Occurrence and susceptibility to antibiotics of Shigella species in stools of hospitalized children with bloody diarrhea in Pakistan.

The aim of the present investigation was to study the frequency of Shigella spp. in patients with bloody diarrhea in Pakistan and the susceptibility of isolated Shigella to three antibiotics: ampicillin, cotrimoxazole and nalidixic acid. In addition, the frequency of Campylobacter and Salmonella was also determined. Stool samples (n = 152) were collected from 152 diarrheic children less than six years of age passing blood and mucus in their stools who were admitted to Paediatric Department of Mayo Hospital in Lahore, Pakistan from June to September 1990. The samples were cultivated on standard media for Shigella, Campylobacter, and Salmonella. Susceptibility of Shigella isolates was tested by disk diffusion method. The frequency of isolation was 19.1% for Shigella spp., 7.9% for Campylobacter, and 4.6% for Salmonella. Shigella flexneri (7.9%) was the most frequently isolated species, followed by S. dysenteriae (6.6%), S. boydii, (3.3%) and S. sonnei (1.3%). All Shigella isolates were susceptible to nalidixic acid (100%), while only a few were susceptible to cotrimoxazole (7.0%) and ampicillin (3.5%). In Pakistan, self-medication and purchases of drugs without a prescription are commonly practiced. Thus, there is a greater possibility of development of resistant strains due to over use of antibiotics.

Norfloxacin resistance in Campylobacter jejuni and Campylobacter coli isolates from Swedish patients.

This study focused on the frequency of Campylobacter jejuni and Campylobacter coli strains resistant to norfloxacin. Included were 1659 consecutive stool specimens isolated between 1992 and 1995, from as many Swedish patients with diarrhoea. The patients were divided with regard to place of infection and age. All strains were tested for susceptibility to norfloxacin by means of disc diffusion test on blood-agar plates. Norfloxacin-resistant strains (n = 310) were furthermore tested for resistance to doxycycline and erythromycin. The Etest was used for determination of MIC values of doxycyclin, erythromycin and norfloxacin of 81 of the strains. C. jejuni and C. coli isolations resistant to norfloxacin were significantly more frequent among patients infected abroad, especially in Spain and Thailand, compared with those infected in Sweden, adults more often than children. The number of resistant strains showed a yearly increase, and the difference between children and adults was equalized in 1995. The MIC50 and MIC90 values for doxycycline and erythromycin have increased markedly through the 4 years studied. This study shows that norfloxacin, because of increased resistance, may have limited utility for treatment of gastrointestinal infections caused by C. jejuni and C. coli.

In vitro studies of Campylobacter jejuni/coli strains from hens and humans regarding adherence, invasiveness, and toxigenicity.

Campylobacter jejuni/coli strains from hens and humans were compared for their ability to adhere to and invade HEp-2-cells and for toxigenicity to CHO-cells. In both hen and human strains, invasiveness was higher among non-toxigenic strains than among toxigenic ones. The frequency of adherence, invasiveness, and toxigenicity was the same in hen and human strains.

Campylobacter jejuni/coli and enterotoxigenic Escherichia coli (ETEC) in faeces from children and adults in Tanzania.

The occurrence of Campylobacter and enterotoxigenic E. coli (ETEC) was studied in faecal samples from Tanzanian children (< 5 years of age), adolescents and adults (only Campylobacter) with and without diarrhoea. The Campylobacter strains isolated were tested for subspecies, enterotoxigenicity and serotype. Out of 394 children with diarrhoea 18% were infected with Campylobacter and 20% with ETEC. In 278 samples tested for Campylobacter and 136 tested for ETEC from asymptomatic children the corresponding numbers were 12 and 5%, respectively. In children < 18 months with diarrhoea Campylobacter was noted in 22% and ETEC in 18%, whereas the figures were 11 and 4% respectively in asymptomatic children. In the age group 18 months to 5 years Campylobacter was demonstrated in 2% of the children with diarrhoea and 27% had ETEC, while the figures were 15 and 8% for asymptomatic children. Among adults the prevalence of Campylobacter-positive samples was 1% both for symptomatic and asymptomatic individuals. There were no seasonal differences in the prevalences of both Campylobacter and ETEC either in the symptomatic or the asymptomatic group. Campylobacter jejuni was the dominating Campylobacter species among both symptomatic and asymptomatic individuals. C. jejuni strains from patients with diarrhoea were significantly more often enterotoxigenic than were C. coli strains. The serotype pattern regarding Campylobacter was in general similar for symptomatic and asymptomatic individuals. We conclude that Campylobacter and ETEC are common causes of bacterial diarrhoea in Tanzanian children, and that Campylobacter infections are more important in children younger than 18 months, than in older ones.

Flies and water as reservoirs for bacterial enteropathogens in urban and rural areas in and around Lahore, Pakistan.

The study was conducted to isolate and characterize campylobacter, enterotoxigenic Escherichia coli-labile toxin (ETEC-LT), shigella and salmonella in flies and water. The material for the study, flies (n = 300) and water samples (n = 148), was collected from different localities in and around Lahore, Pakistan. Cultivation of the samples was performed on conventional standard media. Membrane filtration technique was used for water prior to culture. Determination of ETEC-LT was done by GM1 ELISA. Results of our study showed that flies and water were reservoirs for all the four pathogens, campylobacter, ETEC-LT, shigella and salmonella. Flies from the village were carrying fewer enteropathogens, while water from the village was found to be more contaminated as compared to the city. Campylobacter and ETEC-LT were the most frequently isolated pathogens in both flies and water. Thus the incidence of diarrhoeal disease in children of developing countries may be decreased by providing plenty of safe drinking water, improving excreta disposal, toilet facilities and giving education in personal hygiene.

Early child health in Lahore, Pakistan: VIII. Microbiology.

The causative agents of acute diarrhoea were investigated in children under the age of five years from 1985-1991 in three socio-economically different areas in Lahore, Pakistan. The aim was to determine the frequencies of the most common enteropathogens in faeces. The total isolation rate was 73.4%; two thirds (53.5%) were of bacterial and one third (19.9%) of viral origin. ETEC-LT (23.4%), Rota virus (19.9%), EPEC (15%) and Campylobacter (12%) were the single most frequent pathogens in all age groups and areas of living. The positivity of Campylobacter increased from 1.6% to 12% after the change of isolation technique. Shigella was isolated significantly (p > 0.001) more in children over than under one year of age. The incidence of bacterial infections was high during all seasons, while in certain years Rota virus was relatively low in the summer compared to the cooler months. The prevalence of ETEC-LT diarrhoea was higher in the periurban slum as compared to the village and the urban slum. The study is the first of its kind in Pakistan, and the results are similar to other comparable studies. It is thus possible to establish a well functioning and reliable microbiological laboratory in developing countries in a setting with restricted trained personnel and material resources.

Frequency and enterotoxigenicity of Campylobacter jejuni and C. coli in domestic animals in Pakistan as compared to Sweden.

Domestic and wild animals have been considered to be the most common reservoirs of Campylobacter for human infections. This study has been conducted to establish the frequency and enterotoxigenicity of C. jejuni and C. coli in the stools of domestic animals in a developing versus a developed country. Faecal samples of 300 chickens and laying hens, 102 horses, 180 cows, 92 goats and sheep were collected from Pakistan and Sweden. The hippurate hydrolysis test and CHO-cell test were performed for species differentiation and enterotoxigenicity determination respectively. The pattern and sequence of frequency distribution of Campylobacter in animals in Pakistan and Sweden were similar, the main difference being a significantly higher frequency in Pakistani samples. C. jejuni was the dominant species from both countries and Pakistani strains were more enterotoxigenic than Swedish strains. We conclude that the spread of Campylobacter infection in developing countries originates from different sources in the neighbourhood, for example animals, and unless hygienic conditions improve no change can be expected.

Adherence, enterotoxigenicity, invasiveness and serogroups in Campylobacter jejuni and Campylobacter coli strains from adult humans with acute enterocolitis.

Two hundred Campylobacter jejuni and Campylobacter coli strains from the same number of adult Swedish patients with acute enterocolitis were tested regarding adherence to and invasiveness in HEp-2 cells and for enterotoxigenicity by the CHO-cell assay. The serogroup characteristics, heat-stable and heat-labile, for each strain were also investigated. Eighty-four percent of the strains were classified as C. jejuni and 16 percent as C. coli. All of the strains were adherent to HEp-2 cells, 39% were invasive and 31.5% enterotoxigenic. We found significantly more invasive strains in the non-enterotoxigenic group than in the enterotoxigenic one. There would seem to be no correlation between enterotoxigenicity or invasiveness and serogroup. The results of this study suggest the existence of multiple mechanisms for C. jejuni- and C. coli-induced diarrhoea and that the mechanisms may differ from one strain to another.

Enterotoxigenicity and frequency of Campylobacter jejuni, C. coli and C. laridis in human and animal stool isolates from different countries.

Campylobacter jejuni and C. coli strains were collected during three different years from adult patients with enterocolitis in Sweden (n = 372) from 49 patients in Kuwait, and Campylobacter strains from hens from Mexico, Pakistan and Sweden (n = 107) and Swedish pigs (n = 47). C. jejuni was the predominant species in human and hen isolates, and C. coli in pigs C. coli was significantly more common in human isolates from Sweden, and more common in hen isolates from Pakistan, than in hens from Sweden and Mexico. C. laridis was only isolated from pigs (17%) and was in no case enterotoxigenic. Both in human and hen isolates, C. jejuni strains were more enterotoxigenic than C. coli strains. C. jejuni strains from Swedish hens were less enterotoxigenic than those from Pakistan and Mexico (P less than 0.001), and strains from pigs were less enterotoxigenic than those from hens (P less than 0.001). We conclude that C. jejuni are more often enterotoxigenic and possibly more virulent than c. coli and C. laridis. The relative frequency of C. jejuni and C. coli in humans and animals differs from one country to another.

Enterotoxin production and serogroups of Campylobacter jejuni and Campylobacter coli from patients with diarrhea and from healthy laying hens.

Enterotoxin production, a possible virulence factor, was determined in Campylobacter jejuni and Campylobacter coli by two different techniques, the CHO cell test and the GM1 enzyme-linked immunosorbent assay. The frequency of enterotoxigenic Campylobacter strains was 32% in strains from both humans with acute enteritis and healthy laying hens, as measured by the CHO cell test. The CHO cell test was significantly more sensitive than the GM1 enzyme-linked immunosorbent assay in the detection of enterotoxigenic strains. Enterotoxin production was compared with the presence of heat-stable and heat-labile antigens. There was no significant correlation between enterotoxin production and serogroups for C. jejuni or C. coli. The difference in enterotoxigenicity between C. jejuni (34.1%) and C. coli (21.9%) was not significant.

Comparison of different procedures, transport media, and enrichment media for isolation of Campylobacter species from healthy laying hens and humans with diarrhea.

A comparison was made with some of the available transport and enrichment media for the isolation of Campylobacter species. The merits of duplicate versus single sampling were also investigated. Fecal samples from 721 healthy laying hens and 328 patients with diarrhea were included in the study. The media compared were modified Stuart medium, Campythio-glycolate medium, semisolid motility test medium, and Campylobacter enrichment broth. The isolation rate of Campylobacter species was significantly increased by taking duplicate instead of single samples. The highest outcome was found when one of the two samples was an enrichment medium, such as semisolid motility test medium.

Natural campylobacter colonization in chickens raised under different environmental conditions.

A cross-sectional study of 447 laying hens (age range 0-65 weeks) and a longitudinal study of 164 similar birds showed that Campylobacter jejuni was not present in the faeces of newly hatched chicks, but that colonization arose after 5-9 weeks. A survey of 250 broilers obtained from four breeders showed that all were negative for C. jejuni before and after slaughter at the age of 5 weeks. Once C. jejuni had appeared in a flock, it rapidly spread to virtually all birds, but at the age of 42 weeks only 20-46% of birds remained colonized, possibly as a result of having developed immunity. Birds housed in the protective environment of a laboratory still became colonized (after 9 weeks). The mode of infection is unknown, but water and food were bacteriologically negative and were deemed to be unlikely sources. Transmission via attendants, flies or other insects remain possibilities. It is concluded that prevention of colonization might be possible within the life-span of broiler chickens (5-7 weeks), but that it would be difficult to extend this period. There is a need to define how colonization arises so that the feasibility and cost of possible preventive measures can be assessed.

Methodological aspects of an agar plate technique for determination of biologically active 5-fluorouracil in blood, urine and bile.

A microbiological agar plate technique for estimation of 5-fluorouracil concentrations in blood, urine and bile from man, dog and pig was evaluated. Different bacterial test strains, media modifications and techniques for inoculation were studied. The strain Streptococcus faecalis ATCC 8043, recommended previously by Clarkson et al., was found to be the most suitable. The influence of prediffusion, dilution, antibiotics and chemotherapeutic agents and their antagonists, as well as the effect of storage of samples containing 5-fluorouracil were examined. A detailed methodological description is presented. The method seems to be sufficiently sensitive and practical for routine determination of cytotoxic compounds from 5-fluorouracil in serum, plasma and urine.