Cryptococcosis is a fungal disease affecting more than one million people per year worldwide. Its... more Cryptococcosis is a fungal disease affecting more than one million people per year worldwide. Its main etiological agents are Cryptococcus neoformans species complex and Cryptococcus gattii species complex. Cryptococcal meningitis (CM) is considered an AIDS-defining condition. Rapid diagnosis by cryptococcal antigen assays, either the latex agglutination test (LA) or the lateral flow assay, is key to decreasing mortality due to cryptococcal disease. The aim of the study was to develop a latex agglutination reagent (LA-ANLIS) for the rapid and reliable diagnosis of cryptococcosis in Argentina. This reagent will be produced in order to supply the NMLN (National Mycology Laboratory Network). The evaluation of LA-ANLIS performance and its comparison with the Cryptococcus Antigen Latex Agglutination Test System (LA-IMMY) (Immuno-Mycologics, Inc., USA) were conducted in 94 samples of cerebrospinal fluid. LA-ANLIS and LA-IMMY compared exhibited 100% positive agreement and 97% negative agreement. LA-ANLIS showed 94% sensitivity and 97% specificity with the positive and negative predictive values of 94% and 97%, respectively. The LA-ANLIS is a reliable, reproducible and cost-effective reagent, especially useful in countries where the commercial kit is not generally available and must be obtained at a high cost. National production of reagents is the best choice for a reliable access to the rapid diagnosis of CM in Argentina.
New technologies for rapid detection and drug susceptibility testing of mycobacteria are needed i... more New technologies for rapid detection and drug susceptibility testing of mycobacteria are needed in clinical laboratories in order to attempt a rapid diagnosis and effective treatment of tuberculosis (TB) specially of cases resistant to isoniazid (H) and rifampin (RMP) (MDR-TB). A total of 218 pulmonary specimens from 132 HIV coinfected patients were processed and inoculated into the Mycobacterial Growth Indicator Tube system (MGIT, Becton Dickinson, MD) and on Lowenstein-Jensen (L-J) and Stonebrink (SB) solid media. The average time for recovering Mycobacterium tuberculosis (MTB) from MGIT was 18.3 days and 31.0 days from solid media. Of the patients 14.4% (19/132) were only diagnosed by MGIT. In another experiment susceptibility tests by the classical proportion method (PM) in L-J medium with H, RMP, streptomycin (SM), para-aminosalycilic-acid and ethambutol (EMB) were carried out on 120 isolates. The results were later compared with those obtained by determining the minimal inhibi...
Microscopy with the Ziehl-Neelsen (ZN) stain is frequently negative in paucibacillary tuberculosi... more Microscopy with the Ziehl-Neelsen (ZN) stain is frequently negative in paucibacillary tuberculosis (TB) so that the treatment must be started and continued until the culture results confirm or not the disease. LCx Mycobacterium tuberculosis Assay (Abbott, Lab.) uses the ligase chain reaction for direct amplification of DNA and rapid detection of M. tuberculosis Complex (MTB) in clinical specimens. We evaluated the usefulness of the LCx assay on 1,203 clinical samples: 737 respiratory and 466 extrapulmonary specimens. The LCx results were compared with those obtained by ZN and cultures on solid media and Mycobacteria Growth Indicator Tube (MGIT, Becton Dickinson, Argentina). Since detection and identification of MTB are simultaneously made by the LCx assay, a total of 145 out of 183 patients (79.2%) had a confirmed TB diagnosis in two working days. Positive culture results were predicted in 122 out of 160 cases (76.3%) by LCx and in 70 (43.8%) by ZN as well. The sensitivity (S) and s...
In the city of Buenos Aires, Argentina, Cryptococcus gattii genotype AFLP4/VGI was found to be as... more In the city of Buenos Aires, Argentina, Cryptococcus gattii genotype AFLP4/VGI was found to be associated with decaying wood in hollows of different tree species. The aim of this study was to investigate the presence of C. gattii in the environment of riverside cities of the river Paraná, and to describe its serotypes and molecular types. Five hundred samples were collected in 50 parks by swabbing tree hollows. The samples were inoculated on caffeic acid agar supplemented with chloramphenicol, and incubated at 28 °C for 1 week with a daily observation. The isolates were identified by conventional methods. The serotype was determined by slide agglutination with specific antisera. Molecular typing was carried out by PCR-RFLP of the URA5 gene. Four isolates of C. gattii were recovered: Cryptococcus gattii serotype B, genotype AFLP4/VGI, isolated from Eucalyptus sp. in the city of Rosario and from Grevillea robusta in the city of La Paz; and C. gattii serotype C, genotype AFLP5/VGIII, isolated from two different Tipuana tipu trees in the city of Resistencia. Here, we report for the first time the isolation of C. gattii serotype C, genotype AFLP5/VGIII, from environmental samples in Argentina.
The bacillus Calmette-Guérin (BCG) was obtained in 1920 after successive passages leading to the ... more The bacillus Calmette-Guérin (BCG) was obtained in 1920 after successive passages leading to the attenuation of a Mycobacterium bovis strain. For the following 40 years, BCG had been replicated, resulting in substrains with genotypic and phenotypic differences. Several genomic studies have compared two BCG strains, M. bovis and Mycobacterium tuberculosis, and observed that deleted regions in the different strains could be related to differences in antigenic properties. In this work, a working seed lot was obtained from a lyophilized secondary seed lot from the BCG Pasteur strain 1173 P2 and genetically characterized. The genome was analyzed by PCR directed to five regions (RD1, RD2, RD14, RD15, DU2), using the seed lot and different available strains as templates. No genetic differences were found in the fragments studied as compared to the Pasteur strain. A total of 20 passages were carried out and no differences were found in the size of the fragments amplified by PCR. In conclusi...
Cryptococcosis is a fungal disease affecting more than one million people per year worldwide. Its... more Cryptococcosis is a fungal disease affecting more than one million people per year worldwide. Its main etiological agents are Cryptococcus neoformans species complex and Cryptococcus gattii species complex. Cryptococcal meningitis (CM) is considered an AIDS-defining condition. Rapid diagnosis by cryptococcal antigen assays, either the latex agglutination test (LA) or the lateral flow assay, is key to decreasing mortality due to cryptococcal disease. The aim of the study was to develop a latex agglutination reagent (LA-ANLIS) for the rapid and reliable diagnosis of cryptococcosis in Argentina. This reagent will be produced in order to supply the NMLN (National Mycology Laboratory Network). The evaluation of LA-ANLIS performance and its comparison with the Cryptococcus Antigen Latex Agglutination Test System (LA-IMMY) (Immuno-Mycologics, Inc., USA) were conducted in 94 samples of cerebrospinal fluid. LA-ANLIS and LA-IMMY compared exhibited 100% positive agreement and 97% negative agreement. LA-ANLIS showed 94% sensitivity and 97% specificity with the positive and negative predictive values of 94% and 97%, respectively. The LA-ANLIS is a reliable, reproducible and cost-effective reagent, especially useful in countries where the commercial kit is not generally available and must be obtained at a high cost. National production of reagents is the best choice for a reliable access to the rapid diagnosis of CM in Argentina.
New technologies for rapid detection and drug susceptibility testing of mycobacteria are needed i... more New technologies for rapid detection and drug susceptibility testing of mycobacteria are needed in clinical laboratories in order to attempt a rapid diagnosis and effective treatment of tuberculosis (TB) specially of cases resistant to isoniazid (H) and rifampin (RMP) (MDR-TB). A total of 218 pulmonary specimens from 132 HIV coinfected patients were processed and inoculated into the Mycobacterial Growth Indicator Tube system (MGIT, Becton Dickinson, MD) and on Lowenstein-Jensen (L-J) and Stonebrink (SB) solid media. The average time for recovering Mycobacterium tuberculosis (MTB) from MGIT was 18.3 days and 31.0 days from solid media. Of the patients 14.4% (19/132) were only diagnosed by MGIT. In another experiment susceptibility tests by the classical proportion method (PM) in L-J medium with H, RMP, streptomycin (SM), para-aminosalycilic-acid and ethambutol (EMB) were carried out on 120 isolates. The results were later compared with those obtained by determining the minimal inhibi...
Microscopy with the Ziehl-Neelsen (ZN) stain is frequently negative in paucibacillary tuberculosi... more Microscopy with the Ziehl-Neelsen (ZN) stain is frequently negative in paucibacillary tuberculosis (TB) so that the treatment must be started and continued until the culture results confirm or not the disease. LCx Mycobacterium tuberculosis Assay (Abbott, Lab.) uses the ligase chain reaction for direct amplification of DNA and rapid detection of M. tuberculosis Complex (MTB) in clinical specimens. We evaluated the usefulness of the LCx assay on 1,203 clinical samples: 737 respiratory and 466 extrapulmonary specimens. The LCx results were compared with those obtained by ZN and cultures on solid media and Mycobacteria Growth Indicator Tube (MGIT, Becton Dickinson, Argentina). Since detection and identification of MTB are simultaneously made by the LCx assay, a total of 145 out of 183 patients (79.2%) had a confirmed TB diagnosis in two working days. Positive culture results were predicted in 122 out of 160 cases (76.3%) by LCx and in 70 (43.8%) by ZN as well. The sensitivity (S) and s...
In the city of Buenos Aires, Argentina, Cryptococcus gattii genotype AFLP4/VGI was found to be as... more In the city of Buenos Aires, Argentina, Cryptococcus gattii genotype AFLP4/VGI was found to be associated with decaying wood in hollows of different tree species. The aim of this study was to investigate the presence of C. gattii in the environment of riverside cities of the river Paraná, and to describe its serotypes and molecular types. Five hundred samples were collected in 50 parks by swabbing tree hollows. The samples were inoculated on caffeic acid agar supplemented with chloramphenicol, and incubated at 28 °C for 1 week with a daily observation. The isolates were identified by conventional methods. The serotype was determined by slide agglutination with specific antisera. Molecular typing was carried out by PCR-RFLP of the URA5 gene. Four isolates of C. gattii were recovered: Cryptococcus gattii serotype B, genotype AFLP4/VGI, isolated from Eucalyptus sp. in the city of Rosario and from Grevillea robusta in the city of La Paz; and C. gattii serotype C, genotype AFLP5/VGIII, isolated from two different Tipuana tipu trees in the city of Resistencia. Here, we report for the first time the isolation of C. gattii serotype C, genotype AFLP5/VGIII, from environmental samples in Argentina.
The bacillus Calmette-Guérin (BCG) was obtained in 1920 after successive passages leading to the ... more The bacillus Calmette-Guérin (BCG) was obtained in 1920 after successive passages leading to the attenuation of a Mycobacterium bovis strain. For the following 40 years, BCG had been replicated, resulting in substrains with genotypic and phenotypic differences. Several genomic studies have compared two BCG strains, M. bovis and Mycobacterium tuberculosis, and observed that deleted regions in the different strains could be related to differences in antigenic properties. In this work, a working seed lot was obtained from a lyophilized secondary seed lot from the BCG Pasteur strain 1173 P2 and genetically characterized. The genome was analyzed by PCR directed to five regions (RD1, RD2, RD14, RD15, DU2), using the seed lot and different available strains as templates. No genetic differences were found in the fragments studied as compared to the Pasteur strain. A total of 20 passages were carried out and no differences were found in the size of the fragments amplified by PCR. In conclusi...
Uploads
Papers by Alicia Trovero