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N Rastogi

Publications and source records attributed to N Rastogi.

At least 19 recordsLinked to original sources
PubMed2006

Spoligotype signatures in the Mycobacterium tuberculosis complex.

Evolution of the direct repeat region in Mycobacterium tuberculosis has created unique spoligotype signatures specifically associated with IS6110-defined strain families. Spoligotyping signatures may enable the analysis of the strain population structure in different settings and will enable the rapid identification of strain families that acquire drug resistance or escape protective immunity in drug and vaccine trials.

Bacterial Typing Techniques↗
PubMed2006

[Molecular epidemiology of tuberculosis in Guadeloupe from 1994 to 2000].

In Guadeloupe, the incidence of tuberculosis decreased between 1994 and 2000. The rate of resistance to at least one antibiotic remained constant at 11%, whereas the rate of multiple-drug resistance increased from 0.9 to 2.4% in 2000. The proportion of patients of foreign origin (mainly from Haiti and the Dominican Republic) increased whereas the number of French patients decreased. These results show that the epidemiology of tuberculosis in Guadeloupe is similar to industrialized countries as older people, foreigners from countries where TB is endemic, and HIV+ patients are at a higher risk to declare tuberculosis disease. Molecular typing realized by spoligotyping showed the importance of previous successive colonizations and migrations as characterized by the presence of major phylogenetic families originating essentially from Northern Europe (Haarlem), Latin America and Mediterranean (LAM) and from Anglo-Saxon countries (X). The sub-typing of clustered strains by IS6110-RFLP and by a PCR method based on the variable number of tandem DNA repeats (VNTR), highlighted 29 clusters, corresponding to 44.8% of clustered strains, and allowed to estimate the rate of recent transmission at 32.2%. The epidemiologic data associated with fingerprinting results underlined the importance of reactivation cases among older people, a significant number of imported TB cases without evident links, and casual contacts.

Adolescent↗
PubMed2005

An audit of postoperative radiotherapy after non-curative resection for cancer of the oesophagus.

AIMS: The role of postoperative radiotherapy (PORT) after non-curative resections for cancer oesophagus is not well defined. A policy of offering PORT after non-curative resections for cancer oesophagus has been followed at our institution, and we report an audit of our experience. MATERIAL AND METHODS: Between March 1990 and September 2002, 139 patients underwent resections for cancer oesophagus. Of these, 86 patients received PORT to a dose of 45-50.4 Gy/25-28 fractions. Eleven of these patients also received concurrent and adjuvant 5-fluorouracil (5-FU). Disease-free survival and overall survival were computed from the day of surgery using the Kaplan-Meier method. RESULTS: Seventy-six per cent (65/86) of patients had squamous cell carcinoma and 69% (59/86) of patients had tumours in the lower-third of the oesophagus. The median interval between surgery and PORT was 41 days, and 93% of patients received doses as planned. Strictures at the anastomotic site and ulcerations in the stomach mucosa were seen in 17% and 5% of patients, respectively. The median and 5-year disease-free survival was 12 months (95% CI 9.9-14.1) and 14%; whereas the median and 5-year overall survival was 17 months (95% CI 12.4-21.6) and 17%, respectively. Local and distant failures were seen in 29% and 45% of patients, respectively. CONCLUSIONS: PORT, after a non-curative resection of cancer oesophagus, is well tolerated with acceptable morbidity and survival.

Adenocarcinoma↗
PubMed2005

Adjuvant chemo-radiotherapy in ampullary cancers.

PURPOSE: Patterns of failure following surgical treatment of ampullary cancers indicate that up to 45% of patients develop loco-regional recurrence. The effect of adjuvant chemo-radiotherapy on survival and loco-regional control is not yet established in this malignancy. PATIENTS AND METHODS: From January 1989 to December 2000, 113 patients underwent pancreatico-duodenectomy for ampullary cancer. One hundred and four patients who survived the operation were available for analysis to study the effect of adjuvant chemo-radiotherapy on survival and loco-regional control. Forty-nine patients received adjuvant chemo-radiotherapy (median dose 50.4 Gy with concurrent 5-Flurouracil) and long-term outcome in these patients was compared with those 55 who did not receive adjuvant therapy. RESULTS: The overall median survival was 30.1 (range 1.6-140.0) months with actuarial 1, 3 and 5-year survival rates of 79, 43 and 33%, respectively. No significant difference in median survival (34.6 vs 24.5 months; P=0.3) and actuarial 5-year survival rates (38 vs 28%) was seen between those who received and those who did not receive adjuvant therapy. Adjuvant chemo-radiotherapy did not influence the survival in high-risk patients (P=0.84), in various T and N stages and had no impact on loco-regional recurrence (P=0.6). CONCLUSIONS: Adjuvant chemo-radiotherapy did not improve the long-term survival or decrease recurrence rates in patients with ampullary cancers who had undergone pancreatico-duodenectomy.

Adult↗
PubMed2004

Genotypic and phenotypic characterization of drug-resistant Mycobacterium tuberculosis isolates from rural districts of the Western Cape Province of South Africa.

Genotypic and phenotypic analysis of drug-resistant Mycobacterium tuberculosis isolates from the Western Cape Province of South Africa showed that drug resistance is widespread and recently transmitted. Multidrug-resistant (MDR) isolates comprise 40% of this collection, and a large pool of isoniazid monoresistance may be a future source of MDR tuberculosis.

DNA Primers↗
PubMed2004

Molecular epidemiology of drug-resistant Mycobacterium tuberculosis strains isolated from patients with pulmonary tuberculosis in Poland: a 1-year study.

OBJECTIVE: To characterise drug-resistant Mycobacterium tuberculosis strains isolated in Poland and to estimate the amount of recent transmission in the population. DESIGN: M. tuberculosis strains isolated from 251 patients with resistant pulmonary tuberculosis in Poland in 2000 were analysed by spoligotyping and IS6110 DNA fingerprinting. Part of the strains was also characterised by sequencing of the rpoB, katG and/or the regulatory region of the inhA gene. RESULTS: Using combined spoligotyping/IS6110-RFLP defined clusters, 29% of the strains were clustered, suggesting possible recent transmission. In some cases, transmission links among strains in clusters could be confirmed by epidemiological data and in addition, for most of the strains, by analysis of the mutations associated with resistance to rifampicin and/or isoniazid. Younger age, sex, immigration and history of previous treatment were not associated with clustering, whereas multidrug-resistant disease was more likely to cluster. Strains of the Beijing family could also be found in Poland, although with a much lower frequency than in the neighbouring countries. CONCLUSION: Transmission of drug-resistant M. tuberculosis strains was demonstrated, which might contribute to the emergence of drug-resistant tuberculosis in Poland.

Adolescent↗
PubMed2003

PCR-based methodology for detecting multidrug-resistant strains of Mycobacterium tuberculosis Beijing family circulating in Russia.

The Beijing genotype of Mycobacterium tuberculosis has been identified in 40-50% of the clinical isolates studied in Russia during the last decade. This genotype has been reported to be associated with multiple drug resistance and possesses some significant pathogenic properties. Therefore, early identification of such strains is of extreme importance in the timely detection of drug resistance. The present study was performed on 354 strains isolated in Russia from 1996 to 2002 and previously characterised by IS 6110-restriction fragment length polymorphism (RFLP) typing and spoligotyping. These strains included 198 Beijing family strains and 156 strains of other genotypes (IS 6110-RFLP profiles). A subsequent polymerase chain reaction (PCR) analysis with IS 6110-derived outwardly oriented primers (IS 6110-PCR) easily discriminated the Beijing strains from non-Beijing strains. The multiplex allele-specific (MAS)-PCR assays were further used to detect mutations in katG315 and rpoB531, associated with resistance to isoniazid and rifampin, respectively. The katG315 and rpoB531 mutations were found to be more prevalent among Beijing (96.8% and 77.3%) than among non-Beijing strains (85.7% and 28%). Consequently, we propose a two-step methodology based on routine PCR and simple agarose gel electrophoresis in order to detect (i) a Beijing family strain using IS 6110-PCR, and, (ii) its possible resistance to the major anti-tuberculosis drugs using specific MAS-PCR assays.

Antitubercular Agents↗
PubMed2003

[Epidemiological study of tuberculosis in the area of Angers, France, as studied by 3 PCR-based fingerprinting methods].

The new genotyping methods efficiently complement classical epidemiological investigation in order to attempt a global approach to TB control. In the present work, we have studied the genomic diversity of Mycobacterium tuberculosis isolated during the year 1998 within the district of Angers, France (260,000 inhabitants distributed in 29 districts), in order to identify recent transmission events and any related risk factors. The methods used included "spacer oligonucleotide typing" or spoligotyping, "variable number of DNA tandem repeats" or VNTR, and "double repetitive element PCR" or DRE-PCR. The resulting spoligotyping and VNTR results were also feeded to international databases and compared with >10,000 isolates for spoligotyping and 500 isolates for VNTR, representative of about 60 countries. The results obtained underlined that most of the TB cases in our setting probably reflected reactivation cases, as clustered cases indicative of potential events of recent transmission were rare. Furthermore, interrogation of international databases showed that most of the isolates from the Angers region belonged to major conserved families of TB isolates representative of Europe, with only rare cases of Asian origin, or those previously reported in specific epidemies reported from elsewhere.

DNA Fingerprinting↗
PubMed2003

[Molecular characterization and biodiversity of Mycobacterium tuberculosis in the Antilles-Guiana region and comparative analysis in a metropolitan region, Aquitaine].

Tuberculosis is a highly contagious infectious disease in recrudescence whose epidemiologic monitoring is reinforced by molecular biology. In this context, we were particularly interested in the cases of tuberculosis of French West Indies and French Guiana (FWI-FG). This study covered a period of two years (1997 and 1998) and focused on the demographical and epidemiological characteristics of the cases diagnosed by an analysis of their genotypes. Our results were confronted with a French metropolitan area (Aquitaine) with similar demographic background. Moreover, Aquitaine area has privileged links with FWI-FG region and also has a similar network for monitoring tuberculosis as ours. So we used a PCR method called spoligotyping as a first line method to optimize the alternative IS6110-RFLP method which remains cumbersome. A total of 105 strains of FWI-FG and 172 strains of Aquitaine were typed by spoligotyping and by the standard IS6110-RFLP method. The results of the first grouping by spoligotyping were analyzed in comparison with IS6110-RFLP. The results obtained showed a rate of recent transmission of tuberculosis being 34.3% in FWI-FG and 10.5% in Aquitaine. These observations underlined a high degree of polymorphism in the Aquitaine region as compared to the FWI-FG region. Thanks to the various profiles obtained by spoligotyping, we could study their distribution in the three areas and highlight common types like type 53, 50 and 42 and types found locally like the types 33 and 14 found respectively in Aquitaine and FWI as well as endemic types like type 76 found only in FG. These results are discussed in the context of the evolution of clinical isolates of tubercle bacilli with time.

Biodiversity↗
PubMed2002

A data-mining approach to spacer oligonucleotide typing of Mycobacterium tuberculosis.

MOTIVATION: The Direct Repeat (DR) locus of Mycobacterium tuberculosis is a suitable model to study (i) molecular epidemiology and (ii) the evolutionary genetics of tuberculosis. This is achieved by a DNA analysis technique (genotyping), called sp acer oligo nucleotide typing (spoligotyping ). In this paper, we investigated data analysis methods to discover intelligible knowledge rules from spoligotyping, that has not yet been applied on such representation. This processing was achieved by applying the C4.5 induction algorithm and knowledge rules were produced. Finally, a Prototype Selection (PS) procedure was applied to eliminate noisy data. This both simplified decision rules, as well as the number of spacers to be tested to solve classification tasks. In the second part of this paper, the contribution of 25 new additional spacers and the knowledge rules inferred were studied from a machine learning point of view. From a statistical point of view, the correlations between spacers were analyzed and suggested that both negative and positive ones may be related to potential structural constraints within the DR locus that may shape its evolution directly or indirectly. RESULTS: By generating knowledge rules induced from decision trees, it was shown that not only the expert knowledge may be modeled but also improved and simplified to solve automatic classification tasks on unknown patterns. A practical consequence of this study may be a simplification of the spoligotyping technique, resulting in a reduction of the experimental constraints and an increase in the number of samples processed.

Algorithms↗
PubMed2001

Integration of tuberculosis screening at an HIV voluntary counselling and testing centre in Haiti.

OBJECTIVE: To describe the integration of tuberculosis screening into the activities of an HIV voluntary counselling and testing (VCT) centre in a country with endemic tuberculosis. SETTING: An HIV VCT centre in Port au Prince, Haiti. DESIGN: All patients presenting for HIV VCT who reported cough received same-day evaluation for active tuberculosis. Of the 1327 adults presenting to the centre for the first time between January and April 1997, 263 (20%) reported cough and of these 241 (92%) were evaluated. RESULTS: Of the 241 patients evaluated for cough, 76 (32%) were diagnosed with pulmonary tuberculosis. Of the 76 patients diagnosed with pulmonary tuberculosis, 28 (37%) had a positive smear for acid-fast bacilli (AFB), 14 (18%) had a negative AFB smear but a positive sputum culture for Mycobacterium tuberculosis, and 34 (45%) had culture-negative tuberculosis. Also, 31 out of 241 (13%) VCT clients evaluated for cough were diagnosed with bacterial pneumonia. CONCLUSION: This report confirms that in areas with a high HIV and tuberculosis prevalence, a high proportion of VCT clients have active pulmonary tuberculosis. The integration of tuberculosis screening offers several benefits, including the diagnosis and treatment of large numbers of individuals with tuberculosis, a decreased risk of nosocomial tuberculosis transmission, and the opportunity to provide tuberculosis prophylaxis to HIV-positive patients in whom tuberculosis has been excluded. Future studies are needed to determine the cost-effectiveness of integrated tuberculosis and HIV VCT services, and whether integration should be recommended in all countries with high HIV and tuberculosis rates.

AIDS Serodiagnosis↗
PubMed2001

Mycobacterium tuberculosis phylogeny reconstruction based on combined numerical analysis with IS1081, IS6110, VNTR, and DR-based spoligotyping suggests the existence of two new phylogeographical clades.

This paper deals with phylogenetic relationships among a set of 90 clinical strains representative of the worldwide diversity of the Mycobacterium tuberculosis complex (Kremer et al. 1999) using eight independent genetic markers: IS6110, IS1081, the direct repeat (DR) locus, and five variable number of tandem DNA repeat loci (VNTR). In a preliminary experiment, phylogenetic trees based on single markers were constructed that led to the detection of some similarities between the VNTR-based and the spoligotyping-based phylogenetic trees. In the second step, a more global phenetic approach based on pairwise comparison of strains within each typing system was used, followed by calculations of mean genetic distances based on all the eight loci and the use of the neighbor-joining algorithm for tree reconstruction. This analysis confirmed our preliminary observations and suggested the existence of at least two new phylogeographical clades of M. tuberculosis, one defined as the "East African-Indian family" (EA-I), which may find its origin on the African or Asian continents, and the other as the "Latin American and Mediterranean" (LA-M) family. The existence of these two families was also validated by an independent phylogenetic analysis of spoligotyping on a larger set of shared types (n = 252) and further corroborated by VNTR and katG-gyrA results. The potential origin of these families of bacilli is discussed based on cattle domestication and human migration history. In conclusion, the information contained in insertion sequence and repetitive DNAs may serve as a model for the phylogenetic reconstruction of the M. tuberculosis complex.

Animals↗
PubMed2001

Genetic diversity of Mycobacterium africanum clinical isolates based on IS6110-restriction fragment length polymorphism analysis, spoligotyping, and variable number of tandem DNA repeats.

A collection of 105 clinical isolates originally identified as Mycobacterium africanum were characterized using both phenotypic and genotyping methods. The phenotypic methods included routine determination of cultural properties and biochemical tests used to discriminate among the members of the M. tuberculosis complex, whereas genotypic characterization was based on IS6110-restriction fragment length polymorphism (IS6110-RFLP) analysis, IS1081-RFLP analysis, direct repeat-based spacer oligonucleotide typing (spoligotyping), variable number of tandem DNA repeats (VNTR), and the polymorphism of the oxyR, pncA, and mtp40 loci. The results obtained showed that a majority of M. africanum isolates were characterized by a specific spoligotyping pattern that was intermediate between those of M. tuberculosis and M. bovis, which do not hybridize with spacers 33 to 36 and spacers 39 to 43, respectively. A tentative M. africanum-specific spoligotyping signature appeared to be absence of spacers 8, 9, and 39. Based on spoligotyping, as well as the polymorphism of oxyR and pncA, a total of 24 isolates were excluded from the final study (19 were identified as M. tuberculosis, 2 were identified as M. canetti, and 3 were identified as M. bovis). The remaining 81 M. africanum isolates were efficiently subtyped in three distinct subtypes (A1 to A3) by IS6110-RFLP analysis and spoligotyping. The A1 and A2 subgroups were relatively more homogeneous upon spoligotyping than A3. Further analysis of the three subtypes by VNTR corroborated the highly homogeneous nature of the A2 subtype but showed significant variations for subtypes A1 and A3. A phylogenetic tree based on a selection of isolates representing the three subtypes using VNTR and spoligotyping alone or in combination confirmed the subtypes described as well as the heterogeneity of subtype A3.

Alleles↗
PubMed2001

Rapid differentiation of "Mycobacterium canettii" from other Mycobacterium tuberculosis complex organisms by PCR-restriction analysis of the hsp65 gene.

A total of 102 isolates of the Mycobacterium tuberculosis complex, including available "M. canettii" isolates, were studied by PCR-restriction analysis of a 441-bp fragment of the hsp65 gene. PRA upon HhaI enzyme digestion (GCGC) allowed easy differentiation of "M. canettii" from other members of the M. tuberculosis complex (three bands of 260, 105, and 60 bp for "M. canetti," compared to four bands of 185, 105, 75, and 60 bp for other members of the M. tuberculosis complex). Sequencing of the 441-bp hsp65 fragment of "M. canettii" isolates showed the disappearance of an HhaI site at position 235 due to a C-to-T transition that corresponded to position 631 of the homologous hsp65 gene of M. tuberculosis H37Rv. Considering that "M. canettii" may also exist as a stable rough morphotype, we suggest that the true number of "M. canettii" isolates may be underestimated in clinical microbiology laboratories.

Bacterial Proteins↗
PubMed2001

Burden of unidentifiable mycobacteria in a reference laboratory.

Modern identification techniques at the genomic level have greatly improved the taxonomic knowledge of mycobacteria. In adjunct to nucleic acid sequences, mycobacterial identification has been endorsed by investigation of the lipidic patterns of unique mycolic acids in such organisms. In the present investigation, the routine use of high-performance liquid chromatography (HPLC) of mycolic acids, followed by the sequencing of the 16S rRNA, allowed us to select 72 mycobacterial strains, out of 1,035 screened, that do not belong to any of the officially recognized mycobacterial species. Most strains (i.e., 47) were isolated from humans, 13 were from the environment, 3 were from animals, and 9 were from unknown sources. The majority of human isolates were grown from the respiratory tract and were therefore most likely not clinically significant. Some, however, were isolated from sterile sites (blood, pleural biopsy, central venous catheter, or pus). Many isolates, including several clusters of two or more strains, mostly slow growers and scotochromogenic, presented unique genetic and lipidic features. We hope the data reported here, including the results of major conventional identification tests, the HPLC profiles of strains isolated several times, and the whole sequences of the 16S rRNA hypervariable regions of all 72 mycobacteria, may encourage reporting of new cases. The taxonomy of the genus Mycobacterium is, in our opinion, still far from being fully elucidated, and the reporting of unusual strains provides the best background for the recognition of new species. Our report also shows the usefulness of the integration of novel technology to routine diagnosis, especially in cases involving slow-growing microorganisms such as mycobacteria.

Bacterial Typing Techniques↗
PubMed2001

Genetic diversity of Mycobacterium tuberculosis in Sicily based on spoligotyping and variable number of tandem DNA repeats and comparison with a spoligotyping database for population-based analysis.

In a previous study, we proposed to associate spoligotyping and typing with the variable number of tandem DNA repeats (VNTR) as an alternative strategy to IS6110-restriction fragment length polymorphism (RFLP) for molecular epidemiological studies on tuberculosis. The aim of the present study was to further evaluate this PCR-based typing strategy and to describe the population structure of Mycobacterium tuberculosis in another insular setting, Sicily. A collection of 106 DNA samples from M. tuberculosis patient isolates was characterized by spoligotyping and VNTR typing. All isolates were independently genotyped by the standard IS6110-RFLP method, and clustering results between the three methods were compared. The totals for the clustered isolates were, respectively, 15, 60, and 82% by IS6110-RFLP, spoligotyping, and VNTR typing. The most frequent spoligotype included type 42 that missed spacers 21 to 24 and spacers 33 to 36 and derived types 33, 213, and 273 that, together represented as much as 26% of all isolates, whereas the Haarlem clade of strains (types 47 and 50, VNTR allele 32333) accounted for 9% of the total strains. The combination of spoligotyping and VNTR typing results reduced the number of clusters to 43% but remained superior to the level of IS6110-RFLP clustering (ca. 15%). All but one IS6110-defined cluster were identified by the combination of spoligotyping and VNTR clustering results, whereas 9 of 15 spoligotyping-defined clusters could be further subdivided by IS6110-RFLP. Reinterpretation of previous IS6110-RFLP results in the light of spoligotyping-VNTR typing results allowed us to detect an additional cluster that was previously missed. Although less discriminative than IS6110-RFLP, our results suggest that the use of the combination of spoligotyping and VNTR typing is a good screening strategy for detecting epidemiological links for the study of tuberculosis epidemiology at the molecular level.

Adult↗
PubMed2001

Use of spoligotyping to study the evolution of the direct repeat locus by IS6110 transposition in Mycobacterium tuberculosis.

Based on the variability of 43 spacers within the direct repeat (DR) locus of Mycobacterium tuberculosis complex organisms, spoligotyping is a rapid method that aids in the study of the epidemiology of tuberculosis. It was recently hypothesized that despite its presence in the DR locus, spacer 31 could not be amplified in M. tuberculosis clinical isolates belonging to spoligotype 50 due to the insertion of an extra copy of IS6110 between spacers 31 and 32 that could lead to an asymmetrical split of the primer targets (I. Filliol, C. Sola, and N. Rastogi, J. Clin. Microbiol. 38:1231--1234, 2000). In the present investigation, previous observations were extended to 25 clinical isolates of type 50 showing that the primer set IS6-DRb that selectively amplified the left and central DR regions was indeed able to demonstrate the presence of spacer 31. IS6110-restriction fragment length polymorphism (RFLP) and DR-RFLP showed that type 50 isolates were characterized by the presence of two copies of IS6110 associated with the DR locus and an additional double IS6110 band of 1.4 kb. The primer set IS3-IS6 was then used to selectively amplify a 750-bp inter-IS6110 fragment within the DR locus. The sequencing of the central DR region corroborated our previous findings and showed that the absence of spacer 31 among the type 50 isolates was due to the asymmetric insertion of an extra copy of IS6110 between spacers 31 and 32, leading to an unequal split of the DRa-DRb target into two portions, of 6 and 30 bp, respectively. These results show that the DR locus constitutes an ideal IS6110 preferential locus (ipl), permitting the insertion of two or more copies of IS6110, and provide new clues for epidemiological and phylogenetic interpretation of changes in IS6110-RFLP and spoligotyping profiles.

Bacterial Typing Techniques↗