Genotyping of Pneumocystis jirovecii isolates obtained from clinical samples by multilocus sequencing: a molecular epidemiology study conducted in Turkey

Sürgeç, Ecem; Can, Hüseyin; Döşkaya, Mert; Karakavuk, Muhammet; Şahar, Esra Atalay; Döşkaya, Aysu Değirmenci; Pullukçu, Hüsnü; Taşbakan, Meltem; Taşbakan, Mehmet Sezai; Akyol, Deniz; Zihni, Figen Yargucu; Ün, Cemal; Gürüz, Adnan Yüksel; Demir, Samiye

doi:10.1007/s00203-020-01874-3

Cited by 4 publications

(1 citation statement)

References 20 publications

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“…( Acanthamoeba sp. strain DP/5A with T4 genotype), Giardia intestinalis (microscopy confirmed positive sample), Escherichia coli (BL21 strain) , P. jirovecii (PCR confirmed positive control sample) [ 25 ], and Trichomonas vaginalis (microscopy confirmed positive control sample) [ 26 ]. These DNA samples were investigated by “Rapid-Crypto Colorimetric LAMP test” as described in Sect. "…”

Section: Methodsmentioning

confidence: 99%

Development of a “Rapid-Crypto Colorimetric LAMP Test” to Detect Cryptosporidiosis in Feces of Newborns Calves

Karakavuk,

Can,

Can

et al. 2024

Acta Parasit.

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Background Cryptosporidiosis is a disease that causes major intestinal damage in humans and animals. The causative agents of the disease are Cryptosporidium species. In newborn calves, diarrhea can lead to death, resulting in significant economic losses for the farms. Therefore, accurate, rapid, and cost-effective diagnosis of the disease is very important. Material and methods In this study, a novel colorimetric loop-mediated isothermal amplification (LAMP) test named “Rapid-Crypto Colorimetric LAMP test” targeting Cryptosporidium spp. 18S rRNA gene was developed to detect cryptosporidiosis in the feces of newborn calves. The analytical sensitivity of the test was determined by plasmid controls. Clinical sensitivity was determined using the feces of 127 calves collected from farms in İzmir and Manisa provinces. All of the samples were also investigated with Real-Time PCR targeting the Cryptosporidium spp. COWP gene. Cross-reactivity was tested using the DNA of other parasites and bacteria. Results According to the results, the analytical sensitivity of the “Rapid-Crypto Colorimetric LAMP test” was found as 1 copy plasmid/reaction. When the results were compared with the Real-Time PCR test, the sensitivity of the “Rapid-Crypto Colorimetric LAMP test” was 100% and the specificity was 97.4%. The test did not cross-react with other parasites and bacteria. Conclusion The “Rapid-Crypto Colorimetric LAMP test” developed in this study provides an advantage in the diagnosis of Cryptosporidium spp. in calf stool samples since it can be applied in basic laboratories or in the field, does not require experienced personnel, and has high sensitivity. Moreover, diagnosis can be made with the naked eye without using any device.

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Section: Methodsmentioning

confidence: 99%

Development of a “Rapid-Crypto Colorimetric LAMP Test” to Detect Cryptosporidiosis in Feces of Newborns Calves

Karakavuk,

Can,

Can

et al. 2024

Acta Parasit.

Self Cite

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Genetic diversity of Pneumocystis jirovecii isolates among Turkish population based on mitochondrial large subunit ribosomal RNA and dihydropteroate synthase gene typing

et al. 2021

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Pneumocystis jirovecii (P. jirovecii) is an atypical fungus that can cause severe interstitial pneumonia in immunocompromised patients. In this study, mitochondrial large subunit ribosomal RNA (mtLSU-rRNA) and dihydropteroate synthase (DHPS) gene polymorphism in P. jirovecii isolates were investigated in Western Turkey's Izmir province and its surroundings. For this purpose, a total of 157 P. jirovecii isolates obtained from bronchoalveolar lavage samples of hospitalized cases and lung tissue samples of autopsy cases who died outside hospital were examined. Genotypes were identified by direct sequencing of mtLSU-rRNA restriction fragment length polymorphism analysis of the DHPS gene amplicons. The mtLSU-rRNA analysis revealed that genotype 2 was the most common genotype with 58%. The following genotypes were genotype 3 (13%), genotype 1 (11.6%) and genotype 4 (5.1%), while genotype 5 (0.7%) was detected in only one autopsy case. In addition, 16 (11.6%) cases had dual or triple different genotypes (mixed infection). It was observed that the genotype distribution was not affected by characteristics such as age, gender and immune status. However, the predominance of genotype 2 in solid organ tumors and the predominance of mixed infection in patients with chronic pulmonary disease were statistically significant. On the other hand, DHPS gene amplification was positive in 137 (87.3%) of 157 samples. While no mutation was observed in 135 samples, the association of wild-type and 57th codon mutation was detected in two hospitalized cases (1.5%). In this study, important epidemiological data on the distribution of mtLSU-rRNA genotypes were obtained. Also the existence of DHPS gene mutations associated with potential drug resistance in our community was shown for the first time. Further studies are needed to evaluate the possible effects of genotypes on the prognosis of the disease to help with the clinician's treatment decisions. Lay abstract Pneumocystis jirovecii (P. jirovecii) is an atypical fungus that can cause life-threatening pneumonia in immunocompromised patients. In this study, we investigated the mtLSU-rRNA and DHPS gene polymorphisms in P. jirovecii isolates from both hospital and autopsy cases.

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Pneumocystis jirovecii genetic diversity in a Spanish tertiary hospital

et al. 2021

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Pneumocystis jirovecii is associated with non-noxious colonization or severe pneumonia in immunocompromised hosts. Epidemiological investigations have been hampered by the lack of a standardized typing scheme. Thus, only partial molecular data on Spanish P. jirovecii cases are available. Recently a new ISHAM consensus multilocus sequence typing scheme (MLST) targeting β-TUB, mt26S, CYB, and SOD with a publicly accessible database has been launched to overcome this problem. The molecular epidemiology of P. jirovecii from immunocompromised patients either colonized (n = 50) or having pneumonia (n = 36) seen between 2014 and 2018 at a single center in Barcelona, Spain, was studied. The new ISHAM consensus MSLT scheme was used to investigate the local epidemiology and identify possible unnoticed outbreaks. Mutations in the DHPS gene, not included in the scheme but giving information about potential sulpha treatment failure, were also studied. The study assigned 32 sequence types (ST) to 72.2% pneumonia and 56% colonization cases. The most frequent STs were ST21 (18.5%), ST22 (14.8%), and ST37(14.8%). For non-unique STs, ST3, ST30 and ST31 were found only in pneumonia cases, whereas ST27 was associated exclusively to colonization's. Despite 38 patients sharing similar STs, only two were involved in a potential cross transmission event. No DHPS mutations were identified. The new consensus typing scheme was useful to ascertain the molecular epidemiology of P. jirovecii in our center revealing a high genetic diversity and the potential association of specific STs to colonization and pneumonia cases. Lay Summary A newly described MLST scheme aims at providing a standardized tool to study and compare Pneumocystis jirovecii epidemiology. A high diversity amongst P. jirovecii isolates from patients in Barcelona, Spain, and a potential association between specific STs and infection/colonization were identified.

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Genotyping of Pneumocystis jirovecii isolates obtained from clinical samples by multilocus sequencing: a molecular epidemiology study conducted in Turkey

Cited by 4 publications

References 20 publications

Development of a “Rapid-Crypto Colorimetric LAMP Test” to Detect Cryptosporidiosis in Feces of Newborns Calves

Development of a “Rapid-Crypto Colorimetric LAMP Test” to Detect Cryptosporidiosis in Feces of Newborns Calves

Genetic diversity of Pneumocystis jirovecii isolates among Turkish population based on mitochondrial large subunit ribosomal RNA and dihydropteroate synthase gene typing

Pneumocystis jirovecii genetic diversity in a Spanish tertiary hospital

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