to either Neorickettsia helminthoeca, an intracellular parasite of a fluke and the cause of Salmon Poisoning Disease in dogs, or E. sennetsu, the causative agent of human sennetsu ehrlichiosis in Japan and Malaysia. In order to determine the phylogenetic relationship between the SF agent and other ehrlichial species, the 16S rRNA gene was amplified by the PCR and sequenced. The SF agent sequence was most closely related to the sequences of Ehrlichia risticii (level of sequence similarity, 99.1%), the causative agent of Potomac horse fever, and E. sennetsu (level of sequence similarity, 98.7%). The next most similar sequence was that of N. helminthoeca, but the level of sequence similarity was only 93.7%. E. sennetsu, E. risticii, the SF agent, and N. helminthoeca formed a distinct cluster that was separated from all other ehrlichial species. As determined by immunofluorescence labeling, antiserum against the SF agent cross-reacted strongly with E. sennetsu, E. risticii, and N. helminthoeca. When three genetically distinct ehrlichial isolates obtained from horses with Potomac horse fever were compared with the SF agent, we found that the SF agent was most closely related to Ohio isolate 081, followed by IllinoisT (T = type strain) and a Kentucky isolate. We observed strong antigenic cross-reactivities and similarities in Western blot (immunoblot) reaction profiles when we compared the SF agent, E. risticii, and E. sennetsu; however, weaker antigenic cross-reactivity was observed when the SF agent and N. helminthoeca were compared. Our results indicate that the SF agent is antigenically more closely related to E. risticii and E. sennetsu than to N. helminthoeca. The biological and antigenic characteristics and the 16S rRNA sequence data suggest that the SF agent is a new species that belongs to the genus Ehrlichia.
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16S rRNA Gene Sequence of Neorickettsia helminthoeca and Its Phylogenetic Alignment with Members of the Genus Ehrlichia
International journal of systematic bacteriology, 1995Co-Authors: Charles I. Pretzman, Paul A. Fuerst, David Ralph, Diane R. Stothard, Yasuko RikihisaAbstract:Neorickettsia helminthoeca (tribe Ehrlichieae, family Rickettsiaceae) is the agent of Salmon Poisoning Disease, which affects members of the family Canidae. This bacterium is unusual in that it is the only known obligately intracellular bacterium that is transmitted via a helminth vector. The nucleotide sequence of the N. helminthoeca 16S rRNA gene was determined and compared with the sequences of intracellular bacteria belonging to the alpha subgroup of the Proteobacteria. The N. helminthoeca sequence was most similar to the sequences of two Ehrlichia species, Ehrlichia risticii and Ehrlichia sennetsu (levels of sequence similarity, >95%). All other members of the tribe Ehrlichieae, including members of the other Ehrlichia species, and the related species Cowdria ruminantium and Anaplasma marginale, were only distantly related phylogenetically (levels of sequence similarity, 84 to 86%). Our results corroborate the results of previous ultrastructural and Western blot (immunoblot) comparisons of N. helminthoeca with other ehrlichial species. The genus Ehrlichia is phylogenetically incoherent and can be separated into three identifiable clusters of species. Each cluster is closely associated with a species classified in another non-Ehrlichia bacterial genus. The close relationships among N. helminthoeca, E. risticii, and E. sennetsu and the striking differences between these organisms and other members of the tribe Ehrlichieae suggest that in the future, these organisms should be treated as members of a new bacterial genus separate from the genus Ehrlichia.
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Isolation andContinuous Culture ofNeorickettsia helminthoeca inaMacrophage CellLinet
1991Co-Authors: Yasuko Rikihisa, Harold Stills, Andgary ZimmermanAbstract:kidneys ofSalmon caught in ariver whereinfection ofmetacercariae withNeorickettsia helminthoeca hasbeencircumstantially knownfor decades. Clinically, theclassic course ofSalmon Poisoning Disease developed inallofthedogs. Parasitemia began onday8to11postinfection, whenthedogsdeveloped afebrile peak, andcontinued until euthanasia. At necropsy, characteristic gross andmicroscopic lesions oftheDisease werepresent. ANeorickettsia sp.wasalso isolated fromliver andspleen samples ofthese animals. Theisolates havebeencontinuously propagated and passed inDH82cells formorethan6months. Electron microscopic examination confirmed that therickettsial organisms multiplied inthemembrane-bound compartment ofDH82cells andthattheymorphologically closely resembled rickettsia belonging tothegenusEhrlichia. Anindirect fluorescent antibody test using Neorickettsia organisms cultured inDH82cells showed that alldogsseroconverted 13to15dayspostinfection. Finally, inoculation ofthecell-cultured Neorickettsia organisms into anaive dogreproduced clinically typical Salmon Poisoning Disease whichwasofgreater severity andhadamorerapid timecourse thanthat inthedogs fromwhichtheoriginal isolation wasmade.Onthebasis oftheclinical andpathologic responses ofthedogs inourstudy, webelieve thatvirulent N.helminthoeca wasisolated andcultured inacontinuous cell line.
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Isolation and continuous culture of Neorickettsia helminthoeca in a macrophage cell line.
Journal of clinical microbiology, 1991Co-Authors: Yasuko Rikihisa, Harold Stills, G ZimmermanAbstract:Experimental evidence is presented supporting the development of a system for the isolation and propagation of a Neorickettsia sp. in a continuous canine macrophage cell line (DH82). To isolate a Neorickettsia sp. pathogenic to the canine species, three naive dogs were fed metacercaria-encysted kidneys of Salmon caught in a river where infection of metacercariae with Neorickettsia helminthoeca has been circumstantially known for decades. Clinically, the classic course of Salmon Poisoning Disease developed in all of the dogs. Parasitemia began on day 8 to 11 postinfection, when the dogs developed a febrile peak, and continued until euthanasia. At necropsy, characteristic gross and microscopic lesions of the Disease were present. A Neorickettsia sp. was also isolated from liver and spleen samples of these animals. The isolates have been continuously propagated and passed in DH82 cells for more than 6 months. Electron microscopic examination confirmed that the rickettsial organisms multiplied in the membrane-bound compartment of DH82 cells and that they morphologically closely resembled rickettsia belonging to the genus Ehrlichia. An indirect fluorescent antibody test using Neorickettsia organisms cultured in DH82 cells showed that all dogs seroconverted 13 to 15 days postinfection. Finally, inoculation of the cell-cultured Neorickettsia organisms into a naive dog reproduced clinically typical Salmon Poisoning Disease which was of greater severity and had a more rapid time course than that in the dogs from which the original isolation was made. On the basis of the clinical and pathologic responses of the dogs in our study, we believe that virulent N. helminthoeca was isolated and cultured in a continuous cell line. Images
J. Stephen Dumler - One of the best experts on this subject based on the ideXlab platform.
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Neorickettsia helminthoeca and Salmon Poisoning Disease: A review
Veterinary journal (London England : 1997), 2009Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen DumlerAbstract:Neorickettsia helminthoeca is an obligate intra-cytoplasmic bacterium that causes Salmon Poisoning Disease (SPD), an acute, febrile, fatal Disease of dogs. The complex life-cycle of this pathogen involves stages in an intestinal fluke (Nanophyetus salmincola), a river snail (Oxytrema silicula), in fish, and in fish-eating mammals. This complexity has created confusion with respect to the various bacterial and parasitic infections associated with the Disease and its significance in dogs in specific geographical locations has likely to have previously been under-estimated. This paper addresses the history, taxonomy, microbiology of N. helminthoeca and summarises the pathogenesis, clinical signs and pathological features associated with infection. Furthermore, the biological cycles, treatment, control, and both public and veterinary health impacts associated with this pathogen and the intestinal fluke N. salmincola are discussed.
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Neorickettsia helminthoeca in dog, Brazil.
Emerging Infectious Diseases, 2006Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, Nicole C. Barat, J. Stephen DumlerAbstract:To the Editor: Neorickettsia helminthoeca causes Salmon Poisoning Disease (SPD) in canids. SPD has been described only in the United States and the northwestern Pacific region of Canada (1). This report complements previous pathologic findings (2) and identifies SPD beyond the known Disease-endemic region. From 2001 to 2005, 20 dogs (5 mongrels and 15 beagles) showed pathologic lesions consistent with SPD. All beagles were born in coastal Florianopolis, Santa Catarina, Brazil, and later transferred to Maringa, Parana, Brazil, for the last 3–4 years of life. Lymph nodes, spleen, liver, and intestines from 10 beagles were aseptically obtained at necropsy in Maringa and frozen at -20°C until used at the Johns Hopkins Medical Institutions in Baltimore, Maryland. Genomic DNA was extracted from frozen tissues with QIAamp DNA Mini Kits (Qiagen, Valencia, CA, USA). DNA from N. helminthoeca and Anaplasma phagocytophilum was used as a positive control. Nuclease-free water was used as a negative control. We used gene-specific primers for Neorickettsia spp. 16S rRNA (rrs) (NeoSH-F; 5´-TAGGCCCGCGTTAGATTAGCTTGT-3´ and NeoSH-R; 5´-TACAACCCAAGGGCCTTCATCACT-3´) and N. helminthoeca RNA polymerase β-subunit (rpoB) (NH-rpoB-F: 5´-TGTCTTCGAAGGCCCAAAGACAGA-3´ and NH-rpoB-R: 5´-AGAACCGATAGAGCGGGCATGAAT-3´) (3) and heat-shock protein groESL (NH-groESL-F: 5´-AGGCTACTTCGCAGGCAAATGAGA-3´ and NH-groESL-R: 5´-CACGCTTCATTCCGCCCTTTAACT-3´) (4,5). Citrate synthase (gltA) gene primers (6) were also used. Two PCRs were conducted to maximize sensitivity. Specificity of N. helminthoeca–specific primers was shown by amplification studies of genomic DNA of A. phagocytophilum, Ehrlichia chaffeensis, E. canis, N. risticii, N. sennetsu, and N. helminthoeca. All amplicons were separated by electrophoresis in 1% agarose gels and purified before cloning (pGEM-T and pGEM-T Easy Vector Systems, Promega, Madison, WI, USA) and sequencing. The Maringa sequences obtained were compared with those in GenBank by using BLAST (http://www.ncbi.nlm.nih.gov/BLAST). Phylogenetic trees, sequence alignments, and identity tables were created by using Vector NTI Advance10 Software (Invitrogen, Carlsbad, CA, USA). GenBank accession numbers of Anaplasmataceae and their phylogenetic relationships are shown in the Figure. Figure Neighbor-joining phylogenetic trees of the 16S rRNA (rrs), RNA polymerase β-subunit (rpoB), and heat-shock protein (groESL) gene sequences of Anaplasmataceae families. Trees were constructed with Vector NTI Advance10 Software (Invitrogen, Carlsbad, ... Two dogs (N40–05, mesenteric lymph node, Maringa 1 and N20–04, Peyer's patch, Maringa 2) contained Neorickettsia spp. rrs, rpoB, or groESL genes. Both samples produced partial sequences for Neorickettsia spp. rrs gene; a similarity of 99% was observed for the 2 Maringa dog rrs sequences with N. sennetsu, N. risticii, and the Stellantchasmus falcatus (SF) agent. However, N. helminthoeca rpoB and groESL partial sequences were obtained only from dog 1. DNA identities of 100%, 82%, and 81% were observed between Maringa dog 1 sequences and N. helminthoeca, N. risticii, and N. sennetsu for the rpoB genes, respectively. All dogs were negative when tested with gltA gene primers. We observed 100% identity between the Maringa dog 1 sequence and N. helminthoeca groESL gene sequences. Similarities of 84%, 80%, and 79% were observed with N. sennetsu, the SF agent, and N. risticii, respectively. All positive controls showed bands of appropriate sizes, whereas negative controls yielded no products, confirming lack of amplicon contamination. This study demonstrates that 2 dogs from Maringa, Brazil, with pathologic lesions consistent with SPD (7) were infected with a Neorickettsia sp. The partial sequences from dog 1 were identical to N. helminthoeca rrs, groESL, and rpoB genes, confirming infection with this organism (2). To our knowledge, this is the first confirmed description of this organism beyond the known geographic area of SPD. The organism identified in Brazil has been named N. helminthoeca Maringa strain. Because of difficulty in recovering DNA from samples, need for a highly efficient PCR targeting small DNA regions, and limited sensitivity of the amplifications, sequences obtained for N. helminthoeca Maringa dog 1 (112 bp for rrs, 92 bp for groESL, 143 bp for rpoB) were short compared with those in GenBank (rrs 1,453 bp, groESL 1,914 bp, rpoB, 464 bp). Efficiency and sensitivity of targeting small DNA regions was necessary since storage and shipment of frozen samples were not optimal. Small DNA sequences are often suboptimal for delineation of phylogenetic relationships. Bootstrapping analyses showed poor resolution (
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Suspected cases of Neorickettsia-like organisms in Brazilian dogs.
Annals of the New York Academy of Sciences, 2004Co-Authors: Selwyn Arlington Headley, Odilon Vidotto, Diana G. Scorpio, J. Stephen Dumler, Joseph L. MankowskiAbstract:Preliminary findings of gross and histopathological lesions consistent with Salmon Poisoning Disease in 10 dogs from southern Brazil are described. Lesions were restricted to the spleen, lymph nodes, and intestinal lymphoid tissues. Grossly, there was marked hyperplasia of mesenteric lymph nodes and Peyer's patches. Microscopic alterations were characterized by diffuse hyperplasia of intestinal lymphoid tissues and Peyer's patches. Intracytoplasmic organisms consistent with Neorickettsia helminthoeca were demonstrated by Giemsa stain in reticuloendothelial cells of the intestine, spleen, Peyer's patches, and lymph nodes. We have named this organism Neorickettsia helminthoeca-like because of its marked similarity with the agent described in the United States.
