Pradeep K. Dutta

Sensitivity and performance characteristics of a direct PCR with stool samples in comparison to conventional techniques for diagnosis of Shigella and enteroinvasive Escherichia coli infection in children with acute diarrhoea in Calcutta, India

As the sensitivity of the conventional techniques for identifying Shigella spp. and enteroinvasive Escherichia coli (EIEC) causing dysentery cases is low, a PCR assay was evaluated in this study. Analytical sensitivity (2 x 10(2) cfu) of the PCR technique was obtained by artificially spiking negative stool samples with a standard strain of S. flexneri type 2, then determining the detection limit. Specificity (100%) of the method was determined by testing a number of known Shigella and EIEC strains and organisms other than Shigella spp. A total of 300 stool samples collected from children with acute diarrhoea was plated on to two selective agar media after enrichment in Luria broth. Shigella spp. were isolated from 7.7% (23 of 300) and EIEC from 1% (3 of 300) patients. All enriched stool samples were subjected to PCR to amplify the target sequence of invasive plasmid antigen (ipa)H locus, a multicopy element found on the chromosome and invasion plasmid. The stool PCR was positive in 24 of the 26 culture-positive and in 22 culture-negative stools, thus detecting the presence of Shigella spp. or EIEC in 15.3% (46 of 300) of diarrhoea cases. When an ial probe was used for colony hybridistion with enriched stool cultures blotted on to membranes, 9.6% (29 of 300) of dysentery cases were identified as being caused by Shigella spp. or EIEC. Thus the sensitivity of enriched stool culture, colony hybridisation and enriched stool PCR was found to be 54%, 60% and 96%, respectively, when each of the methods was compared to the total microbiologically confirmed cases of dysentery. It was also observed that only 38% (48 of 126) of acute bloody dysentery cases actually had shigella or EIEC infection, as confirmed by laboratory methods. Moreover, this PCR assay could identify a number of untypable Shigella strains (Sh OUT), which would have remained undiagnosed had this assay not been used.

A clerodane diterpene and other constituents of Clerodendron inerme

Abstract A new clerodane diterpene clerodermic acid was isolated from Clerodendron inerme and the structure deduced from spectral data. The known compounds friedelin, 5-hydroxy-7,4′-dimethoxyflavone, salvigenin, acacetin and apigenin were also found.

Two isomeric flavanones from vitex negundo

Abstract Two new flavanoids, characterized as 5,3′-dihydroxy-7,8,4′-trimethoxyflavanone and 5,3′-dihydroxy-6,7,4′-trimethoxyflavanone, have been isolated from the leaves of Vitex negundo.

Use of PCR to identify enteroaggregative Escherichia coli as an important cause of acute diarrhoea among children living in Calcutta, India.

The importance of enteroaggregative Escherichia coli (EAggEC) as a possible aetiological agent of acute diarrhoea among children in Calcutta, India, was investigated. Simultaneously the use of a previously described PCR diagnostic system was assessed for its ability to identify EAggEC infection. E. coli strains isolated during a 1-year case-control study from faecal samples of 388 children aged <5 years, with or without diarrhoea, were examined for EAggEC by HeLa cell adherence assay in parallel with a PCR assay with primers generated from an EAggEC DNA probe. A blind comparison was made between the two methods to determine their diagnostic potential. E. coli isolates that adhered to HeLa cells in an aggregative pattern were the sole isolates significantly more often in 254 cases (9%) than in 134 control (2%) children. Age stratification showed that EAggEC were isolated more frequently from children aged <36 months. The EAggEC isolates belonged to several O serogroups and showed multiple drug resistance. Both methods were positive for 26 samples, nine samples were positive by PCR alone and seven samples were positive by culture alone, thus indicating a 78% sensitivity and 97% specificity for the PCR assay. EAggEC is an important aetiological agent of acute diarrhoea among infants in and around Calcutta, and the PCR diagnostic system may be useful to identify such infection in epidemiological studies.

A tetracyclic triterpenoid from Musa paradisiaca

Abstract The structure of a new tetracyclic triterpene isolated from the flowers of Musa paradisiaca was determined as (24 R )-4α,14α,24-trimethyl-5α-cholesta-8,25(27)-dien-3β-ol.

A Novel Galacto-Glycerolipid from Oxalis corniculata Kills Entamoeba histolytica and Giardia lamblia

ABSTRACT Oxalis corniculata is a naturally occurring weed that has been used in traditional medicine for the cure of dysentery and diarrhea in India. One of the common causes of dysentery is due to infection by the protist pathogen Entamoeba histolytica. Bioactivity profiling of extracts from O. corniculata identified several compounds that showed antiamoebic activity in axenic cultures of E. histolytica. These were characterized by nuclear magnetic resonance, infrared, and mass spectrometry as (i) Oc-1, a mixture of saturated fatty acids C24 to C28; (ii) Oc-2, a mixture of long-chain alcohols C18 to C28; and (iii) Oc-3, a single compound that was a galacto-glycerolipid (GGL). Of the different compounds that were obtained, the strongest antiamoebic activity was found in GGL. The addition of GGL to E. histolytica xenic cultures containing other microbial flora from the large intestine did not affect its antiamoebic activity. Amoebicidal concentrations of GGL had no effect on intestinal microbial flora or on the mammalian cell line HEK-293. GGL was also found to be equally effective in killing another protist pathogen, Giardia lamblia, that causes diarrhea in humans. The importance of this study is based on the identification of novel natural products and the possibility of developing these compounds as active agents to treat at least two pathogenic parasitic intestinal infections endemic to tropical regions.

Two isomeric benzodipyranone derivatives from Calophyllum inophyllum

Abstract The new compound (2 S ,3 R )-2,3-dihydro-5-hydroxy-2,3,8,8-tetramethyl-6-(1-phenylethenyl)-4H,8H-benzo [1,2- b :3,4- b ′] dipyran-4-one along with (2 R ,3 R )-2,3-dihydro-5-hydroxy-2,3,8,8-tetramethyl-6-(1-phenylethenyl)-4H,8H-benzo [1,2- b :3,4- b ′] dipyran-4-one were isolated from the leaves of Calophyllum inophyllum and identified from their spectral and chemical data.

A neo-clerodane diterpene from Clerodendron inerme

Abstract The structure of a new diterpene isolated from the leaves of Clerodendron inerme was established by spectroscopic methods as 15-methoxy-14,15-dihydro-3-epicaryoptin.

Fluorescent Pigment and Phenol Glucosides from the Heartwood of Pterocarpus marsupium

The fluorescence shown by extracts of the heartwood of Pterocarpus marsupium is attributed to salts of the new compound 1, whose structure was elaborated using detailed spectroscopic/spectrometric studies. The plant material also contains the nonfluorescent compounds 2 and 3. The absolute configuration of 1 was determined by experimental and theoretically calculated electronic CD spectra, while that of 3 was deduced from ECD comparison with reported results in the α-hydroxydihydrochalcone series.

Marine products from bay of Bengal: constituents of the sponge Psammaplysilla purpurea☆

Abstract New bromotyrosine derived secondary metabolites have been isolated from the marine sponge Psammaplysilla purpurea , and the structures assigned on the basis of spectroscopic analyses of their acetates 1a′ and 1b′ .