The Experts below are selected from a list of 651 Experts worldwide ranked by ideXlab platform
Hirotomo Kato - One of the best experts on this subject based on the ideXlab platform.
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nuclear and kinetoplast dna analyses reveal genetically complex leishmania strains with hybrid and mito nuclear discordance in peru
PLOS Neglected Tropical Diseases, 2020Co-Authors: Ahmed Tabbabi, Abraham G. Cáceres, Yoshihisa Hashiguchi, Pershing Bustamante T Chauca, Chisato Seki, Yanisa Choochartpong, Daiki Mizushima, Daisuke S Yamamoto, Hirotomo KatoAbstract:Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the Mannose Phosphate Isomerase (mpi) gene was applied to 134 skin samples collected from patients with cutaneous leishmaniasis (CL) in Peru for identification of the infecting parasite at the species level, and the results were compared with those of cytochrome b (cyt b) gene sequencing obtained in previous studies. Although most results (121/134) including 4 hybrids of Leishmania (Viannia) braziliensis and L. (V.) peruviana corresponded to those obtained in the previous study, PCR-RFLP analyses revealed the distribution of putative hybrid strains between L. (V.) peruviana and L. (V.) lainsoni in two samples, which has never been reported. Moreover, parasite strains showing discordance between kinetoplast and nuclear genes (kDNA and nDNA), so-called mito-nuclear discordance, were identified in 11 samples. Of these, six strains had the kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) guyanensis, and three strains had the kDNAs of L. (V.) shawi and nDNAs of L. (V.) braziliensis. The rest were identified as mito-nuclear discordance strains having kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) lainsoni, and kDNAs of L. (V.) lainsoni and nDNAs of L. (V.) braziliensis. The results demonstrate that Leishmania strains in Peru are genetically more complex than previously considered.
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first evidence of a hybrid of leishmania viannia braziliensis l v peruviana dna detected from the phlebotomine sand fly lutzomyia tejadai in peru
PLOS Neglected Tropical Diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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First Evidence of a Hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana DNA Detected from the Phlebotomine Sand Fly Lutzomyia tejadai in Peru.
PLoS neglected tropical diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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Short Report: Molecular Mass Screening to Incriminate Sand Fly Vectors of Andean-type Cutaneous Leishmaniasis in Ecuador and Peru
2014Co-Authors: Hirotomo Kato, Hiroshi Uezato, Tatsuyuki Mimori, Abraham G. Cáceres, Hiroyuki Iwata, Eduardo A. Gomez, Jorge D. Marco, Paola A. Barroso, Yoshihisa HashiguchiAbstract:Abstract. Sand flies from the Andean areas of Ecuador and Peru were examined for Leishmania infections by using our recently established molecular mass screening method. Leishmanial minicircle DNA-positive sand flies were de-tected in 3 of 192 and 1 of 462 samples from Ecuador and Peru, respectively. Sand fly species were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 18S ribosomal RNA (rRNA) gene, and the positive flies were Lutzomyia (Lu.) ayacuchensis and Lu. peruensis, respectively. Furthermore, cyto-chrome b and Mannose-Phosphate Isomerase gene sequence analyses identified the parasites from Ecuador and Peru as Leishmania (Leishmania) mexicana and L. (Viannia) peruviana, respectively. Thus, the mass screening method was confirmed to be a powerful tool for sand fly research. Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies of the genus Phlebotomus in the Old World and Lutzomyia in the New World.1,2 It is well known that the prevalent sand fly species and infection rate of sand flies with Leishmania spe-cies are important risk factors in each endemic area, because only part of sand fly species transmit each particular specie
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Use of FTA cards for direct sampling of patients' lesions in the ecological study of cutaneous leishmaniasis
Journal of Clinical Microbiology, 2010Co-Authors: Hirotomo Kato, Yuka Ishimaru, Amal S M Sayed, Lenin N. Velez, Megumi Fujita, Hiroshi Uezato, Tatsuyuki Mimori, Abraham G. Cáceres, Hiroyuki Iwata, Eduardo A L GomezAbstract:The FTA card (Whatman) was assessed for its utility as a molecular epidemiological tool in collecting samples from patients with leishmaniasis in Peru because the card has a variety of merits; it is less invasive for patients and easy to handle for both physicians and other medical personnel for sample collection or diagnosis, in addition to its simplicity and easy countrywide and/or intercountry transportation for analysis. Samples were collected from 132 patients suspected of having leishmaniasis, and Leishmania species were successfully identified in samples from 81 patients in 15 departments of Peru by cytochrome b and Mannose Phosphate Isomerase gene analyses. Of these, 61.7% were identified as Leishmania (Viannia) peruviana, 22.2% as L. (V.) braziliensis, 12.3% as L. (V.) guyanensis, 2.5% as L. (V.) shawi, and 1.2% as L. (V.) lainsoni. The three predominant species, L. (V.) peruviana, L. (V.) braziliensis, and L. (V.) guyanensis, were mainly found in the Andean highlands, in the tropical rainforest, and in northern and central rainforest regions, respectively. This is the first time L. (V.) shawi has been identified outside Brazil. The present study showed that the FTA card will be a useful tool for the ecological study of different forms of leishmaniasis. Furthermore, collecting samples directly from patients' lesions by using the FTA card eliminates (i) the possibility of contamination of Leishmania isolates during short- and/or long-term passages of culture in vitro in each laboratory and (ii) pain and suffering of patients from taking samples by skin biopsy.
Yoshihisa Hashiguchi - One of the best experts on this subject based on the ideXlab platform.
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nuclear and kinetoplast dna analyses reveal genetically complex leishmania strains with hybrid and mito nuclear discordance in peru
PLOS Neglected Tropical Diseases, 2020Co-Authors: Ahmed Tabbabi, Abraham G. Cáceres, Yoshihisa Hashiguchi, Pershing Bustamante T Chauca, Chisato Seki, Yanisa Choochartpong, Daiki Mizushima, Daisuke S Yamamoto, Hirotomo KatoAbstract:Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the Mannose Phosphate Isomerase (mpi) gene was applied to 134 skin samples collected from patients with cutaneous leishmaniasis (CL) in Peru for identification of the infecting parasite at the species level, and the results were compared with those of cytochrome b (cyt b) gene sequencing obtained in previous studies. Although most results (121/134) including 4 hybrids of Leishmania (Viannia) braziliensis and L. (V.) peruviana corresponded to those obtained in the previous study, PCR-RFLP analyses revealed the distribution of putative hybrid strains between L. (V.) peruviana and L. (V.) lainsoni in two samples, which has never been reported. Moreover, parasite strains showing discordance between kinetoplast and nuclear genes (kDNA and nDNA), so-called mito-nuclear discordance, were identified in 11 samples. Of these, six strains had the kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) guyanensis, and three strains had the kDNAs of L. (V.) shawi and nDNAs of L. (V.) braziliensis. The rest were identified as mito-nuclear discordance strains having kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) lainsoni, and kDNAs of L. (V.) lainsoni and nDNAs of L. (V.) braziliensis. The results demonstrate that Leishmania strains in Peru are genetically more complex than previously considered.
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First Evidence of a Hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana DNA Detected from the Phlebotomine Sand Fly Lutzomyia tejadai in Peru.
PLoS neglected tropical diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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first evidence of a hybrid of leishmania viannia braziliensis l v peruviana dna detected from the phlebotomine sand fly lutzomyia tejadai in peru
PLOS Neglected Tropical Diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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Short Report: Molecular Mass Screening to Incriminate Sand Fly Vectors of Andean-type Cutaneous Leishmaniasis in Ecuador and Peru
2014Co-Authors: Hirotomo Kato, Hiroshi Uezato, Tatsuyuki Mimori, Abraham G. Cáceres, Hiroyuki Iwata, Eduardo A. Gomez, Jorge D. Marco, Paola A. Barroso, Yoshihisa HashiguchiAbstract:Abstract. Sand flies from the Andean areas of Ecuador and Peru were examined for Leishmania infections by using our recently established molecular mass screening method. Leishmanial minicircle DNA-positive sand flies were de-tected in 3 of 192 and 1 of 462 samples from Ecuador and Peru, respectively. Sand fly species were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 18S ribosomal RNA (rRNA) gene, and the positive flies were Lutzomyia (Lu.) ayacuchensis and Lu. peruensis, respectively. Furthermore, cyto-chrome b and Mannose-Phosphate Isomerase gene sequence analyses identified the parasites from Ecuador and Peru as Leishmania (Leishmania) mexicana and L. (Viannia) peruviana, respectively. Thus, the mass screening method was confirmed to be a powerful tool for sand fly research. Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies of the genus Phlebotomus in the Old World and Lutzomyia in the New World.1,2 It is well known that the prevalent sand fly species and infection rate of sand flies with Leishmania spe-cies are important risk factors in each endemic area, because only part of sand fly species transmit each particular specie
Abraham G. Cáceres - One of the best experts on this subject based on the ideXlab platform.
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nuclear and kinetoplast dna analyses reveal genetically complex leishmania strains with hybrid and mito nuclear discordance in peru
PLOS Neglected Tropical Diseases, 2020Co-Authors: Ahmed Tabbabi, Abraham G. Cáceres, Yoshihisa Hashiguchi, Pershing Bustamante T Chauca, Chisato Seki, Yanisa Choochartpong, Daiki Mizushima, Daisuke S Yamamoto, Hirotomo KatoAbstract:Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the Mannose Phosphate Isomerase (mpi) gene was applied to 134 skin samples collected from patients with cutaneous leishmaniasis (CL) in Peru for identification of the infecting parasite at the species level, and the results were compared with those of cytochrome b (cyt b) gene sequencing obtained in previous studies. Although most results (121/134) including 4 hybrids of Leishmania (Viannia) braziliensis and L. (V.) peruviana corresponded to those obtained in the previous study, PCR-RFLP analyses revealed the distribution of putative hybrid strains between L. (V.) peruviana and L. (V.) lainsoni in two samples, which has never been reported. Moreover, parasite strains showing discordance between kinetoplast and nuclear genes (kDNA and nDNA), so-called mito-nuclear discordance, were identified in 11 samples. Of these, six strains had the kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) guyanensis, and three strains had the kDNAs of L. (V.) shawi and nDNAs of L. (V.) braziliensis. The rest were identified as mito-nuclear discordance strains having kDNAs of L. (V.) braziliensis or L. (V.) peruviana and nDNAs of L. (V.) lainsoni, and kDNAs of L. (V.) lainsoni and nDNAs of L. (V.) braziliensis. The results demonstrate that Leishmania strains in Peru are genetically more complex than previously considered.
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first evidence of a hybrid of leishmania viannia braziliensis l v peruviana dna detected from the phlebotomine sand fly lutzomyia tejadai in peru
PLOS Neglected Tropical Diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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First Evidence of a Hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana DNA Detected from the Phlebotomine Sand Fly Lutzomyia tejadai in Peru.
PLoS neglected tropical diseases, 2016Co-Authors: Hirotomo Kato, Abraham G. Cáceres, Yoshihisa HashiguchiAbstract:The natural infection of sand flies by Leishmania was examined in the Department of Huanuco of Peru, where cutaneous leishmaniasis caused by a hybrid of Leishmania (Viannia) braziliensis/L. (V.) peruviana is endemic. A total of 2,997 female sand flies were captured by CDC light traps and Shannon traps, of which 2,931 and 66 flies were identified as Lutzomyia tejadai and Lu fischeri, respectively. Using crude DNA extracted from individual sand flies as a template, Leishmania DNA was detected from one Lu. tejadai. The parasite species was identified as a hybrid of L. (V.) braziliensis/L. (V.) peruviana on the basis of cytochrome b and Mannose Phosphate Isomerase gene analyses. The result suggested that Lu. tejadai is responsible for the transmission of the hybrid Leishmania circulating in this area.
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Short Report: Molecular Mass Screening to Incriminate Sand Fly Vectors of Andean-type Cutaneous Leishmaniasis in Ecuador and Peru
2014Co-Authors: Hirotomo Kato, Hiroshi Uezato, Tatsuyuki Mimori, Abraham G. Cáceres, Hiroyuki Iwata, Eduardo A. Gomez, Jorge D. Marco, Paola A. Barroso, Yoshihisa HashiguchiAbstract:Abstract. Sand flies from the Andean areas of Ecuador and Peru were examined for Leishmania infections by using our recently established molecular mass screening method. Leishmanial minicircle DNA-positive sand flies were de-tected in 3 of 192 and 1 of 462 samples from Ecuador and Peru, respectively. Sand fly species were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 18S ribosomal RNA (rRNA) gene, and the positive flies were Lutzomyia (Lu.) ayacuchensis and Lu. peruensis, respectively. Furthermore, cyto-chrome b and Mannose-Phosphate Isomerase gene sequence analyses identified the parasites from Ecuador and Peru as Leishmania (Leishmania) mexicana and L. (Viannia) peruviana, respectively. Thus, the mass screening method was confirmed to be a powerful tool for sand fly research. Leishmaniasis is a protozoan disease caused by the genus Leishmania transmitted by female phlebotomine sand flies of the genus Phlebotomus in the Old World and Lutzomyia in the New World.1,2 It is well known that the prevalent sand fly species and infection rate of sand flies with Leishmania spe-cies are important risk factors in each endemic area, because only part of sand fly species transmit each particular specie
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Use of FTA cards for direct sampling of patients' lesions in the ecological study of cutaneous leishmaniasis
Journal of Clinical Microbiology, 2010Co-Authors: Hirotomo Kato, Yuka Ishimaru, Amal S M Sayed, Lenin N. Velez, Megumi Fujita, Hiroshi Uezato, Tatsuyuki Mimori, Abraham G. Cáceres, Hiroyuki Iwata, Eduardo A L GomezAbstract:The FTA card (Whatman) was assessed for its utility as a molecular epidemiological tool in collecting samples from patients with leishmaniasis in Peru because the card has a variety of merits; it is less invasive for patients and easy to handle for both physicians and other medical personnel for sample collection or diagnosis, in addition to its simplicity and easy countrywide and/or intercountry transportation for analysis. Samples were collected from 132 patients suspected of having leishmaniasis, and Leishmania species were successfully identified in samples from 81 patients in 15 departments of Peru by cytochrome b and Mannose Phosphate Isomerase gene analyses. Of these, 61.7% were identified as Leishmania (Viannia) peruviana, 22.2% as L. (V.) braziliensis, 12.3% as L. (V.) guyanensis, 2.5% as L. (V.) shawi, and 1.2% as L. (V.) lainsoni. The three predominant species, L. (V.) peruviana, L. (V.) braziliensis, and L. (V.) guyanensis, were mainly found in the Andean highlands, in the tropical rainforest, and in northern and central rainforest regions, respectively. This is the first time L. (V.) shawi has been identified outside Brazil. The present study showed that the FTA card will be a useful tool for the ecological study of different forms of leishmaniasis. Furthermore, collecting samples directly from patients' lesions by using the FTA card eliminates (i) the possibility of contamination of Leishmania isolates during short- and/or long-term passages of culture in vitro in each laboratory and (ii) pain and suffering of patients from taking samples by skin biopsy.
Marco, Jorge D. - One of the best experts on this subject based on the ideXlab platform.
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Short Report: Molecular Mass Screening to Incriminate Sand Fly Vectors of Andean-type Cutaneous Leishmaniasis in Ecuador and Peru
American Society of Tropical Medicine and Hygiene, 2010Co-Authors: Kato Hirotomo., Cáceres, Abraham G., Gómez, Eduardo A., Mimori Tatsuyuki., Uezato Hiroshi., Marco, Jorge D.Abstract:KATO, Hirotomo [et al.]. Short Report: Molecular Mass Screening to Incriminate Sand Fly Vectors of Andean-type Cutaneous Leishmaniasis in Ecuador and Peru.American Journal of Tropical Medicine and Hygiene. [en línea]. 2008, 79, 5, p. 719-721. ISSN 0002-9637.Sand flies from the Andean areas of Ecuador and Peru were examined for Leishmania infections by using our recently established molecular mass screening method. Leishmanial minicircle DNA-positive sand flies were de- tected in 3 of 192 and 1 of 462 samples from Ecuador and Peru, respectively. Sand fly species were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 18S ribosomal RNA (rRNA)gene, and the positive flies were Lutzomyia (Lu.) ayacuchensis and Lu. peruensis, respectively. Furthermore, cytochrome b and Mannose-Phosphate Isomerase gene sequence analyses identified the parasites from Ecuador and Peru as Leishmania (Leishmania) mexicana and L. (Viannia) peruviana, respectively. Thus, the mass screening method was confirmed to be a powerful tool for sand fly research
Rosimeire Takaki Rosa - One of the best experts on this subject based on the ideXlab platform.
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Detecção do polimorfismo genetico de Streptococcus mutans isolados de individuos livres de carie (CPOD=0)
2017Co-Authors: Rosimeire Takaki RosaAbstract:Resumo: A presente pesquisa teve por objetivo analisar a variabilidade infraespecífica de cepas de Streptococcus mutans isoladas de indivíduos livres de cárie. Foram coletadas amostras de saliva total não estimulada, placa dental e de dorso de língua de indivíduos que apresentavam índice CPOD igual a zero. Após dispersão e diluição seriada das amostras, alíquotas de cada diluição foram inoculadas em meio Mitis Salivarius Bacitracina Agar (MSB). Após incubação em estufa de pC02 10% a 37°C, foram isoladas do meio colônias com morfologia típica de estreptococos, para posterior identificação por provas bioquímicas. As amostras foram incubadas em caldo de Infusão de Cérebro e Coração (BHI), centrifugadas e lavadas, sendo os sedimentos obtidos submetidos ao processo de extração das proteínas intracelulares. Os extratos protéicos foram separados por eletroforese em gel de amido e corados especificamente para a detecção do polimorfismo isoenzimático. Os géis foram analisados através dos sistemas enzimáticos: leucina aminopeptidase (LAP), manitol 1-fosfato desidrogenase (MIP), manose fosfato Isomerase (MPI), nucleosídio fosforilase (NSP), fenilalanil leucina peptidase (PLP) e transaminase glutâmico-oxalacética (GOT). A análise comparativa dos perfis da eletroforese de isoenzimas (Multilocus Enzyme Electrophoresis-MLEE) dos isolados, acessada pela soma de todos sistemas enzimáticos, permitiu observar a ocorrência de dois ou mais clones de S. mutans nos diferentes sítios nestes voluntários. Os resultados obtidos mostram que pode ser encontrado mais de um tipo genético de S. mutans colonizando os diferentes sítios em um mesmo indivíduoAbstract: The current research had as aim to analyze the infra-specific variability of Streptococcus mutans strains obtained from caries-free individuals. Non-stimulated whole saliva, dental biofilm and tongue dorsum samples were collected from subjects with DMFT = O. After dispersing and decimal dilutions, aliquots from each dilution were inoculated in Mitis Salivarius Bacitracin (MSB) agar plates, in duplicate. After growing at pCO2 100,/0 and 37°C, it was taken some S. mutans suspect colonies that were biochemically identified. S. mutans-positive colonies were grown in Brain Heart Infusion (BHI) broth, they were centrifuged and washed. The pellets were submitted to cell disruption and protein extraction. Protein extracts were separated by starch gel electrophoresis and specifically stained in order to obtain the isoenzymic polymorphism. The gels were analyzed for enzymatic systems: leucine amino peptidase (LAP), mannitol-l-Phosphate dehydrogenase (MIP), Mannose Phosphate Isomerase (MPI), nucleoside phosphorylase (NSP), phenylalanyl leucine peptidase (PLP) e glutamic-oxalacetic transaminase (GOT). The comparative analysis of MLEE profiles assessed by the sum of all enzymatic systems allowed the observation of occurrence of one or more S. mutans dones, varying in thedifferent intra oral sites among these volunteers. The obtained results showed that it may be found more than one genetic type of such S. mutans colonizing different sites in a same subjec
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Detecção do polimorfismo genetico de Streptococcus mutans isolados de individuos livres de carie (CPOD=0)
Universidade Estadual de Campinas. Faculdade de Odontologia de Piracicaba, 2001Co-Authors: Rosimeire Takaki RosaAbstract:A presente pesquisa teve por objetivo analisar a variabilidade infraespecífica de cepas de Streptococcus mutans isoladas de indivíduos livres de cárie. Foram coletadas amostras de saliva total não estimulada, placa dental e de dorso de língua de indivíduos que apresentavam índice CPOD igual a zero. Após dispersão e diluição seriada das amostras, alíquotas de cada diluição foram inoculadas em meio Mitis Salivarius Bacitracina Agar (MSB). Após incubação em estufa de pC02 10% a 37°C, foram isoladas do meio colônias com morfologia típica de estreptococos, para posterior identificação por provas bioquímicas. As amostras foram incubadas em caldo de Infusão de Cérebro e Coração (BHI), centrifugadas e lavadas, sendo os sedimentos obtidos submetidos ao processo de extração das proteínas intracelulares. Os extratos protéicos foram separados por eletroforese em gel de amido e corados especificamente para a detecção do polimorfismo isoenzimático. Os géis foram analisados através dos sistemas enzimáticos: leucina aminopeptidase (LAP), manitol 1-fosfato desidrogenase (MIP), manose fosfato Isomerase (MPI), nucleosídio fosforilase (NSP), fenilalanil leucina peptidase (PLP) e transaminase glutâmico-oxalacética (GOT). A análise comparativa dos perfis da eletroforese de isoenzimas (Multilocus Enzyme Electrophoresis-MLEE) dos isolados, acessada pela soma de todos sistemas enzimáticos, permitiu observar a ocorrência de dois ou mais clones de S. mutans nos diferentes sítios nestes voluntários. Os resultados obtidos mostram que pode ser encontrado mais de um tipo genético de S. mutans colonizando os diferentes sítios em um mesmo indivíduoThe current research had as aim to analyze the infra-specific variability of Streptococcus mutans strains obtained from caries-free individuals. Non-stimulated whole saliva, dental biofilm and tongue dorsum samples were collected from subjects with DMFT = O. After dispersing and decimal dilutions, aliquots from each dilution were inoculated in Mitis Salivarius Bacitracin (MSB) agar plates, in duplicate. After growing at pCO2 100,/0 and 37°C, it was taken some S. mutans suspect colonies that were biochemically identified. S. mutans-positive colonies were grown in Brain Heart Infusion (BHI) broth, they were centrifuged and washed. The pellets were submitted to cell disruption and protein extraction. Protein extracts were separated by starch gel electrophoresis and specifically stained in order to obtain the isoenzymic polymorphism. The gels were analyzed for enzymatic systems: leucine amino peptidase (LAP), mannitol-l-Phosphate dehydrogenase (MIP), Mannose Phosphate Isomerase (MPI), nucleoside phosphorylase (NSP), phenylalanyl leucine peptidase (PLP) e glutamic-oxalacetic transaminase (GOT). The comparative analysis of MLEE profiles assessed by the sum of all enzymatic systems allowed the observation of occurrence of one or more S. mutans dones, varying in the different intra oral sites among these volunteers. The obtained results showed that it may be found more than one genetic type of such S. mutans colonizing different sites in a same subjec