Cytogenetic abnormalities in patients with hematological malignancies in Lahore city, Pakistan / Anormalidades citogenéticas em pacientes com neoplasias hematológicas na cidade de Lahore, Paquistão

Abstract Hematological and hematopoietic cells malignancies of the genes and hematopoietic cells are associated with the genetic mutation, often at the chromosomal level. The standard cytogenetic study is widely accepted as one of the main diagnostics and prognostic determinants in patients. Therefore, the current descriptive and cross-sectional study sought to determine the cytogenetic analysis of frequent hematological malignancies in Pakistan. A total of 202 peripheral bone marrow or blood samples from patients with benign and malignant hematological malignancy were taken using a conventional G-banding technique. Among enrolled patients, the mean age was 21.5 years ± 23.4, and gender-wise distribution showed a marked predominance of the male 147 (73%) population compared to the female 55 (27%). Patients in the age group (2-10 years) had the highest frequency, 48 (24%), of hematological neoplasms, followed by age (11-20 years) with 40 (20%). Normal karyotypes (46, XX/46, XY) was found in 51% (n=103) patients. Furthermore, the frequency of complex karyotype was 30 (15%), while normal was seen in 171 (85%) patients. Pre-B Acute Lymphoblastic Leukemia (Pre-B ALL) was the most prevalent malignancy of 66 (33%), followed by Chronic Myelogenous Leukemia (CML) of 41 (20%) and Acute Lymphocytic Leukemia of 29 (14%). Translocation was the most prevalent 50 (25%), followed by hypotriploidy 14 (7%) and monosomy 8 (4%) on chromosome aberration analysis. In addition, t(9:22) translocation was found to be 20 (10%) in CML, with the majority in the age group (31-40 years). This study recommends that karyotyping should be tested frequently in hematological conditions because it may provide insight into the relative chromosomal changes associated with particular malignancies.

Resumo As neoplasias hematológicas e de células hematopoiéticas dos genes e as células hematopoiéticas estão associadas à mutação genética, geralmente em nível cromossômico. O estudo citogenético padrão é amplamente aceito como um dos principais determinantes diagnósticos e prognósticos em pacientes. Portanto, o presente estudo descritivo e transversal buscou determinar a análise citogenética de neoplasias hematológicas frequentes no Paquistão. Um total de 202 amostras de medula óssea periférica ou sangue de pacientes com malignidade hematológica benigna e maligna foi coletado usando uma técnica convencional de banda G. Entre os pacientes inscritos, a média de idade foi de 21,5 anos ± 23,4, e a distribuição por gênero mostrou uma marcada predominância da população masculina de 147 (73%) em comparação com a feminina de 55 (27%). Pacientes na faixa etária (2-10 anos) tiveram a maior frequência, 48 (24%), de neoplasias hematológicas, seguida da idade (11-20 anos) com 40 (20%). Cariótipos normais (46, XX / 46, XY) foram encontrados em 51% (n = 103) dos pacientes. Além disso, a frequência de cariótipo complexo foi de 30 (15%), enquanto normal foi observada em 171 (85%) pacientes. Leucemia linfoblástica aguda pré-B (LLA Pré-B) foi a doença maligna mais prevalente de 66 (33%), seguida por leucemia mieloide crônica (LMC) de 41 (20%) e leucemia linfocítica aguda de 29 (14%). A translocação foi o 50 mais prevalente (25%), seguido por hipotriploidia 14 (7%) e monossomia 8 (4%) na análise de aberração cromossômica. Além disso, a translocação t (9:22) encontrada foi de 20 (10%) na LMC, com a maioria na faixa etária (31-40 anos). Este estudo recomenda que o cariótipo deve ser testado com frequência em condições hematológicas porque pode fornecer informações sobre as alterações cromossômicas relativas associadas a doenças malignas específicas.

Comparison of different diagnostic modalities for isolation of Mycobacterium Tuberculosis among suspected tuberculous lymphadenitis patients / Comparação de diferentes modalidades de diagnóstico para isolamento de Mycobacterium tuberculosis entre pacientes com suspeita de linfadenite tuberculosa

Tuberculosis is a communicable disease with high morbidity and mortality rates in developing countries. The study's primary objective is to compare conventional methods such as acid-fast bacillus (AFB) culture and microscopy with rapid diagnostic methods. The secondary objective is to compare histopathological and microbiological findings in suspected patients with tubercular lymphadenitis. A total of 111 samples (August 2018 to September 2019) of lymph nodes were processed for AFB microscopy, AFB cultures, drug-susceptibility testing (DST), histopathology, and Xpert Mycobacterium Tuberculosis (MTB)/resistance to Rifampin (RIF) assays. Out of 111 lymph node samples, 6 (5.4%) were positive for AFB smear microscopy, 84 (75.6%) were positive for AFB culture, 80 (70.7%) were positive on Gene Xpert, and 102 (91.8%) were indicative of tuberculosis for histopathology studies. Mycobacteria growth indicator tube (MGIT) culture positivity was 84 (75.6%) higher than solid Lowenstein-Jensen (LJ) culture 74 (66.6%). Positive cultures underwent phenotypic DST. Two cases were Multidrug-resistant (MDR) on DST, while three cases were Rifampicin resistant on Gene Xpert. The sensitivity of Genexpert was (62%) against the conventional AFB culture method. The poor performance of conventional lymphadenitis diagnostic methods requires early and accurate diagnostic methodology. Xpert MTB/RIF test can help in the treatment of multidrug-resistant TB cases. Nonetheless, rapid and conventional methods should be used for complete isolation of Mycobacterium tuberculosis.

A tuberculose é uma doença transmissível com altas taxas de morbimortalidade nos países em desenvolvimento. O objetivo principal do estudo é comparar métodos convencionais, como cultura de bacilo álcool-ácido resistente (BAAR) e microscopia, com métodos de diagnóstico rápido. O objetivo secundário é comparar os achados histopatológicos e microbiológicos em pacientes com suspeita de linfadenite tubercular. Um total de 111 amostras (agosto de 2018 a setembro de 2019) de gânglios linfáticos foi processado para microscopia de AFB, culturas de AFB, teste de susceptibilidade a drogas (DST), histopatologia e Xpert Mycobacterium tuberculosis (MTB)/ensaios de resistência à rifampicina (RIF). Das 111 amostras de linfonodos, 6 (5,4%) foram positivas para baciloscopia de AFB, 84 (75,6%) foram positivas para cultura de AFB, 80 (70,7%) foram positivas para o GeneXpert e 102 (91,8%) foram indicativas de tuberculose para estudos histopatológicos. A positividade da cultura do tubo indicador de crescimento de micobactérias (MGIT) foi 84 (75,6%), maior que a cultura sólida de Lowenstein-Jensen (LJ), 74 (66,6%). As culturas positivas foram submetidas a DST fenotípico. Dois casos eram multirresistentes (MDR) ao DST, enquanto três casos eram resistentes à rifampicina no GeneXpert. A sensibilidade do GeneXpert foi 62% contra o método convencional de cultura AFB. O fraco desempenho dos métodos convencionais de diagnóstico de linfadenite requer metodologia de diagnóstico precoce e precisa. O teste Xpert MTB/RIF pode ajudar no tratamento de casos de tuberculose multirresistente. No entanto, métodos rápidos e convencionais devem ser usados para o isolamento completo do Mycobacterium tuberculosis.

Fixation of pfcrt chloroquine resistance alleles in Plasmodium falciparum clinical isolates collected from unrest tribal agencies of Pakistan / Fixação de alelos de resistência à cloroquina pfcrt em isolados clínicos de Plasmodium falciparum coletados de agitações de agências tribais do Paquistão

Plasmodium falciparum resistance to Chloroquine (CQ) is a significant cause of mortality and morbidity worldwide. There is a paucity of documented data on the prevalence of CQ-resistant mutant haplotypes of Pfcrt and Pfmdr1 genes from malaria-endemic war effected Federally Administered Tribal Areas of Pakistan. The objective of this study was to investigate the prevalence of P. falciparum CQ-resistance in this area. Clinical isolates were collected between May 2017 and May 2018 from North Waziristan and South Waziristan agencies of Federally Administrated Trial Area. Subsequently, Giemsa-stained blood smears were examined to detect Plasmodium falciparum. Extraction of malarial DNA was done from microscopy positive P. falciparum samples, and P. falciparum infections were confirmed by nested PCR (targeting Plasmodium small subunit ribosomal ribonucleic acid (ssrRNA) genes). All PCR confirmed P. falciparum samples were sequenced by pyrosequencing to find out mutation in Pfcrt gene at codon K76T and in pfmdr1 at codons N86Y, Y184F, N1042D, and D1246Y. Out of 121 microscopies positive P. falciparum cases, 109 samples were positive for P. falciparum by nested PCR. Pfcrt K76T mutation was found in 96% of isolates, Pfmdr1 N86Y mutation was observed in 20%, and 11% harboured Y184F mutation. All samples were wild type for Pfmdr1 codon N1042D and D1246Y. In the FATA, Pakistan, the frequency of resistant allele 76T remained high despite the removal of CQ. However, current findings of the study suggest complete fixation of P. falciparum CQ‑resistant genotype in the study area.

A resistência do Plasmodium falciparum à cloroquina (CQ) é uma causa significativa de mortalidade e morbidade em todo o mundo. Há uma escassez de dados documentados sobre a prevalência de haplótipos mutantes CQ-resistentes dos genes Pfcrt e Pfmdr1 da guerra endêmica da malária em áreas tribais administradas pelo governo federal do Paquistão. O objetivo deste estudo foi investigar a prevalência de resistência a CQ de P. falciparum nesta área. Isolados clínicos foram coletados entre maio de 2017 e maio de 2018 nas agências do Waziristão do Norte e do Waziristão do Sul da Área de Ensaio Administrada Federalmente. Posteriormente, esfregaços de sangue corados com Giemsa foram examinados para detectar Plasmodium falciparum. A extração do DNA da malária foi feita a partir de amostras de P. falciparum positivas para microscopia, e as infecções por P. falciparum foram confirmadas por nested PCR (visando genes de ácido ribonucleico ribossômico de subunidade pequena de Plasmodium (ssrRNA)). Todas as amostras de P. falciparum confirmadas por PCR foram sequenciadas por pirosequenciamento para descobrir a mutação no gene Pfcrt no códon K76T e em pfmdr1 nos códons N86Y, Y184F, N1042D e D1246Y. De 121 microscopias de casos positivos de P. falciparum, 109 amostras foram positivas para P. falciparum por nested PCR. A mutação Pfcrt K76T foi encontrada em 96% dos isolados, a mutação Pfmdr1 N86Y foi observada em 20% e 11% abrigou a mutação Y184F. Todas as amostras eram do tipo selvagem para o códon N1042D e D1246Y de Pfmdr1. No FATA, Paquistão, a frequência do alelo resistente 76T permaneceu alta apesar da remoção de CQ. No entanto, as descobertas atuais do estudo sugerem a fixação completa do genótipo resistente a CQ de P. falciparum na área de estudo.

Molecular Epidemiology of Plasmodium species in Conflicted Federally Administered Tribal Area (FATA) Pakistan / Epidemiologia molecular de espécies de plasmódio em área tribal sob administração federal em conflito do Paquistão

Military conflicts have been significant obstacles in detecting and treating infectious disease diseases due to the diminished public health infrastructure, resulting in malaria endemicity. A variety of violent and destructive incidents were experienced by FATA (Federally Administered Tribal Areas). It was a struggle to pursue an epidemiological analysis due to continuing conflict and Talibanization. Clinical isolates were collected from Bajaur, Mohmand, Khyber, Orakzai agencies from May 2017 to May 2018. For Giemsa staining, full blood EDTA blood samples have been collected from symptomatic participants. Malaria-positive microscopy isolates were spotted on filter papers for future Plasmodial molecular detection by nested polymerase chain reaction (nPCR) of small subunit ribosomal ribonucleic acid (ssrRNA) genes specific primers. Since reconfirming the nPCR, a malariometric study of 762 patients found 679 positive malaria cases. Plasmodium vivax was 523 (77%), Plasmodium falciparum 121 (18%), 35 (5%) were with mixed-species infection (P. vivax plus P. falciparum), and 83 were declared negative by PCR. Among the five agencies of FATA, Khyber agency has the highest malaria incidence (19%) with followed by P. vivax (19%) and P. falciparum (4.1%). In contrast, Kurram has about (14%), including (10.8%) P. vivax and (2.7%) P. falciparum cases, the lowest malaria epidemiology. Surprisingly, no significant differences in the distribution of mixed-species infection among all five agencies. P. falciparum and P. vivax were two prevalent FATA malaria species in Pakistan's war-torn area. To overcome this rising incidence of malaria, this study recommends that initiating malaria awareness campaigns in school should be supported by public health agencies and malaria related education locally, targeting children and parents alike.

Os conflitos militares têm sido obstáculos significativos na detecção e tratamento de doenças infecciosas devido à diminuição da infraestrutura de saúde pública, resultando na endemicidade da malária. Uma variedade de incidentes violentos e destrutivos foi vivida pelas FATA (áreas tribais administradas pelo governo federal). Foi uma luta busca ruma análise epidemiológica devido ao conflito contínuo e à talibanização. Isolados clínicos foram coletados de agências Bajaur, Mohmand, Khyber e Orakzai, de maio de 2017 a maio de 2018. Para a coloração de Giemsa, amostras de sangue completo com EDTA foram coletadas de participantes sintomáticos. Isolados de microscopia positivos para malária foram colocados em papéis de filtro para futura detecção molecular plasmódica por reação em cadeia da polimerase aninhada (nPCR) de primers específicos de genes de subunidade ribossômica de ácido ribonucleico (ssrRNA). Desde a reconfirmação do nPCR, um estudo malariométrico de 762 pacientes encontrou 679 casos positivos de malária. Plasmodium vivax foi 523 (77%), Plasmodium falciparum 121 (18%), 35 (5%) eram com infecção de espécies mistas (P. vivax mais P. falciparum) e 83 foram declarados negativos por PCR. Entre as cinco agências da FATA, a agência Khyber tem a maior incidência de malária (19%), seguida por P. vivax (19%) e P. falciparum (4,1%). Em contraste, Kurram tem cerca de 14%, incluindo 10,8% casos de P. vivax e 2,7% P. falciparum, a epidemiologia de malária mais baixa. Surpreendentemente, não há diferenças significativas na distribuição da infecção de espécies mistas entre todas as cinco agências. P. falciparum e P. vivax foram duas espécies prevalentes de malária FATA na área devastada pela guerra no Paquistão. Para superar essa incidência crescente de malária, este estudo recomenda que o início de campanhas de conscientização sobre a malária na escola deve ser apoiado por agências de saúde pública e educação relacionada com a malária localmente, visando crianças e pais.

Cytogenetic abnormalities in patients with hematological malignancies in Lahore city, Pakistan / Anormalidades citogenéticas em pacientes com neoplasias hematológicas na cidade de Lahore, Paquistão

Hematological and hematopoietic cells malignancies of the genes and hematopoietic cells are associated with the genetic mutation, often at the chromosomal level. The standard cytogenetic study is widely accepted as one of the main diagnostics and prognostic determinants in patients. Therefore, the current descriptive and cross sectional study sought to determine the cytogenetic analysis of frequent hematological malignancies in Pakistan. A total of 202 peripheral bone marrow or blood samples from patients with benign and malignant hematological malignancy were taken using a conventional G-banding technique. Among enrolled patients, the mean age was 21.5 years ± 23.4, and gender-wise distribution showed a marked predominance of the male 147 (73%) population compared to the female 55 (27%). Patients in the age group (2-10 years) had the highest frequency, 48 (24%), of hematological neoplasms, followed by age (11-20 years) with 40 (20%). Normal karyotypes (46, XX/46, XY) was found in 51% (n=103) patients. Furthermore, the frequency of complex karyotype was 30 (15%), while normal was seen in 171 (85%) patients. Pre-B Acute Lymphoblastic Leukemia (Pre-B ALL) was the most prevalent malignancy of 66 (33%), followed by Chronic Myelogenous Leukemia (CML) of 41 (20%) and Acute Lymphocytic Leukemia of 29 (14%). Translocation was the most prevalent 50 (25%), followed by hypotriploidy 14 (7%) and monosomy 8 (4%) on chromosome aberration analysis. In addition, t(9:22) translocation was found to be 20 (10%) in CML, with the majority in the age group (31-40 years). This study recommends that karyotyping should be tested frequently in hematological conditions because it may provide insight into the relative chromosomal changes associated with particular malignancies.

As neoplasias hematológicas e de células hematopoiéticas dos genes e as células hematopoiéticas estão associadas à mutação genética, geralmente em nível cromossômico. O estudo citogenético padrão é amplamente aceito como um dos principais determinantes diagnósticos e prognósticos em pacientes. Portanto, o presente estudo descritivo e transversal buscou determinar a análise citogenética de neoplasias hematológicas frequentes no Paquistão. Um total de 202 amostras de medula óssea periférica ou sangue de pacientes com malignidade hematológica benigna e maligna foi coletado usando uma técnica convencional de banda G. Entre os pacientes inscritos, a média de idade foi de 21,5 anos ± 23,4, e a distribuição por gênero mostrou uma marcada predominância da população masculina de 147 (73%) em comparação com a feminina de 55 (27%). Pacientes na faixa etária (2-10 anos) tiveram a maior frequência, 48 (24%), de neoplasias hematológicas, seguida da idade (11-20 anos) com 40 (20%). Cariótipos normais (46, XX / 46, XY) foram encontrados em 51% (n = 103) dos pacientes. Além disso, a frequência de cariótipo complexo foi de 30 (15%), enquanto normal foi observada em 171 (85%) pacientes. Leucemia linfoblástica aguda pré-B (LLA Pré-B) foi a doença maligna mais prevalente de 66 (33%), seguida por leucemia mieloide crônica (LMC) de 41 (20%) e leucemia linfocítica aguda de 29 (14%). A translocação foi o 50 mais prevalente (25%), seguido por hipotriploidia 14 (7%) e monossomia 8 (4%) na análise de aberração cromossômica. Além disso, a translocação t (9:22) encontrada foi de 20 (10%) na LMC, com a maioria na faixa etária (31-40 anos). Este estudo recomenda que o cariótipo deve ser testado com frequência em condições hematológicas porque pode fornecer informações sobre as alterações cromossômicas relativas associadas a doenças malignas específicas.

Comparison of different diagnostic modalities for isolation of Mycobacterium Tuberculosis among suspected tuberculous lymphadenitis patients

Abstract Tuberculosis is a communicable disease with high morbidity and mortality rates in developing countries. The study's primary objective is to compare conventional methods such as acid-fast bacillus (AFB) culture and microscopy with rapid diagnostic methods. The secondary objective is to compare histopathological and microbiological findings in suspected patients with tubercular lymphadenitis. A total of 111 samples (August 2018 to September 2019) of lymph nodes were processed for AFB microscopy, AFB cultures, drug-susceptibility testing (DST), histopathology, and Xpert Mycobacterium Tuberculosis (MTB)/resistance to Rifampin (RIF) assays. Out of 111 lymph node samples, 6 (5.4%) were positive for AFB smear microscopy, 84 (75.6%) were positive for AFB culture, 80 (70.7%) were positive on Gene Xpert, and 102 (91.8%) were indicative of tuberculosis for histopathology studies. Mycobacteria growth indicator tube (MGIT) culture positivity was 84 (75.6%) higher than solid Lowenstein-Jensen (LJ) culture 74 (66.6%). Positive cultures underwent phenotypic DST. Two cases were Multidrug-resistant (MDR) on DST, while three cases were Rifampicin resistant on Gene Xpert. The sensitivity of Genexpert was (62%) against the conventional AFB culture method. The poor performance of conventional lymphadenitis diagnostic methods requires early and accurate diagnostic methodology. Xpert MTB/RIF test can help in the treatment of multidrug-resistant TB cases. Nonetheless, rapid and conventional methods should be used for complete isolation of Mycobacterium tuberculosis.

Resumo A tuberculose é uma doença transmissível com altas taxas de morbimortalidade nos países em desenvolvimento. O objetivo principal do estudo é comparar métodos convencionais, como cultura de bacilo álcool-ácido resistente (BAAR) e microscopia, com métodos de diagnóstico rápido. O objetivo secundário é comparar os achados histopatológicos e microbiológicos em pacientes com suspeita de linfadenite tubercular. Um total de 111 amostras (agosto de 2018 a setembro de 2019) de gânglios linfáticos foi processado para microscopia de AFB, culturas de AFB, teste de susceptibilidade a drogas (DST), histopatologia e Xpert Mycobacterium tuberculosis (MTB)/ensaios de resistência à rifampicina (RIF). Das 111 amostras de linfonodos, 6 (5,4%) foram positivas para baciloscopia de AFB, 84 (75,6%) foram positivas para cultura de AFB, 80 (70,7%) foram positivas para o GeneXpert e 102 (91,8%) foram indicativas de tuberculose para estudos histopatológicos. A positividade da cultura do tubo indicador de crescimento de micobactérias (MGIT) foi 84 (75,6%), maior que a cultura sólida de Lowenstein-Jensen (LJ), 74 (66,6%). As culturas positivas foram submetidas a DST fenotípico. Dois casos eram multirresistentes (MDR) ao DST, enquanto três casos eram resistentes à rifampicina no GeneXpert. A sensibilidade do GeneXpert foi 62% contra o método convencional de cultura AFB. O fraco desempenho dos métodos convencionais de diagnóstico de linfadenite requer metodologia de diagnóstico precoce e precisa. O teste Xpert MTB/RIF pode ajudar no tratamento de casos de tuberculose multirresistente. No entanto, métodos rápidos e convencionais devem ser usados para o isolamento completo do Mycobacterium tuberculosis.

Fixation of pfcrt chloroquine resistance alleles in Plasmodium falciparum clinical isolates collected from unrest tribal agencies of Pakistan

Abstract Plasmodium falciparum resistance to Chloroquine (CQ) is a significant cause of mortality and morbidity worldwide. There is a paucity of documented data on the prevalence of CQ-resistant mutant haplotypes of Pfcrt and Pfmdr1 genes from malaria-endemic war effected Federally Administered Tribal Areas of Pakistan. The objective of this study was to investigate the prevalence of P. falciparum CQ-resistance in this area. Clinical isolates were collected between May 2017 and May 2018 from North Waziristan and South Waziristan agencies of Federally Administrated Trial Area. Subsequently, Giemsa-stained blood smears were examined to detect Plasmodium falciparum. Extraction of malarial DNA was done from microscopy positive P. falciparum samples, and P. falciparum infections were confirmed by nested PCR (targeting Plasmodium small subunit ribosomal ribonucleic acid (ssrRNA) genes). All PCR confirmed P. falciparum samples were sequenced by pyrosequencing to find out mutation in Pfcrt gene at codon K76T and in pfmdr1 at codons N86Y, Y184F, N1042D, and D1246Y. Out of 121 microscopies positive P. falciparum cases, 109 samples were positive for P. falciparum by nested PCR. Pfcrt K76T mutation was found in 96% of isolates, Pfmdr1 N86Y mutation was observed in 20%, and 11% harboured Y184F mutation. All samples were wild type for Pfmdr1 codon N1042D and D1246Y. In the FATA, Pakistan, the frequency of resistant allele 76T remained high despite the removal of CQ. However, current findings of the study suggest complete fixation of P. falciparum CQ-resistant genotype in the study area.

Resumo A resistência do Plasmodium falciparum à cloroquina (CQ) é uma causa significativa de mortalidade e morbidade em todo o mundo. Há uma escassez de dados documentados sobre a prevalência de haplótipos mutantes CQ-resistentes dos genes Pfcrt e Pfmdr1 da guerra endêmica da malária em áreas tribais administradas pelo governo federal do Paquistão. O objetivo deste estudo foi investigar a prevalência de resistência a CQ de P. falciparum nesta área. Isolados clínicos foram coletados entre maio de 2017 e maio de 2018 nas agências do Waziristão do Norte e do Waziristão do Sul da Área de Ensaio Administrada Federalmente. Posteriormente, esfregaços de sangue corados com Giemsa foram examinados para detectar Plasmodium falciparum. A extração do DNA da malária foi feita a partir de amostras de P. falciparum positivas para microscopia, e as infecções por P. falciparum foram confirmadas por nested PCR (visando genes de ácido ribonucleico ribossômico de subunidade pequena de Plasmodium (ssrRNA)). Todas as amostras de P. falciparum confirmadas por PCR foram sequenciadas por pirosequenciamento para descobrir a mutação no gene Pfcrt no códon K76T e em pfmdr1 nos códons N86Y, Y184F, N1042D e D1246Y. De 121 microscopias de casos positivos de P. falciparum, 109 amostras foram positivas para P. falciparum por nested PCR. A mutação Pfcrt K76T foi encontrada em 96% dos isolados, a mutação Pfmdr1 N86Y foi observada em 20% e 11% abrigou a mutação Y184F. Todas as amostras eram do tipo selvagem para o códon N1042D e D1246Y de Pfmdr1. No FATA, Paquistão, a frequência do alelo resistente 76T permaneceu alta apesar da remoção de CQ. No entanto, as descobertas atuais do estudo sugerem a fixação completa do genótipo resistente a CQ de P. falciparum na área de estudo.

Molecular Epidemiology of Plasmodium species in Conflicted Federally Administered Tribal Area (FATA) Pakistan

Abstract Military conflicts have been significant obstacles in detecting and treating infectious disease diseases due to the diminished public health infrastructure, resulting in malaria endemicity. A variety of violent and destructive incidents were experienced by FATA (Federally Administered Tribal Areas). It was a struggle to pursue an epidemiological analysis due to continuing conflict and Talibanization. Clinical isolates were collected from Bajaur, Mohmand, Khyber, Orakzai agencies from May 2017 to May 2018. For Giemsa staining, full blood EDTA blood samples have been collected from symptomatic participants. Malaria-positive microscopy isolates were spotted on filter papers for future Plasmodial molecular detection by nested polymerase chain reaction (nPCR) of small subunit ribosomal ribonucleic acid (ssrRNA) genes specific primers. Since reconfirming the nPCR, a malariometric study of 762 patients found 679 positive malaria cases. Plasmodium vivax was 523 (77%), Plasmodium falciparum 121 (18%), 35 (5%) were with mixed-species infection (P. vivax plus P. falciparum), and 83 were declared negative by PCR. Among the five agencies of FATA, Khyber agency has the highest malaria incidence (19%) with followed by P. vivax (19%) and P. falciparum (4.1%). In contrast, Kurram has about (14%), including (10.8%) P. vivax and (2.7%) P. falciparum cases, the lowest malaria epidemiology. Surprisingly, no significant differences in the distribution of mixed-species infection among all five agencies. P. falciparum and P. vivax were two prevalent FATA malaria species in Pakistans war-torn area. To overcome this rising incidence of malaria, this study recommends that initiating malaria awareness campaigns in school should be supported by public health agencies and malaria-related education locally, targeting children and parents alike.

Resumo Os conflitos militares têm sido obstáculos significativos na detecção e tratamento de doenças infecciosas devido à diminuição da infraestrutura de saúde pública, resultando na endemicidade da malária. Uma variedade de incidentes violentos e destrutivos foi vivida pelas FATA (áreas tribais administradas pelo governo federal). Foi uma luta buscar uma análise epidemiológica devido ao conflito contínuo e à talibanização. Isolados clínicos foram coletados de agências Bajaur, Mohmand, Khyber e Orakzai, de maio de 2017 a maio de 2018. Para a coloração de Giemsa, amostras de sangue completo com EDTA foram coletadas de participantes sintomáticos. Isolados de microscopia positivos para malária foram colocados em papéis de filtro para futura detecção molecular plasmódica por reação em cadeia da polimerase aninhada (nPCR) de primers específicos de genes de subunidade ribossômica de ácido ribonucleico (ssrRNA). Desde a reconfirmação do nPCR, um estudo malariométrico de 762 pacientes encontrou 679 casos positivos de malária. Plasmodium vivax foi 523 (77%), Plasmodium falciparum 121 (18%), 35 (5%) eram com infecção de espécies mistas (P. vivax mais P. falciparum) e 83 foram declarados negativos por PCR. Entre as cinco agências da FATA, a agência Khyber tem a maior incidência de malária (19%), seguida por P. vivax (19%) e P. falciparum (4,1%). Em contraste, Kurram tem cerca de 14%, incluindo 10,8% casos de P. vivax e 2,7% P. falciparum, a epidemiologia de malária mais baixa. Surpreendentemente, não há diferenças significativas na distribuição da infecção de espécies mistas entre todas as cinco agências. P. falciparum e P. vivax foram duas espécies prevalentes de malária FATA na área devastada pela guerra no Paquistão. Para superar essa incidência crescente de malária, este estudo recomenda que o início de campanhas de conscientização sobre a malária na escola deve ser apoiado por agências de saúde pública e educação relacionada com a malária localmente, visando crianças e pais.

Cytogenetic abnormalities in patients with hematological malignancies in Lahore city, Pakistan

Abstract Hematological and hematopoietic cells malignancies of the genes and hematopoietic cells are associated with the genetic mutation, often at the chromosomal level. The standard cytogenetic study is widely accepted as one of the main diagnostics and prognostic determinants in patients. Therefore, the current descriptive and cross-sectional study sought to determine the cytogenetic analysis of frequent hematological malignancies in Pakistan. A total of 202 peripheral bone marrow or blood samples from patients with benign and malignant hematological malignancy were taken using a conventional G-banding technique. Among enrolled patients, the mean age was 21.5 years ± 23.4, and gender-wise distribution showed a marked predominance of the male 147 (73%) population compared to the female 55 (27%). Patients in the age group (2-10 years) had the highest frequency, 48 (24%), of hematological neoplasms, followed by age (11-20 years) with 40 (20%). Normal karyotypes (46, XX/46, XY) was found in 51% (n=103) patients. Furthermore, the frequency of complex karyotype was 30 (15%), while normal was seen in 171 (85%) patients. Pre-B Acute Lymphoblastic Leukemia (Pre-B ALL) was the most prevalent malignancy of 66 (33%), followed by Chronic Myelogenous Leukemia (CML) of 41 (20%) and Acute Lymphocytic Leukemia of 29 (14%). Translocation was the most prevalent 50 (25%), followed by hypotriploidy 14 (7%) and monosomy 8 (4%) on chromosome aberration analysis. In addition, t(9:22) translocation was found to be 20 (10%) in CML, with the majority in the age group (31-40 years). This study recommends that karyotyping should be tested frequently in hematological conditions because it may provide insight into the relative chromosomal changes associated with particular malignancies.

Resumo As neoplasias hematológicas e de células hematopoiéticas dos genes e as células hematopoiéticas estão associadas à mutação genética, geralmente em nível cromossômico. O estudo citogenético padrão é amplamente aceito como um dos principais determinantes diagnósticos e prognósticos em pacientes. Portanto, o presente estudo descritivo e transversal buscou determinar a análise citogenética de neoplasias hematológicas frequentes no Paquistão. Um total de 202 amostras de medula óssea periférica ou sangue de pacientes com malignidade hematológica benigna e maligna foi coletado usando uma técnica convencional de banda G. Entre os pacientes inscritos, a média de idade foi de 21,5 anos ± 23,4, e a distribuição por gênero mostrou uma marcada predominância da população masculina de 147 (73%) em comparação com a feminina de 55 (27%). Pacientes na faixa etária (2-10 anos) tiveram a maior frequência, 48 (24%), de neoplasias hematológicas, seguida da idade (11-20 anos) com 40 (20%). Cariótipos normais (46, XX / 46, XY) foram encontrados em 51% (n = 103) dos pacientes. Além disso, a frequência de cariótipo complexo foi de 30 (15%), enquanto normal foi observada em 171 (85%) pacientes. Leucemia linfoblástica aguda pré-B (LLA Pré-B) foi a doença maligna mais prevalente de 66 (33%), seguida por leucemia mieloide crônica (LMC) de 41 (20%) e leucemia linfocítica aguda de 29 (14%). A translocação foi o 50 mais prevalente (25%), seguido por hipotriploidia 14 (7%) e monossomia 8 (4%) na análise de aberração cromossômica. Além disso, a translocação t (9:22) encontrada foi de 20 (10%) na LMC, com a maioria na faixa etária (31-40 anos). Este estudo recomenda que o cariótipo deve ser testado com frequência em condições hematológicas porque pode fornecer informações sobre as alterações cromossômicas relativas associadas a doenças malignas específicas.

Comparison of different diagnostic modalities for isolation of Mycobacterium Tuberculosis among suspected tuberculous lymphadenitis patients / Comparação de diferentes modalidades de diagnóstico para isolamento de Mycobacterium tuberculosis entre pacientes com suspeita de linfadenite tuberculosa

Abstract Tuberculosis is a communicable disease with high morbidity and mortality rates in developing countries. The study's primary objective is to compare conventional methods such as acid-fast bacillus (AFB) culture and microscopy with rapid diagnostic methods. The secondary objective is to compare histopathological and microbiological findings in suspected patients with tubercular lymphadenitis. A total of 111 samples (August 2018 to September 2019) of lymph nodes were processed for AFB microscopy, AFB cultures, drug-susceptibility testing (DST), histopathology, and Xpert Mycobacterium Tuberculosis (MTB)/resistance to Rifampin (RIF) assays. Out of 111 lymph node samples, 6 (5.4%) were positive for AFB smear microscopy, 84 (75.6%) were positive for AFB culture, 80 (70.7%) were positive on Gene Xpert, and 102 (91.8%) were indicative of tuberculosis for histopathology studies. Mycobacteria growth indicator tube (MGIT) culture positivity was 84 (75.6%) higher than solid Lowenstein-Jensen (LJ) culture 74 (66.6%). Positive cultures underwent phenotypic DST. Two cases were Multidrug-resistant (MDR) on DST, while three cases were Rifampicin resistant on Gene Xpert. The sensitivity of Genexpert was (62%) against the conventional AFB culture method. The poor performance of conventional lymphadenitis diagnostic methods requires early and accurate diagnostic methodology. Xpert MTB/RIF test can help in the treatment of multidrug-resistant TB cases. Nonetheless, rapid and conventional methods should be used for complete isolation of Mycobacterium tuberculosis.

Resumo A tuberculose é uma doença transmissível com altas taxas de morbimortalidade nos países em desenvolvimento. O objetivo principal do estudo é comparar métodos convencionais, como cultura de bacilo álcool-ácido resistente (BAAR) e microscopia, com métodos de diagnóstico rápido. O objetivo secundário é comparar os achados histopatológicos e microbiológicos em pacientes com suspeita de linfadenite tubercular. Um total de 111 amostras (agosto de 2018 a setembro de 2019) de gânglios linfáticos foi processado ​​para microscopia de AFB, culturas de AFB, teste de susceptibilidade a drogas (DST), histopatologia e Xpert Mycobacterium tuberculosis (MTB)/ensaios de resistência à rifampicina (RIF). Das 111 amostras de linfonodos, 6 (5,4%) foram positivas para baciloscopia de AFB, 84 (75,6%) foram positivas para cultura de AFB, 80 (70,7%) foram positivas para o GeneXpert e 102 (91,8%) foram indicativas de tuberculose para estudos histopatológicos. A positividade da cultura do tubo indicador de crescimento de micobactérias (MGIT) foi 84 (75,6%), maior que a cultura sólida de Lowenstein-Jensen (LJ), 74 (66,6%). As culturas positivas foram submetidas a DST fenotípico. Dois casos eram multirresistentes (MDR) ao DST, enquanto três casos eram resistentes à rifampicina no GeneXpert. A sensibilidade do GeneXpert foi 62% contra o método convencional de cultura AFB. O fraco desempenho dos métodos convencionais de diagnóstico de linfadenite requer metodologia de diagnóstico precoce e precisa. O teste Xpert MTB/RIF pode ajudar no tratamento de casos de tuberculose multirresistente. No entanto, métodos rápidos e convencionais devem ser usados ​​para o isolamento completo do Mycobacterium tuberculosis.

Fixation of pfcrt chloroquine resistance alleles in Plasmodium falciparum clinical isolates collected from unrest tribal agencies of Pakistan / Fixação de alelos de resistência à cloroquina pfcrt em isolados clínicos de Plasmodium falciparum coletados de agitações de agências tribais do Paquistão

Abstract Plasmodium falciparum resistance to Chloroquine (CQ) is a significant cause of mortality and morbidity worldwide. There is a paucity of documented data on the prevalence of CQ-resistant mutant haplotypes of Pfcrt and Pfmdr1 genes from malaria-endemic war effected Federally Administered Tribal Areas of Pakistan. The objective of this study was to investigate the prevalence of P. falciparum CQ-resistance in this area. Clinical isolates were collected between May 2017 and May 2018 from North Waziristan and South Waziristan agencies of Federally Administrated Trial Area. Subsequently, Giemsa-stained blood smears were examined to detect Plasmodium falciparum. Extraction of malarial DNA was done from microscopy positive P. falciparum samples, and P. falciparum infections were confirmed by nested PCR (targeting Plasmodium small subunit ribosomal ribonucleic acid (ssrRNA) genes). All PCR confirmed P. falciparum samples were sequenced by pyrosequencing to find out mutation in Pfcrt gene at codon K76T and in pfmdr1 at codons N86Y, Y184F, N1042D, and D1246Y. Out of 121 microscopies positive P. falciparum cases, 109 samples were positive for P. falciparum by nested PCR. Pfcrt K76T mutation was found in 96% of isolates, Pfmdr1 N86Y mutation was observed in 20%, and 11% harboured Y184F mutation. All samples were wild type for Pfmdr1 codon N1042D and D1246Y. In the FATA, Pakistan, the frequency of resistant allele 76T remained high despite the removal of CQ. However, current findings of the study suggest complete fixation of P. falciparum CQ-resistant genotype in the study area.

Resumo A resistência do Plasmodium falciparum à cloroquina (CQ) é uma causa significativa de mortalidade e morbidade em todo o mundo. Há uma escassez de dados documentados sobre a prevalência de haplótipos mutantes CQ-resistentes dos genes Pfcrt e Pfmdr1 da guerra endêmica da malária em áreas tribais administradas pelo governo federal do Paquistão. O objetivo deste estudo foi investigar a prevalência de resistência a CQ de P. falciparum nesta área. Isolados clínicos foram coletados entre maio de 2017 e maio de 2018 nas agências do Waziristão do Norte e do Waziristão do Sul da Área de Ensaio Administrada Federalmente. Posteriormente, esfregaços de sangue corados com Giemsa foram examinados para detectar Plasmodium falciparum. A extração do DNA da malária foi feita a partir de amostras de P. falciparum positivas para microscopia, e as infecções por P. falciparum foram confirmadas por nested PCR (visando genes de ácido ribonucleico ribossômico de subunidade pequena de Plasmodium (ssrRNA)). Todas as amostras de P. falciparum confirmadas por PCR foram sequenciadas por pirosequenciamento para descobrir a mutação no gene Pfcrt no códon K76T e em pfmdr1 nos códons N86Y, Y184F, N1042D e D1246Y. De 121 microscopias de casos positivos de P. falciparum, 109 amostras foram positivas para P. falciparum por nested PCR. A mutação Pfcrt K76T foi encontrada em 96% dos isolados, a mutação Pfmdr1 N86Y foi observada em 20% e 11% abrigou a mutação Y184F. Todas as amostras eram do tipo selvagem para o códon N1042D e D1246Y de Pfmdr1. No FATA, Paquistão, a frequência do alelo resistente 76T permaneceu alta apesar da remoção de CQ. No entanto, as descobertas atuais do estudo sugerem a fixação completa do genótipo resistente a CQ de P. falciparum na área de estudo.

Frequency, distribution and determinants of Helicobacter pylori infection in adults and adolescents with gastric symptoms: cross-sectional epidemiological inquiry in district Haripur, Pakistan.

Helicobacter pylori (HP) is a vital element in the etiology of peptic ulcers and gastric cancer. This research aimed to determine the frequency, distribution, and determinants of HP infection in adults and adolescents with gastric symptoms in district Haripur, Khyber Pakhtunkhwa, Pakistan. This cross-sectional study was performed from June 2018 to June 2020 at the Medical Laboratory Technology Department, The University of Haripur, Pakistan. Presence of HP was a research variable, while sex, age groups, education status, overcrowding, dining habits, milk intake, drinking water source and animal contact were grouping variables. Immuno-chromatographic technique (ICT) was used to for serological detection of HP antibodies. All variables were represented by frequency and percentage with 95%CI. Prevalence of HP and its distribution by eight socio-demographic variables was testified by the chi-square goodness-of-fit test while association was testified by chi-square test of association. Out of total 1160 cases, 557 (48%) were positive for HP. Population prevalence was higher in men, in the age group 20-40 years, illiterate, family size ≤ 10 persons, taking restaurant food, using tetra pack, using municipal water, and having animal contact. The observed prevalence of HP was similar to its expected prevalence in the population. The observed distribution of HP in the sample was different from its expected distribution in population by eight socio-demographic variables. Presence of HP was associated with all eight socio-demographic variables besides age groups.

Cytogenetic abnormalities in patients with hematological malignancies in Lahore city, Pakistan.

Hematological and hematopoietic cells malignancies of the genes and hematopoietic cells are associated with the genetic mutation, often at the chromosomal level. The standard cytogenetic study is widely accepted as one of the main diagnostics and prognostic determinants in patients. Therefore, the current descriptive and cross-sectional study sought to determine the cytogenetic analysis of frequent hematological malignancies in Pakistan. A total of 202 peripheral bone marrow or blood samples from patients with benign and malignant hematological malignancy were taken using a conventional G-banding technique. Among enrolled patients, the mean age was 21.5 years ± 23.4, and gender-wise distribution showed a marked predominance of the male 147 (73%) population compared to the female 55 (27%). Patients in the age group (2-10 years) had the highest frequency, 48 (24%), of hematological neoplasms, followed by age (11-20 years) with 40 (20%). Normal karyotypes (46, XX/46, XY) was found in 51% (n=103) patients. Furthermore, the frequency of complex karyotype was 30 (15%), while normal was seen in 171 (85%) patients. Pre-B Acute Lymphoblastic Leukemia (Pre-B ALL) was the most prevalent malignancy of 66 (33%), followed by Chronic Myelogenous Leukemia (CML) of 41 (20%) and Acute Lymphocytic Leukemia of 29 (14%). Translocation was the most prevalent 50 (25%), followed by hypotriploidy 14 (7%) and monosomy 8 (4%) on chromosome aberration analysis. In addition, t(9:22) translocation was found to be 20 (10%) in CML, with the majority in the age group (31-40 years). This study recommends that karyotyping should be tested frequently in hematological conditions because it may provide insight into the relative chromosomal changes associated with particular malignancies.

Molecular Epidemiology of Plasmodium species in Conflicted Federally Administered Tribal Area (FATA) Pakistan.

Military conflicts have been significant obstacles in detecting and treating infectious disease diseases due to the diminished public health infrastructure, resulting in malaria endemicity. A variety of violent and destructive incidents were experienced by FATA (Federally Administered Tribal Areas). It was a struggle to pursue an epidemiological analysis due to continuing conflict and Talibanization. Clinical isolates were collected from Bajaur, Mohmand, Khyber, Orakzai agencies from May 2017 to May 2018. For Giemsa staining, full blood EDTA blood samples have been collected from symptomatic participants. Malaria-positive microscopy isolates were spotted on filter papers for future Plasmodial molecular detection by nested polymerase chain reaction (nPCR) of small subunit ribosomal ribonucleic acid (ssrRNA) genes specific primers. Since reconfirming the nPCR, a malariometric study of 762 patients found 679 positive malaria cases. Plasmodium vivax was 523 (77%), Plasmodium falciparum 121 (18%), 35 (5%) were with mixed-species infection (P. vivax plus P. falciparum), and 83 were declared negative by PCR. Among the five agencies of FATA, Khyber agency has the highest malaria incidence (19%) with followed by P. vivax (19%) and P. falciparum (4.1%). In contrast, Kurram has about (14%), including (10.8%) P. vivax and (2.7%) P. falciparum cases, the lowest malaria epidemiology. Surprisingly, no significant differences in the distribution of mixed-species infection among all five agencies. P. falciparum and P. vivax were two prevalent FATA malaria species in Pakistan's war-torn area. To overcome this rising incidence of malaria, this study recommends that initiating malaria awareness campaigns in school should be supported by public health agencies and malaria-related education locally, targeting children and parents alike.

Comparison of different diagnostic modalities for isolation of Mycobacterium Tuberculosis among suspected tuberculous lymphadenitis patients.

Tuberculosis is a communicable disease with high morbidity and mortality rates in developing countries. The study's primary objective is to compare conventional methods such as acid-fast bacillus (AFB) culture and microscopy with rapid diagnostic methods. The secondary objective is to compare histopathological and microbiological findings in suspected patients with tubercular lymphadenitis. A total of 111 samples (August 2018 to September 2019) of lymph nodes were processed for AFB microscopy, AFB cultures, drug-susceptibility testing (DST), histopathology, and Xpert Mycobacterium Tuberculosis (MTB)/resistance to Rifampin (RIF) assays. Out of 111 lymph node samples, 6 (5.4%) were positive for AFB smear microscopy, 84 (75.6%) were positive for AFB culture, 80 (70.7%) were positive on Gene Xpert, and 102 (91.8%) were indicative of tuberculosis for histopathology studies. Mycobacteria growth indicator tube (MGIT) culture positivity was 84 (75.6%) higher than solid Lowenstein-Jensen (LJ) culture 74 (66.6%). Positive cultures underwent phenotypic DST. Two cases were Multidrug-resistant (MDR) on DST, while three cases were Rifampicin resistant on Gene Xpert. The sensitivity of Genexpert was (62%) against the conventional AFB culture method. The poor performance of conventional lymphadenitis diagnostic methods requires early and accurate diagnostic methodology. Xpert MTB/RIF test can help in the treatment of multidrug-resistant TB cases. Nonetheless, rapid and conventional methods should be used for complete isolation of Mycobacterium tuberculosis.

Fixation of pfcrt chloroquine resistance alleles in Plasmodium falciparum clinical isolates collected from unrest tribal agencies of Pakistan.

Plasmodium falciparum resistance to Chloroquine (CQ) is a significant cause of mortality and morbidity worldwide. There is a paucity of documented data on the prevalence of CQ-resistant mutant haplotypes of Pfcrt and Pfmdr1 genes from malaria-endemic war effected Federally Administered Tribal Areas of Pakistan. The objective of this study was to investigate the prevalence of P. falciparum CQ-resistance in this area. Clinical isolates were collected between May 2017 and May 2018 from North Waziristan and South Waziristan agencies of Federally Administrated Trial Area. Subsequently, Giemsa-stained blood smears were examined to detect Plasmodium falciparum. Extraction of malarial DNA was done from microscopy positive P. falciparum samples, and P. falciparum infections were confirmed by nested PCR (targeting Plasmodium small subunit ribosomal ribonucleic acid (ssrRNA) genes). All PCR confirmed P. falciparum samples were sequenced by pyrosequencing to find out mutation in Pfcrt gene at codon K76T and in pfmdr1 at codons N86Y, Y184F, N1042D, and D1246Y. Out of 121 microscopies positive P. falciparum cases, 109 samples were positive for P. falciparum by nested PCR. Pfcrt K76T mutation was found in 96% of isolates, Pfmdr1 N86Y mutation was observed in 20%, and 11% harboured Y184F mutation. All samples were wild type for Pfmdr1 codon N1042D and D1246Y. In the FATA, Pakistan, the frequency of resistant allele 76T remained high despite the removal of CQ. However, current findings of the study suggest complete fixation of P. falciparum CQ-resistant genotype in the study area.

Causes of childhood deafness in Pukhtoonkhwa Province of Pakistan and the role of consanguinity.

BACKGROUND: Deafness is the hidden disability of childhood, and leads to poor educational and employment prospects. There is little published information on deafness in Pakistan. Profound hearing impairment is more prevalent in countries where consanguineous marriages are common, such as Pakistan. This study aimed to assess causes of childhood deafness and association with parental consanguinity, within deaf and hearing children in the Peshawar district of Pukhtoonkhwa Province, Pakistan. METHODS: One hundred and forty deaf children were identified from two schools for deaf children within the Peshawar district. These children were assessed via audiology, otoscopic examination, case note review and parental history, in order to attempt to ascertain the cause of their deafness. Two hundred and twenty-one attendees at a local immunisation clinic (taken as representative of the local childhood population) were also screened for hearing impairment. Parents of both groups of children were assessed by interview and questionnaire in order to ascertain the mother and father's family relationship (i.e. whether cousins or unrelated). RESULTS: Of the 140 deaf school pupils, 92.1 per cent were profoundly hearing impaired and 7.9 per cent were severely hearing impaired. All these children had bilateral sensorineural hearing loss. A possible cause of deafness was identified in only six of these children. Parental consanguinity (i.e. first or second cousins) was established for 86.4 per cent of deaf school pupils and 59.7 per cent of immunisation clinic attendees. None of the control children were identified as having a hearing problem. CONCLUSION: The prevalence of parental consanguinity was significantly higher in deaf children compared with non-hearing impaired children. However, the study also confirmed a high rate of consanguinity within the general Peshawar community. In this setting, prevention of consanguineous unions is the only means of reducing levels of congenital hearing impairment. The current levels of hearing disability represent both a prominent public health problem and an important, potentially preventable childhood disability.