Waterborne transmission of Giardia and Cryptosporidium

Giardia and Cryptosporidium spp. are parasitic protozoa which are frequent etiologic agents of waterborne diseases. This lecture will summarize the main biological and environmental factors involved in the potential risk for waterborne transmission of giardiosis and cryptosporidiosis, which have caused many outbreaks in different geographical areas. In particular, the current epidemiological situation of these parasitoses in Italy will be analysed, on the basis of research carried out on humans and on the environment. Finally, current methods for evaluating the presence of Giardia cysts and Cryptosporidium oocysts in water and new methods for cyst/oocyst removal from drinking water and wastewater will be examined.     

Characterization ofCampylobacter jejuni lipopolysaccharide

Lipopolysaccharides were isolated from dehydratedCampylobacter jejuni by combination of the phenol-chloroform-petroleum ether and phenol-water extraction techniques. Biochemical characterizations of lipopolysaccharide were performed on the two fractions of highest purity. Neutral sugar analyses detected galactose, glucose, trace amounts of mannose, and an unidentified deoxy-hexose. The primary amino sugars were galactosamine, glucosamine, and glucosamine-phosphate. Chemical analyses of other lipopolysaccharide components included phosphate, 3-deoxy-d-manno-octulosonic acid (KDO), and fatty acids. The predominant fatty acids were 3-hydroxytetradecanoic and hexadecanoic acids with lesser amounts of tetradecanoic acid. 3-Hydroxytetradecanoic acids were bound to lipid A by both amide and ester linkages.     

Pathogenicity ofCampylobacter jejuni in intraperitoneally or intravenously inoculated ferrets

Ferret kits inoculated intravenously (IV) withCampylobacter jejuni after pretreatment with parenteral iron developed more severe systemic signs and more prolonged bacteremia than untreated inoculated controls. Watery diarrhea began in both groups 2–16 h after inoculation and lasted less than 48 h.C. jejuni was cultured from rectal swabs 2–8 h after inoculation, and gut colonization persisted up to 15 days, suggesting that colonization does not necessarily induce diarrhea. Gut colonization occurred as rapidly after IV inoculation of ferrets in which the common bile duct had been ligated as it did in unligated controls.C. jejuni apparently reached the intestinal lumen by mucosal invasion from the bloodstream. Bacteremia following natural infection could thus result in repeated passages ofC. jejuni across the gut wall, exposing the mucosa to both the bacterial cells and their metabolic products. Histological evidence of an inflammatory response in the mucosa, without severe epithelial damage, suggests a toxin-mediated secretory diarrhea.     

Antigenic differences in congenic chicken-colonizing and noncolonizing strains ofCampylobacter jejuni

A congenic pair ofCampylobacter jejuni has been previously developed in our laboratory that will (strain A74/C) and will not (strain A74/O) colonize 2-day-old chicks dosed with 10⁵ colony forming units. Outer membrane protein (OMP) extracts of these organisms were prepared and studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analyses. No consistent differences between the colonizer and noncolonizer were detected by SDS-PAGE. However, an antigen of 28 kD molecular weight was consistently found in the noncolonizer, but only at greatly reduced levels or not at all in the colonizer by Western blot analysis with rabbit anti-OMP serum. After affinity purification and cross absorption of serum against OMP from the colonizer, an antigen of 69 kD molecular weight was found unique to the colonizing strain. Exclusive association of the 69 kD antigen with the colonizing strain suggests that it may be a colonization factor.     

Genomic characterization ofCampylobacter jejuni by field inversion gel electrophoresis

The genome ofCampylobacter jejuni was characterized by field inversion gel electrophoresis (FIGE) after digestion with three rare-cutting restriction endonucleases. The restriction enzymesSac II (5-CCGCGG),Sal I (5-GTCGAC), andSma I (5-CCCGGG) were found to produce 13, 5, and 8 fragments respectively from theC. jejuni genome. The fragment sizes ranged from 1.6 kb to 1300 kb, which gaveC. jejuni a genome size of approximately 1900 kb. Furthermore, thegly A and rRNA genes ofC. jejuni were localized to specific fragments by use of Southern analysis, and thegly A gene was shown to be closely linked to one of the three rRNA genes.     

Waterborne cryptosporidiosis

Awareness of the importance of Cryptosporidium as a gastrointestinal parasite of developed countries not only stems from its prevalence in AIDS patients but also from its recent recognition as a possible contaminant of drinking water supplies. The importance of Cryptosporidium to public health has recently been revealed by a series of major epidemics of diarrhoeal disease in the USA and UK. In this review, Huw Smith and Joan Rose document what is known of the causes of some of these outbreaks and explain why this parasite can escape the battery of treatment processes normally used for drinking water supplies in these countries.     

Cloning and sequence analysis of the flagellin gene ofCampylobacter jejuni TGH9011

Flagella are essential for motility and have been implicated to be one of the pathogenic determinants. The flagellum ofCampylobacter jejuni is a polymeric structure of a 62-kd protein. Using a high-affinity flagellin antibody to screen a lambda gt 11 phage genomic expression library ofC. jejuni strain TGH9011 (Serotype LIO36), a recombinant phage clone lambda gt 11RK that expresses theC. jejuni flagellin protein was isolated. The recombinant lambda gt 11 RK produced a 56-kd protein upon induction with isopropylthiogalactoside, which reacted specifically with anti-flagellin antibody. The flagellin gene was sequenced, and comparative analysis of the nucleotide and amino acid sequence identified a region of the flagellin that shows hypervariability among differentCampylobacter species and strains.     

Detection and identification ofCampylobacter coli andCampylobacter jejuni by two-step polymerase chain reaction

Flagellin gene was used as target sequence to detect and distinguishC. coli andC. jejuni by a “nested PCR” technique. The method shows a high level of sensitivity and specificity. Application of this rapid diagnostic tool could provide further information about epidemiological and pathogenetic implications of each of these two microorganisms.     

Southern blotting analyses of strains ofCampylobacter fetus using the conserved region ofsapA

Chromosomal DNA of 27 strains of Campylobacter fetus was analyzed by Southern blotting with a probe of the conserved region of sapA. The probe hybridized with 23 strains that produced type A lipopolysaccharide. These strains had more than six sapA homologs. In Southern blots of SalI-digested chromosomal DNA separated by pulsed-field gel electrophoresis, one fragment from 19 strains and two fragments from 4 strains hybridized. These data indicate that multiple sapA homologs are localized to a limited region on the chromosomal DNA of C. fetus and thus suggest the possibility of developing a typing system using this method. Received: 28 June 1995 / Accepted: 19 September 1995     

Characterization of the antigens involved in serotyping strains ofCampylobacter jejuni by passive hemagglutination

Components of outer membrane preparations, heated saline extracts, and phenolwater lipopolysaccharide extracts obtained from strains ofCampylobacter jejuni representing seven passive hemagglutination serotypes (Penner serotypes 1–4, 13, 16, and 50) were electrophoresed in sodium dodecyl sulfate-polyacrylamide gels. Tests of gel eluates demonstrated that lipopolysaccharide antigens are involved in serotypingC. jejuni by passive hemagglutination and that other cell surface components have no activity. This finding was confirmed by hemagglutination inhibition. In the typing ofC. jejuni by passive hemagglutination, each serotype is probably defined by the presence of one or more specific lipopolysaccharides. These findings may lead to a clarification of the serotyping nomenclature for those systems that depend on passive hemagglutination. It is recommended that a single internationally agreed numbering system be adopted for lipopolysaccharides derived fromC. jejuni.     

Waterborne cryptosporidiosis: current status

In the past ten years Cryptosporidium oocysts have been shown to be common contaminants of water, causing at least 19 waterborne outbreaks of cryptosporidiosis which have affected more than 427 000 individuals. Recommended methods for oocyst isolation and enumeration are time-consuming and inefficient and experts state that the absence of Cryptosporidium oocysts in drinking water can never be guaranteed. In the UK alone, a National Research Programme costing pound3 million has been undertaken. Here, Huw Smith and Joan Rose review the current status of knowledge and identify approaches taken by UK and USA Government regulatory bodies in order to reduce the likelihood of waterborne transmission.     

Waterborne toxoplasmosis - Recent developments

Humans become infected with Toxoplasma gondii mainly by ingesting uncooked meat containing viable tissue cysts or by ingesting food or water contaminated with oocysts from the feces of infected cats. Circumstantial evidence suggests that oocyst-induced infections in humans are clinically more severe than tissue cyst-acquired infections. Until recently, waterborne transmission of T. gondii was considered uncommon, but a large human outbreak linked to contamination of a municipal water reservoir in Canada by wild felids and the widespread infection of marine mammals in the USA provided reasons to question this view. The present paper examines the possible importance of T. gondii transmission by water.     

Waterborne Escherichia coli O157

The waterborne route of Vero cytotoxin-producing E. coli (VTEC) O157 infection was first suggested in two unconnected human cases in 1985. Since then, waterborne VTEC O157 has been identified in sporadic cases and in outbreaks of illness. Recreational waters, private and municipal supplies have been implicated from microbiological, environmental and epidemiological studies of cases. In addition, a research cohort study of farm workers identified exposure to private water supplies as a risk factor for having antibodies to E. coli O157. Sources of contamination are thought to be animal and human faeces or sewage. The presence of low numbers of target organisms in water makes microbiological confirmation difficult, therefore epidemiological evidence has been essential in outbreak investigations. Despite the potential for contamination of water with VTEC O157, waterborne infection is relatively rare largely due to the susceptibility of the organism to water treatment processes. This paper presents the evidence for waterborne VTEC O157 infection, considering current microbiological, environmental and particularly epidemiological information.     

Rotaviral enteritis in a raccoon

A hand-reared raccoon (Procyon lotor) kit had severe diarrhea and died within 24 hr. Gross and histopathologic findings were compatible with a diagnosis of viral enteritis. The immunoperoxidase test revealed rotavirus group A antigen in the intestinal mucosa. This is the first record of rotaviral enteritis in a raccoon.