Detection of Enterocytozoon bieneusi (Microsporidia) by polymerase chain reaction (PCR) using species-specific primer in stool samples of HIV patients.

BACKGROUND & OBJECTIVE: Microsporidia are obligate intracellular parasites causing infections predominantly in immunocompromised patients. Enterocytozoon bieneusi is the most important microsporidian causing chronic diarrhoea in AIDS patients. The current method used for diagnosing the microsporidia spores is based on light microscopy using stained smears, which do not differentiate spores at species level. The present study was undertaken to detect microsporidia and confirm at species level (E. bieneusi) by PCR from stool samples of HIV positive patients. METHODS: During September 2002 to April 2003, stool samples from 153 HIV-positive patients (with chronic diarrhoea n = 105; without diarrhoea n=48) were collected and examined microscopically for microsporidia spores using modified Weber's chromotrope stain. Stool samples were subjected to PCR assay using species-specific primer EBIEFI/EBIER1, which amplifies small subunit ribosomal RNA (SSU rRNA) of this microsporidian RESULTS: A total of 10 HIV positive patients with chronic diarrhoea were positive for microsporidia by microscopic analysis and confirmed as Enterocytozoon bieneusi by PCR. No false positive results were observed. A diagnostic DNA fragment of 607 bp of the unique SSU rRNA was amplified from all samples infected with E. bieneusi. INTERPRETATION & CONCLUSION: The study revealed that polymerase chain reaction is a useful tool for accurate species identification of microsporidia in stool samples, which serves the benefit of treatment to the patients.

Cefoxitin resistance mediated by loss of a porin in clinical strains of Klebsiella pneumoniae and Escherichia coli.

PURPOSE: Porins are outer membrane protein (OMP) that form water filled channels that permit the diffusion of small hydrophilic solutes like beta-lactam antibiotics across the outer membrane. Two major porins that facilitate diffusion of antimicrobials have been described in Klebsiella spp. and Escherichia coli. The present study was carried out to examine the role of porins among Extended Spectrum beta-Lactamase (ESBL) and AmpC beta-Lactamase positive strains of Klebsiella spp. and E.coli. METHODS: Preparation of OMP from phenotypically characterized clinical isolates K.pneumoniae and E.coli and the separation of the proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were performed as per a previously described procedure. RESULTS: OMP analysis revealed that cefoxitin and ceftazidime resistance was mediated by loss of a porin Omp K35 in the isolates of K.pneumoniae and E.coli. CONCLUSIONS: Loss of porin mediated resistance mechanism against cefoxitin was observed among the multidrug resistant K.pneumoniae and E.coli.

Rotavirus infection among infants and young children in Chennai, South India.

BACKGROUND: Group A rotavirus has been recognized as the major etiologic agent of childhood gastroenteritis in infants and young children worldwide. Rapid progress towards the development of an efficacious rotavirus vaccine has warranted extensive epidemiological studies on rotavirus serotypes that cause severe disease in developing and developed countries and to monitor the emergence of newer and unusual strains in different geographical settings that could represent variants not covered by existing vaccines. METHODS: In this study, we determined the prevalence of rotavirus infection and characterised group A rotavirus in stool samples by using monoclonal antibody (MAb) based ELISA and polyacrylamide gel electrophoresis (PAGE). Stool samples were collected from 745 children of 0-3 years of age presenting to the hospital with acute diarrhea between March 1995 and August 1999. These were assayed for antigenic (group, subgroup, serotype) and genomic (viral RNA profile and VP7 and VP4 genotype) characterization by ELISA and PAGE. RESULTS: Out of 745 stool samples analysed 168 were found to be positive for rotavirus. Among these 118 could be assigned a subgroup (SG), serotype and electropherotype (E-type). The study has evidenced the predominant occurrence of strains with short E-type, SGI and serotype G2 in 66.1% of the samples. The presence of strains representing 10 different E-types and mixed genotype specificities with G2 P[4,8] and G1-G2 P[4,8] has documented the prevailing high genomic diversity of rotaviruses in this geographical area. CONCLUSION: This study has described the predominant strains of rotavirus in south India. There is a need for further detailed studies on the molecular characterization of rotaviruses which would have important implications in vaccine evaluation programmes.

Electron microscopy identification of microsporidia (enterocytozoon bieneusi) and cyclospora cayetanensis from stool samples of HIV infected patients.

Microsporidia (Enterocytozoon bieneusi) and Cyclospora cayetanensis have been reported worldwide causing diarrhoea in AIDS patients. Stool samples from HIV infected patients were subjected to routine examination for parasites, followed by special staining techniques to detect microsporidia and Cyclospora cayetanensis. Confirmed positive cases of these parasites were further processed for electron microscopy identity of the parasites and characteristic details. Scanning and transmission electron microscopy showed better morphological and structural details of the parasites.

Biochemical characteristics, serogroups, and virulence factors of aeromonas species isolated from cases of diarrhoea and domestic water samples in Chennai.

PURPOSE: The objective of the present study was to delineate the differences between the clinical and environmental Aeromonas species with respect to their biochemical characteristics, serogrouping and virulence factors, in order to find a phenotypic marker of enteropathogenicity. METHODS: A total of 55 Aeromonas spp. inclusive of 19 isolates from cases of diarrhoea, and 36 from water samples comprising, 10 isolates of A. hydrophila, 21 isolates each of A. sobria, and A. caviae, two isolates of A. jandaei and one isolate of A. veronii were subjected to analysis of their biochemical characteristics, serogrouping, and virulence factors. RESULTS: Among the differences recorded in the biochemical characteristics in the three major species, the most striking characteristic was fermentation of lactose, which was observed in all the 11 A. caviae isolates recovered from water samples. None of the 10 clinical isolates of A. caviae tested fermented lactose. The clinical Aeromonas isolates belonged to seven typable serogroups, O:13, O:14, O:16, O:21, O:27, O:32 and O:35. The environmental isolates belonged to eight different serogroups, such as, O:3, O:11, O:14, O:16, O:18, O:28, O:64 and O:78 and were predominated by serotypes O:18 and O:64. Among the virulence factors tested, 89% of the environmental isolates produced b haemolysin, while only 62.3% of clinical isolates were able to do so. There was no significant difference between the clinical and environmental aeromonads with respect to their enterotoxigenicity in suckling mice in vivo, cytotoxicity in vitro in Vero cell monolayers, and ability to produce siderophores. CONCLUSION: Efforts to delineate the differences between the clinical and environmental Aeromonas spp. did not reveal significant difference between them. However, difference was observed with respect to their ability to produce b haemolysin, wherein, higher percentage of environmental isolates was haemolytic. The results also suggest that all the haemolytic environmental isolates need not be enteropathogenic. Further, serogroups O:18 and O:64 may not be involved in aeromonal diarrhoea in children in this geographic region.

AmpC beta-lactamase producing multidrug resistant strains of Klebsiella spp. & Escherichia coli isolated from children under five in Chennai.

BACKGROUND & OBJECTIVES: AmpC beta-lactamases are Group I cephalosporinases that confer resistance to a wide variety of beta-lactam drugs. Plasmid mediated AmpC beta-lactamases has been discovered most frequently in isolates of Klebsiella pneumoniae, K. oxytoca, Salmonella, Proteus mirabilis and Escherichia coli. The present study was undertaken to study the occurrence of multidrug resistant and AmpC beta-lactamase producing Klebsiella spp. and Escherichia coli in children less than five years of age as this age group is very susceptible to intestinal and extraintestinal infections. METHODS: A total of 116 isolates of Klebsiella species and 32 isolates of Esch. coli were tested for resistance to cephamycin such as cefoxitin, third generation cephalosporin (3GC) antibiotics (ceftazidime, cefotaxime, ceftriaxone), ampicillin, amikacin, cephaloridine, cefuroxime, co-trimoxazole, gentamycin, imipenem and tetracycline by disc diffusion method. Isolates found resistant to cefoxitin were tested for the production of AmpC beta-lactamases by three dimensional extract method. Transconjugation experiments were done to study the transfer of drug resistance and AmpC beta lactamase production from AmpC producing Klebsiella and Esch. coli isolates to a recipient Esch. coli strain (K12 J62-2). RESULTS: Twenty eight isolates (24.1%) of Klebsiella spp. and 12 (37.5%) of Esch. coli were found to be AmpC beta-lactamase producers; 66.6 per cent and 81 per cent of Klebsiella and Esch. coli isolates respectively showed resistance to all the 3GCs. All the strains were found to be sensitive to imipenem. Eighty four (72%) of Klebsiella isolates and 20 (62.5%) of Esch. coli were found to be resistant to cefoxitin. Transfer of cefoxitin resistance to the recipient strain was observed in all the AmpC producing strains of Klebsiella spp. Of the 12 AmpC producing strains of Esch. coli, only 4 (33.3%) showed the transfer of cefoxitin resistance to the recipient strain. INTERPRETATION & CONCLUSION: This study has shown the occurrence of AmpC beta-lactamase producing Klebsiella and Esch. coli strains in children in Chennai. Since AmpC beta-lactamase production is frequently accompanied by multiresistance to antibiotics, therapeutic options become limited resulting a need for new measures for the management of Klebsiella and Esch. coli infections. Also failure to identify AmpC beta-lactamase producers may lead to inappropriate antimicrobial treatment and may result in increased mortality. Detecting plasmid mediated AmpC beta-lactamase producing strains is technically difficult and the phenotypic tests for AmpC detection are not well defined. If an investigational AmpC beta-lactamase inhibitor was made available for diagnostic testing, it could be useful in combination with a suitable cephamycin to confirm AmpC production.

Role of coccidian parasites in causation of diarrhoea in HIV infected patients in Chennai.

BACKGROUND & OBJECTIVES: Diarrhoea is one of the major problems of HIV positive patients. A coproparasitological study was carried out to assess the role of coccidian parasites in the causation of diarrhoea in HIV infected patients in Chennai. METHODS: During May 2000 to January 2001, 152 stool samples from HIV seropositive individuals (43 with acute diarrhoea, 59 with chronic diarrhoea, 50 without diarrhoea) and 50 normal individuals without diarrhoea were examined for enteric coccidian and other intestinal parasites by microscopy and special staining methods. RESULTS: A total of 52 enteric parasites, 15 from patients with acute diarrhoea and 24 from patients with chronic diarrhoea, 7 from patients infected with HIV without diarrhoea and 6 from normal individuals without diarrhoea were detected from 49 patients. Isospora belli was detected in 14 of 102 (13.7%) patients with acute and chronic diarrhoea. The association with diarrhoea among HIV positive individuals was significant (P < 0.001). Cryptosporidium was detected in 7 patients each with acute and chronic diarrhoea and 4 patients with HIV infection without diarrhoea, its association with diarrhoea among HIV patients was found to be not significant in the present study. Cyclospora and Microsporidia each were detected in only one HIV positive patient with chronic diarrhoea. INTERPRETATION & CONCLUSION: The study revealed that the coccidian parasites are one of the important etiologic agents of diarrhoea (P < 0.001) especially of chronic diarrhoea among HIV positive patients. Isospora belli was found to be a frequent enteric parasite associated with diarrhoea among HIV positive patients in Chennai.

Extended spectrum beta lactamase (ESBL) mediated resistance to third generation cephalosporins among klebsiella pneumoniae in Chennai.

PURPOSE: To examine the incidence of extended spectrum b lactamase (ESbL) producing strains and multidrug resistant strains of Klebsiella pneumoniae isolated from children between 0-5 years of age. METHODS: Multidrug resistance and ESbL production was studied in a total of 120 isolates of Klebsiella pneumoniae obtained from patients aged 0-5 years. RESULTS: 95% of the isolates showed resistance or decreased susceptibility to atleast one of the three third generation cephalosporins [3GC (ceftazidime, cefotaxime, ceftriaxone)] used for the study. 87% of the isolates showed resistance to all the three 3GC antibiotics and this resistance to all the three 3GC was found to coexist with resistance to other antibiotics. All the isolates were found sensitive to the antibiotic imipenem. ESbL production was detected in 8 strains of Klebsiella pneumoniae. The ESbL activity could be experimentally transferred to recipient E.coli (K12 J62-2). Resistance to b-lactam antibiotics was co-transferred with resistance to gentamicin. CONCLUSIONS: This study has shown the incidence of ESbL producing Klebsiella pneumoniae strains among children in Chennai. Tests for the detection of ESbL producing Klebsiella strains should be carried out in all diagnostic centers routinely and the therapeutic use of all the 3GC should be avoided against Klebsiella strains that appear resistant to any third generation antibiotic.

Intestinal parasitic infection in HIV infected patients with diarrhoea in Chennai.

PURPOSE: To determine the prevalence of intestinal parasites in HIV patients with and without diarrhoea in Chennai. METHODS: A total of 150 stool samples, 41 - acute diarrhoea, 59 - chronic diarrhoea and 50 control samples without diarrhoea were collected and examined for enteric parasites by microscopy. RESULTS: Enteric parasites were detected in 39% patients with diarrhoea compared to 14% in patients without diarrhoea. Isospora belli was found in 18.6% (11/59) of chronic diarrhoea and 7.3% (3/41) in acute diarrhoea (P > 0.2). Cryptosporidium was detected in 7 cases each in acute and chronic diarrhoea, which was statistically insignificant as compared to the control group (P> 0.05). Microsporidia and Cyclospora cayetanensis associated diarrhoea were detected in only one chronic case each 1/59 (1.69 %). CONCLUSIONS: Isospora belli appeared to be a predominant parasite associated with diarrhoea among HIV patients. Detection rate of Microsporidia and Cyclospora was found to be very low.

Extended spectrum beta lactamase production & multidrug resistance in Klebsiella species isolated from children under five with intestinal & extraintestinal infections.

BACKGROUND & OBJECTIVES: Extended spectrum beta-lactamases (ES beta L) are enzymes produced in some Gram-negative bacilli that mediate resistance to third generation cephalosporins (3GC) and aztreonam. These are common in Klebsiella spp. and Escherichia coli and in other members of the family enterobacteriaceae. ES beta L production is accompanied by resistance to other antibiotics as these are encoded by multi drug resistance conjugative plasmids. The present study was undertaken to study the incidence of multi drug resistant and ES beta L producing Klebsiella spp. in children under five years of age suffering from intestinal and extraintestinal infections. METHODS: A total of 90 strains of Klebsiella spp. (76 isolates of K. pneumoniae and 14 of K. oxytoca) were tested for resistance to 3GC antibiotics (ceftazidime, cefotaxime, ceftriaxone), amikacin, ampicillin, erythromycin, gentamycin and streptomycin by disc diffusion method. Isolates found resistant to 3GC antibiotics were tested for the production of ES beta L by double disc diffusion synergy test. Transconjugation experiments were done to study the transfer of drug resistance and ES beta L production from Klebsiella isolates to an Esch. coli strain (K12 J62-2). RESULTS: All the 90 isolates showed multi drug resistance; 87 (96.6%) isolates showed resistance or decreased susceptibility to at least one of the three 3GC. ES beta L production was detected in four strains of K. pneumoniae and two K. oxytoca. ES beta L activity could be experimentally transferred to recipient Esch. coli in all the 6 isolates. Resistance to beta-lactam antibiotics was co-transferred along with resistance to gentamycin. INTERPRETATION & CONCLUSION: This study has shown the incidence of ES beta L producing Klebsiella strains in children in Chennai, and possibly poses a threat in the treatment and management of Klebsiella associated infections. The incidence of ES beta L producing strains of Klebsiella and other members of enterobacteriaceae should be carefully monitored in children to prevent unnecessary use of antibiotics especially 3GC and aminoglycoside antibiotics. Hence, tests for the detection of ES beta L producing Klebsiella strains should be carried out routinely for better therapeutic management.

Typing of Aeromonas isolates from children with diarrhoea & water samples by randomly amplified polymorphic DNA polymerase chain reaction & whole cell protein fingerprinting.

BACKGROUND & OBJECTIVES: Aeromonas spp. are water-borne organisms, often associated with childhood diarrhoea. The present study was conducted to examine the epidemiological relationship among the Aeromonas spp. isolated from water and children with acute diarrhoea in Chennai. METHODS: Thirty six Aeromonas isolates inclusive of 16 from children with diarrhoea, 15 from domestic water samples and 5 reference strains were studied by randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR). Twenty eight Aeromonas isolates, 15 from children with diarrhoea, 10 from domestic water samples and three reference strains were analysed by SDS-PAGE for their whole cell protein profiles. RESULTS: The 36 Aeromonas isolates examined by RAPD-PCR generated RAPD fingerprints with majority of the bands ranging from about 250 to 2800 bp. The RAPD fingerprints did not correspond with the phenospecies and varied greatly among the strains within the phenospecies. Cluster analysis revealed two major groups at 75 per cent hierarchical level, comprising 18 Aeromonas isolates, mainly recovered from domestic water samples, while the clinical isolates were scattered in different hierarchical levels in the dendrogram. The whole cell protein fingerprints examined by SDS-PAGE did not correspond with the phenospecies. Only four isolates of A. caviae were found to produce similar protein fingerprints allowing them to form a cluster at about 90 per cent hierarchical level, while the rest of the isolates were scattered at various hierarchical levels in the dendrogram. INTERPRETATION & CONCLUSIONS: In the present study, RAPD fingerprinting was found to be useful in distinguishing Aeromonas isolates recovered from clinical and domestic water supplies. However, RAPD-PCR could not distinguish the phenospecies of the genus Aeromonas. Whole cell protein fingerprinting and cluster analysis could neither differentiate isolates from clinical and domestic water sources nor the phenospecies of the genus Aeromonas.

Comparative analysis of monoclonal antibody based enzyme immunoassay, modified genome electrophoresis & electron microscopy procedures for rotavirus diagnosis from faecal specimens.

BACKGROUND & OBJECTIVES: Reliable and rapid diagnosis of rotavirus infection is necessary for patient management. Several newly introduced commercial enzyme immunoassays (ELISAs) have been evaluated using direct electron microscopy (DEM) with or without direct ultracentrifugation as the standard reference method, and have shown varying results. METHODS: In the present study we compared the diagnostic efficacy of the three methods viz., monoclonal antibody (MAb) ELISA, modified polyacrylamide gel electrophoresis (PAGE) and DEM without ultracentrifugation in the detection of rotaviruses from 211 stool specimens. The data were analysed by two latent class model (2LC) in the absence of a gold standard reference method. RESULTS: Rotavirus was detected in 42 specimens by MAb-ELISA; in 40 specimens by PAGE and in 33 specimens by DEM. The estimates of sensitivities and specificities of the three methods were analysed by 2LC method. The analysis revealed no significant variation among the three methods. However, DEM was found with a comparatively lesser sensitivity over the other two methods. INTERPRETATION & CONCLUSIONS: Though DEM was found to be relatively less sensitive than the other two methods, the differences were not significant, and all the three methods were highly specific. Moreover, DEM has the additional advantage of detecting non-group A and other gastroenteritis viruses. The findings suggest the use of highly sensitive and specific MAb-ELISA and PAGE in parallel to detect group A, non-group A and atypical rotavirus infection in the population.

Production of siderophores & effect of iron restriction on the protein profiles of Aeromonas species isolated from water & patients suffering from acute diarrhoeal disease.

Among 48 strains of Aeromonas species, 21 isolates from patients suffering from acute diarrhoea and 27 from metropolitan water samples grown under iron-restricted conditions, 45 strains produced siderophores. Forty one of the 46 strains tested produced siderophores on chrome azurol S (CAS) agar, while 43 isolates did so when the culture supernatants of the bacterial isolates grown in minimal medium were assayed with chrome azurol S assay solution. The whole cell protein profiles of A. hydrophila strains grown under iron restricted conditions expressed new proteins that were not detected in those cultured in iron rich conditions. Five high molecular weight proteins ranging from 70 to 96 kDa were distinctly absent in cultures grown in the presence of iron, indicating their role in iron acquisition by the aeromonads.

Genomic diversity of group A rotavirus RNA from children with acute diarrhoea in Chennai, south India.

Group A rotavirus was identified in 51 of 245 (20.8%) cases with acute diarrhoea in Chennai analysed between December 1997 and March 1999. Forty eight of the 51 specimens were subgrouped and serotyped. A total of 110 rotavirus positive specimens (inclusive of 62 rotavirus positive cases reported earlier) were analysed for their subgroup (SG) specificity and genomic profiles. SGI and SGII specificity were detected in 60 per cent and 20 per cent of the cases studied. Twenty two cases showed dual SG specificity (SGI + II). Nine electropherotypic patterns (7 'short' and 2 'long') were observed with a predominance of short pattern in 87 of the 110 (79.1%) positive cases studied. Long electropherotypes were found in 23 (20.9%). Serotyping of the 48 rotavirus positives revealed a higher proportion of serotype-2 (68.8%) followed by serotype-1 (14.6%) and serotype-3 in 1 case. Mixed infection of G1-G2 was observed among 7 cases analysed, which revealed G[2,1], P[4,8] genotype specificity. Dual infection of P[4]-P[8] genotypes was observed in 12 cases with G[2] specificity.

Occurrence of haemolytic & cytotoxic Aeromonas species in domestic water supplies in Chennai.

A study on the occurrence of Aeromonas species in the domestic water supplies in Chennai showed that as much as 37.9 per cent of the water samples analyzed from various sources harbored Aeromonas spp. Majority of the isolates belonged to Aeromonas sobria (13.7%), A. caviae (11.6%) and A. hydrophila (9.5%). Among the 37 metropolitan water samples analyzed, 11 samples yielded Aeromonas spp. inclusive of three isolates of A. hydrophila, four of A. sobria and two isolates each of A. caviae and A. jandaei. From a total of 28 bore well water samples analyzed, Aeromonas spp. were recovered from 15 samples, comprising five isolates of A. hydrophila, six of A. sobria and four isolates of A. caviae. Aeromonas spp. inclusive of one isolate of A. hydrophila, five of A. caviae, three of A. sobria and one isolate of A. veronii were isolated from 10 of the 30 water packets of various commercial brands sold in Chennai. Of a total of 36 isolates obtained, 32 (89%) produced beta-haemolysin with the titres ranging from 2-32 and 20 isolates (56%) were cytotoxic to vero cell monolayers. All the Aeromonas isolates were resistant to ampicillin and polymyxin B. All A. hydrophila and A. caviae isolates were also resistant to cephalothin and erythromycin and 83.3 per cent of Aeromonas isolates were resistant to erythromycin. Aeromonads resistant to tetracycline, gentamycin, co-trimoxazole and nalidixic acid appear to be emerging. The study revealed that Aeromonas spp. occur in the potable and domestic water supplies and even in the chlorinated water supplies in Chennai city, which are potentially enteropathogenic and hence may be hazardous to public health. In view of these findings drinking and domestic water quality standards need to be re-evaluated.

Biochemical characteristics & secretory activity of Aeromonas species isolated from children with gastroenteritis in Chennai.

An attempt was made to delineate the phenotypic markers for the detection of enterotoxigenic strains of Aeromonas. Eighteen Aeromonas species comprising one isolate of A. hydrophila, six isolates of A. sobria and 11 isolates of A. caviae were obtained from 379 children suffering from acute diarrhoea in Chennai. Nine of these isolates inclusive of three A. sobria and six A. caviae were found to produce secretory response in vitro in the rabbit intestinal mucosa mounted in the Ussing chambers as revealed by significant increases in the short circuit current. Eleven strains hydrolysed aesculin, 8 fermented arabinose, 6 produced acetyl methyl carbinol, 14 produced lysine decarboxylase, 3 fermented salicin, 9 produced beta-haemolysin, 9 produced CAMP-like factor and only two isolates took up congo red dye. None of these phenotypic traits were found to correlate with the in vitro secretory activity.

Isolation of nalidixic acid resistant Campylobacters from cases of paediatric diarrhoea in Chennai.

A short term investigation on the Campylobacter enteritis among children under 10 years of age was carried out in Chennai. The study revealed an isolation rate of 11 per cent in 100 patients suffering from acute diarrhoea comprising C. jejuni (8%) and C. coli. (3%). Among the two culture methods used, the candle jar method was found to be superior to plastic bag incubation system in recovering campylobacters on charcoal cefeperazone deoxycholate agar. While all the isolates were sensitive to ciprofloxacin, all of them exhibited resistance to nalidixic acid.

Analysis of human rotavirus serotypes in children with acute diarrhoea in Chennai by monoclonal antibody based ELISA.

To determine the individual human rotavirus serotypes prevailing in Chennai, 345 stool specimens obtained from children with acute diarrhoea between March 1996 and November 1997, were screened for the presence of rotavirus by the standardized enzyme linked immunosorbent assay (ELISA). Of the 90 (26%) rotavirus positive specimens, 75 (83.3%) were subgrouped and 65 (72.2%) were serotyped with monoclonal antibody based ELISA. Of the 65 specimens that could be serotyped, 52.3 per cent belonged to serotype 2, 24.6 per cent were serotype 4, 15.4 per cent were serotype 1 and 7.7 per cent were serotype 3. Of the 75 specimens typed for their subgroup specificity, 50.7 per cent were subgroup I, 26.7 per cent were subgroup II, 18.7 per cent were equally specific to both subgroups I and II, 4 per cent belonged to nonsubgroups I and II. Our results indicate a predominance of serotype 2 virus. Unusual strain having both subgroups I and II specificity or neither specificity were also encountered.

Incidence & enteropathogenicity of Aeromonas spp in children suffering from acute diarrhoea in Chennai.

A total of 200 stool samples from children below 10 yr suffering from diarrhoea were screened for enteric pathogens with special interest on Aeromonas. Aeromonas spp were isolated from 6.5 per cent of the patients, comprising 4 per cent A. hydrophila, 2 per cent A. sobria and 0.5 per cent A. caviae. Among the 13 isolates obtained, 10 isolates produced enterotoxin in ligated rabbit ileal loops, and 11 produced cytotoxin in HEp 2 cells. Many of the Aeromonas isolates exhibited resistance to commonly used antibiotics such as trimethoprim, sulphdiazine, chloramphenicol and tetracycline. None of the stool samples obtained from 52 age matched control children yielded Aeromonas species. Four isolates of Salmonella typhi, 7 of S. paratyphi A, 6 of Shigella flexneri, 4 of Sh. dysenteriae and 3 isolates of Vibrio cholerae (Ogawa) were also recovered during the study. Among the samples analyzed, one from a 7 yr old female patient, had A. hydrophila with S. paratyphi A. The results of this study indicate that drug resistant enteropathogenic Aeromonas is also an important etiological agent of childhood diarrhoea in Chennai.

Cryptosporidium parvum--propagation of oocyst in neonatal calves.

Shedding of Cryptosporidium parvum oocyst was studied in experimentally infected Jersey-Sindhi cross bred calves. Three 7 day old bull calves housed in isolation were orally infected with 10(8) oocysts of Cryptosporidium parvum. The prepatent and patent period of the experimental infection were 5 and 4 days respectively. Maximum oocyst output [2 x 10(5) oocyst per gram (OPG) was observed on the 7th day post inoculation (PI). The mean total oocyst output was 2.5 x 10(7). Diarrhoea started on the second day of oocyst shedding.