Sample collection is an important aspect of scientific work because it shapes, to a great extent, the study design and methodology, both of which may influence the outcomes of scientific research. However, often in scientific evaluations of studies which involve both field sample collection and laboratory work, only the laboratory research aspect receives serious attention, while other factors such as the socio-cultural, ecological and belief values of subjects who donate samples for laboratory studies are much less emphasised. These factors and how they play out in any particular study area are critical determinants of successful field sample collection especially in the developing countries.
by Dr Olufunmilola Ibironke, Dr Samuel Asaolu and Dr Clive Shiff
Urinary schistosomiasis is caused by a trematode worm, Schistosoma haematobium [1]. Infection with this parasite has been shown to be the commonest cause of haematuria and urogenital diseases in endemic areas. Thus, detection of haematuria in urine has been proposed as a valid indicator of schistosome infection, and has been widely adopted in many national schistosomiasis control programmes [2,3]. Diagnostic procedures in control programmes accordingly involve collection of urine samples from patients.
Most studies of urinary schistosomiasis in Nigeria and other endemic countries have targeted schoolchildren [4-8], because they represent the prime reservoir for the parasite, and children are amenable to mass chemotherapy [9]. However, studies have shown the debilitating effect of the parasite among adults in communities where it is endemic [10-13] and so this population also needs to be studied. As opposed to urine sample collection from children which is mostly done in schools, collection of urine from adults is difficult, particularly among persons who do not consider schistosomiasis as their major health problem when compared to malaria. In a school-based setting, after obtaining clearance from government health and school administrative authorities, researchers usually work with school teachers to get permission from pupils’ parents, and to educate the children involved in the study about how to follow urine sample collection instructions. However, for studies which involve adults, researchers, with the help of local health officers, would have to deal with patients directly to seek their individual involvement in the study, the acceptance of which depends on a number of the above mentioned factors.
Few studies have investigated the sociology of communities involved in such studies. We present here a study on urinary schistosomiasis in two villages in Ogun State, Nigeria, involving collection of urine samples from adults, to investigate the factors that drive their acceptance or refusal for inclusion in the study.
Methods and study sites
The study involved adults between the ages of 20 and 55 years who were mobilised to school halls in each village through the respective heads of the villages. Participants were informed of their right to accept or reject inclusion in the study. Many adults refused to come to school halls, many others who came rejected inclusion in the study. Some others accepted inclusion and collected urine sample containers but never came back while others accepted full participation. People in endemic communities show negative attitudes to urine sample collection for different reasons. To find out villagers’ attitudes to the urine sample collection process, we asked consenting participants why their friends or family refused to participate and in the process we identified some factors responsible for their attitudes. We also visited some households either to seek consent for inclusion or to understand reasons for refusing inclusion in the study.
This study was conducted in July, 2010, in Ogun State, Nigeria as a part of a study on the diagnosis of urinary schistosomiasis in six villages. For the purpose of comparison, two villages, Apojola located in Odeda Local Government Area (LGA), and Ogbere in Ijebu-east LGA, were selected. Apojola is located on Oyan Dam Reservoir. The inhabitants are all immigrant fishermen and their families, and are a mixture of Moslem Hausas and Christian Idomas. Awawa River serves Ogbere community. The inhabitants are mainly Christian Yorubas, and a mixture of farmers and Local Government Area civil servants. Ethical consideration, the data collection process, the population of each village, vegetation types and locations of each local government area have been reported previously [14].
Observations and discussion
Socio-cultural aspect
Several urinary schistosomiasis studies had been conducted in Nigeria, most of which involved urine sample collection, so there is a high level of awareness about the importance of control programmes. However, in the process of field studies there is often confusion in the minds of the participants leading to fear of exposure to strangers which was found to prevail among the villagers. Frequently researchers are mistaken for government agents visiting for revenue collections. If the researcher can work with members of the community to change these opinions it would likely improve level of cooperation for inclusion in the study. We explored this aspect in Apojola, a community located on the heavily schistosome-infested Oyan dam reservoir. We made the first attempt to recruit participants through the community leader, followed by the religious leader, a nurse and a school teacher. The number of participants recruited through the assistance of the different leaders according to age and gender are shown in Table 1a. In Table 1b, it was shown that the community leader is the most effective in helping to mobilise the villagers of both genders for urine collection.
There is also an increasing cynicism about the disease among adult patients in endemic communities. Many members of the communities who admit passing blood in the urine do not perceive it as an indication of a serious disease. They consider it as a sign of virility and puberty which is a familiar sign among adults in other villages around them. A few others who have experienced some discomfort and thought it might be a major health problem were either ashamed of their disease status or ashamed of bringing their ‘red’ urine. Past studies have noted that individuals’ perceptions on the aetiology and impact of urinary schistosomiasis differed with their levels of education and gender [13]. Lack of knowledge about the cause and effect of the disease affects patient’s turnout for sample collection and this in turn has a direct influence on field data coverage and research quality.
Apart from lack of health education on the cause of the disease, the willingness to participate in the urine sample collection process is seemingly greater among patients with some level of education than among the uneducated. We investigated how patient’s level of education impacts turnout for urine sample collection in Ogbere community. Ogbere inhabitants are a mixture of uneducated farmers, who have nought to six years of formal education, and the educated comprising teachers and Local Government Area civil servants, who have from seven to 16 years of formal education. In Table 2, data from both groups are presented for comparison to show turnout according to education level and gender.
This Table shows the percentage contributions by the Community Leader (CL), Nurse (N), Teacher (T) and Religious Leader (RL) on the total number of respondents. CL is best for mobilising males in the community (P = 0.00155). CL is also best for mobilising male and female with calculated P = 0.052 just higher than 0.05. N is best for mobilising females but this is not statistically significant.
Ecological aspect
Transmission of urinary schistosomiasis is through freshwater snails, Bulinus species, as intermediate hosts and varies with different ecological factors. In many endemic communities, the ecological factors which favour disease transmission also promote agricultural practices such as farming, cattle rearing and fishing. Therefore, transmission to humans often occurs as a result of irrigation systems for agricultural purposes or when visits are made to the rivers for washing and swimming. As such, the rate of transmission to humans varies, to a great extent, with occupation.
However, since diagnosis is by urine testing, many peasant farmers and fishermen who are thought to be the most impacted with S. haematobium because of frequent water contact may remain undiagnosed and untreated. Urine sample collection for the diagnosis of urinary schistosomiasis is preferably done between the hours of 10:00 and 14:00 for optimum egg passage [9]. These hours coincide with the time during which farmers go to farm and fishermen set nets for fish catching. This coincidence might affect turnout for sample collection and estimation of overall disease prevalence in the community.
To evaluate the impact of patient’s occupation on turnout for urine sample collection, we compared turnout of farmers and civil servants in Ogbere community. For statistical purpose, farmers, cattle rearers and fishermen are classified as farming, while students, teachers and local government workers are classified as civil servants, see Table 2. In total, there are 84 participants out of which 33 are farmers (39.2%) and 51 are civil servants (60.7%). In all, more women (79.8%) turned out for sample collection.
According to the community leader, the total adults’ population in Ogbere is 3121 and the ratio of farmers to civil servants is approximately 20:1.
Z- Distribution test was used to compare the response level between the two groups using the formula:
(see picture number 4)
where p is the difference of proportions, N1 = 149 = Educated population and N2 = 2972 = Uneducated population. At all levels of significance 0.05, 0.01 and 0.001, response from the educated civil servant population was significantly higher than response from the uneducated farmer population.
Belief structures
Christians in Apojola and Ogbere communities were relatively unhindered by religious belief regarding their willingness to come forward for education about the project and provision of their urine samples. However there was gender problem with urine collection among the Muslim families at Apojola. The Muslim families at Apojola have the culture of restricting married women within the family household compounds and forbiding male visitors of adolescent age and older from entering the compounds or visiting the women. In order to be able to collect urine samples from these Muslim women, the local community nurse and a female member of our research team were accompanied by a local female Muslim field assistant and interpreter before being allowed access to the compounds to explain the importance of the disease and purpose of the study.
Conclusion
This study attempts to find out patients‘ attitudes to scientific research especially during a field sample collection process and suggests possible reasons for rejection of inclusion in scientific research by patients. In general, this study showed that social and ecological values including educational background, occupation, religious practices and poor knowledge about the aims and objectives of the study, strongly influence turnout for urine sample collection. Therefore, such values are worth considering for a holistic understanding of the scientific study results.
References
1. Edungbola LD, Asaolu SO, Omonisi MK, Aiyedun BA. Schistosoma haematobium infection among schoolchildren in the Babana district, Kwara State, Nigeria. Afr J Med Sci 1988; 7: 187-193.
2. Koukounari A, Gabrielli AF, Toure S, Bosque-Oliva E, Zhang Y, Sellin B, Donnelly CA, Fenwick A, Webster JP. Schistosoma haematobium infection and morbidity before and after large-scale administration of praziquantel in Burkina Faso. J Infect Dis 2007; 196: 659-669.
3. Webster JP, Koukounari A, Lamberton PH, Stothard JR, Fenwick A. Evaluation and application of potential schistosome-associated morbidity markers within large-scale mass chemotherapy programmes. Parasitology 2009; 136: 1789-1799.
4. Abdel-Wahab MF, Esmat G, Ramzy I, Fouad R, Abdel-Rahman M, Yosery A, Narooz S, Strickland GT. Schistosoma haematobium infection in Egyptian schoolchildren: demonstration of both hepatic and urinary tract morbidity by ultrasonography. Trans R Soc Trop Med Hyg 1992; 86: 406-409.
5. Fenwick A, Webster JP, Bosque-Oliva E, Blair L, Fleming FM, Zhang Y, Garba A, Stothard JR, Gabrielli AF, Clements AC, Kabatereine NB, Toure S, Dembele R, Nyandindi U, Mwansa J et al. The Schistosomiasis Control Initiative (SCI): rationale, development and implementation from 2002-2008. Parasitology 2009; 136: 1719-1730.
6. French MD, Rollinson D, Basanez MG, Mgeni AF, Khamis IS, Stothard JR. School-based control of urinary schistosomiasis on Zanzibar, Tanzania: monitoring micro-haematuria with reagent strips as a rapid urological assessment. J Pediatr Urol 2007; 3: 364-368.
7. Nduka FO, Ajaero CM, Nwoke BE. Urinary schistosomiasis among school children in an endemic community in south-eastern Nigeria. Appl Parasitol 1995; 36: 34-40.
8. Okoli EI, Odaibo AB. Urinary schistosomiasis among schoolchildren in Ibadan, an urban community in south-western Nigeria. Trop Med Int Health 1999; 4: 308-315.
9. Ibironke OA, Phillips AE, Garba A, Lamine SM, Shiff C. Diagnosis of Schistosoma haematobium by detection of specific DNA fragments from filtered urine samples. Am J Trop Med Hyg 2011; 84: 998-1001.
10. Koukounari A, Webster JP, Donnelly CA, Bray BC, Naples J, Bosompem K, Shiff C. Sensitivities and specificities of diagnostic tests and infection prevalence of Schistosoma haematobium estimated from data on adults in villages northwest of Accra, Ghana. Am J Trop Med Hyg 2009; 80: 435-441.
11. Mostafa MH, Sheweita SA, O’Connor PJ. Relationship between schistosomiasis and bladder cancer. Clin Microbiol Rev 1999; 12: 97-111.
12. Mungadi IA,.Malami SA. Urinary bladder cancer and schistosomiasis in North-Western Nigeria. West Afr J Med 2007; 26: 226-229.
13. Sarkinfada F, Oyebanji AA, Sadiq IA, Ilyasu Z. Urinary schistosomiasis in the Danjarima community in Kano, Nigeria. J Infect Dev Ctries 2009; 3: 452-457.
14. Ibironke O, Koukounari A, Asaolu S, Moustaki I, Shiff C. Validation of a new test for Schistosoma haematobium based on detection of Dra1 DNA fragments in urine: evaluation through latent class analysis. PLoS Negl Trop Dis 2012; 6: e1464.
The authors
Dr Olufunmiola Ibironke*
Cell and DNA Repository
Rutgers, The State University of New Jersey
New Brunswick
New Jersey, USA
e-mail: oai5@rutgers.edu
Dr Clive Shiff
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health
Baltimore, MD, USA
e-mail: cshiff@jhsph.edu
Dr Samuel Asaolu
Department of Zoology
Obafemi Awolowo University
Ile-Ife
Nigeria
*Corresponding author
Malaria: rapid and precise diagnosis saves lives
, /in Featured Articles /by 3wmediaMalaria is an acute and life threatening infection in individuls with no previous immunity. Symptoms are nonspecific and cannot be distinguished from those of influenza or severe bacterial infections. All febrile patients should thus be asked if they have been travelling over the past six months and if so whether the journey was to a malaria endemic area.
Microscopic examination of Giemsa stained thick and thin blood films remains the gold standard, but rapid tests using antigen-capture assays are increasingly used where access to expert microscopy is not available. The appropriate use of rapid tests and their limits are discussed.
by Dr Eskild Petersen
Malaria is caused by a protozoan parasite and five species can infect humans: Plasmodium falciparum, P. vivax, P. ovale, P. malariae and P. knowlesi. Humans are infected by bites of Anopheles mosquitoes and humans are the reservoir hosts except in the case of P. knowlesi, which is transmitted from monkeys and is only seen in South East Asia. Infection with P. falciparum shows the highest mortality and drug resistance is much more common in P. falciparum compared to P. vivax, and not a problem in the other malaria species. P. ovale and P. vivax have persistent liver forms, hypnozoites, which may reactivate, usually within six months after infection, and give rise to a malaria attack.
Malaria in Europe
Malaria was endemic in Europe up to the middle of the 20th century [Table 1]. Presently malaria is almost exclusively imported, although a number of Plasmodium vivax cases were seen in Greece in 2011, probably introduced with migrant workers from endemic areas [1]. It is estimated that between 10,000 and 15,000 cases of malaria are imported into Europe every year, which makes the recognition of symptoms and knowledge of appropriate diagnosis important.
A special risk group is immigrants resident in Europe who visit their home countries where malaria is found. The proportion of imported malaria cases in immigrants in Europe has increased from a reported 14% more than 10 years ago to 86% in more recent studies [2]. More than five million African immigrants could be living in Europe, one third of whom are from Sub-Saharan Africa [3], and children of immigrants are particularly at risk [4].
Mortality of imported malaria
The mortality from imported Plasmodium falciparum malaria cases varies from 0.4% in a large cohort from France [5], up to 5% in a recent cluster of cases imported from The Gambia [6]. Malaria infection in non-immunes is an emergency which requires prompt diagnosis and treatment while asymptomatic malaria in immigrants raises other public health issues.
Clinical symptoms
Individuals without immunity, i.e. persons who have not lived in malaria endemic countries for a long time, normally have a febrile illness with an acute onset. The symptoms include fever, malaise, muscle and joint pains, headache and rarely respiratory distress and diarrhoea. Malaria infection can be complicated by bacterial septicaemia. As the infection progresses there can be drowsiness, coma, kidney failure, disseminated intravascular coagulation and low blood pressure, and in the non-immune the mortality of untreated P. falciparum malaria is probably more than 50%.
P. falciparum in non-immunes does not usually follow a regular cyclic pattern and the fact that fever is not cyclic with a 48 or 72 hour cycle cannot be used to exclude malaria. Malaria in non-immunes is a medical emergency and diagnosis should be performed without delay.
In semi-immunes the clinical symptoms may be much more discrete and the development more subtle. Immunity to malaria is not a sterile immunity and a low level parasitaemia is seen in semi-immune individuals, ie. individuals from malaria endemic areas [7]. A special risk group is pregnant women from malaria endemic areas who are at greater risk of clinical malaria during pregnancy [8].
Malaria parasites may persist in asymptomatic immigrants long after their arrival in the host country, and malaria can be transmitted, for instance by blood transfusion or organ transplantation.
Who should be tested for malaria?
Diagnostic tests for malaria infection should be performed in any febrile patients who have a history of exposure, which includes patients with a history of travel in malaria endemic areas, as defined by the WHO.
However, rare modes of transmission mean that patients with fever but without a travel history to endemic areas should be tested. This includes so called ‘airport malaria’ where Anopheles mosquitoes carrying malaria parasites are transported in an airplane, leave the destination and take a blood meal from someone living close to the airport [9,10]. Malaria parasites can be transmitted in blood when sharing instruments used for intravenous drug abuse [11]. Transmission of malaria by blood transfusions from asymptomatic carriers is a huge problem in tropical Africa [12] and febrile patients with a history of receiving blood transfusion from a donor in a malaria endemic area should be suspected of having malaria until it is proven otherwise.
Diagnostic procedures for detecting malaria parasites
Traditionally malaria diagnosis rests on the microscopic examination of thick and thin blood films, but over the past decades, rapid tests based on antigen capture are increasingly used. However, rapid test have pitfalls and parasite density must be measured and followed to monitor the response to treatment. Thus microscopy is still a mandatory skill in institutions taking care of malaria patients.
Microscopic examination of Giemsa stained thick blood films remains the gold standard because it is rapid, easy to perform and sensitive [13] with a sensitivity down to five parasites per microlitre of blood [14]. Microscopy and counting of malaria parasites in patients are mandatory to assess the response to treatment and must be available at centres managing patients with malaria.
Rapid test are available which show a 100% sensitivity down to a parasite density level of 200 parasites per microlitre, equivalent to a parasitaemia of approximately 0.004% [15]. Molecular diagnosis by polymerase chain reaction (PCR) can detect parasites down to a density of 0.01 parasites per microlitre after a lysis procedure, and 1 parasite per microlitre without lysis [16]. However, PCR analysis is not instantly available around the clock so in practice diagnosis relies on rapid diagnostic tests and microscopy of Giemsa stained thick blood films.
Rapid tests are increasingly used in medical centres with limited access to experienced microscopists. However, a rapid test cannot determine the parasite density and rapid tests have limitations. False negative results in patients with very high parasite densities have been described, probably due to the so called ‘pro-zone’ phenomena known from other diagnostic tests [17, 18]. The problem seems to be limited to tests based on detection of the Histidine Rich protein 2, HRP2, and not tests based of detection of Plasmodia LDH, Lactate Dehydrogenase [15, 17]. Mutations in the HRP2 gene may also result is false negative results [19, 20]. All species ie. P. falciparum, vivax, ovale and malariae and P. knowlesi, will be found with tests based on the detection of pan-malarial aldolase antigen aldolase and LDH antigens [21]. P. ovale can be divided in variant and classic P. ovale [22], and variant P. ovale is not picked up in HRP2 based rapid diagnostic tests [23].
Thus clinicians using rapid tests should be instructed that no test so far is 100% reliable. In order to reduce the risk of false negative results, testing should be performed at least twice with 24 hours in between and preferable three times within a 24 hours interval. Variant P. ovale and P. knowlesi infections will be detected by rapid tests, which include those incorporating the pan plasmodia antigens Aldolase or Lactate Dehydrogenase antigens [15, 24]. The latest results of the WHO multicentre evaluation of different rapid diagnostic tests showed that the best performance was found in tests based on a combination of the HRP2 and PLDH proteins [15].
References
1. Danis K et al. Euro Surveill 2011;16:19993.
2. Jelinek T et al. Clin Infect Dis 2002; 34:572-576.
3. Eurostat. European Commission. Katya Vasileva. Population and social conditions. 34/2011. Available at: http://epp.eurostat.ec.europa.eu/cache/ITY_OFFPUB/KS-SF-11-034/EN/KS-SF-11-034-EN.PDF
4. Stäger K et al. Emerg Infect Dis 2009; 15:185–91.
5. Bruneel F et al. PLoS One 2010; 5(10):e13236.
6. Jelinek T et al. Euro Surveill.- 2008;13:19077.
7. Wertheimer ER et al. Emerg Infect Dis 2011;17:1701-3.
8. D’Ortenzio E et al. Emerg Infect Dis 2008;14:323-6.
9. Thang HD et al. Neth J Med 2002;60:441-3.
10. Tatem AJ et al. Malar J 2006;5:57.
11. Chau TT et al. Clin Infect Dis 2002;34:1317-22.
12. Noubouossie D et al. Transfus Med 2012;22:63-7
13. Bowers KM et al. Malar J 2009;8:267.
14. Petersen E et al. Am J Trop Med Hyg 1996; 55:485-489.
15. WHO. Rapid Diagnostic Tests. Results of round 3.http://www.who.int/tdr/publications/tdr-research-publications/rdt_round3/en/index.html Geneva 2011 (Accessed 17th March 2012).
16. Mahajan B et al. Transfusion 2012 Feb 10. doi: 10.1111/j.1537-2995.2011.03541.x. [Epub ahead of print].
17. Luchavez J et al. Malar J 2011;10:286.
18. Gillet P et al. Malar J 2011;10:166.
19. Koita OA et al. Am J Trop Med Hyg 2012;86:194-8.
20. Baker J et al. PLoS One 2011;6:e22593.
21. Chiodini PL et al. Trans R Soc Trop Med Hyg 2007; 101:331-337.
22. Sutherland CJ et al. J Infect Dis 2010;201:1544-1550.
23. Tordrup D et al. Malar J 2011;10:15.
24. Hellemond JJ van et al. Emerg Infect Dis 2009;15:1478–1480.
25. Bruce-Chwatt LJ, Zulueta J de. The rise and fall of malaria in Europe. Oxford University Press. 1980.
The author
Dr Eskild Petersen
Department of Infectious Diseases
Aarhus University Hospital
Skejby
Aarhus
Denmark
Tel +45 7845 2817
e-mail: joepeter@rm.dk
Malaria: a global threat
, /in Featured Articles /by 3wmediaMalaria threatens the existence of large numbers of children in tropical and subtropical areas of the world. Increasing malaria parasite drug-unresponsiveness and insecticides-unresponsive mosquitoes lead to emergence of new malaria foci. Insecticide-impregnated bed nets and case detection/prompt treatment with artesunate-based drug combinations offer the most effective control measures. Counterfeit antimalarial drugs pose a serious threat to malaria control. No effective vaccine has been introduced into clinical practice to date.
by Prof. E.A.G Khalil and Dr M.E.E. Elfaki
Malaria is a febrile parasitic disease that is transmitted by female mosquitoes with no known intermediate host except in the case of Plasmodium knowlesi. Malaria is prevalent over most areas of Asia, Africa, eastern Europe, south America and South Pacific. Hot climate and low socio economic conditions make malaria prevalent in these areas. Malaria affects 300 to 700 million people annually with 1-2 million deaths, mostly of children [1]. The malaria parasite can infect all age groups, but children and pregnant women are at an increased risk for developing the severe form of the disease. The red blood cells are the principal cells affected, the parasite usually affects red blood cells of all ages. There are five species of malaria parasite that cause human disease: Plasmodium falciparum, Plasmdium vivax, Plasmodium ovale, Plasmodium malariae and Plasmodium knowlesi.
Malaria can present in a mild uncomplicated form that is characterised by fever, headache, arthralgia, vomiting, malaise, sweating and splenomegaly. On the other hand a severe and complicated form exists that presents as severe anaemia, pulmonary oedema, seizures, coma and renal/respiratory effects. Brain effects can result in the death of 20% of even optimally treated individuals with residuals brain damage in some surviving children. Large numbers of individuals in endemic areas may harbour the parasite without obvious symptoms (subclinical infection); these individuals represent a reservoir during the dry season [2,3].
Under-nutrition is an underlying cause of malaria morbidity in children under the age of five [4]. Nutritional supplementation with vitamin A, zinc, selenium, iron and folate are reported to reduce malaria morbidity in children, probably through their effects on the immune system [5].
Immunity against malaria
The ability of humans to fight malaria relies on the presence of specialised immune cells that produce antibodies against malaria parasite proteins expressed on the surface of infected red blood cells (humoral immunity). Human immunity also relies on the production of cytokines, specialised proteins that arm immune cells and make them more capable of killing malaria parasites. In addition, specialised T-lymphocytes, namely CD8+ cells help the body to eliminate the parasite through destruction of infected cells (cellular immunity) [6,7].
Treatment of simple and complicated malaria
Chloroquine was the drug of choice for malaria treatment for some time, but this has dramatically changed due to the emergence of resistance in different parts of the globe. The same problem has occurred with other antimalarial drugs such as mefloquine, quinine and sulphadoxine [7,8]. Artesunate-based combinations are now used as first line treatment of simple malaria by many control programmes [10]. In addition, fixed-combination anti-malarials such as Dihydroartemsinin-piperaquine (DP) can effectively treat uncomplicated, multidrug-resistant falciparum malaria [11].
Intermittent preventive malaria treatment (IPT) using sulphadoxine/pyrimethamine has been shown to reduce the burden of malaria effectively in children in areas of seasonal transmission [12]. Supportive treatment is an important adjunct to antimalarial treatment (antipyretic, anticonvulsant and exchange blood transfusion) in severe P. falciparum malaria [13,14].
Control of malaria
Malaria morbidity and mortality can be markedly reduced with a sum of money not exceeding $ 3.0/adult. At the present time case detection and prompt treatment with artesunate-based combination drugs and the use of insecticide-treated bed nets (ITN) are the most effective control measures. ITN have proven to reduce malaria morbidity and mortality [10,15,16].
Counterfeit drugs
Counterfeit drugs present a major obstacle to malaria control programmes by prolonging morbidity and increasing mortality. About a third to one half of drugs sold in Africa and Asia are counterfeit drugs. There is some evidence that the problem of counterfeit drugs is increasing, especially in countries where regulatory authorities do not have the will to investigate and take action or do not have the necessary resources. However there is a lot of pressure not to publicise the issue of counterfeit anti-malaria drugs [17,18,19,20].
Vaccines against malaria
The ability of the malaria parasite evade the immune system is the main reason that no really effective vaccine has been produced to date. A number of the parasite molecules have been targeted as vaccine candidates in vain. Recently, the RTS,S/AS01 vaccine has been shown to provide protection against clinical and severe malaria in African children [21,22].
Conclusion
Better use of ITN, rapid and accurate diagnostic tests and the use of artesunate-based drug combinations can effectively control malaria. Counterfeit anti-malarials are a serious and under-estimated problem that could definitely cripple malaria control programmes in Africa and Asia.
References
1. WHO 2005. World Malaria Report.
2. Looareesuwan S et al. Lancet 1985; 2: 4-8
3. Reuben R. Soc Sci Med 1993; 37: 473–480.
4. Caulfield LE et al. Am J Trop Med Hyg 2004; 71 suppl 55-63.
5. Shankar AH. J Infect Dis 2000 182 (Supplement 1): S37-S53. doi: 10.1086/315906.
6. Goodhttp MF & Doolan DL. Curr Opin Immunol 1999; 11, 4, 412–419.
7. Stevenson M & Riley EM. Nature Rev Immunol 2004; 4, 169-180.
8. al-Yaman F et al. P N G Med J 1996; 39 :16-22.
9. Le Bras J & Durand R. Fundam Clin Pharmacol 2003; 17 :147-53.
10. WHO/MAL/94.1067. The role of artemisinin and its derivatives in the current treatment of malaria (1994-1995): report of an informal consultation convened by WHO in Geneva, 27-29 September 1993. Geneva: WHO 1994.
11. Ashley EA et al.. Clin Infect Dis 2005; 41 : 425-432. doi: 10.1086/432011.
12. Dicko A et al. Mal J 2008; 7:123 doi:10. 1186/ 1475-2875-7-123.
13. World Health Organization, Division of Control of Tropical Diseases. Severe and complicated malaria. Trans R Soc Trop Med Hyg 1990; 84: Suppl 2:1-65.
14. Hien TT et al. Trans R Soc Trop Med Hyg 1992; 86:582-583
15. Guerin PJ et al. Lancet Infect Dis 2002; 2 :564-573.
16. Frey C et al. Mal J 2006; 5:70.
17. World Health Organization. Report of the International Workshop on Counterfeit Drugs. 1998; WHO/DRS/CFD/98.1. Geneva: WHO.
18. Newton PN et al. BMJ 2002; 324: 800–801.
19. Dondorp AM et al. Trop Med Int Health 2004; 9: 1241–1246.
20. Rudolf PMM & Bernstein IBG. N Engl J Med 2004; 350: 1384–1386.
21. Plassmeyer ML et al. J Biol Chem 2009; 284 : 26951–63.
22. Agnandji ST et al. N Engl J Med 2011; 365: 1863-1875.
The authors
Prof. E.A.G. Khalil and Dr M.E.E. Elfaki
Department of Clinical Pathology
& Immunology
Institute of Endemic Diseases
University of Khartoum
Khartoum
Sudan
Rapid diagnostic tests for malaria
, /in Featured Articles /by 3wmediaTogether with HIV/AIDS and TB, malaria is one of the major public health challenges of the developing world. Prompt diagnosis is a priority. Rapid diagnostic tests are readily available, quick to yield results and can be effectively used in resource-limited settings.
by Meghna Patel
Malaria is a tropical disease caused by parasites of the genus Plasmodium and transmitted by Anopheles mosquitoes. Being endemic in more than 100 countries, half the world’s population is at risk for malaria. Children are at particular risk, accounting for most malaria deaths globally [1]. Each year roughly 250 million people are infected and nearly a million people die from the disease [2]. Malaria causes significant morbidity and mortality, particularly in resource-poor regions. Sub-Saharan Africa is the hardest hit region in the world and parts of Asia and Latin America also face significant malaria epidemics [3]. Four major species of malarial parasite infect humans: Plasmodium falciparum, P. vivax, P. ovale and P. malariae. The first two species cause the most infections worldwide. On the continent of Africa, P. falciparum malaria predominates, whereas in parts of Asia and Latin America, P. vivax is more prevalent. Two other species, P. ovale and P. malariae, are also capable of causing human disease. A fifth species, Plasmodium knowlesi, is found in Southeast Asia; it mainly causes malaria in simians but it can also infect humans.
Since malaria is preventable and treatable, such high incidences point to inappropriate management of the condition in some cases, with incorrect or inefficient diagnosis and/or treatment. Rapid and accurate diagnosis of malaria before treatment is essential for effective and timely treatment of patients and to minimise the spread of drug resistance and thus the requirement of more expensive drugs, frequently unaffordable for resource-poor countries [4]. This review discusses the currently available techniques for malaria diagnosis
focusing on rapid diagnostic tests (RDT).
Diagnosis
As in other pathological conditions malarial diagnosis is based on clinical investigations and pathological laboratory analysis. Diagnosis based on clinical symptoms is the least expensive, most commonly used method in resource poor conditions. However, the overlapping of malaria symptoms with other tropical diseases impairs its specificity and therefore encourages the indiscriminate use of anti-malarials for managing febrile conditions in endemic areas.
Laboratory diagnosis of malaria includes identifying malarial parasites or their antigens/products in patient blood. Although this may seem simple, diagnostic efficacy depends on various factors such as stage and forms of the various malarial species, endemicity of different species, density of parasitaemia etc.
In the laboratory, malaria is diagnosed using different techniques e.g. conventional microscopic diagnosis by examining stained thin and thick peripheral blood smears, other concentration techniques, e.g. quantitative buffy coat (QBC), rapid diagnostic tests and molecular diagnostic methods, such as PCR. The pros and cons of these methods have also been described, chiefly related to sensitivity, specificity, accuracy, precision, time consumed, cost-effectiveness, labour intensiveness, the need for skilled microscopists etc.
Malaria is conventionally diagnosed by microscopic examination of stained blood films using Giemsa, Wright’s or Field’s stains [5]. Even though microscopic examination is considered to be the gold standard method, the most important limitation is its relatively low sensitivity, thus the generation of false negative results, particulary when microscopy is carried out using a low quality microscope and/or by less experienced personnel, and with low parasitaemias as in asymptomatic malaria. Furthermore the technique is laborious and not really suitable for remote rural settings, with no electricity or health facility resources.
The QBC technique was designed to enhance microscopic detection of malaria parasites [6]. This technique utilises micro-haematocrit tubes, fluorescent dyes and an appropriate fluorescence microscope for detection. Although simple, reliable and user-friendly, QBC is not widely applicable as it is costly, requires specialised instrumentation and is far from ideal for determining species and numbers of parasites.
Serological methods to diagnose malaria usually target antibodies against asexual blood stage malarial parasites. Immunofluorescence antibody testing (IFA) has proved a reliable serological test for malaria [7]. Although IFA is sensitive and specific, it is time-consuming and subjective. Furthermore the reliability greatly depends on the use of standardised reagents, in turn dependent on the expertise of laboratory workers.
Recent developments in malaria diagnosis suggest the use of PCR-based techniques. These techniques have proven to be one of the most specific and sensitive diagnostic methods, especially in malaria cases with low parasitaemia or mixed infections [8]. PCR was found to be more sensitive than QBC and some RDTs [9,10]. Compared with the gold standard method for malaria diagnosis, PCR has exhibited higher sensitivity and specificity [8]. Moreover, PCR can also help detect drug-resistant parasites, and is compatible with automation so that large numbers of samples can be processed. Some modified PCR methods e.g., nested PCR, real-time PCR and reverse transcription PCR are reliable and appear to be useful second-line techniques. Although PCR appears to offer the paramount sensitivity and specificity, its adoption in labs is limited due to the complex methodology, high cost and the demand for specialised instruments, the complex quality control and the difficulty of recruiting trained technicians especially in resource-poor conditions.
As the majority of malaria cases are found in countries where cost-effectiveness is an especially important factor and the ease of diagnostic test performance and training of personnel are also major considerations, new technology has given due attention to these points and utilised techniques that comply with diagnostic need without being very demanding. This has mainly resulted in the
development of RDTs.
Rapid diagnostic tests
RDT are largely based on the principle of immunochromatograpy, in which either monoclonal or polyclonal antibodies against the parasite antigen are immobilised to capture the parasite antigens from the peripheral blood. Currently, immunochromatographic tests target the histidine-rich protein-II of P. falciparum, a pan-malarial Plasmodium aldolase and the parasite-specific
lactate dehydrogenase.
Histidine-rich protein II of P. falciparum (PfHRP-II) is a water soluble protein that is produced by the asexual stages and young gametocytes of P. falciparum. It is abundantly expressed on the red cell membrane surface [11].
Parasite lactate dehydrogenase (pLDH) is a soluble glycolytic enzyme produced by the asexual and sexual stages of the live malarial parasites [9]. It is present in and released from the parasite-infected erythrocytes. It has been found in all four major species causing malaria in humans as their respective isoforms.
Plasmodium aldolase is an enzyme of the glycolytic pathway expressed by sexual and asexual stages of malaria parasites. RDTs have been developed in different test formats such as dipstick, card, well and cassette. The test procedure varies between different test kits. In general, the blood sample is mixed with a buffer solution that contains a haemolysing compound and a specific antibody that is labelled with a visually detectable marker such as colloidal gold. If the target antigen is present in the blood, a labelled antigen-antibody complex is formed and it migrates forward in the test strip and is captured at the test line. It is essential to include a control line to check on test validity. A washing buffer is then added to clear the background for easy
visualisation of the coloured lines.
RDTs are available in kit form with all the necessary reagents so they can be utilised even in remote places by less skilled personnel to generate results within a short period of time, usually within 15-20 minutes.
WHO recommended a few desirable characteristics for RDTs regarding their accuracy and sensitivity (WHO/MAL/2000.1091). According to this RDTs should be at least as accurate as results derived from microscopy performed by an average technician under routine field conditions, the sensitivity should be above 95% compared to microscopy, and the detection of parasitaemia should be such that levels of 100 parasites /µL (0.002% parasitaemia) should be detected reliably with a sensitivity of 100%. One product received U.S. FDA clearance in June 2007.
Today most RDTs have achieved this goal for P. falciparum, but not for other species. Roughly, RDT sensitivity declines at parasite densities < 500/µL blood for P. falciparum and < 5,000/µL blood for P. vivax [12]. RDT consumption, especially in developing countries, has increased over the past few years.
SPAN diagnostics offers RDTs i.e. ParaHIT-Total and ParaHIT-f in both dip stick, as well as in device format, for rapid and reliable diagnosis of malaria. ParaHIT-f is intended to diagnose malaria caused by P. falciparum with the use of P. falciparum specific HRP-II, wheareas ParaHIT-Total explores HRP-II and pan malarial species specific aldolase, as separate lines to screen malaria and for
differential determination of P. falciparum.
References
1. WHO, World Malaria Report 2010; December 2010.
2. WHO 10 facts on malaria
3. CDC, Malaria
4. Barnish G et al. Newer drug combinations for malaria. BMJ 2004; 328: 1511–1512
5. Warhurst DC et al. Laboratory diagnosis of malaria. J Clin Pathol 1996; 49: 533-538
6. Clendennen TE 3rd et al. QBC and Giemsa stained thick blood films: diagnostic performance of laboratory technologists. Trans R Soc Trop Med Hyg 1995; 89: 183-184
7. She RC et al. Comparison of immune fluorescence antibody testing and two enzyme immunoassays in the serologic diagnosis of malaria. J Travel Med 2007; 14: 105-111
8. Morassin B et al. One year’s experience with the polymerase chain reaction as a routine method for the diagnosis of imported malaria. Am J Trop Med Hyg 2002; 66: 503- 508
9. Makler MT et al. A review of practical techniques for the diagnosis of malaria. Ann Trop Med Parasitol 1998; 92: 419-433
10. Rakotonirina H et al. Accuracy and reliability of malaria diagnostic techniques for guiding febrile outpatient treatment in malaria-endemic countries. Am J Trop Med Hyg 2008; 78: 217-221
11. Rock EP et al. Comparative analysis of the Plasmodium falciparum histidine-rich proteins HRP1, HRP2 and HRP3 in malaria diagnosis of diverse origin. Parasitology 1987; 95: 209–227.
12. Wongsrichanalai C et al. A Review of Malaria Diagnostic Tools: Microscopy and Rapid Diagnostic Test (RDT). Am J Trop Med Hyg 2007; 77: 119–12.
The author
Meghna Patel
SPAN Diagnostics Ltd
Udhna, Surat, India
HCC biomarker, a proteomics approach: the journey from bench to bedside
, /in Featured Articles /by 3wmediaA goal of clinical proteomics is to find a disease indicator (biomarker) to identify the presence of, or monitor, a disease. It may be surprising that approximately one-third of all cancer cases could be effectively treated if detected at an early enough stage. As a heterogeneous disease, cancer evolves via multiple pathways and is a culmination of a variety of genetic, molecular and clinical events. Given that there is significant variation in the risk of developing cancer and that early detection often results in increased survival, developing technologies capable of identifying patients at highest risk and detecting tumours in the earliest stages of development is a pressing need.
by Dr Gul M. Mustafa, Prof. Cornelis Elferink and Prof. John R. Petersen
Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer, ranking sixth among cancers in incidence worldwide and is the 3rd leading cause of cancer death. Despite some significant improvements in diagnosis and treatment of human liver diseases over the last decade, the HCC mortality rate has not changed to any extent. Currently there are approximately 20,000 new case in the US annually with millions world-wide [1]. The projected rise in the new HCC cases in the US and the world is mainly due to latent hepatitis C virus (HCV) infections in the general population, accounting for approximately 80% of HCC cases several decades after initial infection. The less than 5% survival rate of patients with HCC is primarily due to the disease eluding early detection and diagnosis, when options for effective treatment still remain. Surveillance of patients at highest risk for developing HCC, notably patients with cirrhosis, would benefit greatly from a biomarker assay capable of accurately detecting HCC in its earliest stages when it is still possible to intervene. One of the most widely used markers for HCC is alpha fetoprotein (AFP) although it is non-specific, providing low sensitivity and poor specificity, especially for early detection of HCC [2]. The false-negative rate with AFP level can be 40% for tumours < 3 cm in diameter. More reliable methods such as triple phase Computed Tomography (CT) imaging and liver biopsies exist, but these are expensive and not conducive to long-term surveillance. Therefore, the identification of superior biomarkers will be of huge clinical significance to
at-risk populations.
The ideal biomarker for this type of application would be one where HCC is detected with a high sensitivity and specificity in easily obtained biological samples in a non-invasive, or minimally invasive, manner. Blood represents the best source for detection of HCC related biomarkers, as every cell in the body leaves a record of its physiological state by the products it sheds into the blood, either as a waste or as a signal to neighbouring cells. What some may view as cellular refuse in is reality a diagnostic gold mine. Because of its easy accessibility from patients on a regular basis and because it is in contact with all the tissues in the body, it is an excellent choice for a proteomics approach as it may reveal when changes, such as development of HCC, occur. The systematic analysis of the whole serum or plasma proteome may thus provide a functional meaning to the information provided by genome expression studies. Expression of proteins, their isoforms or post-translational modifications, can be detected by proteomic analysis and these data can provide precious information to better understand the pathologic/molecular basis of HCC [3]. Proteomic analysis may also allow monitoring of the course of the disease process from cirrhosis to HCC, eventually leading to earlier diagnosis which is essential in determining the best course of treatment options and possible outcomes. In addition to earlier diagnosis proteomic analysis may also be useful in measuring the efficacy/progress of treatment or detecting tumour reoccurrence both of which are missing in HCC treatment.
Proteomics analysis
Proteomics analysis is currently considered to be the best tool for the global evaluation of protein expression, and has been widely applied in the analysis of diseases, especially cancer research. For us the approach was to compare the serum/plasma protein profile from patients infected with HCV against the sera from patients with confirmed HCC. Proteins found to be consistently altered between the two patient populations can then be identified and further characterised to determine if they can be used as biomarkers of HCC. While on the surface this sounds simple, due the complexity of the proteome and the wide dynamic concentration range (9 orders of magnitude from pg/mL to mg/mL) of constituent protein/peptide species it is an extremely challenging task. Because the serum/plasma proteome is predominated by high abundance proteins such as albumin and immunoglobulins, extensive fractionation prior to analysis is required. To reduce the few over-represented (i.e. abundant) proteins, without losing any valuable information, existing fractionation methodologies often discard the high abundance carrier proteins, such as albumin, and thus fail to capture the information associated with this valuable resource. We have used aptamer-based technology (Bio-Rad) a technology that reduces the dynamic range and thus retains the complexity of the serum peptidome, in contrast to strategies that just deplete carrier proteins.
Quantitative protein expression profiling
Because proteins entering the blood from surrounding tissue are much less abundant, it is this fraction that is likely to contain most of the undiscovered biomarkers. Quantitative protein expression profiling is a crucial part of proteomics, and such profiling requires methods that are able to efficiently provide accurate and reproducible differential expression values for proteins in two or more biological samples. Thousands of different protein species present in the biological fluid or tissue must be separated, identified and characterised, which cannot be accomplished by a single experimental approach. An effective approach is two-dimensional differential in gel electrophoresis (2D-DIGE) and mass spectrometry [4]. While two-dimensional electrophoresis (2DE) has been widely used for proteomics research, the inter-gel variation along with excessive time/labour costs are major problems. Two-dimensional differential in gel electrophoresis (2D-DIGE) is a modification of 2DE and is considered as one of the most significant advances in quantitative proteomics. Using 2D-DIGE, two samples that are to be compared are pre-labelled with mass- and charge-matched fluorescent cyanine dyes and co-separated on the same 2D gel. The use of internal standards in every gel minimises problems associated with technical variability. Moreover with the great sensitivity and dynamic range that is afforded by the fluorescent dyes, 2D-DIGE can give greater accuracy of quantitation than traditional silver staining. The data captured from these gels using the Imagers, such as the Typhoon trio, along with and proprietary (Decyder) software can be configured to give inter-gel and intra-gel statistical analysis providing both a quantitative and qualitative analysis. We and others are using this approach to identify differentially expressed proteins for differential expression between the pre-cancerous and cancerous patient groups.
Stable isotope labelling
Another technique that can be useful in the analysis of the whole serum proteome is stable isotope labeling using O16/O18. This is a quantitative proteomic technique that distinguishes individual peptides during LC-MS/MS on the basis of a 4 Dalton m/z change after differential O16/O18 labelling that takes place at the C-terminal carboxyl group of tryptic fragments [5]. It is then possible to determine the ratio of individual protein expression levels between the two samples. Alternatively it is possible to use O16/O18 stable isotope labeling to determine the differential expression between two patient groups. In this way the low molecular weight serum peptidome (<20kDa), suspected of harbouring metabolites and degradation products reflecting HCC, can also be interrogated Selected reaction monitoring
Selected reaction monitoring (SRM), which is used to monitor a precursor and its product ion m/z, is another powerful proteomic tool using tandem mass spectrometry to monitor target peptides within a complex protein digest. The specificity and sensitivity of the approach, as well as its capability to multiplex the measurement of many analytes in parallel, renders it amenable to biomarker discovery and validation proteomics. Using the selectivity of multiple stages of mass selection of tandem mass spectrometers, these targeted SRM assays are the mass spectrometry equivalent of a Western blot. An advantage of using a targeted mass spectrometry-based assay over a traditional Western blot is that it does not rely on the creation of highly selective immunoaffinity reagents. Thus, targeted SRM assays using heavy isotope-labelled internal standards can be multiplexed in quantitative assays that can be directly applicable to clinical settings. A targeted proteomics workflow based on SRM on a triple Quadrupole mass spectrometry platform shows the potential of fast verification of biomarker candidates reducing the gap between discovery and validation in the biomarker pipeline. Although useful, due diligence needs to be exercised in developing and validating SRM assays.
Sample handling
Biomarker research necessitates a clear, rational framework. Technologically, the platform needs to be able to detect low abundant plasma/serum proteins and reproducibly measure them in a high throughput manner. Conceptually, the choice of the technological platform and availability of quality samples should be part of an overall study design that integrates basic and clinical research. Sample preparation is an important and very critical part of clinical proteomics as the collection, sample handling and storage can have a significant impact on the integrity of the proteins being detected. It is so important that a standard operating procedure outlining the steps that should be followed in collecting and storing clinical samples was recently published [6]. In addition to a standardised collection procedure, biological samples need to be carefully chosen based on well-established guidelines either for candidate discovery in the form of controls and the disease being detected or for validation of the candidate biomarkers using well characterised samples.
Most importantly, the samples should be representative of the target population and directly address the clinical question. A conceptual structure of a biomarker study can be provided in the form of sequential phases, each having clear objectives and predefined goals [Figure 1]. Furthermore, guidelines for reporting the outcome of biomarker studies are critical to adequately assess the quality of the research, interpretation and generalisation of the results. By being attentive to and applying these considerations, biomarker research should become more efficient and lead to biomarkers that are translatable into the clinical arena.
Aknowledgements
This research was supported by a pilot grant from the Clinical Translational Sciences Award (5UL1RR029876) and the Mary Gibb Jones endowment.
References
1. Kim WR. The burden of hepatitis C in the United States. Hepatology 2002; 36: 30-34.
2. Sterling RK, Wright EC, Morgan TR, Seeff LB, Hoefs JC, Di Bisceglie AM, Dienstag JL, Lok AS. Frequency of elevated hepatocellular carcinoma (HCC) biomarkers in patients with advanced hepatitis C. Am J Gastroenterol 2012; 107(1): 64-74.
3. Maria P, Laura ML, Antonio RA, Jose LM, Javier B, Ruben C, Jordi M and Manuel de la Mata. Proteomic analysis for developing new biomarkers of hepatocellular carcinoma. World J Hepatol 2010; 2(3): 127-135.
4. Sun W, Xing B, Sun Y, Du X, Lu M, Hao C, Lu Z, Mi W, Wu S, Wei H, Gao X, Zhu Y, Jiang Y, Qian X, He F. Proteome analysis of hepatocellular carcinoma by two-dimensional difference gel electrophoresis: novel protein markers in hepatocellular carcinoma tissues. Mol Cell Proteomics 2007; 6(10): 1798-808.
5. Miyagi M, Rao KC. Proteolytic 18O-labeling strategies for quantitative proteomics. Mass Spectrom Rev 2007; 26(1):121-36.
6. Tuck MK et al. Standard operating procedures for serum and plasma collection: early detection research network consensus statement standard operating procedure integration working group. J Proteome Res 2009; 1: 113-117.
The authors
Gul M. Mustafa, Ph.D. Postdoctoral Fellow, Department of Pharmacology
Cornelis Elferink, Ph.D., Professor, Department of Pharmacology, Director Sealy Center Environmental Health and Medicine
John R. Petersen, Ph.D., Professor and Director Victory Lakes Clinical Laboratory, Department of Pathology,
University of Texas Medical Branch
301 University Boulevard
Galveston, Texas 77555, USA
e-mail: jrpeters@utmb.edu
Determining the likelihood of malignancy in women with pelvic masses
, /in Featured Articles /by 3wmediaThe ROMA (Risk of Ovarian Malignacy) algorithm uses the CA-125 and HE4 blood markers to determine the likelihood that a pelvic mass is malignant. The test has been shown to aid gynaecologists in referring women to gynaecologic oncologists for surgery.
by Dr Zivjena Vucetic
Ovarian cancer is the leading cause of death from gynaecologic malignancies in the United States with annual incidence of 22,000 cases. Estimated annual mortality rate is approximately 15,460 cases [1]. Ovarian cancer has a good prognosis if detected in its early stages and if treated by specialised gynaecologic oncology surgeons [2], however more than three-quarters of cases are diagnosed in the advanced stage and are associated with poor survival rates of 10-30% [3]. These poor outcomes reflect the lack of effective tools for early detection of ovarian cancer and the limitations of current treatment options for ovarian cancer, which generally include cytoreductive surgery followed by adjuvant chemotherapy.
Recent studies have shown that women with ovarian cancer develop non-specific symptoms, including pelvic or abdominal pain, increased abdominal size, bloating, urinary urgency and difficulty eating or feeling full quickly, months before diagnosis [4]. However, ovarian cancer is commonly discovered on surgery for an adnexal mass. It is estimated that 5–10% of women at some point in their lives will undergo surgical evaluation of an adnexal mass and up to one fifth of surgically removed masses will have a diagnosis of ovarian cancer [5]. In premenopausal women, the risk of a mass being malignant is 7-13%, while in the postmenopausal women is 30-40% [6]. Thus, the presence of symptoms and the findings of an adnexal mass increase the risk of malignancy and should prompt thorough diagnostic evaluation.
The primary goals of diagnostic evaluation of women who present with adnexal masses are to confirm that adnexal mass is of ovarian origin and to differentiate whether it is benign or malignant. In order to determine the most appropriate management strategy that would ensure the optimal outcome for the woman with adnexal mass it is essential to effectively triage the risk for malignancy. Combination of multiple diagnostic modalities improves the physician’s ability to preoperatively assess women with adnexal mass. Diagnostic techniques that are commonly used are: clinical exam and thorough medical history, imaging (e.g. transvaginal ultrasound) and serum tumour maker (e.g. CA125) measurements. According to a study by the US Agency for Healthcare Research and Quality, which assessed diagnostic strategies for distinguishing benign from malignant masses, all current diagnostic modalities showed significant trade-offs between sensitivity and specificity [7]. Although the serum CA125 test does not have FDA-cleared indication as preoperative diagnostic aid in women with ovarian masses that are suspected to be malignant, CA125 is commonly used and recommended by the American Congress of Obstetricians and Gynecologists (ACOG) and the Society of Gynecologic Oncologists (SGO) for this indication [8,9]. The main clinical disadvantage of CA125 for adnexal mass assessment is its insufficient sensitivity for detecting early stage cancer and decreased specificity, due to false elevations in benign obstetric-gynaecologic conditions such as endometriosis, leiomyomas, pelvic inflammatory disease and pregnancy [10].
HE4 – ovarian cancer specific biomarker
HE4 (Human epididymis protein 4) is a member of a family of four disulphide core (WFDC) domain proteins and the function of this protein is unknown [11]. The HE4 gene is elevated in serum from women with ovarian cancer and its expression in normal tissues, including ovary, is low [12]. Several studies have indicated that using HE4 alone or in combination with CA125 may improve the accuracy for detection of ovarian cancer. In a study by Moore et al that evaluated nine known biomarkers for ovarian cancer, HE4 showed the highest sensitivity at a set specificity for the detection of ovarian cancer, particularly in early stage disease [13]. In this study, the combination of HE4 and CA 125 was a more accurate predictor of malignancy than either marker alone, with a sensitivity of 76% and a specificity of 95%. Additional studies confirmed that measuring serum HE4 concentrations along with CA 125 concentrations may provide higher accuracy for detecting ovarian cancer, and may improve the accuracy for detection of ovarian cancer at an earlier stage.
Additionally, a number of studies demonstrated improved specificity of HE4 for discriminating ovarian cancers from benign gynaecologic disease. Huhtinen et al was first to report that serum concentration of HE4 was significantly higher in patients with endometrial and ovarian cancer than in patients with ovarian endometriomas or other types of endometriosis [14]. These results were later confirmed in studies reported by Montagnana et al and Holcomb et al [15,16]. Recently, in a large study of 1042 pre- and postmenopausal women with benign gynaecological disorders HE4 was found to be less frequently elevated than CA125 in several benign diseases [17]. For example, HE4 was elevated in only 3% of premenopausal women with endometriosis, while in the same group CA125 was elevated in 72% of women. Unlike CA125, which can be elevated in one fourth of pregnant women and a third of patients with pelvic inflammatory diseases (PID), HE4 is not elevated in pregnancy and PID [16,18]. In addition, in healthy premenopausal women HE4 does not appear to oscillate during the menstrual cycle [19].
ROMA test: an aid in determining the likelihood of malignancy in women who
present with an adnexal mass
In September 2011, the ROMA test received clearance from the FDA as an aid in assessing whether a premenopausal or postmenopausal woman who presents with an adnexal mass is at high or low likelihood of having a malignancy. ROMA is a qualitative serum test that combines the results of two biomarkers, HE4 + CA 125, and menopausal status into a single score and is indicated for women who meet the following criteria: over age 18 and adnexal mass present for which surgery is planned.
The effectiveness of ROMA to aid in estimating the risk of malignancy was determined in a prospective, multi-centre, blinded clinical trial of 461 women over 18 years old (240 pre- and 221 post-menopausal) presenting with an adnexal mass that required surgical intervention [20]. For each patient, an initial cancer risk assessment (ICRA) was completed by a non-gynaecological oncologist, providing the generalist’s assessment of the patient’s mass as benign (negative) or malignant (positive) based upon the information available to the generalist during his/her work-up of the patient. The corresponding histopathology reports were collected and the stratification into low and a high risk groups for finding malignancy on surgery was determined using ROMA. The incidence of ovarian cancers was 10%. ROMA achieved 100% sensitivity at 74.5% specificity, a positive predictive value (PPV) of 13.8% and a negative predictive value (NPV) of 100% for stratification of premenopausal women with epithelial ovarian cancer into low likelihood and high likelihood groups of having malignancy. In postmenopausal women, ROMA had 92.3% sensitivity at 76.8% specificity, a PPV of 50.0% and NPV of 97.5% for stratification into low and high likelihood groups of having malignancy. When considering all women together ROMA had a sensitivity of 93.8%, a specificity of 74.9% and a NPV of 99.0%.
In a separate prospective, multicentre trial conducted at 12 US tertiary care institutions, 566 women undergoing surgery for adnexal mass were classified using ROMA into high and low likelihood groups for having epithelial ovarian cancer [21]. The incidence of ovarian cancers in this cohort was 23%. In the postmenopausal group at specificity of 75.0%, ROMA had sensitivity of 92.3%. In the premenopausal group at the specificity of 74.8% ROMA provided a sensitivity of 76.5% for classifying into high likelihood and low likelihood groups for having malignancy.
Additionally, seven distinct, single centre, multinational studies were published that validated the use of ROMA for adnexal mass risk stratification [22-28]. Combined, these studies assessed over 4,000 women with adnexal mass that were scheduled to undergo surgery in the United States, Europe and Asia. The range of sensitivity for ROMA test was from 75 % – 94%, at specificity from 75% – 95%. ROMA demonstrated consistent and reliable performance for classifying women with adnexal mass into high risk and low likelihood groups for epithelial ovarian cancer.
Conclusions
In the US, women with adnexal masses present primarily to gynaecologists, primary care physicians or general surgeons for initial diagnostic evaluation. According to a Practice Bulletin from the American Congress of Obstetrics and Gynecology (ACOG) an important dilemma is faced by these physicians as to which patients are appropriate for referral to a gynaecologic oncologist, and/or to an institution experienced in gynaecologic cancer surgery. Several recent studies have demonstrated that ovarian cancer patients managed by gyneacologic oncologists and at high volume institutions are more likely to undergo complete surgical staging, and optimal cytoreductive surgery with fewer complications and better survival rates than patients treated by surgeons less familiar with the management of ovarian cancer. Based on the available clinical evidence, ROMA test represents an important tool for improved triage of women diagnosed with an adnexal mass which can ultimately lead to improved patient outcomes.
References
1. Jemal A et al. CA Cancer J Clin 2010; 60(5): 277-300.
2. Giede KC et al. Gynecol Oncol 2005; 99(2):447-61.
3. 1999-2006 National Cancer Institute –Surveillance Epidemiology and End Results (NCI-SEER)
4. Goff BA et al. Cancer 2007; 109: 221-27.
5. Trimble EL. Gynecol Oncol 1994 ; 55(3 Pt 2): S1-3.
6. Danforth’s Obstetrics and Gynecology, ed. B.Y.K. Ronald S. Gibbs, Arthur F. Haney, Ingrid Nygaard. 2008: Lippincott Williams & Wilkins.
7. Myers ER et al. 2006; 130: 1-145.
8. ACOG Practice Bulletin. Obstet Gynecol. 2007; 110: 201-213.
9. The Society of Gynecologic Oncologists. Gynecol Oncol 2000; 78(3 Pt 2): S1-13
10. Jacobs I, Bast RC Jr. Hum Reprod 1989; 4(1): 1-12
11. Bouchard D et al. Lancet Oncol 2006; 7: 167-74.
12. Drapkin R et al. Cancer Res 2006; 65: 2162-69.
13. Moore RG et al. Gynecologic Oncol 2008; 108: 402-408.
14. Hutinen K et al. Br J Cancer 2009; 100: 1315-1319.
15. Montagnana M et al. Br J Cancer 2009; 101(3): 548.
16. Holcomb K et al. Am J Obstet Gynecol Am J Obstet Gynecol 2011; 205(4): 358.e1-6.
17. Moore RG et al. Am J Obstet Gynecol 2012; 206(4): 351.e1-8.
18. Moore RG et al. Am J Obstet Gynecol 2012; 206(4): 349.e1-7.
19. Hallamaa M et al. Gynecol Oncol 2012 Mar 14. [Epub ahead of print]
20. Moore RG et al. Obstet Gynecol 2011; 118(2, Part 1): 280-288.
21. Moore RG et al. Gynecol Oncol 2009; 112(1): 40-6.
22. Van Gorp T et al. Br J Cancer 2011; 104(5): 863-870.
23. Jacob F et al. Gynecol Oncol 2011; 121(3): 487-491.
24. Lenhard M et al. Clin Chem Lab Med 2011 Sep 16.
25. Molina R et al. Tumour Biol 2011; 32(6): 1087-95.
26. Montagnana M et al. Clin Chem Lab Med 2011; 49(3): 521-525.
27. Ruggeri G et al. Clin Chim Acta 2011; 412(15-16): 1447-1453.
28. Kim YM et al. Clin Chem Lab Med 2011; 49(3): 527-534.
The author
Zivjena Vucetic, PhD
Fujirebio Diagnostics, Inc.
Factors impacting on sample collection for urinary schistosomiasis research in Abeokuta, Nigeria
, /in Featured Articles /by 3wmediaSample collection is an important aspect of scientific work because it shapes, to a great extent, the study design and methodology, both of which may influence the outcomes of scientific research. However, often in scientific evaluations of studies which involve both field sample collection and laboratory work, only the laboratory research aspect receives serious attention, while other factors such as the socio-cultural, ecological and belief values of subjects who donate samples for laboratory studies are much less emphasised. These factors and how they play out in any particular study area are critical determinants of successful field sample collection especially in the developing countries.
by Dr Olufunmilola Ibironke, Dr Samuel Asaolu and Dr Clive Shiff
Urinary schistosomiasis is caused by a trematode worm, Schistosoma haematobium [1]. Infection with this parasite has been shown to be the commonest cause of haematuria and urogenital diseases in endemic areas. Thus, detection of haematuria in urine has been proposed as a valid indicator of schistosome infection, and has been widely adopted in many national schistosomiasis control programmes [2,3]. Diagnostic procedures in control programmes accordingly involve collection of urine samples from patients.
Most studies of urinary schistosomiasis in Nigeria and other endemic countries have targeted schoolchildren [4-8], because they represent the prime reservoir for the parasite, and children are amenable to mass chemotherapy [9]. However, studies have shown the debilitating effect of the parasite among adults in communities where it is endemic [10-13] and so this population also needs to be studied. As opposed to urine sample collection from children which is mostly done in schools, collection of urine from adults is difficult, particularly among persons who do not consider schistosomiasis as their major health problem when compared to malaria. In a school-based setting, after obtaining clearance from government health and school administrative authorities, researchers usually work with school teachers to get permission from pupils’ parents, and to educate the children involved in the study about how to follow urine sample collection instructions. However, for studies which involve adults, researchers, with the help of local health officers, would have to deal with patients directly to seek their individual involvement in the study, the acceptance of which depends on a number of the above mentioned factors.
Few studies have investigated the sociology of communities involved in such studies. We present here a study on urinary schistosomiasis in two villages in Ogun State, Nigeria, involving collection of urine samples from adults, to investigate the factors that drive their acceptance or refusal for inclusion in the study.
Methods and study sites
The study involved adults between the ages of 20 and 55 years who were mobilised to school halls in each village through the respective heads of the villages. Participants were informed of their right to accept or reject inclusion in the study. Many adults refused to come to school halls, many others who came rejected inclusion in the study. Some others accepted inclusion and collected urine sample containers but never came back while others accepted full participation. People in endemic communities show negative attitudes to urine sample collection for different reasons. To find out villagers’ attitudes to the urine sample collection process, we asked consenting participants why their friends or family refused to participate and in the process we identified some factors responsible for their attitudes. We also visited some households either to seek consent for inclusion or to understand reasons for refusing inclusion in the study.
This study was conducted in July, 2010, in Ogun State, Nigeria as a part of a study on the diagnosis of urinary schistosomiasis in six villages. For the purpose of comparison, two villages, Apojola located in Odeda Local Government Area (LGA), and Ogbere in Ijebu-east LGA, were selected. Apojola is located on Oyan Dam Reservoir. The inhabitants are all immigrant fishermen and their families, and are a mixture of Moslem Hausas and Christian Idomas. Awawa River serves Ogbere community. The inhabitants are mainly Christian Yorubas, and a mixture of farmers and Local Government Area civil servants. Ethical consideration, the data collection process, the population of each village, vegetation types and locations of each local government area have been reported previously [14].
Observations and discussion
Socio-cultural aspect
Several urinary schistosomiasis studies had been conducted in Nigeria, most of which involved urine sample collection, so there is a high level of awareness about the importance of control programmes. However, in the process of field studies there is often confusion in the minds of the participants leading to fear of exposure to strangers which was found to prevail among the villagers. Frequently researchers are mistaken for government agents visiting for revenue collections. If the researcher can work with members of the community to change these opinions it would likely improve level of cooperation for inclusion in the study. We explored this aspect in Apojola, a community located on the heavily schistosome-infested Oyan dam reservoir. We made the first attempt to recruit participants through the community leader, followed by the religious leader, a nurse and a school teacher. The number of participants recruited through the assistance of the different leaders according to age and gender are shown in Table 1a. In Table 1b, it was shown that the community leader is the most effective in helping to mobilise the villagers of both genders for urine collection.
There is also an increasing cynicism about the disease among adult patients in endemic communities. Many members of the communities who admit passing blood in the urine do not perceive it as an indication of a serious disease. They consider it as a sign of virility and puberty which is a familiar sign among adults in other villages around them. A few others who have experienced some discomfort and thought it might be a major health problem were either ashamed of their disease status or ashamed of bringing their ‘red’ urine. Past studies have noted that individuals’ perceptions on the aetiology and impact of urinary schistosomiasis differed with their levels of education and gender [13]. Lack of knowledge about the cause and effect of the disease affects patient’s turnout for sample collection and this in turn has a direct influence on field data coverage and research quality.
Apart from lack of health education on the cause of the disease, the willingness to participate in the urine sample collection process is seemingly greater among patients with some level of education than among the uneducated. We investigated how patient’s level of education impacts turnout for urine sample collection in Ogbere community. Ogbere inhabitants are a mixture of uneducated farmers, who have nought to six years of formal education, and the educated comprising teachers and Local Government Area civil servants, who have from seven to 16 years of formal education. In Table 2, data from both groups are presented for comparison to show turnout according to education level and gender.
This Table shows the percentage contributions by the Community Leader (CL), Nurse (N), Teacher (T) and Religious Leader (RL) on the total number of respondents. CL is best for mobilising males in the community (P = 0.00155). CL is also best for mobilising male and female with calculated P = 0.052 just higher than 0.05. N is best for mobilising females but this is not statistically significant.
Ecological aspect
Transmission of urinary schistosomiasis is through freshwater snails, Bulinus species, as intermediate hosts and varies with different ecological factors. In many endemic communities, the ecological factors which favour disease transmission also promote agricultural practices such as farming, cattle rearing and fishing. Therefore, transmission to humans often occurs as a result of irrigation systems for agricultural purposes or when visits are made to the rivers for washing and swimming. As such, the rate of transmission to humans varies, to a great extent, with occupation.
However, since diagnosis is by urine testing, many peasant farmers and fishermen who are thought to be the most impacted with S. haematobium because of frequent water contact may remain undiagnosed and untreated. Urine sample collection for the diagnosis of urinary schistosomiasis is preferably done between the hours of 10:00 and 14:00 for optimum egg passage [9]. These hours coincide with the time during which farmers go to farm and fishermen set nets for fish catching. This coincidence might affect turnout for sample collection and estimation of overall disease prevalence in the community.
To evaluate the impact of patient’s occupation on turnout for urine sample collection, we compared turnout of farmers and civil servants in Ogbere community. For statistical purpose, farmers, cattle rearers and fishermen are classified as farming, while students, teachers and local government workers are classified as civil servants, see Table 2. In total, there are 84 participants out of which 33 are farmers (39.2%) and 51 are civil servants (60.7%). In all, more women (79.8%) turned out for sample collection.
According to the community leader, the total adults’ population in Ogbere is 3121 and the ratio of farmers to civil servants is approximately 20:1.
Z- Distribution test was used to compare the response level between the two groups using the formula:
(see picture number 4)
where p is the difference of proportions, N1 = 149 = Educated population and N2 = 2972 = Uneducated population. At all levels of significance 0.05, 0.01 and 0.001, response from the educated civil servant population was significantly higher than response from the uneducated farmer population.
Belief structures
Christians in Apojola and Ogbere communities were relatively unhindered by religious belief regarding their willingness to come forward for education about the project and provision of their urine samples. However there was gender problem with urine collection among the Muslim families at Apojola. The Muslim families at Apojola have the culture of restricting married women within the family household compounds and forbiding male visitors of adolescent age and older from entering the compounds or visiting the women. In order to be able to collect urine samples from these Muslim women, the local community nurse and a female member of our research team were accompanied by a local female Muslim field assistant and interpreter before being allowed access to the compounds to explain the importance of the disease and purpose of the study.
Conclusion
This study attempts to find out patients‘ attitudes to scientific research especially during a field sample collection process and suggests possible reasons for rejection of inclusion in scientific research by patients. In general, this study showed that social and ecological values including educational background, occupation, religious practices and poor knowledge about the aims and objectives of the study, strongly influence turnout for urine sample collection. Therefore, such values are worth considering for a holistic understanding of the scientific study results.
References
1. Edungbola LD, Asaolu SO, Omonisi MK, Aiyedun BA. Schistosoma haematobium infection among schoolchildren in the Babana district, Kwara State, Nigeria. Afr J Med Sci 1988; 7: 187-193.
2. Koukounari A, Gabrielli AF, Toure S, Bosque-Oliva E, Zhang Y, Sellin B, Donnelly CA, Fenwick A, Webster JP. Schistosoma haematobium infection and morbidity before and after large-scale administration of praziquantel in Burkina Faso. J Infect Dis 2007; 196: 659-669.
3. Webster JP, Koukounari A, Lamberton PH, Stothard JR, Fenwick A. Evaluation and application of potential schistosome-associated morbidity markers within large-scale mass chemotherapy programmes. Parasitology 2009; 136: 1789-1799.
4. Abdel-Wahab MF, Esmat G, Ramzy I, Fouad R, Abdel-Rahman M, Yosery A, Narooz S, Strickland GT. Schistosoma haematobium infection in Egyptian schoolchildren: demonstration of both hepatic and urinary tract morbidity by ultrasonography. Trans R Soc Trop Med Hyg 1992; 86: 406-409.
5. Fenwick A, Webster JP, Bosque-Oliva E, Blair L, Fleming FM, Zhang Y, Garba A, Stothard JR, Gabrielli AF, Clements AC, Kabatereine NB, Toure S, Dembele R, Nyandindi U, Mwansa J et al. The Schistosomiasis Control Initiative (SCI): rationale, development and implementation from 2002-2008. Parasitology 2009; 136: 1719-1730.
6. French MD, Rollinson D, Basanez MG, Mgeni AF, Khamis IS, Stothard JR. School-based control of urinary schistosomiasis on Zanzibar, Tanzania: monitoring micro-haematuria with reagent strips as a rapid urological assessment. J Pediatr Urol 2007; 3: 364-368.
7. Nduka FO, Ajaero CM, Nwoke BE. Urinary schistosomiasis among school children in an endemic community in south-eastern Nigeria. Appl Parasitol 1995; 36: 34-40.
8. Okoli EI, Odaibo AB. Urinary schistosomiasis among schoolchildren in Ibadan, an urban community in south-western Nigeria. Trop Med Int Health 1999; 4: 308-315.
9. Ibironke OA, Phillips AE, Garba A, Lamine SM, Shiff C. Diagnosis of Schistosoma haematobium by detection of specific DNA fragments from filtered urine samples. Am J Trop Med Hyg 2011; 84: 998-1001.
10. Koukounari A, Webster JP, Donnelly CA, Bray BC, Naples J, Bosompem K, Shiff C. Sensitivities and specificities of diagnostic tests and infection prevalence of Schistosoma haematobium estimated from data on adults in villages northwest of Accra, Ghana. Am J Trop Med Hyg 2009; 80: 435-441.
11. Mostafa MH, Sheweita SA, O’Connor PJ. Relationship between schistosomiasis and bladder cancer. Clin Microbiol Rev 1999; 12: 97-111.
12. Mungadi IA,.Malami SA. Urinary bladder cancer and schistosomiasis in North-Western Nigeria. West Afr J Med 2007; 26: 226-229.
13. Sarkinfada F, Oyebanji AA, Sadiq IA, Ilyasu Z. Urinary schistosomiasis in the Danjarima community in Kano, Nigeria. J Infect Dev Ctries 2009; 3: 452-457.
14. Ibironke O, Koukounari A, Asaolu S, Moustaki I, Shiff C. Validation of a new test for Schistosoma haematobium based on detection of Dra1 DNA fragments in urine: evaluation through latent class analysis. PLoS Negl Trop Dis 2012; 6: e1464.
The authors
Dr Olufunmiola Ibironke*
Cell and DNA Repository
Rutgers, The State University of New Jersey
New Brunswick
New Jersey, USA
e-mail: oai5@rutgers.edu
Dr Clive Shiff
Department of Molecular Microbiology and Immunology
Johns Hopkins Bloomberg School of Public Health
Baltimore, MD, USA
e-mail: cshiff@jhsph.edu
Dr Samuel Asaolu
Department of Zoology
Obafemi Awolowo University
Ile-Ife
Nigeria
*Corresponding author
The ‘Virtual Patient’ in healthcare: IT Future of Medicine
, /in Featured Articles /by 3wmediaTo be able to mobilise our healthcare system to treat patients as individuals rather than as members of larger, divergent groups, the IT Future of Medicine (ITFoM) initiative proposes to develop a new, data rich computation-based individualised medicine of the future, based on integrated molecular, physiological and anatomical models of every person (‘Virtual Patient’) in the healthcare system. The establishment of such ‘Virtual Patient’ models is now possible due to the enormous progress in analytical techniques, particularly in the ‘omics’ technology areas and in imaging, as well as sensor technologies. Complemented by continuing developments in ICT, these technological developments could, over the coming years, make the ‘Virtual Patient’ a key component in healthcare and disease therapy and prevention. ITFoM is an European consortium combining unparalleled expertise in medicine, analytics and ICT to develop the ‘Virtual patient’.
by the ITFoM consortium
Today´s medicine
Currently medicine assesses patients as parts of large, often inhomogeneous groups. Rather than as individuals, patients are treated as members of a group for which a specific therapy has been statistically shown to be more effective than other therapies. This is even regardless of the fact that this therapy might very well make the majority of patients more ill than they would be without treatment.
Today’s medicine does not take into account the tremendous diversity between human individuals. Moreover, diseases are not homogenous either in regard to clinical manifestation or underlying causative effects. In cancer this is taken to an extreme with each tumour being different, because each of these tumours is the product of a specific and unique accumulation of mutational events.
Symptoms and signs of disease often appear only late in disease progression when a large portion of the involved organ has already failed. The symptoms might be non-specific, making a diagnosis difficult. Today´s routine clinical workup of sick patients can be extensive, expensive and can have side effects. For these reasons, many advocate preventive measures that mandate predefined checkups to be carried out by primary care physicians. Only a few preventive measures are currently useful including blood pressure control, blood sugar and lipid measurements, colonoscopy in older people, gynaecological tests in women and last but not least weight control. Both in the presence or absence of symptoms and signs of diseases, the knowledge of the full genome, the metabolome, the proteome, the microbiome and the total exposure to toxins from the environment, would have a tremendous impact on both disease workup and preventive measures.
Tomorrow´s medicine
The medicine of the future will use a ‘Virtual Patient’ system that can integrate all molecular, physiological and anatomical data into personalised models of individual people, enabling prediction of the result of lifestyle choices and medical interventions on a tailored case-by-case basis. This innovative approach will revolutionise healthcare systems, with enormous benefits for prevention, diagnosis and therapy of patients. The possibility to personalise the models allows tailor-made therapy and treatment strategies for each individual. With the model-based decision of which drug or which doses of drugs will have the optimal effect in an individual patient, the model approach will help to optimise treatment and reduce side-effects dramatically. A model-based approach will also serve as a research tool to discover and validate new compounds for drug development, potential drug treatments and applications, but also new commercial opportunities in ICT, analytics and healthcare.
ITFoM: IT Future of Medicine
ITFoM – one of the six pilot initiatives within the European Future and Emerging Technologies Flagship scheme competing for a total of 1 billion EUR over a time span of 10 years – will lay the groundwork for a project that will integrate medicine, analytical techniques and IT hardware and software development for the IT driven, data-rich, individualised medicine of the future.
By now, it has become quite conceivable to develop sequencing strategies allowing the determination of the genome, epigenome and transcriptome of a tumour, for instance, in parallel to its surgical removal, allowing the surgeon to scale the extent of the operation based on the real time computational modelling of its detailed genomic, epigenomic and transcriptomic characterisation. Dramatic improvements are also expected in the capabilities of other molecular analysis techniques, such as proteomics and metabolomics.
Why ITFoM makes the difference in ‘personalised medicine’: next generation of molecular analytics
The generation of the first draft of the human genome was a worldwide concerted action that had a strong impact on the development of new technologies for molecular biology. During the last ten years high throughput technologies have been emerging not only for DNA sequencing, but also for protein and metabolite analysis. These high throughput technologies are called ‘omics’ technologies, highly parallelised approaches aiming at the generation of information on complete sets of molecules in organelles, cells, whole pathways or even organs in order to get a comprehensive view of a biological system. A variety of ‘omics’ subdisciplines have emerged, each developing its own instruments, techniques and processes. With the increasing amount of data generated by the ‘omics’ technologies, development of tools for intelligent mathematical analysis and data mining are needed. This demand has developed into a completely new area in biology, namely bioinformatics.
For the first driver of the ‘omics’ technologies, DNA sequencing, currently the so-called ‘third generation’ sequencing technology is already appearing on the market. This innovation will allow the sequencing of a whole genome within one day, the costs for sequencing are in almost free fall, it can be anticipated that very soon the goal of sequencing a whole genome for less than 1.000 $ will be reached. These innovations open the door to allowing the sequencing of the genome of each single patient and using this information for truly personalised medicine. DNA sequencing is also used to study transcriptional expression, microRNA, DNA methylation, hydroxymethylation, transcription factor
occupancy, histone modification at specific sites in the genome and overall organisation of genomes in cells.
The personal genome information will be a very important basis for future medicine, but more ‘omics’ information will be integrated: information about proteins and metabolites will allow a much more precise picture of the physiological status of a person. The aim for protein and metabolite analysis now is to apply a method that allows the detection of all proteins and all metabolites in a given sample or tissue. The same holds true for the information about protein modifications and interactions.
Other lab technologies for molecular analysis including imaging and sensor technology are also starting to increase in speed, precision, application range and information output.
Another level of complexity takes into account life style and environmental factors, and more specifically the microorganisms interacting with the human body.
All these technologies allow the generation of highly detailed information about an individual’s genetic make-up and physiological status to give an unprecedented insight into the functioning of a person’s cells, tissues, organs and even the individual as a whole.
Systems biology is a solution that provides the methodologies and tools for mathematical analysis, integration and interpretation of biological data, employing mathematical models of biological processes. Mathematical models support the understanding of data sets on a large scale and integrate existing knowledge for interpretation. Model approaches in the ITFoM will drive the development further into models that are able to generate computational simulations to predict what cannot be measured directly. The translation of these novel approaches into clinical application will allow identification of the optimal therapy or medical treatment for each person based on the individual data available.
To generate the models and implement the ‘Virtual Patient’ model into clinical practice, substantial advances must be made in underpinning hardware and software infrastructures, computational paradigms, human computer interfaces and visualisation, as well as in the instrumentation and automation of techniques required to gather and process all relevant information. Examples of the major challenges in the information and communication technologies are interoperability, data storage and processing, efficient use of computing power, statistics and medical informatics. Integration of the individual datasets is realised via the ITFoM ‘Virtual Patient’ models enabling the provision of concrete health advice on a personal basis.
The authors
IT Future of Medicine Consortium (ITFoM)
Max Planck Institute for Molecular Genetics
Ihnestrasse 63-73
14195 Berlin
Germany
A Step Closer – BC-6800: Closer is clearer
, /in Featured Articles /by 3wmediaDon’t Miss the signs – Freelite
, /in Featured Articles /by 3wmediaSTA®-Liquid Anti-Xa
, /in Featured Articles /by 3wmedia