David B. Elkins

Liver fluke infection and cholangiocarcinoma : Model of endogenous nitric oxide and extragastric nitrosation in human carcinogenesis

Cancers arising during bacterial, viral and parasitic infection provide useful models to investigate the link between inflammation and carcinogenesis. Because the inflammatory agent is known, relationships between immune responses, the production of DNA-damaging agents, such as nitric oxide, oxygen radicles and N-nitroso compounds, and cancer risk can be explored. This paper first describes the close relationship between infection with the liver fluke, Opisthorchis viverrini, and cholangiocarcinoma in humans. Data are then presented which demonstrate an elevation in levels of salivary nitrite and urinary and plasma nitrate among men with moderate and heavy liver fluke infections compared to uninfected controls which was absent 4 months after the parasites were cleared with praziquantel. Because of the strict control over subject selection and dietary intake plus the absence of the increase following treatment, we conclude that the higher levels of nitrate and nitrite reflect endogenous generation of nitric oxide resulting from liver fluke infection. Excess nitric oxide generation in the inflamed tissue is likely to lead directly to the formation of N-nitroso compounds mediated by activated macrophages. Further work will attempt to demonstrate a link between this increase and both parasite-specific immune responses and the risk of cancer.

A high frequency of hepatobiliary disease and suspected cholangiocarcinoma associated with heavy Opisthorchis viverrini infection in a small community in north-east Thailand

A group of 87 adults from a small village in north-east Thailand was chosen to undergo ultrasound investigation based on their intensity of infection with the liver fluke, Opisthorchis viverrini, or clinical status (history of jaundice, current hepatomegaly). From this group, 8 cases of suspected early cholangiocarcinoma were found, and the diagnostic features of 6 of the 8 were confirmed by computerized tomography scan and endoscopic retrograde cholangiopancreatography. In addition, several cases of mild gall-bladder disease, chronic cholecystitis, cholelithiasis and parenchymal liver disease were detected. A highly significant positive relationship between the intensity of liver fluke worm burden and the severity of biliary tract disease within individuals is reported. These results indicate that Opisthorchis is associated with moderate to severe hepatobiliary disease in a considerable proportion of infected individuals.

Opisthorchis viverrini : relationships between egg counts, worms recovered and antibody levels within an endemic community in Northeast Thailand

Three techniques for estimating the intensity of Opisthorchis viverrini infection in individuals from a Northeast Thai community are compared. Egg counts were determined using a quantitative formalin/ethyl acetate technique, worm burdens were estimated by expulsion chemotherapy and antibody levels were measured by ELISA. Log-transformed worm and egg counts were closely correlated (r = 0.80), suggesting that both measurements provide good assessments of relative intensity of infection. However, no Opisthorchis worms were recovered from 34 people with high egg counts; probably due to problems with the expulsion technique in some individuals. Examination of egg production per fluke indicated that each fluke contributed an average of 180 eggs per gram (epg) of faeces and fecundity was negatively associated with total worm burden. Serum IgG levels correlated significantly with Opisthorchis egg count (r = 0.61) at two independent assessments. Although significant associations were observed between antibody levels and echinostome infection, analysis suggested that these reflected independent associations between these two variables and Opisthorchis infection and age. We conclude that all three measurements are useful for epidemiological studies.

Relationship between intensity of Opisthorchis viverrini infection and hepatobiliary disease detected by ultrasonography

Twenty‐four locality‐, age‐ and sex‐matched groups of village residents with no light, moderate and heavy Opisthorchis viverrini infection were examined by ultransonography. Highly significant differences were observed between the groups in the relative size of the left lobe of the liver and the fasting and post‐meal size of the gall‐bladder. In addition, indistinct gall‐bladder wall, the presence of gall‐bladder sludge and strongly enhanced portal vein radicle echoes were most frequently observed in the heavily infected group. Two suspected cases of cholangiocarcinoma were identified from the heavy group. The results highlight the importance of intensity of infection on the frequency and severity of fluke‐associated hepatobiliary disease.

Immune responsiveness and parasite-specific antibody levels in human hepatobiliary disease associated with Opisthorchis viverrini infection.

Opisthorchis viverrini infection is associated with human hepatobiliary disease and cholangiocarcinoma, but the role of the immune response in the pathogenesis of infection is unclear. Here ultrasonography was used to examine the biliary tracts of residents from an endemic community. Delayed‐type hypersensitivity responses to unrelated antigens, and fluke‐specific IgG and IgA levels in serum of this group were also examined. Relationships between immunological parameters, intensity of infection and radiologically measured variables are reported. Immune responsiveness to unrelated antigens did not vary with intensity of parasite infection or disease status. Of all the variables, IgG levels were most markedly elevated in disease cases compared with normal subjects and were closely associated with gall bladder size and dysfunction. This is consistent with the hypothesis that an immunopathologic mechanism is involved in opisthorchiasis and suggests that antibody levels may be useful in screening populations for fluke‐associated hepatobiliary disease.

Opisthorchis viverrini infection in northeast Thailand and its relationship to cholangiocarcinoma

There is strong epidemiological and experimental evidence supporting an important aetiological role for liver fluke infection in the development of cholangiocarcinoma (CHCA) in humans.’-24 However, compared with other experimental cancer models, little research has focused on the mechanisms of this relationship since the first inductions of CHCA in infected hamsters by Thamavit et ~ l . ~ . ’ ~ and Flavell and L ~ c a s . ” ’ ~ Fluke-associated CHCA is not widely discussed, although it has several advantages as a model for carcinogenesis. First, the influence of fluke infection on the susceptibility of bile ducts to malignancy appears to be very strong compared with most other known risk factors of human cancer. Cholangiocarcinoma occurs with a high frequency in infected p e ~ p l e , ’ ~ ’ ~ but only rarely in the absence of i n f e c t i ~ n . ~ ~ ? ’ ~ Bile duct carcinogenesis can be elicited relatively easily in fluke-infected hamsters under conditions where it does not occur in uninfected animals.’-15 Second, since Opisthorchis per se probably does not alter DNA directly, the enhanced susceptibility to carcinogens may closely mimic general pathways of malignancy not associated with biological agents. Research may therefore be widely applicable to human cancer. Third, unlike viruses and unknown aetiological agents, Opisthorchis can be eliminated by simple treatment with the drug, praziq~antel.”-’~ Thus the conditions thought to enhance cancer susceptibility can be assessed in the same individual in the presence and absence of one aetiological agent. This review highlights recent epidemiological literature concerning Opisthorchis and CHCA in humans in Northeast Thailand. An excellent review of Clonorchis sinensis was recently published by Rim.30 Since experimentally induced CHCA, which clearly identifies a role for Opisthorchis in carcinogenesis, is only briefly mentioned here, interested readers are directed to Flavell’ ’ and recent papers of Thamavit et al.14.15 In closing this review, we suggest areas for further research to elucidate the pathogenesis of this cancer by considering recent studies of inflammatory responses and carcinogenesis.

Parasite-associated morbidity: Liver fluke infection and bile duct cancer in Northeast Thailand

Infection with the liver fluke, Opisthorchis viverrini, remains a major public health problem in Northeast Thailand, where approximately one-third of the population is infected. The northeast region is largely populated by Laos-descendent Thais who enjoy eating raw fish, which harbour the infective stage of the fluke. The parasite has maintained its presence in the population despite the widespread use of praziquantel and dissemination of health education material throughout the region by vigorous government-sponsored programs in recent years. The most severe consequence of liver fluke infection is cholangiocarcinoma, i.e. cancer of the bile duct epithelium. Although mortality due to the parasites alone appears to be uncommon, cholangiocarcinoma arising as a result of infection is one of the leading causes of death in the region. This paper reviews the pathogenesis of infection and the geographic, hospital-based and community studies which demonstrate the close relationship between infection and cancer. In addition, data from the Cancer Registry of Khon Kaen, Northeast Thailand and population-based studies using ultrasonography to visualize early tumours which illuminate the very high frequency of the cancer among heavily infected individuals and communities are discussed. Finally, the paper will close with a brief commentary on the prospects for control of the parasite and its likely impact on the frequency of cancer given the current epidemiological situation of liver fluke infection.

Contexts and patterns of men's commercial sexual partnerships in northeastern Thailand: Implications for AIDS prevention

Results of an exploratory research project elaborating the contexts, patterns and specific scenarios of the commercial sexual activity of northeastern Thai men are reported. Data were collected using face-to-face surveys, focus groups, key informant interviews and observations in 32 northeastern villages (n = 744 men), 18 migrant labour camps housing sugarcane workers (n = 219 men), and five cattlemarkets in northeast Thailand. Fifty percent of married men and 43% of single men had visited female sex workers (FSW). Female sex worker visits occurred primarily prior to marriage, though 13% of married men had purchased sexual services within the past year. Nonmarital sexual activity was set within the socio-cultural frameworks of poverty, circular migration, a large commercial sex sector, and a belief system about mens sexuality and mens and womens gender roles. Sexual services were typically purchased as part of friendship group partying (paiy tiaow) and generally included heavy alcohol consumption. The most common scenario for visiting FSWs involved brothels, though cattlemarkets, festivals, and migrant labour situations were also scenarios for FSW contact. These each had unique characteristics that affected the likelihood that condoms would be used. The further the specifics of a scenario (as evaluated by men) diverged from those of brothel contact with an FSW, the less likely men were to identify this as having the potential for HIV transmission and the less likely they were to use a condom. AIDS prevention campaigns must be developed that are sensitive to the socio-cultural framework, contexts and specific scenarios within which nonmarital sexual contacts occur.

Reversal of biliary tract abnormalities associated with Opisthorchis viverrini infection following praziquantel treatment

We recently demonstrated a number of biliary tract abnormalities associated with moderate to heavy Opisthorchis viverrini infection among 95 selected village residents in north-east Thailand, who were treated with praziquantel immediately after initial examination and re-examined by abdominal ultrasonography 10 months later. Dramatic improvement in the gall-bladder status of previously infected individuals was observed, including reduction of gall-bladder length and width and regained contractibility of the organ following fatty meal consumption. The frequency of severe enhanced portal vein radicle echoes and gall-bladder sludge and wall irregularities also decreased following treatment, but persisted in many individuals. Two masses were observed in the right hepatic lobe of one individual who was previously considered normal. The findings suggest that gall-bladder abnormalities are reversible following elimination of liver fluke infection, but malignancies, once initiated, are not likely to be affected by treatment.

Distribution patterns of Opisthorchis viverrini within a human community

This paper describes patterns of infection with Opisthorchis viverrini within a human community assessed by egg count, parasite-specific antibody levels and worm burden determined by expulsion chemotherapy. All three measurements increased significantly with age, reaching a plateau in young adults. There were no significant sex-associated differences in the prevalence or intensity of infection, antibody levels, or in the frequency of heavy worm burdens. A striking 81.5% of the total Opisthorchis population and 74% of the total egg output were expelled by the most heavily infected 10% of the humans sampled. This extreme over-dispersion, together with recent findings that hepatobiliary disease frequently accompanies heavy infections, argues that the priority for community-based parasite control programmes should be to maximize treatment and health promotion efforts toward heavily infected individuals.