Genetic relatedness among Trypanosoma evansi stocks by random amplification of polymorphic DNA and evaluation of a synapomorphic DNA fragment for species-specific diagnosis

Introduction: Klebsiella pneumoniae is a common cause of nosocomial infections; however, there is limited information in Iran regarding nosocomial outbreaks due to extended-spectrum &#946;–lactamase (ESBL) producing K pneumoniae strains, particularly using molecular methods. The present study focused on the molecular mechanism of ESBL resistance and genetic relatedness in K. pneumoniae isolates causing nosocomial infections in an Iranian referral hospital. Material and Methods: This study evaluated the antimicrobial resistance and molecular epidemiology of K. pneumoniae causing nosocomial infections in children between October 2013 and March 2014. The ESBL detection was carried out for all the isolates by the CLSI method and PCR was carried out for the detection of the blaSHV, blaTEM, and blaCTX-M genes among ESBL-producing K. pneumonia. Molecular typing of the K. pneumoniae was performed using random amplification of polymorphic DNA-polymerase chain reaction (RAPD-PCR). Results: A total of 30 isolates of K. pneumoniae were used for epidemiological analysis. High rates of resistance to cefotaxime (n=29, 97%), cefazolin (n=29, 97%), cefepime (n=25, 83%) and gentamicin (n=23, 77%) were observed. A total of 29 strains (97%) produced ESBLs. The frequency of blaSHV, blaCTX-M and blaTEM genes among these isolates was 83% (n=25), 70% (n=21) and 57% (n=17), respectively. Surprisingly 11 isolated (37%) carried blaSHV, blaCTX-M and blaTEM genes simultaneously. Moreover, the concurrent presence of “blaSHV and blaCTX-M” and “blaSHV and blaTEM” was seen in 8 (27%) and 4 (13%) isolates, respectively. RAPDPCR analyses revealed that K. pneumoniae isolates belonged to 2 RAPD-PCR types among which one cluster counted for 28 isolates. Conclusion: To our knowledge, this is the first published report of a nosocomial outbreak of ESBL-producing K. pneumoniae in children in Iran. Although the epidemiology of nosocomial infections with ESBL-producing organisms has not yet been explored in depth in Iran, our findings suggest that ESBL-producing organisms are already an established public health threat in our country.

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Strain Typing of Trichosporon asahii Clinical Isolates by Random Amplification of Polymorphic DNA (RAPD) Analysis

Journal of Laboratory Physicians ◽

10.1055/s-0041-1731111 ◽

2021 ◽

Author(s):

Thayanidhi Premamalini ◽

Vijayaraman Rajyoganandh ◽

Ramaraj Vijayakumar ◽

Hemanth Veena ◽

Anupma Jyoti Kindo ◽

...

Keyword(s):

Genetic Relatedness ◽

Rapd Analysis ◽

Clinical Isolates ◽

Clinical Samples ◽

Strain Typing ◽

Trichosporon Asahii ◽

Pcr Fingerprinting ◽

Specific Pcr ◽

Rapd Primer ◽

Random Amplification

Abstract Objective The aim of this study was to identify and isolate Trichosporon asahii (T. asahii) from clinical samples and to assess the genetic relatedness of the most frequently isolated strains of T. asahii using random amplification of polymorphic DNA (RAPD) primers GAC-1 and M13. Methods All the clinical samples that grew Trichosporon species, identified and confirmed by polymerase chain reaction (PCR) using Trichosporon genus-specific primers, were considered for the study. Confirmation of the species T. asahii was carried out by T. asahii-specific PCR. Fingerprinting of the most frequently isolated T. asahii isolates was carried out by RAPD using random primers GAC-1 and M13. Results Among the 72 clinical isolates of Trichosporon sp. confirmed by Trichosporon-specific PCR, 65 were found to be T. asahii as identified by T. asahii-specific PCR. Fingerprinting of the 65 isolates confirmed as T. asahii using GAC-1 RAPD primer yielded 11 different patterns, whereas that of M13 primer produced only 5 patterns. The pattern I was found to be the most predominant type (29.2%) followed by pattern III (16.9%) by GAC-1 primer. Conclusions This study being the first of its kind in India on strain typing of T. asahii isolates by adopting RAPD analysis throws light on genetic diversity among the T. asahii isolates from clinical samples. Fingerprinting by RAPD primer GAC-1 identified more heterogeneity among the T. asahii isolates than M13.

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Molecular Diversity and Genetic Relatedness of Candida albicans Isolates from Birds in Hungary

Mycopathologia ◽

10.1007/s11046-021-00527-3 ◽

2021 ◽

Vol 186 (2) ◽

pp. 237-244

Author(s):

M. Domán ◽

L. Makrai ◽

Gy. Lengyel ◽

R. Kovács ◽

L. Majoros ◽

...

Keyword(s):

Candida Albicans ◽

Molecular Diversity ◽

Genetic Relatedness ◽

Group Method ◽

Pair Group ◽

Degree Of Separation ◽

Avian Origin ◽

Species Specific ◽

Sequence Types ◽

Unweighted Pair Group Method

AbstractThe molecular epidemiology of Candida albicans infections in animals has been rarely studied. In this study, multilocus sequence typing was used to characterise the genetic diversity and population structure of 24 avian origin C. albicans isolates collected from different birds with candidiasis and compared to human isolates. Fourteen diploid sequence types (DSTs) including six new DSTs were determined. Cluster analysis revealed that isolates grouped into 8 clades. Bird isolates mainly belonged to minor clades and Clade 15 with DST 172 was the most common (11 isolates; 45.8%). The remaining isolates were clustered into Clade 7 (5 isolates; 20.8%), Clade 10 (4 isolates; 16.6%), Clade 8 (2 isolates; 8.3%), Clade 4 (1 isolate; 4.2%) and Clade 16 (1 isolate; 4.2%). Unweighted pair group method with arithmetic averages (UPGMA) and eBURST analyses showed that the genetic construction of avian origin C. albicans population is fairly diverse. Although species-specific lineages were not found, some degree of separation in the evolution of bird and human strains could be observed.

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Characterization of a species-specific DNA fragment originating from theCandida albicansmitochondrial genome

Medical Mycology ◽

10.1080/02681219480000101 ◽

1994 ◽

Vol 32 (1) ◽

pp. 71-75 ◽

Cited By ~ 3

Author(s):

Y. Miyakawa ◽

T. Mabuchi

Keyword(s):

Dna Fragment ◽

Species Specific

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Hospital Specificity, Region Specificity, and Fluconazole Resistance of Candida albicans Bloodstream Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.36.6.1518-1529.1998 ◽

1998 ◽

Vol 36 (6) ◽

pp. 1518-1529 ◽

Cited By ~ 57

Author(s):

M. A. Pfaller ◽

S. R. Lockhart ◽

C. Pujol ◽

J. A. Swails-Wenger ◽

S. A. Messer ◽

...

Keyword(s):

United States ◽

Drug Resistance ◽

Candida Albicans ◽

Genetic Relatedness ◽

Computer Assisted ◽

Specific Probe ◽

Fluconazole Resistance ◽

Fluconazole Resistant ◽

Species Specific ◽

Close Genetic Relationship

In a survey of bloodstream infection (BSI) isolates across the continental United States, 162 Candida albicans isolates were fingerprinted with the species-specific probe Ca3 and the patterns were analyzed for relatedness with a computer-assisted system. The results demonstrate that particular BSI strains are more highly concentrated in particular geographic locales and that established BSI strains are endemic in some, but not all, hospitals in the study and undergo microevolution in hospital settings. The results, however, indicate no close genetic relationship among fluconazole-resistant BSI isolates in the collection, either from the same geographic locale or the same hospital. This study represents the first of three fingerprinting studies designed to analyze the origin, genetic relatedness, and drug resistance of Candida isolates responsible for BSI.

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Molecular detection of equine trypanosomiasis in the Riyadh Province of Saudi Arabia

Journal of Veterinary Diagnostic Investigation ◽

10.1177/1040638718798688 ◽

2018 ◽

Vol 30 (6) ◽

pp. 942-945 ◽

Cited By ~ 3

Author(s):

Abdullah D. Alanazi ◽

Robert Puschendorf ◽

Bashir Salim ◽

Mohamed S. Alyousif ◽

Ibrahim O. Alanazi ◽

...

Keyword(s):

Saudi Arabia ◽

Internal Transcribed Spacer ◽

Molecular Detection ◽

Cross Sectional Study ◽

Trypanosoma Evansi ◽

Type B ◽

Sectional Study ◽

Internal Transcribed Spacer Region ◽

Cross Sectional ◽

Species Specific

We conducted a cross-sectional study to detect trypanosome infections of horses and donkeys in the Riyadh Province of Saudi Arabia. DNA was extracted from blood samples collected from 368 horses and 142 donkeys, and subjected to universal first ribosomal internal transcribed spacer region (ITS1)-PCR followed by Trypanosoma evansi species–specific RoTat1.2-PCR. The universal ITS1-PCR revealed T. evansi infection in horses ( n = 12; 3.3%) and donkeys ( n = 4; 2.8%). There was no significant effect of sex or age on the prevalence of trypanosomiasis in horses or donkeys. Application of the RoTat1.2-PCR revealed that the RoTat1.2 VSG gene was absent from the positive ITS1-PCR samples of 3 horses and 1 donkey. This discrepancy could be explained by the circulation of T. evansi type B in Saudi Arabia; however, this suspicion requires confirmation.

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Identification of Members of the Burkholderia cepacia Complex by Species-Specific PCR

Journal of Clinical Microbiology ◽

10.1128/jcm.38.8.2962-2965.2000 ◽

2000 ◽

Vol 38 (8) ◽

pp. 2962-2965 ◽

Cited By ~ 33

Author(s):

Paul W. Whitby ◽

Karen B. Carter ◽

Kenneth L. Hatter ◽

John J. LiPuma ◽

Terrence L. Stull

Keyword(s):

Burkholderia Cepacia ◽

Growth Characteristics ◽

Rrna Operon ◽

Burkholderia Cepacia Complex ◽

Pcr Analysis ◽

Biochemical Tests ◽

Specific Pcr ◽

Dna Fragment ◽

Species Specific ◽

Phenotypic Tests

Definitive identification of the species in the Burkholderia cepacia complex by routine clinical microbiology methods is difficult. Phenotypic tests to identify B. multivorans andB. vietnamiensis have been established; more recent work indicates B. stabilis may also be identified by growth characteristics and biochemical tests. However, attempts to identify genomovars I and III have, thus far, proved unsuccessful. Previously, we demonstrated the utility of two primer pairs, directed to the rRNA operon, to specifically identify the B. cepacia complex in a PCR. One of these primer pairs, G1-G2, only amplified a DNA fragment from genomovars I and III and B. stabilis in a PCR with genomic DNA isolated from prototypical strains representing the five genomovars. Sequence analysis of the rRNA operon for all the genomovars indicated that this primer pair targeted a region shared by these isolates. Further analysis revealed a region of heterogeneity between genomovar III and B. stabilis internal to the amplified product of G1-G2. Primers designed to target this region were tested with prototypical strains following an initial amplification with the G1-G2 primer pair. New primers specific for the prototypical genomovar III and B. stabilis were designated SPR3 and SPR4, respectively. Analysis of 93 isolates representing 18 genomovar I, 13B. multivorans, 36 genomovar III, 11 B. stabilis, and 15 B. vietnamiensis isolates was performed. DNA from all isolates of genomovars I and III and B. stabilis was amplified by G1-G2. Genomovar III isolates yielded a product with SPR3/G1 while B. stabilis amplified with SPR4-G1. Genomovar I isolates were amplified by either SPR3-G1 or SPR4-G1, but not both. B. multivorans yielded a product with SPR3-G1 but not G1-G2, and B. vietnamiensis isolates were negative in all PCRs. Thus using an algorithm with G1-G2, SPR3-G1, and SPR4-G1 primers in a PCR analysis, genomovar III isolates can be separated from B. stabilis and the identity of B. multivorans and B. vietnamiensis can be confirmed.

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